Evaluation of Nonpoint-Source Contamination, Wisconsin: Selected Topics for Water Year 1995

USGS Publications Warehouse

Owens, D.W.; Corsi, Steven R.; Rappold, K.F.

1997-01-01

The objective of the watershed-management evaluation monitoring program in Wisconsin is to evaluate the effectiveness of best-management practices (BMP's) for controlling nonpoint-source contamination in eight rural and four urban watersheds. This report, the fourth in an annual series of reports, presents a summary of the data collected for the program by the U.S. Geological Survey and the results of several detailed analyses of the data. To complement assessments of water quality, a land-use and BMP inventory is ongoing for 12 evaluation monitoring projects to track nonpoint sources of contamination in each watershed and to document implementation of BMP's that were designed to cause changes in the water quality of streams. Each year, updated information is gathered, mapped, and stored in a geographic-information-system data base. Summaries of land-use, BMP implementation, and water-quality data collected during water years 1989-95 are presented. Storm loads, snowmelt-period loads, and annual loads of suspended sediment and total phosphorus are summarized for eight rural sites. Storm-load data for suspended solids, total phosphorus, total recoverable lead, copper, zinc, and cadmium are summarized for four urban sites. Quality-assurance and quality-control (QA/QC) samples were collected at the eight rural sites to evaluate inorganic sample contamination and at one urban site to evaluate sample-collection and filtration techniques for polycyclic aromatic hydrocarbons (PAR's). Some suspended solids and fecal coliform contamination was detected at the rural sites. Corrective actions will be taken to address this contamination. Evaluation of PAR sample-collection techniques did not uncover any deficiencies, but the small amount of data collected was not sufficient to draw any definite conclusions. Evaluation of PAR filtration techniques indicate that water-sample filtration with O.7-um glass-fiber filters in an aluminum filter unit does not result in significant loss of PAR.

BMP7 Induces Dormancy of Prostatic Tumor Stem Cell in Bone

DTIC Science & Technology

2012-10-01

the upper back of nude mice. Recombinant human BMP7 was peritumorally injected daily after implantation. Tumor growth was monitored weekly by...of BMP7 was administrated daily through i.v. after intracardiac injection of CSCs from PC3mm or C4-2B cells to the mice. As shown in Figure 5A...mice, and then BMP7 was administrated daily . BLI of representative mice in each group six weeks after implantation (A). Normalized BLI signals

OCP TECD Report - TARDEC Blast Mitigation Program (BMP) and National Defense Industrial Association (NDIA) Michigan (MI) Chapter Cooperative Research and Development Agreement (CRADA) Summary

DTIC Science & Technology

2017-09-26

on the desire of both parties to collaborate to develop a practical and useable set of Blast Mitigation Program (BMP) design guidelines and...standards. The effort focused on the following topics: occupant-centric design philosophy and terms, test procedures, Military performance specifications...NDIA) was created based on the desire of both parties to collaborate to develop a practical and useable set of Blast Mitigation Program (BMP) design

Bmp signaling regulates a dose-dependent transcriptional program to control facial skeletal development.

PubMed

Bonilla-Claudio, Margarita; Wang, Jun; Bai, Yan; Klysik, Elzbieta; Selever, Jennifer; Martin, James F

2012-02-01

We performed an in depth analysis of Bmp4, a critical regulator of development, disease, and evolution, in cranial neural crest (CNC). Conditional Bmp4 overexpression, using a tetracycline-regulated Bmp4 gain-of-function allele, resulted in facial skeletal changes that were most dramatic after an E10.5 Bmp4 induction. Expression profiling uncovered a signature of Bmp4-induced genes (BIG) composed predominantly of transcriptional regulators that control self-renewal, osteoblast differentiation and negative Bmp autoregulation. The complimentary experiment, CNC inactivation of Bmp2, Bmp4 and Bmp7, resulted in complete or partial loss of multiple CNC-derived skeletal elements, revealing a crucial requirement for Bmp signaling in membranous bone and cartilage development. Importantly, the BIG signature was reduced in Bmp loss-of-function mutants, indicating Bmp-regulated target genes are modulated by Bmp dose. Chromatin immunoprecipitation (ChIP) revealed a subset of the BIG signature, including Satb2, Smad6, Hand1, Gadd45γ and Gata3, that was bound by Smad1/5 in the developing mandible, revealing direct Smad-mediated regulation. These data support the hypothesis that Bmp signaling regulates craniofacial skeletal development by balancing self-renewal and differentiation pathways in CNC progenitors.

Status of state forestry best management practices for the southeastern United States

Treesearch

R. Cristan; W.M. Aust; M.C. Bolding; S.M. Barrett; J.F. Munsell

2016-01-01

Forestry Best Management Practices (BMPs) are important measures for protecting the waters of the U.S., but few studies have compared monitoring strategies and implementation success of forestry BMPs across states. In order to assess the status of state forestry BMPs, a survey was sent to the state forestry agency in each U.S. state regarding their forestry BMP program...

Di-22:6-bis(monoacylglycerol)phosphate: A clinical biomarker of drug-induced phospholipidosis for drug development and safety assessment

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Nanjun; Tengstrand, Elizabeth A.; Chourb, Lisa

The inability to routinely monitor drug-induced phospholipidosis (DIPL) presents a challenge in pharmaceutical drug development and in the clinic. Several nonclinical studies have shown di-docosahexaenoyl (22:6) bis(monoacylglycerol) phosphate (di-22:6-BMP) to be a reliable biomarker of tissue DIPL that can be monitored in the plasma/serum and urine. The aim of this study was to show the relevance of di-22:6-BMP as a DIPL biomarker for drug development and safety assessment in humans. DIPL shares many similarities with the inherited lysosomal storage disorder Niemann–Pick type C (NPC) disease. DIPL and NPC result in similar changes in lysosomal function and cholesterol status that leadmore » to the accumulation of multi-lamellar bodies (myeloid bodies) in cells and tissues. To validate di-22:6-BMP as a biomarker of DIPL for clinical studies, NPC patients and healthy donors were classified by receiver operator curve analysis based on urinary di-22:6-BMP concentrations. By showing 96.7-specificity and 100-sensitivity to identify NPC disease, di-22:6-BMP can be used to assess DIPL in human studies. The mean concentration of di-22:6-BMP in the urine of NPC patients was 51.4-fold (p ≤ 0.05) above the healthy baseline range. Additionally, baseline levels of di-22:6-BMP were assessed in healthy non-medicated laboratory animals (rats, mice, dogs, and monkeys) and human subjects to define normal reference ranges for nonclinical/clinical studies. The baseline ranges of di-22:6-BMP in the plasma, serum, and urine of humans and laboratory animals were species dependent. The results of this study support the role of di-22:6-BMP as a biomarker of DIPL for pharmaceutical drug development and health care settings. - Highlights: • A reliable biomarker of drug-induced phospholipidosis (DIPL) is needed for humans. • Di-22:6-BMP is specific/sensitive for DIPL in animals as published in literatures. • The di-22:6-BMP biomarker can be validated for humans via NPC patients. • DIPL shares morphologic/mechanistic similarities with Niemann–Pick type C disease. • Di-22:6-BMP is an effective DIPL biomarker in humans via NPC patient validation.« less

Evaluation and Analysis of Regional Best Management Practices in San Diego, California (USA)

NASA Astrophysics Data System (ADS)

Flint, K.; Kinoshita, A. M.

2017-12-01

In urban areas, surface water quality is often impaired due to pollutants transported by stormwater runoff. To maintain and improve surface water quality, the United States Clean Water Act (CWA) requires an evaluation of available water quality information to develop a list of impaired water bodies and establish contaminant restrictions. Structural Best Management Practices (BMPs) are designed to reduce runoff volume and/or pollutant concentrations to comply with CWA requirements. Local level policy makers and managers require an improved understanding of the costs and benefits associated with BMP installation, performance, and maintenance. The International Stormwater BMP Database (Database) is an online platform for submittal of information about existing BMPs, such as cost, design details, and statistical analysis of influent and effluent pollutant concentrations. While the Database provides an aggregation of data which supports analysis of overall BMP performance at international and national scales, the sparse spatial distribution of the data is not suitable for regional and local analysis. This research conducts an extensive review of local inventory and spatial analysis of existing permanent BMPs throughout the San Diego River watershed in California, USA. Information collected from cities within the San Diego River watershed will include BMP types, locations, dates of installation, costs, expected removal efficiencies, monitoring data, and records of maintenance. Aggregating and mapping this information will facilitate BMP evaluation. Specifically, the identification of spatial trends, inconsistencies in BMP performances, and gaps in current records. Regression analysis will provide insight into the nature and significance of correlations between BMP performance and physical characteristics such as land use, soil type, and proximity to impaired waters. This analysis will also result in a metric of relative BMP performance and will provide a basis for future predictions of BMP effectiveness. Ultimately, results from this work will provide information to local governments and agencies for prioritizing, maintaining and monitoring BMPs, and improvement of hydrologic and water quality modeling in urban systems subject to compliance.

Host cell recruitment patterns by bone morphogenetic protein-2 releasing hyaluronic acid hydrogels in a mouse subcutaneous environment.

PubMed

Todeschi, Maria R; El Backly, Rania M; Varghese, Oommen P; Hilborn, Jöns; Cancedda, Ranieri; Mastrogiacomo, Maddalena

2017-07-01

This study aimed to identify host cell recruitment patterns in a mouse model in response to rhBMP-2 releasing hyaluronic acid hydrogels and influence of added nano-hydroxyapatite particles on rhBMP-2 release and pattern of bone formation. Implanted gels were retrieved after implantation and cells were enzymatically dissociated for flow cytometric analysis. Percentages of macrophages, progenitor endothelial cells and putative mesenchymal stem cells were measured. Implants were evaluated for BMP-2 release by ELISA and by histology to monitor tissue formation. Hyaluronic acid+BMP-2 gels influenced the inflammatory response in the bone healing microenvironment. Host-derived putative mesenchymal stem cells were major contributors. Addition of hydroxyapatite nanoparticles modified the release pattern of rhBMP-2, resulting in enhanced bone formation.

Detecting channel riparian vegetation response to best-management-practices implementation in ephemeral streams with the use of spot high-resolution visible imagery

USGS Publications Warehouse

Kamp, Kendall Vande; Rigge, Matthew B.; Troelstrup, Nels H.; Smart, Alexander J.; Wylie, Bruce

2013-01-01

Heavily grazed riparian areas are commonly subject to channel incision, a lower water table, and reduced vegetation, resulting in sediment delivery above normal regimes. Riparian and in-channel vegetation functions as a roughness element and dissipates flow energy, maintaining stable channel geometry. Ash Creek, a tributary of the Bad River in western South Dakota contains a high proportion of incised channels, remnants of historically high grazing pressure. Best management practices (BMP), including off-stream watering sources and cross fencing, were implemented throughout the Bad River watershed during an Environmental Protection Agency (EPA) 319 effort to address high sediment loads. We monitored prairie cordgrass (Spartina pectinata Link) establishment within stream channels for 16 yr following BMP implementation. Photos were used to group stream reaches (n = 103) subjectively into three classes; absent (estimated  40% cover; n = 16) based on the relative amount of prairie cordgrass during 2010 assessments of ephemeral channels. Reaches containing drainage areas of 0.54 to 692 ha were delineated with the use of 2010 National Agriculture Imagery Program (NAIP) imagery. Normalized difference vegetation index (NDVI) values were extracted from 5 to 39 sample points proportional to reach length using a series of Satellite Pour l'Observation de la Terre (SPOT) satellite imagery. Normalized NDVI (nNDVI) of 2 152 sample points were determined from pre- and post-BMP images. Mean nNDVI values for each reach ranged from 0.33 to 1.77. ANOVA revealed significant increase in nNDVI in locations classified as present prairie cordgrass cover following BMP implementation. Establishment of prairie cordgrass following BMP implementation was successfully detected remotely. Riparian vegetation such as prairie cordgrass adds channel roughness that reduces the flow energy responsible for channel degradation.

Green Tool

EPA Pesticide Factsheets

The Green Tool represents infiltration-based stormwater control practices. It allows modelers to select a BMP type, channel shape and BMP unit dimensions, outflow control devices, and infiltration method. The program generates an HSPF-formatted FTABLE.

Essential Roles of Epithelial Bone Morphogenetic Protein Signaling During Prostatic Development

PubMed Central

Omori, Akiko; Miyagawa, Shinichi; Ogino, Yukiko; Harada, Masayo; Ishii, Kenichiro; Sugimura, Yoshiki; Ogino, Hajime; Nakagata, Naomi

2014-01-01

Prostate is a male sex-accessory organ. The prostatic epithelia consist primarily of basal and luminal cells that differentiate from embryonic urogenital sinus epithelia. Prostate tumors are believed to originate in the basal and luminal cells. However, factors that promote normal epithelial differentiation have not been well elucidated, particularly for bone morphogenetic protein (Bmp) signaling. This study shows that Bmp signaling prominently increases during prostatic differentiation in the luminal epithelia, which is monitored by the expression of phosphorylated Smad1/5/8. To elucidate the mechanism of epithelial differentiation and the function of Bmp signaling during prostatic development, conditional male mutant mouse analysis for the epithelial-specific Bmp receptor 1a (Bmpr1a) was performed. We demonstrate that Bmp signaling is indispensable for luminal cell maturation, which regulates basal cell proliferation. Expression of the prostatic epithelial regulatory gene Nkx3.1 was significantly reduced in the Bmpr1a mutants. These results indicate that Bmp signaling is a key factor for prostatic epithelial differentiation, possibly by controlling the prostatic regulatory gene Nkx3.1. PMID:24731097

Dissecting BMP signaling input into the gene regulatory networks driving specification of the blood stem cell lineage

PubMed Central

Kirmizitas, Arif; Meiklejohn, Stuart; Ciau-Uitz, Aldo; Stephenson, Rachel; Patient, Roger

2017-01-01

Hematopoietic stem cells (HSCs) that sustain lifelong blood production are created during embryogenesis. They emerge from a specialized endothelial population, termed hemogenic endothelium (HE), located in the ventral wall of the dorsal aorta (DA). In Xenopus, we have been studying the gene regulatory networks (GRNs) required for the formation of HSCs, and critically found that the hemogenic potential is defined at an earlier time point when precursors to the DA express hematopoietic as well as endothelial genes, in the definitive hemangioblasts (DHs). The GRN for DH programming has been constructed and, here, we show that bone morphogenetic protein (BMP) signaling is essential for the initiation of this GRN. BMP2, -4, and -7 are the principal ligands expressed in the lineage forming the HE. To investigate the requirement and timing of all BMP signaling in HSC ontogeny, we have used a transgenic line, which inducibly expresses an inhibitor of BMP signaling, Noggin, as well as a chemical inhibitor of BMP receptors, DMH1, and described the inputs from BMP signaling into the DH GRN and the HE, as well as into primitive hematopoiesis. BMP signaling is required in at least three points in DH programming: first to initiate the DH GRN through gata2 expression, then for kdr expression to enable the DH to respond to vascular endothelial growth factor A (VEGFA) ligand from the somites, and finally for gata2 expression in the DA, but is dispensable for HE specification after hemangioblasts have been formed. PMID:28584091

Mechanisms of impaired nephrogenesis with fetal growth restriction: altered renal transcription and growth factor expression

PubMed Central

Abdel-Hakeem, Ahmed K; Henry, Tasmia Q; Magee, Thomas R; Desai, Mina; Ross, Michael; Mansano, Roy; Torday, John; Nast, Cynthia C.

2010-01-01

Objective Maternal food restriction during pregnancy results in growth restricted newborns and reduced glomerular number, contributing to programmed offspring hypertension. We investigated whether reduced nephrogenesis may be programmed by dysregulation of factors controlling ureteric bud branching and mesenchyme to epithelial transformation. Study Design 10 to 20 days gestation, Sprague Dawley pregnant rats (n=6/group) received ad libitum food; FR rats were 50% food restricted. At embryonic day 20, mRNA and protein expression of WT1, Pax2, FGF2, GDNF, cRET, WNT4, WNT11, BMP4, BMP7, and FGF7 were determined by real-time PCR and Western blotting. Results Maternal FR resulted in up-regulated mRNA expression for WT1, FGF2, and BMP7 whereas Pax2, GDNF, FGF7, BMP4, WNT4, and WNT11 mRNAs were down-regulated. Protein expression was concordant for WT1, GDNF, Pax2, FGF7, BMP4 and WNT4. Conclusion Maternal FR altered gene expression of fetal renal transcription and growth factors, and likely contributes to development of offspring hypertension. PMID:18639218

Breast Cancer Research Program (BCRP) - Predoctoral Traineeship - Elucidating the Role of the Type III Transforming Growth Factor-beta Receptor in Bone Morphogenetic Signaling in Breast Cancer

DTIC Science & Technology

2008-03-01

phosphorylation in the pancreatic cancer cell model, Panc -1 (data not shown). This data emphasize that TβRIII’s role in BMP signaling is likely to be cell... Panc -1 cells were adenovirally infected with TβRIII, followed by BMP-4-induced EMT. The cells were then plated in Matrigel invasion chambers. A...BMP-2, while loss of TβRIII in Panc -1 (pancreatic cancer cells) increased cell sensitivity to BMP-2 and had no effect on MDA-MB-231 (breast cancer

Effects of best-management practices in Bower Creek in the East River priority watershed, Wisconsin, 1991-2009

USGS Publications Warehouse

Corsi, Steven R.; Horwatich, Judy A.; Rutter, Troy D.; Bannerman, Roger T.

2013-01-01

Hydrologic and water-quality data were collected at Bower Creek during the periods before best-management practices (BMPs), and after BMPs were installed for evaluation of water-quality improvements. The monitoring was done between 1990 and 2009 with the pre-BMP period ending in July 1994 and the post-BMP period beginning in October 2006. BMPs installed in this basin included streambank protection and fencing, stream crossings, grade stabilization, buffer strips, various barnyard-runoff controls, nutrient management, and a low degree of upland BMPs. Water-quality evaluations included base-flow concentrations and storm loads for total suspended solids, total phosphorus, and ammonia nitrogen. The only reductions detected between the base-flow samples of the pre- and post-BMP periods were in median concentrations of total phosphorus from base-flow samples, but not for total suspended solids or dissolved ammonia nitrogen. Differences in storm loads for the three water-quality constituents monitored were not observed during the study period.

Implementation of a best management practice (BMP) system for a clay mining facility in Taiwan.

PubMed

Lin, Jen-Yang; Chen, Yen-Chang; Chen, Walter; Lee, Tsu-Chuan; Yu, Shaw L

2006-01-01

The present paper describes the planning and implementation of a best management practice (BMP) system for a clay mining facility in Northern Taiwan. It is a challenge to plan and design BMPs for mitigating the impact of clay mining operations due to the fact that clay mining drainage typically contains very high concentrations of suspended solids (SS), Fe-ions, and [H+] concentrations. In the present study, a field monitoring effort was conducted to collect data for runoff quality and quantity from a clay mining area in Northern Taiwan. A BMP system including holding ponds connected in series was designed and implemented and its pollutant removal performance was assessed. The assessment was based on mass balance computations and an analysis of the relationship between BMP design parameters such as pond depth, detention time, surface loading rate, etc. and the pollutant removal efficiency. Field sampling results showed that the surface-loading rate is exponential related to the removing rate. The results provide the basis for a more comprehensive and efficient BMP implementation plan for clay mining operations.

Generation of a mouse with conditionally activated signaling through the BMP receptor, ALK2.

PubMed

Fukuda, Tomokazu; Scott, Gregory; Komatsu, Yoshihiro; Araya, Runa; Kawano, Masako; Ray, Manas K; Yamada, Masahisa; Mishina, Yuji

2006-04-01

BMP signaling plays pleiotropic roles in various tissues. Transgenic mouse lines that overexpress BMP signaling in a tissue-specific manner would be beneficial; however, production of each tissue-specific transgenic mouse line is labor-intensive. Here, using a Cre-loxP system, we generated a conditionally overexpressing mouse line for BMP signaling through the type I receptor ALK2 (alternatively known as AVCRI, ActRI, or ActRIA). By mating this line with Cre-expression mouse lines, Cre-mediated recombination removes an intervening floxed lacZ expression cassette and thereby permits the expression of a constitutively active form of Alk2 (caAlk2) driven by a ubiquitous promoter, CAG. Tissue specificity of Cre recombination was monitored by a bicistronically expressed EGFP following Alk2 cDNA. Increased BMP signaling was confirmed by ectopic phosphorylation of SMAD1/5/8 in the areas where Cre recombination had occurred. The conditional overexpression system described here provides versatility in investigating gene functions in a tissue-specific manner without having to generate independent tissue-specific transgenic lines. Published 2006 Wiley-Liss, Inc.

Maintenance and Monitoring of BMPS

EPA Science Inventory

Two best management practice (BMP) sites in the Staten Island Bluebelt in Richmond Creek Watershed are Richmond Creek 5 (RC-5) and Richmond Creek 4 (RC-4). This presentation includes site description, briefing of initial monitoring activity, representative maintenance activity, ...

Evaluation of nonpoint-source contamination, Wisconsin: Selected data for 1992 water year

USGS Publications Warehouse

Graczyk, D.J.; Walker, J.F.; Greb, S.R.; Corsi, Steven R.; Owens, D.W.

1993-01-01

This report presents the annual results of the U.S. Geological Survey's (USGS) watershed-management evaluation monitoring program in Wisconsin. The overall objective of each individual project in the program is to determine if the water chemistry in the receiving stream has changed as a result of the implementation of land-management practices in the watershed. This is accomplished through monitoring of water chemistry and ancillary variables before best-management practices (BMP's) are installed ('pre-BMP'), during installation ('transitional'), and after ('post-BMP') watershed- management plans have been completely implemented. Fecal-coliform (FC) counts ranged between 10 and 310,00/100 mL. A large range of values occurred within duplicate and triplicate samples as well as over time. The median percentage difference between duplicate and triplicate samples was 17 percent although 4 out of the total 60 duplicate and triplicate samples had differences greater than 100 percent. A decrease in FC counts generally occurred over the duration of the 4-day analyses. Linear regression models of the log-concentration values (dependent variable) with respect to time (independent variable) were calculated for all samples. Negative slopes were found for 14 of the 15 samples. Slopes varied from +0.5 to -38.4 percent gain/loss/day, with a median slope of -8.5 percent/day. A t-test was applied to the data to examine whether or not significant differences in FC counts exist with respect to holding times. Because the T-test only compares two treatments, the test was conducted 3 times (0 versus 24-hr holding time, 0 versus 48-hr holding time, and 0 versus 72-hr holding time). Setting the level of significance at p less than 0.05 and assuming equal variances, 27 percent (all from Bower and Otter Creeks) of the samples demonstrated a significant difference in colony count over the first 24 hr, 40 percent over 48 hr, and 47 percent over 72 hr. All samples that exhibited a significant change in colony count were because of a decrease in colony count of the sample.

[Reconstruction of maxillary sinus superior wall fractures with calcium phosphate cement/recombinant human bonemorphogenetic protein 7 compound implanted material in rabbit].

PubMed

Zhang, Qunhui; Yu, Feng; Zhang, Haoliang; Gong, Huicheng; Lin, Ying

2015-11-01

To evaluate the osteogenetic character and repairing maxillary sinus superior wall fractures capability of calcium phosphate cement (CPC) before and after combined with recombinant human bone morphogenetie protein-7(rhBMP-7). A 10 mmX5 mm bone defect in the maxillary sinus superior wall was induced by surgery in all 24 New Zealand white rabbits. These 24 rabbits were randomly divided into two groups. The defects were repaired with CPC group (n = 12) and CPC/rhBMP-7 group (n = 12). The osteogenesis of bone defect was monitored by gro'ss observation, histological examination, observation under scanning electron microscope and measurement of ALP activity at 6 and 12 weeks after the implantation. In group CPC,new bone was found to form slowly and little by little. In group CPC/rhBMP-7, however, new bone was observed to form early and massively. The ALP activity in group CPC showed significant statistical difference with that of group CPC/rhBMP-7 (P < 0.05). The CPC/rhBMP-7 composite has osteoconductibility and osteoinductibility, comparing the use of CPC/rhBMP-7 with CPC for the repair of orbital fracture, the former show obvious advantage repairing ability in maxillary sinus superior wall defect.

A synthetic compound that potentiates bone morphogenetic protein-2-induced transdifferentiation of myoblasts into the osteoblastic phenotype

PubMed Central

Kato, Satoshi; Tomita, Katsuro; Titus, Louisa; Boden, Scott D.

2011-01-01

There is an urgent need to develop methods that lower costs of using recombinant human bone morphogenetic proteins (BMPs) to promote bone induction. In this study, we demonstrate the osteogenic effect of a low-molecular weight compound, SVAK-12, that potentiated the effects of BMP-2 in inducing transdifferentiation of C2C12 myoblasts into the osteoblastic phenotype. Here, we report a specific compound, SVAK-12, which was selected based on in silico screenings of small-molecule databases using the homology modeled interaction motif of Smurf1-WW2 domain. The enhancement of BMP-2 activity by SVAK-12 was characterized by evaluating a BMP-specific reporter activity and by monitoring the BMP-2-induced expression of mRNA for osteocalcin and alkaline phosphatase (ALP), which are widely accepted marker genes of osteoblast differentiation. Finally, we confirmed these results by also measuring the enhancement of BMP-2-induced activity of ALP. Smurf1 is an E3 ligase that targets osteogenic Smads for ubiquitin-mediated proteasomal degradation. Smurf1 is an interesting potential target to enhance bone formation based on the positive effects on bone of proteins that block Smurf1-binding to Smad targets or in Smurf1−/− knockout mice. Since Smads bind Smurf1 via its WW2 domain, we performed in silico screening to identify compounds that might interact with the Smurf1-WW2 domain. We recently reported the activity of a compound, SVAK-3. However, SVAK-3, while exhibiting BMP-potentiating activity, was not stable and thus warranted a new search for a more stable and efficacious compound among a selected group of candidates. In addition to being more stable, SVAK-12 exhibited a dose-dependent activity in inducing osteoblastic differentiation of myoblastic C2C12 cells even when multiple markers of the osteoblastic phenotype were parallelly monitored. PMID:21110071

A synthetic compound that potentiates bone morphogenetic protein-2-induced transdifferentiation of myoblasts into the osteoblastic phenotype.

PubMed

Kato, Satoshi; Sangadala, Sreedhara; Tomita, Katsuro; Titus, Louisa; Boden, Scott D

2011-03-01

There is an urgent need to develop methods that lower costs of using recombinant human bone morphogenetic proteins (BMPs) to promote bone induction. In this study, we demonstrate the osteogenic effect of a low-molecular weight compound, SVAK-12, that potentiated the effects of BMP-2 in inducing transdifferentiation of C2C12 myoblasts into the osteoblastic phenotype. Here, we report a specific compound, SVAK-12, which was selected based on in silico screenings of small-molecule databases using the homology modeled interaction motif of Smurf1-WW2 domain. The enhancement of BMP-2 activity by SVAK-12 was characterized by evaluating a BMP-specific reporter activity and by monitoring the BMP-2-induced expression of mRNA for osteocalcin and alkaline phosphatase (ALP), which are widely accepted marker genes of osteoblast differentiation. Finally, we confirmed these results by also measuring the enhancement of BMP-2-induced activity of ALP. Smurf1 is an E3 ligase that targets osteogenic Smads for ubiquitin-mediated proteasomal degradation. Smurf1 is an interesting potential target to enhance bone formation based on the positive effects on bone of proteins that block Smurf1-binding to Smad targets or in Smurf1-/- knockout mice. Since Smads bind Smurf1 via its WW2 domain, we performed in silico screening to identify compounds that might interact with the Smurf1-WW2 domain. We recently reported the activity of a compound, SVAK-3. However, SVAK-3, while exhibiting BMP-potentiating activity, was not stable and thus warranted a new search for a more stable and efficacious compound among a selected group of candidates. In addition to being more stable, SVAK-12 exhibited a dose-dependent activity in inducing osteoblastic differentiation of myoblastic C2C12 cells even when multiple markers of the osteoblastic phenotype were parallelly monitored.

Probabilistic Determination of Green Infrastructure Pollutant Removal Rates from the International Stormwater BMP Database

NASA Astrophysics Data System (ADS)

Gilliom, R.; Hogue, T. S.; McCray, J. E.

2017-12-01

There is a need for improved parameterization of stormwater best management practices (BMP) performance estimates to improve modeling of urban hydrology, planning and design of green infrastructure projects, and water quality crediting for stormwater management. Percent removal is commonly used to estimate BMP pollutant removal efficiency, but there is general agreement that this approach has significant uncertainties and is easily affected by site-specific factors. Additionally, some fraction of monitored BMPs have negative percent removal, so it is important to understand the probability that a BMP will provide the desired water quality function versus exacerbating water quality problems. The widely used k-C* equation has shown to provide a more adaptable and accurate method to model BMP contaminant attenuation, and previous work has begun to evaluate the strengths and weaknesses of the k-C* method. However, no systematic method exists for obtaining first-order removal rate constants needed to use the k-C* equation for stormwater BMPs; thus there is minimal application of the method. The current research analyzes existing water quality data in the International Stormwater BMP Database to provide screening-level parameterization of the k-C* equation for selected BMP types and analysis of factors that skew the distribution of efficiency estimates from the database. Results illustrate that while certain BMPs are more likely to provide desired contaminant removal than others, site- and design-specific factors strongly influence performance. For example, bioretention systems show both the highest and lowest removal rates of dissolved copper, total phosphorous, and total nitrogen. Exploration and discussion of this and other findings will inform the application of the probabilistic pollutant removal rate constants. Though data limitations exist, this research will facilitate improved accuracy of BMP modeling and ultimately aid decision-making for stormwater quality management in urban systems.

Evaluation and implementation of BMPs for NCDOT's highway and industrial facilities : final report, May 2006.

DOT National Transportation Integrated Search

2006-05-01

This research has provided NCDOT with (1) scientific observations to validate the pollutant removal : performance of selected structural BMPs, (2) a database management option for BMP monitoring and : non-monitoring sites, (3) pollution prevention pl...

Innovative Stormwater Quality Tools by SARA for Holistic Watershed Master Planning

NASA Astrophysics Data System (ADS)

Thomas, S. M.; Su, Y. C.; Hummel, P. R.

2016-12-01

Stormwater management strategies such as Best Management Practices (BMP) and Low-Impact Development (LID) have increasingly gained attention in urban runoff control, becoming vital to holistic watershed master plans. These strategies can help address existing water quality impairments and support regulatory compliance, as well as guide planning and management of future development when substantial population growth and urbanization is projected to occur. However, past efforts have been limited to qualitative planning due to the lack of suitable tools to conduct quantitative assessment. The San Antonio River Authority (SARA), with the assistance of Lockwood, Andrews & Newnam, Inc. (LAN) and AQUA TERRA Consultants (a division of RESPEC), developed comprehensive hydrodynamic and water quality models using the Hydrological Simulation Program-FORTRAN (HSPF) for several urban watersheds in the San Antonio River Basin. These models enabled watershed management to look at water quality issues on a more refined temporal and spatial scale than the limited monitoring data. They also provided a means to locate and quantify potential water quality impairments and evaluate the effects of mitigation measures. To support the models, a suite of software tools were developed. including: 1) SARA Timeseries Utility Tool for managing and processing of large model timeseries files, 2) SARA Load Reduction Tool to determine load reductions needed to achieve screening levels for each modeled constituent on a sub-basin basis, and 3) SARA Enhanced BMP Tool to determine the optimal combination of BMP types and units needed to achieve the required load reductions. Using these SARA models and tools, water quality agencies and stormwater professionals can determine the optimal combinations of BMP/LID to accomplish their goals and save substantial stormwater infrastructure and management costs. The tools can also help regulators and permittees evaluate the feasibility of achieving compliance using BMP/LID. The project has gained national attention, being showcased in multiple newsletters, professional magazines, and conference presentations. The project also won the Texas American Council of Engineering Companies (ACEC) Gold Medal Award and the ACEC National Recognition Award in 2016.

Evaluating changes in water quality with respect to nonpoint source nutrient management strategies in the Chesapeake Bay Watershed

NASA Astrophysics Data System (ADS)

Keisman, J.; Sekellick, A.; Blomquist, J.; Devereux, O. H.; Hively, W. D.; Johnston, M.; Moyer, D.; Sweeney, J.

2014-12-01

Chesapeake Bay is a eutrophic ecosystem with periodic hypoxia and anoxia, algal blooms, diminished submerged aquatic vegetation, and degraded stocks of marine life. Knowledge of the effectiveness of actions taken across the watershed to reduce nitrogen (N) and phosphorus (P) loads to the bay (i.e. "best management practices" or BMPs) is essential to its restoration. While nutrient inputs from point sources (e.g. wastewater treatment plants and other industrial and municipal operations) are tracked, inputs from nonpoint sources, including atmospheric deposition, farms, lawns, septic systems, and stormwater, are difficult to measure. Estimating reductions in nonpoint source inputs attributable to BMPs requires compilation and comparison of data on water quality, climate, land use, point source discharges, and BMP implementation. To explore the relation of changes in nonpoint source inputs and BMP implementation to changes in water quality, a subset of small watersheds (those containing at least 10 years of water quality monitoring data) within the Chesapeake Watershed were selected for study. For these watersheds, data were compiled on geomorphology, demographics, land use, point source discharges, atmospheric deposition, and agricultural practices such as livestock populations, crop acres, and manure and fertilizer application. In addition, data on BMP implementation for 1985-2012 were provided by the Environmental Protection Agency Chesapeake Bay Program Office (CBPO) and the U.S. Department of Agriculture. A spatially referenced nonlinear regression model (SPARROW) provided estimates attributing N and P loads associated with receiving waters to different nutrient sources. A recently developed multiple regression technique ("Weighted Regressions on Time, Discharge and Season" or WRTDS) provided an enhanced understanding of long-term trends in N and P loads and concentrations. A suite of deterministic models developed by the CBPO was used to estimate expected nutrient load reductions attributable to BMPs. Further quantification of the relation of land-based nutrient sources and BMPs to water quality in the bay and its tributaries must account for inconsistency in BMP data over time and uncertainty regarding BMP locations and effectiveness.

Rockfall Milepoint 49 Monitoring Plan for Check Dams on Mt. Hood Highway (US 26) (M.P. 49.10 to M.P. 49.23)

DOT National Transportation Integrated Search

2003-07-31

This Stormwater BMP Monitoring Plan details the approach to be used for monitoring : roadside ditch sediment traps located on Highway 26 in the Mt. Hood National Forest. : These sediment traps were designed and installed by ODOT for the capture of se...

Improved Protocol for Chondrogenic Differentiation of Bone Marrow Derived Mesenchymal Stem Cells -Effect of PTHrP and FGF-2 on TGFβ1/BMP2-Induced Chondrocytes Hypertrophy.

PubMed

Nasrabadi, Davood; Rezaeiani, Siamak; Eslaminejad, Mohamadreza Baghaban; Shabani, Aliakbar

2018-04-24

Growth factors have a pivotal role in chondrogenic differentiation of stem cells. The differential effects of known growth factors involved in the maintenance and homeostasis of cartilage tissue have been previously studied in vitro. However, there are few reported researches about the interactional effects of growth factors on chondrogenic differentiation of stem cells. The aim of this study is to examine the combined effects of four key growth factors on chondrogenic differentiation of mesenchymal stem cells (MSCs). Isolated and expanded rabbit bone marrow-derived MSCs underwent chondrogenic differentiation in a micromass cell culture system that used a combination of the following growth factors: transforming growth factor beta 1 (TGF-β1), bone morphogenetic protein 2 (BMP2), parathyroid hormone related protein (PTHrP), and fibroblast growth factor 2 (FGF2) according to a defined program. The chondrogenic differentiation program was analyzed by histochemistry methods, quantitative RT-PCR (qRT-PCR), and measurement of matrix deposition of sulfated glycosaminoglycan (sGAG) and collagen content at days 16, 23, and 30. The results showed that the short-term combination of TGF-β1 and BMP-2 increased sGAG and collagen content, Alkaline phosphates (ALP) activity, and type X collagen (COL X) expression. Application of either PTHrP or FGF2 simultaneously decreased TGF-β1/BMP-2 induced hypertrophy and chondrogenic markers (at least for FGF2). However, successive application of PTHrP and FGF2 dramatically maintained the synergistic effects of TGF-β1/BMP-2 on the chondrogenic differentiation potential of MSCs and decreased unwanted hypertrophic markers. This new method can be used effectively in chondrogenic differentiation programs.

Review of wildlife resources of Vandenberg Air Force Base, California

NASA Technical Reports Server (NTRS)

Breininger, David R.

1989-01-01

Wildlife resources are reviewed for purposes of developing a Base Biological Monitoring Program (BMP) for Vandenberg Air Force Base (VAFB) in Santa Barbara County, California. The review and recommendations were prepared by review of applicable scientific literature and environmental documents for VAFB, discussing information needs with natural resource management professionals at VAFB, and observations of base field conditions. This process found that there are 29 federally listed vertebrates (endangered, threatened, or Category 2) that occur or may occur in the vicinity of VAFB. There are also 63 other state listed or regionally declining species that may occur in the vicinity of VAFB. Habitats of VAFB represent a very valuable environmental resource for rare and declining wildlife in California. However, little information is available on VAFB wildlife resources other than lists of species that occur or are expected to occur. Recommendations are presented to initiate a long-term wildlife monitoring program at VAFB to provide information for environmental impact assessment and wise land use planning.

MONITORING OF A RETENTION POND BEFORE AND AFTER MAINTENANCE

EPA Science Inventory

The USEPA’s Urban Watershed Management Branch monitored a retention pond with wetland plantings in the Richmond Creek (RC) watershed. This BMP, designated RC-5, is owned and operated by the New York City Department of Environmental Protection’s (DEP) as part of the Bluebelt progr...

A structured bowel management program for patients with severe functional constipation can help decrease emergency department visits, hospital admissions, and healthcare costs.

PubMed

Reck-Burneo, Carlos A; Vilanova-Sanchez, Alejandra; Gasior, Alessandra C; Dingemans, Alexander J M; Lane, Victoria A; Dyckes, Robert; Nash, Onnalisa; Weaver, Laura; Maloof, Tassiana; Wood, Richard J; Zobell, Sarah; Rollins, Michael D; Levitt, Marc A

2018-03-24

Published health-care costs related to constipation in children in the USA are estimated at $3.9 billion/year. We sought to assess the effect of a bowel management program (BMP) on health-care utilization and costs. At two collaborating centers, BMP involves an outpatient week during which a treatment plan is implemented and objective assessment of stool burden is performed with daily radiography. We reviewed all patients with severe functional constipation who participated in the program from March 2011 to June 2015 in center 1 and from April 2014 to April 2016 in center 2. ED visits, hospital admissions, and constipation-related morbidities (abdominal pain, fecal impaction, urinary retention, urinary tract infections) 12 months before and 12 months after completion of the BMP were recorded. One hundred eighty-four patients were included (center 1 = 96, center 2 = 88). Sixty-three (34.2%) patients had at least one unplanned visit to the ED before treatment. ED visits decreased to 23 (12.5%) or by 64% (p < 0.0005). Unplanned hospital admissions decreased from 65 to 28, i.e., a 56.9% reduction (p < 0.0005). In children with severe functional constipation, a structured BMP decreases unplanned visits to the ED, hospital admissions, and costs for constipation-related health care. 3. Copyright © 2018. Published by Elsevier Inc.

pHEMA-nHA encapsulation and delivery of vancomycin and rhBMP-2 enhances its role as a bone graft substitute.

PubMed

Li, Xinning; Xu, Jianwen; Filion, Tera M; Ayers, David C; Song, Jie

2013-08-01

Bone grafts are widely used in orthopaedic procedures. Autografts are limited by donor site morbidity while allografts are known for considerable infection and failure rates. A synthetic composite bone graft substitute poly(2-hydroxyethyl methacrylate)-nanocrystalline hydroxyapatite (pHEMA-nHA) was previously developed to stably press-fit in and functionally repair critical-sized rat femoral segmental defects when it was preabsorbed with a single low dose of 300 ng recombinant human bone morphogenetic protein-2/7 (rhBMP-2/7). To facilitate clinical translation of pHEMA-nHA as a synthetic structural bone graft substitute, we examined its ability to encapsulate and release rhBMP-2 and the antibiotic vancomycin. We analyzed the compressive behavior and microstructure of pHEMA-nHA as a function of vancomycin incorporation doses using a dynamic mechanical analyzer and a scanning electron microscope. In vitro release of vancomycin was monitored by ultraviolet-visible spectroscopy. Release of rhBMP-2 from pHEMA-nHA-vancomycin was determined by ELISA. Bioactivity of the released vancomycin and rhBMP-2 was examined by bacterial inhibition and osteogenic transdifferentiation capabilities in cell culture, respectively. Up to 4.8 wt% of vancomycin was incorporated into pHEMA-nHA without compromising its structural integrity and compressive modulus. Encapsulated vancomycin was released in a dose-dependent and sustained manner in phosphate-buffered saline over 2 weeks, and the released vancomycin inhibited Escherichia coli culture. The pHEMA-nHA-vancomycin composite released preabsorbed rhBMP-2 in a sustained manner over 8 days and locally induced osteogenic transdifferentiation of C2C12 cells in culture. pHEMA-nHA can encapsulate and deliver vancomycin and rhBMP-2 in a sustained and localized manner with reduced loading doses. The elasticity, osteoconductivity, and rhBMP-2/vancomycin delivery characteristics of pHEMA-nHA may benefit orthopaedic reconstructions or fusions with enhanced safety and efficiency and reduced infection risk.

Development of a Repeatable Regional Protocol for Performance-Based Monitoring of Forestry Best Management Practices

Treesearch

Roger Ryder; Pamela J. Edwards; Pamela J. Edwards

2005-01-01

There has been a long-standing interest in improving Best Management Practice (BMP) monitoring within and among states. States monitoring the implementation and effectiveness of BMPs for forest operations take a variety of approaches. This creates inconsistencies in data collection and how results are reported. Since 1990 attempts have been made to develop a consistent...

Acceleration of Alveolar Ridge Augmentation Using a Low Dose of Recombinant Human Bone Morphogenetic Protein-2 Loaded on a Resorbable Bioactive Ceramic.

PubMed

Fahmy, Rania A; Mahmoud, Naguiba; Soliman, Samia; Nouh, Samir R; Cunningham, Larry; El-Ghannam, Ahmed

2015-12-01

The aim of the present study was to evaluate the effect of a porous silica-calcium phosphate composite (SCPC50) loaded with and without recombinant human bone morphogenetic protein-2 (rhBMP-2) on alveolar ridge augmentation in saddle-type defects. Micro-granules of SCPC50 resorbable bioactive ceramic were coated with rhBMP-2 10 mg and then implanted into a saddle-type defect (12 × 7 mm) in a dog mandible and covered with a collagen membrane. Control groups included defects grafted with SCPC50 granules without rhBMP-2 and un-grafted defects. Bone healing was evaluated at 8 and 16 weeks using histologic and histomorphometric techniques. The increase in bone height and total defect fill were assessed for each specimen using the ImageJ 1.46 program. The release kinetics of rhBMP-2 was determined in vitro. The height of the bone in the grafted defects and the total defect fill were statistically analyzed. SCPC50 enhanced alveolar ridge augmentation as indicated by the increased vertical bone height, bone surface area, and bone volume after 16 weeks. SCPC50-rhBMP-2 provided a sustained release profile of a low effective dose (BMP-2 4.6 ± 1.34 pg/mL per hour) during the 1- to 21-day period. The slow rate of release of rhBMP-2 from SCPC50 accelerated synchronized complete bone regeneration and graft material resorption in 8 weeks. Successful rapid reconstruction of the alveolar ridge by SCPC50 and SCPC50-rhBMP-2 occurred without any adverse excessive bone formation, inflammation, or fluid-filled voids. Results of this study suggest that SCPC50 is an effective graft material to preserve the alveolar ridge after tooth extraction. Coating SCPC50-rhBMP-2 further accelerated bone regeneration and a considerable increase in vertical bone height. These findings make SCPC50 the primary choice as a carrier for rhBMP-2. SCPC50-rhBMP-2 can serve as an alternative to autologous bone grafting. Published by Elsevier Inc.

Quantification of BMPs Selection and Spatial Placement Impact on Water Quality Controlling Plans in Lower Bear River Watershed, Utah

NASA Astrophysics Data System (ADS)

Salha, A. A.; Stevens, D. K.

2016-12-01

The aim of the watershed-management program in Box Elder County, Utah set by Utah Division of Water Quality (UDEQ) is to evaluate the effectiveness and spatial placement of the implemented best-management practices (BMP) for controlling nonpoint-source contamination at watershed scale. The need to evaluate the performance of BMPs would help future policy and program decisions making as desired end results. The environmental and costs benefits of BMPs in Lower Bear River watershed have seldom been measured beyond field experiments. Yet, implemented practices have rarely been evaluated at the watershed scale where the combined effects of variable soils, climatic conditions, topography and land use/covers and management conditions may significantly change anticipated results and reductions loads. Such evaluation requires distributed watershed models that are necessary for quantifying and reproducing the movement of water, sediments and nutrients. Soil and Water Assessment Tool (SWAT) model is selected as a watershed level tool to identify contaminant nonpoint sources (critical zones) and areas of high pollution risks. Water quality concerns have been documented and are primarily attributed to high phosphorus and total suspended sediment concentrations caused by agricultural and farming practices (required load is 460 kg/day of total phosphorus based on 0.075 mg/l and an average of total suspended solids of 90 mg/l). Input data such as digital elevation model (DEM), land use/Land cover (LULC), soils, and climate data for 10 years (2000-2010) is utilized along with observed water quality at the watershed outlet (USGS) and some discrete monitoring points within the watershed. Statistical and spatial analysis of scenarios of management practices (BMP's) are not implemented (before implementation), during implementation, and after BMP's have been studied to determine whether water quality of the two main water bodies has improved as required by the LBMR watershed's TMDL and if the BMPs are cost-effectively targeting the critical zones.

The effectiveness and resilience of phosphorus management practices in the Lake Simcoe watershed, Ontario, Canada

NASA Astrophysics Data System (ADS)

Crossman, J.; Futter, M. N.; Palmer, M.; Whitehead, P. G.; Baulch, H. M.; Woods, D.; Jin, L.; Oni, S. K.; Dillon, P. J.

2016-09-01

Uncertainty surrounding future climate makes it difficult to have confidence that current nutrient management strategies will remain effective. This study used monitoring and modeling to assess current effectiveness (% phosphorus reduction) and resilience (defined as continued effectiveness under a changing climate) of best management practices (BMPs) within five catchments of the Lake Simcoe watershed, Ontario. The Integrated Catchment Phosphorus model (INCA-P) was used, and monitoring data were used to calibrate and validate a series of management scenarios. To assess current BMP effectiveness, models were run over a baseline period 1985-2014 with and without management scenarios. Climate simulations were run (2070-2099), and BMP resilience was calculated as the percent change in effectiveness between the baseline and future period. Results demonstrated that livestock removal from water courses was the most effective BMP, while manure storage adjustments were the least. Effectiveness varied between catchments, influenced by the dominant hydrological and nutrient transport pathways. Resilience of individual BMPs was associated with catchment sensitivity to climate change. BMPs were most resilient in catchments with high soil water storage capacity and small projected changes in frozen-water availability and in soil moisture deficits. Conversely, BMPs were less resilient in catchments with larger changes in spring melt magnitude and in overland flow proportions. Results indicated that BMPs implemented are not always those most suited to catchment flow pathways, and a more site-specific approach would enhance prospects for maintaining P reduction targets. Furthermore, BMP resilience to climate change can be predicted from catchment physical properties and present-day hydrochemical sensitivity to climate forcing.

Investigation on the effectiveness of pretreatment in stormwater management technologies.

PubMed

Maniquiz-Redillas, Marla C; Geronimo, Franz Kevin F; Kim, Lee-Hyung

2014-09-01

The effectiveness of presettling basins as component of stormwater best management practice (BMP) technologies was investigated. Storm event monitoring and sediment collection were conducted from May 2009 to November 2012 on the presettling basins of the three BMP technologies designed to capture and treat stormwater runoff from highly impervious roads and parking lots. Data on captured runoff and sediment, total suspended solids (TSS) loadings, rainfall and runoff rate, sediment accumulation rate, as well as particle distribution and pollutant concentrations of sediment were gathered and analyzed along with the physical design characteristics of the presettling basins such as surface area and storage volume. Regression models were generated to determine significant relationships between design parameters. Results revealed that the storage volume ratio (ratio of storage volume of presettling basin to BMP) was an important parameter in designing the presettling basin of the BMP. For practicality, optimizing the design of the presettling basin means that the storage volume ratio should be determined based on the desired captured amount of runoff and sediment from runoff to limit the frequency of maintenance caused by the accumulation of sediment. It was recommended that pretreatment of runoff should be employed when the site in which the BMP is to be sited has high TSS loading and runoff rate, and is subjected to high intensity rainfall. Copyright © 2014. Published by Elsevier B.V.

BMP signaling in dermal papilla cells is required for their hair follicle-inductive properties

PubMed Central

Rendl, Michael; Polak, Lisa; Fuchs, Elaine

2008-01-01

Hair follicle (HF) formation is initiated when epithelial stem cells receive cues from specialized mesenchymal dermal papilla (DP) cells. In culture, DP cells lose their HF-inducing properties, but during hair growth in vivo, they reside within the HF bulb and instruct surrounding epithelial progenitors to orchestrate the complex hair differentiation program. To gain insights into the molecular program that maintains DP cell fate, we previously purified DP cells and four neighboring populations and defined their cell-type-specific molecular signatures. Here, we exploit this information to show that the bulb microenvironment is rich in bone morphogenetic proteins (BMPs) that act on DP cells to maintain key signature features in vitro and hair-inducing activity in vivo. By employing a novel in vitro/in vivo hybrid knockout assay, we ablate BMP receptor 1a in purified DP cells. When DPs cannot receive BMP signals, they lose signature characteristics in vitro and fail to generate HFs when engrafted with epithelial stem cells in vivo. These results reveal that BMP signaling, in addition to its key role in epithelial stem cell maintenance and progenitor cell differentiation, is essential for DP cell function, and suggest that it is a critical feature of the complex epithelial–mesenchymal cross-talk necessary to make hair. PMID:18281466

BMP Effectiveness/Efficiency Monitoring Annotated Bibliography

DOT National Transportation Integrated Search

2002-05-17

The Oregon Department of Transportation (ODOT) has installed several stormwater treatment facilities to : improve the quality of runoff discharged from the States highway network. These facilities include a : variety of both above ground and below...

BMP7 and SHH regulate Pax2 in mouse retinal astrocytes by relieving TLX repression.

PubMed

Sehgal, Rachna; Sheibani, Nader; Rhodes, Simon J; Belecky Adams, Teri L

2009-08-15

Pax2 is essential for development of the neural tube, urogenital system, optic vesicle, optic cup and optic tract. In the eye, Pax2 deficiency is associated with coloboma, a loss of astrocytes in the optic nerve and retina, and abnormal axonal pathfinding of the ganglion cell axons at the optic chiasm. Thus, appropriate expression of Pax2 is essential for astrocyte determination and differentiation. Although BMP7 and SHH have been shown to regulate Pax2 expression, the molecular mechanism by which this regulation occurs is not well understood. In this study, we determined that BMP7 and SHH activate Pax2 expression in mouse retinal astrocyte precursors in vitro. SHH appeared to play a dual role in Pax2 regulation; 1) SHH may regulate BMP7 expression, and 2) the SHH pathway cooperates with the BMP pathway to regulate Pax2 expression. BMP and SHH pathway members can interact separately or together with TLX, a repressor protein in the tailless transcription factor family. Here we show that the interaction of both pathways with TLX relieves the repression of Pax2 expression in mouse retinal astrocytes. Together these data reveal a new mechanism for the cooperative actions of signaling pathways in astrocyte determination and differentiation and suggest interactions of regulatory pathways that are applicable to other developmental programs.

Spatially-Distributed Cost-Effectiveness Analysis Framework to Control Phosphorus from Agricultural Diffuse Pollution.

PubMed

Geng, Runzhe; Wang, Xiaoyan; Sharpley, Andrew N; Meng, Fande

2015-01-01

Best management practices (BMPs) for agricultural diffuse pollution control are implemented at the field or small-watershed scale. However, the benefits of BMP implementation on receiving water quality at multiple spatial is an ongoing challenge. In this paper, we introduce an integrated approach that combines risk assessment (i.e., Phosphorus (P) index), model simulation techniques (Hydrological Simulation Program-FORTRAN), and a BMP placement tool at various scales to identify the optimal location for implementing multiple BMPs and estimate BMP effectiveness after implementation. A statistically significant decrease in nutrient discharge from watersheds is proposed to evaluate the effectiveness of BMPs, strategically targeted within watersheds. Specifically, we estimate two types of cost-effectiveness curves (total pollution reduction and proportion of watersheds improved) for four allocation approaches. Selection of a ''best approach" depends on the relative importance of the two types of effectiveness, which involves a value judgment based on the random/aggregated degree of BMP distribution among and within sub-watersheds. A statistical optimization framework is developed and evaluated in Chaohe River Watershed located in the northern mountain area of Beijing. Results show that BMP implementation significantly (p >0.001) decrease P loss from the watershed. Remedial strategies where BMPs were targeted to areas of high risk of P loss, deceased P loads compared with strategies where BMPs were randomly located across watersheds. Sensitivity analysis indicated that aggregated BMP placement in particular watershed is the most cost-effective scenario to decrease P loss. The optimization approach outlined in this paper is a spatially hierarchical method for targeting nonpoint source controls across a range of scales from field to farm, to watersheds, to regions. Further, model estimates showed targeting at multiple scales is necessary to optimize program efficiency. The integrated model approach described that selects and places BMPs at varying levels of implementation, provides a new theoretical basis and technical guidance for diffuse pollution management in agricultural watersheds.

Single bead-based electrochemical biosensor.

PubMed

Liu, Changchun; Schrlau, Michael G; Bau, Haim H

2009-12-15

A simple, robust, single bead-based electrochemical biosensor was fabricated and characterized. The sensor's working electrode consists of an electrochemically etched platinum wire, with a nominal diameter of 25 microm, hermetically heat-fusion sealed in a pulled glass capillary (micropipette). The sealing process does not require any epoxy or glue. A commercially available, densely functionalized agarose bead was mounted on the tip of the etched platinum wire. The use of a pre-functionalized bead eliminates the tedious and complicated surface functionalization process that is often the bottleneck in the development of electrochemical biosensors. We report on the use of a biotin agarose bead-based, micropipette, electrochemical (Bio-BMP) biosensor to monitor H(2)O(2) concentration and the use of a streptavidin bead-based, micropipette, electrochemical (SA-BMP) biosensor to detect DNA amplicons. The Bio-BMP biosensor's response increased linearly as the H(2)O(2) concentration increased in the range from 1 x 10(-6) to 1.2 x10(-4)M with a detection limit of 5 x 10(-7)M. The SA-BMP was able to detect the amplicons of 1pg DNA template of B. Cereus bacteria, thus providing better detection sensitivity than conventional gel-based electropherograms.

Effects of best-management practices in Eagle and Joos Valley Creeks in the Waumandee Creek Priority Watershed, Wisconsin, 1990-2007

USGS Publications Warehouse

Graczyk, David J.; Walker, John F.; Bannerman, Roger T.; Rutter, Troy D.

2012-01-01

In many watersheds, nonpoint-source contamination is a major contributor to water-quality problems. In response to the recognition of the importance of nonpoint sources, the Wisconsin Nonpoint Source Water Pollution Abatement Program (Nonpoint Program) was enacted in 1978. This report summarizes the results of a study to assess the effectiveness of watershed-management practices for controlling nonpoint-source contamination for the Eagle Creek and Joos Valley Creek Watersheds. Streamflow-gaging stations equipped for automated sample collection and continuous recording of stream stage were installed in July 1990 at Eagle and Joos Valley Creeks and were operated through September 2007. In October 1990, three rain gages were installed in each watershed and were operated through September 2007. Best-Management Practices (BMPs) were installed during 1993 to 2000 in Eagle and Joos Valley Creeks and were tracked throughout the study period. By the year 2000, a majority of the BMPs were implemented in the two watersheds and goals set by the Wisconsin Department of Natural Resources and the local Land Conservation Department had been achieved for the two study watersheds (Wisconsin Department of Natural Resources, 1990). The distributions of the rainstorms that produced surface runoff and storm loads were similar in the pre-BMP (1990-93) and post-BMP implementation (2000-07) periods for both Eagle and Joos Valley Creeks. The highest annual streamflow occurred at both sites in water year 1993, which corresponded to the greatest above normal nonfrozen precipitation measured at two nearby NOAA weather stations. The minimum streamflow occurred in water year 2007 at both sites. Base-flow and stormwater samples were collected and analyzed for suspended solids, total phosphorus, and ammonia nitrogen. For both Eagle and Joos Valley Creeks the median concentrations of suspended solids and total phosphorus in base flow were lower during the post-BMP period compared to the pre-BMP period and were statistically significant at the 0.05 significance level. The decrease in median concentrations of ammonia nitrogen at both sites was not statistically significant at the 0.05 significance level. Multiple linear regression analyses were used to remove the effects of climatologic conditions and seasonality from computed storm loads. For both Eagle and Joos Valley Creeks, the median storm loads for suspended solids, total phosphorus, and ammonia nitrogen were lower during the post-BMP period compared to the pre-BMP period and were statistically significant at the 0.05 significance level. The decreases in storm-load regression residuals from the pre- to the post-BMP periods for both Eagle and Joos Valley Creeks were statistically significant for all three constituents at the 0.05 significance level and indicated an apparent improvement in water-quality in the post-BMP period. Because the rainfall characteristics for individual storms in the pre- and post-BMP periods are likely to be different, separate pre- and post-BMP regressions were used to estimate the theoretical pre- and post-BMP storm loads to allow estimates of precent reductions between the pre- and post-BMP periods. The estimated percent reductions in storm loads for suspended solids, total phosphorus, and ammonia nitrogen were 89, 77, and 66 respectively for Eagle Creek and 84, 67, and 60 respectively for Joos Valley Creek. The apparent improvement in water quality is attributed to the implemented BMPs and to a reduction in the number of cattle in the watersheds.

Morphologic and molecular alteration during tibia fracture healing in rat.

PubMed

Yu, M-D; Su, B-H; Zhang, X-X

2018-03-01

To monitor morphological feature and related osteogenic and bone metabolic change during healing of tibia fracture in a rat model. Tibia density and trabecular thickness were evaluated. Histopathology was examined by HE staining. Serous inflammatory factors IL-4, IL-6, TNF-α and metabolic biomarkers ALP, β-CTX, P1NP, were determined by ELISA. The expression of RUNX2, TGF-β1, VEGF-α, BMP-2, BMP-4, and BMP-7 in callus tissue were qualified by RT-PCR. Bone density decreased until week 4 and then increased post-operation. Trabeculae in callus were thickened over time with active osteogenesis. ELISA indicated the most severe inflammation at week 2, with the highest level of TNF-α, IL-6, and the lowest level of IL-4. After 4 weeks, the inflammation was alleviated accompanying with the decline of TNF-α and IL-6, while there was the elevation of IL-4. Bone metabolism showed active osteogenesis and resorption at week 6 with high P1NP and β-CTX. The expression of RUNX2, TGF-β1, VEGF-α, BMP-2, BMP-4, and BMP-7 increased progressively from week 1 to 6. The major lesions at week 2 in sham were tissue necrosis, periosteal reactive hyperplasia, inflammatory cell infiltration, capillary hyperplasia and slight fibro-blast cytopoiesis. At week 4, proliferation was greatly activated, fibrous callus shaped and chondrogenesis and some osteogenesis occurred at week 8. In rat model, bone density started to increase at week 6 after fracture, accompanied with trabeculae thickening, serous inflammatory factors decline, and peaked bone morphogenetic protein/growth factors, which indicated active osteogenesis was conforming to the classical phase of secondary fracture healing.

Evaluating the biochemical methane potential (BMP) of low-organic waste at Danish landfills.

PubMed

Mou, Zishen; Scheutz, Charlotte; Kjeldsen, Peter

2014-11-01

The biochemical methane potential (BMP) is an essential parameter when using first order decay (FOD) landfill gas (LFG) generation models to estimate methane (CH4) generation from landfills. Different categories of waste (mixed, shredder and sludge waste) with a low-organic content and temporarily stored combustible waste were sampled from four Danish landfills. The waste was characterized in terms of physical characteristics (TS, VS, TC and TOC) and the BMP was analyzed in batch tests. The experiment was set up in triplicate, including blank and control tests. Waste samples were incubated at 55°C for more than 60 days, with continuous monitoring of the cumulative CH4 generation. Results showed that samples of mixed waste and shredder waste had similar BMP results, which was in the range of 5.4-9.1 kg CH4/ton waste (wet weight) on average. As a calculated consequence, their degradable organic carbon content (DOCC) was in the range of 0.44-0.70% of total weight (wet waste). Numeric values of both parameters were much lower than values of traditional municipal solid waste (MSW), as well as default numeric values in current FOD models. The sludge waste and temporarily stored combustible waste showed BMP values of 51.8-69.6 and 106.6-117.3 kg CH4/ton waste on average, respectively, and DOCC values of 3.84-5.12% and 7.96-8.74% of total weight. The same category of waste from different Danish landfills did not show significant variation. This research studied the BMP of Danish low-organic waste for the first time, which is important and valuable for using current FOD LFG generation models to estimate realistic CH4 emissions from modern landfills receiving low-organic waste. Copyright © 2014 Elsevier Ltd. All rights reserved.

Development and optimization of a cell-based assay for the selection of synthetic compounds that potentiate bone morphogenetic protein-2 activity‡

PubMed Central

Okada, Motohiro; Sangadala, Sreedhara; Liu, Yunshan; Yoshida, Munehito; Reddy, Boojala Vijay B.; Titus, Louisa; Boden, Scott D.

2010-01-01

The requirement of large amounts of the recombinant human bone morphogenetic protein-2 (BMP-2) produces a huge translational barrier for its routine clinical use due to high cost. This leads to an urgent need to develop alternative methods to lower costs and/or increase efficacies for using BMP-2. In this study, we describe the development and optimization of a cell-based assay that is sensitive, reproducible, and reliable in identifying reagents that potentiate the effects of BMP-2 in inducing transdifferentiation of C2C12 myoblasts into the osteoblastic phenotype. The assay is based on a BMP-responsive Smad1-driven luciferase reporter gene. LIM mineralization protein-1 (LMP-1) is a novel intracellular LIM domain protein that has been shown by our group to enhance cellular responsiveness to BMP-2. Our previous report elucidated that the binding of LMP-1 with the WW2 domain in Smad ubiquitin regulatory factor-1 (Smurf1) rescues the osteogenic Smads from degradation. Here, using the optimized cell-based assay, we first evaluated the activity of the recombinantly prepared proteins, LMP-1, and its mutant (LMP-1ΔSmurf1) that lacks the Smurf1-WW2 domain-binding motif. Both the wild type and the mutant proteins were engineered to contain an 11-amino acid HIV-TAT protein derived membrane transduction domain to aid the cellular delivery of recombinant proteins. The cell-based reporter assay confirmed that LMP-1 potentiates the BMP-induced stimulation of C2C12 cells towards the osteoblastic phenotype. The potentiating effect of LMP-1 was significantly reduced when a specific-motif known to interact with Smurf1 was mutated. We validated the results obtained in the reporter assay by also monitoring the expression of mRNA for osteocalcin and alkaline phosphatase (ALP) which is widely accepted osteoblast differentiation marker genes. Finally, we provide further confirmation of our results by measuring the activity of alkaline phosphatase in support of the accuracy and reliability of our cell-based assay. Direct delivery of synthesized protein can be limited by high cost, instability or inadequate post-translational modifications. Thus, there would be a clear benefit for a low cost, cell penetrable chemical compound. We successfully used our gene expression-based assay to choose an active compound from a select group of compounds that were identified by computational screenings as the most likely candidates for mimicking the function of LMP-1. Among them, we selected SVAK-3, a compound that showed a dose-dependent potentiation of BMP-2 activity in inducing osteoblastic differentiation of C2C12 cells. We show that either the full length LMP-1 protein or its potential mimetic compound consistently exhibit similar potentiation of BMP-2 activity even when multiple markers of the osteoblastic phenotype were parallely monitored. PMID:19862690

Development and optimization of a cell-based assay for the selection of synthetic compounds that potentiate bone morphogenetic protein-2 activity.

PubMed

Okada, Motohiro; Sangadala, Sreedhara; Liu, Yunshan; Yoshida, Munehito; Reddy, Boojala Vijay B; Titus, Louisa; Boden, Scott D

2009-12-01

The requirement of large amounts of the recombinant human bone morphogenetic protein-2 (BMP-2) produces a huge translational barrier for its routine clinical use due to high cost. This leads to an urgent need to develop alternative methods to lower costs and/or increase efficacies for using BMP-2. In this study, we describe the development and optimization of a cell-based assay that is sensitive, reproducible, and reliable in identifying reagents that potentiate the effects of BMP-2 in inducing transdifferentiation of C2C12 myoblasts into the osteoblastic phenotype. The assay is based on a BMP-responsive Smad1-driven luciferase reporter gene. LIM mineralization protein-1 (LMP-1) is a novel intracellular LIM domain protein that has been shown by our group to enhance cellular responsiveness to BMP-2. Our previous report elucidated that the binding of LMP-1 with the WW2 domain in Smad ubiquitin regulatory factor-1 (Smurf1) rescues the osteogenic Smads from degradation. Here, using the optimized cell-based assay, we first evaluated the activity of the recombinantly prepared proteins, LMP-1, and its mutant (LMP-1DeltaSmurf1) that lacks the Smurf1-WW2 domain-binding motif. Both the wild type and the mutant proteins were engineered to contain an 11-amino acid HIV-TAT protein derived membrane transduction domain to aid the cellular delivery of recombinant proteins. The cell-based reporter assay confirmed that LMP-1 potentiates the BMP-induced stimulation of C2C12 cells towards the osteoblastic phenotype. The potentiating effect of LMP-1 was significantly reduced when a specific-motif known to interact with Smurf1 was mutated. We validated the results obtained in the reporter assay by also monitoring the expression of mRNA for osteocalcin and alkaline phosphatase (ALP) which is widely accepted osteoblast differentiation marker genes. Finally, we provide further confirmation of our results by measuring the activity of alkaline phosphatase in support of the accuracy and reliability of our cell-based assay. Direct delivery of synthesized protein can be limited by high cost, instability or inadequate post-translational modifications. Thus, there would be a clear benefit for a low cost, cell penetrable chemical compound. We successfully used our gene expression-based assay to choose an active compound from a select group of compounds that were identified by computational screenings as the most likely candidates for mimicking the function of LMP-1. Among them, we selected SVAK-3, a compound that showed a dose-dependent potentiation of BMP-2 activity in inducing osteoblastic differentiation of C2C12 cells. We show that either the full length LMP-1 protein or its potential mimetic compound consistently exhibit similar potentiation of BMP-2 activity even when multiple markers of the osteoblastic phenotype were parallely monitored.

BMP UNIT PROCESS DESIGN, MONITORING, AND PERFORMANCE

EPA Science Inventory

Urbanization results in more impervious areas that cause larger quantities of stormwater runoff. This runoff can contribute significant amounts of pollutants (e.g., litter, oils, microorganisms, sediments, nutrients, organic matter, and heavy metals) to receiving waters. To impro...

Benchmark study on glyphosate-resistant cropping systems in the United States. Part 6: Timeliness of economic decision-making in implementing weed resistance management strategies.

PubMed

Weirich, Jason W; Shaw, David R; Coble, Keith H; Owen, Micheal D K; Dixon, Philip M; Weller, Stephen C; Young, Bryan G; Wilson, Robert G; Jordan, David L

2011-07-01

The introduction of glyphosate-resistant (GR) crops in the late 1990s made weed control in maize, cotton and soybean simple. With the rapid adoption of GR crops, many growers began to rely solely on glyphosate for weed control. This eventually led to the evolution of GR weeds. Growers are often reluctant to adopt a weed resistance best management practice (BMP) because of the added cost of additional herbicides to weed control programs which would reduce short-term revenue. This study was designed to evaluate when a grower that is risk neutral (profit maximizing) or risk averse should adopt a weed resistance BMP. Whether a grower is risk neutral or risk averse, the optimal decision would be to adopt a weed resistance BMP when the expected loss in revenue is greater than 30% and the probability of resistance evolution is 0.1 or greater. However, if the probability of developing resistance increases to 0.3, then the best decision would be to adopt a weed resistance BMP when the expected loss is 10% or greater. Given the scenarios analyzed, risk-neutral or risk-averse growers should implement a weed resistance BMP with confidence that they have made the right decision economically and avoided the risk of lost revenue from resistance. If the grower wants to continue to see the same level of return, adoption of BMP is required. Copyright © 2011 Society of Chemical Industry.

Identification and genetic effect of haplotype in the bovine BMP7 gene.

PubMed

Huang, Yong-Zhen; Wang, Xin-Lei; He, Hua; Lan, Xian-Yong; Lei, Chu-Zhao; Zhang, Chun-Lei; Chen, Hong

2013-12-15

Bone morphogenetic proteins (BMPs) are peptide growth factors belonging to the transforming growth factor-beta (TGF-β) superfamily, and some members of the BMP family support white adipocyte differentiation. In this study, we focused on the BMP7 which singularly promotes the differentiation of brown preadipocytes. Haplotypes involving 5 single nucleotide polymorphism (SNP) sites in the bovine BMP7 gene were identified and their effect on body weight was analyzed. 16 haplotypes and 18 combined haplotypes were revealed and the linkage disequilibrium was assessed in the cattle population with 602 individuals representing three main cattle breeds from China. The results showed that haplotypes 3, 10 and 14 were predominant and accounted for 75.64%, 69.85%, and 83.36% in Nanyang, Qinchuan and Jiaxian cattle breeds, respectively. The statistical analyses indicated that the SNP 1, 4, and 5 are associated with the body weight, body length, and heart girth at 12 and 24 months in Nanyang cattle population (P<0.05), whereas there is no significant association between their 16 haplotypes and 18 combined haplotypes. Our results provide evidence that some SNPs and haplotypes in BMP7 are associated with growth traits, and may be utilized as a genetic marker in marker-assisted selection for beef cattle breeding programs. Copyright © 2013. Published by Elsevier B.V.

7 CFR 634.1 - Purpose and scope.

Code of Federal Regulations, 2011 CFR

2011-01-01

... best management practices (BMP's) in project areas which have critical water quality problems resulting... approved agricultural portion of a 208 water quality management plan. Participation in RCWP is voluntary. (c) The program is a new USDA program and an extension of existing water-quality management programs...

7 CFR 634.1 - Purpose and scope.

Code of Federal Regulations, 2010 CFR

2010-01-01

... best management practices (BMP's) in project areas which have critical water quality problems resulting... approved agricultural portion of a 208 water quality management plan. Participation in RCWP is voluntary. (c) The program is a new USDA program and an extension of existing water-quality management programs...

Optimizing farm landscape by two decision-support tools for present and future: A case study in a mountainous farm of Taiwan

NASA Astrophysics Data System (ADS)

Chou, S.; Lin, Y.

2013-12-01

Rapid expansion of agricultural land-use has been identified as the main factor degrading biodiversity. Many studies have indicated that habitat quality and connectivity for multiple species can be preserved by applying the systematic conservation planning and software programs for spatial conservation prioritizations are usually used by planners to solve conservation problems for present and future. However, each conservation software program uses different algorithms and may not be suitable or efficient for all case studies. Therefore, in this study we compared the performance of two commonly used decision-support tools, Marxan and Zonation, on identifying priority areas as reserve region for 16 bird species in the mountain area of Taiwan. The priority areas are considered as the results of the tradeoff between bird presence (biological factor) and agricultural products (economic factor). Marxan uses the minimum set approach to identify priority areas for meeting specific targets while Zonation uses the maximum coverage approach to identify priority areas given a fixed budget. Therefore, we design the scenario with the most comparable setting, which selects target-based planning as the removal rule and boundary length penalty option in zonation. The landscape composition and configuration of the simulated priority areas were further evaluated by using landscape metrics and their similarity were examined by using Spearman's rank tests. The results showed that Marxan performed more efficiently while Zonation generated the priority areas in better connectivity. As the selection of conservation programs depends on users objectives and needs for present and future, this study provides useful information on determining suitable and efficient decision-support tools for future bird conservation. Conservation maps for Zonation based on different BLP parameter. The conservation value for Zonation is based on the hierarchical solution output. (a)BLP =1000 (b)BLP =3000 (c)BLP =5000 (d)BLP =7000 Conservation maps for Marxan based on different BMP parameter. The conservation value for Marxan is based on the selection frequency. (a)BMP =2500 (b)BMP =5000 (c)BMP =7500 (d)BMP =10000

Bone Morphogenetic Protein 15 (BMP15) Acts as a BMP and Wnt Inhibitor during Early Embryogenesis*

PubMed Central

Di Pasquale, Elisa; Brivanlou, Ali H.

2009-01-01

Bone morphogenetic protein 15 (BMP15) belongs to an unusual subgroup of the transforming growth factor β (TGFβ) superfamily of signaling ligands as it lacks a key cysteine residue in the mature region required for proper intermolecular dimerization. Naturally occurring BMP15 mutation leads to early ovarian failure in humans, and BMP15 has been shown to activate the Smad1/5/8 pathway in that context. Despite its important role in germ cell specification, the embryological function of BMP15 remains unknown. Surprisingly, we find that during early Xenopus embryogenesis BMP15 acts solely as an inhibitor of the Smad1/5/8 pathway and the Wnt pathway. BMP15 gain-of-function leads to embryos with secondary ectopic heads and to direct neural induction in intact explants. BMP15 inhibits BMP4-mediated epidermal induction in dissociated explants. BMP15 strongly inhibits BRE response induced by BMP4 and blocks phosphorylation and activation of Smad1/5/8 MH2-domain. Mechanistically, BMP15 protein specifically interacts with BMP4 protein, suggesting inhibition upstream of receptor binding. Loss-of-function experiments using morpholinos or a naturally occurring human BMP15 dominant-negative mutant (BMP15-Y235C) leads to embryos lacking head. BMP15-Y235C also eliminates the inhibitory activity of BMP15 on BRE (BMP-responsive element). Finally, we show that BMP15 inhibits the canonical branch of the Wnt pathway, upstream of β-catenin. We, thus, demonstrate that BMP15 is necessary and sufficient for the specification of dorso-anterior structures and highlight novel mechanisms of BMP15 function that strongly suggest a reinterpretation of its function in ovaries specially for ovarian failure. PMID:19553676

Effectiveness of a sediment time critical removal action-PCB reduction in fish tissue, surface water, and sediment via wet excavation.

PubMed

Santini, Andrew D; King, Todd; Krawczyk, Keith; Kern, John W

2015-01-01

Documenting successful remediation of polychlorinated biphenyl (PCB)-contaminated sediments is limited; potentially due to inadequate monitoring methods, complexities associated with the environment, and selected remedial techniques. At some sites, absence of appropriate baseline and postremoval monitoring limits proper evaluation of remedial efficacy. Accurate interpretation of interactions between media, space, time, species, lipid content, and remedial technique requires robust study design and data. This article presents baseline and postremoval data documenting reduced PCB concentrations in fish tissue, surface water, and sediment in response to the US Environmental Protection Agency (USEPA) time-critical removal action (TCRA) that was conducted at the former Bryant Mill Pond (BMP) on Portage Creek in Kalamazoo, Michigan. The BMP is part of an operable unit (OU) within the Allied Paper, Inc./Portage Creek/Kalamazoo River Superfund Site. PCBs discharged to the creek as a byproduct of carbonless copy paper recycling are the primary contaminant of concern. Paper waste residuals commonly appear as gray to light gray clays in river sediments and floodplain soils. The cleanup criterion was 10 mg/kg, with a residual PCB concentration goal of 1 mg/kg. Because the PCB-containing waste is (generally) associated with readily visible light gray clay, excavation of all visibly contaminated current or formerly impounded sediment served as a surrogate for the cleanup criteria and goal. Sediment was wet excavated and backfilled after diversion of the creek. After confirmation that PCB concentrations met cleanup criteria, the stream was diverted to the excavated side, and excavation and backfilling were completed. Overall, 146000 cubic yards of material including PCB-contaminated sediments were removed from the BMP. The long-term monitoring (LTM) program implemented by the Michigan Department of Environmental Quality (MDEQ), and historic data from a variety of sources (federal, state, and local agencies and responsible parties) provided data demonstrating TCRA effectiveness. Before TCRA, surface sediment PCB concentration in instream and formerly inundated sediment combined was 83 mg/kg (with a maximum of 700 mg/kg). Instream sediments that are more representative of fish exposures had a pre-TCRA surface weighted average concentration (SWAC) of 27 mg/kg. The post-TCRA SWAC for instream and floodplain sediments combined was 0.26 mg/kg. Average surface water concentrations at the downstream end of the BMP were reduced from 0.11 µg/L pre-TCRA to 0.0025 µg/L post-TCRA. Tissue samples for adult carp fillets decreased from 4 mg/kg pre-TCRA to 0.26 mg/kg post-TCRA; whole body white suckers from 3 mg/kg pre-TCRA to 0.1 mg/kg post-TCRA; whole body channel catfish from 39 mg/kg-L pre-TCRA to 2.6 mg/kg-L post-TCRA. Concentrations of PCBs in 2 species of resident fish (carp and white suckers), caged channel catfish, surface water, and sediment were reduced by over 1 order of magnitude within 1 year of completion, substantively accelerating natural recovery processes. A slight increase in PCB concentration was observed in both whole body suckers and adult carp fillets in the second monitoring period post-TCRA; however, these concentrations are still near an order of magnitude less than pre-TCRA concentrations and appear to be currently stable or on a slight downward trend. © 2014 SETAC.

Synergistic effects of BMP-2, BMP-6 or BMP-7 with human plasma fibronectin onto hydroxyapatite coatings: A comparative study.

PubMed

Brigaud, Isabelle; Agniel, Rémy; Leroy-Dudal, Johanne; Kellouche, Sabrina; Ponche, Arnaud; Bouceba, Tahar; Mihailescu, Natalia; Sopronyi, Mihai; Viguier, Eric; Ristoscu, Carmen; Sima, Felix; Mihailescu, Ion N; Carreira, Ana Claudia O; Sogayar, Mari Cleide; Gallet, Olivier; Anselme, Karine

2017-06-01

Design of new osteoinductive biomaterials to reproduce an optimized physiological environment capable of recruiting stem cells and instructing their fate towards the osteoblastic lineage has become a priority in orthopaedic surgery. This work aims at evaluating the bioactivity of BMP combined with human plasma fibronectin (FN/BMP) delivered in solution or coated onto titanium-hydroxyapatite (TiHA) surfaces. Herein, we focus on the comparison of in vitro osteogenic efficacy in mouse C2C12 pre-osteoblasts of three BMP members, namely: BMP-2, BMP-6 and BMP-7. In parallel, we evaluated the molecular binding strength between each BMP with FN using the Surface Plasmon Resonance (SPR) technology. The affinity of BMPs for FN was found totally different and dependent on BMP type. Indeed, the combination of FN with BMP-2 on TiHA surfaces potentiates the burst of gene-mediated osteogenic induction, while it prolongs the osteogenic activity of BMP-6 and surprisingly annihilates the BMP-7 one. These results correlate with FN/BMP affinity for TiHA, since BMP-6>BMP-2>BMP-7. In addition, by analyzing the osteogenic activity in the peri-implant environment, we showed that osteoinductive paracrine effects were significantly decreased upon (FN/BMP-6), as opposed to (FN/BMP-2) coatings. Altogether, our results support the use of FN/BMP-6 to develop a biomimetic microenvironment capable to induce osteogenic activity under physiological conditions, with minimum paracrine signalization. The originality of our paper relies on the first direct comparison of the in vitro osteogenic potential of three osteogenic BMPs (BMP-2, -6 and -7) combined with native human plasma fibronectin delivered in solution or coated by laser transfer onto titanium hydroxyapatite surfaces. We confirm that BMP association with fibronectin enhances the osteogenic activity of BMP-2, -6 and -7, but with essential discrepancies, depending on the BMP member, and in agreement with the affinity of BMPs for fibronectin. Moreover, we bring elements to explain the origin of the BMP-2 medical life-threatening side-effects by analyzing in vitro paracrine effects. Finally, this work supports the alternative use of FN/BMP-6 to induce osteogenic activity under physiological conditions, with minimum side effects. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Concentrations and Loads of Nutrients and Suspended Sediments in Englesby Brook and Little Otter Creek, Lake Champlain Basin, Vermont, 2000-2005

USGS Publications Warehouse

Medalie, Laura

2007-01-01

The effectiveness of best-management practices (BMPs) in improving water quality in Lake Champlain tributaries was evaluated from 2000 through 2005 on the basis of analysis of data collected on concentrations of total phosphorus and suspended sediment in Englesby Brook, an urban stream in Burlington, and Little Otter Creek, an agricultural stream in Ferrisburg. Data also were collected on concentrations of total nitrogen in the Englesby Brook watershed. In the winter of 2001-2002, one of three planned structural BMPs was installed in the urban watershed. At approximately the same time, a set of barnyard BMPs was installed in the agricultural watershed; however, the other planned BMPs, which included streambank fencing and nutrient management, were not implemented within the study period. At Englesby Brook, concentrations of phosphorus ranged from 0.024 to 0.3 milligrams per liter (mg/L) during base-flow and from 0.032 to 11.8 mg/L during high-flow conditions. Concentrations of suspended sediment ranged from 3 to 189 mg/L during base-flow and from 5 to 6,880 mg/L during high-flow conditions. An assessment of the effectiveness of an urban BMP was made by comparing concentrations and loads of phosphorus and suspended sediment before and after a golf-course irrigation pond in the Englesby Brook watershed was retrofitted with the objective of reducing sediment transport. Results from a modified paired watershed study design showed that the BMP reduced concentrations of phosphorus and suspended sediment during high-flow events - when average streamflow was greater than 3 cubic feet per second. While construction of the BMP did not reduce storm loads of phosphorus or suspended sediment, an evaluation of changes in slope of double-mass curves showing cumulative monthly streamflow plotted against cumulative monthly loads indicated a possible reduction in cumulative loads of phosphorus and suspended sediment after BMP construction. Results from the Little Otter Creek assessment of agricultural BMPs showed that concentrations of phosphorus ranged from 0.016 to 0.141 mg/L during base-flow and from 0.019 to 0.565 mg/L during high-flow conditions at the upstream monitoring station. Concentrations of suspended sediment ranged from 2 to 13 mg/L during base-flow and from 1 to 473 mg/L during high-flow conditions at the upstream monitoring station. Concentrations of phosphorus ranged from 0.018 to 0.233 mg/L during base-flow and from 0.019 to 1.95 mg/L during high-flow conditions at the downstream monitoring station. Concentrations of suspended sediment ranged from 10 to 132 mg/L during base-flow and from 8 to 1,190 mg/L during high-flow conditions at the downstream monitoring station. Annual loads of phosphorus at the downstream monitoring station were significantly larger than loads at the upstream monitoring station, and annual loads of suspended sediment at the downstream monitoring station were larger than loads at the upstream monitoring station for 4 out of 6 years. On a monthly basis, loads of phosphorus and suspended sediment at the downstream monitoring station were significantly larger than loads at the upstream monitoring station. Pairs of concentrations of phosphorus and monthly loads of phosphorus and suspended sediment from the upstream and downstream monitoring stations were evaluated using the paired watershed study design. The only significant reduction between the calibration and treatment periods was for monthly loads of phosphorus; all other evaluations showed no change between periods.

Bone Morphogenetic Protein 4 Promotes Vascular Smooth Muscle Contractility by Activating MicroRNA-21 (miR-21), which Down-regulates Expression of Family of Dedicator of Cytokinesis (DOCK) Proteins*

PubMed Central

Kang, Hara; Davis-Dusenbery, Brandi N.; Nguyen, Peter H.; Lal, Ashish; Lieberman, Judy; Van Aelst, Linda; Lagna, Giorgio; Hata, Akiko

2012-01-01

The bone morphogenetic protein 4 (BMP4) signaling pathway plays a critical role in the promotion and maintenance of the contractile phenotype in vascular smooth muscle cell (vSMC). Misexpression or inactivating mutations of the BMP receptor gene can lead to dedifferentiation of vSMC characterized by increased migration and proliferation that is linked to vascular proliferative disorders. Previously we demonstrated that vSMCs increase microRNA-21 (miR-21) biogenesis upon BMP4 treatment, which induces contractile gene expression by targeting programmed cell death 4 (PDCD4). To identify novel targets of miR-21 that are critical for induction of the contractile phenotype by BMP4, biotinylated miR-21 was expressed in vSMCs followed by an affinity purification of mRNAs associated with miR-21. Nearly all members of the dedicator of cytokinesis (DOCK) 180-related protein superfamily were identified as targets of miR-21. Down-regulation of DOCK4, -5, and -7 by miR-21 inhibited cell migration and promoted cytoskeletal organization by modulating an activity of small GTPase. Thus, this study uncovers a regulatory mechanism of the vSMC phenotype by the BMP4-miR-21 axis through DOCK family proteins. PMID:22158624

Statistics for stochastic modeling of volume reduction, hydrograph extension, and water-quality treatment by structural stormwater runoff best management practices (BMPs)

USGS Publications Warehouse

Granato, Gregory E.

2014-01-01

The U.S. Geological Survey (USGS) developed the Stochastic Empirical Loading and Dilution Model (SELDM) in cooperation with the Federal Highway Administration (FHWA) to indicate the risk for stormwater concentrations, flows, and loads to be above user-selected water-quality goals and the potential effectiveness of mitigation measures to reduce such risks. SELDM models the potential effect of mitigation measures by using Monte Carlo methods with statistics that approximate the net effects of structural and nonstructural best management practices (BMPs). In this report, structural BMPs are defined as the components of the drainage pathway between the source of runoff and a stormwater discharge location that affect the volume, timing, or quality of runoff. SELDM uses a simple stochastic statistical model of BMP performance to develop planning-level estimates of runoff-event characteristics. This statistical approach can be used to represent a single BMP or an assemblage of BMPs. The SELDM BMP-treatment module has provisions for stochastic modeling of three stormwater treatments: volume reduction, hydrograph extension, and water-quality treatment. In SELDM, these three treatment variables are modeled by using the trapezoidal distribution and the rank correlation with the associated highway-runoff variables. This report describes methods for calculating the trapezoidal-distribution statistics and rank correlation coefficients for stochastic modeling of volume reduction, hydrograph extension, and water-quality treatment by structural stormwater BMPs and provides the calculated values for these variables. This report also provides robust methods for estimating the minimum irreducible concentration (MIC), which is the lowest expected effluent concentration from a particular BMP site or a class of BMPs. These statistics are different from the statistics commonly used to characterize or compare BMPs. They are designed to provide a stochastic transfer function to approximate the quantity, duration, and quality of BMP effluent given the associated inflow values for a population of storm events. A database application and several spreadsheet tools are included in the digital media accompanying this report for further documentation of methods and for future use. In this study, analyses were done with data extracted from a modified copy of the January 2012 version of International Stormwater Best Management Practices Database, designated herein as the January 2012a version. Statistics for volume reduction, hydrograph extension, and water-quality treatment were developed with selected data. Sufficient data were available to estimate statistics for 5 to 10 BMP categories by using data from 40 to more than 165 monitoring sites. Water-quality treatment statistics were developed for 13 runoff-quality constituents commonly measured in highway and urban runoff studies including turbidity, sediment and solids; nutrients; total metals; organic carbon; and fecal coliforms. The medians of the best-fit statistics for each category were selected to construct generalized cumulative distribution functions for the three treatment variables. For volume reduction and hydrograph extension, interpretation of available data indicates that selection of a Spearman’s rho value that is the average of the median and maximum values for the BMP category may help generate realistic simulation results in SELDM. The median rho value may be selected to help generate realistic simulation results for water-quality treatment variables. MIC statistics were developed for 12 runoff-quality constituents commonly measured in highway and urban runoff studies by using data from 11 BMP categories and more than 167 monitoring sites. Four statistical techniques were applied for estimating MIC values with monitoring data from each site. These techniques produce a range of lower-bound estimates for each site. Four MIC estimators are proposed as alternatives for selecting a value from among the estimates from multiple sites. Correlation analysis indicates that the MIC estimates from multiple sites were weakly correlated with the geometric mean of inflow values, which indicates that there may be a qualitative or semiquantitative link between the inflow quality and the MIC. Correlations probably are weak because the MIC is influenced by the inflow water quality and the capability of each individual BMP site to reduce inflow concentrations.

Receptor-selective retinoids implicate retinoic acid receptor alpha and gamma in the regulation of bmp-2 and bmp-4 in F9 embryonal carcinoma cells.

PubMed

Rogers, M B

1996-01-01

The effect of retinoids on malignant cells and embryos indicates that retinoids influence the expression of growth factors or alter the response of cells to growth factors. The bone morphogenetic proteins, Bmp-2 and Bmp-4, are candidates for such growth factors because retinoic acid (RA) treatment of F9 embryonal carcinoma cells induced Bmp-2 mRNA, while simultaneously repressing Bmp-4 levels. Also, recombinant Bmp-2 affected the growth and differentiation of these cells. Regulation of each gene was concentration dependent and required continuous RA treatment. The short half-lives of the Bmp-2 (75 +/- 11 min) and Bmp-4 (70 +/- 4 min) mRNAs suggest that their abundance is primarily controlled at the transcriptional level. To determine which RA receptor (RAR) controls bmp-2 and bmp-4 expression, F9 cells were exposed to various receptor-selective retinoids. RAR alpha- and gamma-selective retinoids induced Bmp-2 and repressed Bmp-4 equally as well as all-trans RA. In contrast, a RAR beta-selective retinoid had little effect on Bmp-2 induction but repressed Bmp-4. A RAR alpha-selective antagonist inhibited all-trans RA stimulation of Bmp-2, although not as dramatically as a RAR beta gamma-selective antagonist. No differences were observed between Bmp levels in all-trans RA and 9-cis RA-treated cells, indicating that the RXRs play little part in controlling these genes. The results are consistent with RAR alpha and gamma-controlled Bmp-2 and Bmp-4 regulation.

Development of an efficient, non-viral transfection method for studying gene function and bone growth in human primary cranial suture mesenchymal cells reveals that the cells respond to BMP2 and BMP3.

PubMed

Dwivedi, Prem P; Anderson, Peter J; Powell, Barry C

2012-08-03

Achieving efficient introduction of plasmid DNA into primary cultures of mammalian cells is a common problem in biomedical research. Human primary cranial suture cells are derived from the connective mesenchymal tissue between the bone forming regions at the edges of the calvarial plates of the skull. Typically they are referred to as suture mesenchymal cells and are a heterogeneous population responsible for driving the rapid skull growth that occurs in utero and postnatally. To better understand the molecular mechanisms involved in skull growth, and in abnormal growth conditions, such as craniosynostosis, caused by premature bony fusion, it is essential to be able to easily introduce genes into primary bone forming cells to study their function. A comparison of several lipid-based techniques with two electroporation-based techniques demonstrated that the electroporation method known as nucleofection produced the best transfection efficiency. The parameters of nucleofection, including cell number, amount of DNA and nucleofection program, were optimized for transfection efficiency and cell survival. Two different genes and two promoter reporter vectors were used to validate the nucleofection method and the responses of human primary suture mesenchymal cells by fluorescence microscopy, RT-PCR and the dual luciferase assay. Quantification of bone morphogenetic protein (BMP) signalling using luciferase reporters demonstrated robust responses of the cells to both osteogenic BMP2 and to the anti-osteogenic BMP3. A nucleofection protocol has been developed that provides a simple and efficient, non-viral alternative method for in vitro studies of gene and protein function in human skull growth. Human primary suture mesenchymal cells exhibit robust responses to BMP2 and BMP3, and thus nucleofection can be a valuable method for studying the potential competing action of these two bone growth factors in a model system of cranial bone growth.

Effect of Escherichia coli-produced recombinant human bone morphogenetic protein 2 on the regeneration of canine segmental ulnar defects.

PubMed

Harada, Yasuji; Itoi, Takamasa; Wakitani, Shigeyuki; Irie, Hiroyuki; Sakamoto, Michiko; Zhao, Dongwei; Nezu, Yoshinori; Yogo, Takuya; Hara, Yasushi; Tagawa, Masahiro

2012-07-01

Because bone morphogenetic protein 2 gene transfected Escherichia coli (E-BMP-2) produce recombinant human BMP-2 (rhBMP-2) more efficiently than mammalian cells (Chinese hamster ovary [CHO]-BMP-2), they may be a more cost-effective source of rhBMP-2 for clinical use. However, use of E-BMP-2 for regenerating long bones in large animals has not been reported. In the current study, we evaluated the healing efficacy of E-BMP-2 in a canine model. We created 2.5-cm critical-size segmental ulnar defects in test animals, then implanted E-BMP-2 and 700 mg of artificial bone (beta-tricalcium phosphate; β-TCP) into the wounds. We examined the differential effects of 5 E-BMP-2 treatments (0, 35, 140, 560, and 2240 μg) across 5 experimental groups (control, BMP35, BMP140, BMP560, and BMP2240). Radiography and computed tomography were used to observe the regeneration process. The groups in which higher doses of E-BMP-2 were administered (BMP560 and BMP2240) displayed more pronounced bone regeneration; the regenerated tissues connected to the host bone, and the cross-sectional areas of the regenerated bone were larger than those of the originals. The groups in which lower doses of E-BMP-2 were administered (BMP35 and BMP140) experienced relatively less bone regeneration; furthermore, the regenerated tissues failed to connect to the host bone. In these groups, the cross-sectional areas of the regenerated bone were equal to or smaller than those of the originals. No regeneration was observed in the control group. These findings suggest that, like CHO-BMP-2, E-BMP-2 can be used for the regeneration of large defects in long bones and that its clinical use might decrease the cost of bone regeneration treatments.

BMP4 and BMP7 Suppress StAR and Progesterone Production via ALK3 and SMAD1/5/8-SMAD4 in Human Granulosa-Lutein Cells.

PubMed

Zhang, Han; Klausen, Christian; Zhu, Hua; Chang, Hsun-Ming; Leung, Peter C K

2015-11-01

Adequate production of progesterone by the corpus luteum is critical to the successful establishment of pregnancy. In animal models, bone morphogenetic protein (BMP) 4 and BMP7 have been shown to suppress either basal or gonadotropin-induced progesterone production, depending on the species examined. However, the effects of BMP4 and BMP7 on progesterone production in human granulosa cells are unknown. In the present study, we used immortalized (SVOG) and primary human granulosa-lutein cells to investigate the effects of BMP4 and BMP7 on steroidogenic acute regulatory protein (StAR) expression and progesterone production and to examine the underlying molecular mechanism. Treatment of primary and immortalized human granulosa cells with recombinant BMP4 or BMP7 decreased StAR expression and progesterone accumulation. In SVOG cells, the suppressive effects of BMP4 and BMP7 on StAR expression were blocked by pretreatment with inhibitors of activin receptor-like kinase (ALK)2/3/6 (dorsomorphin) or ALK2/3 (DMH1) but not ALK4/5/7 (SB-431542). Moreover, small interfering RNA-mediated depletion of ALK3, but not ALK2 or ALK6, reversed the effects of BMP4 and BMP7 on StAR expression. Likewise, BMP4- and BMP7-induced phosphorylation of SMAD 1/5/8 was reversed by treatment with DMH1 or small interfering RNA targeting ALK3. Knockdown of SMAD4, the essential common SMAD for BMP/TGF-β signaling, abolished the effects of BMP4 and BMP7 on StAR expression. Our results suggest that BMP4 and BMP7 down-regulate StAR and progesterone production via ALK3 and SMAD1/5/8-SMAD4 signaling in human granulosa-lutein cells.

Dual-source dual-power electrospinning and characteristics of multifunctional scaffolds for bone tissue engineering.

PubMed

Wang, Chong; Wang, Min

2012-10-01

Electrospun tissue engineering scaffolds are attractive due to their distinctive advantages over other types of scaffolds. As both osteoinductivity and osteoconductivity play crucial roles in bone tissue engineering, scaffolds possessing both properties are desirable. In this investigation, novel bicomponent scaffolds were constructed via dual-source dual-power electrospinning (DSDPES). One scaffold component was emulsion electrospun poly(D,L-lactic acid) (PDLLA) nanofibers containing recombinant human bone morphogenetic protein (rhBMP-2), and the other scaffold component was electrospun calcium phosphate (Ca-P) particle/poly(lactic-co-glycolic acid) (PLGA) nanocomposite fibers. The mass ratio of rhBMP-2/PDLLA fibers to Ca-P/PLGA fibers in bicomponent scaffolds could be controlled in the DSDPES process by adjusting the number of syringes used to supply solutions for electrospinning. Through process optimization, both types of fibers could be evenly distributed in bicomponent scaffolds. The structure and properties of each type of fibers in the scaffolds were studied. The morphological and structural properties and wettability of scaffolds were assessed. The effects of emulsion composition for rhBMP-2/PDLLA fibers and mass ratio of fibrous components in bicomponent scaffolds on in vitro release of rhBMP-2 from scaffolds were investigated. In vitro degradation of scaffolds was also studied by monitoring their morphological changes, weight losses and decreases in average molecular weight of fiber matrix polymers.

Zygotic LvBMP5-8 is required for skeletal patterning and for left-right but not dorsal-ventral specification in the sea urchin embryo.

PubMed

Piacentino, Michael L; Chung, Oliver; Ramachandran, Janani; Zuch, Daniel T; Yu, Jia; Conaway, Evan A; Reyna, Arlene E; Bradham, Cynthia A

2016-04-01

Skeletal patterning in the sea urchin embryo requires coordinated signaling between the pattern-dictating ectoderm and the skeletogenic primary mesenchyme cells (PMCs); recent studies have begun to uncover the molecular basis for this process. Using an unbiased RNA-Seq-based screen, we have previously identified the TGF-ß superfamily ligand, LvBMP5-8, as a skeletal patterning gene in Lytechinus variegatus embryos. This result is surprising, since both BMP5-8 and BMP2/4 ligands have been implicated in sea urchin dorsal-ventral (DV) and left-right (LR) axis specification. Here, we demonstrate that zygotic LvBMP5-8 is required for normal skeletal patterning on the left side, as well as for normal PMC positioning during gastrulation. Zygotic LvBMP5-8 is required for expression of the left-side marker soxE, suggesting that LvBMP5-8 is required for left-side specification. Interestingly, we also find that LvBMP5-8 knockdown suppresses serotonergic neurogenesis on the left side. While LvBMP5-8 overexpression is sufficient to dorsalize embryos, we find that zygotic LvBMP5-8 is not required for normal DV specification or development. In addition, ectopic LvBMP5-8 does not dorsalize LvBMP2/4 morphant embryos, indicating that, in the absence of BMP2/4, BMP5-8 is insufficient to specify dorsal. Taken together, our data demonstrate that zygotic LvBMP5-8 signaling is essential for left-side specification, and for normal left-side skeletal and neural patterning, but not for DV specification. Thus, while both BMP2/4 and BMP5-8 regulate LR axis specification, BMP2/4 but not zygotic BMP5-8 regulates DV axis specification in sea urchin embryos. Copyright © 2016 Elsevier Inc. All rights reserved.

Nanostructured hydroxyapatite surfaces-mediated adsorption alters recognition of BMP receptor IA and bioactivity of bone morphogenetic protein-2.

PubMed

Huang, Baolin; Yuan, Yuan; Ding, Sai; Li, Jianbo; Ren, Jie; Feng, Bo; Li, Tong; Gu, Yuantong; Liu, Changsheng

2015-11-01

Highly efficient loading of bone morphogenetic protein-2 (BMP-2) onto carriers with desirable performance is still a major challenge in the field of bone regeneration. Till now, the nanoscaled surface-induced changes of the structure and bioactivity of BMP-2 remains poorly understood. Here, the effect of nanoscaled surface on the adsorption and bioactivity of BMP-2 was investigated with a series of hydroxyapatite surfaces (HAPs): HAP crystal-coated surface (HAP), HAP crystal-coated polished surface (HAP-Pol), and sintered HAP crystal-coated surface (HAP-Sin). The adsorption dynamics of recombinant human BMP-2 (rhBMP-2) and the accessibility of the binding epitopes of adsorbed rhBMP-2 for BMP receptors (BMPRs) were examined by a quartz crystal microbalance with dissipation. Moreover, the bioactivity of adsorbed rhBMP-2 and the BMP-induced Smad signaling were investigated with C2C12 model cells. A noticeably high mass-uptake of rhBMP-2 and enhanced recognition of BMPR-IA to adsorbed rhBMP-2 were found on the HAP-Pol surface. For the rhBMP-2-adsorbed HAPs, both ALP activity and Smad signaling increased in the order of HAP-Sin

Radiographic comparison of different concentrations of recombinant human bone morphogenetic protein with allogenic bone compared with the use of 100% mineralized cancellous bone allograft in maxillary sinus grafting.

PubMed

Froum, Stuart J; Wallace, Stephen; Cho, Sang-Choon; Khouly, Ismael; Rosenberg, Edwin; Corby, Patricia; Froum, Scott; Mascarenhas, Patrick; Tarnow, Dennis P

2014-01-01

The purpose of this study was to radiographically evaluate, then analyze, bone height, volume, and density with reference to percentage of vital bone after maxillary sinuses were grafted using two different doses of recombinant human bone morphogenetic protein 2/acellular collagen sponge (rhBMP-2/ACS) combined with mineralized cancellous bone allograft (MCBA) and a control sinus grafted with MCBA only. A total of 18 patients (36 sinuses) were used for analysis of height and volume measurements, having two of three graft combinations (one in each sinus): (1) control, MCBA only; (2) test 1, MCBA + 5.6 mL of rhBMP-2/ACS (containing 8.4 mg of rhBMP-2); and (3) test 2, MCBA + 2.8 mL of rhBMP-2/ACS (containing 4.2 mg of rhBMP-2). The study was completed with 16 patients who also had bilateral cores removed 6 to 9 months following sinus augmentation. A computer software system was used to evaluate 36 computed tomography scans. Two time points where selected for measurements of height: The results indicated that height of the grafted sinus was significantly greater in the treatment groups compared with the control. However, by the second time point, there were no statistically significant differences. Three weeks post-surgery bone volume measurements showed similar statistically significant differences between test and controls. However, prior to core removal, test group 1 with the greater dose of rhBMP-2 showed a statistically significant greater increase compared with test group 2 and the control. There was no statistically significant difference between the latter two groups. All three groups had similar volume and shrinkage. Density measurements varied from the above results, with the control showing statistically significant greater density at both time points. By contrast, the density increase over time in both rhBMP groups was similar and statistically higher than in the control group. There were strong associations between height and volume in all groups and between volume and new vital bone only in the control group. There were no statistically significant relationships observed between height and bone density or between volume and bone density for any parameter measured. More cases and monitoring of the future survival of implants placed in these augmented sinuses are needed to verify these results.

Dose reduction of bone morphogenetic protein-2 for bone regeneration using a delivery system based on lyophilization with trehalose.

PubMed

Zhang, Xiaochen; Yu, Quan; Wang, Yan-An; Zhao, Jun

2018-01-01

To induce sufficient new bone formation, high doses of bone morphogenetic protein-2 (BMP-2) are applied in regenerative medicine that often induce serious side effects. Therefore, improved treatment strategies are required. Here, we investigate whether the delivery of BMP-2 lyophilized in the presence of trehalose reduced the dose of BMP-2 required for bone regeneration. A new growth factor delivery system was fabricated using BMP-2-loaded TiO 2 nanotubes by lyophilization with trehalose (TiO 2 -Lyo-Tre-BMP-2). We measured BMP-2 release characteristics, bioactivity, and stability, and determined the effects on the osteogenic differentiation of bone marrow stromal cells in vitro. Additionally, we evaluated the ability of this formulation to regenerate new bone around implants in rat femur defects by micro-computed tomography (micro-CT), sequential fluorescent labelling, and histological analysis. Compared with absorbed BMP-2-loaded TiO 2 nanotubes (TiO 2 -BMP-2), TiO 2 -Lyo-Tre-BMP-2 exhibited sustained release, consistent bioactivity, and higher stability of BMP-2, and resulted in greater osteogenic differentiation of BMSCs. Eight weeks post-operation, TiO 2 -Lyo-Tre-BMP-2 nanotubes, with various dosages of BMP-2, regenerated larger amounts of new bone than TiO 2 -BMP-2 nanotubes. Our findings indicate that delivery of BMP-2 lyophilized with trehalose may be a promising method to reduce the dose of BMP-2 and avoid the associated side effects.

Periosteal BMP2 activity drives bone graft healing.

PubMed

Chappuis, Vivianne; Gamer, Laura; Cox, Karen; Lowery, Jonathan W; Bosshardt, Dieter D; Rosen, Vicki

2012-10-01

Bone graft incorporation depends on the orchestrated activation of numerous growth factors and cytokines in both the host and the graft. Prominent in this signaling cascade is BMP2. Although BMP2 is dispensable for bone formation, it is required for the initiation of bone repair; thus understanding the cellular mechanisms underlying bone regeneration driven by BMP2 is essential for improving bone graft therapies. In the present study, we assessed the role of Bmp2 in bone graft incorporation using mice in which Bmp2 has been removed from the limb prior to skeletal formation (Bmp2(cKO)). When autograft transplantations were performed in Bmp2cKO mice, callus formation and bone healing were absent. Transplantation of either a vital wild type (WT) bone graft into a Bmp2(cKO) host or a vital Bmp2(cKO) graft into a WT host also resulted in the inhibition of bone graft incorporation. Histological analyses of these transplants show that in the absence of BMP2, periosteal progenitors remain quiescent and healing is not initiated. When we analyzed the expression of Sox9, a marker of chondrogenesis, on the graft surface, we found it significantly reduced when BMP2 was absent in either the graft itself or the host, suggesting that local BMP2 levels drive periosteal cell condensation and subsequent callus cell differentiation. The lack of integrated healing in the absence of BMP2 was not due to the inability of periosteal cells to respond to BMP2. Healing was achieved when grafts were pre-soaked in rhBMP2 protein, indicating that periosteal progenitors remain responsive in the absence of BMP2. In contrast to the requirement for BMP2 in periosteal progenitor activation in vital bone grafts, we found that bone matrix-derived BMP2 does not significantly enhance bone graft incorporation. Taken together, our data show that BMP2 signaling is not essential for the maintenance of periosteal progenitors, but is required for the activation of these progenitors and their subsequent differentiation along the osteo-chondrogenic pathway. These results indicate that BMP2 will be among the signaling molecules whose presence will determine success or failure of new bone graft strategies. Copyright © 2012 Elsevier Inc. All rights reserved.

Neural crest apoptosis and the establishment of craniofacial pattern: an honorable death.

PubMed

Graham, A; Koentges, G; Lumsden, A

1996-01-01

During development of the vertebrate head neural crest cells emigrate from the hindbrain and populate the branchial arches, giving rise to distinct skeletal elements and muscle connective tissues in each arch. The production of neural crest from the hindbrain is discontinuous and crest cells destined for different arches, carrying different positional cues, are separated by regions of apoptosis centered on rhombomeres (r) 3 and r5. This cell death program is under the interactive control of the neighboring hindbrain segments. Both r3 and r5 produce large numbers of crest cells when freed from their flanking rhombomere, but when conjoined with their neighbor the cell death program is restored. Two key components of this program are Bmp 4 and msx-2, both of which are expressed in the apoptotic foci of r3 and r5 and which are also regulated by neighbor interactions. Importantly, the addition of recombinant Bmp 4 to isolated cultures of r3 and r5 induces the expression of Bmp 4 and msx-2 and restores the cell death program. This early neural crest segregation is maintained during development and it has profound effects upon the final craniofacial pattern. Even though crest cells from different axial origins will contribute to compound skeletal elements, these distinct populations do not intermingle. Furthermore head muscle connective tissues are exclusively anchored to skeletal domains arising from neural crest from the same axial level. Thus the discontinuous production of neural crest sculpts the crest into nonmixing streams and consequently ensures the fidelity of patterning.

Systems biology derived source-sink mechanism of BMP gradient formation

PubMed Central

Zinski, Joseph; Bu, Ye; Wang, Xu; Dou, Wei

2017-01-01

A morphogen gradient of Bone Morphogenetic Protein (BMP) signaling patterns the dorsoventral embryonic axis of vertebrates and invertebrates. The prevailing view in vertebrates for BMP gradient formation is through a counter-gradient of BMP antagonists, often along with ligand shuttling to generate peak signaling levels. To delineate the mechanism in zebrafish, we precisely quantified the BMP activity gradient in wild-type and mutant embryos and combined these data with a mathematical model-based computational screen to test hypotheses for gradient formation. Our analysis ruled out a BMP shuttling mechanism and a bmp transcriptionally-informed gradient mechanism. Surprisingly, rather than supporting a counter-gradient mechanism, our analyses support a fourth model, a source-sink mechanism, which relies on a restricted BMP antagonist distribution acting as a sink that drives BMP flux dorsally and gradient formation. We measured Bmp2 diffusion and found that it supports the source-sink model, suggesting a new mechanism to shape BMP gradients during development. PMID:28826472

Systems biology derived source-sink mechanism of BMP gradient formation.

PubMed

Zinski, Joseph; Bu, Ye; Wang, Xu; Dou, Wei; Umulis, David; Mullins, Mary C

2017-08-09

A morphogen gradient of Bone Morphogenetic Protein (BMP) signaling patterns the dorsoventral embryonic axis of vertebrates and invertebrates. The prevailing view in vertebrates for BMP gradient formation is through a counter-gradient of BMP antagonists, often along with ligand shuttling to generate peak signaling levels. To delineate the mechanism in zebrafish, we precisely quantified the BMP activity gradient in wild-type and mutant embryos and combined these data with a mathematical model-based computational screen to test hypotheses for gradient formation. Our analysis ruled out a BMP shuttling mechanism and a bmp transcriptionally-informed gradient mechanism. Surprisingly, rather than supporting a counter-gradient mechanism, our analyses support a fourth model, a source-sink mechanism, which relies on a restricted BMP antagonist distribution acting as a sink that drives BMP flux dorsally and gradient formation. We measured Bmp2 diffusion and found that it supports the source-sink model, suggesting a new mechanism to shape BMP gradients during development.

Bmp 2 and bmp 7 induce odonto- and osteogenesis of human tooth germ stem cells.

PubMed

Taşlı, P Neslihan; Aydın, Safa; Yalvaç, Mehmet Emir; Sahin, Fikrettin

2014-03-01

Bone morphogenetic proteins (BMPs) initiate, promote, and maintain odontogenesis and osteogenesis. In this study, we studied the effect of bone morphogenic protein 2 (BMP 2) and bone morphogenic protein 7 (BMP 7) as differentiation inducers in tooth and bone regeneration. We compared the effect of BMP 2 and BMP 7 on odontogenic and osteogenic differentiation of human tooth germ stem cells (hTGSCs). Third molar-derived hTGSCs were characterized with mesenchymal stem cell surface markers by flow cytometry. BMP 2 and BMP 7 were transfected into hTGSCs and the cells were seeded onto six-well plates. One day after the transfection, hTGSCs were treated with odontogenic and osteogenic mediums for 14 days. For confirmation of odontogenic and osteogenic differentiation, mRNA levels of BMP2, BMP 7, collagen type 1 (COL1A), osteocalsin (OCN), and dentin sialophosphoprotein (DSPP) genes were measured by quantitative real-time PCR. In addition to this, immunocytochemistry was performed by odontogenic and osteogenic antibodies and mineralization obtained by von Kossa staining. Our results showed that the BMP 2 and BMP 7 both promoted odontogenic and osteogenic differentiation of hTGSCs. Data indicated that BMP 2 treatment and BMP 7 treatment induce odontogenic differentiation without affecting each other, whereas they induce osteogenic differentiation by triggering expression of each other. These findings provide a feasible tool for tooth and bone tissue engineering.

TNF-α Upregulates Expression of BMP-2 and BMP-3 Genes in the Rat Dental Follicle – Implications for Tooth Eruption

PubMed Central

Yao, Shaomian; Prpic, Veronica; Pan, Fenghui; Wise, Gary E.

2011-01-01

The dental follicle appears to regulate both the alveolar bone resorption and bone formation needed for tooth eruption. Tumor necrosis factor-alpha ( TNF-α) gene expression is maximally upregulated at postnatal day 9 in the rat dental follicle of the 1st mandibular molar, a time that correlates with rapid bone growth at the base of the tooth crypt, as well as a minor burst of osteoclastogenesis. TNF-α expression is correlated with the expression of bone morphogenetic protein-2 (BMP-2), a molecule expressed in the dental follicle that can promote bone formation. Because BMP-2 signaling may be augmented by bone morphogenetic protein-3 (BMP-3), it was the objective of this study to determine 1) if the dental follicle expresses BMP-3 and 2) if TNF-α stimulates the dental follicle cells to express BMP-2 and BMP-3. Dental follicles were collected from different postnatal ages of rat pups. Dental follicle cells were incubated with TNF-α to study its dosage and time-course effects on gene expression of BMP-2 and BMP-3, as determined by real-time RT-PCR. Next, immunostaining was conducted to confirm if the protein was synthesized and ELISA of the conditioned medium was conducted to determine if BMP-2 was secreted. We found that BMP-3 expression is correlated with the expression of TNF-α in the dental follicle and TNF-α significantly increased BMP-2 and BMP-3 expression in vitro. Immunostaining and ELISA showed that BMP-2 and BMP-3 were synthesized and secreted. This study suggests that TNF-α can upregulate the expression of bone formation genes that may be needed for tooth eruption. PMID:20067418

BMP type I receptor ALK2 is required for angiotensin II-induced cardiac hypertrophy

PubMed Central

Spagnolli, Ester; Ernande, Laura; Thoonen, Robrecht; Kolodziej, Starsha A.; Leyton, Patricio A.; Cheng, Juan; Tainsh, Robert E. T.; Mayeur, Claire; Rhee, David K.; Wu, Mei. X.; Scherrer-Crosbie, Marielle; Buys, Emmanuel S.; Zapol, Warren M.; Bloch, Kenneth D.; Bloch, Donald B.

2016-01-01

Bone morphogenetic protein (BMP) signaling contributes to the development of cardiac hypertrophy. However, the identity of the BMP type I receptor involved in cardiac hypertrophy and the underlying molecular mechanisms are poorly understood. By using quantitative PCR and immunoblotting, we demonstrated that BMP signaling increased during phenylephrine-induced hypertrophy in cultured neonatal rat cardiomyocytes (NRCs), as evidenced by increased phosphorylation of Smads 1 and 5 and induction of Id1 gene expression. Inhibition of BMP signaling with LDN193189 or noggin, and silencing of Smad 1 or 4 using small interfering RNA diminished the ability of phenylephrine to induce hypertrophy in NRCs. Conversely, activation of BMP signaling with BMP2 or BMP4 induced hypertrophy in NRCs. Luciferase reporter assay further showed that BMP2 or BMP4 treatment of NRCs repressed atrogin-1 gene expression concomitant with an increase in calcineurin protein levels and enhanced activity of nuclear factor of activated T cells, providing a mechanism by which BMP signaling contributes to cardiac hypertrophy. In a model of cardiac hypertrophy, C57BL/6 mice treated with angiotensin II (A2) had increased BMP signaling in the left ventricle. Treatment with LDN193189 attenuated A2-induced cardiac hypertrophy and collagen deposition in left ventricles. Cardiomyocyte-specific deletion of BMP type I receptor ALK2 (activin-like kinase 2), but not ALK1 or ALK3, inhibited BMP signaling and mitigated A2-induced cardiac hypertrophy and left ventricular fibrosis in mice. The results suggest that BMP signaling upregulates the calcineurin/nuclear factor of activated T cell pathway via BMP type I receptor ALK2, contributing to cardiac hypertrophy and fibrosis. PMID:26873969

Bone Morphogenetic Proteins, Antagonists and Receptors in Prostate Cancer

DTIC Science & Technology

2005-01-01

expressed in prostate. This work investigates BMP receptors and BMP antagonists to understand the basic mechanisms to inhibit the BMP signaling in...during embryoge- nesis, and prostate cancer metastases to bone. BMP functions can be inhibited by antagonists such as Noggin or DAN. DAN is a protein...protein along with a constant 0-6 -1 10 100 1000 1O0ng/ml of BMP-6, we were able to show a ng/ml BMP-6 dose-dependent inhibition of BMP-6 activity in DU

Evaluation of nonpoint-source contamination, Wisconsin: Land-use and Best-Management-Practices inventory, selected streamwater-quality data, urban-watershed quality assurance and quality control, constituent loads in rural streams, and snowmelt-runoff analysis, water year 1994

USGS Publications Warehouse

Walker, J.F.; Graczyk, D.J.; Corsi, S.R.; Owens, D.W.; Wierl, J.A.

1995-01-01

The objective of the watershed-management evaluation monitoring program in Wisconsin is to evaluate the effectiveness of best-management practices (BMP) for controlling nonpoint-source contamination in rural and urban watersheds. This report is an annual summary of the data collected for the program by the U.S Geological Survey and a report of the results of several different detailed analyses of the data. A land-use and BMP inventory is ongoing for 12 evaluation monitoring projects to track the sources of nonpoint-source pollution in each watershed and to document implementation of BMP's that may cause changes in the water quality of streams. Updated information is gathered each year, mapped, and stored in a geographic-information-system data base. Summaries of data collected during water years 1989-94 are presented. A water year is the period beginning October 1 and ending September 30; the water year is designated by the calendar year in which it ends. Suspended-sediment and total-phosphorus data (storm loads and annual loads) are summarized for eight rural sites. For all sites, the annual suspended-sediment or suspended-solids load for water year 1993 exceeded the average for the period of data collection; the minimum annual loads were transported in water year 1991 or 1992. Continuous dissolved-oxygen data were collected at seven rural sites during water year 1994. Data for water years 1990-93 are summarized and plotted in terms of percentage of time that a particular concentration is equaled or exceeded. Dissolved-oxygen concentrations in four streams were less than 9 mg/L at least 50 percent of the time, a condition that fails to meet suggested criterion for coldwater streams. The dissolved-oxygen probability curve for one of the coldwater streams is markedly different than the curves for the other streams, perhaps because of differences in aquatic biomass. Blank quality-assurance samples were collected at two of the urban evaluation monitoring sites to isolate contamination in the sample bottle, the automatic sampler and splitter, and the filtration system. Significant contamination caused excessive concentrations of dissolved chloride, alkalinity, and biochemical oxygen demand. The level of contamination may be large enough to affect data for water samples in which these analytes are present at low concentration. Further investigation is being done to determine the source of contamination and take measures to minimize its effect on the sampling. A preliminary regression analysis was done for the rural sites using data collected during water years 1989-93. Loads of suspended solids and total phosphorus in stormflow were regressed against various precipitation-related measures. The results indicate that, for most sites, changes in constituent load on the order of 40 to 50 percent could be detected with a statistical test. For two sites, the change would have to be 60 to 70 percent to be detected. A detailed comparison of snowmelt runoff and rainfall stormflow in urban and rural areas was done using data collected during water years 1985-93. For the rural sites where statistically significant differences were found between constituent loads in snowmelt and storm runoff, the loads of suspended solids and total phosphorus in snowmelt runoff were greater than those in storm runoff. For the urban sites where statistically significant differences were found between snowmelt and storm runoff, the loads of suspended solids and total phosphorus in storm runoff were greater than those in snowmelt runoff. The importance of including snowmelt runoff in designing and analyzing the effects of BMP's on streamwater quality, particularly in rural areas, is emphasized by these results.

Establishment of Immortalized Mouse Bmp2 Knock-Out Dental Papilla Mesenchymal Cells Necessary for Study of Odontoblastic Differentiation and Odontogenesis.

PubMed

Wu, Lian; Wang, Feng; Donly, Kevin J; Wan, Chunyan; Luo, Daoshu; Harris, Stephen E; MacDougall, Mary; Chen, Shuo

2015-11-01

Bmp2 is essential for dentin formation. Bmp2 cKO mice exhibited similar phenotype to dentinogenesis imperfecta, showing dental pulp exposure, hypomineralized dentin, and delayed odontoblast differentiation. As it is relatively difficult to obtain lot of primary Bmp2 cKO dental papilla mesenchymal cells and to maintain a long-term culture of these primary cells, availability of immortalized deleted Bmp2 dental papilla mesenchymal cells is critical for studying the underlying mechanism of Bmp2 signal in odontogenesis. In this study, our goal was to generate an immortalized deleted Bmp2 dental papilla mesenchymal (iBmp2(ko/ko)dp) cell line by introducing Cre recombinase and green fluorescent protein (GFP) into the immortalized mouse floxed Bmp2 dental papilla mesenchymal (iBmp2(fx/fx)dp) cells. iBmp2(ko/ko)dp cells were confirmed by GFP and PCR. The deleted Bmp2 cells exhibited slow cell proliferation rate and cell growth was arrested in G2 phase. Expression of tooth-related marker genes and cell differentiation were decreased in the deleted cells. Importantly, extracellular matrix remodeling was impaired in the iBmp2(ko/ko)dp cells as reflected by the decreased Mmp-9 expression. In addition, with exogenous Bmp2 induction, these cell differentiation and mineralization were rescued as well as extracellular matrix remodeling was enhanced. Therefore, we for the first time described establishment of iBmp(ko/ko) cells that are useful for study of mechanisms in regulating dental papilla mesenchymal cell lineages. © 2015 Wiley Periodicals, Inc.

Biological activity of a genetically modified BMP-2 variant with inhibitory activity

PubMed Central

Klammert, Uwe; Nickel, Joachim; Würzler, Kristian; Klingelhöffer, Christoph; Sebald, Walter; Kübler, Alexander C; Reuther, Tobias

2009-01-01

Background Alterations of the binding epitopes of bone morphogenetic protein-2 (BMP-2) lead to a modified interaction with the ectodomains of BMP receptors. In the present study the biological effect of a BMP-2 double mutant with antagonistic activity was evaluated in vivo. Methods Equine-derived collagenous carriers were loaded with recombinant human BMP-2 (rhBMP-2) in a well-known dose to provide an osteoinductive stimulus. The study was performed in a split animal design: carriers only coupled with rhBMP-2 (control) were implanted into prepared cavities of lower limb muscle of rats, specimens coupled with rhBMP-2 as well as BMP-2 double mutant were placed into the opposite limb in the same way. After 28 days the carriers were explanted, measured radiographically and characterized histologically. Results As expected, the BMP-2 loaded implants showed a typical heterotopic bone formation. The specimens coupled with both proteins showed a significant decreased bone formation in a dose dependent manner. Conclusion The antagonistic effect of a specific BMP-2 double mutant could be demonstrated in vivo. The dose dependent influence on heterotopic bone formation by preventing rhBMP-2 induced osteoinduction suggests a competitive receptor antagonism. PMID:19187528

DMH1, a Highly Selective Small Molecule BMP Inhibitor Promotes Neurogenesis of hiPSCs: Comparison of PAX6 and SOX1 Expression during Neural Induction

PubMed Central

2012-01-01

Recent successes in deriving human-induced pluripotent stem cells (hiPSCs) allow for the possibility of studying human neurons derived from patients with neurological diseases. Concomitant inhibition of the BMP and TGF-β1 branches of the TGF-β signaling pathways by the endogenous antagonist, Noggin, and the small molecule SB431542, respectively, induces efficient neuralization of hiPSCs, a method known as dual-SMAD inhibition. The use of small molecule inhibitors instead of their endogenous counterparts has several advantages including lower cost, consistent activity, and the maintenance of xeno-free culture conditions. We tested the efficacy of DMH1, a highly selective small molecule BMP-inhibitor for its potential to replace Noggin in the neuralization of hiPSCs. We compare Noggin and DMH1-induced neuralization of hiPSCs by measuring protein and mRNA levels of pluripotency and neural precursor markers over a period of seven days. The regulation of five of the six markers assessed was indistinguishable in the presence of concentrations of Noggin or DMH1 that have been shown to effectively inhibit BMP signaling in other systems. We observed that by varying the DMH1 or Noggin concentration, we could selectively modulate the number of SOX1 expressing cells, whereas PAX6, another neural precursor marker, remained the same. The level and timing of SOX1 expression have been shown to affect neural induction as well as neural lineage. Our observations, therefore, suggest that BMP-inhibitor concentrations need to be carefully monitored to ensure appropriate expression levels of all transcription factors necessary for the induction of a particular neuronal lineage. We further demonstrate that DMH1-induced neural progenitors can be differentiated into β3-tubulin expressing neurons, a subset of which also express tyrosine hydroxylase. Thus, the combined use of DMH1, a highly specific BMP-pathway inhibitor, and SB431542, a TGF-β1-pathway specific inhibitor, provides us with the tools to independently regulate these two pathways through the exclusive use of small molecule inhibitors. PMID:22860217

Signaling via the transcriptionally regulated activin receptor 2B is a novel mediator of neuronal cell death during chicken ciliary ganglion development.

PubMed

Koszinowski, S; Buss, K; Kaehlcke, K; Krieglstein, K

2015-04-01

The TGF-β ligand superfamily members activin A and BMP control important aspects of embryonic neuronal development and differentiation. Both are known to bind to activin receptor subtypes IIA (ActRIIA) and IIB, while in the avian ciliary ganglion (CG), so far only ActRIIA-expression has been described. We show that the expression of ACVR2B, coding for the ActRIIB, is tightly regulated during CG development and the knockdown of ACVR2B expression leads to a deregulation in the execution of neuronal apoptosis and therefore affects ontogenetic programmed cell death in vivo. While the differentiation of choroid neurons was impeded in the knockdown, pointing toward a reduction in activin A-mediated neural differentiation signaling, naturally occurring neuronal cell death in the CG was not prevented by follistatin treatment. Systemic injections of the BMP antagonist noggin, on the other hand, reduced the number of apoptotic neurons to a similar extent as ACVR2B knockdown. We therefore propose a novel pathway in the regulation of CG neuron ontogenetic programmed cell death, which could be mediated by BMP and signals via the ActRIIB. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

Bone morphogenetic protein 9 (BMP9) and BMP10 enhance tumor necrosis factor-α-induced monocyte recruitment to the vascular endothelium mainly via activin receptor-like kinase 2.

PubMed

Mitrofan, Claudia-Gabriela; Appleby, Sarah L; Nash, Gerard B; Mallat, Ziad; Chilvers, Edwin R; Upton, Paul D; Morrell, Nicholas W

2017-08-18

Bone morphogenetic proteins 9 and 10 (BMP9/BMP10) are circulating cytokines with important roles in endothelial homeostasis. The aim of this study was to investigate the roles of BMP9 and BMP10 in mediating monocyte-endothelial interactions using an in vitro flow adhesion assay. Herein, we report that whereas BMP9/BMP10 alone had no effect on monocyte recruitment, at higher concentrations both cytokines synergized with tumor necrosis factor-α (TNFα) to increase recruitment to the vascular endothelium. The BMP9/BMP10-mediated increase in monocyte recruitment in the presence of TNFα was associated with up-regulated expression levels of E-selectin, vascular cell adhesion molecule (VCAM-1), and intercellular adhesion molecule 1 (ICAM-1) on endothelial cells. Using siRNAs to type I and II BMP receptors and the signaling intermediaries (Smads), we demonstrated a key role for ALK2 in the BMP9/BMP10-induced surface expression of E-selectin, and both ALK1 and ALK2 in the up-regulation of VCAM-1 and ICAM-1. The type II receptors, BMPR-II and ACTR-IIA were both required for this response, as was Smad1/5. The up-regulation of cell surface adhesion molecules by BMP9/10 in the presence of TNFα was inhibited by LDN193189, which inhibits ALK2 but not ALK1. Furthermore, LDN193189 inhibited monocyte recruitment induced by TNFα and BMP9/10. BMP9/10 increased basal IκBα protein expression, but did not alter p65/RelA levels. Our findings suggest that higher concentrations of BMP9/BMP10 synergize with TNFα to induce the up-regulation of endothelial selectins and adhesion molecules, ultimately resulting in increased monocyte recruitment to the vascular endothelium. This process is mediated mainly via the ALK2 type I receptor, BMPR-II/ACTR-IIA type II receptors, and downstream Smad1/5 signaling. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Comparison on batch anaerobic digestion of five different livestock manures and prediction of biochemical methane potential (BMP) using different statistical models.

PubMed

Kafle, Gopi Krishna; Chen, Lide

2016-02-01

There is a lack of literature reporting the methane potential of several livestock manures under the same anaerobic digestion conditions (same inoculum, temperature, time, and size of the digester). To the best of our knowledge, no previous study has reported biochemical methane potential (BMP) predicting models developed and evaluated by solely using at least five different livestock manure tests results. The goal of this study was to evaluate the BMP of five different livestock manures (dairy manure (DM), horse manure (HM), goat manure (GM), chicken manure (CM) and swine manure (SM)) and to predict the BMP using different statistical models. Nutrients of the digested different manures were also monitored. The BMP tests were conducted under mesophilic temperatures with a manure loading factor of 3.5g volatile solids (VS)/L and a feed to inoculum ratio (F/I) of 0.5. Single variable and multiple variable regression models were developed using manure total carbohydrate (TC), crude protein (CP), total fat (TF), lignin (LIG) and acid detergent fiber (ADF), and measured BMP data. Three different kinetic models (first order kinetic model, modified Gompertz model and Chen and Hashimoto model) were evaluated for BMP predictions. The BMPs of DM, HM, GM, CM and SM were measured to be 204, 155, 159, 259, and 323mL/g VS, respectively and the VS removals were calculated to be 58.6%, 52.9%, 46.4%, 81.4%, 81.4%, respectively. The technical digestion time (T80-90, time required to produce 80-90% of total biogas production) for DM, HM, GM, CM and SM was calculated to be in the ranges of 19-28, 27-37, 31-44, 13-18, 12-17days, respectively. The effluents from the HM showed the lowest nitrogen, phosphorus and potassium concentrations. The effluents from the CM digesters showed highest nitrogen and phosphorus concentrations and digested SM showed highest potassium concentration. Based on the results of the regression analysis, the model using the variable of LIG showed the best (R(2)=0.851, p=0.026) for BMP prediction among the single variable models, and the model including variables of TC and TF showed the best prediction for BMPs (R(2)=0.913, p=0.068-0.075) comparing with other two-variable models, while the model including variables of CP, LIG and ADF performed the best in BMP prediction (R(2)=0.999, p=0.009-0.017) if three-variable models were compared. Among the three kinetic models used, the first order kinetic model fitted the measured BMPs data best (R(2)=0.996-0.998, rRMSE=0.171-0.381) and deviations between measured and the first order kinetic model predicted BMPs were less than 3.0%. Published by Elsevier Ltd.

BMP effectiveness/efficiency monitoring evaluation of regional information and data : response to Tasks 2 & 3 : information analysis and needs assessment.

DOT National Transportation Integrated Search

2002-10-04

The second task, Task 2, includes analyzing the information obtained in Task 1, expanding it if necessary, and identifying the extent of regional information and data with respect to the types of water quality facilities typically designed for the tr...

Establishment of Immortalized BMP2/4 Double Knock-Out Osteoblastic Cells Is Essential for Study of Osteoblast Growth, Differentiation, and Osteogenesis.

PubMed

Wu, Li-An; Wang, Feng; Donly, Kevin J; Baker, Andrew; Wan, Chunyan; Luo, Daoshu; MacDougall, Mary; Chen, Shuo

2016-06-01

Bone morphogenetic proteins 2 and 4 (BMP2/4) are essential for osteoblast differentiation and osteogenesis. Generation of a BMP2/4 dual knock-out ((ko/ko)) osteoblastic cell line is a valuable asset for studying effects of BMP2/4 on skeletal development. In this study, our goal was to create immortalized mouse deleted BMP2/4 osteoblasts by infecting adenoviruses with Cre recombinase and green fluorescent protein genes into immortalized murine floxed BMP2/4 osteoblasts. Transduced BMP2/4(ko/ko) cells were verified by green immunofluorescence and PCR. BMP2/4(ko/ko) osteoblasts exhibited small size, slow cell proliferation rate and cell growth was arrested in G1 and G2 phases. Expression of bone-relate genes was reduced in the BMP2/4(ko/ko) cells, resulting in delay of cell differentiation and mineralization. Importantly, extracellular matrix remodeling was impaired in the BMP2/4(ko/ko) osteoblasts as reflected by decreased Mmp-2 and Mmp-9 expressions. Cell differentiation and mineralization were rescued by exogenous BMP2 and/or BMP4. Therefore, we for the first time described establishment of an immortalized deleted BMP2/4 osteoblast line useful for study of mechanisms in regulating osteoblast lineages. © 2015 The Authors. Journal of Cellular Physiology Published by Wiley Periodicals, Inc.

Inactivation of bone morphogenetic protein 2 may predict clinical outcome and poor overall survival for renal cell carcinoma through epigenetic pathways

PubMed Central

Mitsui, Yozo; Hirata, Hiroshi; Arichi, Naoko; Hiraki, Miho; Yasumoto, Hiroaki; Chang, Inik; Fukuhara, Shinichiro; Yamamura, Soichiro; Shahryari, Varahram; Deng, Guoren; Saini, Sharanjot; Majid, Shahana; Dahiya, Rajvir; Tanaka, Yuichiro; Shiina, Hiroaki

2015-01-01

We investigated whether impaired regulation of bone morphogenetic protein-2 (BMP-2) via epigenetic pathways is associated with renal cell carcinoma (RCC) pathogenesis. Expression and CpG methylation of the BMP-2 gene were analyzed using RCC cell lines, and 96 matched RCC and normal renal tissues. We also performed functional analysis using BMP-2 restored RCC cells. A significant association of BMP-2 mRNA expression was also found with advanced tumor stage and lymph node involvement, while lower BMP-2 mRNA expression was significantly associated with poor overall survival after radical nephrectomy. In RCC cells, BMP-2 restoration significantly inhibited cell proliferation, migration, invasion, and colony formation. In addition, BMP-2 overexpression induced p21WAF1/CIP1 and p27KIP1 expression, and cellular apoptosis in RCC cells. BMP-2 mRNA expression was significantly enhanced in RCC cells by 5-aza-2′-deoxycitidine treatment. The prevalence of BMP-2 promoter methylation was significantly greater and BMP-2 mRNA expression was significantly lower in RCC samples as compared to normal kidney samples. Furthermore, a significant correlation was found between BMP-2 promoter methylation and mRNA transcription in tumors. Aberrant BMP-2 methylation and the resultant loss of BMP-2 expression may be a useful molecular marker for designing improved diagnostic and therapeutic strategies for RCC. PMID:25797254

2,2'-Bis(monoacylglycero) PO4 (BMP), but Not 3,1'-BMP, increases membrane curvature stress to enhance α-tocopherol transfer protein binding to membranes.

PubMed

Baptist, Matilda; Panagabko, Candace; Nickels, Jonathan D; Katsaras, John; Atkinson, Jeffrey

2015-03-01

Previous work revealed that α-tocopherol transfer protein (α-TTP) co-localizes with bis(monoacylglycero)phosphate (BMP) in late endosomes. BMP is a lipid unique to late endosomes and is believed to induce membrane curvature and support the multivesicular nature of this organelle. We examined the effect of BMP on α-TTP binding to membranes using dual polarization interferometry and vesicle-binding assay. α-TTP binding to membranes is increased by the curvature-inducing lipid BMP. α-TTP binds to membranes with greater affinity when they contain the 2,2'-BMP versus 3,1'-BMP isomers.

2,2'-Bis(monoacylglycero) PO 4 (BMP), but Not 3,1'-BMP, Increases Membrane Curvature Stress to Enhance α-Tocopherol Transfer Protein Binding to Membranes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Baptist, Matilda; Panagabko, Candace; Nickels, Jonathan D.

2015-01-21

Previous work revealed that α-tocopherol transfer protein (α-TTP) co-localizes with bis(monoacylglycero)phosphate (BMP) in late endosomes. BMP is a lipid unique to late endosomes and is believed to induce membrane curvature and support the multivesicular nature of this organelle. In this paper, we examined the effect of BMP on α-TTP binding to membranes using dual polarization interferometry and vesicle-binding assay. α-TTP binding to membranes is increased by the curvature-inducing lipid BMP. Finally, α-TTP binds to membranes with greater affinity when they contain the 2,2'-BMP versus 3,1'-BMP isomers.

BMP2 and BMP7 play antagonistic roles in feather induction

PubMed Central

Michon, Frédéric; Forest, Loïc; Collomb, Elodie; Demongeot, Jacques; Dhouailly, Danielle

2008-01-01

Summary During embryonic development, feathers first appear as primordia consisting of an epidermal placode associated with a dermal condensation. In most previous studies, the BMPs have been proposed to function as inhibitors of the formation of cutaneous appendages. We showed that the function of BMPs is quite nuanced: BMP-2 and BMP-7, which are expressed in both skin components, act antagonistically and yet are both involved in the dermal condensations formation. BMP-7, the first to be expressed, is implicated in chemotaxis which leads to cell recruitment to the condensation, whereas BMP-2, which is expressed later, leads to an arrest of cell migration, likely via its modulation of EIIIA Fibronectin domain and α4-Integrin expression. We also propose a mathematical model, a reaction-diffusion system, based on cell proliferation, chemotaxis and the timing of BMP-2 and BMP-7 expression, which simulates the endogenous situation and reproduces the negative effects of excess BMP-2 or BMP-7 on feather patterning. PMID:18635609

Comparison of the effects of recombinant human bone morphogenetic protein-2 and -9 on bone formation in rat calvarial critical-size defects.

PubMed

Nakamura, Toshiaki; Shirakata, Yoshinori; Shinohara, Yukiya; Miron, Richard J; Hasegawa-Nakamura, Kozue; Fujioka-Kobayashi, Masako; Noguchi, Kazuyuki

2017-12-01

Among bone morphogenetic protein (BMP) family members, BMP-2 and BMP-9 have demonstrated potent osteoinductive potential. However, in vivo differences in their potential for bone regeneration remain unclear. The present study aimed to compare the effects of recombinant human (rh) BMP-2 and rhBMP-9 on bone formation in rat calvarial critical-size defects (CSD). Twenty-eight Wistar rats surgically received two calvarial defects bilaterally in each parietal bone. Defects (n = 56) were allocated into four groups: absorbable collagen sponge (ACS) alone, rhBMP-2 with ACS (rhBMP-2/ACS), rhBMP-9/ACS, or sham surgery (control), on the condition that the treatments of rhBMP-2/ACS and rhBMP-9/ACS, or the same treatments were not included in the same animal. Animals were sacrificed at 2 and 8 weeks post-surgery. The calvarial defects were analyzed for bone volume (BV) by micro-computed tomography and for percentages of defect closure (DC/DL), newly formed bone area (NBA/TA), bone marrow area (BMA/NBA), adipose tissue area (ATA/NBA), central bone height (CBH), and marginal bone height (MBH) by histomorphometric analysis. The BV in the rhBMP-2/ACS group (5.44 ± 3.65 mm 3 , n = 7) was greater than the other groups at 2 weeks post-surgery, and the rhBMP-2/ACS and rhBMP-9/ACS groups (18.17 ± 2.51 and 16.30 ± 2.46 mm 3 , n = 7, respectively) demonstrated significantly greater amounts of BV compared with the control and ACS groups (6.02 ± 2.90 and 9.30 ± 2.75 mm 3 , n = 7, respectively) at 8 weeks post-surgery. The rhBMP-2/ACS and rhBMP-9/ACS groups significantly induced new bone formation compared to the control and ACS groups at 8 weeks post-surgery. However, there were no statistically significant differences found between the rhBMP-2/ACS and rhBMP-9/ACS groups in any of the histomorphometric parameters. The ATA/NBA in the rhBMP-2/ACS group (9.24 ± 3.72%, n = 7) was the highest among the treatment groups at 8 weeks post-surgery. Within the limits of this study, it can be concluded that rhBMP-2/ACS induced a slight early increase in new bone formation at 2 weeks and that rhBMP-9/ACS provided comparable new bone formation to rhBMP-2/ACS with less adipose tissues after a healing period of 8 weeks in rat CSD. RhBMP-9/ACS treatment provided new bone formation with less adipose tissues compared with rhBMP-2/ACS.

Covalent Binding of BMP-2 on Surfaces Using a Self-assembled Monolayer Approach

PubMed Central

Pohl, Theresa L. M.; Schwab, Elisabeth H.; Cavalcanti-Adam, Elisabetta A.

2013-01-01

Bone morphogenetic protein 2 (BMP-2) is a growth factor embedded in the extracellular matrix of bone tissue. BMP-2 acts as trigger of mesenchymal cell differentiation into osteoblasts, thus stimulating healing and de novo bone formation. The clinical use of recombinant human BMP-2 (rhBMP-2) in conjunction with scaffolds has raised recent controversies, based on the mode of presentation and the amount to be delivered. The protocol presented here provides a simple and efficient way to deliver BMP-2 for in vitro studies on cells. We describe how to form a self-assembled monolayer consisting of a heterobifunctional linker, and show the subsequent binding step to obtain covalent immobilization of rhBMP-2. With this approach it is possible to achieve a sustained presentation of BMP-2 while maintaining the biological activity of the protein. In fact, the surface immobilization of BMP-2 allows targeted investigations by preventing unspecific adsorption, while reducing the amount of growth factor and, most notably, hindering uncontrolled release from the surface. Both short- and long-term signaling events triggered by BMP-2 are taking place when cells are exposed to surfaces presenting covalently immobilized rhBMP-2, making this approach suitable for in vitro studies on cell responses to BMP-2 stimulation. PMID:24021994

Characterization of wise protein and its molecular mechanism to interact with both Wnt and BMP signals.

PubMed

Lintern, Katherine B; Guidato, Sonia; Rowe, Alison; Saldanha, José W; Itasaki, Nobue

2009-08-21

Cross-talk of BMP and Wnt signaling pathways has been implicated in many aspects of biological events during embryogenesis and in adulthood. A secreted protein Wise and its orthologs (Sostdc1, USAG-1, and Ectodin) have been shown to modulate Wnt signaling and also inhibit BMP signals. Modulation of Wnt signaling activity by Wise is brought about by an interaction with the Wnt co-receptor LRP6, whereas BMP inhibition is by binding to BMP ligands. Here we have investigated the mode of action of Wise on Wnt and BMP signals. It was found that Wise binds LRP6 through one of three loops formed by the cystine knot. The Wise deletion construct lacking the LRP6-interacting loop domain nevertheless binds BMP4 and inhibits BMP signals. Moreover, BMP4 does not interfere with Wise-LRP6 binding, suggesting separate domains for the physical interaction. Functional assays also show that the ability of Wise to block Wnt1 activity through LRP6 is not impeded by BMP4. In contrast, the ability of Wise to inhibit BMP4 is prevented by additional LRP6, implying a preference of Wise in binding LRP6 over BMP4. In addition to the interaction of Wise with BMP4 and LRP6, the molecular characteristics of Wise, such as glycosylation and association with heparan sulfate proteoglycans on the cell surface, are suggested. This study helps to understand the multiple functions of Wise at the molecular level and suggests a possible role for Wise in balancing Wnt and BMP signals.

Efficient delivery of recombinant human bone morphogenetic protein (rhBMP-2) with dextran sulfate-chitosan microspheres.

PubMed

Xia, Yuan-Jun; Xia, Hong; Chen, Ling; Ying, Qing-Shui; Yu, Xiang; Li, Li-Hua; Wang, Jian-Hua; Zhang, Ying

2018-04-01

Bone morphogenetic protein-2 (BMP-2) serves an important role in the development of bone and cartilage. However, administration of BMP-2 protein alone by intravenous delivery is not very effective. Sustained delivery of stabilized BMP-2 by carriers has been proven necessary to improve the osteogenesis effect of BMP-2. The present study constructed a novel drug delivery system using dextran sulfate (DS)-chitosan (CS) microspheres and investigated the efficiency of the delivery system on recombinant human bone morphogenetic protein (rhBMP-2). The microsphere morphology, optimal ratio of DS/CS/rhBMP-2, and drug loading rate and entrapment efficiency of rhBMP-2 CS nanoparticles were determined. L929 cells were used to evaluate the cytotoxicity and effect of DS/CS/rhBMP-2 microspheres on cell proliferation. Differentiation study was conducted using bone marrow mesenchymal stem cells (BMSCs-C57) cells treated with DS/CS/rhBMP-2 microspheres or the control microspheres. The DS/CS/rhBMP-2 microspheres delivery system was successfully established. Subsequent complexation of rhBMP-2-bound DS with polycations afforded well defined microspheres with a diameter of ~250 nm. High protein entrapment efficiency (85.6%) and loading ratio (47.245) µg/mg were achieved. Release of rhBMP-2 from resultant microspheres persisted for over 20 days as determined by ELISA assay. The bioactivity of rhBMP-2 encapsulated in the CS/DS microsphere was observed to be well preserved as evidenced by the alkaline phosphatase activity assay and calcium nodule formation of BMSCs-C57 incubated with rhBMP-2-loaded microspheres. The results demonstrated that microspheres based on CS-DS polyion complexes were a highly efficient vehicle for delivery of rhBMP-2 protein. The present study may provide novel orientation for bone tissue engineering for repairing and regenerating bone defects.

A computational model for BMP movement in sea urchin embryos.

PubMed

van Heijster, Peter; Hardway, Heather; Kaper, Tasso J; Bradham, Cynthia A

2014-12-21

Bone morphogen proteins (BMPs) are distributed along a dorsal-ventral (DV) gradient in many developing embryos. The spatial distribution of this signaling ligand is critical for correct DV axis specification. In various species, BMP expression is spatially localized, and BMP gradient formation relies on BMP transport, which in turn requires interactions with the extracellular proteins Short gastrulation/Chordin (Chd) and Twisted gastrulation (Tsg). These binding interactions promote BMP movement and concomitantly inhibit BMP signaling. The protease Tolloid (Tld) cleaves Chd, which releases BMP from the complex and permits it to bind the BMP receptor and signal. In sea urchin embryos, BMP is produced in the ventral ectoderm, but signals in the dorsal ectoderm. The transport of BMP from the ventral ectoderm to the dorsal ectoderm in sea urchin embryos is not understood. Therefore, using information from a series of experiments, we adapt the mathematical model of Mizutani et al. (2005) and embed it as the reaction part of a one-dimensional reaction-diffusion model. We use it to study aspects of this transport process in sea urchin embryos. We demonstrate that the receptor-bound BMP concentration exhibits dorsally centered peaks of the same type as those observed experimentally when the ternary transport complex (Chd-Tsg-BMP) forms relatively quickly and BMP receptor binding is relatively slow. Similarly, dorsally centered peaks are created when the diffusivities of BMP, Chd, and Chd-Tsg are relatively low and that of Chd-Tsg-BMP is relatively high, and the model dynamics also suggest that Tld is a principal regulator of the system. At the end of this paper, we briefly compare the observed dynamics in the sea urchin model to a version that applies to the fly embryo, and we find that the same conditions can account for BMP transport in the two types of embryos only if Tld levels are reduced in sea urchin compared to fly. Copyright © 2014 Elsevier Ltd. All rights reserved.

The role of the BMP signaling cascade in regulation of stem cell activity following massive small bowel resection in a rat.

PubMed

Sukhotnik, I; Berkowitz, D; Dorfman, T; Halabi, Salim; Pollak, Y; Bejar, J; Bitterman, A; Coran, A G

2016-02-01

Bone morphogenetic proteins (BMPs) are a group of growth factors that are implicated in intestinal growth, morphogenesis, differentiation, and homeostasis. The role of the BMP signaling cascade in stimulation of cell proliferation after massive small bowel resection is unknown. The purpose of this study was to evaluate the role of BMP signaling during intestinal adaptation in a rat model of short bowel syndrome (SBS). Male rats were divided into two groups: Sham rats underwent bowel transection and SBS rats underwent a 75 % bowel resection. Parameters of intestinal adaptation, enterocyte proliferation and apoptosis were determined 2 weeks after operation. Illumina's Digital Gene Expression analysis was used to determine the BMP signaling gene expression profiling. BMP-related genes and protein expression were determined using real-time PCR, Western blotting and immunohistochemistry. From the total number of 20,000 probes, 8 genes related to BMP signaling were investigated. From these genes, five genes were found to be up-regulated in jejunum (BMP1-10 %, BMP2-twofold increase, BMP3-10 %, BMP2R-12 % and STAT3-28 %) and four genes to be up-regulated in ileum (BMP1-16 %, BMP2-27 %, BMP3-10 %, and STAT3-20 %) in SBS vs sham animals with a relative change in gene expression level of 10 % or more. SBS rats also demonstrated a significant increase in BMP2 and STAT3 mRNA and protein levels (determined by real-time PCR and Western blot) compared to control animals. Two weeks following massive bowel resection in rats, the BMP signaling pathway is stimulated. BMP signaling may serve as an important mediator of reciprocal interactions between the epithelium and the underlying mesenchymal stroma during intestinal adaptation following massive bowel resection in a rat.

Uncovering the Role of BMP Signaling in Melanocyte Development and Melanoma Tumorigenesis

DTIC Science & Technology

2015-06-01

Graduate Program Nicola Kearns, UMass Medical School, Interdisciplinary Graduate Program Qualifying examination committees: Christopher Clark, UMass...Dutta, UMass Medical School, Interdisciplinary Graduate Program Nicola Kearns, UMass Medical School, Interdisciplinary Graduate Program Hsi-Ju Chen...Pigment Cell Melanoma Research, 24, 378-81. 13. Lian, C.G., Xu. Y., Ceol, C.J., Wu, F., Larson, A., Dresser, K., Xu, W., Tan , L., Zhan, Q., Lee, C., Hu, D

The level of BMP4 signaling is critical for the regulation of distinct T-box gene expression domains and growth along the dorso-ventral axis of the optic cup

PubMed Central

Behesti, Hourinaz; Holt, James KL; Sowden, Jane C

2006-01-01

Background Polarised gene expression is thought to lead to the graded distribution of signaling molecules providing a patterning mechanism across the embryonic eye. Bone morphogenetic protein 4 (Bmp4) is expressed in the dorsal optic vesicle as it transforms into the optic cup. Bmp4 deletions in human and mouse result in failure of eye development, but little attempt has been made to investigate mammalian targets of BMP4 signaling. In chick, retroviral gene overexpression studies indicate that Bmp4 activates the dorsally expressed Tbx5 gene, which represses ventrally expressed cVax. It is not known whether the Tbx5 related genes, Tbx2 and Tbx3, are BMP4 targets in the mammalian retina and whether BMP4 acts at a distance from its site of expression. Although it is established that Drosophila Dpp (homologue of vertebrate Bmp4) acts as a morphogen, there is little evidence that BMP4 gradients are interpreted to create domains of BMP4 target gene expression in the mouse. Results Our data show that the level of BMP4 signaling is critical for the regulation of distinct Tbx2, Tbx3, Tbx5 and Vax2 gene expression domains along the dorso-ventral axis of the mouse optic cup. BMP4 signaling gradients were manipulated in whole mouse embryo cultures during optic cup development, by implantation of beads soaked in BMP4, or the BMP antagonist Noggin, to provide a local signaling source. Tbx2, Tbx3 and Tbx5, showed a differential response to alterations in the level of BMP4 along the entire dorso-ventral axis of the optic cup, suggesting that BMP4 acts across a distance. Increased levels of BMP4 caused expansion of Tbx2 and Tbx3, but not Tbx5, into the ventral retina and repression of the ventral marker Vax2. Conversely, Noggin abolished Tbx5 expression but only shifted Tbx2 expression dorsally. Increased levels of BMP4 signaling caused decreased proliferation, reduced retinal volume and altered the shape of the optic cup. Conclusion Our findings suggest the existence of a dorsal-high, ventral-low BMP4 signaling gradient across which distinct domains of Tbx2, Tbx3, Tbx5 and Vax2 transcription factor gene expression are set up. Furthermore we show that the correct level of BMP4 signaling is critical for normal growth of the mammalian embryonic eye. PMID:17173667

Bone morphogenetic protein (BMP)1-3 enhances bone repair.

PubMed

Grgurevic, Lovorka; Macek, Boris; Mercep, Mladen; Jelic, Mislav; Smoljanovic, Tomislav; Erjavec, Igor; Dumic-Cule, Ivo; Prgomet, Stefan; Durdevic, Dragan; Vnuk, Drazen; Lipar, Marija; Stejskal, Marko; Kufner, Vera; Brkljacic, Jelena; Maticic, Drazen; Vukicevic, Slobodan

2011-04-29

Members of the astacin family of metalloproteinases such as human bone morphogenetic protein 1 (BMP-1) regulate morphogenesis by processing precursors to mature functional extracellular matrix (ECM) proteins and several growth factors including TGFβ, BMP2, BMP4 and GFD8. We have recently discovered that BMP1-3 isoform of the Bmp-1 gene circulates in the human plasma and is significantly increased in patients with acute bone fracture. We hypothesized that circulating BMP1-3 might have an important role in bone repair and serve as a novel bone biomarker. When administered systemically to rats with a long bone fracture and locally to rabbits with a critical size defect of the ulna, recombinant human BMP1-3 enhanced bone healing. In contrast, neutralization of the endogenous BMP1-3 by a specific polyclonal antibody delayed the bone union. Invitro BMP1-3 increased the expression of collagen type I and osteocalcin in MC3T3-E(1) osteoblast like cells, and enhanced the formation of mineralized bone nodules from bone marrow mesenchymal stem cells. We suggest that BMP1-3 is a novel systemic regulator of bone repair. Copyright © 2011 Elsevier Inc. All rights reserved.

Ectopic application of recombinant BMP-2 and BMP-4 can change patterning of developing chick facial primordia.

PubMed

Barlow, A J; Francis-West, P H

1997-01-01

The facial primordia initially consist of buds of undifferentiated mesenchyme, which give rise to a variety of tissues including cartilage, muscle and nerve. These must be arranged in a precise spatial order for correct function. The signals that control facial outgrowth and patterning are largely unknown. The bone morphogenetic proteins Bmp-2 and Bmp-4 are expressed in discrete regions at the distal tips of the early facial primordia suggesting possible roles for BMP-2 and BMP-4 during chick facial development. We show that expression of Bmp-4 and Bmp-2 is correlated with the expression of Msx-1 and Msx-2 and that ectopic application of BMP-2 and BMP-4 can activate Msx-1 and Msx-2 gene expression in the developing facial primordia. We correlate this activation of gene expression with changes in skeletal development. For example, activation of Msx-1 gene expression across the distal tip of the mandibular primordium is associated with an extension of Fgf-4 expression in the epithelium and bifurcation of Meckel's cartilage. In the maxillary primordium, extension of the normal domain of Msx-1 gene expression is correlated with extended epithelial expression of shh and bifurcation of the palatine bone. We also show that application of BMP-2 can increase cell proliferation of the mandibular primordia. Our data suggest that BMP-2 and BMP-4 are part of a signalling cascade that controls outgrowth and patterning of the facial primordia.

Dual Delivery of EPO and BMP2 from a Novel Modular Poly-ɛ-Caprolactone Construct to Increase the Bone Formation in Prefabricated Bone Flaps

PubMed Central

Patel, Janki Jayesh; Modes, Jane E.; Flanagan, Colleen L.; Krebsbach, Paul H.; Edwards, Sean P.

2015-01-01

Poly-ɛ-caprolactone (PCL) is a biocompatible polymer that has mechanical properties suitable for bone tissue engineering; however, it must be integrated with biologics to stimulate bone formation. Bone morphogenetic protein-2 (BMP2) delivered from PCL produces bone when implanted subcutaneously, and erythropoietin (EPO) works synergistically with BMP2. In this study, EPO and BMP2 are adsorbed separately on two 3D-printed PCL scaffold modules that are assembled for codelivery on a single scaffold structure. This assembled modular PCL scaffold with dual BMP2 and EPO delivery was shown to increase bone growth in an ectopic location when compared with BMP2 delivery along a replicate scaffold structure. EPO (200 IU/mL) and BMP2 (65 μg/mL) were adsorbed onto the outer and inner portions of a modular scaffold, respectively. Protein binding and release studies were first quantified. Subsequently, EPO+BMP2 and BMP2 scaffolds were implanted subcutaneously in mice for 4 and 8 weeks, and the regenerated bone was analyzed with microcomputed tomography and histology; 8.6±1.4 μg BMP2 (22%) and 140±29 IU EPO (69.8%) bound to the scaffold and <1% BMP2 and 83% EPO was released in 7 days. Increased endothelial cell proliferation on EPO-adsorbed PCL discs indicated protein bioactivity. At 4 and 8 weeks, dual BMP2 and EPO delivery regenerated more bone (5.1±1.1 and 5.5±1.6 mm3) than BMP2 alone (3.8±1.1 and 4.3±1.7 mm3). BMP2 and EPO scaffolds had more ingrowth (1.4%±0.6%) in the outer module when compared with BMP2 (0.8%±0.3%) at 4 weeks. Dual delivery produced more dense cellular marrow, while BMP2 had more fatty marrow. Dual EPO and BMP2 delivery is a potential method to regenerate bone faster for prefabricated flaps. PMID:25809081

Dual Delivery of EPO and BMP2 from a Novel Modular Poly-ɛ-Caprolactone Construct to Increase the Bone Formation in Prefabricated Bone Flaps.

PubMed

Patel, Janki Jayesh; Modes, Jane E; Flanagan, Colleen L; Krebsbach, Paul H; Edwards, Sean P; Hollister, Scott J

2015-09-01

Poly-ɛ-caprolactone (PCL) is a biocompatible polymer that has mechanical properties suitable for bone tissue engineering; however, it must be integrated with biologics to stimulate bone formation. Bone morphogenetic protein-2 (BMP2) delivered from PCL produces bone when implanted subcutaneously, and erythropoietin (EPO) works synergistically with BMP2. In this study, EPO and BMP2 are adsorbed separately on two 3D-printed PCL scaffold modules that are assembled for codelivery on a single scaffold structure. This assembled modular PCL scaffold with dual BMP2 and EPO delivery was shown to increase bone growth in an ectopic location when compared with BMP2 delivery along a replicate scaffold structure. EPO (200 IU/mL) and BMP2 (65 μg/mL) were adsorbed onto the outer and inner portions of a modular scaffold, respectively. Protein binding and release studies were first quantified. Subsequently, EPO+BMP2 and BMP2 scaffolds were implanted subcutaneously in mice for 4 and 8 weeks, and the regenerated bone was analyzed with microcomputed tomography and histology; 8.6±1.4 μg BMP2 (22%) and 140±29 IU EPO (69.8%) bound to the scaffold and <1% BMP2 and 83% EPO was released in 7 days. Increased endothelial cell proliferation on EPO-adsorbed PCL discs indicated protein bioactivity. At 4 and 8 weeks, dual BMP2 and EPO delivery regenerated more bone (5.1±1.1 and 5.5±1.6 mm(3)) than BMP2 alone (3.8±1.1 and 4.3±1.7 mm(3)). BMP2 and EPO scaffolds had more ingrowth (1.4%±0.6%) in the outer module when compared with BMP2 (0.8%±0.3%) at 4 weeks. Dual delivery produced more dense cellular marrow, while BMP2 had more fatty marrow. Dual EPO and BMP2 delivery is a potential method to regenerate bone faster for prefabricated flaps.

Addition of bone morphogenetic protein type 2 to ascorbate and β-glycerophosphate supplementation did not enhance osteogenic differentiation of human adipose-derived stem cells.

PubMed

Cruz, Ariadne Cristiane Cabral; Silva, Mariana Lúcia; Caon, Thiago; Simões, Cláudia Maria Oliveira

2012-01-01

Bone morphogenetic protein type 2 (BMP-2) is a potent local factor, which promotes bone formation and has been used as an osteogenic supplement for mesenchymal stem cells. This study evaluated the effect of a recombinant BMP-2 as well as the endogenous BMP-4 and BMP-7 in the osteogenic differentiation of adipose-derived stem cells (ASCs) in medium supplemented with ascorbate and β-glycerophosphate. Human ASCs were treated with osteogenic medium in the presence (ASCs+OM+BMP-2) or absence (ASCs+OM) of BMP-2. The alkaline phosphatase (ALP) activity was determined and the extracellular matrix mineralization was evaluated by Von Kossa staining and calcium quantification. The expressions of BMP-4, BMP-7, Smad1, Smad4, and phosphorylated Smad1/5/8 were analyzed by western blotting. Relative mRNA expressions of Smad1, BMP receptor type II (BMPR-II), osteonectin, and osteocalcin were evaluated by qPCR. ASCs+OM demonstrated the highest expression of BMP-4 and BMP-7 at days 21 and 7, respectively, the highest levels of BMPR-II mRNA expression at day 28, and the highest levels of Smad1 mRNA at days 14 and 28. ASCs+OM+BMP-2 demonstrated the highest levels of Smad1 mRNA expression at days 1, 7, and 21, the highest expression of Smad1 at day 7, the highest expression of Smad4 at day 14, the highest ALP activity at days 14 and 21, and expression of phosphorylated Smad1/5/8 at day 7. ASCs+OM and ASCs+OM+BMP2 showed similar ALP activity at days 7 and 28, similar osteonectin and osteocalcin mRNA expression at all time periods, and similar calcium depositions at all time periods. We concluded that human ASCs expressed endogenous BMP-4 and BMP-7. Moreover, the supplementation of ASCs with BMP-2 did not increase the level of osteogenic markers in the initial (ALP activity), intermediate (osteonectin and osteocalcin), or final (calcium deposition) phases, suggesting that the exogenous addition of BMP-2 did not improve the in vitro osteogenesis process of human ASCs.

Stiffness-dependent cellular internalization of matrix-bound BMP-2 and its relation to Smad and non-Smad signaling.

PubMed

Gilde, Flora; Fourel, Laure; Guillot, Raphael; Pignot-Paintrand, Isabelle; Okada, Takaharu; Fitzpatrick, Vincent; Boudou, Thomas; Albiges-Rizo, Corinne; Picart, Catherine

2016-12-01

Surface coatings delivering BMP are a promising approach to render biomaterials osteoinductive. In contrast to soluble BMPs which can interact with their receptors at the dorsal side of the cell, BMPs presented as an insoluble cue physically bound to a biomimetic matrix, called here matrix-bound (bBMP-2), are presented to cells by their ventral side. To date, BMP-2 internalization and signaling studies in cell biology have always been performed by adding soluble (sBMP-2) to cells adhered on cell culture plates or glass slides, which will be considered here as a "reference" condition. However, whether and how matrix-bound BMP-2 can be internalized by cells and its relation to canonical (SMAD) and non-canonical signaling (ALP) remain open questions. In this study, we investigated the uptake and processing of BMP-2 by C2C12 myoblasts. This BMP-2 was presented either embedded in polyelectrolyte multilayer films (matrix-bound presentation) or as soluble form. Using fluorescently labeled BMP-2, we showed that the amount of matrix-bound BMP-2 internalized is dependent on the level of crosslinking of the polyelectrolyte films. Cav-1-mediated internalization is related to both SMAD and ALP signaling, while clathrin-mediated is only related to ALP signaling. BMP-2 internalization was independent of the presentation mode (sBMP-2 versus bBMP-2) for low crosslinked films (soft, EDC10) in striking contrast with high crosslinked (stiff, EDC70) films where internalization was much lower and slower for bBMP-2. As anticipated, internalization of sBMP-2 barely depended on the underlying matrix. Taken together, these results indicate that BMP-2 internalization can be tuned by the underlying matrix and activates downstream BMP-2 signaling, which is key for the effective formation of bone tissue. The presentation of growth factors from material surfaces currently presents significant challenges in academic research, clinics and industry. Being able to deliver efficiently these growth factors by a biomaterial will open new perspectives for regenerative medicine. However, to date, very little is known about how matrix-bound growth factors are delivered to cells, especially whether they are internalized and how they are signaling to drive key differentiation events. These initial steps are crucial as they will guide the subsequent processes leading to tissue regeneration. In this work, we investigate the uptake and processing by cells of BMP-2 ligands embedded in polyelectrolyte multilayer films in comparison to soluble BMP-2. We show that BMP-2 responsive cells can internalize matrix-bound BMP-2 and that internalization is dependent on the cross-linking level of the polyelectrolyte films. In addition, we show that internalization is mediated by both clathrin- and caveolin-dependent pathways. While inhibiting clathrin-dependent endocytosis affects only non-canonical signaling, blocking caveolin-1-dependent endocytosis reduces both canonical and non-canonical BMP signaling. The signaling pathways found for matrix-bound BMP-2 are similar to those found for soluble BMP-2. These results highlight that BMP-2 presented by a biomaterial at the ventral side of the cell can trigger major endocytic and associated signaling pathways leading to bone regeneration. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

A review on effectiveness of best management practices in improving hydrology and water quality: Needs and opportunities.

PubMed

Liu, Yaoze; Engel, Bernard A; Flanagan, Dennis C; Gitau, Margaret W; McMillan, Sara K; Chaubey, Indrajeet

2017-12-01

Best management practices (BMPs) have been widely used to address hydrology and water quality issues in both agricultural and urban areas. Increasing numbers of BMPs have been studied in research projects and implemented in watershed management projects, but a gap remains in quantifying their effectiveness through time. In this paper, we review the current knowledge about BMP efficiencies, which indicates that most empirical studies have focused on short-term efficiencies, while few have explored long-term efficiencies. Most simulation efforts that consider BMPs assume constant performance irrespective of ages of the practices, generally based on anticipated maintenance activities or the expected performance over the life of the BMP(s). However, efficiencies of BMPs likely change over time irrespective of maintenance due to factors such as degradation of structures and accumulation of pollutants. Generally, the impacts of BMPs implemented in water quality protection programs at watershed levels have not been as rapid or large as expected, possibly due to overly high expectations for practice long-term efficiency, with BMPs even being sources of pollutants under some conditions and during some time periods. The review of available datasets reveals that current data are limited regarding both short-term and long-term BMP efficiency. Based on this review, this paper provides suggestions regarding needs and opportunities. Existing practice efficiency data need to be compiled. New data on BMP efficiencies that consider important factors, such as maintenance activities, also need to be collected. Then, the existing and new data need to be analyzed. Further research is needed to create a framework, as well as modeling approaches built on the framework, to simulate changes in BMP efficiencies with time. The research community needs to work together in addressing these needs and opportunities, which will assist decision makers in formulating better decisions regarding BMP implementation in watershed management projects. Copyright © 2017 Elsevier B.V. All rights reserved.

Spatially-Distributed Cost–Effectiveness Analysis Framework to Control Phosphorus from Agricultural Diffuse Pollution

PubMed Central

Geng, Runzhe; Wang, Xiaoyan; Sharpley, Andrew N.; Meng, Fande

2015-01-01

Best management practices (BMPs) for agricultural diffuse pollution control are implemented at the field or small-watershed scale. However, the benefits of BMP implementation on receiving water quality at multiple spatial is an ongoing challenge. In this paper, we introduce an integrated approach that combines risk assessment (i.e., Phosphorus (P) index), model simulation techniques (Hydrological Simulation Program–FORTRAN), and a BMP placement tool at various scales to identify the optimal location for implementing multiple BMPs and estimate BMP effectiveness after implementation. A statistically significant decrease in nutrient discharge from watersheds is proposed to evaluate the effectiveness of BMPs, strategically targeted within watersheds. Specifically, we estimate two types of cost-effectiveness curves (total pollution reduction and proportion of watersheds improved) for four allocation approaches. Selection of a ‘‘best approach” depends on the relative importance of the two types of effectiveness, which involves a value judgment based on the random/aggregated degree of BMP distribution among and within sub-watersheds. A statistical optimization framework is developed and evaluated in Chaohe River Watershed located in the northern mountain area of Beijing. Results show that BMP implementation significantly (p >0.001) decrease P loss from the watershed. Remedial strategies where BMPs were targeted to areas of high risk of P loss, deceased P loads compared with strategies where BMPs were randomly located across watersheds. Sensitivity analysis indicated that aggregated BMP placement in particular watershed is the most cost-effective scenario to decrease P loss. The optimization approach outlined in this paper is a spatially hierarchical method for targeting nonpoint source controls across a range of scales from field to farm, to watersheds, to regions. Further, model estimates showed targeting at multiple scales is necessary to optimize program efficiency. The integrated model approach described that selects and places BMPs at varying levels of implementation, provides a new theoretical basis and technical guidance for diffuse pollution management in agricultural watersheds. PMID:26313561

Differential expression of a BMP4 reporter allele in anterior fungiform versus posterior circumvallate taste buds of mice.

PubMed

Nguyen, Ha M; Barlow, Linda A

2010-10-13

Bone Morphogenetic Protein 4 (BMP4) is a diffusible factor which regulates embryonic taste organ development. However, the role of BMP4 in taste buds of adult mice is unknown. We utilized transgenic mice with LacZ under the control of the BMP4 promoter to reveal the expression of BMP4 in the tongues of adult mice. Further we evaluate the pattern of BMP4 expression with that of markers of specific taste bud cell types and cell proliferation to define and compare the cell populations expressing BMP4 in anterior (fungiform papillae) and posterior (circumvallate papilla) tongue. BMP4 is expressed in adult fungiform and circumvallate papillae, i.e., lingual structures composed of non-taste epithelium and taste buds. Unexpectedly, we find both differences and similarities with respect to expression of BMP4-driven ß-galactosidase. In circumvallate papillae, many fusiform cells within taste buds are BMP4-ß-gal positive. Further, a low percentage of BMP4-expressing cells within circumvallate taste buds is immunopositive for markers of each of the three differentiated taste cell types (I, II and III). BMP4-positive intragemmal cells also expressed a putative marker of immature taste cells, Sox2, and consistent with this finding, intragemmal cells expressed BMP4-ß-gal within 24 hours after their final mitosis, as determined by BrdU birthdating. By contrast, in fungiform papillae, BMP4-ß-gal positive cells are never encountered within taste buds. However, in both circumvallate and fungiform papillae, BMP4-ß-gal expressing cells are located in the perigemmal region, comprising basal and edge epithelial cells adjacent to taste buds proper. This region houses the proliferative cell population that gives rise to adult taste cells. However, perigemmal BMP4-ß-gal cells appear mitotically silent in both fungiform and circumvallate taste papillae, as we do not find evidence of their active proliferation using cell cycle immunomarkers and BrdU birthdating. Our data suggest that intragemmal BMP4-ß-gal cells in circumvallate papillae are immature taste cells which eventually differentiate into each of the 3 taste cell types, whereas perigemmal BMP4-ß-gal cells in both circumvallate and fungiform papillae may be slow cycling stem cells, or belong to the stem cell niche to regulate taste cell renewal from the proliferative cell population.

Bone morphogenetic protein and Notch signalling crosstalk in poor-prognosis, mesenchymal-subtype colorectal cancer.

PubMed

Irshad, Shazia; Bansal, Mukesh; Guarnieri, Paolo; Davis, Hayley; Al Haj Zen, Ayman; Baran, Brygida; Pinna, Claudia Maria Assunta; Rahman, Haseeb; Biswas, Sujata; Bardella, Chiara; Jeffery, Rosemary; Wang, Lai Mun; East, James Edward; Tomlinson, Ian; Lewis, Annabelle; Leedham, Simon John

2017-06-01

The functional role of bone morphogenetic protein (BMP) signalling in colorectal cancer (CRC) is poorly defined, with contradictory results in cancer cell line models reflecting the inherent difficulties of assessing a signalling pathway that is context-dependent and subject to genetic constraints. By assessing the transcriptional response of a diploid human colonic epithelial cell line to BMP ligand stimulation, we generated a prognostic BMP signalling signature, which was applied to multiple CRC datasets to investigate BMP heterogeneity across CRC molecular subtypes. We linked BMP and Notch signalling pathway activity and function in human colonic epithelial cells, and normal and neoplastic tissue. BMP induced Notch through a γ-secretase-independent interaction, regulated by the SMAD proteins. In homeostasis, BMP/Notch co-localization was restricted to cells at the top of the intestinal crypt, with more widespread interaction in some human CRC samples. BMP signalling was downregulated in the majority of CRCs, but was conserved specifically in mesenchymal-subtype tumours, where it interacts with Notch to induce an epithelial-mesenchymal transition (EMT) phenotype. In intestinal homeostasis, BMP-Notch pathway crosstalk is restricted to differentiating cells through stringent pathway segregation. Conserved BMP activity and loss of signalling stringency in mesenchymal-subtype tumours promotes a synergistic BMP-Notch interaction, and this correlates with poor patient prognosis. BMP signalling heterogeneity across CRC subtypes and cell lines can account for previous experimental contradictions. Crosstalk between the BMP and Notch pathways will render mesenchymal-subtype CRC insensitive to γ-secretase inhibition unless BMP activation is concomitantly addressed. © 2017 The Authors. Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland. © 2017 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

Overlapping and differential localization of Bmp-2, Bmp-4, Msx-2 and apoptosis in the endocardial cushion and adjacent tissues of the developing mouse heart.

PubMed

Abdelwahid, E; Rice, D; Pelliniemi, L J; Jokinen, E

2001-07-01

The bone morphogenetic proteins BMP-2 and BMP-4 and the homeobox gene MSX-2 are required for normal development of many embryonic tissues. To elucidate their possible roles during the remodeling of the tubular heart into a fully septated four-chambered heart, we have localized the mRNA of Bmp-2, Bmp-4, Msx-2 and apoptotic cells in the developing mouse heart from embryonic day (E)11 to E17. mRNA was localized by in situ hybridization, and apoptotic cells by TUNEL (TDT-mediated dUTP-biotin nick end-labeling) as well as by transmission electron microscopy. By analyzing adjacent serial sections, we demonstrated that the expression of Msx-2 and Bmp-2 strikingly overlapped in the atrioventricular canal myocardium, in the atrioventricular junctional myocardium, and in the maturing myocardium of the atrioventricular valves. Bmp-4 was expressed in the outflow tract myocardium and in the endocardial cushion of the outflow tract ridges from E12 to E14. Msx-2 appeared in the mesenchyme of the atrioventricular endocardial cushion from E11 to E14, while Bmp-2 and Bmp-4 were detected between E11 and E14. Apoptotic cells were also detected in the mesenchyme of the endocardial cushion between E12 and E14. Our results suggest that BMP-2 and MSX-2 are tightly linked to the formation of the atrioventricular junction and valves and that BMP-4 is involved in the development of the outflow tract myocardium and of the endocardial cushion. In addition, BMP-2, BMP-4 and MSX-2 and apoptosis seem to be associated with differentiation of the endocardial cushion.

Smad, PI3K/Akt, and Wnt-dependent signaling pathways are involved in BMP-4-induced ESC self-renewal.

PubMed

Lee, Min Young; Lim, Hyun Woo; Lee, Sang Hun; Han, Ho Jae

2009-08-01

It is known that bone morphogenetic protein 4 (BMP-4) has a diverse effect on ESCs. However, its precise mechanism in mouse ESCs is not fully understood. We evaluated the effect of BMP-4 on ESC proliferation and its related signal cascades in this study. BMP-4 significantly increased the level of [(3)H]-thymidine incorporation in time- (> or =8 hours) and dose- (> or =10 ng/ml) dependent manners. Additionally, BMP-4 increased cyclin D1 and decreased p27(kip1) expression values in a time-dependent manner. The increases in BMP-4-induced [(3)H]-thymidine incorporation and cyclin D1 expression were inhibited by the BMP-4 receptor antagonist noggin. BMP-4 increased Wnt1 expression. Wnt1 expression was attenuated by Smad4 small interfering RNA (siRNA), and BMP-4-induced cyclin D1 expression was inhibited by Smad4 and Wnt1 siRNAs. BMP-4 also activated beta-catenin, which was blocked by Smad4 and Wnt1 siRNAs. In addition, BMP-4 induced Akt phosphorylation. BMP-4-induced beta-catenin activation and cyclin D1 expression were attenuated by phosphatidyl inositol 3-kinase (PI3K) siRNA and Akt inhibitor. Additionally, downregulation of Smad4, Wnt1, and PI3K expression by siRNA decreased the levels of pluripotency marker mRNAs of ESCs, including Oct4, Sox2, and FoxD3. Our results suggested that BMP-4-induced [(3)H]-thymidine incorporation was significantly attenuated by Smad4, Wnt1, and PI3K knockdown. In conclusion, BMP-4 contributed to the maintenance of cell proliferation and the pluripotent state by Smad, PI3K/Akt, and Wnt1/beta-catenin in mouse ESCs.

EFFECT OF COLLA CORNUS CERVI COMBINED WITH LV-MEDIATED BMP7 TRANSFECTED BMSCs ON ANFH IN RATS.

PubMed

Wang, Ping; Shi, Bin; Gao, Zhi-Hui; Sun, Tie-Feng; Yang, Wu-Bin; Han, Shu-Fang; Liu, Peng; Wang, Lei-Lei; Zhao, Bo-Nian; Wang, Dan-Dan

2016-11-01

In the present study, we investigated the combined effect of Colla Comus Cervi (CCC) and BMP7-overexpressing bone marrow-derived mesenchymal stem cells (BMSCs) on osteogenic induction and the treatment of avascular necrosis of the femoral head (ANFH). BMSCs were isolated from rats. BMP7-overexpressing BMSCs were generated by lentiviral-mediated gene transduction. Cell proliferation, alkaline phosphatase (ALP) activity, osteogenesis related gene expression, osteocalcin levels, and calcified nodules were quantified and compared between four groups: untreated controls, BMSCs cultured with CCC complex medium, BMP7-overexpressing BMSCs, and BMP7-overexpressing BMSCs cultured with CCC complex medium (CCC+BMP7). CCC+BMP7 BMSCs showed higher proliferation rate. ALP activity and osteaocalcin content were significantly increased in CCC+BMP7 BMSCs. The osteogenesis related genes, COLI, and integrin-α2, -α5, and -β1, were expressed significantly higher in CCC+BMP7 BMSCs. The number of calcified nodules in the CCC+BMP7 group was significantly higher than that in other groups. For in vivo assays, ANFH was induced in rats, and BMSCs were injected into the femoral head of the lower left extremity. In rats with induced ANFH, general observation scores of the CCC+BMP7 injected group were significantly higher than the model group. X-ray and microscopic observations revealed that ANFH was significantly improved and femoral head cells gradually recovered in rats treated with CCC+BMP7 BMSCs. Our results suggest that CCC+BMP7 significantly promote the proliferation and osteogenic differentiation of BMSCs in vitm. CCC+BMP7 BMSCs promote the ability of repairing ANFH in rats, providing a new therapeutic paradigm for the treatment of ANFH.

Strontium doping promotes bioactivity of rhBMP-2 upon calcium phosphate cement via elevated recognition and expression of BMPR-IA.

PubMed

Huang, Baolin; Tian, Yu; Zhang, Wenjing; Ma, Yifan; Yuan, Yuan; Liu, Changsheng

2017-11-01

Preserving and improving osteogenic activity of bone morphogenetic protein-2 (BMP-2) upon implants remains one of the key limitations in bone regeneration. With calcium phosphate cement (CPC) as model, we have developed a series of strontium (Sr)-doped CPC (SCPC) to address this issue. The effects of fixed Sr on the bioactivity of recombinant human BMP-2 (rhBMP-2) as well as the underlying mechanism were investigated. The results suggested that the rhBMP-2-induced osteogenic activity was significantly promoted upon SCPCs, especially with a low amount of fixed Sr (SrCO 3 content <10wt%). Further studies demonstrated that the Sr-induced enhancement of bioactivity of rhBMP-2 was related to an elevated recognition of bone morphogenetic protein receptor-IA (BMPR-IA) to rhBMP-2 and an increased expression of BMPR-IA in C2C12 model cells. As a result, the activations of BMP-induced signaling pathways were different in C2C12 cells incubated upon CPC/rhBMP-2 and SCPCs/rhBMP-2. These findings explicitly decipher the mechanism of SCPCs promoting osteogenic bioactivity of rhBMP-2 and signify the promising application of the SCPCs/rhBMP-2 matrix in bone regeneration implants. Copyright © 2017 Elsevier B.V. All rights reserved.

A Bmp/Admp regulatory circuit controls maintenance and regeneration of dorsal-ventral polarity in planarians.

PubMed

Gaviño, Michael A; Reddien, Peter W

2011-02-22

Animal embryos have diverse anatomy and vary greatly in size. It is therefore remarkable that a common signaling pathway, BMP signaling, controls development of the dorsoventral (DV) axis throughout the Bilateria. In vertebrates, spatially opposed expression of the BMP family proteins Bmp4 and Admp (antidorsalizing morphogenetic protein) can promote restoration of DV pattern following tissue removal. bmp4 orthologs have been identified in all three groups of the Bilateria (deuterostomes, ecdysozoans, and lophotrochozoans). By contrast, the absence of admp orthologs in ecdysozoans such as Drosophila and C. elegans has suggested that a regulatory circuit of oppositely expressed bmp4 and admp genes represents a deuterostome-specific innovation. Here we describe the existence of spatially opposed bmp and admp expression in a protostome. An admp ortholog (Smed-admp) is expressed ventrally and laterally in adult Schmidtea mediterranea planarians, opposing the dorsal-pole expression of Smed-bmp4. Smed-admp is required for regeneration following parasagittal amputation. Furthermore, Smed-admp promotes Smed-bmp4 expression and Smed-bmp4 inhibits Smed-admp expression, generating a regulatory circuit that buffers against perturbations of Bmp signaling. These results suggest that a Bmp/Admp regulatory circuit is a central feature of the Bilateria, used broadly for the establishment, maintenance, and regeneration of the DV axis. Copyright © 2011 Elsevier Ltd. All rights reserved.

A Bmp/Admp regulatory circuit controls maintenance and regeneration of dorsal-ventral polarity in planarians

PubMed Central

Gaviño, Michael A.; Reddien, Peter W.

2011-01-01

Animal embryos have diverse anatomy and vary greatly in size. It is therefore remarkable that a common signaling pathway – BMP signaling – controls development of the dorsoventral (DV) axis throughout the Bilateria [1-8]. In vertebrates, spatially opposed expression of the BMP-family signaling proteins Bmp4 and Admp (anti-dorsalizing morphogenetic protein) can promote restoration of DV pattern following tissue removal [9-11]. bmp4 orthologs have been identified in all three groups of the Bilateria (deuterostomes, ecdysozoans, and lophotrochozoans) [12]. By contrast, the absence of admp orthologs in ecdysozoans such as Drosophila and C. elegans has suggested that a DV regulatory circuit of oppositely expressed bmp4 and admp genes represents an innovation specific to deuterostomes. Here we describe the existence of spatially opposed bmp and admp expression in a protostome. An admp ortholog (Smed-admp) is expressed at the ventral pole and laterally in adult Schmidtea mediterranea planarians, spatially opposing the dorsal-pole domain of Smed-bmp4 expression. Smed-admp is required for planarian regeneration following parasagittal amputation. Furthermore, Smed-admp promotes Smed-bmp4 expression and Smed-bmp4 inhibits Smed-admp expression, generating a regulatory circuit that buffers against perturbations of Bmp signaling. These results suggest that a Bmp/Admp regulatory circuit is a central feature of the Bilateria, used broadly for the establishment, maintenance, and regeneration of the DV axis. PMID:21295483

Results of a program to control phosphorus discharges from dairy operations in south-central Florida, USA

NASA Astrophysics Data System (ADS)

Havens, Karl E.; Flaig, Eric G.; James, R. Thomas; Lostal, Sergio; Muszick, Dera

1996-07-01

During 1987 1992, a mandatory program to control phosphorus discharges was implemented at dairy operations located to the north of Lake Okeechobee, Florida, USA. Thirty of 48 dairies participated in this program and implemented best management practices (BMPs), which included the construction of intensive animal waste management systems. Eighteen dairies closed their milkproducing operations under a government-funded buyout program. In this paper, we compare trends in runoff total phosphorus (TP) concentrations among the dairies that remained active and implemented BMPs. A central feature of the dairy waste management system is the high intensity area (HIA), defined as the milking barn and adjacent vegetation-free land, encircled by a drainage ditch and dike. Animal waste from the HIA is diverted into anaerobic lagoons and storage ponds, from which water is periodically removed and used for irrigation of field crops. The impacts of BMP construction on runoff TP concentrations were immediate and, in most cases, dramatic. Average TP concentrations declined significantly ( P < 0.001), from 9.0 to 1.2 mg TP liter-1 at dairies in one basin (Lower Kissimmee River), and from 2.6 to 1.0 mg TP liter-1 in another (Taylor Creek/Nubbin Slough). Some sites experienced greater declines in TP than others. To elucidate possible causes for the difference in response, a multivariate statistical model was utilized. Independent variables included soil pH, soil drainage characteristics, spodic horizon depth, and the areas of different BMP components (pasture, HIA, spray fields). The analysis significantly separated dairies with the highest and lowest runoff TP concentrations. Lowest TP occurred at dairies having particular soil characteristic (shallow spodic horizon) and certain BMP features (large HIA and small heard pastures).

Comparison of transforaminal lumbar interbody fusion outcomes in patients receiving rhBMP-2 versus autograft.

PubMed

Khan, Taleef R; Pearce, Kalin R; McAnany, Steven J; Peters, Colleen M; Gupta, Munish C; Zebala, Lukas P

2018-03-01

Recombinant human bone morphogenetic protein 2 (rhBMP-2) plays a pivotal role in complex spine surgery. Despite its limited approval, the off-label use of rhBMP-2 is prevalent, particularly in transforaminal lumbar interbody fusions (TLIFs). To determine the effectiveness and safety of rhBMP-2 use in TLIF procedures versus autograft. Retrospective cohort study. Patients older than 18 years undergoing spine surgery for lumbar degenerative spine disease at a single academic institution. Clinical outcome was determined according to patient records. Radiographic outcome was determined according to plain X-rays and computed tomography (CT). A retrospective study from 1997 to 2014 was conducted on 191 adults undergoing anterior-posterior instrumented spinal fusion with TLIF at a single academic institution. Patient data were gathered from operative notes, follow-up clinic notes, and imaging studies to determine complications and fusion rates. One hundred eighty-seven patients fit the criteria, which included patients with a minimum of one TLIF, and had a minimum 2-year radiographic and clinical follow-up. Patients were further classified into a BMP group (n=83) or non-BMP group (n=104). Three logistic regression models were run using rhBMP-2 exposure as the independent variable. The respective outcome variables were TLIF-related complications (radiculitis, seroma, osteolysis, and ectopic bone), surgical complications, and all complications. Bone morphogenetic protein (n=83) and non-BMP (n=104) groups had similar baseline demographics (sex, diabetes, pre-existing cancer). On average, the BMP and non-BMP groups were similarly aged (51.9 vs. 47.9 years, p>.05), but the BMP group had a shorter follow-up time (3.03 vs. 4.06 years; p<.001) and fewer smokers (8 vs. 21 patients; p<.048). The fusion rate for the BMP and non-BMP groups was 92.7% and 92.3%, respectively. The pseudoarthrosis rate was 7.5% (14 of 187 patients). Radiculitis was observed in seven patients in the BMP group (8.4%) and two patients in the non-BMP group (1.9%). Seroma was observed in two patients in the BMP group (2.4%) and none in the non-BMP group. No deep infections were observed in the BMP group, and in one patient in the non-BMP group (0.96%). Although patients exposed to BMP were at a significantlygreater risk of developing radiculitis and seroma (odds ratio [OR]=4.53, confidence interval [CI]=1.42-14.5), BMP exposure was not a significant predictor of surgical complications (OR=0.32, CI=0.10-1.00) or overall complications (OR=1.11, CI=0.53-2.34). The outcome of TLIF-related complications was too rare and the confidence interval too wide for practical significance of the first model. Evidence supports the hypothesis that off-label use of rhBMP-2 in TLIF procedures is relatively effective for achieving bone fusion at rates similar to patients receiving autograft. Patients exhibited similar complication rates between the two groups, with the BMP group exhibiting slightly higher rates of radiculitis and seroma. Copyright © 2017 Elsevier Inc. All rights reserved.

The ventralizing activity of Radar, a maternally expressed bone morphogenetic protein, reveals complex bone morphogenetic protein interactions controlling dorso-ventral patterning in zebrafish.

PubMed

Goutel, C; Kishimoto, Y; Schulte-Merker, S; Rosa, F

2000-12-01

In Xenopus and zebrafish, BMP2, 4 and 7 have been implicated, after the onset of zygotic expression, in inducing and maintaining ventro-lateral cell fate during early development. We provide evidence here that a maternally expressed bone morphogenetic protein (BMP), Radar, may control early ventral specification in zebrafish. We show that Radar ventralizes zebrafish embryos and induces the early expression of bmp2b and bmp4. The analysis of Radar overexpression in both swirl/bmp2b mutants and embryos expressing truncated BMP receptors shows that Radar-induced ventralization is dependent on functional BMP2/4 pathways, and may initially rely on an Alk6-related signaling pathway. Finally, we show that while radar-injected swirl embryos still exhibit a strongly dorsalized phenotype, the overexpression of Radar into swirl/bmp2b mutant embryos restores ventral marker expression, including bmp4 expression. Our results suggest that a complex regulation of different BMP pathways controls dorso-ventral (DV) patterning from early cleavage stages until somitogenesis.

In vitro study on the osteogenesis enhancement effect of BMP-2 incorporated biomimetic apatite coating on titanium surfaces.

PubMed

Zhu, Xiaojing; Zhang, Hui; Zhang, Xinchun; Ning, Chengyun; Wang, Yan

2017-09-26

To fabricate a sustained-release delivery system of bone morphogenetic protein (BMP-2) on titanium surface, explore the effect of BMP-2 concentration on the loading/release behavior of BMP-2 and evaluate the cell compatibility of the system in vitro, pure titanium specimens were immersed into supersaturated calcium phosphate solutions (SCP) containing 4 different concentrations of BMP-2: 0, 50, 100, 200 and 400 ng/mL. Biomimetic calcium phosphate coating was formed on titanium surface and BMP-2 was incorporated into the coating through co-deposition. The release profile of BMP-2 suggested that BMP-2 were delivered sustainably up to 20 days. CCK-8 and ALP assay showed that 200 group and 400 ng/mL BMP-2 group have significant effect on promoting MC3T3-E1 cell proliferation and differentiation. The BMP-2 incorporated into the hybrid coating released in a sustained manner and significantly promoted the proliferation and differentiation of MC3T3-E1 on the titanium surface.

Bmp2 Deletion Causes an Amelogenesis Imperfecta Phenotype Via Regulating Enamel Gene Expression

PubMed Central

GUO, FENG; FENG, JUNSHENG; WANG, FENG; LI, WENTONG; GAO, QINGPING; CHEN, ZHUO; SHOFF, LISA; DONLY, KEVIN J.; GLUHAK-HEINRICH, JELICA; CHUN, YONG HEE PATRICIA; HARRIS, STEPHEN E.; MACDOUGALL, MARY; CHEN, SHUO

2015-01-01

Although Bmp2 is essential for tooth formation, the role of Bmp2 during enamel formation remains unknown in vivo. In this study, the role of Bmp2 in regulation of enamel formation was investigated by the Bmp2 conditional knock out (Bmp2 cKO) mice. Teeth of Bmp2 cKO mice displayed severe and profound phenotypes with asymmetric and misshaped incisors as well as abrasion of incisors and molars. Scanning electron microscopy analysis showed that the enamel layer was hypoplastic and enamel lacked a typical prismatic pattern. Teeth from null mice were much more brittle as tested by shear and compressive moduli. Expression of enamel matrix protein genes, amelogenin, enamelin, and enamel-processing proteases, Mmp-20 and Klk4 was reduced in the Bmp2 cKO teeth as reflected in a reduced enamel formation. Exogenous Bmp2 up-regulated those gene expressions in mouse enamel organ epithelial cells. This result for the first time indicates Bmp2 signaling is essential for proper enamel development and mineralization in vivo. PMID:25545831

Bmp2 deletion causes an amelogenesis imperfecta phenotype via regulating enamel gene expression.

PubMed

Guo, Feng; Feng, Junsheng; Wang, Feng; Li, Wentong; Gao, Qingping; Chen, Zhuo; Shoff, Lisa; Donly, Kevin J; Gluhak-Heinrich, Jelica; Chun, Yong Hee Patricia; Harris, Stephen E; MacDougall, Mary; Chen, Shuo

2015-08-01

Although Bmp2 is essential for tooth formation, the role of Bmp2 during enamel formation remains unknown in vivo. In this study, the role of Bmp2 in regulation of enamel formation was investigated by the Bmp2 conditional knock out (Bmp2 cKO) mice. Teeth of Bmp2 cKO mice displayed severe and profound phenotypes with asymmetric and misshaped incisors as well as abrasion of incisors and molars. Scanning electron microscopy analysis showed that the enamel layer was hypoplastic and enamel lacked a typical prismatic pattern. Teeth from null mice were much more brittle as tested by shear and compressive moduli. Expression of enamel matrix protein genes, amelogenin, enamelin, and enamel-processing proteases, Mmp-20 and Klk4 was reduced in the Bmp2 cKO teeth as reflected in a reduced enamel formation. Exogenous Bmp2 up-regulated those gene expressions in mouse enamel organ epithelial cells. This result for the first time indicates Bmp2 signaling is essential for proper enamel development and mineralization in vivo. © 2015 Wiley Periodicals, Inc.

Overexpression of BMP3 in the developing skeleton alters endochondral bone formation resulting in spontaneous rib fractures.

PubMed

Gamer, Laura W; Cox, Karen; Carlo, Joelle M; Rosen, Vicki

2009-09-01

Bone morphogenetic protein-3 (BMP) has been identified as a negative regulator in the skeleton as mice lacking BMP3 have increased bone mass. To further understand how BMP3 mediates bone formation, we created transgenic mice overexpressing BMP3 using the type I collagen promoter. BMP3 transgenic mice displayed spontaneous rib fractures that were first detected at E17.0. The fractures were due to defects in differentiation of the periosteum and late hypertrophic chondrocytes resulting in thinner cortical bone with decreased mineralization. As BMP3 modulates BMP and activin signaling through ActRIIB, we examined the ribs of ActRIIB receptor knockout mice and found they had defects in late chondrogenesis and mineralization similar to BMP3 transgenic mice. These data suggest that BMP3 exerts its effects in the skeleton by altering signaling through ActRIIB in chondrocytes and the periosteum, and this results in defects in bone collar formation and late hypertrophic chondrocyte maturation leading to decreased mineralization and less bone. 2009 Wiley-Liss, Inc.

Addition of BMP-2 or BMP-6 to dexamethasone, ascorbic acid, and β-glycerophosphate may not enhance osteogenic differentiation of human periodontal ligament cells.

PubMed

Khanna-Jain, Rashi; Agata, Hideki; Vuorinen, Annukka; Sándor, George K B; Suuronen, Riitta; Miettinen, Susanna

2010-12-01

This study was designed to investigate the potential merits of the combined use of bone morphogenetic protein (BMP)-2 or BMP-6 and osteogenic supplements (OS) [dexamethasone, ascorbic acid (AA), and β-glycerophosphate] on osteogenic differentiation of periodontal ligament cells (PDLCs). Osteogenic differentiation was evaluated by quantitative alkaline phosphatase (ALP) assay, alizarin red staining, quantitative calcium assay, and the qRT-PCR analysis for the expression of collagen type I, runt-related transcription factor-2, osteopontin (OPN), and osteocalcin in PDLCs. Culture with BMP-2 or BMP-6+AA increased ALP activity of PDLCs, suggesting their osteo-inductive effects. However, longer duration of culture showed neither of the BMPs induced in vitro mineralization. In contrast, OS were able to increase ALP activity and OPN expressions, and also induced in vitro mineralization. The mineralization ability was not enhanced by the addition of BMP-2 or BMP-6. These findings suggest that the addition of BMP-2 or BMP-6 to OS may not enhance an osteogenic differentiation of hPDLCs.

Insulin-like growth factor-1 (IGF-1) enhances the osteogenic activity of bone morphogenetic protein-6 (BMP-6) in vitro and in vivo, and together have a stronger osteogenic effect than when IGF-1 is combined with BMP-2.

PubMed

Rico-Llanos, Gustavo A; Becerra, Jose; Visser, Rick

2017-07-01

Bone morphogenetic protein-2 (BMP-2) is widely used in orthopedic surgery and bone tissue engineering because of its strong osteogenic activity. However, BMP-2 treatments have several drawbacks and many groups are actively exploring alternatives. Since BMP-6 has been demonstrated to be more osteoinductive, its use, either alone or together with other growth factors, might be an interesting option. In this work, we have compared the effect of BMP-2, BMP-6, or insulin-like growth factor-1 (IGF-1), either alone or in combination. Murine preosteoblasts were treated with 15 nM IGF-1 and/or 6 nM BMP-2 or -6 and the expression of osteogenic marker genes, proliferation, and alkaline phosphatase (ALP) activity in vitro were analyzed. The results showed that IGF-1 greatly enhanced the BMP-induced osteogenic differentiation of these cells in general and that the ALP activity in the cultures was higher when the combination was made with BMP-6 than with BMP-2. Furthermore, we tested the osteogenic potential of these treatments in vivo by loading 25 pmoles of IGF-1 and/or 10 pmoles of BMP-2 or -6 onto absorbable collagen sponges and implanting them into an ectopic bone formation model in rats. This study revealed that only BMP-6 was able to induce bone formation at the used dose and that the addition of IGF-1 contributed to an increase of the mineralization in the implants. Hence, the combination of BMP-6 with IGF-1 might be a better alternative than BMP-2 for orthopedic surgery or bone tissue engineering approaches. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 1867-1875, 2017. © 2017 Wiley Periodicals, Inc.

An intermediate level of BMP signaling directly specifies cranial neural crest progenitor cells in zebrafish.

PubMed

Schumacher, Jennifer A; Hashiguchi, Megumi; Nguyen, Vu H; Mullins, Mary C

2011-01-01

The specification of the neural crest progenitor cell (NCPC) population in the early vertebrate embryo requires an elaborate network of signaling pathways, one of which is the Bone Morphogenetic Protein (BMP) pathway. Based on alterations in neural crest gene expression in zebrafish BMP pathway component mutants, we previously proposed a model in which the gastrula BMP morphogen gradient establishes an intermediate level of BMP activity establishing the future NCPC domain. Here, we tested this model and show that an intermediate level of BMP signaling acts directly to specify the NCPC. We quantified the effects of reducing BMP signaling on the number of neural crest cells and show that neural crest cells are significantly increased when BMP signaling is reduced and that this increase is not due to an increase in cell proliferation. In contrast, when BMP signaling is eliminated, NCPC fail to be specified. We modulated BMP signaling levels in BMP pathway mutants with expanded or no NCPCs to demonstrate that an intermediate level of BMP signaling specifies the NCPC. We further investigated the ability of Smad5 to act in a graded fashion by injecting smad5 antisense morpholinos and show that increasing doses first expand the NCPCs and then cause a loss of NCPCs, consistent with Smad5 acting directly in neural crest progenitor specification. Using Western blot analysis, we show that P-Smad5 levels are dose-dependently reduced in smad5 morphants, consistent with an intermediate level of BMP signaling acting through Smad5 to specify the neural crest progenitors. Finally, we performed chimeric analysis to demonstrate for the first time that BMP signal reception is required directly by NCPCs for their specification. Together these results add substantial evidence to a model in which graded BMP signaling acts as a morphogen to pattern the ectoderm, with an intermediate level acting in neural crest specification.

Low-dose rhBMP2/7 heterodimer to reconstruct peri-implant bone defects: a micro-CT evaluation.

PubMed

Wang, Jingxiao; Zheng, Yuanna; Zhao, Juan; Liu, Tie; Gao, Lixia; Gu, Zhiyuan; Wu, Gang

2012-01-01

To delineate the dynamic micro-architectures of bone induced by low-dose bone morphogenetic protein (BMP)-2/7 heterodimer in peri-implant bone defects compared to BMP2 and BMP7 homodimer. Peri-implant bone defects (8 mm in diameter, 4 mm in depth) were created surrounding SLA-treated titanium implants (3.1 mm in diameter, 10 mm in length) in minipig's calvaria. We administrated collagen sponges with adsorbed low-dose (30 ng/mm(3) ) BMP2/7 to treat the defects using BMP2, BMP7 or no BMP as controls.2, 3 and 6 weeks after implantation, we adopted micro-computer tomography to evaluate the micro-architectures of new bone using the following parameters: relative bone volume (BV/TV), trabecular number (Tb.N), trabecular thickness (Tb.Th), trabecular separation (Tb.Sp), connectivity density, and structure mode index (SMI). Bone implant contact (BIC) was also revealed histologically. Consistent with 2 and 3 weeks, after 6 weeks post-operation, BMP2/7 resulted in significantly higher BV/TV (63.033 ± 2.055%) and significantly lower SMI (-4.405 ± 0.500) than BMP2 (BV/TV: 43.133 ± 2.001%; SMI: -0.086 ± 0.041) and BMP7 (BV/TV: 41.467 ± 1.850%; SMI: -0.044 ± 0.016) respectively. Significant differences were also found in Tb.N, Tb.Th and Tb.Sp at all time points. At 2 weeks, BMP2/7 resulted in significantly higher BIC than the controls. Low-dose BMP2/7 heterodimer facilitated more rapid bone regeneration in better quality in peri-implant bone defects than BMP2 and BMP7 homodimers. © 2011 John Wiley & Sons A/S.

Towards eliminating systematic errors caused by the experimental conditions in Biochemical Methane Potential (BMP) tests

DOE Office of Scientific and Technical Information (OSTI.GOV)

Strömberg, Sten, E-mail: sten.stromberg@biotek.lu.se; Nistor, Mihaela, E-mail: mn@bioprocesscontrol.com; Liu, Jing, E-mail: jing.liu@biotek.lu.se

Highlights: • The evaluated factors introduce significant systematic errors (10–38%) in BMP tests. • Ambient temperature (T) has the most substantial impact (∼10%) at low altitude. • Ambient pressure (p) has the most substantial impact (∼68%) at high altitude. • Continuous monitoring of T and p is not necessary for kinetic calculations. - Abstract: The Biochemical Methane Potential (BMP) test is increasingly recognised as a tool for selecting and pricing biomass material for production of biogas. However, the results for the same substrate often differ between laboratories and much work to standardise such tests is still needed. In the currentmore » study, the effects from four environmental factors (i.e. ambient temperature and pressure, water vapour content and initial gas composition of the reactor headspace) on the degradation kinetics and the determined methane potential were evaluated with a 2{sup 4} full factorial design. Four substrates, with different biodegradation profiles, were investigated and the ambient temperature was found to be the most significant contributor to errors in the methane potential. Concerning the kinetics of the process, the environmental factors’ impact on the calculated rate constants was negligible. The impact of the environmental factors on the kinetic parameters and methane potential from performing a BMP test at different geographical locations around the world was simulated by adjusting the data according to the ambient temperature and pressure of some chosen model sites. The largest effect on the methane potential was registered from tests performed at high altitudes due to a low ambient pressure. The results from this study illustrate the importance of considering the environmental factors’ influence on volumetric gas measurement in BMP tests. This is essential to achieve trustworthy and standardised results that can be used by researchers and end users from all over the world.« less

Tbx20 Transcription Factor Is a Downstream Mediator for Bone Morphogenetic Protein-10 in Regulating Cardiac Ventricular Wall Development and Function*

PubMed Central

Zhang, Wenjun; Chen, Hanying; Wang, Yong; Yong, Weidong; Zhu, Wuqiang; Liu, Yunlong; Wagner, Gregory R.; Payne, R. Mark; Field, Loren J.; Xin, Hongbo; Cai, Chen-Leng; Shou, Weinian

2011-01-01

Bone morphogenetic protein 10 (BMP10) belongs to the TGFβ-superfamily. Previously, we had demonstrated that BMP10 is a key regulator for ventricular chamber formation, growth, and maturation. Ablation of BMP10 leads to hypoplastic ventricular wall formation, and elevated levels of BMP10 are associated with abnormal ventricular trabeculation/compaction and wall maturation. However, the molecular mechanism(s) by which BMP10 regulates ventricle wall growth and maturation is still largely unknown. In this study, we sought to identify the specific transcriptional network that is potentially mediated by BMP10. We analyzed and compared the gene expression profiles between α-myosin heavy chain (αMHC)-BMP10 transgenic hearts and nontransgenic littermate controls using Affymetrix mouse exon arrays. T-box 20 (Tbx20), a cardiac transcription factor, was significantly up-regulated in αMHC-BMP10 transgenic hearts, which was validated by quantitative RT-PCR and in situ hybridization. Ablation of BMP10 reduced Tbx20 expression specifically in the BMP10-expressing region of the developing ventricle. In vitro promoter analysis demonstrated that BMP10 was able to induce Tbx20 promoter activity through a conserved Smad binding site in the Tbx20 promoter proximal region. Furthermore, overexpression of Tbx20 in myocardium led to dilated cardiomyopathy that exhibited ventricular hypertrabeculation and an abnormal muscular septum, which phenocopied genetically modified mice with elevated BMP10 levels. Taken together, our findings demonstrate that the BMP10-Tbx20 signaling cascade is important for ventricular wall development and maturation. PMID:21890625

Rapid Activation of Bone Morphogenic Protein 9 by Receptor-mediated Displacement of Pro-domains*

PubMed Central

Kienast, Yvonne; Jucknischke, Ute; Scheiblich, Stefan; Thier, Martina; de Wouters, Mariana; Haas, Alexander; Lehmann, Christian; Brand, Verena; Bernicke, Dirk; Honold, Konrad; Lorenz, Stefan

2016-01-01

By non-covalent association after proteolytic cleavage, the pro-domains modulate the activities of the mature growth factor domains across the transforming growth factor-β family. In the case of bone morphogenic protein 9 (BMP9), however, the pro-domains do not inhibit the bioactivity of the growth factor, and the BMP9·pro-domain complexes have equivalent biological activities as the BMP9 mature ligand dimers. By using real-time surface plasmon resonance, we could demonstrate that either binding of pro-domain-complexed BMP9 to type I receptor activin receptor-like kinase 1 (ALK1), type II receptors, co-receptor endoglin, or to mature BMP9 domain targeting antibodies leads to immediate and complete displacement of the pro-domains from the complex. Vice versa, pro-domain binding by an anti-pro-domain antibody results in release of the mature BMP9 growth factor. Based on these findings, we adjusted ELISA assays to measure the protein levels of different BMP9 variants. Although mature BMP9 and inactive precursor BMP9 protein were directly detectable by ELISA, BMP9·pro-domain complex could only be measured indirectly as dissociated fragments due to displacement of mature growth factor and pro-domains after antibody binding. Our studies provide a model in which BMP9 can be readily activated upon getting into contact with its receptors. This increases the understanding of the underlying biology of BMP9 activation and also provides guidance for ELISA development for the detection of circulating BMP9 variants. PMID:26677222

Combination of Root Surface Modification with BMP-2 and Collagen Hydrogel Scaffold Implantation for Periodontal Healing in Beagle Dogs

PubMed Central

Kato, Akihito; Miyaji, Hirofumi; Ishizuka, Ryosuke; Tokunaga, Keisuke; Inoue, Kana; Kosen, Yuta; Yokoyama, Hiroyuki; Sugaya, Tsutomu; Tanaka, Saori; Sakagami, Ryuji; Kawanami, Masamitsu

2015-01-01

Objective : Biomodification of the root surface plays a major role in periodontal wound healing. Root surface modification with bone morphogenetic protein (BMP) stimulates bone and cementum-like tissue formation; however, severe ankylosis is simultaneously observed. Bio-safe collagen hydrogel scaffolds may therefore be useful for supplying periodontal ligament cells and preventing ankylosis. We examined the effects of BMP modification in conjunction with collagen hydrogel scaffold implantation on periodontal wound healing in dogs. Material and Methods: The collagen hydrogel scaffold was composed of type I collagen sponge and collagen hydrogel. One-wall infrabony defects (5 mm in depth, 3 mm in width) were surgically created in six beagle dogs. In the BMP/Col group, BMP-2 was applied to the root surface (loading dose; 1 µg/µl), and the defects were filled with collagen hydrogel scaffold. In the BMP or Col group, BMP-2 coating or scaffold implantation was performed. Histometric parameters were evaluated at 4 weeks after surgery. Results: Single use of BMP stimulated formation of alveolar bone and ankylosis. In contrast, the BMP/Col group frequently enhanced reconstruction of periodontal attachment including cementum-like tissue, periodontal ligament and alveolar bone. The amount of new periodontal ligament in the BMP/Col group was significantly greater when compared to all other groups. In addition, ankylosis was rarely observed in the BMP/Col group. Conclusion: The combination method using root surface modification with BMP and collagen hydrogel scaffold implantation facilitated the reestablishment of periodontal attachment. BMP-related ankylosis was suppressed by implantation of collagen hydrogel. PMID:25674172

NF-kappaB specifically activates BMP-2 gene expression in growth plate chondrocytes in vivo and in a chondrocyte cell line in vitro.

PubMed

Feng, Jian Q; Xing, Lianping; Zhang, Jiang-Hong; Zhao, Ming; Horn, Diane; Chan, Jeannie; Boyce, Brendan F; Harris, Stephen E; Mundy, Gregory R; Chen, Di

2003-08-01

Bone morphogenetic protein-2 (BMP-2) regulates growth plate chondrogenesis during development and postnatal bone growth, but the control mechanisms of BMP-2 expression in growth plate chondrocytes are unknown. Here we have used both in vitro and in vivo approaches to demonstrate that transcription factor, NF-kappaB, regulates BMP-2 gene expression in chondrocytes. Two putative NF-kappaB response elements were found in the -2712/+165 region of the BMP-2 gene. Cotransfection of mutant I-kappaBalpha expression plasmids with BMP-2 promoter-luciferase reporters into TMC-23 chondrocyte cell line suppressed BMP-2 transcription. Mutations in NF-kappaB response elements in the BMP-2 gene lead to decreases in BMP-2 promoter activity. Electrophoretic mobility shift assay using nuclear extracts from TMC-23 chondrocytic cells revealed that the NF-kappaB subunits p50 and p65 bound to the NF-kappaB response elements of the BMP-2 gene. Thus, NF-kappaB may positively regulate BMP-2 gene transcription. Consistent with these findings, expression of BMP-2 mRNA was significantly reduced in growth plate chondrocytes in NF-kappaB p50/p52 dKO mice, which associated with decreased numbers of 5-bromo-2'-deoxyuridine (BrdUrd)-positive cells in the proliferating zone of growth plate in these mice. Therefore, in postnatal growth plate chondrocytes, expression of BMP-2 is regulated by NF-kappaB, which may play an important role in chondrogenesis.

Dragon enhances BMP signaling and increases transepithelial resistance in kidney epithelial cells.

PubMed

Xia, Yin; Babitt, Jodie L; Bouley, Richard; Zhang, Ying; Da Silva, Nicolas; Chen, Shanzhuo; Zhuang, Zhenjie; Samad, Tarek A; Brenner, Gary J; Anderson, Jennifer L; Hong, Charles C; Schneyer, Alan L; Brown, Dennis; Lin, Herbert Y

2010-04-01

The neuronal adhesion protein Dragon acts as a bone morphogenetic protein (BMP) coreceptor that enhances BMP signaling. Given the importance of BMP signaling in nephrogenesis and its putative role in the response to injury in the adult kidney, we studied the localization and function of Dragon in the kidney. We observed that Dragon localized predominantly to the apical surfaces of tubular epithelial cells in the thick ascending limbs, distal convoluted tubules, and collecting ducts of mice. Dragon expression was weak in the proximal tubules and glomeruli. In mouse inner medullary collecting duct (mIMCD3) cells, Dragon generated BMP signals in a ligand-dependent manner, and BMP4 is the predominant endogenous ligand for the Dragon coreceptor. In mIMCD3 cells, BMP4 normally signaled through BMPRII, but Dragon enhanced its signaling through the BMP type II receptor ActRIIA. Dragon and BMP4 increased transepithelial resistance (TER) through the Smad1/5/8 pathway. In epithelial cells isolated from the proximal tubule and intercalated cells of collecting ducts, we observed coexpression of ActRIIA, Dragon, and BMP4 but not BMPRII. Taken together, these results suggest that Dragon may enhance BMP signaling in renal tubular epithelial cells and maintain normal renal physiology.

Poly(lactic-co-glycolide) polymer constructs cross-linked with human BMP-6 and VEGF protein significantly enhance rat mandible defect repair.

PubMed

Das, Anusuya; Fishero, Brian A; Christophel, J Jared; Li, Ching-Ju; Kohli, Nikita; Lin, Yong; Dighe, Abhijit S; Cui, Quanjun

2016-04-01

We have previously shown that the combined delivery of mesenchymal stem cells (MSCs), vascular endothelial growth factor (VEGF) and bone morphogenetic protein 6 (BMP-6) induces significantly more bone formation than that induced by the delivery of any single factor or a combination of any two factors. We now determine whether the exogenous addition of VEGF and BMP-6 is sufficient for bone healing when MSCs are not provided. Poly(lactic-co-glycolic acid) (PLAGA) microsphere-based three-dimensional scaffolds (P) were fabricated by thermal sintering of PLAGA microspheres. The scaffolds were chemically cross-linked with 200 ng recombinant human VEGF (P(VEGF)) or BMP-6 (P(BMP-6)) or both (P(VEGF+BMP-6)) by the EDC-NHS-MES method. Release of the proteins from the scaffolds was detected for 21 days in vitro which confirmed their comparable potential to supply the proteins in vivo. The scaffolds were delivered to a critical-sized mandibular defect created in 32 Sprague Dawley rats. Significant bone regeneration was observed only in rats with P(VEGF+BMP-6) scaffolds at weeks 2, 8 and 12 as revealed by micro-computer tomography. Vascular ingrowth was higher in the P(VEGF+BMP-6) group as seen by microfil imaging than in other groups. Trichrome staining revealed that a soft callus formed in P(VEGF), P(BMP-6) and P(VEGF+BMP-6) but not in P. MSCs isolated from rat femurs displayed expression of the bone-specific marker osteocalcin when cultured with P(VEGF), P(BMP-6), or P(VEGF+BMP-6) but not with P. Robust mineralization and increased alkaline phosphatase gene expression were seen in rat MSCs when cultured on P(VEGF+BMP-6) but not on P, P(VEGF), or P(BMP-6). Thus, unlike the delivery of VEGF or BMP-6 alone, the combined delivery of VEGF and BMP-6 to the bone defect significantly enhanced bone repair through the enhancement of angiogenesis and the differentiation of endogenously recruited MSCs into the bone repair site.

[Experimental study of the effect of new bone formation on new type artificial bone composed of bioactive ceramics].

PubMed

Zhu, Minghua; Zeng, Yi; Sun, Tao; Peng, Qiang

2005-03-15

To investigate the osteogenic potential of four kinds of new bioactive ceramics combined with bovine bone morphogenetic proteins (BMP) and to explore the feasibility of using compounds as bone substitute material. Ninety-six rats were divided into 4 groups (24 in each group). BMP was combined with hydroxyapatite (HA), tricalcium phosphate (TCP), fluoridated-HA (FHA), and collagen-HA(CHA) respectively. The left thighs of the rats implanted with HA/BMP, TCP/BMP, FHA/BMP, and CHA/BMP were used as experimental groups. The right thighs of the rats implanted with HA, TCP, CHA, and decalcified dentin matrix (DDM) were used as control groups. The rats were sacrificed 1, 3, 5 and 7 weeks after implantation and bone induction was estimated by alkaline phosphatase (ALP), phosphorus (P), and total protein (TP) measurement. The histological observation and electronic microscope scanning of the implants were also made. The cartilage growth in the 4 experimental groups and the control group implanted with DDM was observed 1 week after operation and fibrous connective tissues were observed in the other 3 control groups. 3 weeks after implantation, lamellar bone with bone marrow and positive reaction in ALP stain were observed in the 4 experimental groups. No bone formation or positive reaction in ALP stain were observed in the control groups. The amount of ALP activity, P value, and new bone formation in the experimental groups were higher than those in the control group(P < 0.05). The amount of ALP activity, P value, and new bone formation in TCP/BMP group were higher than those in HA/BMP, CHA/BMP and FHA/BMP groups (P < 0.05). There was no significant difference in TP between the BMP treatment group and the control groups. From 5th to 7th week, new bone formation, histochemistry evaluation, and the level of ALP, P, TP value were as high as those in the 3rd week. New composite artificial bone of TCP/BMP, HA/BMP, CHA/BMP, and FHA/BMP all prove to be effective, but TCP/BMP is the most effective so that it is the most suitable biomaterial replacement of tissue.

Immunohistochemical localization of bone morphogenetic proteins and the receptors in epiphyseal growth plate.

PubMed

Yazaki, Y; Matsunaga, S; Onishi, T; Nagamine, T; Origuchi, N; Yamamoto, T; Ishidou, Y; Imamura, T; Sakou, T

1998-01-01

The expression of bone morphogenetic proteins (BMPs) and BMP receptors (BMPRs) in the epiphyseal growth plate has not been clarified. In this study, we studied immunohistochemically the spatial and temporal localization of BMP-2/4, osteogenic protein-1 (OP-1, or BMP-7), and BMP receptors (BMPR-IA, BMPR-IB, and BMPR-II) in the epiphyseal plate of growing rats. The proximal parts of tibia in growing rats were observed. At 12 weeks after birth, BMP-2/4 and OP-1 were expressed markedly in proliferating and maturing chondrocytes. BMPR-IA, IB and II were clearly co-expressed in proliferating and maturing chondrocytes, and the expression was decreased in hypertrophic chondrocytes. At 24 weeks, the expression of BMP-2/4 and OP-1 was decreased, but BMPRs were still well-expressed in proliferating chondrocytes. The temporal and spatial expression of BMP and BMPR suggests that BMP and BMP receptors play roles in the multistep cascade of enchondral ossification in the epiphyseal growth plate.

Notch regulates BMP responsiveness and lateral branching in vessel networks via SMAD6

PubMed Central

Mouillesseaux, Kevin P.; Wiley, David S.; Saunders, Lauren M.; Wylie, Lyndsay A.; Kushner, Erich J.; Chong, Diana C.; Citrin, Kathryn M.; Barber, Andrew T.; Park, Youngsook; Kim, Jun-Dae; Samsa, Leigh Ann; Kim, Jongmin; Liu, Jiandong; Jin, Suk-Won; Bautch, Victoria L.

2016-01-01

Functional blood vessel growth depends on generation of distinct but coordinated responses from endothelial cells. Bone morphogenetic proteins (BMP), part of the TGFβ superfamily, bind receptors to induce phosphorylation and nuclear translocation of SMAD transcription factors (R-SMAD1/5/8) and regulate vessel growth. However, SMAD1/5/8 signalling results in both pro- and anti-angiogenic outputs, highlighting a poor understanding of the complexities of BMP signalling in the vasculature. Here we show that BMP6 and BMP2 ligands are pro-angiogenic in vitro and in vivo, and that lateral vessel branching requires threshold levels of R-SMAD phosphorylation. Endothelial cell responsiveness to these pro-angiogenic BMP ligands is regulated by Notch status and Notch sets responsiveness by regulating a cell-intrinsic BMP inhibitor, SMAD6, which affects BMP responses upstream of target gene expression. Thus, we reveal a paradigm for Notch-dependent regulation of angiogenesis: Notch regulates SMAD6 expression to affect BMP responsiveness of endothelial cells and new vessel branch formation. PMID:27834400

Strontium attenuates rhBMP-2-induced osteogenic differentiation via formation of Sr-rhBMP-2 complex and suppression of Smad-dependent signaling pathway.

PubMed

Zhang, Wenjing; Tian, Yu; He, Hongyan; Chen, Rui; Ma, Yifan; Guo, Han; Yuan, Yuan; Liu, Changsheng

2016-03-01

Strontium (Sr(2+)) has pronounced effects on stimulating bone formation and inhibiting bone resorption in bone regeneration. In this current study, the effect and the underlying mechanism involved of Sr(2+) on the biological activity of bone morphogenetic protein-2 (BMP-2) were studied in detail with pluripotent skeletal muscle myogenic progenitor C2C12 model cell line. The results indicated that Sr(2+) could bind recombinant human BMP-2 (rhBMP-2) rapidly, even in the presence of Ca(2+) and Mg(2+), and inhibited rhBMP-2-induced osteogenic differentiation in vitro and osteogenetic efficiency in vivo. Further studies demonstrated that Sr(2+) treatment undermined the binding capacity of rhBMP-2 with its receptor BMPRIA and thus attenuated Smad 1/5/8 phosphorylation without affecting their dephosphorylation in C2C12 cells. Furthermore, circular dichroism spectroscopy, fluorescence spectroscopy and X-ray photoelectron spectroscopy all revealed that the inhibitory effect of Sr(2+) on the rhBMP-2 osteogenic activity was associated with the formation of Sr-rhBMP-2 complex and ensuing enhancement of β-sheet structure. Our work suggests the activity of rhBMP-2 to induce osteogenic differentiation was decreased by directly interaction with free Sr ions in solution, which should provide guide and assist for development of BMP-2-based materials for bone regeneration. Due to easy denaturation and ensuing the reduced activity of rhBMP-2, preserving/enhancing the capacity of rhBMP-2 to induce osteogenic differentiation is of critical importance in developing the protein-based therapy. Cations as effective elements influence the conformation and thereby the bioactivity of protein. Strontium (Sr(2+)), stimulating bone formation and inhibiting bone resorption, has been incorporated into biomaterials/scaffold to improve the bioactivity for bone-regeneration applications. However, Sr(2+)-induced changes in the conformation and bioactivity of BMP-2 have never been investigated. In this study, the formation of Sr-rhBMP-2 complex inhibited the osteogenic differentiation in vitro and osteogenetic efficiency in vivo through the inhibition of BMP/Smad signaling pathway, providing guidance for development of Sr-containing BMP-2-based bone scaffold/matrice and other Sr-dopped protein therapy. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Addition of bone morphogenetic protein type 2 to ascorbate and β-glycerophosphate supplementation did not enhance osteogenic differentiation of human adipose-derived stem cells

PubMed Central

CRUZ, Ariadne Cristiane Cabral; SILVA, Mariana Lúcia; CAON, Thiago; SIMÕES, Cláudia Maria Oliveira

2012-01-01

Bone morphogenetic protein type 2 (BMP-2) is a potent local factor, which promotes bone formation and has been used as an osteogenic supplement for mesenchymal stem cells. Objectives This study evaluated the effect of a recombinant BMP-2 as well as the endogenous BMP-4 and BMP-7 in the osteogenic differentiation of adipose-derived stem cells (ASCs) in medium supplemented with ascorbate and β-glycerophosphate. Material and Methods Human ASCs were treated with osteogenic medium in the presence (ASCs+OM+BMP-2) or absence (ASCs+OM) of BMP-2. The alkaline phosphatase (ALP) activity was determined and the extracellular matrix mineralization was evaluated by Von Kossa staining and calcium quantification. The expressions of BMP-4, BMP-7, Smad1, Smad4, and phosphorylated Smad1/5/8 were analyzed by western blotting. Relative mRNA expressions of Smad1, BMP receptor type II (BMPR-II), osteonectin, and osteocalcin were evaluated by qPCR. Results: ASCs+OM demonstrated the highest expression of BMP-4 and BMP-7 at days 21 and 7, respectively, the highest levels of BMPR-II mRNA expression at day 28, and the highest levels of Smad1 mRNA at days 14 and 28. ASCs+OM+BMP-2 demonstrated the highest levels of Smad1 mRNA expression at days 1, 7, and 21, the highest expression of Smad1 at day 7, the highest expression of Smad4 at day 14, the highest ALP activity at days 14 and 21, and expression of phosphorylated Smad1/5/8 at day 7. ASCs+OM and ASCs+OM+BMP2 showed similar ALP activity at days 7 and 28, similar osteonectin and osteocalcin mRNA expression at all time periods, and similar calcium depositions at all time periods. Conclusions We concluded that human ASCs expressed endogenous BMP-4 and BMP-7. Moreover, the supplementation of ASCs with BMP-2 did not increase the level of osteogenic markers in the initial (ALP activity), intermediate (osteonectin and osteocalcin), or final (calcium deposition) phases, suggesting that the exogenous addition of BMP-2 did not improve the in vitro osteogenesis process of human ASCs. PMID:23329244

BMP9 inhibits the bone metastasis of breast cancer cells by downregulating CCN2 (connective tissue growth factor, CTGF) expression.

PubMed

Ren, Wei; Sun, Xiaoxiao; Wang, Ke; Feng, Honglei; Liu, Yuehong; Fei, Chang; Wan, Shaoheng; Wang, Wei; Luo, Jinyong; Shi, Qiong; Tang, Min; Zuo, Guowei; Weng, Yaguang; He, Tongchuan; Zhang, Yan

2014-03-01

Bone morphogenetic proteins (BMPs), which belong to the transforming growth factor-β superfamily, regulate a wide range of cellular responses including cell proliferation, differentiation, adhesion, migration, and apoptosis. BMP9, the latest BMP to be discovered, is reportedly expressed in a variety of human carcinoma cell lines, but the role of BMP9 in breast cancer has not been fully clarified. In a previous study, BMP9 was found to inhibit the growth, migration, and invasiveness of MDA-MB-231 breast cancer cells. In the current study, the effect of BMP9 on the bone metastasis of breast cancer cells was investigated. After absent or low expression of BMP9 was detected in the MDA-MB-231 breast cancer cells and breast non-tumor adjacent tissues using Western blot and immunohistochemistry, In our previous study, BMP9 could inhibit the proliferation and invasiveness of breast cancer cells MDA-MB-231 in vitro and in vivo. This paper shows that BMP9 inhibit the bone metastasis of breast cancer cells by activating the BMP/Smad signaling pathway and downregulating connective tissue growth factor (CTGF); however, when CTGF expression was maintained, the inhibitory effect of BMP9 on the MDA-MB-231 cells was abolished. Together, these observations indicate that BMP9 is an important mediator of breast cancer bone metastasis and a potential therapeutic target for treating this deadly disease.

Signaling by Bone Morphogenetic Proteins directs formation of an ectodermal signaling center that regulates craniofacial development

PubMed Central

Foppiano, Silvia; Hu, Diane; Marcucio, Ralph S.

2008-01-01

We previously described a signaling center, the Frontonasal Ectodermal Zone (FEZ) that regulates growth and patterning of the frontonasal process (FNP). The FEZ is comprised of FNP ectoderm flanking a boundary between Sonic hedgehog (Shh) and Fibroblast growth factor 8 (Fgf8) expression domains. Our objective was to examine BMP signaling during formation of the FEZ. We blocked BMP signaling throughout the FNP prior to FEZ formation by infecting chick embryos at stage 10 (HH10) with a replication competent avian retrovirus encoding the BMP antagonist Noggin. We assessed gene expression patterns in the FNP 72 hours after infection (~HH22) and observed that Shh expression was reduced or absent. In the mesenchyme we observed that Bmp2 transcripts were absent while the Bmp4 expression domain was expanded proximally. In addition to the molecular changes, infected embryos also exhibited facial malformations at 72 and 96 hours after infection suggesting that the FEZ did not form. Our data indicate that reduced cell proliferation, but not apoptosis, in the mesenchyme contributed to the phenotype that we observed. Additionally, adding exogenous SHH into the mesenchyme of RCAS-Noggin infected embryos did not restore Bmp2 and Bmp4 to a normal pattern of expression. These data indicate that BMP signaling mediates interactions between tissues in the FNP that regulate FEZ formation; and that the correct pattern of Bmp2 and Bmp4, but not Bmp7, expression in the FNP mesenchyme requires signaling by the BMP pathway. PMID:18028903

In vitro and in vivo protein release and anti-ischemia/reperfusion injury properties of bone morphogenetic protein-2-loaded glycyrrhetinic acid-poly(ethylene glycol)-b-poly(l-lysine) nanoparticles

PubMed Central

Shan, Fang; Liu, YuJuan; Jiang, Haiying; Tong, Fei

2017-01-01

Here, we describe a bone morphogenetic protein-2 (BMP-2) nanocarrier based on glycyrrhetinic acid (GA)-poly(ethylene glycol) (PEG)-b-poly(l-lysine) (PLL). A protein nanocarrier was synthesized, characterized and evaluated as a BMP-2 delivery system. The designed nanocarrier was synthesized based on the ring-opening polymerization of amino acid N-carboxyanhydride. The final product was measured with 1H nuclear magnetic resonance. GA-PEG-b-PLL nanocarrier could combine with BMP-2 through electrostatic interaction to form polyion complex (PIC) micelles. BMP-2 could be rapidly and efficiently encapsulated through the GA-PEG-b-PLL nanocarrier under physiological conditions, exhibiting efficient encapsulation and sustained release. In addition, the GA-PEG-b-PLL-mediated BMP-2 delivery system could target the liver against hepatic diseases as it has GA-binding receptors. The anti-hepatic ischemia/reperfusion injury (anti-HI/RI) effect of BMP-2/GA-PEG-b-PLL PIC micelles was investigated in rats using free BMP-2 and BMP-2/PEG-b-PLL PIC micelles as controls, and the results showed that BMP-2/GA-PEG-b-PLL PIC micelles indicated significantly enhanced anti-HI/RI property compared to BMP-2 and BMP-2/PEG-b-PLL. All results suggested that GA-PEG-b-PLL could be used as a potential BMP-2 nanocarrier. PMID:29089759

Evaluation of agricultural best-management practices in the Conestoga River headwaters, Pennsylvania; effects of nutrient management on quality of surface runoff at a small carbonate-rock site near Ephrate, Pennsylvania, 1984-90

USGS Publications Warehouse

Hall, D.W.; Lietman, P.L.; Koerkle, E.J.

1997-01-01

The U.S. Geological Survey and the Pennsylvania Department of Environmental Protection conducted a study from 1984 to 1990 to determine theeffects of the implementation and practice of nutrient management [an agricultural best-management practice (BMP)] on the quality of surface runoff and ground water at a 55-acre crop and livestock farm in carbonate terrain nearEphrata, Pa. Implementation of nutrient management at Field-Site 2 resulted in application decreases of 33 percent for nitrogen and 29 percent for phosphorus. There wereno significant changes in nitrogen or phosphorusloads for a given amount of runoff from the pre-BMP to the post-BMP periods. However, less than 2 percent of the applied nutrients weredischarged with runoff throughout the study period.After the implementation of nutrient management, statistically significant decreases in concentrations of nitrate in ground-water samples occurred at threeof the four wells monitored throughout the pre- and post-BMP periods. The largest decreases in nitrate concentrations occurred at wells where samples hadthe largest nitrate concentrations prior to nutrient management. Changes in nitrogen applications to the contributing areas of five wells were correlated with nitrate concentrations of the well water. The correlations between the timing and amount of applied nitrogen and changes in ground-water quality met the four conditions that are characteristic of a cause-effect relation: an association, consistency, responsiveness, and a mechanism. Changes in ground-water nitrate concentrations lagged behind changes in loading of nitrogen fertilizers (primarily manure) by approximately 4 to 19 months.

Local administration of bone morphogenetic protein-2 and bisphosphonate during non-weight-bearing treatment of ischemic osteonecrosis of the femoral head: an experimental investigation in immature pigs.

PubMed

Kim, Harry K W; Aruwajoye, Olumide; Du, Justin; Kamiya, Nobuhiro

2014-09-17

Non-weight-bearing decreases the femoral head deformity but increases bone resorption without increasing bone formation in an experimental animal model of Legg-Calvé-Perthes disease. We sought to determine if local administration of bone morphogenetic protein (BMP)-2 with or without bisphosphonate can increase the bone formation during the non-weight-bearing treatment in the large animal model of Legg-Calvé-Perthes disease. Eighteen piglets were surgically induced with femoral head ischemia. Immediately following the surgery, all animals received an above-the-knee amputation to enforce local non-weight-bearing (NWB). One to two weeks later, six animals received local BMP-2 to the necrotic head (BMP group), six received local BMP-2 and ibandronate (BMP+IB group), and the remaining six received no treatment (NWB group). All animals were killed at eight weeks after the induction of ischemia. Radiographic, microcomputed tomography (micro-CT), and histomorphometric assessments were performed. Radiographic assessment showed that the femoral heads in the NWB, BMP, and BMP+IB groups had a decrease of 20%, 14%, and 10%, respectively, in their mean epiphyseal quotient in comparison with the normal control group. Micro-CT analyses showed significantly higher femoral head bone volume in the BMP+IB group than in the BMP group (p = 0.02) and the NWB group (p < 0.001). BMP+IB and BMP groups had a significantly higher trabecular number (p < 0.01) and lower trabecular separation (p < 0.02) than the NWB group. In addition, the osteoclast number per bone surface was significantly lower in the BMP+IB group compared with the NWB group. Calcein labeling showed significantly higher bone formation in the BMP and BMP+IB groups than in the NWB group (p < 0.05). Heterotopic ossification was found in the capsule of four hips in the BMP+IB group but not in the BMP group. Administration of BMP-2 with bisphosphonate best decreased bone resorption and increased new bone formation during non-weight-bearing treatment of ischemic osteonecrosis in a pig model, but heterotopic ossification is a concern. This preclinical study provides new evidence that BMP-2 with bisphosphonate can effectively prevent the extreme bone loss associated with the non-weight-bearing treatment and increase new bone formation in the femoral head in this animal model of ischemic osteonecrosis. Copyright © 2014 by The Journal of Bone and Joint Surgery, Incorporated.

Turning Bone Morphogenetic Protein 2 (BMP2) On and Off in Mesenchymal Cells†

PubMed Central

Rogers, Melissa B.; Shah, Tapan A.; Shaikh, Nadia N.

2016-01-01

The concentration, location, and timing of bone morphogenetic protein 2 (BMP2, HGNC:1069, GeneID: 650) gene expression must be precisely regulated. Abnormal BMP2 levels cause congenital anomalies and diseases involving the mesenchymal cells that differentiate into muscle, fat, cartilage, and bone. The molecules and conditions that influence BMP2 synthesis are diverse. Understandably, complex mechanisms control Bmp2 gene expression. This review includes a compilation of agents and conditions that can induce Bmp2. The currently known trans-regulatory factors and cis-regulatory elements that modulate Bmp2 expression are summarized and discussed. This article is protected by copyright. All rights reserved PMID:25776852

The prodomain of BMP4 is necessary and sufficient to generate stable BMP4/7 heterodimers with enhanced bioactivity in vivo.

PubMed

Neugebauer, Judith M; Kwon, Sunjong; Kim, Hyung-Seok; Donley, Nathan; Tilak, Anup; Sopory, Shailaja; Christian, Jan L

2015-05-05

Bone morphogenetic proteins 4 and 7 (BMP4 and BMP7) are morphogens that signal as either homodimers or heterodimers to regulate embryonic development and adult homeostasis. BMP4/7 heterodimers exhibit markedly higher signaling activity than either homodimer, but the mechanism underlying the enhanced activity is unknown. BMPs are synthesized as inactive precursors that dimerize and are then cleaved to generate both the bioactive ligand and prodomain fragments, which lack signaling activity. Our study reveals a previously unknown requirement for the BMP4 prodomain in promoting heterodimer activity. We show that BMP4 and BMP7 precursor proteins preferentially or exclusively form heterodimers when coexpressed in vivo. In addition, we show that the BMP4 prodomain is both necessary and sufficient for generation of stable heterodimeric ligands with enhanced activity and can enable homodimers to signal in a context in which they normally lack activity. Our results suggest that intrinsic properties of the BMP4 prodomain contribute to the relative bioactivities of homodimers versus heterodimers in vivo. These findings have clinical implications for the use of BMPs as regenerative agents for the treatment of bone injury and disease.

Comparison of Two Porcine Collagen Membranes Combined with rhBMP-2 and rhBMP-9 on Osteoblast Behavior In Vitro.

PubMed

Fujioka-Kobayashi, Masako; Schaler, Benoit; Shirakata, Yoshinori; Nakamura, Toshiaki; Noguchi, Kazuyuki; Zhang, Yufeng; Miron, Richard J

To investigate the bone-inducing properties of two types of collagen membranes in combination with recombinant human bone morphogenetic protein (rhBMP)-2 and rhBMP-9 on osteoblast behavior. Porcine pericardium collagen membranes (PPCM) and porcine dermis-derived collagen membranes (PDCM) were coated with either rhBMP-2 or rhBMP-9. The adsorption and release abilities were first investigated via enzyme-linked immunosorbent assay up to 10 days. Moreover, murine bone stromal ST2 cell adhesion, proliferation, and osteoblast differentiation were assessed by MTS assay; real-time polymerase chain reaction for genes encoding runt-related transcription factor 2 (Runx2); alkaline phosphatase (ALP); and osteocalcin, ALP assay, and alizarin red staining. Both rhBMP-2 and rhBMP-9 adsorbed to collagen membranes and were gradually released over time up to 10 days. PPCM showed significantly less cell attachment, whereas PDCM demonstrated comparable cell attachment with the control tissue culture plastic at 8 hours. While both rhBMPs were shown not to affect cell proliferation, collagen membranes combined with rhBMP-9 significantly increased ALP activity at 7 days and ALP mRNA levels at either 3 or 14 days compared with the control tissue culture plastic. Furthermore, rhBMP-9 increased osteocalcin mRNA levels and alizarin red staining at 14 days compared with the control tissue culture plastic. The results from this study suggest that both porcine-derived collagen membranes combined with rhBMP-9 accelerated the osteopromotive potential of ST2 cells. Interestingly, rhBMP-9 demonstrated additional osteogenic differentiation compared with rhBMP-2 and may serve as a suitable growth factor for future clinical use.

Continuous release of bone morphogenetic protein-2 through nano-graphene oxide-based delivery influences the activation of the NF-κB signal transduction pathway

PubMed Central

Zhong, Cheng; Feng, Jun; Lin, Xiangjin; Bao, Qi

2017-01-01

Graphene oxide (GO) has been used as a delivery vehicle for small molecule drugs and nucleotides. To further investigate GO as a smart biomaterial for the controlled release of cargo molecules, we hypothesized that GO may be an appropriate delivery vehicle because it releases bone morphogenetic protein 2 (BMP2). GO characterization indicated that the size distribution of the GO flakes ranged from 81.1 nm to 45,749.7 nm, with an approximate thickness of 2 nm. After BMP2 adsorption onto GO, Fourier-transformed infrared spectroscopy (FTIR) and thermal gravimetric analysis were performed. Compared to GO, BMP2-GO did not induce significant changes in the characteristics of the materials. GO continuously released BMP2 for at least 40 days. Bone marrow stem cells (BMSCs) and chondrocytes were treated with BMP2-GO in interleukin-1 media and assessed in terms of cell viability, flow cytometric characterization, and expression of particular mRNA. Compared to GO, BMP2-GO did not induce any significant changes in biocompatibility. We treated osteoarthritic rats with BMP2 and BMP2-GO, which showed significant differences in Osteoarthritis Research Society International (OARSI) scores (P<0.05). Quantitative assessment revealed significant differences compared to that using BMP2 and BMP2-GO (P<0.05). These findings indicate that GO may be potentially used to control the release of carrier materials. The combination of BMP2 and GO slowed the progression of NF-κB-activated degenerative changes in osteoarthritis. Therefore, we infer that our BMP2-GO strategy could alleviate the NF-κB pathway by inducing continuous BMP2 release. PMID:28243085

Growth differentiation factor 3 is induced by bone morphogenetic protein 6 (BMP-6) and BMP-7 and increases luteinizing hormone receptor messenger RNA expression in human granulosa cells.

PubMed

Shi, Jia; Yoshino, Osamu; Osuga, Yutaka; Akiyama, Ikumi; Harada, Miyuki; Koga, Kaori; Fujimoto, Akihisa; Yano, Tetsu; Taketani, Yuji

2012-04-01

To examine the relevance of growth differentiation factor 3 (GDF-3) and bone morphogenetic protein (BMP) cytokines in human ovary. Molecular studies. Research laboratory. Eight women undergoing salpingo-oophorectomy and 30 women undergoing ovarian stimulation for in vitro fertilization. Localizing GDF-3 protein in human ovaries; granulosa cells (GC) cultured with GDF-3, BMP-6, or BMP-7 followed by RNA extraction. The localization of GDF-3 protein in normal human ovaries via immunohistochemical analysis, GDF-3 messenger RNA (mRNA) expression evaluation via quantitative real-time reverse transcription and polymerase chain reaction (RT-PCR), and evaluation of the effect of GDF-3 on leuteinizing hormone (LH) receptor mRNA expression via quantitative real-time RT-PCR. In the ovary, BMP cytokines, of the transforming growth factor beta (TGF-β) superfamily, are known as a luteinization inhibitor by suppressing LH receptor expression in GC. Growth differentiation factor 3, a TGF-β superfamily cytokine, is recognized as an inhibitor of BMP cytokines in other cells. Immunohistochemical analysis showed that GDF-3 was strongly detected in the GC of antral follicles. An in vitro assay revealed that BMP-6 or BMP-7 induced GDF-3 mRNA in GC. Also, GDF-3 increased LH receptor mRNA expression and inhibited the effect of BMP-7, which suppressed the LH receptor mRNA expression in GC. GDF-3, induced with BMP-6 and BMP-7, might play a role in folliculogenesis by inhibiting the effect of BMP cytokines. Copyright © 2012 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Inductive specification and axonal orientation of spinal neurons mediated by divergent bone morphogenetic protein signaling pathways

PubMed Central

2011-01-01

Background Bone morphogenetic protein (BMP)7 evokes both inductive and axon orienting responses in dorsal interneurons (dI neurons) in the developing spinal cord. These events occur sequentially during the development of spinal neurons but in these and other cell types such inductive and acute chemotactic responses occur concurrently, highlighting the requirement for divergent intracellular signaling. Both type I and type II BMP receptor subtypes have been implicated selectively in orienting responses but it remains unclear how, in a given cell, divergence occurs. We have examined the mechanisms by which disparate BMP7 activities are generated in dorsal spinal neurons. Results We show that widely different threshold concentrations of BMP7 are required to elicit the divergent inductive and axon orienting responses. Type I BMP receptor kinase activity is required for activation of pSmad signaling and induction of dI character by BMP7, a high threshold response. In contrast, neither type I BMP receptor kinase activity nor Smad1/5/8 phosphorylation is involved in the low threshold orienting responses of dI axons to BMP7. Instead, BMP7-evoked axonal repulsion and growth cone collapse are dependent on phosphoinositide-3-kinase (PI3K) activation, plausibly through type II receptor signaling. BMP7 stimulates PI3K-dependent signaling in dI neurons. BMP6, which evokes neural induction but does not have orienting activity, activates Smad signaling but does not stimulate PI3K. Conclusions Divergent signaling through pSmad-dependent and PI3K-dependent (Smad-independent) mechanisms mediates the inductive and orienting responses of dI neurons to BMP7. A model is proposed whereby selective engagement of BMP receptor subunits underlies choice of signaling pathway. PMID:22085733

BMP9 and BMP10 are necessary for proper closure of the ductus arteriosus

PubMed Central

Levet, Sandrine; Ouarné, Marie; Ciais, Delphine; Coutton, Charles; Subileau, Mariela; Mallet, Christine; Ricard, Nicolas; Bidart, Marie; Debillon, Thierry; Faravelli, Francesca; Rooryck, Caroline; Feige, Jean-Jacques; Tillet, Emmanuelle; Bailly, Sabine

2015-01-01

The transition to pulmonary respiration after birth requires rapid alterations in the structure of the mammalian cardiovascular system. One dramatic change that occurs is the closure of the ductus arteriosus (DA), an arterial connection in the fetus that directs blood flow away from the pulmonary circulation. Two members of the TGFβ family, bone morphogenetic protein 9 (BMP9) and BMP10, have been recently involved in postnatal angiogenesis, both being necessary for remodeling of newly formed microvascular beds. The aim of the present work was to study whether BMP9 and BMP10 could be involved in closure of the DA. We found that Bmp9 knockout in mice led to an imperfect closure of the DA. Further, addition of a neutralizing anti-BMP10 antibody at postnatal day 1 (P1) and P3 in these pups exacerbated the remodeling defect and led to a reopening of the DA at P4. Transmission electron microscopy images and immunofluorescence stainings suggested that this effect could be due to a defect in intimal cell differentiation from endothelial to mesenchymal cells, associated with a lack of extracellular matrix deposition within the center of the DA. This result was supported by the identification of the regulation by BMP9 and BMP10 of several genes known to be involved in this process. The involvement of these BMPs was further supported by human genomic data because we could define a critical region in chromosome 2 encoding eight genes including BMP10 that correlated with the presence of a patent DA. Together, these data establish roles for BMP9 and BMP10 in DA closure. PMID:26056270

Identifying biogeochemical processes beneath stormwater infiltration ponds in support of a new best management practice for groundwater protection

USGS Publications Warehouse

O'Reilly, Andrew M.; Chang, Ni-Bin; Wanielista, Martin P.; Xuan, Zhemin; Schirmer, Mario; Hoehn, Eduard; Vogt, Tobias

2011-01-01

 When applying a stormwater infiltration pond best management practice (BMP) for protecting the quality of underlying groundwater, a common constituent of concern is nitrate. Two stormwater infiltration ponds, the SO and HT ponds, in central Florida, USA, were monitored. A temporal succession of biogeochemical processes was identified beneath the SO pond, including oxygen reduction, denitrification, manganese and iron reduction, and methanogenesis. In contrast, aerobic conditions persisted beneath the HT pond, resulting in nitrate leaching into groundwater. Biogeochemical differences likely are related to soil textural and hydraulic properties that control surface/subsurface oxygen exchange. A new infiltration BMP was developed and a full-scale application was implemented for the HT pond. Preliminary results indicate reductions in nitrate concentration exceeding 50% in soil water and shallow groundwater beneath the HT pond.

Effect of early addition of bone morphogenetic protein 5 (BMP5) to embryo culture medium on in vitro development and expression of developmentally important genes in bovine preimplantation embryos.

PubMed

García, Elina V; Miceli, Dora C; Rizo, Gabriela; Valdecantos, Pablo A; Barrera, Antonio D

2015-09-01

Previous studies have reported that bone morphogenetic protein 5 (BMP5) is differentially expressed in the isthmus of bovine oviducts and it is present in the oviductal fluid. However, the specific action of this factor is unknown. To evaluate whether BMP5 exerts some effect during early bovine embryo development, gene expression of BMP5, BMP receptors, and the effect of exogenous BMP5 on in vitro development and expression of developmentally important genes were assessed. In experiment 1, pools of embryos at two-cell, four-cell, eight-cell, and blastocyst stages, derived from in vitro fertilization, were collected for analysis of BMP5 and BMP receptors (BMPR1A, BMPR1B, and BMPR2) messenger RNA (mRNA) expression. On the basis of previous results, in experiment 2, presumptive zygotes were cultured for the first 48 hours after insemination in CR1aa medium assaying three different treatments: (1) control (CR1aa); (2) vehicle control (CR1aa + 0.04 mM HCl), and (3) BMP5 treatment (CR1aa + 100 ng/mL of BMP5). The cleavage rate was evaluated 48 hours after insemination (Day 2), and then, embryos were transferred to CR1aa + 10% fetal bovine serum. The blastocyst rate was determined on Day 7. In experiment 3, pools of embryos at two-cell, four-cell, eight-cell, and blastocyst stages, derived from control and BMP5-treated groups, were collected for analysis of ID2 (BMP target gene), OCT4, NANOG, and SOX2 (pluripotency genes) mRNA expression. BMP5 transcripts were not detectable in any of the embryonic stages examined, whereas the relative mRNA abundance of the three BMP receptors analyzed was greater in early embryo development stages before maternal-embryonic transition, raising the possibility of a direct effect of exogenous BMPs on the embryo during the first developmental period. Although early addition of 100 ng/mL of BMP5 to the embryo culture medium had no effect on the cleavage rate, a significantly higher proportion of cleaved embryos developed to the blastocyst stage in the BMP5 group. Moreover, reverse transcription quantitative real-time polymerase chain reaction analysis showed a significant increase in the relative abundance of SOX2 in two-cell stage embryos, ID2 and OCT4 in eight-cell stage embryos, and NANOG and OCT4 in blastocysts derived from BMP5-treated embryos. In conclusion, our results report that early addition of BMP5 to the embryo culture medium had a positive effect on the blastocyst rate and affected the relative expression of BMP target and pluripotency genes, suggesting that BMP5 could play an important role in the preimplantation development of bovine embryos. Copyright © 2015 Elsevier Inc. All rights reserved.

Dual delivery of rhPDGF-BB and bone marrow mesenchymal stromal cells expressing the BMP2 gene enhance bone formation in a critical-sized defect model.

PubMed

Park, Shin-Young; Kim, Kyoung-Hwa; Shin, Seung-Yun; Koo, Ki-Tae; Lee, Yong-Moo; Seol, Yang-Jo

2013-11-01

Bone tissue healing is a dynamic, orchestrated process that relies on multiple growth factors and cell types. Platelet-derived growth factor-BB (PDGF-BB) is released from platelets at wound sites and induces cellular migration and proliferation necessary for bone regeneration in the early healing process. Bone morphogenetic protein-2 (BMP-2), the most potent osteogenic differentiation inducer, directs new bone formation at the sites of bone defects. This study evaluated a combinatorial treatment protocol of PDGF-BB and BMP-2 on bone healing in a critical-sized defect model. To mimic the bone tissue healing process, a dual delivery approach was designed to deliver the rhPDGF-BB protein transiently during the early healing phase, whereas BMP-2 was supplied by rat bone marrow stromal cells (BMSCs) transfected with an adenoviral vector containing the BMP2 gene (AdBMP2) for prolonged release throughout the healing process. In in vitro experiments, the dual delivery of rhPDGF-BB and BMP2 significantly enhanced cell proliferation. However, the osteogenic differentiation of BMSCs was significantly suppressed even though the amount of BMP-2 secreted by the AdBMP2-transfected BMSCs was not significantly affected by the rhPDGF-BB treatment. In addition, dual delivery inhibited the mRNA expression of BMP receptor type II and Noggin in BMSCs. In in vivo experiments, critical-sized calvarial defects in rats showed enhanced bone regeneration by dual delivery of autologous AdBMP2-transfected BMSCs and rhPDGF-BB in both the amount of new bone formed and the bone mineral density. These enhancements in bone regeneration were greater than those observed in the group treated with AdBMP2-transfected BMSCs alone. In conclusion, the dual delivery of rhPDGF-BB and AdBMP2-transfected BMSCs improved the quality of the regenerated bone, possibly due to the modulation of PDGF-BB on BMP-2-induced osteogenesis.

PELA microspheres with encapsulated arginine-chitosan/pBMP-2 nanoparticles induce pBMP-2 controlled-release, transfected osteoblastic progenitor cells, and promoted osteogenic differentiation.

PubMed

Xu, Xiaolong; Qiu, Sujun; Zhang, Yuxian; Yin, Jie; Min, Shaoxiong

2017-03-01

Repair of the bone injury remains a challenge in clinical practices. Recent progress in tissue engineering and therapeutic gene delivery systems have led to promising new strategies for successful acceleration of bone repair process. The aim of this study was to create a controlled-release system to slowly release the arginine-chitosan/plasmid DNA nanoparticles encoding BMP-2 gene (Arg-CS/pBMP-2 NPs), efficiently transfect osteoblastic progenitor cells, secrete functional BMP-2 protein, and promote osteogenic differentiation. In this study, chitosan was conjugated with arginine to generate arginine-chitosan polymer (Arg-CS) for gene delivery. Mix the Arg-CS with pBMP-2 to condense pBMP-2 into nano-sized particles. In vitro transfection assays demonstrated that the transfection efficiency of Arg-CS/pBMP-2 nanoparticles and the expression level of BMP-2 was obviously exceed control groups. Further, PELA microspheres as the controlled-release carrier for the nanoparticles were used to encapsulate Arg-CS/pBMP-2 NPs. We demonstrated that the Arg-CS/pBMP-2 NPs could slowly release from the PELA microspheres at least for 42 d. During the co-culture with the PELA microspheres, the content of BMP-2 protein secreted by MC3T3-E1 reached the peak at 7 d. After 21d, the secretion of BMP-2 protein still maintain a higher level. The alkaline phosphatase activity, alizarin red staining, and osteogenesis-related gene expression by real-time quantitative PCR analysis all showed the PELA microspheres entrapping with Arg-CS/pBMP-2 NPs can obviously induce the osteogenic differentiation. The results indicated that the Arg-CS is a suitable gene vector which can promote the gene transfection. And the novel PELA microspheres-nanoparticle controlled-release system has potential clinical application in the future after further research.

Arsenite suppression of BMP signaling in human keratinocytes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Phillips, Marjorie A.; Qin, Qin; Hu, Qin

2013-06-15

Arsenic, a human skin carcinogen, suppresses differentiation of cultured keratinocytes. Exploring the mechanism of this suppression revealed that BMP-6 greatly increased levels of mRNA for keratins 1 and 10, two of the earliest differentiation markers expressed, a process prevented by co-treatment with arsenite. BMP also stimulated, and arsenite suppressed, mRNA for FOXN1, an important transcription factor driving early keratinocyte differentiation. Keratin mRNAs increased slowly after BMP-6 addition, suggesting they are indirect transcriptional targets. Inhibition of Notch1 activation blocked BMP induction of keratins 1 and 10, while FOXN1 induction was largely unaffected. Supporting a requirement for Notch1 signaling in keratin induction,more » BMP increased levels of activated Notch1, which was blocked by arsenite. BMP also greatly decreased active ERK, while co-treatment with arsenite maintained active ERK. Inhibition of ERK signaling mimicked BMP by inducing keratin and FOXN1 mRNAs and by increasing active Notch1, effects blocked by arsenite. Of 6 dual-specificity phosphatases (DUSPs) targeting ERK, two were induced by BMP unless prevented by simultaneous exposure to arsenite and EGF. Knockdown of DUSP2 or DUSP14 using shRNAs greatly reduced FOXN1 and keratins 1 and 10 mRNA levels and their induction by BMP. Knockdown also decreased activated Notch1, keratin 1 and keratin 10 protein levels, both in the presence and absence of BMP. Thus, one of the earliest effects of BMP is induction of DUSPs, which increases FOXN1 transcription factor and activates Notch1, both required for keratin gene expression. Arsenite prevents this cascade by maintaining ERK signaling, at least in part by suppressing DUSP expression. - Highlights: • BMP induces FOXN1 transcription. • BMP induces DUSP2 and DUSP14, suppressing ERK activation. • Arsenite suppresses levels of phosphorylated Smad1/5 and FOXN1 and DUSP mRNA. • These actions rationalize arsenite suppression of keratinocyte differentiation.« less

The expression and crucial roles of BMP signaling in development of smooth muscle progenitor cells in the mouse embryonic gut.

PubMed

Torihashi, Shigeko; Hattori, Takako; Hasegawa, Hirotaka; Kurahashi, Masaaki; Ogaeri, Takunori; Fujimoto, Toyoshi

2009-03-01

Bone morphogenetic protein (BMP) signaling is essential for normal development of the gastrointestinal (GI) tract. BMPs also play multiple roles in vascular smooth muscle cells; however, the BMP signaling in the development of the GI musculature remains to be clarified. We investigated the expression of BMPs and their receptors in mouse embryonic GI tracts by immunohistochemistry and in situ hybridization. We demonstrated that BMP2, BMP receptor Ib and BMP receptor II were expressed in the smooth muscle progenitors from E12 to E13 for the first time. BMP signaling on smooth muscle differentiation was examined by implantation of agarose beads soaked with BMPs in the in vitro developmental model that is gut-like structures from mouse embryonic stem (ES) cells. BMP2 rather than BMP4 beads enhanced smooth muscle differentiation, and increased gut-like structures showing spontaneous contractions and expressing intensive alpha-smooth muscle actin immunoreactivity. This increase was confirmed by up-regulation of SM22 mRNA shown by real-time PCR. By addition of noggin beads or noggin to the medium at BMP2 bead implantation, the ratio of contractive gut-like structures decreased. Implantation of BMP2 beads at EB7 (EB--embryoid bodies) (corresponding to E12 or E13 of mouse embryo) showed the highest effects and up-regulation of transcription factors msx-1 after 24h. This increase was blocked by noggin, and msx-1 decreased to almost the control level after 60 h. BMP2 beads at EB7 increased platelet-derived growth factor-A (PDGF-A) in the differentiating smooth muscle cells. We have recently reported that PDGF-A is expressed in the developing inner circular smooth muscle and is crucial for the longitudinal smooth muscle differentiation. Taken together, BMP signaling was expressed for a short window in the smooth muscle progenitors and the signal, especially BMP2, plays an essential role in smooth muscle differentiation in cooperation with PDGF signaling.

A comparative examination of odontogenic gene expression in both toothed and toothless amniotes

PubMed Central

Lainoff, Alexis J.; Moustakas-Verho, Jacqueline E.; Hu, Diane; Kallonen, Aki; Marcucio, Ralph S.; Hlusko, Leslea J.

2015-01-01

A well-known tenet of murine tooth development is that BMP4 and FGF8 antagonistically initiate odontogenesis, but whether this tenet is conserved across amniotes is largely unexplored. Moreover, changes in BMP4-signaling have previously been implicated in evolutionary tooth loss in Aves. Here we demonstrate that Bmp4, Msx1, and Msx2 expression is limited proximally in the red-eared slider turtle (Trachemys scripta) mandible at stages equivalent to those at which odontogenesis is initiated in mice, a similar finding to previously reported results in chicks. To address whether the limited domains in the turtle and the chicken indicate an evolutionary molecular parallelism, or whether the domains simply constitute an ancestral phenotype, we assessed gene expression in a toothed reptile (the American alligator, Alligator mississippiensis) and a toothed non-placental mammal (the gray short-tailed opossum, Monodelphis domestica). We demonstrate that the Bmp4 domain is limited proximally in M. domestica and that the Fgf8 domain is limited distally in A. mississippiensis just preceding odontogenesis. Additionally, we show that Msx1 and Msx2 expression patterns in these species differ from those found in mice. Our data suggest that a limited Bmp4 domain does not necessarily correlate with edentulism, and reveal that the initiation of odontogenesis in non-murine amniotes is more complex than previously imagined. Our data also suggest a partially conserved odontogenic program in T. scripta, as indicated by conserved Pitx2, Pax9, and Barx1 expression patterns and by the presence of a Shh-expressing palatal epithelium, which we hypothesize may represent potential dental rudiments based on the Testudinata fossil record. PMID:25678399

Expression of bone morphogenetic proteins 4, 6 and 7 is downregulated in kidney allografts with interstitial fibrosis and tubular atrophy.

PubMed

Furic-Cunko, Vesna; Kes, Petar; Coric, Marijana; Hudolin, Tvrtko; Kastelan, Zeljko; Basic-Jukic, Nikolina

2015-07-01

Bone morphogenetic proteins (BMPs) are pleiotropic growth factors. This paper investigates the connection between the expression pattern of BMPs in kidney allograft tissue versus the cause of allograft dysfunction. The expression pattern of BMP2, BMP4, BMP6 and BMP7 in 50 kidney allografts obtained by transplant nephrectomy is investigated. Immunohistochemical staining is semiquantitatively evaluated for intensity to identify the expression pattern of BMPs in normal and allograft kidney tissues. The expression of BMP4 is unique between different tubular cell types in grafts without signs of fibrosis. This effect is not found in specimens with high grades of interstitial fibrosis and tubular atrophy (IFTA). In samples with IFTA grades II and III, the BMP7 expression is reduced in a significant fraction of specimens relative to those without signs of IFTA. The expression pattern of BMP6 indicates that its activation may be triggered by the act of transplantation and subsequent reperfusion injury. The expression of BMP2 is strong in all types of tubular epithelial cells and does not differ between the compared allografts and control kidney specimens. The intensity and expression pattern of BMP4, BMP6 and BMP7 in transplanted kidney tissue are found to be dependent upon the length of the transplanted period, the clinical indication for transplant nephrectomy and signs of IFTA in kidney tissue.

Bone morphogenetic protein 9 regulates tumor growth of osteosarcoma cells through the Wnt/β-catenin pathway.

PubMed

Lv, Zilan; Wang, Chuan; Yuan, Taixian; Liu, Yuehong; Song, Tao; Liu, Yueliang; Chen, Chu; Yang, Min; Tang, Zuchuan; Shi, Qiong; Weng, Yaguang

2014-02-01

Bone morphogenetic protein 9 (BMP9) is a member of the transforming growth factor-β (TGF-β) family, which has been shown to regulate the progression of several tumors. Recent studies indicated that BMP9 affects osteosarcoma (OS) processes, but its specific roles and molecular mechanisms have yet to be fully elucidated. The human OS cell lines 143B and MG63 were used for the present study. We found that BMP9 overexpression suppressed the growth of OS cells, whereas inhibition of BMP9 reversed this effect. Our results also showed that BMP9 overexpression induced G0/G1 phase arrest and apoptosis in OS cells. We further investigated the possible molecular mechanisms mediating the biological role of BMP9. We observed that BMP9 overexpression reduced β-catenin mRNA and protein levels, and also downregulated its downstream proteins c-Myc and osteoprotegerin (OPG) and inhibited the phosphorylation levels of GSK-3β (Ser 9) in OS cells, whereas inhibition of BMP9 reversed these effects. Moreover, the suppressive effects of BMP9 overexpression on OS cells was reversed by exogenous β-catenin expression, but augmented by β-catenin silencing. In conclusion, our results revealed that BMP9 can regulate tumor growth of OS cells through the Wnt/β-catenin pathway. Therefore, BMP9 may be a new therapeutic target in OS.

A new molecular logic for BMP-mediated dorsoventral patterning in the leech Helobdella.

PubMed

Kuo, Dian-Han; Weisblat, David A

2011-08-09

Bone morphogenetic protein (BMP) signaling is broadly implicated in dorsoventral (DV) patterning of bilaterally symmetric animals [1-3], and its role in axial patterning apparently predates the birth of Bilateria [4-7]. In fly and vertebrate embryos, BMPs and their antagonists (primarily Sog/chordin) diffuse and interact to generate signaling gradients that pattern fields of cells [8-10]. Work in other species reveals diversity in essential facets of this ancient patterning process, however. Here, we report that BMP signaling patterns the DV axis of segmental ectoderm in the leech Helobdella, a clitellate annelid (superphylum Lophotrochozoa) featuring stereotyped developmental cell lineages, but the detailed mechanisms of DV patterning in Helobdella differ markedly from fly and vertebrates. In Helobdella, BMP2/4s are expressed broadly, rather than in dorsal territory, whereas a dorsally expressed BMP5-8 specifies dorsal fate by short-range signaling. A BMP antagonist, gremlin, is upregulated by BMP5-8 in dorsolateral, rather than ventral territory, and yet the BMP-antagonizing activity of gremlin is required for normal ventral cell fates. Gremlin promotes ventral fates without disrupting dorsal fates by selectively inhibiting BMP2/4s, not BMP5-8. Thus, DV patterning in the development of the leech revealed unexpected evolutionary plasticity of the conserved BMP patterning system, presumably reflecting its adaptation to different modes of embryogenesis. Copyright © 2011 Elsevier Ltd. All rights reserved.

Implant Composed of Demineralized Bone and Mesenchymal Stem Cells Genetically Modified with AdBMP2/AdBMP7 for the Regeneration of Bone Fractures in Ovis aries.

PubMed

Hernandez-Hurtado, Adelina A; Borrego-Soto, Gissela; Marino-Martinez, Ivan A; Lara-Arias, Jorge; Romero-Diaz, Viktor J; Abrego-Guerra, Adalberto; Vilchez-Cavazos, Jose F; Elizondo-Riojas, Guillermo; Martinez-Rodriguez, Herminia G; Espinoza-Juarez, Marcela A; Lopez-Romero, Gloria C; Robles-Zamora, Alejandro; Mendoza Lemus, Oscar F; Ortiz-Lopez, Rocio; Rojas-Martinez, Augusto

2016-01-01

Adipose-derived mesenchymal stem cells (ADMSCs) are inducible to an osteogenic phenotype by the bone morphogenetic proteins (BMPs). This facilitates the generation of implants for bone tissue regeneration. This study evaluated the in vitro osteogenic differentiation of ADMSCs transduced individually and in combination with adenoviral vectors expressing BMP2 and BMP7. Moreover, the effectiveness of the implant containing ADMSCs transduced with the adenoviral vectors AdBMP2/AdBMP7 and embedded in demineralized bone matrix (DBM) was tested in a model of tibial fracture in sheep. This graft was compared to ewes implanted with untransduced ADMSCs embedded in the same matrix and with injured but untreated animals. In vivo results showed accelerated osteogenesis in the group treated with the AdBMP2/AdBMP7 transduced ADMSC graft, which also showed improved restoration of the normal bone morphology.

Regulation of Msx-1, Msx-2, Bmp-2 and Bmp-4 during foetal and postnatal mammary gland development.

PubMed

Phippard, D J; Weber-Hall, S J; Sharpe, P T; Naylor, M S; Jayatalake, H; Maas, R; Woo, I; Roberts-Clark, D; Francis-West, P H; Liu, Y H; Maxson, R; Hill, R E; Dale, T C

1996-09-01

Expression of the Msx-1 and Msx-2 homeobox genes have been shown to be coordinately regulated with the Bmp-2 and Bmp-4 ligands in a variety of developing tissues. Here we report that transcripts from all four genes are developmentally regulated during both foetal and postnatal mammary gland development. The location and time-course of the Bmp and Msx expression point to a role for Msx and Bmp gene products in the control of epithelial-mesenchymal interactions. Expression of Msx-2, but not Msx-1, Bmp-2 or Bmp-4 was decreased following ovariectomy, while expression of the human Msx-2 homologue was regulated by 17beta-oestradiol in the MCF-7 breast cancer cell line. The regulation of Msx-2 expression by oestrogen raises the possibility that hormonal regulation of mammary development is mediated through the control of epithelial-mesenchymal interactions.

BMP4 Signaling Is Able to Induce an Epithelial-Mesenchymal Transition-Like Phenotype in Barrett’s Esophagus and Esophageal Adenocarcinoma through Induction of SNAIL2

PubMed Central

Kestens, Christine; Siersema, Peter D.; Offerhaus, G. Johan A.; van Baal, Jantine W. P. M.

2016-01-01

Background Bone morphogenetic protein 4 (BMP4) signaling is involved in the development of Barrett’s esophagus (BE), a precursor of esophageal adenocarcinoma (EAC). In various cancers, BMP4 has been found to induce epithelial-mesenchymal transition (EMT) but its function in the development of EAC is currently unclear. Aim To investigate the expression of BMP4 and several members of the BMP4 pathway in EAC. Additionally, to determine the effect of BMP4 signaling in a human Barrett’s esophagus (BAR-T) and adenocarcinoma (OE33) cell line. Methods Expression of BMP4, its downstream target ID2 and members of the BMP4 pathway were determined by Q-RT-PCR, immunohistochemistry and Western blot analysis using biopsy samples from EAC patients. BAR-T and OE33 cells were incubated with BMP4 or the BMP4 antagonist, Noggin, and cell viability and migration assays were performed. In addition, expression of factors associated with EMT (SNAIL2, CDH1, CDH2 and Vimentin) was evaluated by Q-RT-PCR and Western blot analysis. Results Compared to squamous epithelium (SQ), BMP4 expression was significantly upregulated in EAC and BE. In addition, the expression of ID2 was significantly upregulated in EAC and BE compared to SQ. Western blot analysis confirmed our results, showing an upregulated expression of BMP4 and ID2 in both BE and EAC. In addition, more phosphorylation of SMAD1/5/8 was observed. BMP4 incubation inhibited cell viability, but induced cell migration in both BAR-T and OE33 cells. Upon BMP4 incubation, SNAIL2 expression was significantly upregulated in BAR-T and OE33 cells while CDH1 expression was significantly downregulated. These results were confirmed by Western blot analysis. Conclusion Our results indicate active BMP4 signaling in BE and EAC and suggest that this results in an invasive phenotype by inducing an EMT-like response through upregulation of SNAIL2 and subsequent downregulation of CDH1. PMID:27191723

Bone Morphogenetic Protein (BMP) signaling in animal reproductive system development and function.

PubMed

Lochab, Amaneet K; Extavour, Cassandra G

2017-07-15

In multicellular organisms, the specification, maintenance, and transmission of the germ cell lineage to subsequent generations are critical processes that ensure species survival. A number of studies suggest that the Bone Morphogenetic Protein (BMP) pathway plays multiple roles in this cell lineage. We wished to use a comparative framework to examine the role of BMP signaling in regulating these processes, to determine if patterns would emerge that might shed light on the evolution of molecular mechanisms that may play germ cell-specific or other reproductive roles across species. To this end, here we review evidence to date from the literature supporting a role for BMP signaling in reproductive processes across Metazoa. We focus on germ line-specific processes, and separately consider somatic reproductive processes. We find that from primordial germ cell (PGC) induction to maintenance of PGC identity and gametogenesis, BMP signaling regulates these processes throughout embryonic development and adult life in multiple deuterostome and protostome clades. In well-studied model organisms, functional genetic evidence suggests that BMP signaling is required in the germ line across all life stages, with the exception of PGC specification in species that do not use inductive signaling to induce germ cell formation. The current evidence is consistent with the hypothesis that BMP signaling is ancestral in bilaterian inductive PGC specification. While BMP4 appears to be the most broadly employed ligand for the reproductive processes considered herein, we also noted evidence for sex-specific usage of different BMP ligands. In gametogenesis, BMP6 and BMP15 seem to have roles restricted to oogenesis, while BMP8 is restricted to spermatogenesis. We hypothesize that a BMP-based mechanism may have been recruited early in metazoan evolution to specify the germ line, and was subsequently co-opted for use in other germ line-specific and somatic reproductive processes. We suggest that if future studies assessing the function of the BMP pathway across extant species were to include a reproductive focus, that we would be likely to find continued evidence in favor of an ancient association between BMP signaling and the reproductive cell lineage in animals. Copyright © 2017 Elsevier Inc. All rights reserved.

Canonical Wnt signaling acts synergistically on BMP9-induced osteo/odontoblastic differentiation of stem cells of dental apical papilla (SCAPs)

PubMed Central

Zhang, Hongmei; Wang, Jinhua; Deng, Fang; Huang, Enyi; Yan, Zhengjian; Wang, Zhongliang; Deng, Youlin; Zhang, Qian; Zhang, Zhonglin; Ye, Jixing; Qiao, Min; Li, Ruifang; Wang, Jing; Wei, Qiang; Zhou, Guolin; Luu, Hue H.; Haydon, Rex C.; He, Tong-Chuan; Deng, Feng

2014-01-01

Dental pulp/dentin regeneration using dental stem cells combined with odontogenic factors may offer great promise to treat and/or prevent premature tooth loss. Here, we investigate if BMP9 and Wnt/β-catenin act synergistically on odontogenic differentiation. Using the immortalized SCAPs (iSCAPs) isolated from mouse apical papilla tissue, we demonstrate that Wnt3A effectively induces early osteogenic marker alkaline phosphatase (ALP) in iSCAPs, which is reduced by β-catenin knockdown. While Wnt3A and BMP9 enhance each other’s ability to induce ALP activity in iSCAPs, silencing β- catenin significantly diminishes BMP9-induced osteo/odontogenic differentiation. Furthermore, silencing β-catenin reduces BMP9-induced expression of osteocalcin and osteopontin and in vitro matrix mineralization of iSCAPs. In vivo stem cell implantation assay reveals that while BMP9-transduced iSCAPs induce robust ectopic bone formation, iSCAPs stimulated with both BMP9 and Wnt3A exhibit more mature and highly mineralized trabecular bone formation. However, knockdown of β-catenin in iSCAPs significantly diminishes BMP9 or BMP9/Wnt3A-induced ectopic bone formation in vivo. Thus, our results strongly suggest that β-catenin may play an important role in BMP9-induced osteo/ondontogenic signaling and that BMP9 and Wnt3A may act synergistically to induce osteo/odontoblastic differentiation of iSCAPs. It’s conceivable that BMP9 and/or Wnt3A may be explored as efficacious biofactors for odontogenic regeneration and tooth engineering. PMID:25468367

Postoperative Complications Associated With rhBMP2 Use in Posterior/Posterolateral Lumbar Fusion.

PubMed

Esmail, Nabil; Buser, Zorica; Cohen, Jeremiah R; Brodke, Darrel S; Meisel, Hans-Joerg; Park, Jong-Beom; Youssef, Jim A; Wang, Jeffrey C; Yoon, S Tim

2018-04-01

Retrospective database review. Posterior/posterolateral lumbar fusion (PLF) is an effective treatment for a variety of spinal disorders; however, variations in surgical technique have different complication profiles. The aim of our study was to quantify the frequency of various complications in patients undergoing PLF with and without human recombinant bone morphogenetic protein 2 (rhBMP2). We queried the orthopedic subset of the Medicare database (PearlDiver) between 2005 and 2011 for patients undergoing PLF procedures with and without rhBMP2. Complication and reoperation rates were analyzed within 1 year of the index procedure. Complications assessed include: acute renal failure, deep vein thrombosis, dural tear, hematoma, heterotopic ossification, incision and drainage, cardiac complications, nervous system complications, osteolysis, pneumonia, pseudarthrosis, pulmonary embolism, radiculopathy, respiratory complications, sepsis, urinary retention, urinary tract infection, mechanical, and wound complications. Chi-square analysis was used to calculate the complication differences between the groups. Our data revealed higher overall complication rates in patients undergoing PLF with rhBMP2 versus no_rhBMP2 (76.9% vs 68.8%, P < .05). Stratified by gender, rhBMP2 males had higher rates of mechanical complications, pseudarthrosis, and reoperations compared with no_rhBMP2 males ( P < .05), whereas rhBMP2 females had higher rates of pseudarthrosis, urinary tract infection, and urinary retention compared with no_rhBMP2 females ( P < .05). Our data revealed higher overall complication rates in PLF patients given rhBMP2 compared with no_rhBMP2. Furthermore, our data suggests that rhBMP2-associated complications may be gender specific.

Sizzled controls dorso-ventral polarity by repressing cleavage of the Chordin protein.

PubMed

Muraoka, Osamu; Shimizu, Takashi; Yabe, Taijiro; Nojima, Hideaki; Bae, Young-Ki; Hashimoto, Hisashi; Hibi, Masahiko

2006-04-01

The Bone morphogenetic protein (Bmp) signalling gradient has a major function in the formation of the dorso-ventral axis. The zebrafish ventralized mutant, ogon, encodes Secreted Frizzled (Sizzled). sizzled is ventrally expressed in a Bmp-dependent manner and is required for the suppression of Bmp signalling on the ventral side of zebrafish embryos. However, it remains unclear how Sizzled inhibits Bmp signalling and controls ventro-lateral cell fate. We found that Sizzled stabilizes Chordin, a Bmp antagonist, by binding and inhibiting the Tolloid-family metalloproteinase, Bmp1a, which cleaves and inactivates Chordin. The cysteine-rich domain of Sizzled is required for inhibition of Bmp1a activity. Loss of both Bmp1a and Tolloid-like1 (Tll1; another Tolloid-family metalloproteinase) function leads to a complete suppression and reversal of the ogon mutant phenotype. These results indicate that Sizzled represses the activities of Tolloid-family proteins, thereby creating the Chordin-Bmp activity gradient along the dorso-ventral axis. Here, we describe a previously unrecognized role for a secreted Frizzled-related protein.

BMP-2 and titanium particles synergistically activate osteoclast formation

PubMed Central

Sun, S.X.; Guo, H.H.; Zhang, J.; Yu, B.; Sun, K.N.; Jin, Q.H.

2014-01-01

A previous study showed that BMP-2 (bone morphogenetic protein-2) and wear debris can separately support osteoclast formation induced by the receptor activator of NF-κB ligand (RANKL). However, the effect of BMP-2 on wear debris-induced osteoclast formation is unclear. In this study, we show that neither titanium particles nor BMP-2 can induce osteoclast formation in RAW 264.7 mouse leukemic monocyte macrophage cells but that BMP-2 synergizes with titanium particles to enhance osteoclast formation in the presence of RANKL, and that at a low concentration, BMP-2 has an optimal effect to stimulate the size and number of multinuclear osteoclasts, expression of osteoclast genes, and resorption area. Our data also clarify that the effects caused by the increase in BMP-2 on phosphorylated SMAD levels such as c-Fos expression increased throughout the early stages of osteoclastogenesis. BMP-2 and titanium particles stimulate the expression of p-JNK, p-P38, p-IkB, and P50 compared with the titanium group. These data suggested that BMP-2 may be a crucial factor in titanium particle-mediated osteoclast formation. PMID:24820069

Ski represses BMP signaling in Xenopus and mammalian cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

kluo@lbl.gov

2001-05-16

The bone morphogenic proteins (BMPs) play important roles in vertebrate development. In Xenopus, BMPs act as epidermal inducers and also as negative regulators of neurogenesis. Antagonism of BMP signaling results in neuralization. BMPs signal through the cell-surface receptors and downstream Smad molecules. Upon stimulation with BMP, Smad1, Smad5, and Smad8 are phosphorylated by the activated BMP receptors, form a complex with Smad4, and translocate into the nucleus, where they regulate the expression of BMP target genes. Here, we show that the Ski oncoprotein can block BMP signaling and the expression of BMP-responsive genes in both Xenopus and mammalian cells bymore » directly interacting with and repressing the activity of BMP-specific Smad complexes. This ability to antagonize BMP signaling results in neuralization by Ski in the Xenopus embryo and blocking of osteoblast differentiation of murine W-20-17 cells. Thus, Ski is able to repress the activity of all receptor-associated Smads and may regulate vertebrate development by modulating the signaling activity of transforming growth factor-{beta} family members.« less

TGF-β but not BMP signaling induces prechondrogenic condensation through ATP oscillations during chondrogenesis.

PubMed

Kwon, Hyuck Joon

2012-08-10

Although both TGF-β and BMP signaling enhance expression of adhesion molecules during chondrogenesis, TGF-β but not BMP signaling can initiate condensation of uncondensed mesenchymal cells. However, it remains unclear what causes the differential effects between TGF-β and BMP signaling on prechondrogenic condensation. Our previous report demonstrated that ATP oscillations play a critical role in prechondrogenic condensation. Thus, the current study examined whether ATP oscillations are associated with the differential actions of TGF-β and BMP signaling on prechondrogenic condensation. The result revealed that while both TGF-β1 and BMP2 stimulated chondrogenic differentiation, TGF-β1 but not BMP2 induced prechondrogenic condensation. It was also found that TGF-β1 but not BMP2 induced ATP oscillations and inhibition of TGF-β but not BMP signaling prevented insulin-induced ATP oscillations. Moreover, blockage of ATP oscillations inhibited TGF-β1-induced prechondrogenic condensation. In addition, TGF-β1-driven ATP oscillations and prechondrogenic condensation depended on Ca(2+) influx via voltage-dependent calcium channels. This study suggests that Ca(2+)-driven ATP oscillations mediate TGF-β-induced the initiation step of prechondrogenic condensation and determine the differential effects between TGF-β and BMP signaling on chondrogenesis. Copyright © 2012 Elsevier Inc. All rights reserved.

Bone Morphogenic Protein-2 (rhBMP2)-Loaded Silk Fibroin Scaffolds to Enhance the Osteoinductivity in Bone Tissue Engineering

NASA Astrophysics Data System (ADS)

Du, Guang-Yu; He, Sheng-Wei; Sun, Chuan-Xiu; Mi, Li-Dong

2017-10-01

There is an increasing demand for formulations of silk fibroin (SF) scaffolds in biomedical applications. SF was crosslinked via glutaraldehyde with osteoinductive recombinant human bone morphogenic protein-2 (rhBMP2) of different ratios viz. (i) 3% SF with no rhBMP2 (SF), (ii) 3% SF with equal amount of rhBMP2 (SF+BMP2), and (iii) 12% SF with 3% of rhBMP2 (4SF+BMP2), and these solutions were used in electrospinning-based fabrication of nanoscaffolds for evaluating increased osteoinductive potential of SF scaffolds with rhBMP2. Stress-strain relationship suggested there is no loss in mechanical strength of fibers with addition of rhBMP2, and mechanical strength of scaffold was improved with increase in concentration of SF. rhBMP2 association increased the water retention capacity of scaffold as evident from swelling studies. Viability of hMSCs was found to be higher in conjugated scaffolds, and scaffolds do not exhibit any cytotoxicity towards guest cells. Cells were found to have higher alkaline phosphatase activity in conjugated scaffolds under in vitro and in vivo conditions which establishes the increased osteoinductivity of the novel construct. The scaffolds were found to be effective for in vivo bone formation as well.

Bmp6 Regulates Retinal Iron Homeostasis and Has Altered Expression in Age-Related Macular Degeneration

PubMed Central

Hadziahmetovic, Majda; Song, Ying; Wolkow, Natalie; Iacovelli, Jared; Kautz, Leon; Roth, Marie-Paule; Dunaief, Joshua L.

2011-01-01

Iron-induced oxidative stress causes hereditary macular degeneration in patients with aceruloplasminemia. Similarly, retinal iron accumulation in age-related macular degeneration (AMD) may exacerbate the disease. The cause of retinal iron accumulation in AMD is poorly understood. Given that bone morphogenetic protein 6 (Bmp6) is a major regulator of systemic iron, we examined the role of Bmp6 in retinal iron regulation and in AMD pathogenesis. Bmp6 was detected in the retinal pigment epithelium (RPE), a major site of pathology in AMD. In cultured RPE cells, Bmp6 was down-regulated by oxidative stress and up-regulated by iron. Intraocular Bmp6 protein injection in mice up-regulated retinal hepcidin, an iron regulatory hormone, and altered retinal labile iron levels. Bmp6−/− mice had age-dependent retinal iron accumulation and degeneration. Postmortem RPE from patients with early AMD exhibited decreased Bmp6 levels. Because oxidative stress is associated with AMD pathogenesis and down-regulates Bmp6 in cultured RPE cells, the diminished Bmp6 levels observed in RPE cells in early AMD may contribute to iron build-up in AMD. This may in turn propagate a vicious cycle of oxidative stress and iron accumulation, exacerbating AMD and other diseases with hereditary or acquired iron excess. PMID:21703414

Monitoring sediment production from forest road approaches to stream crossing in the Virginia Piedmont

Treesearch

Kristopher R. Brown; W. Michael Aust; Kevin J. McGuire

2015-01-01

Reopening of abandoned legacy roads is common in forest operations and represents a reduced cost in comparison to new road construction. However, legacy roads may have lower road standards and require additional best management practice (BMP) implementation upon reopening to protect water quality. Silt fences and elevation measurements of trapped sediment were used to...

In vitro and in vivo evaluation of calcium phosphate composite scaffolds containing BMP-VEGF loaded PLGA microspheres for the treatment of avascular necrosis of the femoral head.

PubMed

Zhang, Hao-Xuan; Zhang, Xiu-Ping; Xiao, Gui-Yong; Hou, Yong; Cheng, Lei; Si, Meng; Wang, Shuai-Shuai; Li, Yu-Hua; Nie, Lin

2016-03-01

Avascular necrosis of the femoral head (ANFH) is difficult to treat due to high pressure and hypoxia, and reduced levels of growth factors such as bone morphogenetic protein (BMP), and vascular endothelial growth factor (VEGF). We generated a novel calcium phosphate (CPC) composite scaffold, which contains BMP-VEGF-loaded poly-lactic-co-glycolic acid (PLGA) microspheres (BMP-VEGF-PLGA-CPC). The BMP-VEGF-loaded microspheres have an encapsulation efficiency of 89.15% for BMP, and 78.55% for VEGF. The BMP-VEGF-PLGA-CPC scaffold also demonstrated a porosity of 62% with interconnected porous structures, and pore sizes of 219 μm and compressive strength of 6.60 MPa. Additionally, bone marrow mesenchymal stem cells (BMSCs) were seeded on scaffolds in vitro. Further characterization showed that the BMP-VEGF-PLGA-CPC scaffolds were biocompatible and enhanced osteogenesis and angiogenesis in vitro. Using a rabbit model of ANFH, BMP-VEGF-PLGA-CPC scaffolds were implanted into the bone tunnels of core decompression in the femoral head for 6 and 12 weeks. Radiographic and histological analysis demonstrated that the BMP-VEGF-PLGA-CPC scaffolds exhibited good biocompatibility, and osteogenic and angiogenic activity in vivo. These results indicate that the BMP-VEGF-PLGA-CPC scaffold may improve the therapeutic effect of core decompression surgery and be used as a treatment for ANFH. Copyright © 2015 Elsevier B.V. All rights reserved.

Hydrolysis and Sulfation Pattern Effects on Release of Bioactive Bone Morphogenetic Protein-2 from Heparin-Based Microparticles.

PubMed

Tellier, Liane E; Miller, Tobias; McDevitt, Todd C; Temenoff, Johnna S

2015-10-28

Glycosaminoglycans (GAGs) such as heparin are promising materials for growth factor delivery due to their ability to efficiently bind positively charged growth factors including bone morphogenetic protein-2 (BMP-2) through their negatively charged sulfate groups. Therefore, the goal of this study was to examine BMP-2 release from heparin-based microparticles (MPs) after first, incorporating a hydrolytically degradable crosslinker and varying heparin content within MPs to alter MP degradation and second, altering the sulfation pattern of heparin within MPs to vary BMP-2 binding and release. Using varied MP formulations, it was found that the time course of MP degradation for 1 wt% heparin MPs was ~4 days slower than 10 wt% heparin MPs, indicating that MP degradation was dependent on heparin content. After incubating 100 ng BMP-2 with 0.1 mg MPs, most MP formulations loaded BMP-2 with ~50% efficiency and significantly more BMP-2 release (60% of loaded BMP-2) was observed from more sulfated heparin MPs (MPs with ~100% and 80% of native sulfation). Similarly, BMP-2 bioactivity in more sulfated heparin MP groups was at least four-fold higher than soluble BMP-2 and less sulfated heparin MP groups, as determined by an established C2C12 cell alkaline phosphatase (ALP) assay. Ultimately, the two most sulfated 10 wt% heparin MP formulations were able to efficiently load and release BMP-2 while enhancing BMP-2 bioactivity, making them promising candidates for future growth factor delivery applications.

BMP-2 Induces Versican and Hyaluronan That Contribute to Post-EMT AV Cushion Cell Migration

PubMed Central

Inai, Kei; Burnside, Jessica L.; Hoffman, Stanley; Toole, Bryan P.; Sugi, Yukiko

2013-01-01

Distal outgrowth and maturation of mesenchymalized endocardial cushions are critical morphogenetic events during post-EMT atrioventricular (AV) valvuloseptal morphogenesis. We explored the role of BMP-2 in the regulation of valvulogenic extracellular matrix (ECM) components, versican and hyaluronan (HA), and cell migration during post-EMT AV cushion distal outgrowth/expansion. We observed intense staining of versican and HA in AV cushion mesenchyme from the early cushion expansion stage, Hamburger and Hamilton (HH) stage-17 to the cushion maturation stage, HH stage-29 in the chick. Based on this expression pattern we examined the role of BMP-2 in regulating versican and HA using 3D AV cushion mesenchymal cell (CMC) aggregate cultures on hydrated collagen gels. BMP-2 induced versican expression and HA deposition as well as mRNA expression of versican and Has2 by CMCs in a dose dependent manner. Noggin, an antagonist of BMP, abolished BMP-2-induced versican and HA as well as mRNA expression of versican and Has2. We further examined whether BMP-2-promoted cell migration was associated with expression of versican and HA. BMP-2- promoted cell migration was significantly impaired by treatments with versican siRNA and HA oligomer. In conclusion, we provide evidence that BMP-2 induces expression of versican and HA by AV CMCs and that these ECM components contribute to BMP-2-induced CMC migration, indicating critical roles for BMP-2 in distal outgrowth/expansion of mesenchymalized AV cushions. PMID:24147033

BMP15 c.-9C>G promoter sequence variant may contribute to the cause of non-syndromic premature ovarian failure.

PubMed

Fonseca, Dora Janeth; Ortega-Recalde, Oscar; Esteban-Perez, Clara; Moreno-Ortiz, Harold; Patiño, Liliana Catherine; Bermúdez, Olga María; Ortiz, Angela María; Restrepo, Carlos M; Lucena, Elkin; Laissue, Paul

2014-11-01

BMP15 has drawn particular attention in the pathophysiology of reproduction, as its mutations in mammalian species have been related to different reproductive phenotypes. In humans, BMP15 coding regions have been sequenced in large panels of women with premature ovarian failure (POF), but only some mutations have been definitely validated as causing the phenotype. A functional association between the BMP15 c.-9C>G promoter polymorphism and cause of POF have been reported. The aim of this study was to determine the potential functional effect of this sequence variant on specific BMP15 promoter transactivation disturbances. Bioinformatics was used to identify transcription factor binding sites located on the promoter region of BMP15. Reverse transcription polymerase chain reaction was used to study specific gene expression in ovarian tissue. Luciferase reporter assays were used to establish transactivation disturbances caused by the BMP15 c.-9C>G variant. The c.-9C>G variant was found to modify the PITX1 transcription factor binding site. PITX1 and BMP15 co-expressed in human and mouse ovarian tissue, and PITX1 transactivated both BMP15 promoter versions (-9C and -9G). It was found that the BMP15 c.-9G allele was related to BMP15 increased transcription, supporting c.-9C>G as a causal agent of POF. Copyright © 2014 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.

Combined effects of dentin sialoprotein and bone morphogenetic protein-2 on differentiation in human cementoblasts.

PubMed

Lee, So-Youn; Auh, Q-Schick; Kang, Soo-Kyung; Kim, Hyung-Joon; Lee, Jung-Woo; Noh, Kwantae; Jang, Jun-Hyeog; Kim, Eun-Cheol

2014-07-01

The aim of this study is to determine the effects of the combination of recombinant human BMP-2 (rh-BMP-2) and dentin sialoprotein (rh-DSP) on growth and differentiation in human cementoblasts and determine the underlying signal transduction mechanism. Compared to treatment of cementoblasts with either rh-BMP-2 or rh-DSP alone, the combination of rh-BMP-2 and rh-DSP synergistically increased cell growth, ALP activity, nodule formation and expression of differentiation markers. The differentiation-promoting effect was also observed in periodontal ligament cells and an osteoblastic cell line. Likewise, combination of rh-DSP and rh-BMP-2 increased BMP-2 mRNA expression and Smad1/5/8 phosphorylation, which was blocked by the BMP antagonist noggin. The expression levels of α2β1 integrin and RhoA, as well as the phosphorylation status of FAK and Akt, were increased by the combination of rh-BMP-2 and rh-DSP in a time-dependent manner. In addition, rh-BMP-2 and rh-DSP enhanced expression of Wnt ligands, β-catenin activation and GSK-3β phosphorylation, all of which were inhibited by the Wnt receptor antagonist DKK1. Furthermore, treatment with rh-DSP plus rh-BMP-2 resulted in phosphorylation of extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK) and p38 and also induced the nuclear translocation of the NF-κB p65 subunit, which was blocked by noggin. This study demonstrates for the first time that rh-DSP and rh-BMP-2 act synergistically, enhancing each other's ability to stimulate cementoblastic cell growth and differentiation in vitro via autocrine BMP, integrin, Wnt/β-catenin, MAP kinase and NF-κB pathways. These results support the therapeutic potential of a combination strategy for aiding periodontal regeneration.

BMP9 Inhibits Proliferation and Metastasis of HER2-Positive SK-BR-3 Breast Cancer Cells through ERK1/2 and PI3K/AKT Pathways

PubMed Central

Ren, Wei; Liu, Yuehong; Wan, Shaoheng; Fei, Chang; Wang, Wei; Chen, Yingying; Zhang, Zhihui; Wang, Ting; Wang, Jinshu; Zhou, Lan; Weng, Yaguang; He, Tongchuan; Zhang, Yan

2014-01-01

Bone morphogenetic protein 9 (BMP9), a member of TGF-β superfamily, is reported to inhibit the growth and migration of prostate cancer, osteosarcoma and triple-negative MDA-MB-231 breast cancer cells. However, little is known about the effect of on the biological behaviors of HER2-positive SK-BR-3 breast cancer cells and the underlying mechanisms. This study aimed to investigate the effects of BMP9 on the proliferation and metastasis of SK-BR-3 cells with BMP9 over-expression or BMP9 down-regulated expression. Results indicated that exogenously expressed BMP9 inhibited the proliferation and metastasis of SK-BR-3 cells while decreased endogenous BMP9 expression in SK-BR-3 cells promoted the proliferation and migration of breast cancer cells in vitro and in vivo. In SK-BR-3 cells with BMP9 over-expression, the phosphorylation of HER2, ERK1/2 and AKT was markedly suppressed and the HER2 expression decreased at both mRNA and protein levels, while opposite results were observed in SK-BR-3 cells with BMP9 knock down. When the phosphorylation of ERK1/2 and PI3K/AKT was inhibited by PD98059 and LY294002, respectively, the decreased proliferation and invasion induced by BMP9 knock down were eliminated. These findings suggest that BMP9 can inhibit the proliferation and metastasis of SK-BR-3 cells via inactivating ERK1/2 and PI3K/AKT signaling pathways. Thus, BMP9 may serve as a useful agent in the treatment of HER-2 positive breast cancer. PMID:24805814

BMP9 inhibits proliferation and metastasis of HER2-positive SK-BR-3 breast cancer cells through ERK1/2 and PI3K/AKT pathways.

PubMed

Ren, Wei; Liu, Yuehong; Wan, Shaoheng; Fei, Chang; Wang, Wei; Chen, Yingying; Zhang, Zhihui; Wang, Ting; Wang, Jinshu; Zhou, Lan; Weng, Yaguang; He, Tongchuan; Zhang, Yan

2014-01-01

Bone morphogenetic protein 9 (BMP9), a member of TGF-β superfamily, is reported to inhibit the growth and migration of prostate cancer, osteosarcoma and triple-negative MDA-MB-231 breast cancer cells. However, little is known about the effect of on the biological behaviors of HER2-positive SK-BR-3 breast cancer cells and the underlying mechanisms. This study aimed to investigate the effects of BMP9 on the proliferation and metastasis of SK-BR-3 cells with BMP9 over-expression or BMP9 down-regulated expression. Results indicated that exogenously expressed BMP9 inhibited the proliferation and metastasis of SK-BR-3 cells while decreased endogenous BMP9 expression in SK-BR-3 cells promoted the proliferation and migration of breast cancer cells in vitro and in vivo. In SK-BR-3 cells with BMP9 over-expression, the phosphorylation of HER2, ERK1/2 and AKT was markedly suppressed and the HER2 expression decreased at both mRNA and protein levels, while opposite results were observed in SK-BR-3 cells with BMP9 knock down. When the phosphorylation of ERK1/2 and PI3K/AKT was inhibited by PD98059 and LY294002, respectively, the decreased proliferation and invasion induced by BMP9 knock down were eliminated. These findings suggest that BMP9 can inhibit the proliferation and metastasis of SK-BR-3 cells via inactivating ERK1/2 and PI3K/AKT signaling pathways. Thus, BMP9 may serve as a useful agent in the treatment of HER-2 positive breast cancer.

Molecular characterization, genomic arrangement, and expression of bmpD, a new member of the bmp class of genes encoding membrane proteins of Borrelia burgdorferi.

PubMed Central

Ramamoorthy, R; Povinelli, L; Philipp M, T

1996-01-01

An expression library made with Borrelia burgdorferi DNA in the vector lambda ZapII was screened with serum from a monkey infected with the Lyme disease agent. This serum killed B. burgdorferi in vitro by an antibody-dependent, complement-mediated mechanism and contained antibodies to at least seven spirochetal antigens, none of which were the major outer surface proteins OspA or OspB. Among several positive clones, a clone containing the B. burgdorferi bmpA gene encoding the immunodominant antigen P39 was obtained. Chromosome walking and DNA sequence analysis permitted the identification of two additional upstream genes homologous to the bmpA gene and its related companion, bmpB. The first of these was the recently characterized bmpC gene, and adjacent to it was the fourth and new member of this class, which has been designated bmpD. The gene product encoded by bmpD is 34l residues long, contains a signal sequence with a potential signal peptidase II cleavage site, and has 26% identity with TmpC of Treponema pallidum. Southern blotting confirmed the tandem arrangement of all four bmp genes in the chromosome of B. burgdorferi JD1. However, Northern (RNA) blotting revealed that bmpD is expressed as a monocistronic transcript, which indicates that it is not part of an operon at the bmp locus. The bmpD gene was found to be conserved in representative members of the three species of the B. burgdorferi sensu lato complex, suggesting that it serves an important biological function in the spirochete. PMID:8606088

Complication Rates in Posterior Lumbar Interbody Fusion (PLIF) Surgery With Human Bone Morphogenetic Protein 2: Medicare Population.

PubMed

Alobaidaan, Raed; Cohen, Jeremiah R; Lord, Elizabeth L; Buser, Zorica; Yoon, S Tim; Youssef, Jim A; Park, Jong-Beom; Brodke, Darrel S; Wang, Jeffrey C; Meisel, Hans-Joerg

2017-12-01

Retrospective cohort study among Medicare beneficiaries who underwent posterior lumbar interbody fusion (PLIF) surgery. To identify the complication rates associated with the use of bone morphogenetic protein 2 (BMP2) in PLIF. Human BMP2 is commonly used in the "off-label" manner for various types of spine fusion procedures, including PLIF. However, recent studies have reported potential complications associated with the recombinant human BMP2 (rhBMP2) use in the posterior approach. Medicare records within the PearlDiver database were queried for patients undergoing PLIF procedure with and without rhBMP2 between 2005 and 2010. We evaluated complications within 1 year postoperatively. Chi-square was used to compare the complication rates between the 2 groups. A total of 8609 patients underwent PLIF procedure with or without rhBMP2. Individual complication rates in the rhBMP2 group ranged from 0.45% to 7.68% compared with 0.65% to 10.99 in the non-rhBMP2 group. Complication rates for cardiac, pulmonary, lumbosacral neuritis, infection, wound, and urinary tract (include acute kidney failure and post-operative complications) were significantly lower in the rhBMP2 group ( P < .05). There was no difference in the rates of central nervous system complications or radiculitis between the 2 groups. Our data showed that the patients who received rhBMP2 had lower complication rates compared to the non-rhBMP2 group. However, use of rhBMP2 was associated with a higher rate of pseudarthrosis. We did not observe any difference in radiculitis and central nervous system complications between the groups.

Complication Rates in Posterior Lumbar Interbody Fusion (PLIF) Surgery With Human Bone Morphogenetic Protein 2: Medicare Population

PubMed Central

Alobaidaan, Raed; Cohen, Jeremiah R.; Lord, Elizabeth L.; Yoon, S. Tim; Youssef, Jim A.; Park, Jong-Beom; Brodke, Darrel S.; Wang, Jeffrey C.; Meisel, Hans-Joerg

2017-01-01

Study Design: Retrospective cohort study among Medicare beneficiaries who underwent posterior lumbar interbody fusion (PLIF) surgery. Objective: To identify the complication rates associated with the use of bone morphogenetic protein 2 (BMP2) in PLIF. Human BMP2 is commonly used in the “off-label” manner for various types of spine fusion procedures, including PLIF. However, recent studies have reported potential complications associated with the recombinant human BMP2 (rhBMP2) use in the posterior approach. Methods: Medicare records within the PearlDiver database were queried for patients undergoing PLIF procedure with and without rhBMP2 between 2005 and 2010. We evaluated complications within 1 year postoperatively. Chi-square was used to compare the complication rates between the 2 groups. Results: A total of 8609 patients underwent PLIF procedure with or without rhBMP2. Individual complication rates in the rhBMP2 group ranged from 0.45% to 7.68% compared with 0.65% to 10.99 in the non-rhBMP2 group. Complication rates for cardiac, pulmonary, lumbosacral neuritis, infection, wound, and urinary tract (include acute kidney failure and post-operative complications) were significantly lower in the rhBMP2 group (P < .05). There was no difference in the rates of central nervous system complications or radiculitis between the 2 groups. Conclusion: Our data showed that the patients who received rhBMP2 had lower complication rates compared to the non-rhBMP2 group. However, use of rhBMP2 was associated with a higher rate of pseudarthrosis. We did not observe any difference in radiculitis and central nervous system complications between the groups. PMID:29238641

Postoperative Complications Associated With rhBMP2 Use in Posterior/Posterolateral Lumbar Fusion

PubMed Central

Esmail, Nabil; Buser, Zorica; Cohen, Jeremiah R.; Brodke, Darrel S.; Meisel, Hans-Joerg; Park, Jong-Beom; Youssef, Jim A.; Wang, Jeffrey C.; Yoon, S. Tim

2017-01-01

Study Design: Retrospective database review. Objective: Posterior/posterolateral lumbar fusion (PLF) is an effective treatment for a variety of spinal disorders; however, variations in surgical technique have different complication profiles. The aim of our study was to quantify the frequency of various complications in patients undergoing PLF with and without human recombinant bone morphogenetic protein 2 (rhBMP2). Methods: We queried the orthopedic subset of the Medicare database (PearlDiver) between 2005 and 2011 for patients undergoing PLF procedures with and without rhBMP2. Complication and reoperation rates were analyzed within 1 year of the index procedure. Complications assessed include: acute renal failure, deep vein thrombosis, dural tear, hematoma, heterotopic ossification, incision and drainage, cardiac complications, nervous system complications, osteolysis, pneumonia, pseudarthrosis, pulmonary embolism, radiculopathy, respiratory complications, sepsis, urinary retention, urinary tract infection, mechanical, and wound complications. Chi-square analysis was used to calculate the complication differences between the groups. Results: Our data revealed higher overall complication rates in patients undergoing PLF with rhBMP2 versus no_rhBMP2 (76.9% vs 68.8%, P < .05). Stratified by gender, rhBMP2 males had higher rates of mechanical complications, pseudarthrosis, and reoperations compared with no_rhBMP2 males (P < .05), whereas rhBMP2 females had higher rates of pseudarthrosis, urinary tract infection, and urinary retention compared with no_rhBMP2 females (P < .05). Conclusion: Our data revealed higher overall complication rates in PLF patients given rhBMP2 compared with no_rhBMP2. Furthermore, our data suggests that rhBMP2-associated complications may be gender specific. PMID:29662744

Hyaline cartilage regeneration by combined therapy of microfracture and long-term bone morphogenetic protein-2 delivery.

PubMed

Yang, Hee Seok; La, Wan-Geun; Bhang, Suk Ho; Kim, Hak-Jun; Im, Gun-Il; Lee, Haeshin; Park, Jung-Ho; Kim, Byung-Soo

2011-07-01

Microfracture of cartilage induces migration of bone-marrow-derived mesenchymal stem cells. However, this treatment often results in fibrocartilage regeneration. Growth factors such as bone morphogenetic protein (BMP)-2 induce the differentiation of bone-marrow-derived mesenchymal stem cells into chondrocytes, which can be used for hyaline cartilage regeneration. Here, we tested the hypothesis that long-term delivery of BMP-2 to cartilage defects subjected to microfracture results in regeneration of high-quality hyaline-like cartilage, as opposed to short-term delivery of BMP-2 or no BMP-2 delivery. Heparin-conjugated fibrin (HCF) and normal fibrin were used as carriers for the long- and short-term delivery of BMP-2, respectively. Rabbit articular cartilage defects were treated with microfracture combined with one of the following: no treatment, fibrin, short-term delivery of BMP-2, HCF, or long-term delivery of BMP-2. Eight weeks after treatment, histological analysis revealed that the long-term delivery of BMP-2 group (microfracture + HCF + BMP-2) showed the most staining with alcian blue. A biochemical assay, real-time polymerase chain reaction assay and Western blot analysis all revealed that the long-term delivery of BMP-2 group had the highest glucosaminoglycan content as well as the highest expression level of collagen type II. Taken together, the long-term delivery of BMP-2 to cartilage defects subjected to microfracture resulted in regeneration of hyaline-like cartilage, as opposed to short-term delivery or no BMP-2 delivery. Therefore, this method could be more convenient for hyaline cartilage regeneration than autologous chondrocyte implantation due to its less invasive nature and lack of cell implantation.

Bone Morphogenetic Protein 15 in the Pro-Mature Complex Form Enhances Bovine Oocyte Developmental Competence

PubMed Central

Sudiman, Jaqueline; Sutton-McDowall, Melanie L.; Ritter, Lesley J.; White, Melissa A.; Mottershead, David G.; Thompson, Jeremy G.; Gilchrist, Robert B.

2014-01-01

Developmental competence of in vitro matured (IVM) oocytes needs to be improved and this can potentially be achieved by adding recombinant bone morphogenetic protein 15 (BMP15) or growth differentiation factor (GDF9) to IVM. The aim of this study was to determine the effect of a purified pro-mature complex form of recombinant human BMP15 versus the commercially available bioactive forms of BMP15 and GDF9 (both isolated mature regions) during IVM on bovine embryo development and metabolic activity. Bovine cumulus oocyte complexes (COCs) were matured in vitro in control medium or treated with 100 ng/ml pro-mature BMP15, mature BMP15 or mature GDF9 +/− FSH. Metabolic measures of glucose uptake and lactate production from COCs and autofluorescence of NAD(P)H, FAD and GSH were measured in oocytes after IVM. Following in vitro fertilisation and embryo culture, day 8 blastocysts were stained for cell numbers. COCs matured in medium +/− FSH containing pro-mature BMP15 displayed significantly improved blastocyst development (57.7±3.9%, 43.5±4.2%) compared to controls (43.3±2.4%, 28.9±3.7%) and to mature GDF9+FSH (36.1±3.0%). The mature form of BMP15 produced intermediate levels of blastocyst development; not significantly different to control or pro-mature BMP15 levels. Pro-mature BMP15 increased intra-oocyte NAD(P)H, and reduced glutathione (GSH) levels were increased by both forms of BMP15 in the absence of FSH. Exogenous BMP15 in its pro-mature form during IVM provides a functional source of oocyte-secreted factors to improve bovine blastocyst development. This form of BMP15 may prove useful for improving cattle and human artificial reproductive technologies. PMID:25058588

Bone morphogenetic protein 15 in the pro-mature complex form enhances bovine oocyte developmental competence.

PubMed

Sudiman, Jaqueline; Sutton-McDowall, Melanie L; Ritter, Lesley J; White, Melissa A; Mottershead, David G; Thompson, Jeremy G; Gilchrist, Robert B

2014-01-01

Developmental competence of in vitro matured (IVM) oocytes needs to be improved and this can potentially be achieved by adding recombinant bone morphogenetic protein 15 (BMP15) or growth differentiation factor (GDF9) to IVM. The aim of this study was to determine the effect of a purified pro-mature complex form of recombinant human BMP15 versus the commercially available bioactive forms of BMP15 and GDF9 (both isolated mature regions) during IVM on bovine embryo development and metabolic activity. Bovine cumulus oocyte complexes (COCs) were matured in vitro in control medium or treated with 100 ng/ml pro-mature BMP15, mature BMP15 or mature GDF9 +/- FSH. Metabolic measures of glucose uptake and lactate production from COCs and autofluorescence of NAD(P)H, FAD and GSH were measured in oocytes after IVM. Following in vitro fertilisation and embryo culture, day 8 blastocysts were stained for cell numbers. COCs matured in medium +/- FSH containing pro-mature BMP15 displayed significantly improved blastocyst development (57.7±3.9%, 43.5±4.2%) compared to controls (43.3±2.4%, 28.9±3.7%) and to mature GDF9+FSH (36.1±3.0%). The mature form of BMP15 produced intermediate levels of blastocyst development; not significantly different to control or pro-mature BMP15 levels. Pro-mature BMP15 increased intra-oocyte NAD(P)H, and reduced glutathione (GSH) levels were increased by both forms of BMP15 in the absence of FSH. Exogenous BMP15 in its pro-mature form during IVM provides a functional source of oocyte-secreted factors to improve bovine blastocyst development. This form of BMP15 may prove useful for improving cattle and human artificial reproductive technologies.

Success of riparian restoration projects in the mountains, piedmont, and coastal plain of Virginia

Treesearch

Benjamin N. Bradburn; W. Michael Aust; Matthew B. Carroll; Dean Cumbia; Jerre Creighton

2010-01-01

Forested riparian buffers are a Best Management Practice (BMP) for protection of water quality and for habitat. Since the 1990s, conservation agencies in Virginia have been involved in establishment of riparian buffers under the auspices of programs such as the Conservation Reserve Enhancement Program (CREP). Although CREP was established for protection of water...

Unbiased RNAi screen for hepcidin regulators links hepcidin suppression to proliferative Ras/RAF and nutrient-dependent mTOR signaling

PubMed Central

Mleczko-Sanecka, Katarzyna; Roche, Franziska; Rita da Silva, Ana; Call, Debora; D’Alessio, Flavia; Ragab, Anan; Lapinski, Philip E.; Ummanni, Ramesh; Korf, Ulrike; Oakes, Christopher; Damm, Georg; D’Alessandro, Lorenza A.; Klingmüller, Ursula; King, Philip D.; Boutros, Michael; Hentze, Matthias W.

2014-01-01

The hepatic hormone hepcidin is a key regulator of systemic iron metabolism. Its expression is largely regulated by 2 signaling pathways: the “iron-regulated” bone morphogenetic protein (BMP) and the inflammatory JAK-STAT pathways. To obtain broader insights into cellular processes that modulate hepcidin transcription and to provide a resource to identify novel genetic modifiers of systemic iron homeostasis, we designed an RNA interference (RNAi) screen that monitors hepcidin promoter activity after the knockdown of 19 599 genes in hepatocarcinoma cells. Interestingly, many of the putative hepcidin activators play roles in signal transduction, inflammation, or transcription, and affect hepcidin transcription through BMP-responsive elements. Furthermore, our work sheds light on new components of the transcriptional machinery that maintain steady-state levels of hepcidin expression and its responses to the BMP- and interleukin-6–triggered signals. Notably, we discover hepcidin suppression mediated via components of Ras/RAF MAPK and mTOR signaling, linking hepcidin transcriptional control to the pathways that respond to mitogen stimulation and nutrient status. Thus using a combination of RNAi screening, reverse phase protein arrays, and small molecules testing, we identify links between the control of systemic iron homeostasis and critical liver processes such as regeneration, response to injury, carcinogenesis, and nutrient metabolism. PMID:24385536

Hydraulic characteristics of low-impact development practices in northeastern Ohio, 2008–2010

USGS Publications Warehouse

Darner, Robert A.; Dumouchelle, Denise H.

2011-01-01

Low-impact development (LID) is an approach to managing stormwater as near to its source as possible; this is accomplished by minimizing impervious surfaces and promoting more natural infiltration and evapotranspiration than is typically associated with developed areas. Two newly constructed LID sites in northeastern Ohio were studied to document their hydraulic characteristics. A roadside best-management practice (BMP) was constructed by replacing about 1,400 linear feet of existing ditches with a bioswale/rain garden BMP consisting of a grassed swale interspersed with rain-garden/overflow structures. The site was monitored in 2008, 2009, and 2010. Although some overflows occurred, numerous precipitation events exceeding the 0.75-inch design storm did not result in overflows. A second study site consists of an 8,200-square-foot parking lot made of a pervious pavers and a rain garden that receives runoff from the roof of a nearby commercial building. A comparison of data from 2009 and 2010 indicates that the median runoff volume in 2010 decreased relative to 2009. The centroid lag times (time difference between centroid of precipitation and centroid of flow) decreased in 2010, most likely due to more intense, shorter duration precipitation events and maturation of the rain garden. Additional data could help quantify the relation between meteorological variables and BMP efficiency.

Intervariability and intravariability of bone morphogenetic proteins in commercially available demineralized bone matrix products.

PubMed

Bae, Hyun W; Zhao, Li; Kanim, Linda E A; Wong, Pamela; Delamarter, Rick B; Dawson, Edgar G

2006-05-20

Enzyme-linked immunosorbent assay was used to detect bone morphogenetic proteins (BMPs) 2, 4, and 7 in 9 commercially available ("off the shelf") demineralized bone matrix (DBM) product formulations using 3 different manufacturer's production lots of each DBM formulation. To evaluate and compare the quantity of BMPs among several different DBM formulations (inter-product variability), as well as examine the variability of these proteins in different production lots within the same DBM formulation (intra-product variability). DBMs are commonly used to augment available bone graft in spinal fusion procedures. Surgeons are presented with an ever-increasing variety of commercially available human DBMs from which to choose. Yet, there is limited information on a specific DBM product's osteoinductive efficacy, potency, and constancy. There were protein extracts from each DBM sample separately dialyzed 4 times against distilled water at 4 degrees C for 48 hours. The amount of BMP-2, BMP-4, and BMP-7 was determined using enzyme-linked immunosorbent assay. RESULTS.: The concentrations of detected BMP-2 and BMP-7 were low for all DBM formulations, only nanograms of BMP were extracted from each gram of DBM (20.2-120.6 ng BMP-2/g DBM product; 54.2-226.8 ng BMP-7/g DBM). The variability of BMP concentrations among different lots of the same DBM formulation, intra-product variability, was higher than the variability of concentrations among different DBM formulations, inter-product variability (coefficient of variation range BMP-2 [16.34% to 76.01%], P < 0.01; BMP-7 [3.71% to 82.08%], P < 0.001). BMP-4 was undetectable. The relative quantities of BMPs in DBMs are low, in the order of 1 x 10(-9) g of BMP/g of DBM. There is higher variability in concentration of BMPs among 3 different lots of the same DBM formulation than among different DBM formulations. This variability questions DBM products' reliability and, possibly, efficacy in providing consistent osteoinduction.

Promising efficacy of Escherichia coli recombinant human bone morphogenetic protein-2 in collagen sponge for ectopic and orthotopic bone formation and comparison with mammalian cell recombinant human bone morphogenetic protein-2.

PubMed

Kim, In Sook; Lee, Eui Nam; Cho, Tae Hyung; Song, Yun Mi; Hwang, Soon Jung; Oh, Ji Hye; Park, Eun Kyung; Koo, Tai Young; Seo, Young-Kwon

2011-02-01

Nonglycosylated recombinant human bone morphogenetic protein (rhBMP)-2 prepared in Escherichia coli (E. coli rhBMP-2) has recently been considered as an alternative to mammalian cell rhBMP-2. However, its clinical use is still limited owing to lack of evidence for osteogenic activity comparable with that of mammalian cell rhBMP-2 via microcomputed tomography-based analysis. Therefore, this study aimed to evaluate the ability of E. coli rhBMP-2 in absorbable collagen sponge to form ectopic and orthotopic bone and to compare it to that of mammalian rhBMP-2. In vitro investigation was performed to study osteoblast differentiation of human mesenchymal stromal cells. Both types of rhBMP-2 enhanced proliferation, alkaline phosphatase activity, and matrix mineralization of human mesenchymal stromal cells at similar levels. Similar tendencies were observed in microcomputed tomography analysis, which determined bone volume, fractional bone volume, trabecular thickness, trabecular separation, bone mineral density, and other characteristics. Histology from an in vivo osteoinductivity test and from a rat calvarial defect model demonstrated a dose-dependent increase in local bone formation. The E. coli rhBMP-2 group (5 μg) not only induced complete regeneration of an 8-mm critical-sized defect at 4 weeks, but also led to new bone with the same bone mineral density as normal bone at 8 weeks, with the same efficiency as that of mammalian cell rhBMP-2 (5 μg). These uniformly favorable results provide evidence that the osteogenic activity of E. coli rhBMP-2 is not inferior to that of mammalian cell rhBMP-2 despite its low solubility and lack of gylcosylation. These results suggest that the application of E. coli rhBMP-2 in absorbable collagen sponge may be a promising equivalent to mammalian cell rhBMP-2 in bone tissue engineering.

Bmp2 conditional knockout in osteoblasts and endothelial cells does not impair bone formation after injury or mechanical loading in adult mice

PubMed Central

McKenzie, Jennifer A.; Buettmann, Evan G.; Gardner, Michael J.; Silva, Matthew J.

2015-01-01

Post-natal osteogenesis after mechanical trauma or stimulus occurs through either endochondral healing, intramembranous healing or lamellar bone formation. Bone morphogenetic protein 2 (BMP2) is up-regulated in each of these osteogenic processes and is expressed by a variety of cells including osteoblasts and vascular cells. It is known that genetic knockout of Bmp2 in all cells or in osteo-chondroprogenitor cells completely abrogates endochondral healing after full fracture. However, the importance of BMP2 from differentiated osteoblasts and endothelial cells is not known. Moreover, the importance of BMP2 in non-endochondral bone formation such as intramembranous healing or lamellar bone formation is not known. Using inducible and tissue-specific Cre-lox mediated targeting of Bmp2 in adult (10–24 week old) mice, we assessed the role of BMP2 expression globally, by osteoblasts, and by vascular endothelial cells in endochondral healing, intramembranous healing and lamellar bone formation. These three osteogenic processes were modeled using full femur fracture, ulnar stress fracture, and ulnar non-damaging cyclic loading, respectively. Our results confirmed the requirement of BMP2 for endochondral fracture healing, as mice in which Bmp2 was knocked out in all cells prior to fracture failed to form a callus. Targeted deletion of Bmp2 in osteoblasts (osterix-expressing) or vascular endothelial cells (vascular endothelial cadherin-expressing) did not impact fracture healing in any way. Regarding non-endochondral bone formation, we found that BMP2 is largely dispensable for intramembranous bone formation after stress fracture and also not required for lamellar bone formation induced by mechanical loading. Taken together our results indicate that osteoblasts and endothelial cells are not a critical source of BMP2 in endochondral fracture healing, and that non-endochondral bone formation in the adult mouse is not as critically dependent on BMP2. PMID:26344756

Bmp signaling mediates endoderm pouch morphogenesis by regulating Fgf signaling in zebrafish.

PubMed

Lovely, C Ben; Swartz, Mary E; McCarthy, Neil; Norrie, Jacqueline L; Eberhart, Johann K

2016-06-01

The endodermal pouches are a series of reiterated structures that segment the pharyngeal arches and help pattern the vertebrate face. Multiple pathways regulate the complex process of endodermal development, including the Bone morphogenetic protein (Bmp) pathway. However, the role of Bmp signaling in pouch morphogenesis is poorly understood. Using genetic and chemical inhibitor approaches, we show that pouch morphogenesis requires Bmp signaling from 10-18 h post-fertilization, immediately following gastrulation. Blocking Bmp signaling during this window results in morphological defects to the pouches and craniofacial skeleton. Using genetic chimeras we show that Bmp signals directly to the endoderm for proper morphogenesis. Time-lapse imaging and analysis of reporter transgenics show that Bmp signaling is necessary for pouch outpocketing via the Fibroblast growth factor (Fgf) pathway. Double loss-of-function analyses demonstrate that Bmp and Fgf signaling interact synergistically in craniofacial development. Collectively, our analyses shed light on the tissue and signaling interactions that regulate development of the vertebrate face. © 2016. Published by The Company of Biologists Ltd.

Implant Composed of Demineralized Bone and Mesenchymal Stem Cells Genetically Modified with AdBMP2/AdBMP7 for the Regeneration of Bone Fractures in Ovis aries

PubMed Central

Hernandez-Hurtado, Adelina A.; Lara-Arias, Jorge; Romero-Diaz, Viktor J.; Abrego-Guerra, Adalberto; Vilchez-Cavazos, Jose F.; Elizondo-Riojas, Guillermo; Martinez-Rodriguez, Herminia G.; Espinoza-Juarez, Marcela A.; Mendoza Lemus, Oscar F.

2016-01-01

Adipose-derived mesenchymal stem cells (ADMSCs) are inducible to an osteogenic phenotype by the bone morphogenetic proteins (BMPs). This facilitates the generation of implants for bone tissue regeneration. This study evaluated the in vitro osteogenic differentiation of ADMSCs transduced individually and in combination with adenoviral vectors expressing BMP2 and BMP7. Moreover, the effectiveness of the implant containing ADMSCs transduced with the adenoviral vectors AdBMP2/AdBMP7 and embedded in demineralized bone matrix (DBM) was tested in a model of tibial fracture in sheep. This graft was compared to ewes implanted with untransduced ADMSCs embedded in the same matrix and with injured but untreated animals. In vivo results showed accelerated osteogenesis in the group treated with the AdBMP2/AdBMP7 transduced ADMSC graft, which also showed improved restoration of the normal bone morphology. PMID:27818692

Repulsive Guidance Molecule is a structural bridge between Neogenin and Bone Morphogenetic Protein

PubMed Central

Healey, Eleanor G.; Bishop, Benjamin; Elegheert, Jonathan; Bell, Christian H.; Padilla-Parra, Sergi; Siebold, Christian

2015-01-01

Repulsive guidance molecules (RGMs) control crucial processes spanning cell motility, adhesion, immune cell regulation and systemic iron metabolism. RGMs signal via two fundamental signaling cascades: the Neogenin (NEO1) and the Bone Morphogenetic Protein (BMP) pathways. Here, we report crystal structures of the N-terminal domains of all human RGM family members in complex with the BMP ligand BMP2, revealing a novel protein fold and a conserved BMP-binding mode. Our structural and functional data suggest a pH-linked mechanism for RGM-activated BMP signaling and offer a rationale for RGM mutations causing juvenile hemochromatosis. We also determined the ternary BMP2–RGM–NEO1 complex crystal structure, which combined with solution scattering and live-cell super-resolution fluorescence microscopy, indicates BMP-induced clustering of the RGM–NEO1 complex. Our results show how RGM acts as the central hub linking BMP and NEO1 and physically connecting these fundamental signaling pathways. PMID:25938661

A conserved post-transcriptional BMP2 switch in lung cells.

PubMed

Jiang, Shan; Fritz, David T; Rogers, Melissa B

2010-05-15

An ultra-conserved sequence in the bone morphogenetic protein 2 (BMP2) 3' untranslated region (UTR) markedly represses BMP2 expression in non-transformed lung cells. In contrast, the ultra-conserved sequence stimulates BMP2 expression in transformed lung cells. The ultra-conserved sequence functions as a post-transcriptional cis-regulatory switch. A common single-nucleotide polymorphism (SNP, rs15705, +A1123C), which has been shown to influence human morphology, disrupts a conserved element within the ultra-conserved sequence and altered reporter gene activity in non-transformed lung cells. This polymorphism changed the affinity of the BMP2 RNA for several proteins including nucleolin, which has an increased affinity for the C allele. Elevated BMP2 synthesis is associated with increased malignancy in mouse models of lung cancer and poor lung cancer patient prognosis. Understanding the cis- and trans-regulatory factors that control BMP2 synthesis is relevant to the initiation or progression of pathologies associated with abnormal BMP2 levels. (c) 2010 Wiley-Liss, Inc.

In vitro and in vivo evaluation of osteoinductivity and bone fusion ability of an activin a/BMP2 chimera (AB204): a comparison study between AB204 and rhBMP-2.

PubMed

Zheng, Guang Bin; Lee, Jae Hyup; Jin, Yuan-Zhe

2017-12-01

This study compared osteoinductivity and osteogenic capacity between AB204 and rhBMP-2 using hMSCs in vitro and a beagle's posterolateral spinal fusion model. Cultured hMSCs were treated with AB204 or rhBMP-2 with low to high doses. Three male beagles were performed posterolateral spinal fusion with biphasic calcium phosphate (2 ml) + AB204 or rhBMP-2 (20, 50 or 200 µg). They were euthanized after 8 weeks. The fusion rate and bone formation of spine samples were examined. AB204 had higher alkaline phosphatase activity, mineralization and osteogenic-related gene expression than rhBMP-2. Fusion rates in all rhBMP-2 groups were 0. They were 100% for 50 μg and 200 μg AB204 groups. Therefore, AB204 showed higher osteogenicity than rhBMP-2. It could be a better bone graft substitute.

Bmp signaling mediates endoderm pouch morphogenesis by regulating Fgf signaling in zebrafish

PubMed Central

Swartz, Mary E.; McCarthy, Neil; Norrie, Jacqueline L.; Eberhart, Johann K.

2016-01-01

The endodermal pouches are a series of reiterated structures that segment the pharyngeal arches and help pattern the vertebrate face. Multiple pathways regulate the complex process of endodermal development, including the Bone morphogenetic protein (Bmp) pathway. However, the role of Bmp signaling in pouch morphogenesis is poorly understood. Using genetic and chemical inhibitor approaches, we show that pouch morphogenesis requires Bmp signaling from 10-18 h post-fertilization, immediately following gastrulation. Blocking Bmp signaling during this window results in morphological defects to the pouches and craniofacial skeleton. Using genetic chimeras we show that Bmp signals directly to the endoderm for proper morphogenesis. Time-lapse imaging and analysis of reporter transgenics show that Bmp signaling is necessary for pouch outpocketing via the Fibroblast growth factor (Fgf) pathway. Double loss-of-function analyses demonstrate that Bmp and Fgf signaling interact synergistically in craniofacial development. Collectively, our analyses shed light on the tissue and signaling interactions that regulate development of the vertebrate face. PMID:27122171

Interactions between Bmp-4 and Msx-1 act to restrict gene expression to odontogenic mesenchyme.

PubMed

Tucker, A S; Al Khamis, A; Sharpe, P T

1998-08-01

Tooth development is regulated by a reciprocal series of epithelial-mesenchymal interactions. Bmp4 has been identified as a candidate signalling molecule in these interactions, initially as an epithelial signal and then later at the bud stage as a mesenchymal signal (Vainio et al. [1993] Cell 75:45-58). A target gene for Bmp4 signalling is the homeobox gene Msx-1, identified by the ability of recombinant Bmp4 protein to induce expression in mesenchyme. There is, however, no evidence that Bmp4 is the endogenous inducer of Msx-1 expression. Msx-1 and Bmp-4 show dynamic, interactive patterns of expression in oral epithelium and ectomesenchyme during the early stages of tooth development. In this study, we compare the temporal and spatial expression of these two genes to determine whether the changing expression patterns of these genes are consistent with interactions between the two molecules. We show that changes in Bmp-4 expression precede changes in Msx-1 expression. At embryonic day (E)10.5-E11.0, expression patterns are consistent with BMP4 from the epithelium, inducing or maintaining Msx-1 in underlying mesenchyme. At E11.5, Bmp-4 expression shifts from epithelium to mesenchyme and is rapidly followed by localised up-regulation of Msx-1 expression at the sites of Bmp-4 expression. Using cultured explants of developing mandibles, we confirm that exogenous BMP4 is capable of replacing the endogenous source in epithelium and inducing Msx-1 gene expression in mesenchyme. By using noggin, a BMP inhibitor, we show that endogenous Msx-1 expression can be inhibited at E10.5 and E11.5, providing the first evidence that endogenous Bmp-4 from the epithelium is responsible for regulating the early spatial expression of Msx-1. We also show that the mesenchymal shift in Bmp-4 is responsible for up-regulating Msx-1 specifically at the sites of future tooth formation. Thus, we establish that a reciprocal series of interactions act to restrict expression of both genes to future sites of tooth formation, creating a positive feedback loop that maintains expression of both genes in tooth mesenchymal cells.

A polymorphism in a conserved posttranscriptional regulatory motif alters bone morphogenetic protein 2 (BMP2) RNA:protein interactions.

PubMed

Fritz, David T; Jiang, Shan; Xu, Junwang; Rogers, Melissa B

2006-07-01

The bone morphogenetic protein (BMP)2 gene has been genetically linked to osteoporosis and osteoarthritis. We have shown that the 3'-untranslated regions (UTR) of BMP2 genes from mammals to fishes are extraordinarily conserved. This indicates that the BMP2 3'-UTR is under stringent selective pressure. We present evidence that the conserved region is a strong posttranscriptional regulator of BMP2 expression. Polymorphisms in cis-regulatory elements have been proven to influence susceptibility to a growing number of diseases. A common single nucleotide polymorphism (SNP) disrupts a putative posttranscriptional regulatory motif, an AU-rich element, within the BMP2 3'-UTR. The affinity of specific proteins for the rs15705 SNP sequence differs from their affinity for the normal human sequence. More importantly, the in vitro decay rate of RNAs with the SNP is higher than that of RNAs with the normal sequence. Such changes in mRNA:protein interactions may influence the posttranscriptional mechanisms that control BMP2 gene expression. The consequent alterations in BMP2 protein levels may influence the development or physiology of bone or other BMP2-influenced tissues.

Effects of Bone Morphogenic Proteins on Engineered Cartilage

NASA Technical Reports Server (NTRS)

Gooch, Keith, J.; Blunk, Torsten; Courter, Donald L.; Sieminski, Alisha; Vunjak-Novakovic, Gordana; Freed, Lisa E.

2007-01-01

A report describes experiments on the effects of bone morphogenic proteins (BMPs) on engineered cartilage grown in vitro. In the experiments, bovine calf articular chondrocytes were seeded onto biodegradable polyglycolic acid scaffolds and cultured in, variously, a control medium or a medium supplemented with BMP-2, BMP-12, or BMP-13 in various concentrations. Under all conditions investigated, cell-polymer constructs cultivated for 4 weeks macroscopically and histologically resembled native cartilage. At a concentration of 100 ng/mL, BMP-2, BMP-12, or BMP-13 caused (1) total masses of the constructs to exceed those of the controls by 121, 80, or 62 percent, respectively; (2) weight percentages of glycosaminoglycans in the constructs to increase by 27, 18, or 15, respectively; and (3) total collagen contents of the constructs to decrease to 63, 89, or 83 percent of the control values, respectively. BMP-2, but not BMP-12 or BMP-13, promoted chondrocyte hypertrophy. These observations were interpreted as suggesting that the three BMPs increase the growth rates and modulate the compositions of engineered cartilage. It was also concluded that in vitro engineered cartilage is a suitable system for studying effects of BMPs on chondrogenesis in a well-defined environment.

Negative regulation of BMP signaling by the ski oncoprotein.

PubMed

Luo, Kunxin

2003-01-01

The bone morphogenetic proteins (BMPs) play important roles in the regulation of multiple aspects of vertebrate development. BMPs signal through the cell surface receptors and downstream Smad molecules. Upon stimulation with BMP, Smad1, Smad5, and Smad8 are phosphorylated by the activated BMP receptors, form a complex with Smad4, and translocate into the nucleus, where they regulate the expression of BMP target genes. The activity of this signal pathway can be modulated both by extracellular factors that regulate the binding of BMPs to the receptor and by intracellular proteins that interact with the Smad proteins. We have shown that Ski is an important negative regulator of the Smad proteins. Ski can bind to the BMP-Smad protein complexes in response to BMP and repress their ability to activate BMP target genes through disruption of a functional Smad complex and through recruitment of transcriptional co-repressors. The antagonism of BMP signaling by Ski results in neural specification in Xenopus embryos and inhibition of osteoblast differentiation in mouse bone-marrow stromal progenitor cells. This ability to modulate BMP signaling by Ski may play an important role in the regulation of craniofacial, neuronal, and skeletal muscle development.

Trends, Costs, and Complications of Anterior Cervical Discectomy and Fusion With and Without Bone Morphogenetic Protein in the United States Medicare Population.

PubMed

Lord, Elizabeth L; Cohen, Jeremiah R; Buser, Zorica; Meisel, Hans-Joerg; Brodke, Darrel S; Yoon, S Tim; Youssef, Jim A; Wang, Jeffrey C; Park, Jong-Beom

2017-10-01

Retrospective database review. After the Food and Drug Administration approved bone morphogenetic protein-2 (BMP) in 2002, BMP was used off-label in the cervical spine to increase bone growth and bony fusion. Since then, concerns have been raised regarding complication rates and safety. This study was conducted to examine the use of BMP in anterior cervical discectomy and fusion (ACDF) in the Medicare population and to determine risk of complications and associated costs within 90 days of surgery. Patients who underwent ACDF were identified using Current Procedural Terminology (CPT) and International Classification of Diseases, Ninth Revision Procedure codes (ICD9-P). Complications were identified using ICD9 diagnostic codes. Charges were calculated as amount billed, and reimbursements were calculated as amounts paid by Medicare. Data for these analyses came from a nationwide claims database. A total of 215 047 patients were identified who had ACDF from 2005 to 2011. For the majority of the procedures (89.0%), BMP was not used. BMP use rose from 11.84% in 2005 to a peak of 16.73% in 2007 before decreasing to 12.01% in 2011. BMP was used 16% more in women than men. BMP use was the highest in the West (13.6%) followed by Midwest (11.8%), South (10.6%), and Northeast (7.5%). There was a higher overall complication rate in the BMP group (2.1%) compared with the non-BMP group (1.9%) (odds ratio [OR] = 1.11, 95% CI = 1.01-1.22). The BMP group also had a higher rate of wound complications (0.98% vs 0.76%, OR = 1.29, 95% CI = 1.12-1.48). In this study population, there was no difference in dysphagia/hoarseness, neurologic, medical, or other complications. During the 90-day perioperative period, BMP surgeries were charged at 17.6% higher than non-BMP surgeries. The use of BMP in ACDF in the Medicare population has decreased since a peak in 2007. The rate of wound and overall complications for BMP use with ACDF was higher than without. Our results regarding dysphagia/hoarseness did not show a statistically meaningful difference, which is in contrast with many other studies. Charges associated with BMP use were higher during the 90-day perioperative period.

Trends, Costs, and Complications of Anterior Cervical Discectomy and Fusion With and Without Bone Morphogenetic Protein in the United States Medicare Population

PubMed Central

Cohen, Jeremiah R.; Buser, Zorica; Meisel, Hans-Joerg; Brodke, Darrel S.; Yoon, S. Tim; Youssef, Jim A.; Wang, Jeffrey C.; Park, Jong-Beom

2017-01-01

Study Design: Retrospective database review. Objectives: After the Food and Drug Administration approved bone morphogenetic protein–2 (BMP) in 2002, BMP was used off-label in the cervical spine to increase bone growth and bony fusion. Since then, concerns have been raised regarding complication rates and safety. This study was conducted to examine the use of BMP in anterior cervical discectomy and fusion (ACDF) in the Medicare population and to determine risk of complications and associated costs within 90 days of surgery. Methods: Patients who underwent ACDF were identified using Current Procedural Terminology (CPT) and International Classification of Diseases, Ninth Revision Procedure codes (ICD9-P). Complications were identified using ICD9 diagnostic codes. Charges were calculated as amount billed, and reimbursements were calculated as amounts paid by Medicare. Data for these analyses came from a nationwide claims database. Results: A total of 215 047 patients were identified who had ACDF from 2005 to 2011. For the majority of the procedures (89.0%), BMP was not used. BMP use rose from 11.84% in 2005 to a peak of 16.73% in 2007 before decreasing to 12.01% in 2011. BMP was used 16% more in women than men. BMP use was the highest in the West (13.6%) followed by Midwest (11.8%), South (10.6%), and Northeast (7.5%). There was a higher overall complication rate in the BMP group (2.1%) compared with the non-BMP group (1.9%) (odds ratio [OR] = 1.11, 95% CI = 1.01-1.22). The BMP group also had a higher rate of wound complications (0.98% vs 0.76%, OR = 1.29, 95% CI = 1.12-1.48). In this study population, there was no difference in dysphagia/hoarseness, neurologic, medical, or other complications. During the 90-day perioperative period, BMP surgeries were charged at 17.6% higher than non-BMP surgeries. Conclusions: The use of BMP in ACDF in the Medicare population has decreased since a peak in 2007. The rate of wound and overall complications for BMP use with ACDF was higher than without. Our results regarding dysphagia/hoarseness did not show a statistically meaningful difference, which is in contrast with many other studies. Charges associated with BMP use were higher during the 90-day perioperative period. PMID:28989837

Aggressive melanoma cells escape from BMP7-mediated autocrine growth inhibition through coordinated Noggin upregulation

PubMed Central

Hsu, Mei-Yu; Rovinsky, Sherry; Lai, Chiou-Yan; Qasem, Shadi; Liu, Xiaoming; How, Joan; Engelhardt, John F.; Murphy, George F.

2009-01-01

Bone morphogenetic proteins (BMPs) are members of the TGF-β superfamily responsible for mediating a diverse array of cellular functions both during embryogenesis and in adult life. Previously, we reported that upregulation of BMP7 in human melanoma correlates with tumor progression. However, melanoma cells are either inhibited by or become resistant to BMP7 as a function of tumor progression, with normal melanocytes being most susceptible. Herein, real-time quantitative reverse transcriptase-polymerase chain reactions and Western blotting revealed that the expression of BMP antagonist, Noggin, correlates with resistance to BMP7 in advanced melanoma cells. To test the hypothesis that coordinated upregulation of Noggin protects advanced melanoma cells from autocrine inhibition by BMP7, functional expression of Noggin in susceptible melanoma cells was achieved by adenoviral gene transfer. The Noggin-overexpressing cells exhibited a growth advantage in response to subsequent BMP7 transduction in vitro under anchorage-dependent and -independent conditions, in three-dimensional skin reconstructs, as well as in vivo in severe combined immune-deficiency mice. In concordance, Noggin knockdown by lentiviral shRNA confers sensitivity to BMP7-induced growth inhibition in advanced melanoma cells. Our findings suggest that, like TGF-β, BMP7 acts as an autocrine growth inhibitor in melanocytic cells, and that advanced melanoma cells may escape from BMP7-induced inhibition through concomitant aberrant expression of Noggin. PMID:18560367

Expression of BMP-2 in Vascular Endothelial Cells of Recipient May Predict Delayed Graft Function After Renal Transplantation.

PubMed

Basic-Jukic, Nikolina; Gulin, Marijana; Hudolin, Tvrtko; Kastelan, Zeljko; Katalinic, Lea; Coric, Marijana; Veda, Marija Varnai; Ivkovic, Vanja; Kes, Petar; Jelakovic, Bojan

2016-01-01

Delayed graft function (DGF) is associated with adverse outcomes after renal transplantation. Bone morphogenetic protein-2 (BMP-2) is involved in both endothelial function and immunological events. We compared expression of BMP-2 in epigastric artery of renal transplant recipients with immediate graft function (IGF) and DGF. 79 patients were included in this prospective study. Patients were divided in IGF group (64 patients) and DGF group (15 patients). BMP-2 expression in intima media (BMP2m) and endothelium (BMP2e) of epigastric artery was assessed by immunohistochemistry. Lower intensity of BMP2e staining was recorded in DGF compared to IGF. In DGF patients, 93% had no expression of BMP2e and 7% had 1st grade expression, compared to 45% and 41% in IGF group, respectively (P=0.001) (P<0.001 for no expression and P = 0.015 for 1st grade expression). Patients who had BMP2e staining positive had lower odds for DGF (OR 0.059 [0.007, 0.477]) and this remained significant even after adjustment for donor and recipient variables, cold ischemia time, and immunological matching (OR 0.038 [0.003, 0.492]). Our results demonstrate that BMP-2 expression in endothelial cells of epigastric arteries may predict development of DGF. © 2016 The Author(s) Published by S. Karger AG, Basel.

BMP-driven NRF2 activation in esophageal basal cell differentiation and eosinophilic esophagitis

PubMed Central

Jiang, Ming; Ku, Wei-Yao; Zhou, Zhongren; Dellon, Evan S.; Falk, Gary W.; Nakagawa, Hiroshi; Wang, Mei-Lun; Liu, Kuancan; Wang, Jun; Katzka, David A.; Peters, Jeffrey H.; Lan, Xiaopeng; Que, Jianwen

2015-01-01

Tissue homeostasis requires balanced self-renewal and differentiation of stem/progenitor cells, especially in tissues that are constantly replenished like the esophagus. Disruption of this balance is associated with pathological conditions, including eosinophilic esophagitis (EoE), in which basal progenitor cells become hyperplastic upon proinflammatory stimulation. However, how basal cells respond to the inflammatory environment at the molecular level remains undetermined. We previously reported that the bone morphogenetic protein (BMP) signaling pathway is critical for epithelial morphogenesis in the embryonic esophagus. Here, we address how this pathway regulates tissue homeostasis and EoE development in the adult esophagus. BMP signaling was specifically activated in differentiated squamous epithelium, but not in basal progenitor cells, which express the BMP antagonist follistatin. Previous reports indicate that increased BMP activity promotes Barrett’s intestinal differentiation; however, in mice, basal progenitor cell–specific expression of constitutively active BMP promoted squamous differentiation. Moreover, BMP activation increased intracellular ROS levels, initiating an NRF2-mediated oxidative response during basal progenitor cell differentiation. In both a mouse EoE model and human biopsies, reduced squamous differentiation was associated with high levels of follistatin and disrupted BMP/NRF2 pathways. We therefore propose a model in which normal squamous differentiation of basal progenitor cells is mediated by BMP-driven NRF2 activation and basal cell hyperplasia is promoted by disruption of BMP signaling in EoE. PMID:25774506

Immunohistological Localization of BMP-2, BMP-7, and Their Receptors in Knee Joints with Focal Cartilage Lesions

PubMed Central

Schmal, Hagen; Mehlhorn, Alexander T.; Pilz, Ingo H.; Dovi-Akue, David; Kirchhoff, Christina; Südkamp, Norbert P.; Gerlach, Ulrike; Lohrmann, Christian; Niemeyer, Philipp

2012-01-01

Introduction. Although it is well known that BMP-2 and BMP-7 play significant roles in cartilage metabolism, data about intra-articular expression and localization of these proteins and their receptors in humans are rare. Methods. Biopsies of synovia and debrided cartilage were taken in patients undergoing autologous chondrocyte implantation. Expression of BMP-2, BMP-7, and their receptors BMPR-1A, BMPR-1B and BMPR-2 were semiquantitatively evaluated by immunohistological staining. Results. BMP-7 was equally highly expressed in all cartilage and synovial biopsies. Increased levels of BMPR-1A, but not of BMPR-1B, and BMPR-2, were found in all synovial and 47% of all cartilage samples (P = 0.002). BMP-2 was positively scored in 47% of all cartilage and 40% of all synovial specimens. Defect size, KOSS, Henderson or Kellgren-Lawrence score did not statistically significant correlate with the expression of the analyzed proteins or Mankin and Pritzker scores. Duration of symptoms and localization of lesions were associated with KOSS (P < 0.02), but there was no influence of these parameters on protein expression. Conclusions. BMP-2, BMP-7, and BMPR-1A were expressed in cartilage and synovia of knees with focal cartilage lesions. Although defect localization and duration of symptoms decisively influence KOSS, there was no associated alteration of protein expression observed. PMID:22272175

Facilitated receptor-recognition and enhanced bioactivity of bone morphogenetic protein-2 on magnesium-substituted hydroxyapatite surface

PubMed Central

Huang, Baolin; Yuan, Yuan; Li, Tong; Ding, Sai; Zhang, Wenjing; Gu, Yuantong; Liu, Changsheng

2016-01-01

Biomaterial surface functionalized with bone morphogenetic protein-2 (BMP-2) is a promising approach to fabricating successful orthopedic implants/scaffolds. However, the bioactivity of BMP-2 on material surfaces is still far from satisfactory and the mechanism of related protein-surface interaction remains elusive. Based on the most widely used bone-implants/scaffolds material, hydroxyapatite (HAP), we developed a matrix of magnesium-substituted HAP (Mg-HAP, 2.2 at% substitution) to address these issues. Further, we investigated the adsorption dynamics, BMPRs-recruitment, and bioactivity of recombinant human BMP-2 (rhBMP-2) on the HAP and Mg-HAP surfaces. To elucidate the mechanism, molecular dynamic simulations were performed to calculate the preferred orientations, conformation changes, and cysteine-knot stabilities of adsorbed BMP-2 molecules. The results showed that rhBMP-2 on the Mg-HAP surface exhibited greater bioactivity, evidenced by more facilitated BMPRs-recognition and higher ALP activity than on the HAP surface. Moreover, molecular simulations indicated that BMP-2 favoured distinct side-on orientations on the HAP and Mg-HAP surfaces. Intriguingly, BMP-2 on the Mg-HAP surface largely preserved the active protein structure evidenced by more stable cysteine-knots than on the HAP surface. These findings explicitly clarify the mechanism of BMP-2-HAP/Mg-HAP interactions and highlight the promising application of Mg-HAP/BMP-2 matrixes in bone regeneration implants/scaffolds. PMID:27075233

Facilitated receptor-recognition and enhanced bioactivity of bone morphogenetic protein-2 on magnesium-substituted hydroxyapatite surface

NASA Astrophysics Data System (ADS)

Huang, Baolin; Yuan, Yuan; Li, Tong; Ding, Sai; Zhang, Wenjing; Gu, Yuantong; Liu, Changsheng

2016-04-01

Biomaterial surface functionalized with bone morphogenetic protein-2 (BMP-2) is a promising approach to fabricating successful orthopedic implants/scaffolds. However, the bioactivity of BMP-2 on material surfaces is still far from satisfactory and the mechanism of related protein-surface interaction remains elusive. Based on the most widely used bone-implants/scaffolds material, hydroxyapatite (HAP), we developed a matrix of magnesium-substituted HAP (Mg-HAP, 2.2 at% substitution) to address these issues. Further, we investigated the adsorption dynamics, BMPRs-recruitment, and bioactivity of recombinant human BMP-2 (rhBMP-2) on the HAP and Mg-HAP surfaces. To elucidate the mechanism, molecular dynamic simulations were performed to calculate the preferred orientations, conformation changes, and cysteine-knot stabilities of adsorbed BMP-2 molecules. The results showed that rhBMP-2 on the Mg-HAP surface exhibited greater bioactivity, evidenced by more facilitated BMPRs-recognition and higher ALP activity than on the HAP surface. Moreover, molecular simulations indicated that BMP-2 favoured distinct side-on orientations on the HAP and Mg-HAP surfaces. Intriguingly, BMP-2 on the Mg-HAP surface largely preserved the active protein structure evidenced by more stable cysteine-knots than on the HAP surface. These findings explicitly clarify the mechanism of BMP-2-HAP/Mg-HAP interactions and highlight the promising application of Mg-HAP/BMP-2 matrixes in bone regeneration implants/scaffolds.

β₂ adrenergic receptor activation suppresses bone morphogenetic protein (BMP)-induced alkaline phosphatase expression in osteoblast-like MC3T3E1 cells.

PubMed

Yamada, Takayuki; Ezura, Yoichi; Hayata, Tadayoshi; Moriya, Shuichi; Shirakawa, Jumpei; Notomi, Takuya; Arayal, Smriti; Kawasaki, Makiri; Izu, Yayoi; Harada, Kiyoshi; Noda, Masaki

2015-06-01

β adrenergic stimulation suppresses bone formation in vivo while its actions in osteoblastic differentiation are still incompletely understood. We therefore examined the effects of β2 adrenergic stimulation on osteoblast-like MC3T3-E1 cells focusing on BMP-induced alkaline phosphatase expression. Morphologically, isoproterenol treatment suppresses BMP-induced increase in the numbers of alkaline phosphatase-positive small foci in the cultures of MC3T3-E1 cells. Biochemically, isoproterenol treatment suppresses BMP-induced enzymatic activity of alkaline phosphatase in a dose-dependent manner. Isoproterenol suppression of alkaline phosphatase activity is observed even when the cells are treated with high concentrations of BMP. With respect to cell density, isoproterenol treatment tends to suppress BMP-induced increase in alkaline phosphatase expression more in osteoblasts cultured at higher cell density. In terms of treatment protocol, continuous isoproterenol treatment is compared to cyclic treatment. Continuous isoproterenol treatment is more suppressive against BMP-induced increase in alkaline phosphatase expression than cyclic regimen. At molecular level, isoproterenol treatment suppresses BMP-induced enhancement of alkaline phosphatase mRNA expression. Regarding the mode of isoproterenol action, isoproterenol suppresses BMP-induced BRE-luciferase activity. These data indicate that isoproterenol regulates BMP-induced alkaline phosphatase expression in osteoblast-like MC3T3E1 cells. © 2014 Wiley Periodicals, Inc.

A SHH-FOXF1-BMP4 signaling axis regulating growth and differentiation of epithelial and mesenchymal tissues in ureter development.

PubMed

Bohnenpoll, Tobias; Wittern, Anna B; Mamo, Tamrat M; Weiss, Anna-Carina; Rudat, Carsten; Kleppa, Marc-Jens; Schuster-Gossler, Karin; Wojahn, Irina; Lüdtke, Timo H-W; Trowe, Mark-Oliver; Kispert, Andreas

2017-08-01

The differentiated cell types of the epithelial and mesenchymal tissue compartments of the mature ureter of the mouse arise in a precise temporal and spatial sequence from uncommitted precursor cells of the distal ureteric bud epithelium and its surrounding mesenchyme. Previous genetic efforts identified a member of the Hedgehog (HH) family of secreted proteins, Sonic hedgehog (SHH) as a crucial epithelial signal for growth and differentiation of the ureteric mesenchyme. Here, we used conditional loss- and gain-of-function experiments of the unique HH signal transducer Smoothened (SMO) to further characterize the cellular functions and unravel the effector genes of HH signaling in ureter development. We showed that HH signaling is not only required for proliferation and SMC differentiation of cells of the inner mesenchymal region but also for survival of cells of the outer mesenchymal region, and for epithelial proliferation and differentiation. We identified the Forkhead transcription factor gene Foxf1 as a target of HH signaling in the ureteric mesenchyme. Expression of a repressor version of FOXF1 in this tissue completely recapitulated the mesenchymal and epithelial proliferation and differentiation defects associated with loss of HH signaling while re-expression of a wildtype version of FOXF1 in the inner mesenchymal layer restored these cellular programs when HH signaling was inhibited. We further showed that expression of Bmp4 in the ureteric mesenchyme depends on HH signaling and Foxf1, and that exogenous BMP4 rescued cell proliferation and epithelial differentiation in ureters with abrogated HH signaling or FOXF1 function. We conclude that SHH uses a FOXF1-BMP4 module to coordinate the cellular programs for ureter elongation and differentiation, and suggest that deregulation of this signaling axis occurs in human congenital anomalies of the kidney and urinary tract (CAKUT).

Repair of Cranial Bone Defects Using rhBMP2 and Submicron Particle of Biphasic Calcium Phosphate Ceramics with Through-Hole

PubMed Central

Jeong, Byung-Chul; Choi, Hyuck; Hur, Sung-Woong; Kim, Jung-Woo; Oh, Sin-Hye; Kim, Hyun-Seung; Song, Soo-Chang; Lee, Keun-Bae; Park, Kwang-Bum; Koh, Jeong-Tae

2015-01-01

Recently a submicron particle of biphasic calcium phosphate ceramic (BCP) with through-hole (donut-shaped BCP (d-BCP)) was developed for improving the osteoconductivity. This study was performed to examine the usefulness of d-BCP for the delivery of osteoinductive rhBMP2 and the effectiveness on cranial bone regeneration. The d-BCP was soaked in rhBMP2 solution and then freeze-dried. Scanning electron microscope (SEM), energy dispersive spectroscopy (EDS), and Raman spectroscopy analyses confirmed that rhBMP2 was well delivered onto the d-BCP surface and the through-hole. The bioactivity of the rhBMP2/d-BCP composite was validated in MC3T3-E1 cells as an in vitro model and in critical-sized cranial defects in C57BL/6 mice. When freeze-dried d-BCPs with rhBMP2 were placed in transwell inserts and suspended above MC3T3-E1, alkaline phosphatase activity and osteoblast-specific gene expression were increased compared to non-rhBMP2-containing d-BCPs. For evaluating in vivo effectiveness, freeze-dried d-BCPs with or without rhBMP2 were implanted into critical-sized cranial defects. Microcomputed tomography and histologic analysis showed that rhBMP2-containing d-BCPs significantly enhanced cranial bone regeneration compared to non-rhBMP2-containing control. These results suggest that a combination of d-BCP and rhBMP2 can accelerate bone regeneration, and this could be used to develop therapeutic strategies in hard tissue healing. PMID:26491693

Repair of Cranial Bone Defects Using rhBMP2 and Submicron Particle of Biphasic Calcium Phosphate Ceramics with Through-Hole.

PubMed

Jeong, Byung-Chul; Choi, Hyuck; Hur, Sung-Woong; Kim, Jung-Woo; Oh, Sin-Hye; Kim, Hyun-Seung; Song, Soo-Chang; Lee, Keun-Bae; Park, Kwang-Bum; Koh, Jeong-Tae

2015-01-01

Recently a submicron particle of biphasic calcium phosphate ceramic (BCP) with through-hole (donut-shaped BCP (d-BCP)) was developed for improving the osteoconductivity. This study was performed to examine the usefulness of d-BCP for the delivery of osteoinductive rhBMP2 and the effectiveness on cranial bone regeneration. The d-BCP was soaked in rhBMP2 solution and then freeze-dried. Scanning electron microscope (SEM), energy dispersive spectroscopy (EDS), and Raman spectroscopy analyses confirmed that rhBMP2 was well delivered onto the d-BCP surface and the through-hole. The bioactivity of the rhBMP2/d-BCP composite was validated in MC3T3-E1 cells as an in vitro model and in critical-sized cranial defects in C57BL/6 mice. When freeze-dried d-BCPs with rhBMP2 were placed in transwell inserts and suspended above MC3T3-E1, alkaline phosphatase activity and osteoblast-specific gene expression were increased compared to non-rhBMP2-containing d-BCPs. For evaluating in vivo effectiveness, freeze-dried d-BCPs with or without rhBMP2 were implanted into critical-sized cranial defects. Microcomputed tomography and histologic analysis showed that rhBMP2-containing d-BCPs significantly enhanced cranial bone regeneration compared to non-rhBMP2-containing control. These results suggest that a combination of d-BCP and rhBMP2 can accelerate bone regeneration, and this could be used to develop therapeutic strategies in hard tissue healing.

Influence of Mussel-Derived Bioactive BMP-2-Decorated PLA on MSC Behavior in Vitro and Verification with Osteogenicity at Ectopic Sites in Vivo.

PubMed

Chen, Zhuoyue; Zhang, Zhen; Feng, Juantao; Guo, Yayuan; Yu, Yuan; Cui, Jihong; Li, Hongmin; Shang, Lijun

2018-04-11

Osteoinductive activity of the implant in bone healing and regeneration is still a challenging research topic. Therapeutic application of recombinant human bone morphogenetic protein-2 (BMP-2) is a promising approach to enhance osteogenesis. However, high dose and uncontrolled burst release of BMP-2 may introduce edema, bone overgrowth, cystlike bone formation, and inflammation. In this study, low-dose BMP-2 of 1 μg was used to design PLA-PD-BMP for functionalization of polylactic acid (PLA) implants via mussel-inspired polydopamine (PD) assist. For the first time, the binding property and efficiency of the PD coating with BMP-2 were directly demonstrated and analyzed using an antigen-antibody reaction. The obtained PLA-PD-BMP surface immobilized with this low BMP-2 dose can endow the implants with abilities of introducing strong stem cell adhesion and enhanced osteogenicity. Furthermore, in vivo osteoinduction of the PLA-PD-BMP-2 scaffolds was confirmed by a rat ectopic bone model, which is marked as the "gold standard" for the evidence of osteoinductive activity. The microcomputed tomography, Young's modulus, and histology analyses were also employed to demonstrate that PLA-PD-BMP grafted with 1 μg of BMP-2 can induce bone formation. Therefore, the method in this study can be used as a model system to immobilize other growth factors onto various different types of polymer substrates. The highly biomimetic mussel-derived strategy can therefore improve the clinical outcome of polymer-based medical implants in a facile, safe, and effective way.

Mapping the Interaction Anatomy of BmP02 on Kv1.3 Channel

NASA Astrophysics Data System (ADS)

Wu, B.; Wu, B. F.; Feng, Y. J.; Tao, J.; Ji, Y. H.

2016-07-01

The potassium channel Kv 1.3 plays a vital part in the activation of T lymphocytes and is an attractive pharmacological target for autoimmune diseases. BmP02, a 28-residue peptide isolated from Chinese scorpion (Buthus martensi Karsch) venom, is a potent and selective Kv1.3 channel blocker. However, the mechanism through which BmP02 recognizes and inhibits the Kv1.3 channel is still unclear. In the present study, a complex molecular model of Kv1.3-BmP02 was developed by docking analysis and molecular dynamics simulations. From these simulations, it appears the large β-turn (residues 10-16) of BmP02 might be the binding interface with Kv 1.3. These results were confirmed by scanning alanine mutagenesis of BmP02, which identified His9, Lys11 and Lys13, which lie within BmP02’s β-turn, as key residues for interacting with Kv1.3. Based on these results and molecular modeling, two negatively charged residues of Kv1.3, D421 and D422, located in turret region, were predicted to act as the binding site for BmP02. Mutation of these residues reduced sensitivity of Kv 1.3 to BmP02 inhibition, suggesting that electrostatic interactions play a crucial role in Kv1.3-BmP02 interaction. This study revealed the molecular basis of Kv 1.3 recognition by BmP02 venom, and provides a novel interaction model for Kv channel-specific blocker complex, which may help guide future drug-design for Kv1.3-related channelopathies.

Bone morphogenetic protein signaling is impaired in an Hfe knockout mouse model of hemochromatosis

PubMed Central

Corradini, Elena; Garuti, Cinzia; Montosi, Giuliana; Ventura, Paolo; Andriopoulos, Billy; Lin, Herbert Y.; Pietrangelo, Antonello; Babitt, Jodie L.

2009-01-01

Background and Aims Mutations in HFE are the most common cause of the iron-overload disorder hereditary hemochromatosis (HH). Levels of the main iron regulatory hormone, hepcidin, are inappropriately low in HH mouse models and patients with HFE mutations, indicating that HFE regulates hepcidin. The bone morphogenetic protein 6 (BMP6)-SMAD signaling pathway is an important endogenous regulator of hepcidin expression. We investigated whether HFE is involved in BMP6-SMAD regulation of hepcidin expression. Methods The BMP6-SMAD pathway was examined in Hfe knockout (KO) mice and in wild-type (WT) mice as controls. Mice were placed on diets of varying iron content. Hepcidin induction by BMP6 was examined in primary hepatocytes from Hfe KO mice; data were compared with those of WT mice. Results Liver levels of Bmp6 mRNA were higher in Hfe KO mice; these were appropriate for the increased hepatic levels of iron in these mice, compared with WT mice. However, levels of hepatic phosphorylated Smad 1/5/8 protein (an intracellular mediator of Bmp6 signaling) and Id1 mRNA (a target gene of Bmp6) were inappropriately low for the body iron burden and Bmp6 mRNA levels in Hfe KO, compared with WT mice. BMP6 induction of hepcidin expression was reduced in Hfe KO hepatocytes compared with WT hepatocytes. Conclusions HFE is not involved in regulation of BMP6 by iron, but does regulate the downstream signals of BMP6 that are triggered by iron. PMID:19591830

Prediction of Biological Motion Perception Performance from Intrinsic Brain Network Regional Efficiency

PubMed Central

Wang, Zengjian; Zhang, Delong; Liang, Bishan; Chang, Song; Pan, Jinghua; Huang, Ruiwang; Liu, Ming

2016-01-01

Biological motion perception (BMP) refers to the ability to perceive the moving form of a human figure from a limited amount of stimuli, such as from a few point lights located on the joints of a moving body. BMP is commonplace and important, but there is great inter-individual variability in this ability. This study used multiple regression model analysis to explore the association between BMP performance and intrinsic brain activity, in order to investigate the neural substrates underlying inter-individual variability of BMP performance. The resting-state functional magnetic resonance imaging (rs-fMRI) and BMP performance data were collected from 24 healthy participants, for whom intrinsic brain networks were constructed, and a graph-based network efficiency metric was measured. Then, a multiple linear regression model was used to explore the association between network regional efficiency and BMP performance. We found that the local and global network efficiency of many regions was significantly correlated with BMP performance. Further analysis showed that the local efficiency rather than global efficiency could be used to explain most of the BMP inter-individual variability, and the regions involved were predominately located in the Default Mode Network (DMN). Additionally, discrimination analysis showed that the local efficiency of certain regions such as the thalamus could be used to classify BMP performance across participants. Notably, the association pattern between network nodal efficiency and BMP was different from the association pattern of static directional/gender information perception. Overall, these findings show that intrinsic brain network efficiency may be considered a neural factor that explains BMP inter-individual variability. PMID:27853427

Pasture BMP effectiveness using an HRU-based subarea approach in SWAT.

PubMed

Sheshukov, Aleksey Y; Douglas-Mankin, Kyle R; Sinnathamby, Sumathy; Daggupati, Prasad

2016-01-15

Many conservation programs have been established to motivate producers to adopt best management practices (BMP) to minimize pasture runoff and nutrient loads, but a process is needed to assess BMP effectiveness to help target implementation efforts. A study was conducted to develop and demonstrate a method to evaluate water-quality impacts and the effectiveness of two widely used BMPs on a livestock pasture: off-stream watering site and stream fencing. The Soil and Water Assessment Tool (SWAT) model was built for the Pottawatomie Creek Watershed in eastern Kansas, independently calibrated at the watershed outlet for streamflow and at a pasture site for nutrients and sediment runoff, and also employed to simulate pollutant loads in a synthetic pasture. The pasture was divided into several subareas including stream, riparian zone, and two grazing zones. Five scenarios applied to both a synthetic pasture and a whole watershed were simulated to assess various combinations of widely used pasture BMPs: (1) baseline conditions with an open stream access, (2) an off-stream watering site installed in individual subareas in the pasture, and (3) stream or riparian zone fencing with an off-stream watering site. Results indicated that pollutant loads increase with increasing stocking rates whereas off-stream watering site and/or stream fencing reduce time cattle spend in the stream and nutrient loads. These two BMPs lowered organic P and N loads by more than 59% and nitrate loads by 19%, but TSS and sediment-attached P loads remained practically unchanged. An effectiveness index (EI) quantified impacts from the various combinations of off-stream watering sites and fencing in all scenarios. Stream bank contribution to pollutant loads was not accounted in the methodology due to limitations of the SWAT model, but can be incorporated in the approach if an amount of bank soil loss is known for various stocking rates. The proposed methodology provides an adaptable framework for pasture BMP assessment and was utilized to represent a consistent, defensible process to quantify the effectiveness of BMP proposals in a BMP auction in eastern Kansas. Copyright © 2015 Elsevier Ltd. All rights reserved.

Sox5 is a DNA binding co-factor for BMP R-Smads that directs target specificity during patterning of the early ectoderm

PubMed Central

Nordin, Kara; LaBonne, Carole

2014-01-01

SUMMARY The SoxD factor, Sox5, is expressed in ectodermal cells at times and places where BMP signaling is active, including the cells of the animal hemisphere at blastula stages, and the neural plate border (NPB) and neural crest (NC) at neurula stages. Sox5 is required for proper ectoderm development, and deficient embryos display patterning defects characteristic of perturbations of BMP signaling, including loss of neural crest and epidermis and expansion of the neural plate. We show that Sox5 is essential for activation of BMP target genes in embryos and explants, that it physically interacts with BMP R-Smads, and that it is essential for recruitment of Smad1/4 to BMP regulatory elements. Our findings identify Sox5 as the long sought DNA binding partner for BMP R-Smads essential to plasticity and pattern in the early ectoderm. PMID:25453832

Dynamics of BMP signaling in limb bud mesenchyme and polydactyly.

PubMed

Norrie, Jacqueline L; Lewandowski, Jordan P; Bouldin, Cortney M; Amarnath, Smita; Li, Qiang; Vokes, Martha S; Ehrlich, Lauren I R; Harfe, Brian D; Vokes, Steven A

2014-09-15

Mutations in the Bone Morphogenetic Protein (BMP) pathway are associated with a range of defects in skeletal formation. Genetic analysis of BMP signaling requirements is complicated by the presence of three partially redundant BMPs that are required for multiple stages of limb development. We generated an inducible allele of a BMP inhibitor, Gremlin, which reduces BMP signaling. We show that BMPs act in a dose and time dependent manner in which early reduction of BMPs result in digit loss, while inhibiting overall BMP signaling between E10.5 and E11.5 allows polydactylous digit formation. During this period, inhibiting BMPs extends the duration of FGF signaling. Sox9 is initially expressed in normal digit ray domains but at reduced levels that correlate with the reduction in BMP signaling. The persistence of elevated FGF signaling likely promotes cell proliferation and survival, inhibiting the activation of Sox9 and secondarily, inhibiting the differentiation of Sox9-expressing chondrocytes. Our results provide new insights into the timing and clarify the mechanisms underlying BMP signaling during digit morphogenesis. Copyright © 2014 Elsevier Inc. All rights reserved.

cAMP enhances BMP2-signaling through PKA and MKP1-dependent mechanisms

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ghayor, Chafik; Ehrbar, Martin; Miguel, Blanca San

2009-04-03

Recent studies suggest that the elevation of intracellular cyclic adenosine monophosphate (cAMP) and the activation of the protein kinase A regulate BMP-induced osteogenesis. However, the precise mechanisms underlying the enhancing effect of cAMP on BMP2 signaling were not completely revealed. In this study we investigated the effect of elevated cAMP level and PKA activation on the BMP2-induced osteoblastic differentiation in pluripotent C2C12 cells. Alkaline phosphatase activity and its mRNA were consistently induced by BMP2 treatment. The pretreatment of C2C12 cells with Forskolin, a cAMP generating agent, dbcAMP, an analogue of cAMP, or IBMX (3-isobutyl 1-methyl xanthine), and a nonspecific inhibitormore » of phosphodiesterases elicited further activation of alkaline phosphatase. Furthermore, elevated intracellular cAMP level increased BMP2-induced MKP1. On the other hand, BMP2-induced Erk phosphorylation (p44/p42) and cell proliferation were suppressed in the presence of cAMP. Thus, cAMP might enhance BMP2-induced osteoblastic differentiation by a MKP1-Erk-dependent mechanism.« less

Ski represses bone morphogenic protein signaling in Xenopus and mammalian cells

PubMed Central

Wang, Wei; Mariani, Francesca V.; Harland, Richard M.; Luo, Kunxin

2000-01-01

The bone morphogenic proteins (BMPs) play important roles in vertebrate development. In Xenopus, BMPs act as epidermal inducers and also as negative regulators of neurogenesis. Antagonism of BMP signaling results in neuralization. BMPs signal through the cell-surface receptors and downstream Smad molecules. Upon stimulation with BMP, Smad1, Smad5, and Smad8 are phosphorylated by the activated BMP receptors, form a complex with Smad4, and translocate into the nucleus, where they regulate the expression of BMP target genes. Here, we show that the Ski oncoprotein can block BMP signaling and the expression of BMP-responsive genes in both Xenopus and mammalian cells by directly interacting with and repressing the activity of BMP-specific Smad complexes. This ability to antagonize BMP signaling results in neuralization by Ski in the Xenopus embryo and blocking of osteoblast differentiation of murine W-20-17 cells. Thus, Ski is able to repress the activity of all receptor-associated Smads and may regulate vertebrate development by modulating the signaling activity of transforming growth factor-β family members. PMID:11121043

Ski represses bone morphogenic protein signaling in Xenopus and mammalian cells.

PubMed

Wang, W; Mariani, F V; Harland, R M; Luo, K

2000-12-19

The bone morphogenic proteins (BMPs) play important roles in vertebrate development. In Xenopus, BMPs act as epidermal inducers and also as negative regulators of neurogenesis. Antagonism of BMP signaling results in neuralization. BMPs signal through the cell-surface receptors and downstream Smad molecules. Upon stimulation with BMP, Smad1, Smad5, and Smad8 are phosphorylated by the activated BMP receptors, form a complex with Smad4, and translocate into the nucleus, where they regulate the expression of BMP target genes. Here, we show that the Ski oncoprotein can block BMP signaling and the expression of BMP-responsive genes in both Xenopus and mammalian cells by directly interacting with and repressing the activity of BMP-specific Smad complexes. This ability to antagonize BMP signaling results in neuralization by Ski in the Xenopus embryo and blocking of osteoblast differentiation of murine W-20-17 cells. Thus, Ski is able to repress the activity of all receptor-associated Smads and may regulate vertebrate development by modulating the signaling activity of transforming growth factor-beta family members.

Attenuated BMP1 Function Compromises Osteogenesis, Leading to Bone Fragility in Humans and Zebrafish

PubMed Central

Asharani, P.V.; Keupp, Katharina; Semler, Oliver; Wang, Wenshen; Li, Yun; Thiele, Holger; Yigit, Gökhan; Pohl, Esther; Becker, Jutta; Frommolt, Peter; Sonntag, Carmen; Altmüller, Janine; Zimmermann, Katharina; Greenspan, Daniel S.; Akarsu, Nurten A.; Netzer, Christian; Schönau, Eckhard; Wirth, Radu; Hammerschmidt, Matthias; Nürnberg, Peter; Wollnik, Bernd; Carney, Thomas J.

2012-01-01

Bone morphogenetic protein 1 (BMP1) is an astacin metalloprotease with important cellular functions and diverse substrates, including extracellular-matrix proteins and antagonists of some TGFβ superfamily members. Combining whole-exome sequencing and filtering for homozygous stretches of identified variants, we found a homozygous causative BMP1 mutation, c.34G>C, in a consanguineous family affected by increased bone mineral density and multiple recurrent fractures. The mutation is located within the BMP1 signal peptide and leads to impaired secretion and an alteration in posttranslational modification. We also characterize a zebrafish bone mutant harboring lesions in bmp1a, demonstrating conservation of BMP1 function in osteogenesis across species. Genetic, biochemical, and histological analyses of this mutant and a comparison to a second, similar locus reveal that Bmp1a is critically required for mature-collagen generation, downstream of osteoblast maturation, in bone. We thus define the molecular and cellular bases of BMP1-dependent osteogenesis and show the importance of this protein for bone formation and stability. PMID:22482805

Combinatorial Fgf and Bmp signalling patterns the gastrula ectoderm into prospective neural and epidermal domains

PubMed Central

Kudoh, Tetsuhiro; Concha, Miguel L.; Houart, Corinne; Dawid, Igor B.; Wilson, Stephen W.

2009-01-01

Summary Studies in fish and amphibia have shown that graded Bmp signalling activity regulates dorsal-to-ventral (DV) patterning of the gastrula embryo. In the ectoderm, it is thought that high levels of Bmp activity promote epidermal development ventrally, whereas secreted Bmp antagonists emanating from the organiser induce neural tissue dorsally. However, in zebrafish embryos, the domain of cells destined to contribute to the spinal cord extends all the way to the ventral side of the gastrula, a long way from the organiser. We show that in vegetal (trunk and tail) regions of the zebrafish gastrula, neural specification is initiated at all DV positions of the ectoderm in a manner that is unaffected by levels of Bmp activity and independent of organiser-derived signals. Instead, we find that Fgf activity is required to induce vegetal prospective neural markers and can do so without suppressing Bmp activity. We further show that Bmp signalling does occur within the vegetal prospective neural domain and that Bmp activity promotes the adoption of caudal fate by this tissue. PMID:15262889

Effect of rhBMP-2 on tibial plateau fractures in a canine model.

PubMed

Schaefer, Susan L; Lu, Yan; Seeherman, Howard; Li, X Jian; Lopez, Mandi J; Markel, Mark D

2009-04-01

This study was to determine the efficacy of recombinant human bone morphogenetic protien-2 (rhBMP-2)/calcium phosphate matrix (CPX) paste to accelerate healing in a canine articular fracture model with associated subchondral defect. rhBMP-2/CPX (BMP), CPX alone (CPX) or autogenous bone graft (ABG) was administered to a canine articular tibial plateau osteotomy with a subchondral defect in each of 21 female dogs. The unoperated contralateral limbs served as controls. Ground reaction forces, synovial fluid, radiographic changes, mechanical testing, bone density, and histology of bone and synovium were analyzed at 6 weeks after surgery. Radiographic analysis demonstrated that the BMP and CPX groups showed improved bony healing compared to the ABG group at week 6. Histomorphometric analysis demonstrated that the BMP group had significantly increased trabecular bone volume compared to the CPX and ABG groups. Mechanical testing revealed that the BMP group had significantly greater maximum failure loads than the ABG group. Histological analysis demonstrated that the BMP group had significantly less sub-synovial inflammation than CPX group. This study demonstrated that rhBMP-2/CPX accelerated healing of articular fractures with subchondral defect compared to ABG in most of the parameters evaluated, and had less subsynovial inflammation than the CPX alone in a canine model.

Role of RGM coreceptors in bone morphogenetic protein signaling

PubMed Central

Halbrooks, Peter J; Ding, Ru; Wozney, John M; Bain, Gerard

2007-01-01

Background The repulsive guidance molecule (RGM) proteins, originally discovered for their roles in neuronal development, have been recently identified as co-receptors in the bone morphogenetic protein (BMP) signaling pathway. BMPs are members of the TGFβ superfamily of signaling cytokines, and serve to regulate many aspects of cellular growth and differentiation. Results Here, we investigate whether RGMa, RGMb, and RGMc play required roles in BMP and TGFβ signaling in the mouse myoblast C2C12 cell line. These cells are responsive to BMPs and are frequently used to study BMP/TGFβ signaling pathways. Using siRNA reagents to specifically knock down each RGM protein, we show that the RGM co-receptors are required for significant BMP signaling as reported by two cell-based BMP activity assays: endogenous alkaline phosphatase activity and a luciferase-based BMP reporter assay. Similar cell-based assays using a TGFβ-induced luciferase reporter show that the RGM co-receptors are not required for TGFβ signaling. The binding interaction of each RGM co-receptor to each of BMP2 and BMP12 is observed and quantified, and equilibrium dissociation constants in the low nanomolar range are reported. Conclusion Our results demonstrate that the RGMs play a significant role in BMP signaling and reveal that these molecules cannot functionally compensate for one another. PMID:17615080

Characterization of the post-translational modification of recombinant human BMP-15 mature protein

PubMed Central

Saito, Seiji; Yano, Keiichi; Sharma, Shweta; McMahon, Heather E.; Shimasaki, Shunichi

2008-01-01

Bone morphogenetic protein-15 (BMP-15) is an oocyte-secreted factor critical for the regulation of ovarian physiology. When recombinant human BMP-15 (rhBMP-15) produced in human embryonic kidney 293 cells was subjected to SDS-PAGE analysis, two mature protein forms corresponding to 16 kDa (P16) and 17 kDa (P17) were observed. Despite the physiological relevance and critical function of BMP-15 in female reproduction, little is known about the structure of rhBMP-15. Here, we have analyzed the structure of the rhBMP-15 mature proteins (P16 and P17) using state-of-the-art proteomics technology. Our findings are as follows: (1) the N-terminal amino acid of P16 and P17 is pyroglutamic acid; (2) the Ser residue at the sixth position of P16 is phosphorylated; (3) P17 is O-glycosylated at Thr10; and (4) the C-terminal amino acid of P16 and P17 is truncated. These findings are the first knowledge of the structure of rhBMP-15 mature protein toward understanding the molecular basis of BMP-15 function and could provide an important contribution to the rapidly progressing research area involving oocyte-specific growth factors in modulation of female fertility. PMID:18227435

Expression of BMP-4 in papillary thyroid carcinoma and its correlation with tumor invasion and progression.

PubMed

Meng, Xiaomei; Zhu, Peng; Li, Ning; Hu, Jinchen; Wang, Shaoguang; Pang, Shuguang; Wang, Jiahui

2017-04-01

Bone morphogenetic protein 4 (BMP-4) is a member of the BMP protein family. BMP-4 was reported to induce epithelial-mesenchymal transition (EMT) and promote tumor cell immigration and invasion. This study aimed to investigate the expression of BMP-4 in papillary thyroid carcinoma (PTC) and its correlation with the patients' clinicophathological features and with tumor invasion and metastasis. Surgically resected PTC specimens from 82 patients admitted to the Department of Thyroid Surgery of Yantai Yuhuangding Hospital between Feb 1st and May 31st, 2016 were collected. The expression level of BMP-4 in PTC tissues was examined by immunohistochemical staining. The full clinical records of all patients were collected to analyze the relevance between BMP-4 expression and the clinical pathological features of PTC. Our result showed that BMP-4-positive cell rate and staining intensity were positively correlated with the patient's age (P=0.031, 0.037), tumor size (P=0.033, 0.019), capsular invasion (P=0.001, 0.002) and TNM stage (P=0.001, 0.004), while not correlated with gender, multicentricity of tumor or lymphatic metastasis. In conclusion, this study identified BMP-4 as a potential molecular marker for predicting the invasion and progression of PTC. Copyright © 2017 Elsevier GmbH. All rights reserved.

BMP suppresses PTEN expression via RAS/ERK signaling.

PubMed

Beck, Stayce E; Carethers, John M

2007-08-01

Bone morphogenetic protein (BMP), a member of the transforming growth factor beta family, classically utilizes the SMAD signaling pathway for its growth suppressive effects,and loss of this signaling cascade may accelerate cell growth. In the colon cancer predisposition syndrome Juvenile Polyposis, as well as in the late progression stages of nonsyndromic colorectal cancers, SMAD4 function is typically abrogated. Here, we utilized the SMAD4-null SW480 colon cancer cell line to examine BMPs effect on a potential target gene, PTEN, and how its expression might be regulated. Initial treatment of the SMAD4-null cells with BMP resulted in mild growth suppression, but with prolonged exposure to BMP, the cells become growth stimulatory, which coincided with observed decreases in transcription and translation of PTEN, and with corresponding increases in phospho-AKT protein levels. BMP-induced PTEN suppression was mediated via the RAS/ERK pathway, as pharmacologic inhibition of RAS/ERK, or interference with protein function in the cytosol by DN-RAS prevented BMP-induced growth promotion and changes in PTEN levels, as did treatment with noggin, a BMP ligand inhibitor. Thus, BMP downregulates PTEN via RAS/ERK in a SMAD4-null environment that contributes to cell growth, and constitutes a SMAD4-independent but BMP-responsive signaling pathway.

Dkk1 haploinsufficiency requires expression of Bmp2 for bone anabolic activity.

PubMed

Intini, Giuseppe; Nyman, Jeffry S

2015-06-01

Bone fractures remain a serious health burden and prevention and enhanced healing of fractures have been obtained by augmenting either BMP or Wnt signaling. However, whether BMP and Wnt signaling are both required or are self-sufficient for anabolic and fracture healing activities has never been fully elucidated. Mice haploinsufficient for Dkk1 (Dkk1(+/-)) exhibit a high bone mass phenotype due to an up-regulation of canonical Wnt signaling while mice lacking Bmp2 expression in the limbs (Bmp2(c/c);Prx1::cre) succumb to spontaneous fracture and are unable to initiate fracture healing; combined, these mice offer an opportunity to examine the requirement for activated BMP signaling on the anabolic and fracture healing activity of Wnts. When Dkk1(+/-) mice were crossed with Bmp2(c/c);Prx1::cre mice, the offspring bearing both genetic alterations were unable to increase bone mass and heal fractures, indicating that increased canonical Wnt signaling is unable to exploit its activity in absence of Bmp2. Thus, our data suggest that BMP signaling is required for Wnt-mediated anabolic activity and that therapies aimed at preventing fractures and fostering fracture repair may need to target both pathways for maximal efficacy. Copyright © 2015 Elsevier Inc. All rights reserved.

Evaluation of nonpoint-source contamination, Wisconsin: water year 1999

USGS Publications Warehouse

Walker, John F.; Graczyk, D.J.; Corsi, Steven R.; Wierl, J.A.; Owens, D.W.

2001-01-01

For two of the eight rural streams (Rattlesnake and Kuenster Creeks) minimal BMP implementation has occurred, hence a comparison of pre- BMP and data collected after BMP implementation began is not warranted. For two other rural streams (Brewery and Garfoot Creeks), BMP implementation is complete. For the four remaining rural streams (Bower, Otter, Eagle, and Joos Valley Creeks), the pre-BMP load data were compared to the transitional data to determine if significant reductions in the loads have occurred as a result of the BMP implementation to date. For all sites, the actual constituent loads for suspended solids and total phosphorus exhibit no statistically significant reductions after BMP installation. Multiple regressions were used to remove some of the natural variability in the data. Based on the residual analysis, for Otter Creek, there is a significant difference in the suspended-solids regression residuals between the pre-BMP and transitional periods, indicating a potential reduction as a result of the BMP implementation after accounting for natural variability. For Joos Valley Creek, the residuals for suspended solids and total phosphorus both show a significant reduction after accounting for natural variability. It is possible that the other sites will also show statistically significant reductions in suspended solids and total phosphorus if additional BMPs are implemented.

Dkk1 haploinsufficiency requires expression of Bmp2 for bone anabolic activity

PubMed Central

Intini, Giuseppe; Nyman, Jeffry S.

2015-01-01

Bone fractures remain a serious health burden and prevention and enhanced healing of fractures has been obtained by augmenting either BMP or Wnt signaling. However, whether BMP and Wnt signaling are both required or are self-sufficient for anabolic and fracture healing activities has never been fully elucidated. Mice haploinsufficient for Dkk1 (Dkk1+/−) exhibit a high bone mass phenotype due to an up-regulation of canonical Wnt signaling while mice lacking Bmp2 expression in the limbs (Bmp2c/c;Prx1::cre) succumb to spontaneous fracture and are unable to initiate fracture healing; combined, these mice offer an opportunity to examine the requirement for activated BMP signaling on the anabolic and fracture healing activity of Wnts. When Dkk1+/− mice were crossed with Bmp2c/c;Prx1::cre mice, the offspring bearing both genetic alterations were unable to increase bone mass and heal fractures, indicating that increased canonical Wnt signaling is unable to exploit its activity in absence of Bmp2. Thus, our data suggest that BMP signaling is required for Wnt-mediated anabolic activity and that therapies aimed at preventing fractures and fostering fracture repair may need to target both pathways for maximal efficacy. PMID:25603465

A regional protocol for evaluating the effectiveness of forestry best management practices at controlling erosion and sedimentation

Treesearch

Roger Ryder; Pamela Edwards; Pamela Edwards

2006-01-01

Forestry operations do not have permitting requirements under the Clean Water Act because there is a ccsilvicultural exemption" given in that law, as long as best management practices (BMPs) are used to help control non-point source pollution. However, states' monitoring of BMP effectiveness often has been sporadic and anecdotal, and the procedures used have...

Ex vivo bone morphogenetic protein 2 gene delivery using periodontal ligament stem cells for enhanced re-osseointegration in the regenerative treatment of peri-implantitis.

PubMed

Park, Shin-Young; Kim, Kyoung-Hwa; Gwak, Eun-Hye; Rhee, Sang-Hoon; Lee, Jeong-Cheol; Shin, Seung-Yun; Koo, Ki-Tae; Lee, Yong-Moo; Seol, Yang-Jo

2015-01-01

Peri-implantitis is a chronic inflammatory process with advanced bone loss and impaired healing potential. For peri-implantitis treatment, tissue engineering can be applied to enhance bone regeneration of peri-implant defects. This study aimed to evaluate ex vivo bone morphogenetic protein 2 (BMP2) gene delivery using canine periodontal ligament stem cells (PDLSCs) for regeneration of peri-implantitis defects. Canine PDLSCs were transduced with adenoviral vectors containing BMP2 (BMP2/PDLSCs). After peri-implantitis was induced by ligature placement in six beagle dogs, regenerative procedures were performed; hydroxyapatite (HA) particles and collagen gel with autologous canine PDLSCs (PDLSC group) or BMP2/PDLSCs (BMP/PDLSC group) or without cells (control group) were grafted into the defects and covered by an absorbable membrane. Three months later, the animals were sacrificed. In vitro, BMP2/PDLSCs showed similar levels of stem cell properties to PDLSCs, such as colony-forming efficiency and expression of MSC markers STRO-1 and CD 146. BMP2/PDLSCs produced BMP-2 until day 21 at a concentration of 4-8 ng/mL. In vivo, the BMP2/PDLSC group showed significantly more new bone formation and re-osseointegration in peri-implantitis defects compared to the other groups. In conclusion, ex vivo BMP2 gene delivery using PDLSCs enhanced new bone formation and re-osseointegration in peri-implantitis defects. © 2014 Wiley Periodicals, Inc.

Endocytosis contributes to BMP2-induced Smad signalling and neuronal growth.

PubMed

Hegarty, Shane V; Sullivan, Aideen M; O'Keeffe, Gerard W

2017-03-16

Bone morphogenetic protein 2 (BMP2) is a neurotrophic factor which induces the growth of midbrain dopaminergic (DA) neurons in vitro and in vivo, and its neurotrophic effects have been shown to be dependent on activation of BMP receptors (BMPRs) and Smad 1/5/8 signalling. However, the precise intracellular cascades that regulate BMP2-BMPR-Smad-signalling-induced neurite growth remain unknown. Endocytosis has been shown to regulate Smad 1/5/8 signalling and differentiation induced by BMPs. However, these studies were carried out in non-neural cells. Indeed, there are scant reports regarding the role of endocytosis in BMP-Smad signalling in neurons. To address this, and to further characterise the mechanisms regulating the neurotrophic effects of BMP2, the present study examined the role of dynamin-dependent endocytosis in BMP2-induced Smad signalling and neurite growth in the SH-SY5Y neuronal cell line. The activation, temporal kinetics and magnitude of Smad 1/5/8 signalling induced by BMP2 were significantly attenuated by dynasore-mediated inhibition of endocytosis in SH-SY5Y cells. Furthermore, BMP2-induced increases in neurite length and neurite branching in SH-SY5Y cells were significantly reduced following inhibition of dynamin-dependent endocytosis using dynasore. This study demonstrates that BMP2-induced Smad signalling and neurite growth is regulated by dynamin-dependent endocytosis in a model of human midbrain dopaminergic neurons. Copyright © 2017 Elsevier B.V. All rights reserved.

Injectable hydrogels embedded with alginate microspheres for controlled delivery of bone morphogenetic protein-2.

PubMed

Zhu, Youjia; Wang, Jiulong; Wu, Jingjing; Zhang, Jun; Wan, Ying; Wu, Hua

2016-03-23

Some delivery carriers with injectable characteristics were built using the thermosensitive chitosan/dextran-polylactide/glycerophosphate hydrogel and selected alginate microspheres for the controllable release of bone morphogenetic protein-2 (BMP-2). BMP-2 was first loaded into the microspheres with an average size of around 20 μm and the resulting microspheres were then embedded into the gel in order to achieve well-controlled BMP-2 release. The microsphere-embedded gels show their incipient gelation temperature at around 32 °C and pH at about 7.1. Some gels had their elastic modulus close to 1400 Pa and the ratio of elastic modulus to viscous modulus at around 34, revealing that they behaved like mechanically strong gels. Optimized microsphere-embedded gels were found to be able to administer the BMP-2 release without significant initial burst release in an approximately linear manner over a period of time longer than four weeks. The release rate and the released amount of BMP-2 from these gels could be regulated individually or cooperatively by the initial BMP-2 load and the dextran-polylactide content in the gels. Measurements of the BMP-2 induced alkaline phosphatase activity in C2C12 cells confirmed that C2C12 cells responded to BMP-2 in a dose-dependent way and the released BMP-2 from the microsphere-embedded gels well retained their bioactivity. In vivo assessment of some gels revealed that the released BMP-2 maintained its osteogenesis functions.

BMP2 and BMP4 variations and risk of non-syndromic cleft lip and palate.

PubMed

Saket, Mitra; Saliminejad, Kioomars; Kamali, Koorosh; Moghadam, Fatemeh Aghakhani; Anvar, Nazanin Esmaeili; Khorram Khorshid, Hamid Reza

2016-12-01

Non-syndromic cleft lip with or without cleft palate (CL/P) is one of the most common congenital anomalies and arises from the interaction of environmental and genetic factors. The objective of this study was to investigate the association between the BMP2 (bone morphogenetic protein 2) and BMP4 (bone morphogenetic protein 4) polymorphisms with non-syndromic CL/P to clarify the potential role of these genes in the etiology of CL/P in Iranian population. The allelic and genotypic frequencies of BMP2 rs235768 A>T and BMP4 rs17563 T>C polymorphisms were determined in 107 unrelated Iranian subjects with non-syndromic CL/P and 186 control subjects using PCR and RFLP methods, and the results were compared with healthy controls. A p-value of <0.05 was considered statistically significant. The BMP2 rs235768 AT genotype was significantly higher (P=0.009, OR=3, 95% CI=1.3-7.0) in the CL/P (59.8%) than the control group (33.3%). Similarly, the BMP4 rs17563 TC genotype were significantly higher (P=0.008, OR=3.7, 95% CI=1.4-9.9) in the CL/P (70.0%) than the control group (44.6%). The BMP2 rs235768 A>T and BMP4 rs17563 T>C polymorphisms could be considered as the risk factor for non-syndromic CL/P in Iranian population. Copyright © 2016 Elsevier Ltd. All rights reserved.

Bone morphogenetic protein 9 (BMP9) controls lymphatic vessel maturation and valve formation

PubMed Central

Levet, Sandrine; Ciais, Delphine; Merdzhanova, Galina; Mallet, Christine; Zimmers, Teresa A.; Lee, Se-Jin; Navarro, Fabrice P.; Texier, Isabelle; Feige, Jean-Jacques; Bailly, Sabine

2013-01-01

Lymphatic vessels are critical for the maintenance of tissue fluid homeostasis and their dysfunction contributes to several human diseases. The activin receptor-like kinase 1 (ALK1) is a transforming growth factor-β family type 1 receptor that is expressed on both blood and lymphatic endothelial cells (LECs). Its high-affinity ligand, bone morphogenetic protein 9 (BMP9), has been shown to be critical for retinal angiogenesis. The aim of this work was to investigate whether BMP9 could play a role in lymphatic development. We found that Bmp9 deficiency in mice causes abnormal lymphatic development. Bmp9-knockout (KO) pups presented hyperplastic mesenteric collecting vessels that maintained LYVE-1 expression. In accordance with this result, we found that BMP9 inhibited LYVE-1 expression in LECs in an ALK1-dependent manner. Bmp9-KO pups also presented a significant reduction in the number and in the maturation of mesenteric lymphatic valves at embryonic day 18.5 and at postnatal days 0 and 4. Interestingly, the expression of several genes known to be involved in valve formation (Foxc2, Connexin37, EphrinB2, and Neuropilin1) was upregulated by BMP9 in LECS. Finally, we demonstrated that Bmp9-KO neonates and adult mice had decreased lymphatic draining efficiency. These data identify BMP9 as an important extracellular regulator in the maturation of the lymphatic vascular network affecting valve development and lymphatic vessel function. PMID:23741013

Cross-talk between EGF and BMP9 signalling pathways regulates the osteogenic differentiation of mesenchymal stem cells.

PubMed

Liu, Xing; Qin, Jiaqiang; Luo, Qing; Bi, Yang; Zhu, Gaohui; Jiang, Wei; Kim, Stephanie H; Li, Mi; Su, Yuxi; Nan, Guoxin; Cui, Jing; Zhang, Wenwen; Li, Ruidong; Chen, Xiang; Kong, Yuhan; Zhang, Jiye; Wang, Jinhua; Rogers, Mary Rose; Zhang, Hongyu; Shui, Wei; Zhao, Chen; Wang, Ning; Liang, Xi; Wu, Ningning; He, Yunfeng; Luu, Hue H; Haydon, Rex C; Shi, Lewis L; Li, Tingyu; He, Tong-Chuan; Li, Ming

2013-09-01

Mesenchymal stem cells (MSCs) are multipotent progenitors, which give rise to several lineages, including bone, cartilage and fat. Epidermal growth factor (EGF) stimulates cell growth, proliferation and differentiation. EGF acts by binding with high affinity to epidermal growth factor receptor (EGFR) on the cell surface and stimulating the intrinsic protein tyrosine kinase activity of its receptor, which initiates a signal transduction cascade causing a variety of biochemical changes within the cell and regulating cell proliferation and differentiation. We have identified BMP9 as one of the most osteogenic BMPs in MSCs. In this study, we investigate if EGF signalling cross-talks with BMP9 and regulates BMP9-induced osteogenic differentiation. We find that EGF potentiates BMP9-induced early and late osteogenic markers of MSCs in vitro, which can be effectively blunted by EGFR inhibitors Gefitinib and Erlotinib or receptor tyrosine kinase inhibitors AG-1478 and AG-494 in a dose- and time-dependent manner. Furthermore, EGF significantly augments BMP9-induced bone formation in the cultured mouse foetal limb explants. In vivo stem cell implantation experiment reveals that exogenous expression of EGF in MSCs can effectively potentiate BMP9-induced ectopic bone formation, yielding larger and more mature bone masses. Interestingly, we find that, while EGF can induce BMP9 expression in MSCs, EGFR expression is directly up-regulated by BMP9 through Smad1/5/8 signalling pathway. Thus, the cross-talk between EGF and BMP9 signalling pathways in MSCs may underline their important roles in regulating osteogenic differentiation. Harnessing the synergy between BMP9 and EGF should be beneficial for enhancing osteogenesis in regenerative medicine. © 2013 The Authors. Journal of Cellular and Molecular Medicine Published by Foundation for Cellular and Molecular Medicine/Blackwell Publishing Ltd.

Cross-talk between EGF and BMP9 signalling pathways regulates the osteogenic differentiation of mesenchymal stem cells

PubMed Central

Liu, Xing; Qin, Jiaqiang; Luo, Qing; Bi, Yang; Zhu, Gaohui; Jiang, Wei; Kim, Stephanie H; Li, Mi; Su, Yuxi; Nan, Guoxin; Cui, Jing; Zhang, Wenwen; Li, Ruidong; Chen, Xiang; Kong, Yuhan; Zhang, Jiye; Wang, Jinhua; Rogers, Mary Rose; Zhang, Hongyu; Shui, Wei; Zhao, Chen; Wang, Ning; Liang, Xi; Wu, Ningning; He, Yunfeng; Luu, Hue H; Haydon, Rex C; Shi, Lewis L; Li, Tingyu; He, Tong-Chuan; Li, Ming

2013-01-01

Mesenchymal stem cells (MSCs) are multipotent progenitors, which give rise to several lineages, including bone, cartilage and fat. Epidermal growth factor (EGF) stimulates cell growth, proliferation and differentiation. EGF acts by binding with high affinity to epidermal growth factor receptor (EGFR) on the cell surface and stimulating the intrinsic protein tyrosine kinase activity of its receptor, which initiates a signal transduction cascade causing a variety of biochemical changes within the cell and regulating cell proliferation and differentiation. We have identified BMP9 as one of the most osteogenic BMPs in MSCs. In this study, we investigate if EGF signalling cross-talks with BMP9 and regulates BMP9-induced osteogenic differentiation. We find that EGF potentiates BMP9-induced early and late osteogenic markers of MSCs in vitro, which can be effectively blunted by EGFR inhibitors Gefitinib and Erlotinib or receptor tyrosine kinase inhibitors AG-1478 and AG-494 in a dose- and time-dependent manner. Furthermore, EGF significantly augments BMP9-induced bone formation in the cultured mouse foetal limb explants. In vivo stem cell implantation experiment reveals that exogenous expression of EGF in MSCs can effectively potentiate BMP9-induced ectopic bone formation, yielding larger and more mature bone masses. Interestingly, we find that, while EGF can induce BMP9 expression in MSCs, EGFR expression is directly up-regulated by BMP9 through Smad1/5/8 signalling pathway. Thus, the cross-talk between EGF and BMP9 signalling pathways in MSCs may underline their important roles in regulating osteogenic differentiation. Harnessing the synergy between BMP9 and EGF should be beneficial for enhancing osteogenesis in regenerative medicine. PMID:23844832

Lack of Obvious Influence of PLLA Nanofibers on the Gene Expression of BMP-2 and VEGF during Growth and Differentiation of Human Mesenchymal Stem Cells

PubMed Central

Schofer, Markus D.; Fuchs-Winkelmann, S.; Wack, C.; Rudisile, M.; Dersch, R.; Leifeld, I.; Wendorff, J.; Greiner, A.; Paletta, J. R. J.; Boudriot, U.

2009-01-01

Growth factors like bone morphogenetic protein 2 (BMP-2) and vascular endothelial growth factor (VEGF) play an important role in bone remodeling and fracture repair. Therefore, with respect to tissue engineering, an artificial graft should have no negative impact on the expression of these factors. In this context, the aim of this study was to analyze the impact of poly(L-lactic acid) (PLLA) nanofibers on VEGF and BMP-2 gene expression during the time course of human mesenchymal stem cell (hMSC) differentiation towards osteoblasts. PLLA matrices were seeded with hMSCs and cultivated over a period of 22 days under growth and osteoinductive conditions, and analyzed during the course of culture, with respect to gene expression of VEGF and BMP-2. Furthermore, BMP-2–enwoven PLLA nanofibers were used in order to elucidate whether initial down-regulation of growth factor expression could be compensated. Although there was a great interpatient variability with respect to the expression of VEGF and BMP-2, PLLA nanofibers tend to result in a down-regulation in BMP-2 expression during the early phase of cultivation. This effect was diminished in the case of VEGF gene expression. The initial down-regulation was overcome when BMP-2 was directly incorporated into the PLLA nanofibers by electrospinning. Furthermore, the incorporation of BMP-2 into the PLLA nanofibers resulted in an increase in VEGF gene expression. Summarized, the results indicate that the PLLA nanofibers have little effect on growth factor production. An enhancement in gene expression of BMP-2 and VEGF can be achieved by an incorporation of BMP-2 into the PLLA nanofibers. PMID:19412560

BMP-2/PLGA delayed-release microspheres composite graft, selection of bone particulate diameters, and prevention of aseptic inflammation for bone tissue engineering.

PubMed

Ji, Ye; Xu, Gong Ping; Zhang, Zhi Peng; Xia, Jing Jun; Yan, Jing Long; Pan, Shang Ha

2010-03-01

Autogenous bone grafts are widely used in the repair of bone defects. Growth factors such as bone morphogenetic protein 2 (BMP-2) can induce bone regeneration and enhance bone growth. The combination of an autogenous bone graft and BMP-2 may provide a better osteogenic effect than either treatment alone, but BMP-2 is easily inactivated in body fluid. The objective of this study was to develop a technique that can better preserve the in vivo activity of BMP-2 incorporated in bone grafts. In this study, we first prepared BMP-2/poly(lactic-co-glycolic acid) (PLGA) delayed-release microspheres, and then combined collagen, the delayed-release microspheres, and rat autologous bone particulates to form four groups of composite grafts with different combinations: collagen in group A; collagen combined with bone particulates in group B; collagen combined with BMP-2/PLGA delayed-release microspheres in group C; and collagen combined with both bone particulates and BMP-2/PLGA delayed-release microspheres in group D. The four groups of composite grafts were implanted into the gluteus maximus pockets in rats. The ectopic osteogenesis and ALP level in group D (experimental group) were compared with those in groups A, B, and C (control groups) to study whether it had higher osteogenic capability. Results showed that the composite graft design increased the utility of BMP-2 and reduced the required dose of BMP-2 and volume of autologous bone. The selection of bone particulate diameter had an impact on the osteogenetic potential of bone grafts. Collagen prevented the occurrence of aseptic inflammation and improved the osteoinductivity of BMP-2. These results showed that this composite graft design is effective and feasible for use in bone repair.

Alk2/ACVR1 and Alk3/BMPR1A Provide Essential Function for Bone Morphogenetic Protein-Induced Retinal Angiogenesis.

PubMed

Lee, Heon-Woo; Chong, Diana C; Ola, Roxana; Dunworth, William P; Meadows, Stryder; Ka, Jun; Kaartinen, Vesa M; Qyang, Yibing; Cleaver, Ondine; Bautch, Victoria L; Eichmann, Anne; Jin, Suk-Won

2017-04-01

Increasing evidence suggests that bone morphogenetic protein (BMP) signaling regulates angiogenesis. Here, we aimed to define the function of BMP receptors in regulating early postnatal angiogenesis by analysis of inducible, endothelial-specific deletion of the BMP receptor components Bmpr2 (BMP type 2 receptor), Alk1 (activin receptor-like kinase 1), Alk2 , and Alk3 in mouse retinal vessels. Expression analysis of several BMP ligands showed that proangiogenic BMP ligands are highly expressed in postnatal retinas. Consistently, BMP receptors are also strongly expressed in retina with a distinct pattern. To assess the function of BMP signaling in retinal angiogenesis, we first generated mice carrying an endothelial-specific inducible deletion of Bmpr2 . Postnatal deletion of Bmpr2 in endothelial cells substantially decreased the number of angiogenic sprouts at the vascular front and branch points behind the front, leading to attenuated radial expansion. To identify critical BMPR1s (BMP type 1 receptors) associated with BMPR2 in retinal angiogenesis, we generated endothelial-specific inducible deletion of 3 BMPR1s abundantly expressed in endothelial cells and analyzed the respective phenotypes. Among these, endothelial-specific deletion of either Alk2 / acvr1 or Alk3 / Bmpr1a caused a delay in radial expansion, reminiscent of vascular defects associated with postnatal endothelial-specific deletion of BMPR2, suggesting that ALK2/ACVR1 and ALK3/BMPR1A are likely to be the critical BMPR1s necessary for proangiogenic BMP signaling in retinal vessels. Our data identify BMP signaling mediated by coordination of ALK2/ACVR1, ALK3/BMPR1A, and BMPR2 as an essential proangiogenic cue for retinal vessels. © 2017 The Authors.

Site-Directed Immobilization of BMP-2: Two Approaches for the Production of Innovative Osteoinductive Scaffolds.

PubMed

Tabisz, Barbara; Schmitz, Werner; Schmitz, Michael; Luehmann, Tessa; Heusler, Eva; Rybak, Jens-Christoph; Meinel, Lorenz; Fiebig, Juliane E; Mueller, Thomas D; Nickel, Joachim

2017-03-13

The regenerative potential of bone is strongly impaired in pathological conditions, such as nonunion fractures. To support bone regeneration various scaffolds have been developed in the past, which have been functionalized with osteogenic growth factors such as bone morphogenetic proteins (BMPs). However, most of them required supra-physiological levels of these proteins leading to burst releases, thereby causing severe side effects. Site-specific, covalent coupling of BMP2 to implant materials might be an optimal strategy in order to overcome these problems. Therefore, we created a BMP-2 variant (BMP2-K3Plk) containing a noncanonical amino acid (propargyl-l-lysine) substitution introduced by genetic code expansion that allows for site-specific and covalent immobilization onto polymeric scaffold materials. To directly compare different coupling strategies, we also produced a BMP2 variant containing an additional cysteine residue (BMP2-A2C) allowing covalent coupling by thioether formation. The BMP2-K3Plk mutant was coupled to functionalized beads by a copper-catalyzed azide-alkyne cycloaddition (CuAAC) either directly or via a short biotin-PEG linker both with high specificity. After exposing the BMP-coated beads to C2C12 cells, ALP expression appeared locally restricted in close proximity to these beads, showing that both coupled BMP2 variants trigger cell differentiation. The advantage of our approach over non-site-directed immobilization techniques is the ability to produce fully defined osteogenic surfaces, allowing for lower BMP2 loads and concomitant higher bioactivities, for example, due to controlled orientation toward BMP2 receptors. Such products might provide superior bone healing capabilities with potential safety advantages as of homogeneous product outcome.

Creation of bony microenvironment with CaP and cell-derived ECM to enhance human bone-marrow MSC behavior and delivery of BMP-2

PubMed Central

Kang, Yunqing; Kim, Sungwoo; Khademhosseini, Ali; Yang, Yunzhi

2011-01-01

Extracellular matrix (ECM) comprises a rich meshwork of proteins and proteoglycans, which not only contains biological cues for cell behavior, but is also a reservoir for binding growth factors and controlling their release. Here we aimed to create a suitable bony microenvironment with cell-derived ECM and biodegradable β-tricalcium phosphate (β-TCP). More specifically, we investigated whether the ECM produced by bone marrow-derived mesenchymal stem cells (hBMSC) on a β-TCP scaffold can bind bone morphogenetic protein-2 (BMP-2) and control its release in a sustained manner, and further examined the effect of ECM and the BMP-2 released from ECM on cell behaviors. The ECM was obtained through culturing the hBMSC on a β-TCP porous scaffold and performing decellularization and sterilization. SEM, XPS, FTIR, and immunofluorescent staining results indicated the presence of ECM on the β-TCP and the amount of ECM increased with the incubation time. BMP-2 was loaded onto the β-TCP with and without ECM by immersing the scaffolds in the BMP-2 solution. The loading and release kinetics of the BMP-2 on the β-TCP/ECM were significantly slower than those on the β-TCP. The β-TCP/ECM exhibited a sustained release profile of the BMP-2, which was also affected by the amount of ECM. This is probably because the β-TCP/ECM has different binding mechanisms with BMP-2. The β-TCP/ECM promoted cell proliferation. Furthermore, the BMP-2-loaded β-TCP/ECM stimulated reorganization of the actin cytoskeleton, increased expression of alkaline phosphatase and calcium deposition by the cells compared to those without BMP-2 loading and the β-TCP with BMP-2 loading. PMID:21632105

A BMP7 Variant Inhibits Tumor Angiogenesis In Vitro and In Vivo through Direct Modulation of Endothelial Cell Biology.

PubMed

Tate, Courtney M; Mc Entire, Jacquelyn; Pallini, Roberto; Vakana, Eliza; Wyss, Lisa; Blosser, Wayne; Ricci-Vitiani, Lucia; D'Alessandris, Quintino Giorgio; Morgante, Liliana; Giannetti, Stefano; Larocca, Luigi Maria; Todaro, Matilde; Benfante, Antonina; Colorito, Maria Luisa; Stassi, Giorgio; De Maria, Ruggero; Rowlinson, Scott; Stancato, Louis

2015-01-01

Bone morphogenetic proteins (BMPs), members of the TGF-β superfamily, have numerous biological activities including control of growth, differentiation, and vascular development. Using an in vitro co-culture endothelial cord formation assay, we investigated the role of a BMP7 variant (BMP7v) in VEGF, bFGF, and tumor-driven angiogenesis. BMP7v treatment led to disruption of neo-endothelial cord formation and regression of existing VEGF and bFGF cords in vitro. Using a series of tumor cell models capable of driving angiogenesis in vitro, BMP7v treatment completely blocked cord formation. Pre-treatment of endothelial cells with BMP7v significantly reduced their cord forming ability, indicating a direct effect on endothelial cell function. BMP7v activated the canonical SMAD signaling pathway in endothelial cells but targeted gene knockdown using shRNA directed against SMAD4 suggests this pathway is not required to mediate the anti-angiogenic effect. In contrast to SMAD activation, BMP7v selectively decreased ERK and AKT activation, significantly decreased endothelial cell migration and down-regulated expression of critical RTKs involved in VEGF and FGF angiogenic signaling, VEGFR2 and FGFR1 respectively. Importantly, in an in vivo angiogenic plug assay that serves as a measurement of angiogenesis, BMP7v significantly decreased hemoglobin content indicating inhibition of neoangiogenesis. In addition, BMP7v significantly decreased angiogenesis in glioblastoma stem-like cell (GSLC) Matrigel plugs and significantly impaired in vivo growth of a GSLC xenograft with a concomitant reduction in microvessel density. These data support BMP7v as a potent anti-angiogenic molecule that is effective in the context of tumor angiogenesis.

BMP-2 up-regulates PTEN expression and induces apoptosis of pulmonary artery smooth muscle cells under hypoxia.

PubMed

Pi, Weifeng; Guo, Xuejun; Su, Liping; Xu, Weiguo

2012-01-01

To investigate the role of bone morphogenetic protein 2 (BMP-2) in regulation of phosphatase and tensin homologue deleted on chromosome ten (PTEN) and apoptosis of pulmonary artery smooth muscle cells (PASMCs) under hypoxia. Normal human PASMCs were cultured in growth medium (GM) and treated with BMP-2 from 5-80 ng/ml under hypoxia (5% CO(2)+94% N(2)+1% O(2)) for 72 hours. Gene expression of PTEN, AKT-1 and AKT-2 were determined by quantitative RT-PCR (QRT-PCR). Protein expression levels of PTEN, AKT and phosph-AKT (pAKT) were determined. Apoptosis of PASMCs were determined by measuring activities of caspases-3, -8 and -9. siRNA-smad-4, bpV(HOpic) (PTEN inhibitor) and GW9662 (PPARγ antagonist) were used to determine the signalling pathways. Proliferation of PASMCs showed dose dependence of BMP-2, the lowest proliferation rate was achieved at 60 ng/ml concentration under hypoxia (82.2±2.8%). BMP-2 increased PTEN gene expression level, while AKT-1 and AKT-2 did not change. Consistently, the PTEN protein expression also showed dose dependence of BMP-2. AKT activity significantly reduced in BMP-2 treated PASMCs. Increased activities of caspase-3, -8 and -9 of PASMCs were found after cultured with BMP-2. PTEN expression remained unchanged when Smad-4 expression was inhibited by siRNA-Smad-4. bpV(HOpic) and GW9662 (PPARγ inhibitor) inhibited PTEN protein expression and recovered PASMCs proliferation rate. BMP-2 increased PTEN expression under hypoxia in a dose dependent pattern. BMP-2 reduced AKT activity and increased caspase activity of PASMCs under hypoxia. The increased PTEN expression may be mediated through PPARγ signalling pathway, instead of BMP/Smad signalling pathway.

Regulation of bone morphogenetic proteins in early embryonic development

NASA Astrophysics Data System (ADS)

Yamamoto, Yukiyo; Oelgeschläger, Michael

2004-11-01

Bone morphogenetic proteins (BMPs), a large subgroup of the TGF-β family of secreted growth factors, control fundamental events in early embryonic development, organogenesis and adult tissue homeostasis. The plethora of dose-dependent cellular processes regulated by BMP signalling demand a tight regulation of BMP activity. Over the last decade, a number of proteins have been identified that bind BMPs in the extracellular space and regulate the interaction of BMPs with their cognate receptors, including the secreted BMP antagonist Chordin. In the early vertebrate embryo, the localized secretion of BMP antagonists from the dorsal blastopore lip establishes a functional BMP signalling gradient that is required for the determination of the dorsoventral or back to belly body axis. In particular, inhibition of BMP activity is essential for the formation of neural tissue in the development of vertebrate and invertebrate embryos. Here we review recent studies that have provided new insight into the regulation of BMP signalling in the extracellular space. In particular, we discuss the recently identified Twisted gastrulation protein that modulates, in concert with metalloproteinases of the Tolloid family, the interaction of Chordin with BMP and a family of proteins that share structural similarities with Chordin in the respective BMP binding domains. In addition, genetic and functional studies in zebrafish and frog provide compelling evidence that the secreted protein Sizzled functionally interacts with the Chd BMP pathway, despite being expressed ventrally in the early gastrula-stage embryo. These intriguing discoveries may have important implications, not only for our current concept of early embryonic patterning, but also for the regulation of BMP activity at later developmental stages and tissue homeostasis in the adult.

An FGF3-BMP Signaling Axis Regulates Caudal Neural Tube Closure, Neural Crest Specification and Anterior-Posterior Axis Extension

PubMed Central

Anderson, Matthew J.; Schimmang, Thomas; Lewandoski, Mark

2016-01-01

During vertebrate axis extension, adjacent tissue layers undergo profound morphological changes: within the neuroepithelium, neural tube closure and neural crest formation are occurring, while within the paraxial mesoderm somites are segmenting from the presomitic mesoderm (PSM). Little is known about the signals between these tissues that regulate their coordinated morphogenesis. Here, we analyze the posterior axis truncation of mouse Fgf3 null homozygotes and demonstrate that the earliest role of PSM-derived FGF3 is to regulate BMP signals in the adjacent neuroepithelium. FGF3 loss causes elevated BMP signals leading to increased neuroepithelium proliferation, delay in neural tube closure and premature neural crest specification. We demonstrate that elevated BMP4 depletes PSM progenitors in vitro, phenocopying the Fgf3 mutant, suggesting that excessive BMP signals cause the Fgf3 axis defect. To test this in vivo we increased BMP signaling in Fgf3 mutants by removing one copy of Noggin, which encodes a BMP antagonist. In such mutants, all parameters of the Fgf3 phenotype were exacerbated: neural tube closure delay, premature neural crest specification, and premature axis termination. Conversely, genetically decreasing BMP signaling in Fgf3 mutants, via loss of BMP receptor activity, alleviates morphological defects. Aberrant apoptosis is observed in the Fgf3 mutant tailbud. However, we demonstrate that cell death does not cause the Fgf3 phenotype: blocking apoptosis via deletion of pro-apoptotic genes surprisingly increases all Fgf3 defects including causing spina bifida. We demonstrate that this counterintuitive consequence of blocking apoptosis is caused by the increased survival of BMP-producing cells in the neuroepithelium. Thus, we show that FGF3 in the caudal vertebrate embryo regulates BMP signaling in the neuroepithelium, which in turn regulates neural tube closure, neural crest specification and axis termination. Uncovering this FGF3-BMP signaling axis is a major advance toward understanding how these tissue layers interact during axis extension with important implications in human disease. PMID:27144312

Bmp6 Expression in Murine Liver Non Parenchymal Cells: A Mechanism to Control their High Iron Exporter Activity and Protect Hepatocytes from Iron Overload?

PubMed Central

Rausa, Marco; Pagani, Alessia; Nai, Antonella; Campanella, Alessandro; Gilberti, Maria Enrica; Apostoli, Pietro; Camaschella, Clara; Silvestri, Laura

2015-01-01

Bmp6 is the main activator of hepcidin, the liver hormone that negatively regulates plasma iron influx by degrading the sole iron exporter ferroportin in enterocytes and macrophages. Bmp6 expression is modulated by iron but the molecular mechanisms are unknown. Although hepcidin is expressed almost exclusively by hepatocytes (HCs), Bmp6 is produced also by non-parenchymal cells (NPCs), mainly sinusoidal endothelial cells (LSECs). To investigate the regulation of Bmp6 in HCs and NPCs, liver cells were isolated from adult wild type mice whose diet was modified in iron content in acute or chronic manner and in disease models of iron deficiency (Tmprss6 KO mouse) and overload (Hjv KO mouse). With manipulation of dietary iron in wild-type mice, Bmp6 and Tfr1 expression in both HCs and NPCs was inversely related, as expected. When hepcidin expression is abnormal in murine models of iron overload (Hjv KO mice) and deficiency (Tmprss6 KO mice), Bmp6 expression in NPCs was not related to Tfr1. Despite the low Bmp6 in NPCs from Tmprss6 KO mice, Tfr1 mRNA was also low. Conversely, despite body iron overload and high expression of Bmp6 in NPCs from Hjv KO mice, Tfr1 mRNA and protein were increased. However, in the same cells ferritin L was only slightly increased, but the iron content was not, suggesting that Bmp6 in these cells reflects the high intracellular iron import and export. We propose that NPCs, sensing the iron flux, not only increase hepcidin through Bmp6 with a paracrine mechanism to control systemic iron homeostasis but, controlling hepcidin, they regulate their own ferroportin, inducing iron retention or release and further modulating Bmp6 production in an autocrine manner. This mechanism, that contributes to protect HC from iron loading or deficiency, is lost in disease models of hepcidin production. PMID:25860887

Bone morphogenetic protein 4 induces differentiation of colorectal cancer stem cells and increases their response to chemotherapy in mice.

PubMed

Lombardo, Ylenia; Scopelliti, Alessandro; Cammareri, Patrizia; Todaro, Matilde; Iovino, Flora; Ricci-Vitiani, Lucia; Gulotta, Gaspare; Dieli, Francesco; de Maria, Ruggero; Stassi, Giorgio

2011-01-01

The limited clinical response observed in many patients with colorectal cancer may be related to the presence of chemoresistant colorectal cancer stem cells (CRC-SCs). Bone morphogenetic protein 4 (BMP4) promotes the differentiation of normal colonic stem cells. We investigated whether BMP4 might be used to induce differentiation of CRC-SCs and for therapeutic purposes. CRC-SCs were isolated from 25 tumor samples based on expression of CD133 or using a selection culture medium. BMP4 expression and activity on CRC-SCs were evaluated in vitro; progeny of the stem cells were evaluated by immunofluorescence, immunoblot, and flow cytometry analyses. The potential therapeutic effect of BMP4 was assessed in immunocompromised mice after injection of CRC-SCs that responded to chemotherapy (n = 4) or that did not (n = 2). CRC-SCs did not express BMP4 whereas differentiated cells did. Recombinant BMP4 promoted differentiation and apoptosis of CRC-SCs in 12 of 15 independent experiments; this effect did not depend on Small Mothers against decapentaplegic (Smad)4 expression level or microsatellite stability. BMP4 activated the canonical and noncanonical BMP signaling pathways, including phosphoInositide 3-kinase (PI3K) and PKB (protein kinase B)/AKT. Mutations in PI3K or loss of Phosphatase and Tensin homolog (PTEN) in Smad4-defective tumors made CRC-SCs unresponsive to BMP4. Administration of BMP4 to immunocompromised mice with tumors that arose from CRC-SCs increased the antitumor effects of 5-fluorouracil and oxaliplatin. BMP4 promotes terminal differentiation, apoptosis, and chemosensitization of CRC-SCs in tumors that do not have simultaneous mutations in Smad4 and constitutive activation of PI3K. BMP4 might be developed as a therapeutic agent against cancer stem cells in advanced colorectal tumors. Copyright © 2011 AGA Institute. Published by Elsevier Inc. All rights reserved.

Lactoferricin Enhances BMP7-Stimulated Anabolic Pathways in Intervertebral Disc Cells

PubMed Central

Ellman, Michael B; Kim, Jaesung; An, Howard S; Chen, Di; Kc, Ranjan; Li, Xin; Xiao, Guozhi; Yan, Dongyao; Suh, Joon; van Wijnen, Andre J.; Wang, James H-C; Kim, Su-Gwan; Im, Hee-Jeong

2013-01-01

Bone-morphogenetic protein-7 (BMP7) is a well-known anabolic and anti-catabolic growth factor on intervertebral (IVD) matrix and cell homeostasis. Similarly, lactoferricin B (LfcinB) has recently been shown to have pro-anabolic, anti-catabolic, anti-oxidative and/or anti-inflammatory effects in bovine disc cells in vitro. In this study, we investigated the potential benefits of using combined peptide therapy with LfcinB and BMP7 for intervertebral disc (IVD) matrix repair and to understand cellular and signaling mechanisms controlled by these factors. We studied the effects of BMP7 and LfcinB as individual treatments and combined therapy on bovine nucleus pulposus (NP) cells by assessing proteoglycan (PG) accumulation and synthesis, and the expression of matrix protein aggrecan and transcription factor SOX-9. We also analyzed the role of noggin, a BMP antagonist, in IVD tissue and examined its effect after stimulation with LfcinB. To understand the molecular mechanisms by which LfcinB synergizes with BMP7, we investigated the ERK-SP1 axis as a downstream intracellular signaling regulator involved in BMP7 and LfcinB-mediated activities. Treatment of bovine NP cells cultured in alginate with LfcinB plus BMP7 synergistically stimulates PG synthesis and accumulation in part by upregulation of aggrecan gene expression. The synergism results from LfcinB-mediated activation of Sp1 and SMAD signaling pathways by (i) phosphorylation of SMAD 1/5/8; (ii) downregulation of SMAD inhibitory factors [i.e., noggin (BMP receptor antagonist) and SMAD6 (inhibitory SMAD)]; and (iii) upregulation of SMAD4 (universal co-SMAD). These data indicate that LfcinB-suppression of noggin may eliminate the negative feedback of BMP7, thereby maximizing biological activity of BMP7 and ultimately shifting homeostasis to a pro-anabolic state in disc cells. We propose that combination growth factor therapy using BMP7 and LfcinB may be beneficial for treatment of disc degeneration. PMID:23644135

Synergistic roles of bone morphogenetic protein 15 and growth differentiation factor 9 in ovarian function.

PubMed

Yan, C; Wang, P; DeMayo, J; DeMayo, F J; Elvin, J A; Carino, C; Prasad, S V; Skinner, S S; Dunbar, B S; Dube, J L; Celeste, A J; Matzuk, M M

2001-06-01

Knockout mouse technology has been used over the last decade to define the essential roles of ovarian-expressed genes and uncover genetic interactions. In particular, we have used this technology to study the function of multiple members of the transforming growth factor-beta superfamily including inhibins, activins, and growth differentiation factor 9 (GDF-9 or Gdf9). Knockout mice lacking GDF-9 are infertile due to a block in folliculogenesis at the primary follicle stage. In addition, recombinant GDF-9 regulates multiple cumulus granulosa cell functions in the periovulatory period including hyaluronic acid synthesis and cumulus expansion. We have also cloned an oocyte-specific homolog of GDF-9 from mice and humans, which is termed bone morphogenetic protein 15 (BMP-15 or Bmp15). To define the function of BMP-15 in mice, we generated embryonic stem cells and knockout mice, which have a null mutation in this X-linked gene. Male chimeric and Bmp15 null mice are normal and fertile. In contrast to Bmp15 null males and Gdf9 knockout females, Bmp15 null females (Bmp15(-/-)) are subfertile and usually have minimal ovarian histopathological defects, but demonstrate decreased ovulation and fertilization rates. To further decipher possible direct or indirect genetic interactions between GDF-9 and BMP-15, we have generated double mutant mice lacking one or both alleles of these related homologs. Double homozygote females (Bmp15(-/-)Gdf9(-/-)) display oocyte loss and cysts and resemble Gdf9(-/-) mutants. In contrast, Bmp15(-/-)Gdf9(+/-) female mice have more severe fertility defects than Bmp15(-/-) females, which appear to be due to abnormalities in ovarian folliculogenesis, cumulus cell physiology, and fertilization. Thus, the dosage of intact Bmp15 and Gdf9 alleles directly influences the destiny of the oocyte during folliculogenesis and in the periovulatory period. These studies have important implications for human fertility control and the maintenance of fertility and normal ovarian physiology.

[Preparation of vanilline cross-linked rhBMP-2/chitosan microspheres and its effect on mesenchymal stem cells].

PubMed

Wu, Gui; Wang, Hai; Qiu, Guixing; Yu, Xin; Su, Xinlin; Ma, Pei; Yin, Bo; Wu, Zhihong

2015-06-02

To prepare rhBMP-2/chitosan microspheres (rhBMP-2 CMs) with vanilline as a cross-linking reagent and study the biocompatibility and drug release characteristic of microspheres in vitro. Emulsion cross-linking method was utilized to prepare rhBMP-2 CMs, Scanning electron microscope (SEM) was used to observe the microstructure of microspheres.Leaching solution of microspheres and blank culture medium were designated as experimental and control groups respectively. Both groups were cultured with human mesenchymal stem cells (hMSCs) to determine its cytotoxicity and its effect on the proliferation of hMSCs. Dynamic immersion method was used to examine the in vitro release characteristic of rhBMP-2. And the alkaline phosphatase (ALP) activity of hMSCs was determined to reveal the bioactivity of released rhBMP-2. The rhBMP-2 CMs were spherical under SEM.After treating with leaching solution for 24 and 48 h, there was no inter-group statistical difference in optical density (OD) values at both timepoints (24 h:0.72 ± 0.07 vs 0.73 ± 0.05, P > 0.05; 48 h:1.19 ± 0.11 vs 1.27 ± 0.06, P > 0.05). After culturing with leaching solution for 1, 3 and 7 days, the number of cells increased with time for both groups. And the OD values were not statistically different at each timepoint. Five milligram rhBMP-2 CMs soaked for 19 days with a gradual release of rhBMP-2. The concentration of rhBMP-2 was 216.1 ± 20.0 ng/ml at Day 19. At Days 3 and 7, the ALP activities of hMSCs were (0.50 ± 0.07) and (0.68 ± 0.06) µmol pNPP·min⁻¹·mg⁻¹ protein respectively and both were higher than that of blank culture medium group (0.14 ± 0.01) (P < 0.05). With an excellent biocompatibility, rhBMP-2 CMs may be an ideal carrier for control-released rhBMP-2 and encapsulated rhBMP-2 remains bioactive.

Effect of grafting RGD and BMP-2 protein-derived peptides to a hydrogel substrate on osteogenic differentiation of marrow stromal cells.

PubMed

He, Xuezhong; Ma, Junyu; Jabbari, Esmaiel

2008-11-04

Osteogenic differentiation and mineralization of bone marrow stromal (BMS) cells depends on the cells' interactions with bioactive peptides associated with the matrix proteins. The RGD peptides of ECM proteins interact with BMS cells through integrin surface receptors to facilitate cell spreading and adhesion. The BMP peptide corresponding to residues 73-92 of bone morphogenetic protein-2 promotes differentiation and mineralization of BMS cells. The objective of this work was to investigate the effects of RGD and BMP peptides, grafted to a hydrogel substrate, on osteogenic differentiation and mineralization of BMS cells. RGD peptide was acrylamide-terminated by reacting acrylic acid with the N-terminal amine group of the peptide to produce the functionalized Ac-GRGD peptide. The PEGylated BMP peptide was reacted with 4-carboxybenzenesulfonazide to produce an azide functionalized Az-mPEG-BMP peptide. Poly (lactide-co-ethylene oxide- co-fumarate) (PLEOF) macromer was cross-linked with Ac-GRGD peptide and propargyl acrylate to produce an RGD conjugated hydrogel. Az-mPEG-BMP peptide was grafted to the hydrogel by "click chemistry". The RGD and BMP peptide density on the hydrogel surface was 1.62+/-0.37 and 5.2+/-0.6 pmol/cm2, respectively. BMS cells were seeded on the hydrogels and the effect of RGD and BMP peptides on osteogenesis was evaluated by measuring ALPase activity and calcium content with incubation time. BMS cells cultured on RGD conjugated, BMP peptide grafted, and RGD+BMP peptide modified hydrogels showed 3, 2.5, and 5-fold increase in ALPase activity after 14 days incubation. BMS cells seeded on RGD+BMP peptides modified hydrogel showed 4.9- and 11.8-fold increase in calcium content after 14 and 21 days, respectively, which was significantly higher than RGD conjugated or BMP grafted hydrogels. These results demonstrate that RGD and BMP peptides, grafted to a hydrogel substrate, act synergistically to enhance osteogenic differentiation and mineralization of BMS cells. These findings are potentially useful in developing engineered scaffolds for bone regeneration.

Activin A amplifies dysregulated BMP signaling and induces chondro-osseous differentiation of primary connective tissue progenitor cells in patients with fibrodysplasia ossificans progressiva (FOP).

PubMed

Wang, Haitao; Shore, Eileen M; Pignolo, Robert J; Kaplan, Frederick S

2018-04-01

Fibrodysplasia ossificans progressiva (FOP), is caused by mutations in the type I BMP receptor ACVR1 that lead to increased activation of the BMP-pSmad1/5/8 signaling pathway. Recent findings suggest that Activin A (Act A) promiscuously stimulates the bone morphogenetic protein (BMP) signaling pathway in vitro and mediates heterotopic ossification (HO) in mouse models of FOP, but primary data from FOP patient cells are lacking. To examine BMP-pSmad1/5/8 pathway signaling and chondro-osseous differentiation in response to endogenous and exogenous Act A in primary connective tissue progenitor cells [CTPCs; also known as stem cells from human exfoliated deciduous teeth (SHED) cells] from patients with FOP. These cells express the common FOP mutation, ACVR1 (R206H). We found that Act A amplifies dysregulated BMP pathway signaling in human FOP primary CTPCs cells through the Smad1/5/8 pathway and induces chondro-osseous differentiation. Amplification of BMP-pSmad1/5/8 signaling was inhibited by Follistatin and by a neutralizing antibody to Activin A. The increased basal pSmad1/5/8 activity, as well as the hypoxia-induced stimulation of FOP CTPCs cells, were BMP4 and Act A independent. Importantly, either BMP4 or Act A stimulated pSmad1/5/8 pathway signaling but BMP4 signaling was not dependent on Activin A and vice versa. Circulating plasma levels of Act A or BMP4 are similar in controls compared to FOP patients, and suggest the potential for an autocrine or paracrine route for pathological signaling. The mutated FOP receptor [ACVR1 (R206H)] is hypersensitive to BMP4 and uniquely sensitive (compared to the wild type receptor) to Act A. Both canonical and non-canonical ligands have a synergistic effect on BMP-pSmad1/5/8 signaling in FOP CTPCs and may cooperate to alter the threshold for HO in FOP. Our findings in primary human FOP CTPCs have important implications for the design of clinical trials to inhibit dysregulated BMP pathway signaling in humans who have FOP. Copyright © 2017 Elsevier Inc. All rights reserved.

Use of Bioresorbable Hydrogels and Genetic Engineering to Accomplish Rapid Stabilization and Healing in Segmental Long Bone Defects

DTIC Science & Technology

2013-04-29

transduction of human mesenchymal stem cells (MSCs), BMP2 was not detectable by Western blotting, whereas high levels of the protein were produced by A549 (human... mesenchymal stem cells , generating high levels of BMP2. When Ad5BMP2 or Ad5F35BMP2 were compared in vitro for their ability to induce BMP2 synthesis...in human mesenchymal stem cells and in vivo for their ability to stimulate formation of heterotopic bone, mineralized bone was radiologically

Treatment BMP technology report.

DOT National Transportation Integrated Search

2006-04-01

The Treatment BMP Technology Report consolidates and standardizes information on storm : water quality technologies that are part of the California Department of Transportations : (Departments) BMP identification, and evaluation process describ...

Structural analyses of von Willebrand factor C domains of collagen 2A and CCN3 reveal an alternative mode of binding to bone morphogenetic protein-2.

PubMed

Xu, Emma-Ruoqi; Blythe, Emily E; Fischer, Gerhard; Hyvönen, Marko

2017-07-28

Bone morphogenetic proteins (BMPs) are secreted growth factors that promote differentiation processes in embryogenesis and tissue development. Regulation of BMP signaling involves binding to a variety of extracellular proteins, among which are many von Willebrand factor C (vWC) domain-containing proteins. Although the crystal structure of the complex of crossveinless-2 (CV-2) vWC1 and BMP-2 previously revealed one mode of the vWC/BMP-binding mechanism, other vWC domains may bind to BMP differently. Here, using X-ray crystallography, we present for the first time structures of the vWC domains of two proteins thought to interact with BMP-2: collagen IIA and matricellular protein CCN3. We found that these two vWC domains share a similar N-terminal fold that differs greatly from that in CV-2 vWC, which comprises its BMP-2-binding site. We analyzed the ability of these vWC domains to directly bind to BMP-2 and detected an interaction only between the collagen IIa vWC and BMP-2. Guided by the collagen IIa vWC domain crystal structure and conservation of surface residues among orthologous domains, we mapped the BMP-binding epitope on the subdomain 1 of the vWC domain. This binding site is different from that previously observed in the complex between CV-2 vWC and BMP-2, revealing an alternative mode of interaction between vWC domains and BMPs. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Increased BMP6 levels in the brains of Alzheimer's disease patients and APP transgenic mice are accompanied by impaired neurogenesis.

PubMed

Crews, Leslie; Adame, Anthony; Patrick, Christina; Delaney, Alexandra; Pham, Emiley; Rockenstein, Edward; Hansen, Lawrence; Masliah, Eliezer

2010-09-15

During aging and in the progression of Alzheimer's disease (AD), synaptic plasticity and neuronal integrity are disturbed. In addition to the alterations in plasticity in mature neurons, the neurodegenerative process in AD has been shown to be accompanied by alterations in neurogenesis. Members of the bone morphogenetic protein (BMP) family of growth factors have been implicated as important regulators of neurogenesis and neuronal cell fate determination during development; however, their role in adult neurogenesis and in AD is less clear. We show here by qRT-PCR analysis that BMP6 mRNA levels were significantly increased in the hippocampus of human patients with AD and in APP transgenic mice compared to controls. Immunoblot and immunohistochemical analyses confirmed that BMP6 protein levels were increased in human AD brains and APP transgenic mouse brains compared to controls and accumulated around hippocampal plaques. The increased levels of BMP6 were accompanied by defects in hippocampal neurogenesis in AD patients and APP transgenic mice. In support of a role for BMP6 in defective neurogenesis in AD, we show in an in vitro model of adult neurogenesis that treatment with amyloid-β(1-42) protein (Aβ) resulted in increased expression of BMP6, and that exposure to recombinant BMP6 resulted in reduced proliferation with no toxic effects. Together, these results suggest that Aβ-associated increases in BMP6 expression in AD may have deleterious effects on neurogenesis in the hippocampus, and therapeutic approaches could focus on normalization of BMP6 levels to protect against AD-related neurogenic deficits.

BMP15 Mutations Associated With Primary Ovarian Insufficiency Reduce Expression, Activity, or Synergy With GDF9.

PubMed

Patiño, Liliana C; Walton, Kelly L; Mueller, Thomas D; Johnson, Katharine E; Stocker, William; Richani, Dulama; Agapiou, David; Gilchrist, Robert B; Laissue, Paul; Harrison, Craig A

2017-03-01

Bone morphogenetic protein (BMP)15 is an oocyte-specific growth factor, which, together with growth differentiation factor (GDF) 9, regulates folliculogenesis and ovulation rate. Multiple mutations in BMP15 have been identified in women with primary ovarian insufficiency (POI), supporting a pathogenic role; however, the underlying biological mechanism of many of these mutants remains unresolved. To determine how mutations associated with ovarian dysfunction alter the biological activity of human BMP15. The effects of 10 mutations in BMP15 on protein production, activation of granulosa cells, and synergy with GDF9 were assessed. Sequencing of 35 patients with POI identified both an unrecognized BMP15 variant (c.986G>A, R329H) and a variant (c.581T>C, F194S) previously associated with the condition. Assessing expression and activity of these and 8 other BMP15 mutants identified: (1) multiple variants, including L148P, F194S, and Y235C, with reduced mature protein production; (2) three variants (R138H, A180T, and R329H) with ∼fourfold lower activity than wild-type BMP15; and (3) 3 variants (R68W, F194S, and N196K) with a significantly reduced ability to synergize with GDF9. Mutations in BMP15 associated with POI reduce mature protein production, activity, or synergy with GDF9. The latter effect is perhaps most interesting given that interactions with GDF9 most likely underlie the physiology of BMP15 in the human ovary. Copyright © 2017 by the Endocrine Society

Hedgehog induction of murine vasculogenesis is mediated by Foxf1 and Bmp4.

PubMed

Astorga, Jeanette; Carlsson, Peter

2007-10-01

The first vasculature of the developing vertebrate embryo forms by assembly of endothelial cells into simple tubes from clusters of mesodermal angioblasts. Maturation of this vasculature involves remodeling, pruning and investment with mural cells. Hedgehog proteins are part of the instructive endodermal signal that triggers the assembly of the first primitive vessels in the mesoderm. We used a combination of genetic and in vitro culture methods to investigate the role of hedgehogs and their targets in murine extraembryonic vasculogenesis. We show that Bmps, in particular Bmp4, are crucial for vascular tube formation, that Bmp4 expression in extraembryonic tissues requires the forkhead transcription factor Foxf1 and that the role of hedgehog proteins in this process is to activate Foxf1 expression in the mesoderm. We show in the allantois that genetic disruption of hedgehog signaling (Smo(-/-)) has no effect on Foxf1 expression, and neither Bmp4 expression nor vasculogenesis are disturbed. By contrast, targeted inactivation of Foxf1 leads to loss of allantoic Bmp4 and vasculature. In vitro, the avascular Foxf1(-/-) phenotype can be rescued by exogenous Bmp4, and vasculogenesis in wild-type tissue can be blocked by the Bmp antagonist noggin. Hedgehogs are required for activation of Foxf1, Bmp4 expression and vasculogenesis in the yolk sac. However, vasculogenesis in Smo(-/-) yolk sacs can be rescued by exogenous Bmp4, consistent with the notion that the role of hedgehog signaling in primary vascular tube formation is as an activator of Bmp4, via Foxf1.

DAN (NBL1) specifically antagonizes BMP2 and BMP4 and modulates the actions of GDF9, BMP2, and BMP4 in the rat ovary.

PubMed

Hung, Wei-Ting; Wu, Fang-Ju; Wang, Chun-Jen; Luo, Ching-Wei

2012-05-01

Although differential screening-selected gene aberrative in neuroblastoma (DAN, official symbol NBL1) is the founding member of the DAN subfamily of bone morphogenetic protein (BMP) antagonists, its antagonizing targets, gene regulation, and physiological functions remain unclear. Using diverse cell expression systems, we found that the generation of bioactive DAN is likely to be cell type specific. Unlike other phylogenetically close members, which are covalently linked homodimers, DAN forms a noncovalently linked homodimer during folding. Purified recombinant DAN specifically blocked signaling of BMP2 and BMP4 but not that of other ovarian-expressed transforming growth factor-beta members. Although widely distributed in many organs, DAN transcript level was periodically regulated by gonadotropins. Ovarian microdissection indicated that NBL1 (DAN) mRNA is mainly expressed in granulosa cells, where its transcript level is up-regulated by the gonadotropin-driven cAMP cascade. We further investigated the local regulation and ovarian functions of DAN. NBL1 (DAN) mRNA expression in granulosa cells was up-regulated by oocyte-derived growth differentiation factor 9 (GDF9), whereas treatment with DAN significantly reversed the inhibitory effect of BMP4 on follicle-stimulating hormone-induced progesterone production in cultured granulosa cells. Our findings suggest the DAN gradient in granulosa cells, established by oocyte-derived GDF9, may serve as an antagonist barrier that modulates the actions of theca-derived BMP4 and granulosa/theca-derived BMP2 during folliculogenesis both spatially and temporally.

TGF-β/BMP signaling and other molecular events: regulation of osteoblastogenesis and bone formation

PubMed Central

Rahman, Md Shaifur; Akhtar, Naznin; Jamil, Hossen Mohammad; Banik, Rajat Suvra; Asaduzzaman, Sikder M

2015-01-01

Transforming growth factor-beta (TGF-β)/bone morphogenetic protein (BMP) plays a fundamental role in the regulation of bone organogenesis through the activation of receptor serine/threonine kinases. Perturbations of TGF-β/BMP activity are almost invariably linked to a wide variety of clinical outcomes, i.e., skeletal, extra skeletal anomalies, autoimmune, cancer, and cardiovascular diseases. Phosphorylation of TGF-β (I/II) or BMP receptors activates intracellular downstream Smads, the transducer of TGF-β/BMP signals. This signaling is modulated by various factors and pathways, including transcription factor Runx2. The signaling network in skeletal development and bone formation is overwhelmingly complex and highly time and space specific. Additive, positive, negative, or synergistic effects are observed when TGF-β/BMP interacts with the pathways of MAPK, Wnt, Hedgehog (Hh), Notch, Akt/mTOR, and miRNA to regulate the effects of BMP-induced signaling in bone dynamics. Accumulating evidence indicates that Runx2 is the key integrator, whereas Hh is a possible modulator, miRNAs are regulators, and β-catenin is a mediator/regulator within the extensive intracellular network. This review focuses on the activation of BMP signaling and interaction with other regulatory components and pathways highlighting the molecular mechanisms regarding TGF-β/BMP function and regulation that could allow understanding the complexity of bone tissue dynamics. PMID:26273537

Selective inhibitors of the osteoblast proteasome stimulate bone formation in vivo and in vitro

PubMed Central

Garrett, I.R.; Chen, D.; Gutierrez, G.; Zhao, M.; Escobedo, A.; Rossini, G.; Harris, S.E.; Gallwitz, W.; Kim, K.B.; Hu, S.; Crews, C.M.; Mundy, G.R.

2003-01-01

We have found that the ubiquitin-proteasome pathway exerts exquisite control of osteoblast differentiation and bone formation in vitro and in vivo in rodents. Structurally different inhibitors that bind to specific catalytic β subunits of the 20S proteasome stimulated bone formation in bone organ cultures in concentrations as low as 10 nM. When administered systemically to mice, the proteasome inhibitors epoxomicin and proteasome inhibitor–1 increased bone volume and bone formation rates over 70% after only 5 days of treatment. Since the ubiquitin-proteasome pathway has been shown to modulate expression of the Drosophila homologue of the bone morphogenetic protein-2 and -4 (BMP-2 and BMP-4) genes, we examined the effects of noggin, an endogenous inhibitor of BMP-2 and BMP-4 on bone formation stimulated by these compounds and found that it was abrogated. These compounds increased BMP-2 but not BMP-4 or BMP-6 mRNA expression in osteoblastic cells, suggesting that BMP-2 was responsible for the observed bone formation that was inhibited by noggin. We show proteasome inhibitors regulate BMP-2 gene expression at least in part through inhibiting the proteolytic processing of Gli3 protein. Our results suggest that the ubiquitin-proteasome machinery regulates osteoblast differentiation and bone formation and that inhibition of specific components of this system may be useful therapeutically in common diseases of bone loss. PMID:12782679

TGF-β and BMP Signaling in Osteoblast Differentiation and Bone Formation

PubMed Central

Chen, Guiqian; Deng, Chuxia; Li, Yi-Ping

2012-01-01

Transforming growth factor-beta (TGF-β)/bone morphogenic protein (BMP) signaling is involved in a vast majority of cellular processes and is fundamentally important throughout life. TGF-β/BMPs have widely recognized roles in bone formation during mammalian development and exhibit versatile regulatory functions in the body. Signaling transduction by TGF-β/BMPs is specifically through both canonical Smad-dependent pathways (TGF-β/BMP ligands, receptors and Smads) and non-canonical Smad-independent signaling pathway (e.g. p38 mitogen-activated protein kinase pathway, MAPK). Following TGF-β/BMP induction, both the Smad and p38 MAPK pathways converge at the Runx2 gene to control mesenchymal precursor cell differentiation. The coordinated activity of Runx2 and TGF-β/BMP-activated Smads is critical for formation of the skeleton. Recent advances in molecular and genetic studies using gene targeting in mice enable a better understanding of TGF-β/BMP signaling in bone and in the signaling networks underlying osteoblast differentiation and bone formation. This review summarizes the recent advances in our understanding of TGF-β/BMP signaling in bone from studies of genetic mouse models and human diseases caused by the disruption of TGF-β/BMP signaling. This review also highlights the different modes of cross-talk between TGF-β/BMP signaling and the signaling pathways of MAPK, Wnt, Hedgehog, Notch, and FGF in osteoblast differentiation and bone formation. PMID:22298955

Signal mingle: Micropatterns of BMP-2 and fibronectin on soft biopolymeric films regulate myoblast shape and SMAD signaling.

PubMed

Fitzpatrick, Vincent; Fourel, Laure; Destaing, Olivier; Gilde, Flora; Albigès-Rizo, Corinne; Picart, Catherine; Boudou, Thomas

2017-01-30

In vivo, bone morphogenetic protein 2 (BMP-2) exists both in solution and bound to the extracellular matrix (ECM). While these two modes of presentation are known to influence cell behavior distinctly, their role in the niche microenvironment and their functional relevance in the genesis of a biological response has sparsely been investigated at a cellular level. Here we used the natural affinity of BMP-2 for fibronectin (FN) to engineer cell-sized micropatterns of BMP-2. This technique allowed the simultaneous control of the spatial presentation of fibronectin-bound BMP-2 and cell spreading. These micropatterns induced a specific actin and adhesion organization around the nucleus, and triggered the phosphorylation and nuclear translocation of SMAD1/5/8 in C2C12 myoblasts and mesenchymal stem cells, an early indicator of their osteoblastic trans-differentiation. We found that cell spreading itself potentiated a BMP-2-dependent phosphorylation of SMAD1/5/8. Finally, we demonstrated that FN/BMP-2-mediated early SMAD signaling depended on LIM kinase 2 and ROCK, rather than myosin II activation. Altogether, our results show that FN/BMP-2 micropatterns are a useful tool to study the mechanisms underlying BMP-2-mediated mechanotransduction. More broadly, our approach could be adapted to other combinations of ECM proteins and growth factors, opening an exciting avenue to recreate tissue-specific niches in vitro.

Alveolar ridge and maxillary sinus augmentation using rhBMP-2: a systematic review.

PubMed

Freitas, Rubens Moreno de; Spin-Neto, Rubens; Marcantonio Junior, Elcio; Pereira, Luís Antônio Violin Dias; Wikesjö, Ulf M E; Susin, Cristiano

2015-01-01

The aim of this systematic review was to evaluate clinical and safety data for recombinant human bone morphogenetic protein-2 (rhBMP-2) in an absorbable collagen sponge (ACS) carrier when used for alveolar ridge/maxillary sinus augmentation in humans. Clinical studies/case series published 1980 through June 2012 using rhBMP-2/ACS were searched. Studies meeting the following criteria were considered eligible for inclusion: >10 subjects at baseline and maxillary sinus or alveolar ridge augmentation not concomitant with implant placement. Seven of 69 publications were eligible for review. rhBMP-2/ACS yielded clinically meaningful bone formation for maxillary sinus augmentation that would allow placement of regular dental implants without consistent differences between rhBMP-2 concentrations. Nevertheless, the statistical analysis showed that sinus augmentation following autogenous bone graft was significantly greater (mean bone height: 1.6 mm, 95% CI: 0.5-2.7 mm) than for rhBMP-2/ACS (rhBMP-2 at 1.5 mg/mL). In extraction sockets, rhBMP-2/ACS maintained alveolar ridge height while enhancing alveolar ridge width. Safety reports did not represent concerns for the proposed indications. rhBMP-2/ACS appears a promising alternative to autogenous bone grafts for alveolar ridge/maxillary sinus augmentation; dose and carrier optimization may expand its efficacy, use, and clinical application. © 2013 Wiley Periodicals, Inc.

Development and characterization of a mouse floxed Bmp2 osteoblast cell line that retains osteoblast genotype and phenotype

PubMed Central

Wu, Li-an; Feng, Junsheng; Wang, Lynn; Mu, Yan-dong; Baker, Andrew; Donly, Kevin J.; Harris, Stephen E.; MacDougall, Mary; Chen, Shuo

2011-01-01

Bone morphogenetic protein 2 (Bmp2) is essential for osteoblast differentiation and osteogenesis. Generation of floxed Bmp2 osteoblast cell lines is a valuable tool for studying the effects of Bmp2 on osteoblast differentiation and its signaling pathways during skeletal metabolism. Due to relatively limited sources of primary osteoblasts, we have developed cell lines that serve as good surrogate models for the study of osteoblast cell differentiation and bone mineralization. In this study, we established and characterized immortalized mouse floxed Bmp2 osteoblast cell lines. Primary mouse floxed Bmp2 osteoblasts were transfected with pSV3-neo and clonally selected. These transfected cells were verified by PCR and immunohistochemistry. To determine the genotype and phenotype of the immortalized cells, cell morphology, proliferation, differentiation and mineralization were analyzed. Also, expression of osteoblast-related gene markers including Runx2, Osx, ATF4, Dlx3, bone sialoprotein, dentin matrix protein 1, osteonectin, osteocalcin and osteopontin were examined by quantitative RT-PCR and immunohistochemistry. These results showed that immortalized floxed Bmp2 osteoblasts had a higher proliferation rate but preserved their genotypic and phenotypic characteristics similar to the primary cells. Thus, we, for the first time, describe the development of immortalized mouse floxed Bmp2 osteoblast cell lines and present a useful model to study osteoblast biology mediated by BMP2 and its downstream signaling transduction pathways. PMID:21271257

Suppression of BMP-7 by histone deacetylase 2 promoted apoptosis of renal tubular epithelial cells in acute kidney injury

PubMed Central

Ma, Taotao; Huang, Cheng; Xu, Qingqing; Yang, Yang; Liu, Yaru; Meng, Xiaoming; Li, Jun; Ye, Min; Liang, Hong

2017-01-01

Cisplatin, a highly effective and widely used chemotherapeutic agent, has a major limitation for its nephrotoxicity. Currently, there are no therapies available to treat or prevent cisplatin nephrotoxicity. We recently identified a novel strategy for attenuating its nephrotoxicity in chemotherapy by histone deacetylase (HDAC) inhibitors via epigenetic modification to enhance bone morphogenetic protein 7 (BMP-7) expression. Cisplatin upregulated the activity of HDAC2 in the kidney. Inhibition of HDAC with clinically used trichostatin A (TSA) or valproic acid (VPA) suppressed cisplatin-induced kidney injury and epithelial cell apoptosis. Overexpression of HDAC2 promotes CP-treated tubular epithelium cells apoptosis. Chromatin immunoprecipitation assay clearly detected HDAC2 assosiation with BMP-7 promoter. Western blot and immunofluorescence results demonstrated that the expression of BMP-7 was clearly induced by TSA or VPA in vivo and in vitro. Interestingly, administration of recombinant BMP-7 (rhBMP-7) reduced cisplatin-induced kidney dysfunction. Moreover, BMP-7 treatment suppressed epithelial cell apoptosis and small interfering RNA-based knockdown of BMP-7 expression abolished HDAC inhibitors suppression of epithelial cell apoptosis in vitro. Results of current study indicated that TSA or VPA inhibited apoptosis of renal tubular epithelial cells via promoting the level of BMP-7 epigenetically through targeting HDAC2. Hence, HDAC inhibitors could be useful therapeutic agents for the prevention of cisplatin nephrotoxicity. PMID:29072686

Signal mingle: Micropatterns of BMP-2 and fibronectin on soft biopolymeric films regulate myoblast shape and SMAD signaling

NASA Astrophysics Data System (ADS)

Fitzpatrick, Vincent; Fourel, Laure; Destaing, Olivier; Gilde, Flora; Albigès-Rizo, Corinne; Picart, Catherine; Boudou, Thomas

2017-01-01

In vivo, bone morphogenetic protein 2 (BMP-2) exists both in solution and bound to the extracellular matrix (ECM). While these two modes of presentation are known to influence cell behavior distinctly, their role in the niche microenvironment and their functional relevance in the genesis of a biological response has sparsely been investigated at a cellular level. Here we used the natural affinity of BMP-2 for fibronectin (FN) to engineer cell-sized micropatterns of BMP-2. This technique allowed the simultaneous control of the spatial presentation of fibronectin-bound BMP-2 and cell spreading. These micropatterns induced a specific actin and adhesion organization around the nucleus, and triggered the phosphorylation and nuclear translocation of SMAD1/5/8 in C2C12 myoblasts and mesenchymal stem cells, an early indicator of their osteoblastic trans-differentiation. We found that cell spreading itself potentiated a BMP-2-dependent phosphorylation of SMAD1/5/8. Finally, we demonstrated that FN/BMP-2-mediated early SMAD signaling depended on LIM kinase 2 and ROCK, rather than myosin II activation. Altogether, our results show that FN/BMP-2 micropatterns are a useful tool to study the mechanisms underlying BMP-2-mediated mechanotransduction. More broadly, our approach could be adapted to other combinations of ECM proteins and growth factors, opening an exciting avenue to recreate tissue-specific niches in vitro.

The Bmp signaling pathway regulates development of left-right asymmetry in amphioxus.

PubMed

Soukup, Vladimir; Kozmik, Zbynek

2018-02-01

Establishment of asymmetry along the left-right (LR) body axis in vertebrates requires interplay between Nodal and Bmp signaling pathways. In the basal chordate amphioxus, the left-sided activity of the Nodal signaling has been attributed to the asymmetric morphogenesis of paraxial structures and pharyngeal organs, however the role of Bmp signaling in LR asymmetry establishment has not been addressed to date. Here, we show that Bmp signaling is necessary for the development of LR asymmetric morphogenesis of amphioxus larvae through regulation of Nodal signaling. Loss of Bmp signaling results in loss of the left-sided expression of Nodal, Gdf1/3, Lefty and Pitx and in gain of ectopic expression of Cerberus on the left side. As a consequence, the larvae display loss of the offset arrangement of axial structures, loss of the left-sided pharyngeal organs including the mouth, and ectopic development of the right-sided organs on the left side. Bmp inhibition thus phenocopies inhibition of Nodal signaling and results in the right isomerism. We conclude that Bmp and Nodal pathways act in concert to specify the left side and that Bmp signaling plays a fundamental role during LR development in amphioxus. Copyright © 2017 Elsevier Inc. All rights reserved.

DRAGON, a GPI-anchored membrane protein, inhibits BMP signaling in C2C12 myoblasts.

PubMed

Kanomata, Kazuhiro; Kokabu, Shoichiro; Nojima, Junya; Fukuda, Toru; Katagiri, Takenobu

2009-06-01

Bone morphogenetic proteins (BMPs) induce osteoblastic differentiation of myoblasts via binding to cell surface receptors. Repulsive guidance molecules (RGMs) have been identified as BMP co-receptors. We report here that DRAGON/RGMb, a member of the RGM family, suppressed BMP signaling in C2C12 myoblasts via a novel mechanism. All RGMs were expressed in C2C12 cells that were differentiated into myocytes and osteoblastic cells, but RGMc was not detected in immature cells. In C2C12 cells, only DRAGON suppressed ALP and Id1 promoter activities induced by BMP-4 or by constitutively activated BMP type I receptors. This inhibition by DRAGON was dependent on the secretory form of the von Willbrand factor type D domain. DRAGON even suppressed BMP signaling induced by constitutively activated Smad1. Over-expression of neogenin did not alter the inhibitory capacity of DRAGON. Taken together, these findings indicate that DRAGON may be an inhibitor of BMP signaling in C2C12 myoblasts. We also suggest that a novel molecule(s) expressed on the cell membrane may mediate the signal transduction of DRAGON in order to suppress BMP signaling in C2C12 myoblasts.

BMP Sustains Embryonic Stem Cell Self-Renewal through Distinct Functions of Different Krüppel-like Factors.

PubMed

Morikawa, Masato; Koinuma, Daizo; Mizutani, Anna; Kawasaki, Natsumi; Holmborn, Katarina; Sundqvist, Anders; Tsutsumi, Shuichi; Watabe, Tetsuro; Aburatani, Hiroyuki; Heldin, Carl-Henrik; Miyazono, Kohei

2016-01-12

Bone morphogenetic protein (BMP) signaling exerts paradoxical roles in pluripotent stem cells (PSCs); it sustains self-renewal of mouse embryonic stem cells (ESCs), while it induces differentiation in other PSCs, including human ESCs. Here, we revisit the roles of BMP-4 using mouse ESCs (mESCs) in naive and primed states. SMAD1 and SMAD5, which transduce BMP signals, recognize enhancer regions together with KLF4 and KLF5 in naive mESCs. KLF4 physically interacts with SMAD1 and suppresses its activity. Consistently, a subpopulation of cells with active BMP-SMAD can be ablated without disturbing the naive state of the culture. Moreover, Smad1/5 double-knockout mESCs stay in the naive state, indicating that the BMP-SMAD pathway is dispensable for it. In contrast, the MEK5-ERK5 pathway mediates BMP-4-induced self-renewal of mESCs by inducing Klf2, a critical factor for the ground state pluripotency. Our study illustrates that BMP exerts its self-renewing effect through distinct functions of different Krüppel-like factors. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Ecsit is required for Bmp signaling and mesoderm formation during mouse embryogenesis

PubMed Central

Xiao, Changchun; Shim, Jae-hyuck; Klüppel, Michael; Zhang, Samuel Shao-Min; Dong, Chen; Flavell, Richard A.; Fu, Xin-Yuan; Wrana, Jeffrey L.; Hogan, Brigid L.M.; Ghosh, Sankar

2003-01-01

Bone morphogenetic proteins (Bmps) are members of the transforming growth factor β (TGFβ) superfamily that play critical roles during mouse embryogenesis. Signaling by Bmp receptors is mediated mainly by Smad proteins. In this study, we show that a targeted null mutation of Ecsit, encoding a signaling intermediate of the Toll pathway, leads to reduced cell proliferation, altered epiblast patterning, impairment of mesoderm formation, and embryonic lethality at embryonic day 7.5 (E7.5), phenotypes that mimic the Bmp receptor type1a (Bmpr1a) null mutant. In addition, specific Bmp target gene expression is abolished in the absence of Ecsit. Biochemical analysis demonstrates that Ecsit associates constitutively with Smad4 and associates with Smad1 in a Bmp-inducible manner. Together with Smad1 and Smad4, Ecsit binds to the promoter of specific Bmp target genes. Finally, knock-down of Ecsit with Ecsit-specific short hairpin RNA inhibits both Bmp and Toll signaling. Therefore, these results show that Ecsit functions as an essential component in two important signal transduction pathways and establishes a novel role for Ecsit as a cofactor for Smad proteins in the Bmp signaling pathway. PMID:14633973

Comparison of Measured to Predicted Estimations of Nonpoint Source Contaminants Using Conservation Practices in an Agriculturally-Dominated Watershed in Northeast Arkansas, USA.

PubMed

Frasher, Sarah K; Woodruff, Tracy M; Bouldin, Jennifer L

2016-06-01

In efforts to reduce nonpoint source runoff and improve water quality, Best Management Practices (BMPs) were implemented in the Outlet Larkin Creek Watershed. Farmers need to make scientifically informed decisions concerning BMPs addressing contaminants from agricultural fields. The BMP Tool was developed from previous studies to estimate BMP effectiveness at reducing nonpoint source contaminants. The purpose of this study was to compare the measured percent reduction of dissolved phosphorus (DP) and total suspended solids to the reported percent reductions from the BMP Tool for validation. Similarities were measured between the BMP Tool and the measured water quality parameters. Construction of a sedimentation pond resulted in 74 %-76 % reduction in DP as compared to 80 % as predicted with the BMP Tool. However, further research is needed to validate the tool for additional water quality parameters. The BMP Tool is recommended for future BMP implementation as a useful predictor for farmers.

Serum Proteases Potentiate BMP-Induced Cell Cycle Re-entry of Dedifferentiating Muscle Cells during Newt Limb Regeneration.

PubMed

Wagner, Ines; Wang, Heng; Weissert, Philipp M; Straube, Werner L; Shevchenko, Anna; Gentzel, Marc; Brito, Goncalo; Tazaki, Akira; Oliveira, Catarina; Sugiura, Takuji; Shevchenko, Andrej; Simon, András; Drechsel, David N; Tanaka, Elly M

2017-03-27

Limb amputation in the newt induces myofibers to dedifferentiate and re-enter the cell cycle to generate proliferative myogenic precursors in the regeneration blastema. Here we show that bone morphogenetic proteins (BMPs) and mature BMPs that have been further cleaved by serum proteases induce cell cycle entry by dedifferentiating newt muscle cells. Protease-activated BMP4/7 heterodimers that are present in serum strongly induced myotube cell cycle re-entry with protease cleavage yielding a 30-fold potency increase of BMP4/7 compared with canonical BMP4/7. Inhibition of BMP signaling via muscle-specific dominant-negative receptor expression reduced cell cycle entry in vitro and in vivo. In vivo inhibition of serine protease activity depressed cell cycle re-entry, which in turn was rescued by cleaved-mimic BMP. This work identifies a mechanism of BMP activation that generates blastema cells from differentiated muscle. Copyright © 2017 Elsevier Inc. All rights reserved.

Bone morphogenetic protein-2 and bone therapy: successes and pitfalls.

PubMed

Poon, Bonnie; Kha, Tram; Tran, Sally; Dass, Crispin R

2016-02-01

Bone morphogenetic proteins (BMPs), more specifically BMP-2, are being increasingly used in orthopaedic surgery due to advanced research into osteoinductive factors that may enhance and improve bone therapy. There are many areas in therapy that BMP-2 is being applied to, including dental treatment, open tibial fractures, cancer and spinal surgery. Within these areas of treatment, there are many reports of successes and pitfalls. This review explores the use of BMP-2 and its successes, pitfalls and future prospects in bone therapy. The PubMed database was consulted to compile this review. With successes in therapy, there were descriptions of a more rapid healing time with no signs of rejection or infection attributed to BMP-2 treatment. Pitfalls included BMP-2 'off-label' use, which lead to various adverse effects. Our search highlighted that optimising treatment with BMP-2 is a direction that many researchers are exploring, with areas of current research interest including concentration and dose of BMP-2, carrier type and delivery. © 2015 Royal Pharmaceutical Society.

SMOC can act as both an antagonist and an expander of BMP signaling.

PubMed

Thomas, J Terrig; Eric Dollins, D; Andrykovich, Kristin R; Chu, Tehyen; Stultz, Brian G; Hursh, Deborah A; Moos, Malcolm

2017-03-21

The matricellular protein SMOC (Secreted Modular Calcium binding protein) is conserved phylogenetically from vertebrates to arthropods. We showed previously that SMOC inhibits bone morphogenetic protein (BMP) signaling downstream of its receptor via activation of mitogen-activated protein kinase (MAPK) signaling. In contrast, the most prominent effect of the Drosophila orthologue, pentagone ( pent ), is expanding the range of BMP signaling during wing patterning. Using SMOC deletion constructs we found that SMOC-∆EC, lacking the extracellular calcium binding (EC) domain, inhibited BMP2 signaling, whereas SMOC-EC (EC domain only) enhanced BMP2 signaling. The SMOC-EC domain bound HSPGs with a similar affinity to BMP2 and could expand the range of BMP signaling in an in vitro assay by competition for HSPG-binding. Together with data from studies in vivo we propose a model to explain how these two activities contribute to the function of Pent in Drosophila wing development and SMOC in mammalian joint formation.

Bone regeneration by polyhedral microcrystals from silkworm virus

PubMed Central

Matsumoto, Goichi; Ueda, Takayo; Shimoyama, Junko; Ijiri, Hiroshi; Omi, Yasushi; Yube, Hisato; Sugita, Yoshihiko; Kubo, Katsutoshi; Maeda, Hatsuhiko; Kinoshita, Yukihiko; Arias, Duverney Gaviria; Shimabukuro, Junji; Kotani, Eiji; Kawamata, Shin; Mori, Hajime

2012-01-01

Bombyx mori cypovirus is a major pathogen which causes significant losses in silkworm cocoon harvests because the virus particles are embedded in micrometer-sized protein crystals called polyhedra and can remain infectious in harsh environmental conditions for years. But the remarkable stability of polyhedra can be applied on slow-release carriers of cytokines for tissue engineering. Here we show the complete healing in critical-sized bone defects by bone morphogenetic protein-2 (BMP-2) encapsulated polyhedra. Although absorbable collagen sponge (ACS) safely and effectively delivers recombinant human BMP-2 (rhBMP-2) into healing tissue, the current therapeutic regimens release rhBMP-2 at an initially high rate after which the rate declines rapidly. ACS impregnated with BMP-2 polyhedra had enough osteogenic activity to promote complete healing in critical-sized bone defects, but ACS with a high dose of rhBMP-2 showed incomplete bone healing, indicating that polyhedral microcrystals containing BMP-2 promise to advance the state of the art of bone healing. PMID:23226833

Bone regeneration by implantation of adipose-derived stromal cells expressing BMP-2

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li Huiwu; Health and Science Center, SIBS CAS and SSMU, 225 South Chongqing Road, Shanghai 200025; Dai Kerong

2007-05-18

In this study, we reported that the adipose-derived stromal cells (ADSCs) genetically modified by bone morphogenetic protein 2 (BMP-2) healed critical-sized canine ulnar bone defects. First, the osteogenic and adipogenic differentiation potential of the ADSCs derived from canine adipose tissue were demonstrated. And then the cells were modified by the BMP-2 gene and the expression and bone-induction ability of BMP-2 were identified. Finally, the cells modified by BMP-2 gene were applied to a {beta}-tricalcium phosphate (TCP) carrier and implanted into ulnar bone defects in the canine model. After 16 weeks, radiographic, histological, and histomorphometry analysis showed that ADSCs modified bymore » BMP-2 gene produced a significant increase of newly formed bone area and healed or partly healed all of the bone defects. We conclude that ADSCs modified by the BMP-2 gene can enhance the repair of critical-sized bone defects in large animals.« less

BMP signalling in human fetal ovary somatic cells is modulated in a gene-specific fashion by GREM1 and GREM2

PubMed Central

Bayne, Rosemary A.; Donnachie, Douglas J.; Kinnell, Hazel L.; Childs, Andrew J.; Anderson, Richard A.

2016-01-01

STUDY QUESTION Do changes in the expression of bone morphogenetic proteins (BMPs) 2 and 4, and their antagonists Gremlin1 (GREM1) and Gremlin2 (GREM2) during human fetal ovarian development impact on BMP pathway activity and lead to changes in gene expression that may influence the fate and/or function of ovarian somatic cells? STUDY FINDING BMPs 2 and 4 differentially regulate gene expression in cultured human fetal ovarian somatic cells. Expression of some, but not all BMP target genes is antagonised by GREM1 and GREM2, indicating the existence of a mechanism to fine-tune BMP signal intensity in the ovary. Leucine-rich repeat-containing G-protein coupled receptor 5 (LGR5), a marker of immature ovarian somatic cells, is identified as a novel transcriptional target of BMP4. WHAT IS KNOWN ALREADY Extensive re-organisation of the germ and somatic cell populations in the feto-neonatal ovary culminates in the formation of primordial follicles, which provide the basis for a female's future fertility. BMP growth factors play important roles at many stages of ovarian development and function. GREM1, an extracellular antagonist of BMP signalling, regulates the timing of primordial follicle formation in the mouse ovary, and mRNA levels of BMP4 decrease while those of BMP2 increase prior to follicle formation in the human fetal ovary. STUDY DESIGN, SAMPLES/MATERIALS, METHODS Expression of genes encoding BMP pathway components, BMP antagonists and markers of ovarian somatic cells were determined by quantitative (q)RT-PCR in human fetal ovaries (from 8 to 21 weeks gestation) and fetal ovary-derived somatic cell cultures. Ovarian expression of GREM1 protein was confirmed by immunoblotting. Primary human fetal ovarian somatic cell cultures were derived from disaggregated ovaries by differential adhesion and cultured in the presence of recombinant human BMP2 or BMP4, with or without the addition of GREM1 or GREM2. MAIN RESULTS AND THE ROLE OF CHANCE We demonstrate that the expression of BMP antagonists GREM1, GREM2 and CHRD  increases in the lead-up to primordial follicle formation in the human fetal ovary, and that the BMP pathway is active in cultured ovarian somatic cells. This leads to differential changes in the expression of a number of genes, some of which are further modulated by GREM1 and/or GREM2. The positive transcriptional regulation of LGR5 (a marker of less differentiated somatic cells) by BMP4 in vitro suggests that increasing levels of GREM1 and reduced levels of BMP4 as the ovary develops in vivo may act to reduce LGR5 levels and allow pre-granulosa cell differentiation. LIMITATIONS, REASONS FOR CAUTION While we have demonstrated that markers of different somatic cell types are expressed in the cultured ovarian somatic cells, their proportions may not represent the same cells in the intact ovary which also contains germ cells. WIDER IMPLICATIONS OF THE FINDINGS This study extends previous work identifying germ cells as targets of ovarian BMP signalling, and suggests BMPs may regulate the development of both germ and somatic cells in the developing ovary around the time of follicle formation. LARGE SCALE DATA Not applicable. STUDY FUNDING/COMPETING INTERESTS This work was supported by The UK Medical Research Council (Grant No.: G1100357 to RAA), and Medical Research Scotland (Grant No. 345FRG to AJC). The authors have no competing interests to declare. PMID:27385727

BMP signalling in human fetal ovary somatic cells is modulated in a gene-specific fashion by GREM1 and GREM2.

PubMed

Bayne, Rosemary A; Donnachie, Douglas J; Kinnell, Hazel L; Childs, Andrew J; Anderson, Richard A

2016-09-01

Do changes in the expression of bone morphogenetic proteins (BMPs) 2 and 4, and their antagonists Gremlin1 (GREM1) and Gremlin2 (GREM2) during human fetal ovarian development impact on BMP pathway activity and lead to changes in gene expression that may influence the fate and/or function of ovarian somatic cells? BMPs 2 and 4 differentially regulate gene expression in cultured human fetal ovarian somatic cells. Expression of some, but not all BMP target genes is antagonised by GREM1 and GREM2, indicating the existence of a mechanism to fine-tune BMP signal intensity in the ovary. Leucine-rich repeat-containing G-protein coupled receptor 5 (LGR5), a marker of immature ovarian somatic cells, is identified as a novel transcriptional target of BMP4. Extensive re-organisation of the germ and somatic cell populations in the feto-neonatal ovary culminates in the formation of primordial follicles, which provide the basis for a female's future fertility. BMP growth factors play important roles at many stages of ovarian development and function. GREM1, an extracellular antagonist of BMP signalling, regulates the timing of primordial follicle formation in the mouse ovary, and mRNA levels of BMP4 decrease while those of BMP2 increase prior to follicle formation in the human fetal ovary. Expression of genes encoding BMP pathway components, BMP antagonists and markers of ovarian somatic cells were determined by quantitative (q)RT-PCR in human fetal ovaries (from 8 to 21 weeks gestation) and fetal ovary-derived somatic cell cultures. Ovarian expression of GREM1 protein was confirmed by immunoblotting. Primary human fetal ovarian somatic cell cultures were derived from disaggregated ovaries by differential adhesion and cultured in the presence of recombinant human BMP2 or BMP4, with or without the addition of GREM1 or GREM2. We demonstrate that the expression of BMP antagonists GREM1, GREM2 and CHRD  increases in the lead-up to primordial follicle formation in the human fetal ovary, and that the BMP pathway is active in cultured ovarian somatic cells. This leads to differential changes in the expression of a number of genes, some of which are further modulated by GREM1 and/or GREM2. The positive transcriptional regulation of LGR5 (a marker of less differentiated somatic cells) by BMP4 in vitro suggests that increasing levels of GREM1 and reduced levels of BMP4 as the ovary develops in vivo may act to reduce LGR5 levels and allow pre-granulosa cell differentiation. While we have demonstrated that markers of different somatic cell types are expressed in the cultured ovarian somatic cells, their proportions may not represent the same cells in the intact ovary which also contains germ cells. This study extends previous work identifying germ cells as targets of ovarian BMP signalling, and suggests BMPs may regulate the development of both germ and somatic cells in the developing ovary around the time of follicle formation. Not applicable. This work was supported by The UK Medical Research Council (Grant No.: G1100357 to RAA), and Medical Research Scotland (Grant No. 345FRG to AJC). The authors have no competing interests to declare. © The Author 2016. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology.

Microcontact printing of BMP-2 and its effect on human chondrocytes behavior

NASA Astrophysics Data System (ADS)

Pan, Chang-Jiang; Nie, Yu-Dong

2010-01-01

The present study is to investigate human chondrocytes behavior on microcontact printed bone morphogenetic protein-2 (BMP-2) lines on polystyrene (PS) surface. It was found that the cells aligned with BMP lines and expressed type II and VI collagen. The chondrocytes in vitro cultured on BMP lines were elongated, which resulted in altered cell morphology. Taking all these results into consideration, BMP-2 lines enhance cell adhesion, restrict spreading, and increase type II and VI collagen expression. The results represented in this study may be an approach to the problem of engineering reparative cartilage in vitro.

Smad4 mediated BMP2 signal is essential for the regulation of GATA4 and Nkx2.5 by affecting the histone H3 acetylation in H9c2 cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Si, Lina; Shi, Jin; Gao, Wenqun

2014-07-18

Highlights: • BMP2 can upregulated cardiac related gene GATA4, Nkx2.5, MEF2c and Tbx5. • Inhibition of Smad4 decreased BMP2-induced hyperacetylation of histone H3. • Inhibition of Smad4 diminished BMP2-induced overexpression of GATA4 and Nkx2.5. • Inhibition of Smad4 decreased hyperacetylated H3 in the promoter of GATA4 and Nkx2.5. • Smad4 is essential for BMP2 induced hyperacetylated histone H3. - Abstract: BMP2 signaling pathway plays critical roles during heart development, Smad4 encodes the only common Smad protein in mammals, which is a pivotal nuclear mediator. Our previous studies showed that BMP2 enhanced the expression of cardiac transcription factors in part bymore » increasing histone H3 acetylation. In the present study, we tested the hypothesis that Smad4 mediated BMP2 signaling pathway is essential for the expression of cardiac core transcription factors by affecting the histone H3 acetylation. We successfully constructed a lentivirus-mediated short hairpin RNA interference vector targeting Smad4 (Lv-Smad4) in rat H9c2 embryonic cardiac myocytes (H9c2 cells) and demonstrated that it suppressed the expression of the Smad4 gene. Cultured H9c2 cells were transfected with recombinant adenoviruses expressing human BMP2 (AdBMP2) with or without Lv-Smad4. Quantitative real-time RT-PCR analysis showed that knocking down of Smad4 substantially inhibited both AdBMP2-induced and basal expression levels of cardiac transcription factors GATA4 and Nkx2.5, but not MEF2c and Tbx5. Similarly, chromatin immunoprecipitation (ChIP) analysis showed that knocking down of Smad4 inhibited both AdBMP2-induced and basal histone H3 acetylation levels in the promoter regions of GATA4 and Nkx2.5, but not of Tbx5 and MEF2c. In addition, Lv-Smad4 selectively suppressed AdBMP2-induced expression of HAT p300, but not of HAT GCN5 in H9c2 cells. The data indicated that inhibition of Smad4 diminished both AdBMP2 induced and basal histone acetylation levels in the promoter regions of GATA4 and Nkx2.5, suggesting that Smad4 mediated BMP2 signaling pathway was essential for the regulation of GATA4 and Nkx2.5 by affecting the histone H3 acetylation in H9c2 cells.« less

BMP2-Induced Inflammation Can Be Suppressed by the Osteoinductive Growth Factor NELL-1

PubMed Central

Shen, Jia; James, Aaron W.; Zara, Janette N.; Asatrian, Greg; Khadarian, Kevork; Zhang, James B.; Ho, Stephanie; Kim, Hyun Ju

2013-01-01

Bone-morphogenetic protein 2 (BMP2) is currently the only Food and Drug Administration-approved osteoinductive growth factor used in clinical settings for bone regeneration and repair. However, the use of BMP2 is encumbered by numerous clinical complications, including postoperative inflammation and life-threatening cervical swelling. Thus, methods to prevent BMP2-induced inflammation would have far-reaching clinical implications toward improving current BMP2-based methods for bone regeneration. For the first time, we investigate the potential role of the growth factor Nel-like molecule-1 (NELL-1) in inhibiting BMP2-induced inflammation. Adult rats underwent a femoral bone onlay procedure, treated with either BMP2 protein (4 mg/mL), NELL-1 protein (4 mg/mL), or both proteins combined. Animals were evaluated at 3, 7, and 14 days postoperatively by histology, histomorphometry, immunohistochemistry, and real-time PCR for markers of inflammation (TNFα, IL6). The relative levels of TNFα and IL6 in serum were also detected by ELISA. The mechanism for NELL-1's anti-inflammatory effect was further assessed through examining inflammatory markers and generation of reactive oxygen species (ROS) in the mouse embryonic fibroblast NIH3T3 cells. BMP2 significantly induced local inflammation, including an early and pronounced polymorphonuclear cell infiltration accompanied by increased expression of TNFα and IL6. Treatment with NELL-1 alone elicited no significant inflammatory response. However, NELL-1 significantly attenuated BMP2-induced inflammation by all markers and at all timepoints. These local findings were also confirmed using systemic serum inflammatory biomarkers (TNFα, IL6). In each case, NELL-1 fully reversed BMP2-induced systemic inflammation. Lastly, our findings were recapitulated in vitro, where NELL-1 suppressed BMP2 induced expression of inflammatory markers, as well as NF-κB transcriptional activity and generation of ROS. BMP2-induced inflammation is a serious public health concern with potentially life-threatening complications. In the present study, we observed that the growth factor, NELL-1, significantly attenuates or completely reverses BMP2-induced inflammation. The mechanisms of NELL-1's anti-inflammatory effect are only partially elucidated, and may include reduction of NF-κB transcriptional activity or ROS generation. PMID:23758588

Using Arc/Info GIS to help implement the National Pollutant Discharge Elimination System (NPDES) stormwater permit for Los Angeles County

DOE Office of Scientific and Technical Information (OSTI.GOV)

Levine, D.A.; Pace, P.J.; Woods, J.A.

1997-06-01

One of Los Angeles County Department of Public Works` many responsibilities is to manage non-point pollution that enters the storm drain network within Los Angeles County. The management of this non-point source pollution is mandated by the NPDES guidelines under the Federal Clean Water Act. These guidelines require the County to monitor the drainage network and the storm water and urban runoff flowing through it. The County covers over 3,117 square miles, with the NPDES Permit covering over 3,100 square miles and over 2500 miles of storm drains. A proposed solution to monitor and manage this vast geographic area ismore » centered upon an Arc/Info GIS. Some of the many concerns which need to be addressed include the administration and evaluation of Best Management Practices (BMP`s), storm drain inspection for illegal connections and illicit discharges, and pollutant load assessment and modeling. The storm drain network and other coverages will be related to external data bases currently used for facility management and planning. This system would be used for query purposes to perform spatial modeling and {open_quotes}what if{close_quotes} scenarios needed to create maps and reports required by the permit and to evaluate various BMP implementation strategies.« less

Dorsoventral patterning in hemichordates: insights into early chordate evolution.

PubMed

Lowe, Christopher J; Terasaki, Mark; Wu, Michael; Freeman, Robert M; Runft, Linda; Kwan, Kristen; Haigo, Saori; Aronowicz, Jochanan; Lander, Eric; Gruber, Chris; Smith, Mark; Kirschner, Marc; Gerhart, John

2006-09-01

We have compared the dorsoventral development of hemichordates and chordates to deduce the organization of their common ancestor, and hence to identify the evolutionary modifications of the chordate body axis after the lineages split. In the hemichordate embryo, genes encoding bone morphogenetic proteins (Bmp) 2/4 and 5/8, as well as several genes for modulators of Bmp activity, are expressed in a thin stripe of ectoderm on one midline, historically called "dorsal." On the opposite midline, the genes encoding Chordin and Anti-dorsalizing morphogenetic protein (Admp) are expressed. Thus, we find a Bmp-Chordin developmental axis preceding and underlying the anatomical dorsoventral axis of hemichordates, adding to the evidence from Drosophila and chordates that this axis may be at least as ancient as the first bilateral animals. Numerous genes encoding transcription factors and signaling ligands are expressed in the three germ layers of hemichordate embryos in distinct dorsoventral domains, such as pox neuro, pituitary homeobox, distalless, and tbx2/3 on the Bmp side and netrin, mnx, mox, and single-minded on the Chordin-Admp side. When we expose the embryo to excess Bmp protein, or when we deplete endogenous Bmp by small interfering RNA injections, these expression domains expand or contract, reflecting their activation or repression by Bmp, and the embryos develop as dorsalized or ventralized limit forms. Dorsoventral patterning is independent of anterior/posterior patterning, as in Drosophila but not chordates. Unlike both chordates and Drosophila, neural gene expression in hemichordates is not repressed by high Bmp levels, consistent with their development of a diffuse rather than centralized nervous system. We suggest that the common ancestor of hemichordates and chordates did not use its Bmp-Chordin axis to segregate epidermal and neural ectoderm but to pattern many other dorsoventral aspects of the germ layers, including neural cell fates within a diffuse nervous system. Accordingly, centralization was added in the chordate line by neural-epidermal segregation, mediated by the pre-existing Bmp-Chordin axis. Finally, since hemichordates develop the mouth on the non-Bmp side, like arthropods but opposite to chordates, the mouth and Bmp-Chordin axis may have rearranged in the chordate line, one relative to the other.

Dorsoventral Patterning in Hemichordates: Insights into Early Chordate Evolution

PubMed Central

Lowe, Christopher J; Terasaki, Mark; Wu, Michael; Freeman, Robert M; Runft, Linda; Kwan, Kristen; Haigo, Saori; Aronowicz, Jochanan; Lander, Eric; Gruber, Chris; Smith, Mark; Kirschner, Marc; Gerhart, John

2006-01-01

We have compared the dorsoventral development of hemichordates and chordates to deduce the organization of their common ancestor, and hence to identify the evolutionary modifications of the chordate body axis after the lineages split. In the hemichordate embryo, genes encoding bone morphogenetic proteins (Bmp) 2/4 and 5/8, as well as several genes for modulators of Bmp activity, are expressed in a thin stripe of ectoderm on one midline, historically called “dorsal.” On the opposite midline, the genes encoding Chordin and Anti-dorsalizing morphogenetic protein (Admp) are expressed. Thus, we find a Bmp-Chordin developmental axis preceding and underlying the anatomical dorsoventral axis of hemichordates, adding to the evidence from Drosophila and chordates that this axis may be at least as ancient as the first bilateral animals. Numerous genes encoding transcription factors and signaling ligands are expressed in the three germ layers of hemichordate embryos in distinct dorsoventral domains, such as pox neuro, pituitary homeobox, distalless, and tbx2/3 on the Bmp side and netrin, mnx, mox, and single-minded on the Chordin-Admp side. When we expose the embryo to excess Bmp protein, or when we deplete endogenous Bmp by small interfering RNA injections, these expression domains expand or contract, reflecting their activation or repression by Bmp, and the embryos develop as dorsalized or ventralized limit forms. Dorsoventral patterning is independent of anterior/posterior patterning, as in Drosophila but not chordates. Unlike both chordates and Drosophila, neural gene expression in hemichordates is not repressed by high Bmp levels, consistent with their development of a diffuse rather than centralized nervous system. We suggest that the common ancestor of hemichordates and chordates did not use its Bmp-Chordin axis to segregate epidermal and neural ectoderm but to pattern many other dorsoventral aspects of the germ layers, including neural cell fates within a diffuse nervous system. Accordingly, centralization was added in the chordate line by neural-epidermal segregation, mediated by the pre-existing Bmp-Chordin axis. Finally, since hemichordates develop the mouth on the non-Bmp side, like arthropods but opposite to chordates, the mouth and Bmp-Chordin axis may have rearranged in the chordate line, one relative to the other. PMID:16933975

Differing impact of the deletion of hemochromatosis-associated molecules HFE and transferrin receptor-2 on the iron phenotype of mice lacking bone morphogenetic protein 6 or hemojuvelin.

PubMed

Latour, Chloé; Besson-Fournier, Céline; Meynard, Delphine; Silvestri, Laura; Gourbeyre, Ophélie; Aguilar-Martinez, Patricia; Schmidt, Paul J; Fleming, Mark D; Roth, Marie-Paule; Coppin, Hélène

2016-01-01

Hereditary hemochromatosis, which is characterized by inappropriately low levels of hepcidin, increased dietary iron uptake, and systemic iron accumulation, has been associated with mutations in the HFE, transferrin receptor-2 (TfR2), and hemojuvelin (HJV) genes. However, it is still not clear whether these molecules intersect in vivo with bone morphogenetic protein 6 (BMP6)/mothers against decapentaplegic (SMAD) homolog signaling, the main pathway up-regulating hepcidin expression in response to elevated hepatic iron. To answer this question, we produced double knockout mice for Bmp6 and β2-microglobulin (a surrogate for the loss of Hfe) and for Bmp6 and Tfr2, and we compared their phenotype (hepcidin expression, Bmp/Smad signaling, hepatic and extrahepatic tissue iron accumulation) with that of single Bmp6-deficient mice and that of mice deficient for Hjv, alone or in combination with Hfe or Tfr2. Whereas the phenotype of Hjv-deficient females was not affected by loss of Hfe or Tfr2, that of Bmp6-deficient females was considerably worsened, with decreased Smad5 phosphorylation, compared with single Bmp6-deficient mice, further repression of hepcidin gene expression, undetectable serum hepcidin, and massive iron accumulation not only in the liver but also in the pancreas, the heart, and the kidneys. These results show that (1) BMP6 does not require HJV to transduce signal to hepcidin in response to intracellular iron, even if the loss of HJV partly reduces this signal, (2) another BMP ligand can replace BMP6 and significantly induce hepcidin expression in response to extracellular iron, and (3) BMP6 alone is as efficient at inducing hepcidin as the other BMPs in association with the HJV/HFE/TfR2 complex; they provide an explanation for the compensatory effect of BMP6 treatment on the molecular defect underlying Hfe hemochromatosis in mice. © 2015 by the American Association for the Study of Liver Diseases.

Regulation of bone morphogenetic protein signalling and cranial osteogenesis by Gpc1 and Gpc3.

PubMed

Dwivedi, Prem P; Grose, Randall H; Filmus, Jorge; Hii, Charles S T; Xian, Cory J; Anderson, Peter J; Powell, Barry C

2013-08-01

From birth, the vault of the skull grows at a prodigious rate, driven by the activity of osteoblastic cells at the fibrous joints (sutures) that separate the bony calvarial plates. One in 2500 children is born with a medical condition known as craniosynostosis because of premature bony fusion of the calvarial plates and a cessation of bone growth at the sutures. Bone morphogenetic proteins (BMPs) are potent growth factors that promote bone formation. Previously, we found that Glypican-1 (GPC1) and Glypican-3 (GPC3) are expressed in cranial sutures and are decreased during premature suture fusion in children. Although glypicans are known to regulate BMP signalling, a mechanistic link between GPC1, GPC3 and BMPs and osteogenesis has not yet been investigated. We now report that human primary suture mesenchymal cells coexpress GPC1 and GPC3 on the cell surface and release them into the media. We show that they inhibit BMP2, BMP4 and BMP7 activities, which both physically interact with BMP2 and that immunoblockade of endogenous GPC1 and GPC3 potentiates BMP2 activity. In contrast, increased levels of GPC1 and GPC3 as a result of overexpression or the addition of recombinant protein, inhibit BMP2 signalling and BMP2-mediated osteogenesis. We demonstrate that BMP signalling in suture mesenchymal cells is mediated by both SMAD-dependent and SMAD-independent pathways and that GPC1 and GPC3 inhibit both pathways. GPC3 inhibition of BMP2 activity is independent of attachment of the glypican on the cell surface and post-translational glycanation, and thus appears to be mediated by the core glypican protein. The discovery that GPC1 and GPC3 regulate BMP2-mediated osteogenesis, and that inhibition of endogenous GPC1 and GPC3 potentiates BMP2 responsiveness of human suture mesenchymal cells, indicates how downregulation of glypican expression could lead to the bony suture fusion that characterizes craniosynostosis. Copyright © 2013 Elsevier Inc. All rights reserved.

RhBMP-2-induced radiculitis in patients undergoing transforaminal lumbar interbody fusion: relationship to dose.

PubMed

Villavicencio, Alan T; Burneikiene, Sigita

2016-10-01

Recombinant human bone morphogenetic protein-2 (rhBMP-2) remains the primary synthetic osteoinductive material used in spinal fusion surgery today. The early inflammation reaction to rhBMP-2 manifesting with radicular symptoms has been previously reported in patients undergoing transforaminal lumbar interbody fusion (TLIF). There is a disagreement with regard to the factors affecting its occurrence and whether such symptoms are dose dependent. The purpose of this analysis was to determine the incidence of rhBMP-2-induced radiculitis and its relationship to dose. A retrospective cohort analysis was performed of the prospectively collected data. All consecutive patients (n=204) who underwent one- or two-level TLIF and instrumented posterolateral fusion with an off-label rhBMP-2 use were included in this analysis. The patients who developed new radicular symptoms after initial improvement postoperatively and had sterile fluid collections indicative of inflammatory process, or in the absence of any structural abnormalities that would explain these symptoms on imaging studies, were deemed to have rhBMP-2-induced radiculitis. Magnetic resonance imaging (MRI) scans were obtained for all patients who developed postoperative radicular symptoms. Correlations between the total rhBMP-2 dose, dose per spinal level, and incidence of radiculitis were evaluated while controlling for age, sex, number of TLIF levels, and surgeon. The incidence of postoperative radiculitis was 11.3% (23 out of 204). The average total rhBMP-2 dose was 4.9 mg (range=2.1-12) and the average dose per spinal level was 3.8 mg (range=1.05-12). Logistic regression analysis did not identify any significant correlations between the rhBMP-2 doses and the incidence of radiculitis (p=.6). The incidence of rhBMP-2-induced radiculitis in patients undergoing TLIF is quite high, but there were no dose-related correlations found. The study, however, cannot rule out a possibility that a larger variation in bone morphogenetic protein (BMP) doses could still be a factor in the development of rhBMP-2-associated radiculitis. Copyright © 2016 Elsevier Inc. All rights reserved.

Trends Analysis of rhBMP Utilization in Single-Level Posterior Lumbar Interbody Fusion in the United States

PubMed Central

Lao, Lifeng; Cohen, Jeremiah R.; Brodke, Darrel S.; Youssef, Jim A.; Park, Jong-Beom; Yoon, S. Tim; Wang, Jeffrey C.; Meisel, Hans-Joerg

2017-01-01

Study Design: Retrospective study. Objectives: Recombinant human bone morphogenetic protein-2 (rhBMP-2) has been widely used in spinal fusion surgery, but there is little information on rhBMP-2 utilization in single-level posterior lumbar interbody fusion (PLIF). The purpose of our study was to evaluate the trends and demographics of rhBMP-2 utilization in single-level PLIF. Methods: Patients who underwent single-level PLIF from 2005 to 2011 were identified by searching ICD-9 diagnosis and procedure codes in the PearlDiver Patient Records Database, a national database of orthopedic insurance records. The year of procedure, age, gender, and region of the United States were recorded for each patient. Results were reported for each variable as the incidence of procedures identified per 100 000 patients searched in the database. Results: A total of 2735 patients had single-level PLIF. The average rate of single-level PLIF with rhBMP-2 maintained at a relatively stable level (28% to 31%) from 2005 to 2009, but decreased in 2010 (9.9%) and 2011 (11.8%). The overall incidence of single-level PLIF without rhBMP-2 (0.68 cases per 100 000 patients) was statistically higher (P < .01) compared to single-level PLIF with rhBMP-2 (0.21 cases per 100 000 patients). The average rate of single-level PLIF with rhBMP-2 utilization was the highest in West (30.1%), followed by Midwest (26.9%), South (20.5%), and Northeast (17.8%). The highest incidence of single-level PLIF with rhBMP-2 was observed in the age group <65 years (0.3 per 100 000 patients). Conclusions: To our knowledge, this is the first study to report on the demographics associated with rhBMP-2 use in single-level PLIF. There was a 3-fold increase in the rate of PLIF without rhBMP-2 compared to PLIF with rhBMP-2, with both procedures being mainly done in patients less than 65 years of age. PMID:28989840

Complications Related to the Recombinant Human Bone Morphogenetic Protein 2 Use in Posterior Cervical Fusion.

PubMed

Takahashi, Shinji; Buser, Zorica; Cohen, Jeremiah R; Roe, Allison; Myhre, Sue L; Meisel, Hans-Joerg; Brodke, Darrel S; Yoon, S Tim; Park, Jong-Beom; Wang, Jeffrey C; Youssef, Jim A

2017-11-01

A retrospective cohort study. To compare the complications between posterior cervical fusions with and without recombinant human bone morphogenetic protein 2 (rhBMP2). Use of rhBMP2 in anterior cervical spinal fusion procedures can lead to potential complications such as neck edema, resulting in airway complications or neurological compression. However, there are no data on the complications associated with the "off-label" use of rhBMP2 in upper and lower posterior cervical fusion approaches. Patients from the PearlDiver database who had a posterior cervical fusion between 2005 and 2011 were identified. We evaluated complications within 90 days after fusion and data was divided in 2 groups: (1) posterior cervical fusion including upper cervical spine O-C2 (upper group) and (2) posterior cervical fusion including lower cervical spine C3-C7 (lower group). Complications were divided into: any complication, neck-related complications, wound-related complications, and other complications. Of the 352 patients in the upper group, 73 patients (20.7%) received rhBMP2, and 279 patients (79.3%) did not. Likewise, in the lower group of 2372 patients, 378 patients (15.9%) had surgery with rhBMP2 and 1994 patients (84.1%) without. In the upper group, complications were observed in 7 patients (9.6%) with and 34 patients (12%) without rhBMP2. In the lower group, complications were observed in 42 patients (11%) with and 276 patients (14%) without rhBMP2. Furthermore, in the lower group the wound-related complications were significantly higher in the rhBMP2 group (23 patients, 6.1%) compared with the non-rhBMP2 group (75 patients, 3.8%). Our data showed that the use of rhBMP2 does not increase the risk of complications in upper cervical spine fusion procedures. However, in the lower cervical spine, rhBMP2 may elevate the risk of wound-related complications. Overall, there were no major complications associated with the use of rhBMP2 for posterior cervical fusion approaches. Level III.

Gelatin- hydroxyapatite- calcium sulphate based biomaterial for long term sustained delivery of bone morphogenic protein-2 and zoledronic acid for increased bone formation: In-vitro and in-vivo carrier properties.

PubMed

Raina, Deepak Bushan; Larsson, David; Mrkonjic, Filip; Isaksson, Hanna; Kumar, Ashok; Lidgren, Lars; Tägil, Magnus

2018-02-28

In this study, a novel macroporous composite biomaterial consisting of gelatin-hydroxyapatite-calcium sulphate for delivery of bone morphogenic protein-2 (rhBMP-2) and zoledronic acid (ZA) has been developed. The biomaterial scaffold has a porous structure and functionalization of the scaffold with rhBMP-2 induces osteogenic differentiation of MC3T3-e1 cells seen by a significant increase in biochemical and genetic markers of osteoblastic differentiation. In-vivo muscle pouch experiments showed higher mineralization using scaffold+rhBMP-2 when compared to an approved absorbable collagen sponge (ACS)+rhBMP-2 as verified by micro-CT. Co-delivery of rhBMP-2+ZA via the novel scaffold enabled a reduction in the effective rhBMP-2 doses. The presence of tartrate resistant acid phosphatase staining in the rhBMP-2 group indicates osteoclastic resorption, which could be stalled by adding ZA, which by speculation could explain the net increase in mineralization. The new scaffold allowed for slow release of rhBMP-2 in-vitro (3.3±0.1%) after 4weeks. Using single photon emission computed tomography (SPECT), the release kinetics of 125 I-rhBMP-2 in-vivo was followed for 4weeks and a total of 65.3±15.2% 125 I-rhBMP-2 was released from the scaffolds. In-vitro 14 C-ZA release curve shows an initial burst release on day 1 (8.8±0.7%) followed by a slow release during the following 4weeks (13±0.1%). In-vivo, an initial release of 43.2±7.6% of 14 C-ZA was detected after 1day, after which the scaffold retained the remaining ZA during 4-weeks. Taken together, our results show that the developed biomaterial is an efficient carrier for spatio-temporal delivery of rhBMP-2 and ZA leading to increased bone formation compared to commercially available carrier for rhBMP-2. Copyright © 2018 Elsevier B.V. All rights reserved.

Runx2 mediates epigenetic silencing of the bone morphogenetic protein-3B (BMP-3B/GDF10) in lung cancer cells

PubMed Central

2012-01-01

Background The Runt-related transcription factor Runx2 is essential for bone development but is also implicated in progression of several cancers of breast, prostate and bone, where it activates cancer-related genes and promotes invasive properties. The transforming growth factor β (TGF-β) family member bone morphogenetic protein-3B (BMP-3B/GDF10) is regarded as a tumor growth inhibitor and a gene silenced in lung cancers; however the regulatory mechanisms leading to its silencing have not been identified. Results Here we show that Runx2 is highly expressed in lung cancer cells and downregulates BMP-3B. This inverse relationship between Runx2 and BMP-3B expression is further supported by increased expression of BMP-3B in mesenchymal cells from Runx2 deficient mice. The ectopic expression of Runx2, but not DNA binding mutant Runx2, in normal lung fibroblast cells and lung cancer cells resulted in suppression of BMP-3B levels. The chromatin immunoprecipitation studies identified that the mechanism of Runx2-mediated suppression of BMP-3B is due to the recruitment of Runx2 and histone H3K9-specific methyltransferase Suv39h1 to BMP-3B proximal promoter and a concomitant increase in histone methylation (H3K9) status. The knockdown of Runx2 in H1299 cells resulted in decreased histone H3K9 methylation on BMP-3B promoter and increased BMP-3B expression levels. Furthermore, co-immunoprecipitation studies showed a direct interaction of Runx2 and Suv39h1 proteins. Phenotypically, Runx2 overexpression in H1299 cells increased wound healing response to TGFβ treatment. Conclusions Our studies identified BMP-3B as a new Runx2 target gene and revealed a novel function of Runx2 in silencing of BMP-3B in lung cancers. Our results suggest that Runx2 is a potential therapeutic target to block tumor suppressor gene silencing in lung cancer cells. PMID:22537242

Trends Analysis of rhBMP Utilization in Single-Level Posterior Lumbar Interbody Fusion in the United States.

PubMed

Lao, Lifeng; Cohen, Jeremiah R; Buser, Zorica; Brodke, Darrel S; Youssef, Jim A; Park, Jong-Beom; Yoon, S Tim; Wang, Jeffrey C; Meisel, Hans-Joerg

2017-10-01

Retrospective study. Recombinant human bone morphogenetic protein-2 (rhBMP-2) has been widely used in spinal fusion surgery, but there is little information on rhBMP-2 utilization in single-level posterior lumbar interbody fusion (PLIF). The purpose of our study was to evaluate the trends and demographics of rhBMP-2 utilization in single-level PLIF. Patients who underwent single-level PLIF from 2005 to 2011 were identified by searching ICD-9 diagnosis and procedure codes in the PearlDiver Patient Records Database, a national database of orthopedic insurance records. The year of procedure, age, gender, and region of the United States were recorded for each patient. Results were reported for each variable as the incidence of procedures identified per 100 000 patients searched in the database. A total of 2735 patients had single-level PLIF. The average rate of single-level PLIF with rhBMP-2 maintained at a relatively stable level (28% to 31%) from 2005 to 2009, but decreased in 2010 (9.9%) and 2011 (11.8%). The overall incidence of single-level PLIF without rhBMP-2 (0.68 cases per 100 000 patients) was statistically higher ( P < .01) compared to single-level PLIF with rhBMP-2 (0.21 cases per 100 000 patients). The average rate of single-level PLIF with rhBMP-2 utilization was the highest in West (30.1%), followed by Midwest (26.9%), South (20.5%), and Northeast (17.8%). The highest incidence of single-level PLIF with rhBMP-2 was observed in the age group <65 years (0.3 per 100 000 patients). To our knowledge, this is the first study to report on the demographics associated with rhBMP-2 use in single-level PLIF. There was a 3-fold increase in the rate of PLIF without rhBMP-2 compared to PLIF with rhBMP-2, with both procedures being mainly done in patients less than 65 years of age.

Activation of Bmp2-Smad1 Signal and Its Regulation by Coordinated Alteration of H3K27 Trimethylation in Ras-Induced Senescence

PubMed Central

Kaneda, Atsushi; Fujita, Takanori; Anai, Motonobu; Yamamoto, Shogo; Nagae, Genta; Morikawa, Masato; Tsuji, Shingo; Oshima, Masanobu; Miyazono, Kohei; Aburatani, Hiroyuki

2011-01-01

Cellular senescence involves epigenetic alteration, e.g. loss of H3K27me3 in Ink4a-Arf locus. Using mouse embryonic fibroblast (MEF), we here analyzed transcription and epigenetic alteration during Ras-induced senescence on genome-wide scale by chromatin immunoprecipitation (ChIP)-sequencing and microarray. Bmp2 was the most activated secreted factor with H3K4me3 gain and H3K27me3 loss, whereas H3K4me3 loss and de novo formation of H3K27me3 occurred inversely in repression of nine genes, including two BMP-SMAD inhibitors Smad6 and Noggin. DNA methylation alteration unlikely occurred. Ras-activated cells senesced with nuclear accumulation of phosphorylated SMAD1/5/8. Senescence was bypassed in Ras-activated cells when Bmp2/Smad1 signal was blocked by Bmp2 knockdown, Smad6 induction, or Noggin induction. Senescence was induced when recombinant BMP2 protein was added to Bmp2-knocked-down Ras-activated cells. Downstream Bmp2-Smad1 target genes were then analyzed genome-wide by ChIP-sequencing using anti-Smad1 antibody in MEF that was exposed to BMP2. Smad1 target sites were enriched nearby transcription start sites of genes, which significantly correlated to upregulation by BMP2 stimulation. While Smad6 was one of Smad1 target genes to be upregulated by BMP2 exposure, Smad6 repression in Ras-activated cells with increased enrichment of Ezh2 and gain of H3K27me3 suggested epigenetic disruption of negative feedback by Polycomb. Among Smad1 target genes that were upregulated in Ras-activated cells without increased repressive mark, Parvb was found to contribute to growth inhibition as Parvb knockdown lead to escape from senescence. It was revealed through genome-wide analyses in this study that Bmp2-Smad1 signal and its regulation by harmonized epigenomic alteration play an important role in Ras-induced senescence. PMID:22072987

Heterozygous Mutations in BMP6 Pro-peptide Lead to Inappropriate Hepcidin Synthesis and Moderate Iron Overload in Humans.

PubMed

Daher, Raed; Kannengiesser, Caroline; Houamel, Dounia; Lefebvre, Thibaud; Bardou-Jacquet, Edouard; Ducrot, Nicolas; de Kerguenec, Caroline; Jouanolle, Anne-Marie; Robreau, Anne-Marie; Oudin, Claire; Le Gac, Gerald; Moulouel, Boualem; Loustaud-Ratti, Veronique; Bedossa, Pierre; Valla, Dominique; Gouya, Laurent; Beaumont, Carole; Brissot, Pierre; Puy, Hervé; Karim, Zoubida; Tchernitchko, Dimitri

2016-03-01

Hereditary hemochromatosis is a heterogeneous group of genetic disorders characterized by parenchymal iron overload. It is caused by defective expression of liver hepcidin, the main regulator of iron homeostasis. Iron stimulates the gene encoding hepcidin (HAMP) via the bone morphogenetic protein (BMP)6 signaling to SMAD. Although several genetic factors have been found to cause late-onset hemochromatosis, many patients have unexplained signs of iron overload. We investigated BMP6 function in these individuals. We sequenced the BMP6 gene in 70 consecutive patients with a moderate increase in serum ferritin and liver iron levels who did not carry genetic variants associated with hemochromatosis. We searched for BMP6 mutations in relatives of 5 probands and in 200 healthy individuals (controls), as well as in 2 other independent cohorts of hyperferritinemia patients. We measured serum levels of hepcidin by liquid chromatography-tandem mass spectrometry and analyzed BMP6 in liver biopsy specimens from patients by immunohistochemistry. The functions of mutant and normal BMP6 were assessed in transfected cells using immunofluorescence, real-time quantitative polymerase chain reaction, and immunoblot analyses. We identified 3 heterozygous missense mutations in BMP6 (p.Pro95Ser, p.Leu96Pro, and p.Gln113Glu) in 6 unrelated patients with unexplained iron overload (9% of our cohort). These mutations were detected in less than 1% of controls. p.Leu96Pro also was found in 2 patients from the additional cohorts. Family studies indicated dominant transmission. Serum levels of hepcidin were inappropriately low in patients. A low level of BMP6, compared with controls, was found in a biopsy specimen from 1 patient. In cell lines, the mutated residues in the BMP6 propeptide resulted in defective secretion of BMP6; reduced signaling via SMAD1, SMAD5, and SMAD8; and loss of hepcidin production. We identified 3 heterozygous missense mutations in BMP6 in patients with unexplained iron overload. These mutations lead to loss of signaling to SMAD proteins and reduced hepcidin production. These mutations might increase susceptibility to mild-to-moderate late-onset iron overload. Copyright © 2016 AGA Institute. Published by Elsevier Inc. All rights reserved.

Bone Morphogenic Protein 4-Smad-Induced Upregulation of Platelet-Derived Growth Factor AA Impairs Endothelial Function.

PubMed

Hu, Weining; Zhang, Yang; Wang, Li; Lau, Chi Wai; Xu, Jian; Luo, Jiang-Yun; Gou, Lingshan; Yao, Xiaoqiang; Chen, Zhen-Yu; Ma, Ronald Ching Wan; Tian, Xiao Yu; Huang, Yu

2016-03-01

Bone morphogenic protein 4 (BMP4) is an important mediator of endothelial dysfunction in cardio-metabolic diseases, whereas platelet-derived growth factors (PDGFs) are major angiogenic and proinflammatory mediator, although the functional link between these 2 factors is unknown. The present study investigated whether PDGF mediates BMP4-induced endothelial dysfunction in diabetes mellitus. We generated Ad-Bmp4 to overexpress Bmp4 and Ad-Pdgfa-shRNA to knockdown Pdgfa in mice through tail intravenous injection. SMAD4-shRNA lentivirus, SMAD1-shRNA, and SMAD5 shRNA adenovirus were used for knockdown in human and mouse endothelial cells. We found that PDGF-AA impaired endothelium-dependent vasodilation in aortas and mesenteric resistance arteries. BMP4 upregulated PDGF-AA in human and mouse endothelial cells, which was abolished by BMP4 antagonist noggin or knockdown of SMAD1/5 or SMAD4. BMP4-impared relaxation in mouse aorta was also ameliorated by PDGF-AA neutralizing antibody. Tail injection of Ad-Pdgfa-shRNA ameliorates endothelial dysfunction induced by Bmp4 overexpression (Ad-Bmp4) in vivo. Serum PDGF-AA was elevated in both diabetic patients and diabetic db/db mice compared with nondiabetic controls. Pdgfa-shRNA or Bmp4-shRNA adenovirus reduced serum PDGF-AA concentration in db/db mice. PDGF-AA neutralizing antibody or tail injection with Pdgfa-shRNA adenovirus improved endothelial function in aortas and mesenteric resistance arteries from db/db mice. The effect of PDGF-AA on endothelial function in mouse aorta was also inhibited by Ad-Pdgfra-shRNA to inhibit PDGFRα. The present study provides novel evidences to show that PDGF-AA impairs endothelium-dependent vasodilation and PDGF-AA mediates BMP4-induced adverse effect on endothelial cell function through SMAD1/5- and SMAD4-dependent mechanisms. Inhibition of PGDF-AA ameliorates vascular dysfunction in diabetic mice. © 2016 American Heart Association, Inc.

Construction of doxycycline-mediated BMP-2 transgene combining with APA microcapsules for bone repair.

PubMed

Qian, Dongyang; Bai, Bo; Yan, Guangbin; Zhang, Shujiang; Liu, Qi; Chen, Yi; Tan, Xiaobo; Zeng, Yanjun

2016-01-01

The repairing of large segmental bone defects is difficult for clinical orthopedists at present. Gene therapy is regarded as a promising method for bone defects repair. The present study aimed to construct an effective and controllable Tet-On expression system for transferring hBMP-2 gene into bone marrow mesenchymal progenitor cells (BMSCs). Meanwhile, with combination of alginate-poly-L-lysine-alginate (APA) microencapsulation technology, we attempted to reduce the influence of immunologic rejection and examine the effect of the Tet-On expression system on osteogenesis of BMSCs. The adenovirus encoding hBMP-2 (ADV-hBMP2) was constructed using the means of molecular cloning. The ADV-hBMP2 and Adeno-X Tet-On virus was respectively transfected to the HEK293 for amplification and afterward BMSCs were co-infected with the virus of ADV-hBMP2 and the Adeno-X Tet-On. The expression of hBMP-2 was measured with induction by doxycycline (DOX) at different concentration by means of RT-PCR and ELISA. Combining Tet-On expression system and APA microcapsules with the use of the pulsed high-voltage electrostatic microcapsule instrument, we examined the expression level of hBMP-2 in APA microcapsules by ELISA as well as the osteogenesis by alizarin red S staining. An effective Tet-On expression system for transferring hBMP-2 gene into BMSCs was constructed successfully. Also, the expression of hBMP-2 could be regulated by concentration of DOX. The data exhibited that BMSCs in APA microcapsules maintained the capability of proliferation and differentiation. The combination of Tet-On expression system and APA microcapsules could promote the osteogenesis of BMSCs. According to the results, microencapsulated Tet-On expression system showed the effective characteristics of secreting hBMP-2 and enhancing osteogenesis, which would provide a promising way for bone repair.

Bone morphogenetic protein 7 (BMP-7) influences tendon-bone integration in vitro.

PubMed

Schwarting, Tim; Lechler, Philipp; Struewer, Johannes; Ambrock, Marius; Frangen, Thomas Manfred; Ruchholtz, Steffen; Ziring, Ewgeni; Frink, Michael

2015-01-01

Successful graft ingrowth following reconstruction of the anterior cruciate ligament is governed by complex biological processes at the tendon-bone interface. The aim of this study was to investigate in an in vitro study the effects of bone morphogenetic protein 7 (BMP-7) on tendon-bone integration. To study the biological effects of BMP-7 on the process of tendon-bone-integration, two independent in vitro models were used. The first model involved the mono- and coculture of bovine tendon specimens and primary bovine osteoblasts with and without BMP-7 exposure. The second model comprised the mono- and coculture of primary bovine osteoblasts and fibroblasts. Alkaline phosphatase (ALP), lactate dehydrogenase (LDH), lactate and osteocalcin (OCN) were analyzed by ELISA. Histological analysis and electron microscopy of the tendon specimens were performed. In both models, positive effects of BMP-7 on ALP enzyme activity were observed (p<0.001). Additionally, similar results were noted for LDH activity and lactate concentration. BMP-7 stimulation led to a significant increase in OCN expression. Whereas the effects of BMP-7 on tendon monoculture peaked during an early phase of the experiment (p<0.001), the cocultures showed a maximal increase during the later stages (p<0.001). The histological analysis showed a stimulating effect of BMP-7 on extracellular matrix formation. Organized ossification zones and calcium carbonate-like structures were only observed in the BMP-stimulated cell cultures. This study showed the positive effects of BMP-7 on the biological process of tendon-bone integration in vitro. Histological signs of improved mineralization were paralleled by increased rates of osteoblast-specific protein levels in primary bovine osteoblasts and fibroblasts. Our findings indicated a role for BMP-7 as an adjuvant therapeutic agent in the treatment of ligamentous injuries, and they emphasized the importance of the transdifferentiation process of tendinous fibroblasts at the tendon-bone interface.

Influence of WFIKKN1 on BMP1-mediated activation of latent myostatin.

PubMed

Szláma, György; Vásárhelyi, Viktor; Trexler, Mária; Patthy, László

2016-12-01

The NTR domain of WFIKKN1 protein has been shown to have significant affinity for the prodomain regions of promyostatin and latent myostatin but the biological significance of these interactions remained unclear. In view of its role as a myostatin antagonist, we tested the assumption that WFIKKN1 inhibits the release of myostatin from promyostatin and/or latent myostatin. WFIKKN1 was found to have no effect on processing of promyostatin by furin, the rate of cleavage of latent myostatin by BMP1, however, was significantly enhanced in the presence of WFIKKN1 and this enhancer activity was superstimulated by heparin. Unexpectedly, WFIKKN1 was also cleaved by BMP1 and our studies have shown that the KKN1 fragment generated by BMP1-cleavage of WFIKKN1 contributes most significantly to the observed enhancer activity. Analysis of a pro-TGF-β -based homology model of homodimeric latent myostatin revealed that the BMP1-cleavage sites are buried and not readily accessible to BMP1. In view of this observation, the most plausible explanation for the BMP1-enhancer activity of the KKN1 fragment is that it shifts a conformational equilibrium of latent myostatin from the closed circular structure of the homodimer to a more open form, making the cleavage sites more accessible to BMP1. On the other hand, the observation that the enhancer activity of KKN1 is superstimulated in the presence of heparin is explained by the fact KKN1, latent myostatin, and BMP1 have affinity for heparin and these interactions with heparin increase the local concentrations of the reactants thereby facilitating the action of BMP1. Furin: EC 3.4.21.75; BMP1, bone morphogentic protein 1 or procollagen C-endopeptidase: EC 3.4.24.19. © 2016 The Authors. The FEBS Journal published by John Wiley & Sons Ltd on behalf of Federation of European Biochemical Societies.

Influence of rhBMP-2 on Guided Bone Regeneration for Placement and Functional Loading of Dental Implants: A Radiographic and Histologic Study in Dogs.

PubMed

Ben Amara, Heithem; Lee, Jung-Won; Kim, Jung-Ju; Kang, Yun-Mi; Kang, Eun-Jung; Koo, Ki-Tae

Evidence on the outcomes of functional loading placed in recombinant human bone morphogenetic protein 2 (rhBMP-2)/acellular collagen sponge (ACS)-induced bone is lacking. The aim of this study was to verify whether guided bone regeneration (GBR) with rhBMP-2/ACS enhances regeneration of missing bone and osseointegration of dental implants subject to functional loading. Two bilateral standardized large saddle-type defects (≈10 × 10 × 6 mm) were surgically created in each mandible of seven beagle dogs 2 months after tooth extraction. Defects were immediately reconstructed randomly using rhBMP-2 (O-BMP or InFuse) soaked in ACS, deproteinized bovine bone mineral (DBBM) granules, or ACS alone as surgical control and subsequently covered with collagen membrane. Screw-type sand-blasted, acid-etched dental implants were placed 3 months later into the reconstructed defects and into adjacent bone. Osseointegration was allowed to progress for 3 months before functional loading of 3 months until sacrifice. Significantly more bone fill was radiographically observed for GBR with rhBMP-2/ACS (O-BMP: 92.5%, InFuse: 79%) in comparison to the DBBM (52%) and ACS alone groups (56.6%). Osseointegration was achieved and maintained in all experimental defects challenged by prostheses-driven functional load. The bone density ranged from 37.49% in the ACS group to 64.9% in the rhBMP-2/ACS (InFuse) group with no significance. The highest mean percentage of BIC was found in rhBMP-2/ACS (InFuse: 52.98%) with no statistical difference. Crestal bone resorption was observed around implants placed in reconstructed areas without any significant difference. GBR with rhBMP-2/ACS provided the greatest bone fill among the three treatment procedures. GBR with rhBMP-2/ACS showed efficacy for placement, osseointegration, and functional loading of titanium implants in alveolar ridge defects.

Functional expression of BMP7 receptors in oral epithelial cells. Interleukin-17F production in response to BMP7.

PubMed

Nishio, Kensuke; Ozawa, Yasumasa; Ito, Hisanori; Kifune, Takashi; Narita, Tatsuya; Iinuma, Toshimitsu; Gionhaku, Nobuhito; Asano, Masatake

2017-10-01

Bone morphogenetic proteins (BMPs) are members of the transforming growth factor-β (TGF-β) superfamily. Recently, BMP7 has been demonstrated to be produced by salivary glands and contribute to embryonic branching in mice. The BMP7 in saliva is thought to be delivered to the oral cavity and is expected to contact with stratified squamous epithelial cells which line the surface of oral mucosa. In this study, we attempted to investigate the effects of BMP7 on oral epithelial cells. The expression of BMP receptors was examined by reverse transcriptase-polymerase chain reaction (RT-PCR). OSCCs were stimulated with human recombinant BMP7 (hrBMP7) and the phosphorylation status of Smad1/5/8 was examined by western blotting. For microarray analysis, Ca9-22 cells were stimulated with 100 ng/mL of hrBMP7 and total RNA was extracted and subjected to real-time PCR. The 5'-untranslated region (5'-UTR) of IL-17 F gene was cloned to pGL4-basic vector and used for luciferase assay. Ca9-22 cells were pre-incubated with DM3189, a specific inhibitor of Smad1/5/8, for inhibition assay. All isoforms of type I and type II BMP receptors were expressed in both Ca9-22 and HSC3 cells and BMP7 stimulation resulted in the phosphorylation of Smad1/5/8 in both cell lines. The microarray analysis revealed the induction of interleukin-17 F (IL-17 F), netrin G2 (NTNG2) and hyaluronan synthase 1 (HAS1). Luciferase assay using the 5'-UTR of the IL-17 F gene revealed transcriptional regulation. Induced IL-17 F production was further confirmed at the protein level by ELISA. Smad1/5/8 inhibitor pretreatment decreased IL-17 F expression levels in the cells.

A Retrospective Analysis of Complications Associated With Bone Morphogenetic Protein 2 in Anterior Lumbar Interbody Fusion.

PubMed

Hindoyan, Kevork; Tilan, Justin; Buser, Zorica; Cohen, Jeremiah R; Brodke, Darrel S; Youssef, Jim A; Park, Jong-Beom; Yoon, S Tim; Meisel, Hans-Joerg; Wang, Jeffrey C

2017-04-01

Retrospective review. The aim of our study was to quantify the frequency of complications associated with recombinant human bone morphogenetic protein 2 (rhBMP-2) use in anterior lumbar interbody fusion (ALIF). The orthopedic subset of the Medicare database (PearlDiver) was queried for this retrospective cohort study using International Statistical Classification of Diseases 9 (ICD-9) and Current Procedure Terminology (CPT) codes for ALIF procedures with and without rhBMP-2 between 2005 and 2010. Frequencies of complications and reoperations were then identified within 1 year from the index procedure. Complications included reoperations, pulmonary embolus, deep vein thrombosis, myocardial infarction, nerve-related complications, incision and drainage procedures, wound, sepsis, pneumonia, urinary tract infections, respiratory, heterotopic ossification, retrograde ejaculation, radiculopathy, and other medical complications. Odds ratios (ORs) and 95% confidence intervals (CIs) were used to assess the statistical significance. We identified a total of 41 865 patients who had an ALIF procedure. A total of 14 384 patients received rhBMP-2 while 27 481 did not. Overall, 6016 (41.8%) complications within 1 year from surgery were noted within the group who received rhBMP-2 and 12 950 (47.1%) complications within 1 year from surgery were recorded in those who did not receive rhBMP-2 (OR = 0.81, CI = 0.77-0.84). Overall, exposure to rhBMP-2 was associated with significantly decreased odds of complications with exception to reoperation rates (0.9% rhBMP-2 vs 1.0% no rhBMP-2; OR = 0.88, CI = 0.71-1.09) and radiculopathy (4.4% rhBMP-2 vs 4.3% no rhBMP-2; OR = 1.02, CI = 0.93-1.13). The use of rhBMP-2 in patients undergoing ALIF procedure was associated with a significantly decreased rate of complications. Further studies are needed to elucidate a true incidence of complication.

A Retrospective Analysis of Complications Associated With Bone Morphogenetic Protein 2 in Anterior Lumbar Interbody Fusion

PubMed Central

Hindoyan, Kevork; Tilan, Justin; Cohen, Jeremiah R.; Brodke, Darrel S.; Youssef, Jim A.; Park, Jong-Beom; Yoon, S. Tim; Meisel, Hans-Joerg; Wang, Jeffrey C.

2017-01-01

Study Design: Retrospective review. Objective: The aim of our study was to quantify the frequency of complications associated with recombinant human bone morphogenetic protein 2 (rhBMP-2) use in anterior lumbar interbody fusion (ALIF). Methods: The orthopedic subset of the Medicare database (PearlDiver) was queried for this retrospective cohort study using International Statistical Classification of Diseases 9 (ICD-9) and Current Procedure Terminology (CPT) codes for ALIF procedures with and without rhBMP-2 between 2005 and 2010. Frequencies of complications and reoperations were then identified within 1 year from the index procedure. Complications included reoperations, pulmonary embolus, deep vein thrombosis, myocardial infarction, nerve-related complications, incision and drainage procedures, wound, sepsis, pneumonia, urinary tract infections, respiratory, heterotopic ossification, retrograde ejaculation, radiculopathy, and other medical complications. Odds ratios (ORs) and 95% confidence intervals (CIs) were used to assess the statistical significance. Results: We identified a total of 41 865 patients who had an ALIF procedure. A total of 14 384 patients received rhBMP-2 while 27 481 did not. Overall, 6016 (41.8%) complications within 1 year from surgery were noted within the group who received rhBMP-2 and 12 950 (47.1%) complications within 1 year from surgery were recorded in those who did not receive rhBMP-2 (OR = 0.81, CI = 0.77-0.84). Overall, exposure to rhBMP-2 was associated with significantly decreased odds of complications with exception to reoperation rates (0.9% rhBMP-2 vs 1.0% no rhBMP-2; OR = 0.88, CI = 0.71-1.09) and radiculopathy (4.4% rhBMP-2 vs 4.3% no rhBMP-2; OR = 1.02, CI = 0.93-1.13). Conclusions: The use of rhBMP-2 in patients undergoing ALIF procedure was associated with a significantly decreased rate of complications. Further studies are needed to elucidate a true incidence of complication. PMID:28507884

SNW1 Is a Critical Regulator of Spatial BMP Activity, Neural Plate Border Formation, and Neural Crest Specification in Vertebrate Embryos

PubMed Central

Wu, Mary Y.; Ramel, Marie-Christine; Howell, Michael; Hill, Caroline S.

2011-01-01

Bone morphogenetic protein (BMP) gradients provide positional information to direct cell fate specification, such as patterning of the vertebrate ectoderm into neural, neural crest, and epidermal tissues, with precise borders segregating these domains. However, little is known about how BMP activity is regulated spatially and temporally during vertebrate development to contribute to embryonic patterning, and more specifically to neural crest formation. Through a large-scale in vivo functional screen in Xenopus for neural crest fate, we identified an essential regulator of BMP activity, SNW1. SNW1 is a nuclear protein known to regulate gene expression. Using antisense morpholinos to deplete SNW1 protein in both Xenopus and zebrafish embryos, we demonstrate that dorsally expressed SNW1 is required for neural crest specification, and this is independent of mesoderm formation and gastrulation morphogenetic movements. By exploiting a combination of immunostaining for phosphorylated Smad1 in Xenopus embryos and a BMP-dependent reporter transgenic zebrafish line, we show that SNW1 regulates a specific domain of BMP activity in the dorsal ectoderm at the neural plate border at post-gastrula stages. We use double in situ hybridizations and immunofluorescence to show how this domain of BMP activity is spatially positioned relative to the neural crest domain and that of SNW1 expression. Further in vivo and in vitro assays using cell culture and tissue explants allow us to conclude that SNW1 acts upstream of the BMP receptors. Finally, we show that the requirement of SNW1 for neural crest specification is through its ability to regulate BMP activity, as we demonstrate that targeted overexpression of BMP to the neural plate border is sufficient to restore neural crest formation in Xenopus SNW1 morphants. We conclude that through its ability to regulate a specific domain of BMP activity in the vertebrate embryo, SNW1 is a critical regulator of neural plate border formation and thus neural crest specification. PMID:21358802

Osteoblastic differentiation of human and equine adult bone marrow-derived mesenchymal stem cells when BMP-2 or BMP-7 homodimer genetic modification is compared to BMP-2/7 heterodimer genetic modification in the presence and absence of dexamethasone.

PubMed

Carpenter, Ryan S; Goodrich, Laurie R; Frisbie, David D; Kisiday, John D; Carbone, Beth; McIlwraith, C Wayne; Centeno, Christopher J; Hidaka, Chisa

2010-10-01

Bone marrow-derived mesenchymal stem cells (BMDMSCs) have been targeted for use in enhancement of bone healing; and their osteogenic potential may be further augmented by genes encoding bone morphogenetic proteins (BMP's). The purpose of this study was to compare the effect of genetic modification of human and equine BMDMSCs with BMP-2 or -7 or BMP-2 and -7 on their osteoblastogenic differentiation in the presence or absence of dexamethasone. The BMDMSCs were harvested from the iliac crest of three human donors and tuber coxae of three equine donors. Monolayer cells were genetically modified using adenovirus vectors encoding BMP-2, -7 or both and cultured in the presence or absence of dexamethasone. Expression of BMPs was confirmed by enzyme linked immunosorbent assay (ELISA). To evaluate osteoblastic differentiation, cellular morphology was assessed every other day and expression and secretion of alkaline phosphatase (ALP), as well as expression levels of osteonectin (OSTN), osteocalcin (OCN), and runt-related transcription factor-2 (Runx2) were measured for up to 14 days. Human and equine BMDMSCs showed a capacity for osteogenic differentiation regardless of genetic modification or dexamethasone supplementation. Dexamethasone supplementation was more important for osteoblastogenic differentiation of equine BMDMSCs than human BMDMSCs. Genetic modification of BMDMSCs increased ALP secretion with AdBMP-2 homodimer having the greatest effect in both human and equine cells compared to AdBMP 7 or AdBMP 2/7. BMP protein elution rates reached their maximal concentration between day 4 and 8 and remained relatively stable thereafter, suggesting that genetically modified BMDMSCs could be useful for cell-based delivery of BMPs to a site of bone formation. Published by Wiley Periodicals, Inc. J Orthop Res 28:1330-1337, 2010.

Increased Retinal Expression of the Pro-Angiogenic Receptor GPR91 via BMP6 in a Mouse Model of Juvenile Hemochromatosis

PubMed Central

Arjunan, Pachiappan; Gnanaprakasam, Jaya P.; Ananth, Sudha; Romej, Michelle A.; Rajalakshmi, Veeranan-Karmegam; Prasad, Puttur D.; Martin, Pamela M.; Gurusamy, Mariappan; Thangaraju, Muthusamy; Bhutia, Yangzom D.; Ganapathy, Vadivel

2016-01-01

Purpose Hemochromatosis, an iron-overload disease, occurs as adult and juvenile types. Mutations in hemojuvelin (HJV), an iron-regulatory protein and a bone morphogenetic protein (BMP) coreceptor, underlie most of the juvenile type. Hjv−/− mice accumulate excess iron in retina and exhibit aberrant vascularization and angiomas. A succinate receptor, GPR91, is pro-angiogenic in retina. We hypothesized that Hjv−/− retinas have increased BMP signaling and increased GPR91 expression as the basis of angiomas. Methods Expression of GPR91 was examined by qPCR, immunofluorescence, and Western blot in wild-type and Hjv−/− mouse retinas and pRPE cells. Influence of excess iron and BMP6 on GPR91 expression was investigated in ARPE-19 cells, and wild-type and Hjv−/− pRPE cells. Succinate was used to activate GPR91 and determine the effects of GPR91 signaling on VEGF expression. Signaling of BMP6 was studied by the expression of Smad1/5/8 and pSmad4, and the BMP-target gene Id1. The interaction of pSmad4 with GPR91 promoter was studied by ChIP. Results Expression of GPR91 was higher in Hjv−/− retinas and RPE than in wild-type counterparts. Unexpectedly, BMP signaling was increased, not decreased, in Hjv−/− retinas and RPE. Bone morphogenetic protein 6 induced GPR91 in RPE, suggesting that increased BMP signaling in Hjv−/− retinas was likely responsible for GPR91 upregulation. Exposure of RPE to excess iron and succinate as well as BMP6 and succinate increased VEGF expression. Bone morphogenetic protein 6 promoted the interaction of pSmad4 with GPR91 promoter in RPE. Conclusions G-protein-coupled receptor 91 is a BMP6 target and Hjv deletion enhances BMP signaling in retina, thus underscoring a role for excess iron and hemochromatosis in abnormal retinal vascularization. PMID:27046124

Increased Retinal Expression of the Pro-Angiogenic Receptor GPR91 via BMP6 in a Mouse Model of Juvenile Hemochromatosis.

PubMed

Arjunan, Pachiappan; Gnanaprakasam, Jaya P; Ananth, Sudha; Romej, Michelle A; Rajalakshmi, Veeranan-Karmegam; Prasad, Puttur D; Martin, Pamela M; Gurusamy, Mariappan; Thangaraju, Muthusamy; Bhutia, Yangzom D; Ganapathy, Vadivel

2016-04-01

Hemochromatosis, an iron-overload disease, occurs as adult and juvenile types. Mutations in hemojuvelin (HJV), an iron-regulatory protein and a bone morphogenetic protein (BMP) coreceptor, underlie most of the juvenile type. Hjv(-/-) mice accumulate excess iron in retina and exhibit aberrant vascularization and angiomas. A succinate receptor, GPR91, is pro-angiogenic in retina. We hypothesized that Hjv(-/-) retinas have increased BMP signaling and increased GPR91 expression as the basis of angiomas. Expression of GPR91 was examined by qPCR, immunofluorescence, and Western blot in wild-type and Hjv(-/-) mouse retinas and pRPE cells. Influence of excess iron and BMP6 on GPR91 expression was investigated in ARPE-19 cells, and wild-type and Hjv(-/-) pRPE cells. Succinate was used to activate GPR91 and determine the effects of GPR91 signaling on VEGF expression. Signaling of BMP6 was studied by the expression of Smad1/5/8 and pSmad4, and the BMP-target gene Id1. The interaction of pSmad4 with GPR91 promoter was studied by ChIP. Expression of GPR91 was higher in Hjv(-/-) retinas and RPE than in wild-type counterparts. Unexpectedly, BMP signaling was increased, not decreased, in Hjv(-/-) retinas and RPE. Bone morphogenetic protein 6 induced GPR91 in RPE, suggesting that increased BMP signaling in Hjv(-/-) retinas was likely responsible for GPR91 upregulation. Exposure of RPE to excess iron and succinate as well as BMP6 and succinate increased VEGF expression. Bone morphogenetic protein 6 promoted the interaction of pSmad4 with GPR91 promoter in RPE. G-protein-coupled receptor 91 is a BMP6 target and Hjv deletion enhances BMP signaling in retina, thus underscoring a role for excess iron and hemochromatosis in abnormal retinal vascularization.

Lactoferricin enhances BMP7-stimulated anabolic pathways in intervertebral disc cells.

PubMed

Ellman, Michael B; Kim, Jaesung; An, Howard S; Chen, Di; Kc, Ranjan; Li, Xin; Xiao, Guozhi; Yan, Dongyao; Suh, Joon; van Wjnen, Andre J; Wang, James H-C; Kim, Su-Gwan; Im, Hee-Jeong

2013-07-25

Bone-morphogenetic protein-7 (BMP7) is a well-known anabolic and anti-catabolic growth factor on intervertebral disc (IVD) matrix and cell homeostasis. Similarly, Lactoferricin B (LfcinB) has recently been shown to have pro-anabolic, anti-catabolic, anti-oxidative and/or anti-inflammatory effects in bovine disc cells in vitro. In this study, we investigated the potential benefits of using combined peptide therapy with LfcinB and BMP7 for intervertebral disc matrix repair and to understand cellular and signaling mechanisms controlled by these factors. We studied the effects of BMP7 and LfcinB as individual treatments and combined therapy on bovine nucleus pulposus (NP) cells by assessing proteoglycan (PG) accumulation and synthesis, and the gene expression of matrix protein aggrecan and transcription factor SOX-9. We also analyzed the role of Noggin, a BMP antagonist, in IVD tissue and examined its effect after stimulation with LfcinB. To understand the molecular mechanisms by which LfcinB synergizes with BMP7, we investigated the ERK-SP1 axis as a downstream intracellular signaling regulator involved in BMP7 and LfcinB-mediated activities. Treatment of bovine NP cells cultured in alginate with LfcinB plus BMP7 synergistically stimulates PG synthesis and accumulation in part by upregulation of aggrecan gene expression. The synergism results from LfcinB-mediated activation of Sp1 and SMAD signaling pathways by (i) phosphorylation of SMAD 1/5/8; (ii) downregulation of SMAD inhibitory factors [i.e., noggin and SMAD6 (inhibitory SMAD)]; and (iii) upregulation of SMAD4 (universal co-SMAD). These data indicate that LfcinB-suppression of Noggin may eliminate the negative feedback of BMP7, thereby maximizing biological activity of BMP7 and ultimately shifting homeostasis to a pro-anabolic state in disc cells. We propose that combination growth factor therapy using BMP7 and LfcinB may be beneficial for treatment of disc degeneration. Copyright © 2013 Elsevier B.V. All rights reserved.

Expression characteristics of BMP2, BMPR-IA and Noggin in different stages of hair follicle in yak skin.

PubMed

Song, Liang-Li; Cui, Yan; Yu, Si-Jiu; Liu, Peng-Gang; Liu, Jun; Yang, Xue; He, Jun-Feng; Zhang, Qian

2018-05-01

Bone morphogenetic protein 2 (BMP2), BMP receptor-IA (BMPR-IA), and the BMP2 antagonist Noggin are important proteins involved in regulating the hair follicle (HF) cycle in skin. In order to explore the expression profiles of BMP2, BMPR-IA, and Noggin in the HF cycle of yak skin, we collected adult yak skin in the telogen, proanagen, and midanagen phases of HFs and evaluated gene and protein expression by real-time quantitative polymerase chain reaction (qRT-PCR), western blotting, and immunohistochemistry. qRT-PCR and western blotting results showed that BMP2 and BMPR-IA expression levels were highest in the telogen of HFs and higher than that of Noggin in the same phase. The expression of Noggin was significantly higher in proanagen and midanagen phases of HFs than in the telogen phase, with the highest expression observed in the proanagen phase. Moreover, the expression of Noggin in the proanagen phase was significantly higher than those of BMP2 and BMPR-IA during the same phase. Immunohistochemistry results showed that BMP2, BMPR-IA, and Noggin were expressed in the skin epidermis, sweat glands, sebaceous glands, HF outer root sheath, and hair matrix. In summary, the characteristic expression profiles of BMP2, BMPR-IA, and Noggin suggested that BMP2 and BMPR-IA had inhibitory effects on the growth of HFs in yaks, whereas Noggin promoted the growth of yak HFs, mainly by affecting skin epithelial cell activity. These results provide a basis for further studies of HF development and cycle transition in yak skin. Copyright © 2017. Published by Elsevier Inc.

Leiomyoma-derived transforming growth factor-β impairs bone morphogenetic protein-2-mediated endometrial receptivity.

PubMed

Doherty, Leo F; Taylor, Hugh S

2015-03-01

To determine whether transforming growth factor (TGF)-β3 is a paracrine signal secreted by leiomyoma that inhibits bone morphogenetic protein (BMP)-mediated endometrial receptivity and decidualization. Experimental. Laboratory. Women with symptomatic leiomyomas. Endometrial stromal cells (ESCs) and leiomyoma cells were isolated from surgical specimens. Leiomyoma-conditioned media (LCM) was applied to cultured ESC. The TGF-β was blocked by two approaches: TGF-β pan-specific antibody or transfection with a mutant TGF-β receptor type II. Cells were then treated with recombinant human BMP-2 to assess BMP responsiveness. Expression of BMP receptor types 1A, 1B, 2, as well as endometrial receptivity mediators HOXA10 and leukemia inhibitory factor (LIF). Enzyme-linked immunosorbent assay showed elevated TGF-β levels in LCM. LCM treatment of ESC reduced expression of BMP receptor types 1B and 2 to approximately 60% of pretreatment levels. Preincubation of LCM with TGF-β neutralizing antibody or mutant TGF receptor, but not respective controls, prevented repression of BMP receptors. HOXA10 and LIF expression was repressed in recombinant human BMP-2 treated, LCM exposed ESC. Pretreatment of LCM with TGF-β antibody or transfection with mutant TGF receptor prevented HOXA10 and LIF repression. Leiomyoma-derived TGF-β was necessary and sufficient to alter endometrial BMP-2 responsiveness. Blockade of TGF-β prevents repression of BMP-2 receptors and restores BMP-2-stimulated expression of HOXA10 and LIF. Blockade of TGF signaling is a potential strategy to improve infertility and pregnancy loss associated with uterine leiomyoma. Copyright © 2015 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Alk2/ACVR1 and Alk3/BMPR1A Provide Essential Function for Bone Morphogenetic Protein Induced Retinal Angiogenesis

PubMed Central

Lee, Heon-Woo; Chong, Diana C.; Ola, Roxana; Dunworth, William P.; Meadows, Stryder; Ka, Jun; Kaartinen, Vesa M.; Qyang, Yibing; Cleaver, Ondine; Bautch, Victoria L.; Eichmann, Anne; Jin, Suk-Won

2017-01-01

Objective Increasing evidence suggests that Bone Morphogenetic Protein (BMP) signaling regulates angiogenesis. Here, we aimed to define the function of BMP receptors in regulating early post-natal angiogenesis by analysis of inducible, endothelial specific deletion of the BMP receptor components Bmpr2, Alk1, Alk2 and Alk3 in mouse retinal vessels. Approach and Results Expression analysis of several BMP ligands showed that pro-angiogenic BMP ligands are highly expressed in postnatal retinas. Consistently, BMP receptors are also strongly expressed in retina with a distinct pattern. To assess the function of BMP signaling in retinal angiogenesis, we first generated mice carrying an endothelial-specific inducible deletion of BMP Type 2 receptor (Bmpr2). Postnatal deletion of Bmpr2 in endothelial cells substantially decreased the number of angiogenic sprouts at the vascular front and branchpoints behind the front, leading to attenuated radial expansion. To identify critical BMPR1s associated with BMPR2 in retinal angiogenesis, we generated endothelial-specific inducible deletion of three BMPR1s abundantly expressed in endothelial cells and analyzed the respective phenotypes. Among these, endothelial specific deletion of either Alk2/acvr1 or Alk3/Bmpr1a caused a delay in radial expansion, reminiscent of vascular defects associated with postnatal endothelial specific deletion of BMPR2, suggesting that ALK2/ACVR1 and ALK3/BMPR1A are likely to be the critical BMPR1s necessary for pro-angiogenic BMP signaling in retinal vessels. Conclusions Our data identify BMP signaling mediated by coordination of ALK2/ACVR1, ALK3/BMPR1A, and BMPR2 as an essential pro-angiogenic cue for retinal vessels. PMID:28232325

Bone mesenchymal stem cells co-expressing VEGF and BMP-6 genes to combat avascular necrosis of the femoral head

PubMed Central

Liao, Hongxing; Zhong, Zhixiong; Liu, Zhanliang; Li, Liangping; Ling, Zemin; Zou, Xuenong

2018-01-01

The aim of the present study was to investigate the potential of bone mesenchymal stem cells (BMSCs) treated with a combination of vascular endothelial growth factor (VEGF) and bone morphogenetic protein-6 (BMP-6) genes for the treatment of avascular necrosis of the femoral head (ANFH). Rat BMSCs were isolated and purified using a density gradient centrifugation method. The purity and characteristics of the BMSCs were detected by cell surface antigens identification using flow cytometry. The experimental groups were administered with one of the following adeno-associated virus (AAV) vector constructs: AAV-green fluorescent protein (AAV-GFP), AAV-BMP-6, AAV-VEGF or AAV-VEGF-BMP-6. The expression of VEGF and BMP-6 was detected by reverse transcription-quantitative polymerase chain reaction, western blotting and ELISA assays. The effects of VEGF and BMP-6 on BMSCs were evaluated by angiogenic and osteogenic assays. The transfected BMSCs were combined with a biomimetic synthetic scaffold poly lactide-co-glycolide (PLAGA) and they were then subcutaneously implanted into nude mice. After four weeks, the implants were analyzed with histology and subsequent immunostaining to evaluate the effects of BMSCs on blood vessel and bone formation in vivo. In the AAV-VEGF-BMP-6 group, the expression levels of VEGF and BMP-6 were significantly increased and human umbilical vein endothelial cells tube formation was significantly enhanced compared with other groups. Capillaries and bone formation in the AAV-VEGF-BMP-6 group was significantly higher compared with the other groups. The results of the present study suggest that BMSCs expressing both VEGF and BMP-6 induce an increase in blood vessels and bone formation, which provides theoretical support for ANFH gene therapy. PMID:29399103

Bone mesenchymal stem cells co-expressing VEGF and BMP-6 genes to combat avascular necrosis of the femoral head.

PubMed

Liao, Hongxing; Zhong, Zhixiong; Liu, Zhanliang; Li, Liangping; Ling, Zemin; Zou, Xuenong

2018-01-01

The aim of the present study was to investigate the potential of bone mesenchymal stem cells (BMSCs) treated with a combination of vascular endothelial growth factor (VEGF) and bone morphogenetic protein-6 (BMP-6) genes for the treatment of avascular necrosis of the femoral head (ANFH). Rat BMSCs were isolated and purified using a density gradient centrifugation method. The purity and characteristics of the BMSCs were detected by cell surface antigens identification using flow cytometry. The experimental groups were administered with one of the following adeno-associated virus (AAV) vector constructs: AAV-green fluorescent protein (AAV-GFP), AAV-BMP-6, AAV-VEGF or AAV-VEGF-BMP-6. The expression of VEGF and BMP-6 was detected by reverse transcription-quantitative polymerase chain reaction, western blotting and ELISA assays. The effects of VEGF and BMP-6 on BMSCs were evaluated by angiogenic and osteogenic assays. The transfected BMSCs were combined with a biomimetic synthetic scaffold poly lactide-co-glycolide (PLAGA) and they were then subcutaneously implanted into nude mice. After four weeks, the implants were analyzed with histology and subsequent immunostaining to evaluate the effects of BMSCs on blood vessel and bone formation in vivo . In the AAV-VEGF-BMP-6 group, the expression levels of VEGF and BMP-6 were significantly increased and human umbilical vein endothelial cells tube formation was significantly enhanced compared with other groups. Capillaries and bone formation in the AAV-VEGF-BMP-6 group was significantly higher compared with the other groups. The results of the present study suggest that BMSCs expressing both VEGF and BMP-6 induce an increase in blood vessels and bone formation, which provides theoretical support for ANFH gene therapy.

Bone morphogenetic protein 9 as a key regulator of liver progenitor cells in DDC-induced cholestatic liver injury.

PubMed

Addante, Annalisa; Roncero, Cesáreo; Almalé, Laura; Lazcanoiturburu, Nerea; García-Álvaro, María; Fernández, Margarita; Sanz, Julián; Hammad, Seddik; Nwosu, Zeribe C; Lee, Se-Jin; Fabregat, Isabel; Dooley, Steven; Ten Dijke, Peter; Herrera, Blanca; Sánchez, Aránzazu

2018-05-11

Bone morphogenetic protein 9 (BMP9) interferes with liver regeneration upon acute injury, while promoting fibrosis upon carbon tetrachloride-induced chronic injury. We have now addressed the role of BMP9 in 3,5 diethoxicarbonyl-1,4 dihydrocollidine (DDC)-induced cholestatic liver injury, a model of liver regeneration mediated by hepatic progenitor cell (known as oval cell), exemplified as ductular reaction and oval cell expansion. WT and BMP9KO mice were submitted to DDC diet. Livers were examined for liver injury, fibrosis, inflammation and oval cell expansion by serum biochemistry, histology, RT-qPCR and western blot. BMP9 signalling and effects in oval cells were studied in vitro using western blot and transcriptional assays, plus functional assays of DNA synthesis, cell viability and apoptosis. Crosslinking assays and short hairpin RNA approaches were used to identify the receptors mediating BMP9 effects. Deletion of BMP9 reduces liver damage and fibrosis, but enhances inflammation upon DDC feeding. Molecularly, absence of BMP9 results in overactivation of PI3K/AKT, ERK-MAPKs and c-Met signalling pathways, which together with an enhanced ductular reaction and oval cell expansion evidence an improved regenerative response and decreased damage in response to DDC feeding. Importantly, BMP9 directly targets oval cells, it activates SMAD1,5,8, decreases cell growth and promotes apoptosis, effects that are mediated by Activin Receptor-Like Kinase 2 (ALK2) type I receptor. We identify BMP9 as a negative regulator of oval cell expansion in cholestatic injury, its deletion enhancing liver regeneration. Likewise, our work further supports BMP9 as an attractive therapeutic target for chronic liver diseases. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

BMP-7 Preserves Surface Integrity of Degradable-ceramic Cranioplasty in a Göttingen Minipig Model

PubMed Central

Schulz, Peter; Klünter, Tim; Deisinger, Ulrike; Diez, Claudius; Waiss, Waltraud; Kirschneck, Christian; Reichert, Torsten E.; Detsch, Rainer

2017-01-01

Background: The aim of the study was to evaluate the integrity of a craniotomy grafted site in a minipig model using different highly porous calcium phosphate ceramic scaffolds either loaded or nonloaded with bone morphogenetic protein-7 (BMP-7). Methods: Four craniotomies with a diameter of 15 mm (critical-size defect) were grafted with different highly porous (92–94 vol%) calcium phosphate ceramics [hydroxyapatite (HA), tricalcium phosphate (TCP), and biphasic calcium phosphate (BCP; a mixture of HA and TCP)] in 10 Göttingen minipigs: (a) group I (n = 5): HA versus BCP; (b) group II (n = 5): TCP versus BCP. One scaffold of each composition was supplied with 250 μg of BMP-7. In vivo computed tomography scan and fluorochrome bone labeling were performed. Specimens were evaluated 14 weeks after surgery by environmental scanning electron microscopy, fluorescence microscopy, and Giemsa staining histology. Results: BMP-7 significantly enhanced bone formation in TCP (P = 0.047). Slightly enhanced bone formation was observed in BCP (P = 0.059) but not in HA implants. BMP-7 enhanced ceramic degradation in TCP (P = 0.05) and BCP (P = 0.05) implants but not in HA implants. Surface integrity of grafted site was observed in all BMP-7-loaded implants after successful creeping substitution by the newly formed bone. In 9 of 10 HA implants without BMP-7, partial collapse of the implant site was observed. All TCP implants without BMP-7 collapsed. Fluorescent labeling showed bone formation at week 1 in BMP-7-stimulated implants. Conclusions: BMP-7 supports bone formation, ceramic degradation, implant integration, and surface integrity of the grafted site. PMID:28458969

Trends and Cost of Posterior Cervical Fusions With and Without Recombinant Human Bone Morphogenetic Protein-2 in the US Medicare Population.

PubMed

Myhre, Sue Lynn; Buser, Zorica; Meisel, Hans-Joerg; Brodke, Darrel S; Yoon, S Tim; Wang, Jeffrey C; Park, Jong-Beom; Youssef, Jim A

2017-06-01

Retrospective database review. To analyze and report the trends and cost of posterior cervical fusions (PCFs) with and without off-label recombinant human bone morphogenetic protein-2 (rhBMP-2) in the Medicare population. Patient records from the PearlDiver database were retrospectively reviewed from January 1, 2005, to December 31, 2012, to distinguish individuals who underwent a PCF with or without rhBMP-2. Total numbers, incidence, age, gender, geographic region, reimbursement, and length of stay were analyzed and summarized. The combined total of non-rhBMP-2 (n = 39 479; 85.51%) and rhBMP-2 PCF (n = 6692; 14.49%) procedures performed between 2005 and 2012 was 46 171. In general, the number of PCFs without rhBMP-2 consistently increased over time, while the number of PCFs with rhBMP-2 had only a slight increase from 2005 to 2012. On average, PCFs without rhBMP-2 were associated with $1197 higher cost than those with rhBMP-2, but the average length of stay was similar (6 days). From 2005 to 2012, the average cost for procedures with and without rhBMP-2 increased by $12 605 and $7291, respectively. The percentage of rhBMP-2 use peaked in 2007 and dwindled until 2010, and declined an additional 2.84% from 2011 to 2012. Multiple age, region, and gender tendencies were observed. To our knowledge, this was the first study to use the PearlDiver database to report incidence and cost trends of PCF procedures. This article provides meaningful trend data on PCFs to surgeons and clinicians, researchers, and patients, as well as functions as a beacon for future research questions.

The effect of recombinant bone morphogenetic protein-2 in revision tibiotalocalcaneal arthrodesis: utilization of the Retrograde Arthrodesis Intramedullary Nail database.

PubMed

DeVries, J George; Nguyen, Minh; Berlet, Gregory C; Hyer, Christopher F

2012-01-01

The use of bone morphogenetic protein-2 (BMP-2) has been recommended for high-risk fusions and nonunion. Patients undergoing revisional tibiotalocalcaneal (TTC) arthrodesis via a retrograde arthrodesis nail to evaluate the influence of BMP-2 on rate of fusion in this high-risk population are presented. A retrospective chart and radiographic review were performed on 23 patients with failed prior fusion attempts at the ankle treated with retrograde intramedullary nailing. Sixteen patients were treated without BMP-2 (None group) and 7 were treated with BMP-2 (BMP group). The primary and secondary end-points were successful fusion, and time to fusion, respectively, with a variety of variables evaluated for influence. Other than the use of BMP-2, there were no statistical differences in the patient population. Overall, 11 of 16 ankles (68.8%) in the None group and 5 of 7 ankles (71.4%) in the BMP group resulted in a stable, functional limb. Rate of complication was similar between the 2 groups (p > .05). Time to radiographic ankle union was 115.2 and 184.0 days in the None and BMP groups (p > .05). The effect of BMP-2 on revisional TTC fusions with retrograde nails is reported here. The overall result as a stable, functional limb was 69.6%, which suggests that revision surgery in this high-risk population is a reasonable consideration. Even though this study was unable to demonstrate statistically significant differences, biologic augmentation with BMP-2 did not increase the complication rate and showed a slightly enhanced salvage rate for revision TTC fusions with an intramedullary nail. Copyright © 2012 American College of Foot and Ankle Surgeons. Published by Elsevier Inc. All rights reserved.

Bone Formation in a Rat Tibial Defect Model Using Carboxymethyl Cellulose/BioC/Bone Morphogenic Protein-2 Hybrid Materials

PubMed Central

Kim, Hak-Jun; Park, Kyeongsoon; Kim, Sung Eun; Song, Hae-Ryong

2014-01-01

The objective of this study was to assess whether carboxymethyl cellulose- (CMC-) based hydrogel containing BioC (biphasic calcium phosphate (BCP); tricalcium phosphate (TCP) : hydroxyapatite (Hap) = 70 : 30) and bone morphogenic protein-2 (BMP-2) led to greater bone formation than CMC-based hydrogel containing BioC without BMP-2. In order to demonstrate bone formation at 4 and 8 weeks, plain radiographs, microcomputed tomography (micro-CT) evaluation, and histological studies were performed after implantation of all hybrid materials on an 8 mm defect of the right tibia in rats. The plain radiographs and micro-CT analyses revealed that CMC/BioC/BMP-2 (0.5 mg) led to much greater mineralization at 4 and 8 weeks than did CMC/BioC or CMC/Bio/BMP-2 (0.1 mg). Likewise, bone formation and bone remodeling studies revealed that CMC/BioC/BMP-2 (0.5 mg) led to a significantly greater amount of bone formation and bone remodeling at 4 and 8 weeks than did CMC/BioC or CMC/BioC/BMP-2 (0.1 mg). Histological studies revealed that mineralized bone tissue was present around the whole circumference of the defect site with CMC/BioC/BMP-2 (0.5 mg) but not with CMC/BioC or CMC/BioC/BMP-2 (0.1 mg) at 4 and 8 weeks. These results suggest that CMC/BioC/BMP-2 hybrid materials induced greater bone formation than CMC/BioC hybrid materials. Thus, CMC/BioC/BMP-2 hybrid materials may be used as an injectable substrate to regenerate bone defects. PMID:24804202

Complications of Anterior Cervical Fusion using a Low-dose Recombinant Human Bone Morphogenetic Protein-2

PubMed Central

Kukreja, Sunil; Ahmed, Osama I; Haydel, Justin; Nanda, Anil

2015-01-01

Objective There are several reports, which documented a high incidence of complications following the use of recombinant human bone morphogenetic protein-2 (rhBMP-2) in anterior cervical fusions (ACFs). The objective of this study is to share our experience with low-dose rhBMP-2 in anterior cervical spine. Methods We performed a retrospective analysis of 197 patients who underwent anterior cervical fusion (ACF) with the use of recombinant human bone morphogenetic protein-2 (rhBMP-2) during 2007-2012. A low-dose rhBMP-2 (0.7mg/level) sponge was placed exclusively within the cage. In 102 patients demineralized bone matrix (DBM) was filled around the BMP sponge. Incidence and severity of dysphagia was determined by 5 points SWAL-QOL scale. Results Two patients had prolonged hospitalization due to BMP unrelated causes. Following the discharge, 13.2%(n=26) patients developed dysphagia and 8.6%(n=17) patients complained of neck swelling. More than half of the patients (52.9%, n=9) with neck swelling also had associated dysphagia; however, only 2 of these patients necessitated readmission. Both of these patients responded well to the intravenous dexamethasone. The use of DBM did not affect the incidence and severity of complications (p>0.05). Clinico-radiological evidence of fusion was not observed in 2 patients. Conclusion A low-dose rhBMP-2 in ACFs is not without risk. However, the incidence and severity of complications seem to be lower with low-dose BMP placed exclusively inside the cage. Packing DBM putty around the BMP sponge does not affect the safety profile of rhBMP-2 in ACFs. PMID:26217385

Homeobox protein MSX-1 inhibits expression of bone morphogenetic protein 2, bone morphogenetic protein 4, and lymphoid enhancer-binding factor 1 via Wnt/β-catenin signaling to prevent differentiation of dental mesenchymal cells during the late bell stage.

PubMed

Feng, Xiao-Yu; Wu, Xiao-Shan; Wang, Jin-Song; Zhang, Chun-Mei; Wang, Song-Lin

2018-02-01

Homeobox protein MSX-1 (hereafter referred to as MSX-1) is essential for early tooth-germ development. Tooth-germ development is arrested at bud stage in Msx1 knockout mice, which prompted us to study the functions of MSX-1 beyond this stage. Here, we investigated the roles of MSX-1 during late bell stage. Mesenchymal cells of the mandibular first molar were isolated from mice at embryonic day (E)17.5 and cultured in vitro. We determined the expression levels of β-catenin, bone morphogenetic protein 2 (Bmp2), Bmp4, and lymphoid enhancer-binding factor 1 (Lef1) after knockdown or overexpression of Msx1. Our findings suggest that knockdown of Msx1 promoted expression of Bmp2, Bmp4, and Lef1, resulting in elevated differentiation of odontoblasts, which was rescued by blocking the expression of these genes. In contrast, overexpression of Msx1 decreased the expression of Bmp2, Bmp4, and Lef1, leading to a reduction in odontoblast differentiation. The regulation of Bmp2, Bmp4, and Lef1 by Msx1 was mediated by the Wnt/β-catenin signaling pathway. Additionally, knockdown of Msx1 impaired cell proliferation and slowed S-phase progression, while overexpression of Msx1 also impaired cell proliferation and prolonged G1-phase progression. We therefore conclude that MSX-1 maintains cell proliferation by regulating transition of cells from G1-phase to S-phase and prevents odontoblast differentiation by inhibiting expression of Bmp2, Bmp4, and Lef1 at the late bell stage via the Wnt/β-catenin signaling pathway. © 2017 Eur J Oral Sci.

Effect of Emdogain enamel matrix derivative and BMP-2 on the gene expression and mineralized nodule formation of alveolar bone proper-derived stem/progenitor cells.

PubMed

Fawzy El-Sayed, Karim M; Dörfer, Christof; Ungefroren, Hendrick; Kassem, Neemat; Wiltfang, Jörg; Paris, Sebastian

2014-07-01

The objective of this study was to evaluate the effect of Emdogain (Enamel Matrix Derivative, EMD) and Bone Morphogenetic Protein-2 (BMP-2), either solely or in combination, on the gene expression and mineralized nodule formation of alveolar bone proper-derived stem/progenitor cells. Stem/progenitor cells were isolated from human alveolar bone proper, magnetically sorted using STRO-1 antibodies, characterized flowcytometrically for their surface markers' expression, and examined for colony formation and multilineage differentiation potential. Subsequently, cells were treated over three weeks with 100 μg/ml Emdogain (EMD-Group), or 100 ng/ml BMP-2 (BMP-Group), or a combination of 100 ng/ml BMP-2 and 100 μg/ml Emdogain (BMP/EMD-Group). Unstimulated stem/progenitor cells (MACS(+)-Group) and osteoblasts (OB-Group) served as controls. Osteogenic gene expression was analyzed using RTq-PCR after 1, 2 and 3 weeks (N = 3/group). Mineralized nodule formation was evaluated by Alizarin-Red staining. BMP and EMD up-regulated the osteogenic gene expression. The BMP Group showed significantly higher expression of Collagen-I, III, and V, Alkaline phosphatase and Osteonectin compared to MACS(+)- and OB-Group (p < 0.05; Two-way ANOVA/Bonferroni) with no mineralized nodule formation. Under in-vitro conditions, Emdogain and BMP-2 up-regulate the osteogenic gene expression of stem/progenitor cells. The combination of BMP-2 and Emdogain showed no additive effect and would not be recommended for a combined clinical stimulation. Copyright © 2013 European Association for Cranio-Maxillo-Facial Surgery. Published by Elsevier Ltd. All rights reserved.

Magnesium modification up-regulates the bioactivity of bone morphogenetic protein-2 upon calcium phosphate cement via enhanced BMP receptor recognition and Smad signaling pathway.

PubMed

Ding, Sai; Zhang, Jing; Tian, Yu; Huang, Baolin; Yuan, Yuan; Liu, Changsheng

2016-09-01

Efficient presentation of growth factors is one of the great challenges in tissue engineering. In living systems, bioactive factors exist in soluble as well as in matrix-bound forms, both of which play an integral role in regulating cell behaviors. Herein, effect of magnesium on osteogenic bioactivity of recombinant human bone morphogenetic protein-2 (rhBMP-2) was investigated systematically with a series of Mg modified calcium phosphate cements (xMCPCs, x means the content of magnesium phosphate cement wt%) as matrix model. The results indicated that the MCPC, especially 5MCPC, could promote the rhBMP-2-induced in vitro osteogenic differentiation via Smad signaling of C2C12 cells. Further studies demonstrated that all MCPC substrates exhibited similar rhBMP-2 release rate and preserved comparable conformation and biological activity of the released rhBMP-2. Also, the ionic extracts of MCPC made little difference to the bioactivity of rhBMP-2, either in soluble or in matrix-bound forms. However, with the quartz crystal microbalance (QCM), we observed a noticeable enhancement of rhBMP-2 mass-uptake on 5MCPC as well as a better recognition of the bound rhBMP-2 to BMPR IA and BMPR II. In vivo results demonstrated a better bone regeneration capacity of 5MCPC/rhBMP-2. From the above, our results demonstrated that it was the Mg anchored on the underlying substrates that tailored the way of rhBMP-2 bound on MCPC, and thus facilitated the recognition of BMPRs to stimulate osteogenic differentiation. The study will guide the development of Mg-doped bioactive bone implants for tissue regeneration. Copyright © 2016 Elsevier B.V. All rights reserved.

Cyclooxygenase-2 promotes pulmonary intravascular macrophage accumulation by exacerbating BMP signaling in rat experimental hepatopulmonary syndrome.

PubMed

Liu, Chang; Gao, Jing; Chen, Bing; Chen, Lin; Belguise, Karine; Yu, Weifeng; Lu, Kaizhi; Wang, Xiaobo; Yi, Bin

2017-08-15

One central factor in hepatopulmonary syndrome (HPS) pathogenesis is intravascular accumulation of activated macrophages in small pulmonary arteries. However, molecular mechanism underlying the macrophage accumulation in HPS is unknown. In this study, we aimed to explore whether elevated COX-2 induces the Bone morphogenic protein-2 (BMP-2)/Crossveinless-2 (CV-2) imbalance and then activation of BMP signaling pathway promotes the macrophage accumulation in Common Bile Duct Ligation (CBDL) rat lung. The COX-2/PGE2 signaling activation, the BMP-2/CV-2 imbalance and the activation of Smad1 were evaluated in CBDL rat lung and in cultured pulmonary microvascular endothelial cells (PMVECs) under the HPS serum stimulation. The effects of Parecoxib (COX-2 inhibitor), BMP-2 and CV-2 recombinant proteins on 4-week CBDL rat lung were determined, respectively. The COX-2/PGE2 signaling pathway was activated in CBDL rat lung in vivo and PMVECs in vitro, which was due to the activation of NF-κB P65. The inhibition of COX-2 by Parecoxib reduced macrophage accumulation, decreased lung angiogenesis and improved HPS. Meanwhile, the CBDL rat lung secreted more BMP-2 but less CV-2, and the imbalance between BMP-2 and CV-2 exacerbated the BMP signaling activation thus promoting the macrophage accumulation and lung angiogenesis. The BMP-2/CV-2 imbalance is dependent on the COX-2/PGE2 signaling pathway, and thus the effects of this imbalance can be reversed by adminstration of Parecoxib. Our findings indicate that inhibition of COX-2 by parecoxib can improve the HPS through the repression of BMP signaling and macrophage accumulation. Copyright © 2017 Elsevier Inc. All rights reserved.

Importance of a diffusion-dominant small volume to activate cell-secreted soluble factor signaling in embryonic stem cell culture in microbioreactors: a mathematical model based study.

PubMed

Chowdhury, Mohammad Mahfuz; Fujii, Teruo; Sakai, Yasuyuki

2013-07-01

In our previous studies, we observed that cell-secreted BMP4 had a prominent influence on mouse embryonic stem cell (mESC) behaviors in a membrane-based two-chambered microbioreactor (MB), but not in a macro-scale culture (6-well plate/6WP). In this study, we investigated how the physical aspects of these cultures regulated BMP4 signaling by developing mathematical models of the cultures. The models estimated signaling activity in the cultures by considering size of the undifferentiated mESC colonies and their growth, diffusion of BMP4, and BMP4 trafficking process in the colonies. The models successfully depicted measured profile of BMP4 concentration in the culture medium which was two times higher in the MB than that in the 6WP during 5-day culture. The models estimated that, owing to the small volume and the membrane, cells were exposed to a higher BMP4 concentration in the top chamber of the MB than that in the 6WP culture. The higher concentration of BMP4 induced a higher concentration of BMP4-bound receptor in the colony in the MB than in the 6WP, thereby leading to the higher activation of BMP4 signaling in the MB. The models also predicted that the size of the MB, but not that of the 6WP, was suitable for maximizing BMP4 accumulation and upregulating its signaling. This study will be helpful in analyzing culture systems, designing microfluidic devices for controlling ESC or other cell behavior. Copyright © 2013 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.

Verifying critical control points for Phytophthora introduction into nurseries

Treesearch

N.K. Osterbauer; M. Lujan; G. McAninch; A. Trippe; S. Lane

2013-01-01

The Oregon Department of Agriculture implemented the Grower Assisted Inspection Program (GAIP) for nurseries in 2007. Participants in GAIP adopted best management practices (BMP) for five critical control points (CCP) (used containers, irrigation water, soil substrate, potting media, and incoming plants), where foliar Phytophthora can be introduced...

Injectable chitosan microparticles incorporating bone morphogenetic protein-7 for bone tissue regeneration

PubMed Central

Mantripragada, Venkata P.; Jayasuriya, Ambalangodage C.

2014-01-01

This study investigates the influence of the controlled release of bone morphogenetic protein 7 (BMP-7) from cross-linked chitosan microparticles on pre-osteoblasts (OB-6) in vitro. BMP-7 was incorporated into microparticles by encapsulation during the particle preparation and coating after particle preparation. Chitosan microparticles had an average diameter of 700 μm containing ~100 ng of BMP-7. The release study profile indicates that nearly 98% of the BMP-7 coated on the microparticles was released in a period of 18 days while only 36% of the BMP-7 encapsulated in the microparticles was released in the same time period. Cell attachment study indicated that the BMP-7 coated microparticles have many cells adhered on the microparticles in comparison with microparticles without growth factors on day 10. DNA assay indicated a statistical significant increase (p<0.05) in the amount of DNA obtained from BMP-7 encapsulated and coated microparticles in comparison with microparticles without any growth factors. A real time RT-PCR experiment was performed to determine the expression of a few osteoblast specific genes - Dlx5, runx2, osterix, osteopontin, osteocalcin, and bone sialoprotein. The results thus suggest that chitosan microparticles obtained by coacervation method are biocompatible and helps in improving the encapsulation efficiency of BMP-7. Also BMP-7 incorporated in the microparticles is being released in a controlled fashion to support attachment, proliferation and differentiation of pre-osteoblasts, thus acting as a good scaffold for bone tissue regeneration. PMID:24497318

Unique players in the BMP pathway: Small C-terminal domain phosphatases dephosphorylate Smad1 to attenuate BMP signaling

PubMed Central

Knockaert, Marie; Sapkota, Gopal; Alarcón, Claudio; Massagué, Joan; Brivanlou, Ali H.

2006-01-01

Smad transcription factors are key signal transducers for the TGF-β/bone morphogenetic protein (BMP) family of cytokines and morphogens. C-terminal serine phosphorylation by TGF-β and BMP membrane receptors drives Smads into the nucleus as transcriptional regulators. Dephosphorylation and recycling of activated Smads is an integral part of this process, which is critical for agonist sensing by the cell. However, the nuclear phosphatases involved have remained unknown. Here we provide functional, biochemical, and embryological evidence identifying the SCP (small C-terminal domain phosphatase) family of nuclear phosphatases as mediators of Smad1 dephosphorylation in the BMP signaling pathway in vertebrates. Xenopus SCP2/Os4 inhibits BMP activity in the presumptive ectoderm and leads to neuralization. In Xenopus embryos, SCP2/Os4 and human SCP1, 2, and 3 cause selective dephosphorylation of Smad1 compared with Smad2, inhibiting BMP- and Smad1-dependent transcription and leading to the induction of the secondary dorsal axis. In human cells, RNAi-mediated depletion of SCP1 and SCP2 increases the extent and duration of Smad1 phosphorylation in response to BMP, the transcriptional action of Smad1, and the strength of endogenous BMP gene responses. The present identification of the SCP family as Smad C-terminal phosphatases sheds light on the events that attenuate Smad signaling and reveals unexpected links to the essential phosphatases that control RNA polymerase II in eukaryotes. PMID:16882717

Notch Signaling Augments BMP9-Induced Bone Formation by Promoting the Osteogenesis-Angiogenesis Coupling Process in Mesenchymal Stem Cells (MSCs).

PubMed

Liao, Junyi; Wei, Qiang; Zou, Yulong; Fan, Jiaming; Song, Dongzhe; Cui, Jing; Zhang, Wenwen; Zhu, Yunxiao; Ma, Chao; Hu, Xue; Qu, Xiangyang; Chen, Liqun; Yu, Xinyi; Zhang, Zhicai; Wang, Claire; Zhao, Chen; Zeng, Zongyue; Zhang, Ruyi; Yan, Shujuan; Wu, Tingting; Wu, Xingye; Shu, Yi; Lei, Jiayan; Li, Yasha; Luu, Hue H; Lee, Michael J; Reid, Russell R; Ameer, Guillermo A; Wolf, Jennifer Moriatis; He, Tong-Chuan; Huang, Wei

2017-01-01

Mesenchymal stem cells (MSCs) are multipotent progenitors that can differentiate into several lineages including bone. Successful bone formation requires osteogenesis and angiogenesis coupling of MSCs. Here, we investigate if simultaneous activation of BMP9 and Notch signaling yields effective osteogenesis-angiogenesis coupling in MSCs. Recently-characterized immortalized mouse adipose-derived progenitors (iMADs) were used as MSC source. Transgenes BMP9, NICD and dnNotch1 were expressed by adenoviral vectors. Gene expression was determined by qPCR and immunohistochem¡stry. Osteogenic activity was assessed by in vitro assays and in vivo ectopic bone formation model. BMP9 upregulated expression of Notch receptors and ligands in iMADs. Constitutively-active form of Notch1 NICD1 enhanced BMP9-induced osteogenic differentiation both in vitro and in vivo, which was effectively inhibited by dominant-negative form of Notch1 dnNotch1. BMP9- and NICD1-transduced MSCs implanted with a biocompatible scaffold yielded highly mature bone with extensive vascularization. NICD1 enhanced BMP9-induced expression of key angiogenic regulators in iMADs and Vegfa in ectopic bone, which was blunted by dnNotch1. Notch signaling may play an important role in BMP9-induced osteogenesis and angiogenesis. It's conceivable that simultaneous activation of the BMP9 and Notch pathways should efficiently couple osteogenesis and angiogenesis of MSCs for successful bone tissue engineering. © 2017 The Author(s)Published by S. Karger AG, Basel.

Efficacy of rhBMP-2 Loaded PCL/β-TCP/bdECM Scaffold Fabricated by 3D Printing Technology on Bone Regeneration.

PubMed

Bae, Eun-Bin; Park, Keun-Ho; Shim, Jin-Hyung; Chung, Ho-Yun; Choi, Jae-Won; Lee, Jin-Ju; Kim, Chang-Hwan; Jeon, Ho-Jun; Kang, Seong-Soo; Huh, Jung-Bo

2018-01-01

This study was undertaken to evaluate the effect of 3D printed polycaprolactone (PCL)/ β -tricalcium phosphate ( β -TCP) scaffold containing bone demineralized and decellularized extracellular matrix (bdECM) and human recombinant bone morphogenetic protein-2 (rhBMP-2) on bone regeneration. Scaffolds were divided into PCL/ β -TCP, PCL/ β -TCP/bdECM, and PCL/ β -TCP/bdECM/BMP groups. In vitro release kinetics of rhBMP-2 were determined with respect to cell proliferation and osteogenic differentiation. These three reconstructive materials were implanted into 8 mm diameter calvarial bone defect in male Sprague-Dawley rats. Animals were sacrificed four weeks after implantation for micro-CT, histologic, and histomorphometric analyses. The findings obtained were used to calculate new bone volumes (mm 3 ) and new bone areas (%). Excellent cell bioactivity was observed in the PCL/ β -TCP/bdECM and PCL/ β -TCP/bdECM/BMP groups, and new bone volume and area were significantly higher in the PCL/ β -TCP/bdECM/BMP group than in the other groups ( p < .05). Within the limitations of this study, bdECM printed PCL/ β -TCP scaffolds can reproduce microenvironment for cells and promote adhering and proliferating the cells onto scaffolds. Furthermore, in the rat calvarial defect model, the scaffold which printed rhBMP-2 loaded bdECM stably carries rhBMP-2 and enhances bone regeneration confirming the possibility of bdECM as rhBMP-2 carrier.

Cyst-Like Osteolytic Formations in Recombinant Human Bone Morphogenetic Protein-2 (rhBMP-2) Augmented Sheep Spinal Fusion.

PubMed

Pan, Hsin Chuan; Lee, Soonchul; Ting, Kang; Shen, Jia; Wang, Chenchao; Nguyen, Alan; Berthiaume, Emily A; Zara, Janette N; Turner, A Simon; Seim, Howard B; Kwak, Jin Hee; Zhang, Xinli; Soo, Chia

2017-07-01

Multiple case reports using recombinant human bone morphogenetic protein-2 (rhBMP-2) have reported complications. However, the local adverse effects of rhBMP-2 application are not well documented. In this report we show that, in addition to promoting lumbar spinal fusion through potent osteogenic effects, rhBMP-2 augmentation promotes local cyst-like osteolytic formations in sheep trabecular bones that have undergone anterior lumbar interbody fusion. Three months after operation, conventional computed tomography showed that the trabecular bones of the rhBMP-2 application groups could fuse, whereas no fusion was observed in the control group. Micro-computed tomography analysis revealed that the core implant area's bone volume fraction and bone mineral density increased proportionately with rhBMP-2 dose. Multiple cyst-like bone voids were observed in peri-implant areas when using rhBMP-2 applications, and these sites showed significant bone mineral density decreases in relation to the unaffected regions. Biomechanically, these areas decreased in strength by 32% in comparison with noncystic areas. Histologically, rhBMP-2-affected void sites had an increased amount of fatty marrow, thinner trabecular bones, and significantly more adiponectin- and cathepsin K-positive cells. Despite promoting successful fusion, rhBMP-2 use in clinical applications may result in local adverse structural alterations and compromised biomechanical changes to the bone. Copyright © 2017 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

BMP signaling balances proliferation and differentiation of muscle satellite cell descendants

PubMed Central

2011-01-01

Background The capacity of muscle to grow or to regenerate after damage is provided by adult stem cells, so called satellite cells, which are located under the basement lamina of each myofiber. Upon activation satellite cells enter the cell cycle, proliferate and differentiate into myoblasts, which fuse to injured myofibers or form new fibers. These processes are tightly controlled by many growth factors. Results Here we investigate the role of bone morphogenetic proteins (BMPs) during satellite cell differentiation. Unlike the myogenic C2C12 cell line, primary satellite cells do not differentiate into osteoblasts upon BMP signaling. Instead BMP signaling inhibits myogenic differentiation of primary satellite cells ex vivo. In contrast, inhibition of BMP signaling results in cell cycle exit, followed by enhanced myoblast differentiation and myotube formation. Using an in vivo trauma model we demonstrate that satellite cells respond to BMP signals during the regeneration process. Interestingly, we found the BMP inhibitor Chordin upregulated in primary satellite cell cultures and in regenerating muscles. In both systems Chordin expression follows that of Myogenin, a marker for cells committed to differentiation. Conclusion Our data indicate that BMP signaling plays a critical role in balancing proliferation and differentiation of activated satellite cells and their descendants. Initially, BMP signals maintain satellite cells descendants in a proliferating state thereby expanding cell numbers. After cells are committed to differentiate they upregulate the expression of the BMP inhibitor Chordin thereby supporting terminal differentiation and myotube formation in a negative feedback mechanism. PMID:21645366

A peptide that blocks the interaction of NF-κB p65 subunit with Smad4 enhances BMP2-induced osteogenesis.

PubMed

Urata, Mariko; Kokabu, Shoichiro; Matsubara, Takuma; Sugiyama, Goro; Nakatomi, Chihiro; Takeuchi, Hiroshi; Hirata-Tsuchiya, Shizu; Aoki, Kazuhiro; Tamura, Yukihiko; Moriyama, Yasuko; Ayukawa, Yasunori; Matsuda, Miho; Zhang, Min; Koyano, Kiyoshi; Kitamura, Chiaki; Jimi, Eijiro

2018-09-01

Bone morphogenetic protein (BMP) potentiates bone formation through the Smad signaling pathway in vitro and in vivo. The transcription factor nuclear factor κB (NF-κB) suppresses BMP-induced osteoblast differentiation. Recently, we identified that the transactivation (TA) 2 domain of p65, a main subunit of NF-κB, interacts with the mad homology (MH) 1 domain of Smad4 to inhibit BMP signaling. Therefore, we further attempted to identify the interacting regions of these two molecules at the amino acid level. We identified a region that we term the Smad4-binding domain (SBD), an amino-terminal region of TA2 that associates with the MH1 domain of Smad4. Cell-permeable SBD peptide blocked the association of p65 with Smad4 and enhanced BMP2-induced osteoblast differentiation and mineralization without affecting the phosphorylation of Smad1/5 or the activation of NF-κB signaling. SBD peptide enhanced the binding of the BMP2-inudced phosphorylated Smad1/5 on the promoter region of inhibitor of DNA binding 1 (Id-1) compared with control peptide. Although SBD peptide did not affect BMP2-induced chondrogenesis during ectopic bone formation, the peptide enhanced BMP2-induced ectopic bone formation in subcortical bone. Thus, the SBD peptide is useful for enabling BMP2-induced bone regeneration without inhibiting NF-κB activity. © 2018 Wiley Periodicals, Inc.

Dual role of BMP signaling in the regulation of Drosophila intestinal stem cell self-renewal.

PubMed

Tian, Aiguo; Jiang, Jin

2017-10-02

Many adult organs including Drosophila adult midguts rely on resident stem cells to replenish damaged cells during tissue homeostasis and regeneration. Previous studies have shown that, upon injury, intestinal stem cells (ISCs) in the midguts can increase proliferation and lineage differentiation to meet the demand for tissue repair. Our recent study has demonstrated that, in response to certain injury, midguts can expand ISC population size as an additional regenerative mechanism. We found that injury elicited by bleomycin feeding or bacterial infection increased the production of two BMP ligands (Dpp and Gbb) in enterocytes (ECs), leading to elevated BMP signaling in progenitor cells that drove an expansion of ISCs by promoting their symmetric self-renewing division. Interestingly, we also found that BMP signaling in ECs inhibits the production of Dpp and Gbb, and that this negative feedback mechanism is required to reset ISC pool size to the homeostatic state. Our findings suggest that BMP signaling exerts two opposing influences on stem cell activity depending on where it acts: BMP signaling in progenitor cells promotes ISC self-renewal while BMP signaling in ECs restricts ISC self-renewal by preventing excessive production of BMP ligands. Our results further suggest that transient expansion of ISC population in conjunction with increasing ISC proliferation provides a more effective strategy for tissue regeneration.

Evaluation of a thiolated chitosan scaffold for local delivery of BMP-2 for osteogenic differentiation and ectopic bone formation.

PubMed

Bae, In-Ho; Jeong, Byung-Chul; Kook, Min-Suk; Kim, Sun-Hun; Koh, Jeong-Tae

2013-01-01

Thiolated chitosan (Thio-CS) is a well-established pharmaceutical excipient for drug delivery. However, its use as a scaffold for bone formation has not been investigated. The aim of this study was to evaluate the potential of Thio-CS in bone morphogenetic protein-2 (BMP-2) delivery and bone formation. In vitro study showed that BMP-2 interacted with the Thio-CS and did not affect the swelling behavior. The release kinetics of BMP-2 from the Thio-CS was slightly delayed (70%) within 7 days compared with that from collagen gel (Col-gel, 85%), which is widely used in BMP-2 delivery. The BMP-2 released from Thio-CS increased osteoblastic cell differentiation but did not show any cytotoxicity until 21 days. Analysis of the in vivo ectopic bone formation at 4 weeks of posttransplantation showed that use of Thio-CS for BMP-2 delivery induced more bone formation to a greater extent (1.8 fold) than that of Col-gel. However, bone mineral density in both bones was equivalent, regardless of Thio-CS or Col-gel carrier. Taken together, Thio-CS system might be useful for delivering osteogenic protein BMP-2 and present a promising bone regeneration strategy.

Novel Wnt Regulator NEL-Like Molecule-1 Antagonizes Adipogenesis and Augments Osteogenesis Induced by Bone Morphogenetic Protein 2

PubMed Central

Shen, Jia; James, Aaron W.; Zhang, Xinli; Pang, Shen; Zara, Janette N.; Asatrian, Greg; Chiang, Michael; Lee, Min; Khadarian, Kevork; Nguyen, Alan; Lee, Kevin S.; Siu, Ronald K.; Tetradis, Sotirios; Ting, Kang; Soo, Chia

2017-01-01

The differentiation factor NEL-like molecule-1 (NELL-1) has been reported as osteoinductive in multiple in vivo preclinical models. Bone morphogenetic protein (BMP)-2 is used clinically for skeletal repair, but in vivo administration can induce abnormal, adipose-filled, poor-quality bone. We demonstrate that NELL-1 combined with BMP2 significantly optimizes osteogenesis in a rodent femoral segmental defect model by minimizing the formation of BMP2-induced adipose-filled cystlike bone. In vitro studies using the mouse bone marrow stromal cell line M2-10B4 and human primary bone marrow stromal cells have confirmed that NELL-1 enhances BMP2-induced osteogenesis and inhibits BMP2-induced adipogenesis. Importantly, the ability of NELL-1 to direct BMP2-treated cells toward osteogenesis and away from adipogenesis requires intact canonical Wnt signaling. Overall, these studies establish the feasibility of combining NELL-1 with BMP2 to improve clinical bone regeneration and provide mechanistic insight into canonical Wnt pathway activity during NELL-1 and BMP2 osteogenesis. The novel abilities of NELL-1 to stimulate Wnt signaling and to repress adipogenesis may highlight new treatment approaches for bone loss in osteoporosis. PMID:26772960

Arginine methylation of Smad7 by PRMT1 in TGF-β-induced epithelial-mesenchymal transition and epithelial stem cell generation.

PubMed

Katsuno, Yoko; Qin, Jian; Oses-Prieto, Juan A; Wang, Hongjun; Jackson-Weaver, Olan; Zhang, Tingwei; Lamouille, Samy; Wu, Jian; Burlingame, A L L; Xu, Jian; Derynck, Rik

2018-06-15

The epithelial-to-mesenchymal transdifferentiation (EMT) is crucial for tissue differentiation in development, and drives essential steps in cancer and fibrosis. EMT is accompanied by reprogramming of gene expression, and has been associated with the epithelial stem cell state in normal and carcinoma cells. The cytokine TGF-β drives this program in cooperation with other signaling pathways and through TGF-β-activated Smad3 as major effector. TGF-β-induced Smad3 activation is inhibited by Smad7 and to a lesser extent by Smad6, and Smad6 and Smad7 both inhibit Smad1 and Smad5 activation in response to the TGF-β-related bone morphogenetic proteins (BMPs). We previously reported that, in response to BMP, the protein arginine methyltransferase PRMT1 methylates Smad6 at the BMP receptor complex, thereby promoting its dissociation from the receptors and enabling BMP-induced Smad1 and Smad5 activation. We now provide evidence that PRMT1 also facilitates TGF-β signaling by methylating Smad7, which complements Smad6 methylation. We found that PRMT1 is required for TGF-β-induced Smad3 activation, through a mechanism similar to that of BMP-induced Smad6 methylation, and thus promotes the TGF-β-induced EMT and epithelial stem cell generation. This critical mechanism positions PRMT1 as an essential mediator of TGF-β signaling that controls the EMT and epithelial cell stemness through Smad7 methylation. Published under license by The American Society for Biochemistry and Molecular Biology, Inc.

Multiobjective Optimization Combining BMP Technology and Land Preservation for Watershed-based Stormwater Management

NASA Astrophysics Data System (ADS)

McGarity, A. E.

2009-12-01

Recent progress has been made developing decision-support models for optimal deployment of best management practices (BMP’s) in an urban watershed to achieve water quality goals. One example is the high-level screening model StormWISE, developed by the author (McGarity, 2006) that uses linear and nonlinear programming to narrow the search for optimal solutions to certain land use categories and drainage zones. Another example is the model SUSTAIN developed by USEPA and Tetra Tech (Lai, et al., 2006), which builds on the work of Yu, et al., 2002), that uses a detailed, computationally intensive simulation model driven by a genetic solver to select optimal BMP sites. However, a model that deals only with best management practice (BMP) site selections may fail to consider solutions that avoid future nonpoint pollutant loadings by preserving undeveloped land. This paper presents results of a recently completed research project in which water resource engineers partnered with experienced professionals at a land conservation trust to develop a multiobjective model for watershed management. The result is a revised version of StormWISE that can be used to identify optimal, cost-effective combinations of easements and similar land preservation tools for undeveloped sites along with low impact development (LID) and BMP technologies for developed sites. The goal is to achieve the watershed-wide limits on runoff volume and pollutant loads that are necessary to meet water quality goals as well as ecological benefits associated with habitat preservation and enhancement. A nonlinear programming formulation is presented for the extended StormWISE model that achieves desired levels of environmental benefits at minimum cost. Tradeoffs between different environmental benefits are generated by multiple runs of the model while varying the levels of each environmental benefit obtained. The model is solved using piecewise linearization of environmental benefit functions where each linear segment of represents a different option for reducing stormwater runoff volumes and pollutant loadings. The solutions space is comprised of optimal levels of expenditure for categories of BMP's by land use category and optimal land preservation expenditures by drainage zone. To demonstrate the usefulness of the model, results from its application to the Little Crum Creek watershed in suburban Philadelphia are presented. The model has been used to assist a watershed association and four municipalities to develop an action plan for restoration of water quality on this impaired stream. References Lai, F., J. Zhen, J. Riverson, and L. Shoemaker (2006). "SUSTAIN - An Evaluation and Cost-Optimization Tool for Placement of BMPs," ASCE World Environmental and Water Resource Congress 2006. McGarity, A.E. (2006). A Cost Minimization Model to Priortize Urban Catchments for Stormwater BMP Implementation Projects. American Water Resources Association National Meeting, Baltimore, MD, November, 2006. Yu, S., J. X. Zhen, and S.Y. Zhai, (2002). Development of Stormwater Best Management Practice Placement Strategy for the Virginia Department of Transportation. Final Contract Report, VTRC 04-CR9, Virginia Transportation Research Council.

Orphan nuclear receptor TLX regulates astrogenesis by modulating BMP signaling

PubMed Central

Qin, Song; Niu, Wenze; Iqbal, Nida; Smith, Derek K.; Zhang, Chun-Li

2014-01-01

Neural stem cells (NSCs) are self-renewing multipotent progenitors that generate both neurons and glia. The precise control of NSC behavior is fundamental to the architecture and function of the central nervous system. We previously demonstrated that the orphan nuclear receptor TLX is required for postnatal NSC activation and neurogenesis in the neurogenic niche. Here, we show that TLX modulates bone morphogenetic protein (BMP)-SMAD signaling to control the timing of postnatal astrogenesis. Genes involved in the BMP signaling pathway, such as Bmp4, Hes1, and Id3, are upregulated in postnatal brains lacking Tlx. Chromatin immunoprecipitation and electrophoretic mobility shift assays reveal that TLX can directly bind the enhancer region of Bmp4. In accordance with elevated BMP signaling, the downstream effectors SMAD1/5/8 are activated by phosphorylation in Tlx mutant mice. Consequently, Tlx mutant brains exhibit an early appearance and increased number of astrocytes with marker expression of glial fibrillary acidic protein (GFAP) and S100B. Taken together, these results suggest that TLX tightly controls postnatal astrogenesis through the modulation of BMP-SMAD signaling pathway activity. PMID:24782704

Orphan nuclear receptor TLX regulates astrogenesis by modulating BMP signaling.

PubMed

Qin, Song; Niu, Wenze; Iqbal, Nida; Smith, Derek K; Zhang, Chun-Li

2014-01-01

Neural stem cells (NSCs) are self-renewing multipotent progenitors that generate both neurons and glia. The precise control of NSC behavior is fundamental to the architecture and function of the central nervous system. We previously demonstrated that the orphan nuclear receptor TLX is required for postnatal NSC activation and neurogenesis in the neurogenic niche. Here, we show that TLX modulates bone morphogenetic protein (BMP)-SMAD signaling to control the timing of postnatal astrogenesis. Genes involved in the BMP signaling pathway, such as Bmp4, Hes1, and Id3, are upregulated in postnatal brains lacking Tlx. Chromatin immunoprecipitation and electrophoretic mobility shift assays reveal that TLX can directly bind the enhancer region of Bmp4. In accordance with elevated BMP signaling, the downstream effectors SMAD1/5/8 are activated by phosphorylation in Tlx mutant mice. Consequently, Tlx mutant brains exhibit an early appearance and increased number of astrocytes with marker expression of glial fibrillary acidic protein (GFAP) and S100B. Taken together, these results suggest that TLX tightly controls postnatal astrogenesis through the modulation of BMP-SMAD signaling pathway activity.

The bone morphogenetic protein axis is a positive regulator of skeletal muscle mass

PubMed Central

Chen, Justin L.; Qian, Hongwei; Liu, Yingying; Bernardo, Bianca C.; Beyer, Claudia; Watt, Kevin I.; Thomson, Rachel E.; Connor, Timothy; Turner, Bradley J.; McMullen, Julie R.; Larsson, Lars; McGee, Sean L.; Harrison, Craig A.

2013-01-01

Although the canonical transforming growth factor β signaling pathway represses skeletal muscle growth and promotes muscle wasting, a role in muscle for the parallel bone morphogenetic protein (BMP) signaling pathway has not been defined. We report, for the first time, that the BMP pathway is a positive regulator of muscle mass. Increasing the expression of BMP7 or the activity of BMP receptors in muscles induced hypertrophy that was dependent on Smad1/5-mediated activation of mTOR signaling. In agreement, we observed that BMP signaling is augmented in models of muscle growth. Importantly, stimulation of BMP signaling is essential for conservation of muscle mass after disruption of the neuromuscular junction. Inhibiting the phosphorylation of Smad1/5 exacerbated denervation-induced muscle atrophy via an HDAC4-myogenin–dependent process, whereas increased BMP–Smad1/5 activity protected muscles from denervation-induced wasting. Our studies highlight a novel role for the BMP signaling pathway in promoting muscle growth and inhibiting muscle wasting, which may have significant implications for the development of therapeutics for neuromuscular disorders. PMID:24145169

Expression of bone morphogenetic proteins and Msx genes during root formation.

PubMed

Yamashiro, T; Tummers, M; Thesleff, I

2003-03-01

Like crown development, root formation is also regulated by interactions between epithelial and mesenchymml tissues. Bone morphogenetic proteins (BMPs), together with the transcription factors Msx1 and Msx2, play important roles in these interactions during early tooth morphogenesis. To investigate the involvement of this signaling pathway in root development, we analyzed the expression patterns of Bmp2, Bmp3, Bmp4, and Bmp7 as well as Msx1 and Msx2 in the roots of mouse molars. Bmp4 was expressed in the apical mesenchyme and Msx2 in the root sheath. However, Bmps were not detected in the root sheath epithelium, and Msx transcripts were absent from the underlying mesenchyme. These findings indicate that this Bmp signaling pathway, required for tooth initiation, does not regulate root development, but we suggest that root shape may be regulated by a mechanism similar to that regulating crown shape in cap-stage tooth germs. Msx2 expression continued in the epithelial cell rests of Malassez, and the nearby cementoblasts intensely expressed Bmp3, which may regulate some functions of the fragmented epithelium.

Regulators and effectors of bone morphogenetic protein signalling in the cardiovascular system.

PubMed

Luo, Jiang-Yun; Zhang, Yang; Wang, Li; Huang, Yu

2015-07-15

Bone morphogenetic proteins (BMPs) play key roles in the regulation of cell proliferation, differentiation and apoptosis in various tissues and organs, including the cardiovascular system. BMPs signal through both Smad-dependent and -independent cascades to exert a wide spectrum of biological activities. Cardiovascular disorders such as abnormal angiogenesis, atherosclerosis, pulmonary hypertension and cardiac hypertrophy have been linked to aberrant BMP signalling. To correct the dysregulated BMP signalling in cardiovascular pathogenesis, it is essential to get a better understanding of how the regulators and effectors of BMP signalling control cardiovascular function and how the dysregulated BMP signalling contributes to cardiovascular dysfunction. We hence highlight several key regulators of BMP signalling such as extracellular regulators of ligands, mechanical forces, microRNAs and small molecule drugs as well as typical BMP effectors like direct downstream target genes, mitogen-activated protein kinases, reactive oxygen species and microRNAs. The insights into these molecular processes will help target both the regulators and important effectors to reverse BMP-associated cardiovascular pathogenesis. © 2015 The Authors. The Journal of Physiology © 2015 The Physiological Society.

Increased bone morphogenetic protein signaling contributes to age-related declines in neurogenesis and cognition.

PubMed

Meyers, Emily A; Gobeske, Kevin T; Bond, Allison M; Jarrett, Jennifer C; Peng, Chian-Yu; Kessler, John A

2016-02-01

Aging is associated with decreased neurogenesis in the hippocampus and diminished hippocampus-dependent cognitive functions. Expression of bone morphogenetic protein 4 (BMP4) increases with age by more than 10-fold in the mouse dentate gyrus while levels of the BMP inhibitor, noggin, decrease. This results in a profound 30-fold increase in phosphorylated-SMAD1/5/8, the effector of canonical BMP signaling. Just as observed in mice, a profound increase in expression of BMP4 is observed in the dentate gyrus of humans with no known cognitive abnormalities. Inhibition of BMP signaling either by overexpression of noggin or transgenic manipulation not only increases neurogenesis in aging mice, but remarkably, is associated with a rescue of cognitive deficits to levels comparable to young mice. Additive benefits are observed when combining inhibition of BMP signaling and environmental enrichment. These findings indicate that increased BMP signaling contributes significantly to impairments in neurogenesis and to cognitive decline associated with aging, and identify this pathway as a potential druggable target for reversing age-related changes in cognition. Copyright © 2016 Elsevier Inc. All rights reserved.

Effectiveness of three best management practices for highway-runoff quality along the Southeast Expressway, Boston, Massachusetts

USGS Publications Warehouse

Smith, Kirk P.

2002-01-01

Best management practices (BMPs) near highways are designed to reduce the amount of suspended sediment and associated constituents, including debris and litter, discharged from the roadway surface. The effectiveness of a deep-sumped hooded catch basin, three 2-chambered 1,500-gallon oil-grit separators, and mechanized street sweeping in reducing sediment and associated constituents was examined along the Southeast Expressway (Interstate Route 93) in Boston, Massachusetts. Repeated observations of the volume and distribution of bottom material in the oil-grit separators, including data on particle-size distributions, were compared to data from bottom material deposited during the initial 3 years of operation. The performance of catch-basin hoods and the oil-grit separators in reducing floating debris was assessed by examining the quantity of material retained by each structural BMP compared to the quantity of material retained by and discharged from the oil-grit separators, which received flow from the catch basins. The ability of each structural BMP to reduce suspended-sediment loads was assessed by examining (a) the difference in the concentrations of suspended sediment in samples collected simultaneously from the inlet and outlet of each BMP, and (b) the difference between inlet loads and outlet loads during a 14-month monitoring period for the catch basin and one separator, and a 10-month monitoring period for the second separator. The third separator was not monitored continuously; instead, samples were collected from it during three visits separated in time by several months. Suspended-sediment loads for the entire study area were estimated on the basis of the long-term average annual precipitation and the estimated inlet and outlet loads of two of the separators. The effects of mechanized street sweeping were assessed by evaluating the differences between suspended-sediment loads before and after street sweeping, relative to storm precipitation totals, and by comparing the particle-size distributions of sediment samples collected from the sweepers to bottom-material samples collected from the structural BMPs. A mass-balance calculation was used to quantify the accuracy of the estimated sediment-removal efficiency for each structural BMP. The ability of each structural BMP to reduce concentrations of inorganic and organic constituents was assessed by determining the differences in concentrations between the inlets and outlets of the BMPs for four storms. The inlet flows of the separators were sampled during five storms for analysis of fecal-indicator bacteria. The particle-size distribution of bottom material found in the first and second chambers of the separators was similar for all three separators. Consistent collection of floatable debris at the outlet of one separator during 12 storms suggests that floatable debris were not indefinitely retained.Concentrations of suspended sediment in discrete samples of runoff collected from the inlets of the two separators ranged from 8.5 to 7,110 mg/L. Concentrations of suspended sediment in discrete samples of runoff collected from the outlets of the separators ranged from 5 to 2,170 mg/L. The 14-month sediment-removal efficiency was 35 percent for one separator, and 28 percent for the second separator. In the combined-treatment system in this study, where catch basins provided primary suspended-sediment treatment, the separators reduced the mass of the suspended sediment from the pavement by about an additional 18 percent. The concentrations of suspended sediment in discrete samples of runoff collected from the inlet of the catch basin ranged from 32 to 13,600 mg/L. Concentrations of suspended sediment in discrete samples of runoff collected from the outlet of the catch basin ranged from 25.7 to 7,030 mg/L. The sediment-removal efficiency for individual storms during the 14-month monitoring period for the deep-sumped hooded catch basin was 39 percent.The concentrations of 29 in

Enhancement of Tendon–Bone Healing for Anterior Cruciate Ligament (ACL) Reconstruction Using Bone Marrow-Derived Mesenchymal Stem Cells Infected with BMP-2

PubMed Central

Dong, Yu; Zhang, Qingguo; Li, Yunxia; Jiang, Jia; Chen, Shiyi

2012-01-01

At present, due to the growing attention focused on the issue of tendon–bone healing, we carried out an animal study of the use of genetic intervention combined with cell transplantation for the promotion of this process. Here, the efficacy of bone marrow stromal cells infected with bone morphogenetic protein-2 (BMP-2) on tendon–bone healing was determined. A eukaryotic expression vector containing the BMP-2 gene was constructed and bone marrow-derived mesenchymal stem cells (bMSCs) were infected with a lentivirus. Next, we examined the viability of the infected cells and the mRNA and protein levels of BMP-2-infected bMSCs. Gastrocnemius tendons, gastrocnemius tendons wrapped by bMSCs infected with the control virus (bMSCs+Lv-Control), and gastrocnemius tendons wrapped by bMSCs infected with the recombinant BMP-2 virus (bMSCs+Lv-BMP-2) were used to reconstruct the anterior cruciate ligament (ACL) in New Zealand white rabbits. Specimens from each group were harvested four and eight weeks postoperatively and evaluated using biomechanical and histological methods. The bMSCs were infected with the lentivirus at an efficiency close to 100%. The BMP-2 mRNA and protein levels in bMSCs were significantly increased after lentiviral infection. The bMSCs and BMP-2-infected bMSCs on the gastrocnemius tendon improved the biomechanical properties of the graft in the bone tunnel; specifically, bMSCs infected with BMP-2 had a positive effect on tendon–bone healing. In the four-week and eight-week groups, bMSCs+Lv-BMP-2 group exhibited significantly higher maximum loads of 29.3 ± 7.4 N and 45.5 ± 11.9 N, respectively, compared with the control group (19.9 ± 6.4 N and 21.9 ± 4.9 N) (P = 0.041 and P = 0.001, respectively). In the eight-week groups, the stiffness of the bMSCs+Lv-BMP-2 group (32.5 ± 7.3) was significantly higher than that of the bMSCs+Lv-Control group (22.8 ± 7.4) or control groups (12.4 ± 6.0) (p = 0.036 and 0.001, respectively). Based on the histological findings, there was an increased amount of perpendicular collagen fibers formed between the tendon and bone in the bMSCs+Lv-Control and bMSCs+Lv-BMP-2 group, compared with the gastrocnemius tendons. The proliferation of cartilage-like cells and the formation of fibrocartilage-like tissue were highest within the bone tunnels in the bMSCs+Lv-BMP-2 group. These results suggest that this lentivirus can be used to efficiently infect bMSCs with BMP-2. Furthermore, tendons wrapped by bMSCs+Lv-BMP-2 improved tendon–bone healing. PMID:23202970

Developmentally inspired programming of adult human mesenchymal stromal cells toward stable chondrogenesis.

PubMed

Occhetta, Paola; Pigeot, Sebastien; Rasponi, Marco; Dasen, Boris; Mehrkens, Arne; Ullrich, Thomas; Kramer, Ina; Guth-Gundel, Sabine; Barbero, Andrea; Martin, Ivan

2018-05-01

It is generally accepted that adult human bone marrow-derived mesenchymal stromal cells (hMSCs) are default committed toward osteogenesis. Even when induced to chondrogenesis, hMSCs typically form hypertrophic cartilage that undergoes endochondral ossification. Because embryonic mesenchyme is obviously competent to generate phenotypically stable cartilage, it is questioned whether there is a correspondence between mesenchymal progenitor compartments during development and in adulthood. Here we tested whether forcing specific early events of articular cartilage development can program hMSC fate toward stable chondrogenesis. Inspired by recent findings that spatial restriction of bone morphogenetic protein (BMP) signaling guides embryonic progenitors toward articular cartilage formation, we hypothesized that selective inhibition of BMP drives the phenotypic stability of hMSC-derived chondrocytes. Two BMP type I receptor-biased kinase inhibitors were screened in a microfluidic platform for their time- and dose-dependent effect on hMSC chondrogenesis. The different receptor selectivity profile of tested compounds allowed demonstration that transient blockade of both ALK2 and ALK3 receptors, while permissive to hMSC cartilage formation, is necessary and sufficient to maintain a stable chondrocyte phenotype. Remarkably, even upon compound removal, hMSCs were no longer competent to undergo hypertrophy in vitro and endochondral ossification in vivo, indicating the onset of a constitutive change. Our findings demonstrate that adult hMSCs effectively share properties of embryonic mesenchyme in the formation of transient but also of stable cartilage. This opens potential pharmacological strategies to articular cartilage regeneration and more broadly indicates the relevance of developmentally inspired protocols to control the fate of adult progenitor cell systems.

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Science.gov Websites

Crops Conservation Agriculture Systems Alliance Biotechnology/Sustainability Know Your Watershed technology for conservation agriculture. Celebrates producer achievements. Encourage the formation of local Survey BMP Survey Form BMP Survey Results BMP News Release Conservation Agriculture Systems Alliance CASA

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wu, Li-An; Department of Pediatric Dentistry, School of Stomatology, The Fourth Military Medical University, Xi-an; Yuan, Guohua

Generation of a floxed Bmp2/4 osteoblast cell line is a valuable tool for studying the modulatory effects of Bmp2 and Bmp4 on osteoblast differentiation as well as relevant molecular events. In this study, primary floxed Bmp2/4 mouse osteoblasts were cultured and transfected with simian virus 40 large T-antigen. Transfection was verified by polymerase chain reaction (PCR) and immunohistochemistry. To examine the characteristics of the transfected cells, morphology, proliferation and mineralization were analyzed, expression of cell-specific genes including Runx2, ATF4, Dlx3, Osx, dentin matrix protein 1, bone sialoprotein, osteopontin, osteocalcin, osteonectin and collagen type I was detected. These results show thatmore » transfected floxed Bmp2/4 osteoblasts bypassed senescence with a higher proliferation rate, but retain the genotypic and phenotypic characteristics similar to the primary cells. Thus, we for the first time demonstrate the establishment of an immortalized mouse floxed Bmp2/4 osteoblast cell line.« less

Two Variants of Recombinant Human Bone Morphogenetic Protein-2 (rhBMP-2) with Additional Protein Domains: Synthesis in an Escherichia coli Heterologous Expression System.

PubMed

Karyagina, A S; Boksha, I S; Grunina, T M; Demidenko, A V; Poponova, M S; Sergienko, O V; Lyashchuk, A M; Galushkina, Z M; Soboleva, L A; Osidak, E O; Bartov, M S; Gromov, A V; Lunin, V G

2017-05-01

Two variants of recombinant human bone morphogenetic protein-2 (rhBMP-2) with additional N-terminal protein domains were obtained by expression in E. coli. The N-terminal domains were s-tag (15-a.a. oligopeptide from bovine pancreatic ribonuclease A) and lz (leucine zipper dimerization domain from yeast transcription factor GCN4). The s-tag-BMP-2 and lz-BMP-2 were purified by a procedure that excluded a long refolding stage. The resulting dimeric proteins displayed higher solubility compared to rhBMP-2 without additional protein domains. Biological activity of both proteins was demonstrated in vitro by induction of alkaline phosphatase in C2C12 cells, and the activity of s-tag-BMP-2 in vivo was shown in various experimental animal models.

BMP signaling restricts hemato-vascular development from lateral mesoderm during somitogenesis.

PubMed

Gupta, Sunny; Zhu, Hao; Zon, Leonard I; Evans, Todd

2006-06-01

The bone morphogenetic protein (BMP) signaling pathway is essential during gastrulation for the generation of ventral mesoderm, which makes it a challenge to define functions for this pathway at later stages of development. We have established an approach to disrupt BMP signaling specifically in lateral mesoderm during somitogenesis, by targeting a dominant-negative BMP receptor to Lmo2+ cells in developing zebrafish embryos. This results in expansion of hematopoietic and endothelial cells, while restricting the expression domain of the pronephric marker pax2.1. Expression of a constitutively active receptor and transplantation experiments were used to confirm that BMP signaling in lateral mesoderm restricts subsequent hemato-vascular development. The results show that the BMP signaling pathway continues to function after cells are committed to a lateral mesoderm fate, and influences subsequent lineage decisions by restricting hemato-vascular fate in favor of pronephric development.

Protein-protein recognition control by modulating electrostatic interactions.

PubMed

Han, Song; Yin, Shijin; Yi, Hong; Mouhat, Stéphanie; Qiu, Su; Cao, Zhijian; Sabatier, Jean-Marc; Wu, Yingliang; Li, Wenxin

2010-06-04

Protein-protein control recognition remains a huge challenge, and its development depends on understanding the chemical and biological mechanisms by which these interactions occur. Here we describe a protein-protein control recognition technique based on the dominant electrostatic interactions occurring between the proteins. We designed a potassium channel inhibitor, BmP05-T, that was 90.32% identical to wild-type BmP05. Negatively charged residues were translocated from the nonbinding interface to the binding interface of BmP05 inhibitor, such that BmP05-T now used BmP05 nonbinding interface as the binding interface. This switch demonstrated that nonbinding interfaces were able to control the orientation of protein binding interfaces in the process of protein-protein recognition. The novel function findings of BmP05-T peptide suggested that the control recognition technique described here had the potential for use in designing and utilizing functional proteins in many biological scenarios.

Production of BMP4 by endothelial cells is crucial for endogenous thymic regeneration

PubMed Central

Wertheimer, Tobias; Velardi, Enrico; Tsai, Jennifer; Cooper, Kirsten; Xiao, Shiyun; Kloss, Christopher C.; Ottmüller, Katja J.; Mokhtari, Zeinab; Brede, Christian; deRoos, Paul; Kinsella, Sinéad; Palikuqi, Brisa; Ginsberg, Michael; Young, Lauren F.; Kreines, Fabiana; Lieberman, Sophia R.; Lazrak, Amina; Guo, Peipei; Malard, Florent; Smith, Odette M.; Shono, Yusuke; Jenq, Robert R.; Hanash, Alan M.; Nolan, Daniel J.; Butler, Jason M.; Beilhack, Andreas; Manley, Nancy R.; Rafii, Shahin; Dudakov, Jarrod A; van den Brink, Marcel RM

2018-01-01

The thymus is extremely sensitive to damage but also has a remarkable ability to repair itself. However, the mechanisms underlying this endogenous regeneration remain poorly understood and this capacity diminishes considerably with age. Here we show that thymic endothelial cells (ECs) comprise a critical pathway of regeneration, via their production of BMP4. ECs increased their production of BMP4 after thymic damage, and abrogating BMP4 signalling or production by either pharmacologic or genetic inhibition impaired thymic repair. EC-derived BMP4 acted on thymic epithelial cells (TECs) to increase their expression of Foxn1, a key transcription factor involved in TEC development, maintenance and regeneration; and its downstream targets such as Dll4, itself a key mediator of thymocyte development and regeneration. These studies demonstrate the importance of the BMP4 pathway in endogenous tissue regeneration and offer a potential clinical approach to enhance T cell immunity. PMID:29330161

Regulation of Msx genes by a Bmp gradient is essential for neural crest specification.

PubMed

Tribulo, Celeste; Aybar, Manuel J; Nguyen, Vu H; Mullins, Mary C; Mayor, Roberto

2003-12-01

There is evidence in Xenopus and zebrafish embryos that the neural crest/neural folds are specified at the border of the neural plate by a precise threshold concentration of a Bmp gradient. In order to understand the molecular mechanism by which a gradient of Bmp is able to specify the neural crest, we analyzed how the expression of Bmp targets, the Msx genes, is regulated and the role that Msx genes has in neural crest specification. As Msx genes are directly downstream of Bmp, we analyzed Msx gene expression after experimental modification in the level of Bmp activity by grafting a bead soaked with noggin into Xenopus embryos, by expressing in the ectoderm a dominant-negative Bmp4 or Bmp receptor in Xenopus and zebrafish embryos, and also through Bmp pathway component mutants in the zebrafish. All the results show that a reduction in the level of Bmp activity leads to an increase in the expression of Msx genes in the neural plate border. Interestingly, by reaching different levels of Bmp activity in animal cap ectoderm, we show that a specific concentration of Bmp induces msx1 expression to a level similar to that required to induce neural crest. Our results indicate that an intermediate level of Bmp activity specifies the expression of Msx genes in the neural fold region. In addition, we have analyzed the role that msx1 plays on neural crest specification. As msx1 has a role in dorsoventral pattering, we have carried out conditional gain- and loss-of-function experiments using different msx1 constructs fused to a glucocorticoid receptor element to avoid an early effect of this factor. We show that msx1 expression is able to induce all other early neural crest markers tested (snail, slug, foxd3) at the time of neural crest specification. Furthermore, the expression of a dominant negative of Msx genes leads to the inhibition of all the neural crest markers analyzed. It has been previously shown that snail is one of the earliest genes acting in the neural crest genetic cascade. In order to study the hierarchical relationship between msx1 and snail/slug we performed several rescue experiments using dominant negatives for these genes. The rescuing activity by snail and slug on neural crest development of the msx1 dominant negative, together with the inability of msx1 to rescue the dominant negatives of slug and snail strongly argue that msx1 is upstream of snail and slug in the genetic cascade that specifies the neural crest in the ectoderm. We propose a model where a gradient of Bmp activity specifies the expression of Msx genes in the neural folds, and that this expression is essential for the early specification of the neural crest.

Retention of 125I-labeled recombinant human bone morphogenetic protein-2 by biphasic calcium phosphate or a composite sponge in a rabbit posterolateral spine arthrodesis model.

PubMed

Louis-Ugbo, John; Kim, Hak-Sun; Boden, Scott D; Mayr, Matthew T; Li, Ronald C; Seeherman, Howard; D'Augusta, Darren; Blake, Cara; Jiao, Aiping; Peckham, Steve

2002-09-01

The purpose of this study was to characterize the retention kinetics of recombinant human bone morphogenetic protein-2 (rhBMP-2) applied to two calcium-based delivery matrices. Biphasic calcium phosphate (BCP) and a composite containing BCP in an absorbable collagen sponge (BCP/ACS) were evaluated using a spinal fusion model in rabbits. rhBMP-2 labeled with radioactive iodine (125I) was used as a tracer to assess in vivo retention of rhBMP-2 in the presence of these materials (nine animals per material studied). Over a 36 day study period, animals were assessed for the following: percent administered dose retained at the implant site as measured by scintigraphic imaging (counting) with a gamma camera (all animals), radiography of the implant site (all animals), radioactivity in blood and plasma (all animals), and radioactivity in the urine and feces (three animals for each material). Radioactivity data were corrected for the decay of 125I and the attenuation between the implant in vivo and the gamma camera. Differences observed between the two materials for the area under the retention vs. time profile (AUC; 988%*day for BCP vs. 1070%*day for BCP/ACS, p = 0.57) and the mean residence time (MRT; 10.2 days for BCP vs. 7.6 days for BCP/ACS, p = 0.06) were not statistically significant. Initial retention/incorporation of rhBMP-2 was slightly higher for rhBMP-2/BCP/ACS than for rhBMP-2/BCP (96.8% vs. 86.0%, p < 0.05). Animals receiving rhBMP-2/BCP showed a longer terminal retention half-life (t1/2) than did those receiving rhBMP-2/BCP/ACS (7.5 vs. 4.5 days, p < 0.05). The urinary radioactivity recovery data supported the data obtained by scintigraphy. Over the 36 day collection period, essentially complete recovery of radioactivity (dose) in urine was observed for rhBMP-2/BCP and rhBMP-2/BCP/ACS and the majority of the radioactivity (approximately 95%) was soluble in trichloroacetic acid, suggesting extensive catabolism of rhBMP-2 before renal excretion. Fecal recovery of radioactivity was low, approximately 2-3%. In conclusion, rhBMP-2 was retained at the implant site when delivered with either BCP or BCP/ACS based on mean residence time and area under the retention curve vs. time profile. Use of these matrices resulted in detectable rhBMP-2 levels at the surgical site for over a week in contrast to data reported with several other matrices that lasted less time. Systemic catabolism and elimination of the rhBMP-2 was extensive and systemic presence of the protein was negligible.

Bone morphogenetic protein 4 antagonizes hair cell regeneration in the avian auditory epithelium.

PubMed

Lewis, Rebecca M; Keller, Jesse J; Wan, Liangcai; Stone, Jennifer S

2018-07-01

Permanent hearing loss is often a result of damage to cochlear hair cells, which mammals are unable to regenerate. Non-mammalian vertebrates such as birds replace damaged hair cells and restore hearing function, but mechanisms controlling regeneration are not understood. The secreted protein bone morphogenetic protein 4 (BMP4) regulates inner ear morphogenesis and hair cell development. To investigate mechanisms controlling hair cell regeneration in birds, we examined expression and function of BMP4 in the auditory epithelia (basilar papillae) of chickens of either sex after hair cell destruction by ototoxic antibiotics. In mature basilar papillae, BMP4 mRNA is highly expressed in hair cells, but not in hair cell progenitors (supporting cells). Supporting cells transcribe genes encoding receptors for BMP4 (BMPR1A, BMPR1B, and BMPR2) and effectors of BMP4 signaling (ID transcription factors). Following hair cell destruction, BMP4 transcripts are lost from the sensory epithelium. Using organotypic cultures, we demonstrate that treatments with BMP4 during hair cell destruction prevent supporting cells from upregulating expression of the pro-hair cell transcription factor ATOH1, entering the cell cycle, and fully transdifferentiating into hair cells, but they do not induce cell death. By contrast, noggin, a BMP4 inhibitor, increases numbers of regenerated hair cells. These findings demonstrate that BMP4 antagonizes hair cell regeneration in the chicken basilar papilla, at least in part by preventing accumulation of ATOH1 in hair cell precursors. Copyright © 2018 Elsevier B.V. All rights reserved.

Preliminary evaluation of a load-bearing BMP-2 carrier for segmental defect regeneration.

PubMed

Chu, Tien-Min G; Sargent, Peter; Warden, Stuart J; Turner, Charles H; Stewart, Rena L

2006-01-01

Large segmental defects in bones can result from tumor removal, massive trauma, congenital malformation, or non-union fractures. Such defects often are difficult to manage and require multiple-phase surgery to achieve adequate union and function. In this study, we propose a novel design of bone morphogenetic protein 2 (BMP-2) carrier for tissue engineering of segmental defect regeneration. The tube-shaped BMP-2 carrier was fabrication from a poly(propylene fumarate)/tricalcium phosphate (PPF/TCP) composite via casting technique developed in our laboratory. An in vitro evaluation showed that the compressive strength of the carrier decreased about 48% in 12 weeks while maintained a pH in the 6.8-7.4 range. In vivo study was conducted by implanting carriers loaded with 10 microg of BMP-2 in 5 mm rat femur gap model for 15 weeks. X-ray evidence of bridging was first found in the BMP group at 3 weeks. Bridging in all animals (N = 4) in the BMP group was found at 9 weeks. No x-ray evidence of bridging was found in the No BMP group (N = 3). pQCT analysis indicated that the bone mineral density of the callus in the BMP group has reached the level of native femur at 15 weeks after implantation, while the callus in the No BMP group has a bone mineral density at a lower level of 84% to the native femur. Histology analysis shows that a normal fatty bone marrow was restored and mineralized callus formed and bridged the segmental defect.

Collagen I derived recombinant protein microspheres as novel delivery vehicles for bone morphogenetic protein-2.

PubMed

Mumcuoglu, Didem; de Miguel, Laura; Jekhmane, Shehrazade; Siverino, Claudia; Nickel, Joachim; Mueller, Thomas D; van Leeuwen, Johannes P; van Osch, Gerjo J; Kluijtmans, Sebastiaan G

2018-03-01

Bone morphogenetic protein-2 (BMP-2) is a powerful osteoinductive protein; however, there is a need for the development of a safe and efficient BMP-2 release system for bone regeneration therapies. Recombinant extracellular matrix proteins are promising next generation biomaterials since the proteins are well-defined, reproducible and can be tailored for specific applications. In this study, we have developed a novel and versatile BMP-2 delivery system using microspheres from a recombinant protein based on human collagen I (RCP). In general, a two-phase release pattern was observed while the majority of BMP-2 was retained in the microspheres for at least two weeks. Among different parameters studied, the crosslinking and the size of the RCP microspheres changed the in vitro BMP-2 release kinetics significantly. Increasing the chemical crosslinking (hexamethylene diisocyanide) degree decreased the amount of initial burst release (24h) from 23% to 17%. Crosslinking by dehydrothermal treatment further decreased the burst release to 11%. Interestingly, the 50 and 72μm-sized spheres showed a significant decrease in the burst release compared to 207-μm sized spheres. Very importantly, using a reporter cell line, the released BMP-2 was shown to be bioactive. SPR data showed that N-terminal sequence of BMP-2 was important for the binding and retention of BMP-2 and suggested the presence of a specific binding epitope on RCP (K D : 1.2nM). This study demonstrated that the presented RCP microspheres are promising versatile BMP-2 delivery vehicles. Copyright © 2017 Elsevier B.V. All rights reserved.

The human chondrosarcoma HCS-2/8 cell line is responsive to BMP-7, but not to IL-1beta.

PubMed

Saas, Joachim; Gebauer, Matthias; Jacobi, Carsten; Haag, Jochen; Takigawa, Masaharu; Aigner, Thomas

2005-05-01

Cultures of primary chondrocytes as in vitro model systems for studying the cellular behavior of chondrocytes are notoriously difficult to cultivate and propagate. One way to circumvent these problems appears to be the use of immortalized/immortal chondrocytic cell lines. In the present study, we were interested whether the chondrosarcoma derived HCS-2/8 cells are suitable for studying major cellular reaction pattern in response to key anabolic (BMP-7) and catabolic (IL-1beta) factors. Therefore, we used cDNA array and real-time PCR technology in order to evaluate gene expression triggered by stimulation with IL-1beta (0,1-100 ng/ml) and BMP-7 in confluent monolayer cultures. HCS-2/8 cells hardly responded to IL-1beta, but showed good responsiveness to BMP-7. We found 12 genes up- and 17 significantly down-regulated by BMP-7 (out of 340 investigated genes). Besides the expected activation of anabolic genes chondrocytic cells after BMP-stimulation try to neutralize activation of the BMP-signalling cascade by expressing intra- and extracellular BMP-antagonists. Chondrosarcoma derived cell lines are a potential substitute for primary articular chondrocytes promising consistent expression of a differentiated chondrocyte phenotype with sufficient proliferative capacity. However, as shown by this study one needs to carefully select the cell line depending on the effects which one intends to study. In this respect, HCS-2/8 cells are a validated tool for studying BMP-effects on chondrocytes, but not e.g. effects of interleukin-1.

Thyroid Hormone-Induced Hypertrophy in Mesenchymal Stem Cell Chondrogenesis Is Mediated by Bone Morphogenetic Protein-4

PubMed Central

Karl, Alexandra; Olbrich, Norman; Pfeifer, Christian; Berner, Arne; Zellner, Johannes; Kujat, Richard; Angele, Peter; Nerlich, Michael

2014-01-01

Chondrogenic differentiating mesenchymal stem cells (MSCs) express markers of hypertrophic growth plate chondrocytes. As hypertrophic cartilage undergoes ossification, this is a concern for the application of MSCs in articular cartilage tissue engineering. To identify mechanisms that elicit this phenomenon, we used an in vitro hypertrophy model of chondrifying MSCs for differential gene expression analysis and functional experiments with the focus on bone morphogenetic protein (BMP) signaling. Hypertrophy was induced in chondrogenic MSC pellet cultures by transforming growth factor β (TGFβ) and dexamethasone withdrawal and addition of triiodothyronine. Differential gene expression analysis of BMPs and their receptors was performed. Based on these results, the in vitro hypertrophy model was used to investigate the effect of recombinant BMP4 and the BMP inhibitor Noggin. The enhancement of hypertrophy could be shown clearly by an increased cell size, alkaline phosphatase activity, and collagen type X deposition. Upon induction of hypertrophy, BMP4 and the BMP receptor 1B were upregulated. Addition of BMP4 further enhanced hypertrophy in the absence, but not in the presence of TGFβ and dexamethasone. Thyroid hormone induced hypertrophy by upregulation of BMP4 and this induced enhancement of hypertrophy could be blocked by the BMP antagonist Noggin. BMP signaling is an important modulator of the late differentiation stages in MSC chondrogenesis and the thyroid hormone induces this pathway. As cartilage tissue engineering constructs will be exposed to this factor in vivo, this study provides important insight into the biology of MSC-based cartilage. Furthermore, the possibility to engineer hypertrophic cartilage may be helpful for critical bone defect repair. PMID:23937304

Bone morphogenetic protein 2 and decorin expression in old fracture fragments and surrounding tissues.

PubMed

Han, X G; Wang, D K; Gao, F; Liu, R H; Bi, Z G

2015-09-21

Bone morphogenetic protein 2 (BMP-2) can promote fracture healing. Although the complex role BMP-2 in bone formation is increasingly understood, the role of endogenous BMP-2 in nonunion remains unclear. Decorin (DCN) can promote the formation of bone matrix and calcium deposition to control bone morphogenesis. In this study, tissue composition and expression of BMP-2 and DCN were detected in different parts of old fracture zones to explore inherent anti-fibrotic ability and osteogenesis. Twenty-three patients were selected, including eight cases of delayed union and 15 cases of nonunion. Average duration of delayed union or nonunion was 15 months. Fracture fragments and surrounding tissues, including bone grafts, marrow cavity contents, and sticking scars, were categorically sampled during surgery. Through observation and histological testing, component comparisons were made between fracture fragments and surrounding tissue. The expression levels of DCN and BMP-2 in different tissues were detected by immunohistochemical staining and real-time polymerase chain reaction. The expression of DCN and BMP- 2 in different parts of the nonunion area showed that, compared with bone graft and marrow cavity contents, sticking scars had the highest expression of BMP-2. Compared with the marrow cavity contents and sticking scars, bone grafts had the highest expression of DCN. The low antifibrotic and osteogenic activity of the nonunion area was associated with non-co-expression of BMP-2 and DCN. Therefore, the co-injection of osteogenic factor BMP and DCN into the nonunion area can improve the induction of bone formation and enhance the conversion of the old scar, thereby achieving better nonunion treatment.

BMP6 down-regulates GDNF expression through SMAD1/5 and ERK1/2 signaling pathways in human granulosa-lutein cells.

PubMed

Zhang, Xin-Yue; Chang, Hsun-Ming; Taylor, Elizabeth L; Leung, Peter C K; Liu, Rui-Zhi

2018-05-09

Bone morphogenetic protein 6 (BMP6) is a critical regulator of follicular development that is expressed in mammalian oocytes and granulosa cells. Glial cell line-derived neurotrophic factor (GDNF) is an intraovarian neurotrophic factor that plays an essential role in regulating mammalian oocyte maturation. The aim of this study was to investigate the effect of BMP6 on the regulation of GDNF expression and the potential underlying mechanisms. We used an established immortalized human granulosa cell line (SVOG cells) and primary human granulosa-lutein cells as in vitro cell models. Our results showed that BMP6 significantly down-regulated the expression of GDNF in both SVOG and primary human granulosa-lutein cells. Using dual inhibition approaches (kinase receptor inhibitor and small interfering RNA knockdown), our results showed that both ALK2 and ALK3 are involved in BMP6-induced down-regulation of GDNF. In addition, BMP6 induced the phosphorylation of SMAD1/5/8 and ERK1/2 but not AKT or p38. Among three downstream mediators, both SMAD1 and SMAD5 are involved in BMP6-induced down-regulation of GDNF. Moreover, concomitant knockdown of endogenous SMAD4 and inhibition of ERK1/2 activity completely reversed BMP6-induced down-regulation of GDNF, indicating that both SMAD and ERK1/2 signaling pathways are required for the regulatory effect of BMP6 on GDNF expression. Our findings suggest an additional role for an intrafollicular growth factor in regulating follicular function through their paracrine interactions in human granulosa cells.

Osseous regeneration in the rat calvarium using novel delivery systems for recombinant human bone morphogenetic protein-2 (rhBMP-2).

PubMed

Kenley, R; Marden, L; Turek, T; Jin, L; Ron, E; Hollinger, J O

1994-10-01

In the current investigation, we report osseous regeneration in critical-size rat calvarial defects using recombinant human bone morphogenetic protein-2 (rhBMP-2) and novel delivery systems based on biomaterials. The novel systems combine rhBMP-2 with dry powder microparticles of poly(D,L-lactide-co-glycolide) (PLGA). The mixture of rhBMP-2 with PLGA microparticles is added to an aqueous solution of biopolymer to yield a semisolid paste. The biopolymers tested include autologous blood clot, hydroxypropyl methylcellulose, and sodium alginate cross-linked with calcium ion. Insoluble collageneous bone matrix was also studied as a control. Test articles were made at 0-, 10-, and 30-micrograms doses of rhBMP-2 and imiplanted in 8-mm-diameter rat calvarial defects (which will not heal if left untreated). The animals were examined 21 days after implantation by radiography, radiomorphometry, histology, and histomorphometry. All tested materials containing rhBMP-2 restored radiopacity and normal contouring to the calvarial defects. Samples without added rhBMP-2 yielded only soft tissue within the defects. Histology showed restoration of inner and outer bone tables plus marrow constituents. The PLGA microparticles were significantly resorbed at the 21-day time point. Although small differences between delivery systems were evident at 0- and 10-micrograms rhBMP-2 doses, all test articles performed essentially equivalently at the 30-micrograms dose. Thus, novel delivery systems for rhBMP-2 offer the promise of combining the intrinsic bioactivity of the osteoinductive protein with pharmaceutically acceptable biomaterials.

Posterolateral lumbar fusion using Escherichia coli-derived rhBMP-2/hydroxyapatite in the mini pig.

PubMed

Kong, Chang-Bae; Lee, Jae Hyup; Baek, Hae-Ri; Lee, Choon-Ki; Chang, Bong-Soon

2014-12-01

Hydroxyapatite (HA) is used as a bone graft extender for posterolateral spinal fusion in human. It is also useful as a recombinant human bone morphogenetic protein (rhBMP)-2 carrier because of its high affinity for rhBMP-2. To assess the osteoinductivity of Escherichia coli-derived rhBMP-2 (E-BMP-2) using HA granules as a carrier and to evaluate the bone-forming ability depending on the different dosages of E-BMP-2. A mini-pig lumbar posterolateral fusion model using microcomputed tomography (μCT) scanning. Thirty-one adult male mini pigs were randomized into a single control group (n=8) without E-BMP-2 and two experimental groups with two different doses of E-BMP-2 (1 mg per side, n=8 and 3 mg per side, n=15). Outcome was measured by plain radiography, manual palpation, CT, three-dimensional μCT, and histologic examinations. Bilateral intertransverse process arthrodesis was performed, and E-BMP-2 (0, 1.0, 3.0 mg per side) was implanted into the intertransverse space using HA granules as a carrier. Three mini pigs were removed because of death. Among 28 experimental subjects, 19 animals achieved solid bony union. The fusion rates were 37.5% for control group, 71.4% for 1 mg group, and 84.6% for 3 mg group. Fusion rates were significantly different among groups (p=.031). However, there was no statistically significant difference in fusion rates between 1 and 3 mg groups (p=.587). Thirty-eight intertransverse fusion masses of 19 subjects underwent μCT scanning. The bone volumes determined by μCT were 12,603±3,240 mm(3) for control group, 18,718±3,000 mm(3) for 1 mg group, and 26,768±7,256 mm(3) for 3 mg group, and the difference between groups was statistically significant (p<.001). This study shows that E-BMP-2 has osteoinductive activity in dose-dependent fashion, and porous HA granule is suitable for E-BMP-2 carrier in a porcine posterolateral fusion model. These preliminary findings suggest that E-BMP-2-adsorbed porous HA granules could be a novel effective bone graft substitute. Copyright © 2014 Elsevier Inc. All rights reserved.

BmpR1A is a major type 1 BMP receptor for BMP-Smad signaling during skull development.

PubMed

Pan, Haichun; Zhang, Honghao; Abraham, Ponnu; Komatsu, Yoshihiro; Lyons, Karen; Kaartinen, Vesa; Mishina, Yuji

2017-09-01

Craniosynostosis is caused by premature fusion of one or more sutures in an infant skull, resulting in abnormal facial features. The molecular and cellular mechanisms by which genetic mutations cause craniosynostosis are incompletely characterized, and many of the causative genes for diverse types of syndromic craniosynostosis have not yet been identified. We previously demonstrated that augmentation of BMP signaling mediated by a constitutively active BMP type IA receptor (ca-BmpR1A) in neural crest cells (ca1A hereafter) causes craniosynostosis and superimposition of heterozygous null mutation of Bmpr1a rescues premature suture fusion (ca1A;1aH hereafter). In this study, we superimposed heterozygous null mutations of the other two BMP type I receptors, Bmpr1b and Acvr1 (ca1A;1bH and ca1A;AcH respectively hereafter) to further dissect involvement of BMP-Smad signaling. Unlike caA1;1aH, ca1A;1bH and ca1A;AcH did not restore the craniosynostosis phenotypes. In our in vivo study, Smad-dependent BMP signaling was decreased to normal levels in mut;1aH mice. However, BMP receptor-regulated Smads (R-Smads; pSmad1/5/9 hereafter) levels were comparable between ca1A, ca1A;1bH and ca1A;AcH mice, and elevated compared to control mice. Bmpr1a, Bmpr1b and Acvr1 null cells were used to examine potential mechanisms underlying the differences in ability of heterozygosity for Bmpr1a vs. Bmpr1b or Acvr1 to rescue the mut phenotype. pSmad1/5/9 level was undetectable in Bmpr1a homozygous null cells while pSmad1/5/9 levels did not decrease in Bmpr1b or Acvr1 homozygous null cells. Taken together, our study indicates that different levels of expression and subsequent activation of Smad signaling differentially contribute each BMP type I receptor to BMP-Smad signaling and craniofacial development. These results also suggest differential involvement of each type 1 receptor in pathogenesis of syndromic craniosynostoses. Copyright © 2017 Elsevier Inc. All rights reserved.

BMP2-loaded hollow hydroxyapatite microspheres exhibit enhanced osteoinduction and osteogenicity in large bone defects.

PubMed

Xiong, Long; Zeng, Jianhua; Yao, Aihua; Tu, Qiquan; Li, Jingtang; Yan, Liang; Tang, Zhiming

2015-01-01

The regeneration of large bone defects is an osteoinductive, osteoconductive, and osteogenic process that often requires a bone graft for support. Limitations associated with naturally autogenic or allogenic bone grafts have demonstrated the need for synthetic substitutes. The present study investigates the feasibility of using novel hollow hydroxyapatite microspheres as an osteoconductive matrix and a carrier for controlled local delivery of bone morphogenetic protein 2 (BMP2), a potent osteogenic inducer of bone regeneration. Hollow hydroxyapatite microspheres (100±25 μm) with a core (60±18 μm) and a mesoporous shell (180±42 m(2)/g surface area) were prepared by a glass conversion technique and loaded with recombinant human BMP2 (1 μg/mg). There was a gentle burst release of BMP2 from microspheres into the surrounding phosphate-buffered saline in vitro within the initial 48 hours, and continued at a low rate for over 40 days. In comparison with hollow hydroxyapatite microspheres without BMP2 or soluble BMP2 without a carrier, BMP2-loaded hollow hydroxyapatite microspheres had a significantly enhanced capacity to reconstitute radial bone defects in rabbit, as shown by increased serum alkaline phosphatase; quick and complete new bone formation within 12 weeks; and great biomechanical flexural strength. These results indicate that BMP2-loaded hollow hydroxyapatite microspheres could be a potential new option for bone graft substitutes in bone regeneration.

NOGGIN IS REQUIRED FOR NORMAL LOBE PATTERNING AND DUCTAL BUDDING IN THE MOUSE PROSTATE

PubMed Central

Cook, Crist; Vezina, Chad M.; Hicks, Sarah M.; Shaw, Aubie; Yu, Min; Peterson, Richard E.; Bushman, Wade

2008-01-01

Mesenchymal expression of the BMP antagonist NOGGIN during prostate development plays a critical role in pre-natal ventral prostate development and opposes BMP4-mediated inhibition of cell proliferation during postnatal ductal development. Morphologic examination of newborn Noggin-/- male fetuses revealed genitourinary anomalies including cryptorchidism, incomplete separation of the hindgut from the urogenital sinus (UGS), absence of the ventral mesenchymal pad and a complete loss of ventral prostate (VP) budding. Examination of lobe-specific marker expression in the E14 Noggin-/- UGS rescued by transplantation under the renal capsule of a male nude mouse confirmed a complete loss of VP determination. More modest effects were observed in the other lobes, including decreased number of ductal buds in the dorsal and lateral prostates of newborn Noggin-/- males. BMP4 and BMP7 have been shown to inhibit ductal budding and outgrowth by negatively regulating epithelial cell proliferation. We show here that NOGGIN can neutralize budding inhibition by BMP4 and rescues branching morphogenesis of BMP4-exposed UGS in organ culture and show that the effects of BMP4 and NOGGIN activities converge on P63+ epithelial cells located at nascent duct tips. Together, these studies show that the BMP-NOGGIN axis regulates patterning of the ventral prostate, regulates ductal budding, and controls proliferation of P63+ epithelial cells in the nascent ducts of developing mouse prostate. PMID:18028901

(−)-Epigallocatechin gallate but not chlorogenic acid upregulates osteoprotegerin synthesis through regulation of bone morphogenetic protein-4 in osteoblasts

PubMed Central

Fujita, Kazuhiko; Otsuka, Takanobu; Yamamoto, Naohiro; Kainuma, Shingo; Ohguchi, Reou; Kawabata, Tetsu; Sakai, Go; Kuroyanagi, Gen; Matsushima-Nishiwaki, Rie; Kozawa, Osamu; Tokuda, Haruhiko

2017-01-01

Chlorogenic acid (CGA) is a primary phenolic component of coffee and (−)-epigallocatechin gallate (EGCG) is a primary flavonoid component of green tea, both of which have been documented to possess beneficial health properties. A previous study by the present authors demonstrated that p38 mitogen-activated protein kinase (MAPK) may be associated with osteoprotegerin synthesis stimulated by bone morphogenetic protein-4 (BMP-4) in osteoblast-like MC3T3-E1 cells. In the present study, the effects of CGA and EGCG on BMP-4-stimulated osteoprotegerin synthesis in MC3T3-E1 cells were investigated. It was observed that CGA had no effect on osteoprotegerin release stimulated by BMP-4, whereas EGCG significantly enhanced BMP-4-stimulated osteoprotegerin release (P=0.003). Levels of osteoprotegerin mRNA expression induced by BMP-4 were also significantly increased by EGCG (P=0.03). By contrast, EGCG had no significant effect on phosphorylation of Smad1 or p38 MAPK induced by BMP-4. In addition, EGCG had little effect on BMP-induced phosphorylation of p70 S6 kinase; however rapamycin, as an inhibitor of p70 S6 kinase, significantly suppressed osteoprotegerin release (P=0.007). These data suggest that EGCG but not CGA may upregulate the synthesis of osteoprotegerin induced by BMP-4 in osteoblasts. PMID:28672948

(-)-Epigallocatechin gallate but not chlorogenic acid upregulates osteoprotegerin synthesis through regulation of bone morphogenetic protein-4 in osteoblasts.

PubMed

Fujita, Kazuhiko; Otsuka, Takanobu; Yamamoto, Naohiro; Kainuma, Shingo; Ohguchi, Reou; Kawabata, Tetsu; Sakai, Go; Kuroyanagi, Gen; Matsushima-Nishiwaki, Rie; Kozawa, Osamu; Tokuda, Haruhiko

2017-07-01

Chlorogenic acid (CGA) is a primary phenolic component of coffee and (-)-epigallocatechin gallate (EGCG) is a primary flavonoid component of green tea, both of which have been documented to possess beneficial health properties. A previous study by the present authors demonstrated that p38 mitogen-activated protein kinase (MAPK) may be associated with osteoprotegerin synthesis stimulated by bone morphogenetic protein-4 (BMP-4) in osteoblast-like MC3T3-E1 cells. In the present study, the effects of CGA and EGCG on BMP-4-stimulated osteoprotegerin synthesis in MC3T3-E1 cells were investigated. It was observed that CGA had no effect on osteoprotegerin release stimulated by BMP-4, whereas EGCG significantly enhanced BMP-4-stimulated osteoprotegerin release (P=0.003). Levels of osteoprotegerin mRNA expression induced by BMP-4 were also significantly increased by EGCG (P=0.03). By contrast, EGCG had no significant effect on phosphorylation of Smad1 or p38 MAPK induced by BMP-4. In addition, EGCG had little effect on BMP-induced phosphorylation of p70 S6 kinase; however rapamycin, as an inhibitor of p70 S6 kinase, significantly suppressed osteoprotegerin release (P=0.007). These data suggest that EGCG but not CGA may upregulate the synthesis of osteoprotegerin induced by BMP-4 in osteoblasts.

A Long-Acting BMP-2 Release System Based on Poly(3-hydroxybutyrate) Nanoparticles Modified by Amphiphilic Phospholipid for Osteogenic Differentiation

PubMed Central

Peng, Xiaochun; Chen, Yunsu; Li, Yamin; Wang, Yiming

2016-01-01

We explored a novel poly(3-hydroxybutyrate) (PHB) nanoparticle loaded with hydrophilic recombinant human BMP-2 with amphiphilic phospholipid (BPC-PHB NP) for a rapid-acting and long-acting delivery system of BMP-2 for osteogenic differentiation. The BPC-PHB NPs were prepared by a solvent evaporation method and showed a spherical particle with a mean particle size of 253.4 nm, mean zeta potential of −22.42 mV, and high entrapment efficiency of 77.18%, respectively. For BPC-PHB NPs, a short initial burst release of BMP-2 from NPs in 24 h was found and it has steadily risen to reach about 80% in 20 days for in vitro test. BPC-PHB NPs significantly reduced the burst release of BMP-2, as compared to that of PHB NPs loading BMP-2 without PL (B-PHB NPs). BPC-PHB NPs maintained the content of BMP-2 for a long-term osteogenic differentiation. The OCT-1 cells with BPC-PHB NPs have high ALP activity in comparison with others. The gene markers for osteogenic differentiation were significantly upregulated for sample with BPC-PHB NPs, implying that BPC-PHB NPs can be used as a rapid-acting and long-acting BMP-2 delivery system for osteogenic differentiation. PMID:27379249

Gene Therapy of Bone Morphogenetic Protein for Periodontal Tissue Engineering

PubMed Central

Jin, Q-M.; Anusaksathien, O.; Webb, S.A.; Rutherford, R.B.; Giannobile, W.V.

2009-01-01

Background The reconstruction of lost periodontal support including bone, ligament, and cementum is a major goal of therapy. Bone morphogenetic proteins (BMPs) have shown much potential in the regeneration of the periodontium. Limitations of BMP administration to periodontal lesions include need for high-dose bolus delivery, BMP transient biological activity, and low bioavailability of factors at the wound site. Gene transfer offers promise as an alternative treatment strategy to deliver BMPs to periodontal tissues. Methods This study utilized ex vivo BMP-7 gene transfer to stimulate tissue engineering of alveolar bone wounds. Syngeneic dermal fibroblasts (SDFs) were transduced ex vivo with adenoviruses encoding either green fluorescent protein (Ad-GFP or control virus), BMP-7 (Ad-BMP-7), or an antagonist of BMP bioactivity, noggin (Ad-noggin). Transduced cells were seeded onto gelatin carriers and then transplanted to large mandibular alveolar bone defects in a rat wound repair model. Results Ad-noggin treatment tended to inhibit osteogenesis as compared to the control-treated and Ad-BMP-7-treated specimens. The osseous lesions treated by Ad-BMP-7 gene delivery demonstrated rapid chrondrogenesis, with subsequent osteogenesis, cementogenesis and predictable bridging of the periodontal bone defects. Conclusion These results demonstrate the first successful evidence of periodontal tissue engineering using ex vivo gene transfer of BMPs and offers a new approach for repairing periodontal defects. PMID:12666709

Abrogation of epithelial BMP2 and BMP4 causes Amelogenesis Imperfecta by reducing MMP20 and KLK4 expression.

PubMed

Xie, Xiaohua; Liu, Chao; Zhang, Hua; Jani, Priyam H; Lu, Yongbo; Wang, Xiaofang; Zhang, Bin; Qin, Chunlin

2016-05-05

Amelogenesis Imperfecta (AI) can be caused by the deficiencies of enamel matrix proteins, molecules responsible for the transportation and secretion of enamel matrix components, and proteases processing enamel matrix proteins. In the present study, we discovered the double deletion of bone morphogenetic protein 2 (Bmp2) and bone morphogenetic protein 4 (Bmp4) in the dental epithelium by K14-cre resulted in hypoplastic enamel and reduced density in X-ray radiography as well as shortened enamel rods under scanning electron microscopy. Such enamel phenotype was consistent with the diagnosis of hypoplastic amelogenesis imperfecta. Histological and molecular analyses revealed that the removal of matrix proteins in the mutant enamel was drastically delayed, which was coincided with the greatly reduced expression of matrix metalloproteinase 20 (MMP20) and kallikrein 4 (KLK4). Although the expression of multiple enamel matrix proteins was down-regulated in the mutant ameloblasts, the cleavage of ameloblastin was drastically impaired. Therefore, we attributed the AI primarily to the reduction of MMP20 and KLK4. Further investigation found that BMP/Smad4 signaling pathway was down-regulated in the K14-cre;Bmp2(f/f);Bmp4(f/f)ameloblasts, suggesting that the reduced MMP20 and KLK4 expression may be due to the attenuated epithelial BMP/Smad4 signaling.

Abrogation of epithelial BMP2 and BMP4 causes Amelogenesis Imperfecta by reducing MMP20 and KLK4 expression

PubMed Central

Xie, Xiaohua; Liu, Chao; Zhang, Hua; Jani, Priyam H.; Lu, Yongbo; Wang, Xiaofang; Zhang, Bin; Qin, Chunlin

2016-01-01

Amelogenesis Imperfecta (AI) can be caused by the deficiencies of enamel matrix proteins, molecules responsible for the transportation and secretion of enamel matrix components, and proteases processing enamel matrix proteins. In the present study, we discovered the double deletion of bone morphogenetic protein 2 (Bmp2) and bone morphogenetic protein 4 (Bmp4) in the dental epithelium by K14-cre resulted in hypoplastic enamel and reduced density in X-ray radiography as well as shortened enamel rods under scanning electron microscopy. Such enamel phenotype was consistent with the diagnosis of hypoplastic amelogenesis imperfecta. Histological and molecular analyses revealed that the removal of matrix proteins in the mutant enamel was drastically delayed, which was coincided with the greatly reduced expression of matrix metalloproteinase 20 (MMP20) and kallikrein 4 (KLK4). Although the expression of multiple enamel matrix proteins was down-regulated in the mutant ameloblasts, the cleavage of ameloblastin was drastically impaired. Therefore, we attributed the AI primarily to the reduction of MMP20 and KLK4. Further investigation found that BMP/Smad4 signaling pathway was down-regulated in the K14-cre;Bmp2f/f;Bmp4f/fameloblasts, suggesting that the reduced MMP20 and KLK4 expression may be due to the attenuated epithelial BMP/Smad4 signaling. PMID:27146352

MicroRNA-137 inhibits BMP7 to enhance the epithelial-mesenchymal transition of breast cancer cells

PubMed Central

Ying, Xuexiang; Sun, Yunpo; He, Pingqing

2017-01-01

Bone morphogenetic protein-7 (BMP7) is known to antagonize transforming growth factor β 1 (TGFβ1)-mediated fibrosis through suppressing epithelial-mesenchymal transition (EMT). We recently reported that BMP7 also antagonizes the effects of TGFβ1 in breast cancer (BC) tumorigenesis-related EMT. Nevertheless, the control of BMP7 expression in BC remains ill-defined. Here, we detected significantly lower levels of BMP7 and significantly higher levels of microRNA-137 (miR-137) in the BC specimens, relative to paired adjacent non-tumor breast tissue. BMP7 and miR-137 levels were correlated inversely. Additionally, the high miR-137 levels in BC specimens were correlated with reduced patient survival. In vitro, overexpression of miR-137 significantly increased cell EMT and invasion, while depletion of miR-137 significantly decreased cell EMT and invasion in BC cells. The increases in BC cell invasiveness by miR-137 appeared to result from its suppression of BMP7, through direct binding of miR-137 to the 3'-UTR of BMP7 mRNA, thereby blocking its protein translation in BC cells. This study sheds light on miR-137 as a crucial factor that enhances BC cell EMT and invasiveness, and points to miR-137 as a promising innovative therapeutic target for BC treatment. PMID:28407692

Bone Morphogenetic Protein-2 Promotes Human Mesenchymal Stem Cell Survival and Resultant Bone Formation When Entrapped in Photocrosslinked Alginate Hydrogels.

PubMed

Ho, Steve S; Vollmer, Nina L; Refaat, Motasem I; Jeon, Oju; Alsberg, Eben; Lee, Mark A; Leach, J Kent

2016-10-01

There is a substantial need to prolong cell persistence and enhance functionality in situ to enhance cell-based tissue repair. Bone morphogenetic protein-2 (BMP-2) is often used at high concentrations for osteogenic differentiation of mesenchymal stem cells (MSCs) but can induce apoptosis. Biomaterials facilitate the delivery of lower doses of BMP-2, reducing side effects and localizing materials at target sites. Photocrosslinked alginate hydrogels (PAHs) can deliver osteogenic materials to irregular-sized bone defects, providing improved control over material degradation compared to ionically cross-linked hydrogels. It is hypothesized that the delivery of MSCs and BMP-2 from a PAH increases cell persistence by reducing apoptosis, while promoting osteogenic differentiation and enhancing bone formation compared to MSCs in PAHs without BMP-2. BMP-2 significantly decreases apoptosis and enhances survival of photoencapsulated MSCs, while simultaneously promoting osteogenic differentiation in vitro. Bioluminescence imaging reveals increased MSC survival when implanted in BMP-2 PAHs. Bone defects treated with MSCs in BMP-2 PAHs demonstrate 100% union as early as 8 weeks and significantly higher bone volumes at 12 weeks, while defects with MSC-entrapped PAHs alone do not fully bridge. This study demonstrates that transplantation of MSCs with BMP-2 in PAHs achieves robust bone healing, providing a promising platform for bone repair. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

BMP4 Cooperates with Retinoic Acid to Induce the Expression of Differentiation Markers in Cultured Mouse Spermatogonia

PubMed Central

Feng, Yanmin; Feng, Xue; Wang, Xiuxia; Gan, Haiyun; Wang, Lixian; Lin, Xiwen

2016-01-01

Spermatogenesis is sustained by the proliferation and differentiation of spermatogonial stem cells (SSCs). However, the molecules controlling these processes remain largely unknown. Here, we developed a simplified high concentration serum-containing system for the culture of mouse SSCs. Analysis of SSCs markers and transplantation results revealed that the cultured spermatogonia retained stem cell characteristics after long-term in vitro propagation. Using this culture system, the expression and function of bone morphogenetic protein 4 (BMP4) were explored. Immunostaining showed that BMP4 was predominantly expressed in germ cells and that its level increased as spermatogenesis progresses. BMP4 receptors BMPR1A and BMPRII were present in spermatogonia, spermatocytes, and round spermatids. Moreover, despite the mRNAs of these two genes being present in mouse Sertoli cells, only BMPRII was detected by using Western blotting assays. While exogenous BMP4 by itself did not induce the expression of Stra8 and c-Kit, two marker genes of differentiating spermatogonia, a significant cooperative effect of BMP4 and retinoic acid (RA) was observed. Moreover, pretreatment of cultured spermatogonia with the BMP4 antagonist Noggin could inhibit RA-induced expression of these two marker genes. In conclusion, BMP4 may exert autocrine effects and act cooperatively with RA to induce the differentiation of spermatogonia in vivo. PMID:27795714

Essential roles of zebrafish bmp2a, fgf10, and fgf24 in the specification of the ventral pancreas

PubMed Central

Naye, François; Voz, Marianne L.; Detry, Nathalie; Hammerschmidt, Matthias; Peers, Bernard; Manfroid, Isabelle

2012-01-01

In vertebrates, pancreas and liver arise from bipotential progenitors located in the embryonic gut endoderm. Bone morphogenic protein (BMP) and fibroblast growth factor (FGF) signaling pathways have been shown to induce hepatic specification while repressing pancreatic fate. Here we show that BMP and FGF factors also play crucial function, at slightly later stages, in the specification of the ventral pancreas. By analyzing the pancreatic markers pdx1, ptf1a, and hlxb9la in different zebrafish models of BMP loss of function, we demonstrate that the BMP pathway is required between 20 and 24 h postfertilization to specify the ventral pancreatic bud. Knockdown experiments show that bmp2a, expressed in the lateral plate mesoderm at these stages, is essential for ventral pancreas specification. Bmp2a action is not restricted to the pancreatic domain and is also required for the proper expression of hepatic markers. By contrast, through the analysis of fgf10−/−; fgf24−/− embryos, we reveal the specific role of these two FGF ligands in the induction of the ventral pancreas and in the repression of the hepatic fate. These mutants display ventral pancreas agenesis and ectopic masses of hepatocytes. Overall, these data highlight the dynamic role of BMP and FGF in the patterning of the hepatopancreatic region. PMID:22219376

Bone morphogenetic protein use in spine surgery-complications and outcomes: a systematic review.

PubMed

Faundez, Antonio; Tournier, Clément; Garcia, Matthieu; Aunoble, Stéphane; Le Huec, Jean-Charles

2016-06-01

Because of significant complications related to the use of autologous bone grafts in spinal fusion surgery, bone substitutes and growth factors such as bone morphogenetic protein (BMP) have been developed. One of them, recombinant human (rh) BMP-2, has been approved by the Food and Drug Administration (FDA) for use under precise conditions. However, rhBMP-2-related side effects have been reported, used in FDA-approved procedures, but also in off-label use.A systematic review of clinical data was conducted to analyse the rhBMP-2-related adverse events (AEs), in order to assess their prevalence and the associated surgery practices. Medline search with keywords "bone morphogenetic protein 2", "lumbar spine", "anterolateral interbody fusion" (ALIF) and the filter "clinical trial". FDA published reports were also included. Study assessment was made by authors (experienced spine surgeons), based on quality of study designs and level of evidence. Extensive review of randomised controlled trials (RCTs) and controlled series published up to the present point, reveal no evidence of a significant increase of AEs related to rhBMP-2 use during ALIF surgeries, provided that it is used following FDA guidelines. Two additional RCTs performed with rhBMP-2 in combination with allogenic bone dowels reported increased bone remodelling in BMP-treated patients. This AE was transient and had no consequence on the clinical outcome of the patients. No other BMP-related AEs were reported in these studies. This literature review confirms that the use of rhBMP-2 following FDA-approved recommendations (i.e. one-level ALIF surgery with an LT-cage) is safe. The rate of complications is low and the AEs had been identified by the FDA during the pre-marketing clinical trials. The clinical efficiency of rhBMP-2 is equal or superior to that of allogenic or autologous bone graft in respect to fusion rate, low back pain disability, patient satisfaction and rate of re-operations. For all other off-label use, the safety and effectiveness of rhBMP-2 have not been established, and further RCTs with high level of evidence are required.

Deproteinized bovine bone functionalized with the slow delivery of BMP-2 for the repair of critical-sized bone defects in sheep.

PubMed

Liu, Tie; Wu, Gang; Wismeijer, Daniel; Gu, Zhiyuan; Liu, Yuelian

2013-09-01

As an alternative to an autologous bone graft, deproteinized bovine bone (DBB) is widely used in the clinical dentistry. Although DBB provides an osteoconductive scaffold, it is not capable of enhancing bone regeneration because it is not osteoinductive. In order to render DBB osteoinductive, bone morphogenetic protein 2 (BMP-2) has previously been incorporated into a three dimensional reservoir (a biomimetic calcium phosphate coating) on DBB, which effectively promoted the osteogenic response by the slow delivery of BMP-2. The aim of this study was to investigate the therapeutic effectiveness of such coating on the DBB granules in repairing a large cylindrical bone defect (8 mm diameter, 13 mm depth) in sheep. Eight groups were randomly assigned to the bone defects: (i) no graft material; (ii) autologous bone; (iii) DBB only; (iv) DBB mixed with autologous bone; (v) DBB bearing adsorbed BMP-2; (vi) DBB bearing a coating but no BMP-2; (vii) DBB bearing a coating with adsorbed BMP-2; and (viii) DBB bearing a coating-incorporated depot of BMP-2. 4 and 8 weeks after implantation, samples were withdrawn for a histological and a histomorphometric analysis. Histological results confirmed the excellent biocompatibility and osteoconductivity of all the grafts tested. At 4 weeks, DBB mixed with autologous bone or functionalized with coating-incorporated BMP-2 showed more newly-formed bone than the other groups with DBB. At 8 weeks, the volume of newly-formed bone around DBB that bore a coating-incorporated depot of BMP-2 was greatest among the groups with DBB, and was comparable to the autologous bone group. The use of autologous bone and BMP-2 resulted in more bone marrow formation. Multinucleated giant cells were observed in the resorption process around DBB, whereas histomorphometric analysis revealed no significant degradation of DBB. In conclusion, it was shown that incorporating BMP-2 into the calcium phosphate coating of DBB induced strong bone formation around DBB for repairing a critical-sized bone defect. Copyright © 2013 Elsevier Inc. All rights reserved.

Neurologic impairment from ectopic bone in the lumbar canal: a potential complication of off-label PLIF/TLIF use of bone morphogenetic protein-2 (BMP-2).

PubMed

Wong, David A; Kumar, Anant; Jatana, Sanjay; Ghiselli, Gary; Wong, Katherine

2008-01-01

Bone morphogenetic protein-2 (BMP-2) (Infuse) has been approved for use in anterior lumbar fusion in conjunction with an LT cage. However, off-label use is seen with anterior cervical fusion, posterior lumbar interbody fusion (PLIF), and transforaminal lumbar interbody fusion (TLIF). The Federal Food and Drug Administration trial of BMP-2 in a PLIF application was halted because of a high incidence of ectopic bone forming in the neural canal (75%). The authors did not find a correlation between ectopic bone and increased leg pain. They concluded that the ectopic bone was a radiographic phenomenon and not associated with clinical findings. Complications using BMP in the cervical spine have been reported. Heretofore, there has not been a similar warning voiced for use of BMP in a lumbar PLIF or TLIF. The purpose was to report five cases of ectopic bone in the canal associated with PLIF/TLIF off-label use of BMP-2 potentially contributing to abnormal neurologic findings. This is an observational cohort study of patients referred to a tertiary care private medical center. This was a retrospective chart review of patients referred to a tertiary spine institute with complications after surgery where BMP-2 had been used in an off-label PLIF or TLIF application. Patient demographics, operating room (OR) notes from the index BMP surgery, imaging studies, and current clinical status were reviewed. Five cases of ectopic bone in the spinal canal with potential neurologic compromise were identified. It does appear that ectopic bone in the spinal canal associated with BMP-2 use in PLIF or TLIF may contribute to symptomatic neurologic findings in rare cases. Revision surgeries are difficult. This article challenges a previous publication, which concluded that the high incidence of ectopic bone was of no clinical significance. Isolating BMP anteriorly in the disc space using layered barriers of bone graft between the BMP and the annular defect may reduce the incidence of ectopic bone in the spinal canal. Surgeons need to weigh the benefits versus risks of any technology used off label when making treatment decisions with their patients.

7 CFR 634.23 - Water quality plan.

Code of Federal Regulations, 2010 CFR

2010-01-01

... quality management plan. Such BMP's must reduce the amount of pollutants that enter a stream or lake by... 7 Agriculture 6 2010-01-01 2010-01-01 false Water quality plan. 634.23 Section 634.23 Agriculture... AGRICULTURE LONG TERM CONTRACTING RURAL CLEAN WATER PROGRAM Participant RCWP Contracts § 634.23 Water quality...

[A novel tissue-engineered bone constructed by using human adipose-derived stem cells and biomimetic calcium phosphate scaffold coprecipitated with bone morphogenetic protein-2].

PubMed

Jiang, W R; Zhang, X; Liu, Y S; Wu, G; Ge, Y J; Zhou, Y S

2017-02-18

To construct a novel biomimetic calcium phosphate (BioCaP) scaffold loaded with bone morphogenetic protein-2 (BMP-2), and to investigate its role in the osteogenesis of human adipose-derived stem cells (hASCs) in vitro and in vivo. The BioCaP scaffold coprecipitated with BMP-2 (BMP-2-BioCaP) was constructed in this study. Field emission scanning electron microscopy (SEM) was used to analyze the morphology of the surfaces. The release kinetics was measured to evaluate the slow-release characteristics in vitro. BMP-2-BioCaP was immersed in proliferation medium (PM) or osteogenic medium (OM), respectively. The supernatants were collected and used to culture hASCs in vitro. Cell numbers were determined using the cell-counting kit-8 (CCK-8) to assess the cell proliferation. After 7 and 14 days, alkaline phosphatase (ALP) staining and quantification were performed to test the activity of ALP. After 14 and 21 days, the calcification deposition was determined by alizarin red S (ARS) staining and quantification. The expressions of the osteoblast-related genes were tested on day 4 and day 14. In the in vivo study, 6 nude mice were used and implanted subcutaneously into the back of the nude mice for 4 groups: (1) BioCaP scaffold only, (2) BioCaP scaffold+hASCs, (3) BMP-2-BioCaP scaffold, (4) BMP-2-BioCaP scaffold+hASCs (test group). After 4 weeks of implantation, hematoxylin-eosin (HE) staining was performed to evaluate the in vivo osteogenesis of hASCs. SEM observations showed that BioCaP and BMP-2-BioCaP scaffold were entirely composed of straight, plate-like and sharp-edged crystal units, and the length of the crystal units varied between 5 and 10 μm. Release kinetics analysis demonstrated that BMP-2 incorporated with BioCaP could be released at certain concentration and last for more than 21 days, and the accumulative protein release could reach 20%. CCK-8 assays showed that cell proliferation was not significantly affected by BMP-2-BioCaP. ALP activity was higher by the induction of OM+BMP-2-BioCaP than of the other groups (P<0.01). More mineralization deposition and more expressions of osteoblast-related genes such as Runt-related transcription factor 2 (RUNX2), ALP, osteopontin (OPN) and osteocalcin (OC) were determined in the OM+BMP-2-BioCaP group at different time points (P<0.01). HE staining showed that, in the test group and BMP-2-BioCaP scaffold group, the extracellular matrix (ECM) with eosinophilic staining were observed around hASCs, and newly-formed bone-like tissues could be found in ECM around the scaffold materials. Moreover, compared with the BMP-2-BioCaP scaffold group, more bone-like tissues could be observed in ECM with typical structure of bone tissue in the test groups. No obvious positive results were found in the other groups. BMP-2-BioCaP scaffold could achieve slow-release of BMP-2 and promote the osteogenic differentiation of hASCs in vitro and in vivo. The novel tissue-engineered bone composed of hASCs and BMP-2-BioCaPis promising for the repair of bone defect.

BMP signaling components in embryonic transcriptomes of the hover fly Episyrphus balteatus (Syrphidae)

PubMed Central

2011-01-01

Background In animals, signaling of Bone Morphogenetic Proteins (BMPs) is essential for dorsoventral (DV) patterning of the embryo, but how BMP signaling evolved with changes in embryonic DV differentiation is largely unclear. Based on the extensive knowledge of BMP signaling in Drosophila melanogaster, the morphological diversity of extraembryonic tissues in different fly species provides a comparative system to address this question. The closest relatives of D. melanogaster with clearly distinct DV differentiation are hover flies (Diptera: Syrphidae). The syrphid Episyrphus balteatus is a commercial bio-agent against aphids and has been established as a model organism for developmental studies and chemical ecology. The dorsal blastoderm of E. balteatus gives rise to two extraembryonic tissues (serosa and amnion), whereas in D. melanogaster, the dorsal blastoderm differentiates into a single extraembryonic epithelium (amnioserosa). Recent studies indicate that several BMP signaling components of D. melanogaster, including the BMP ligand Screw (Scw) and other extracellular regulators, evolved in the dipteran lineage through gene duplication and functional divergence. These findings raise the question of whether the complement of BMP signaling components changed with the origin of the amnioserosa. Results To search for BMP signaling components in E. balteatus, we generated and analyzed transcriptomes of freshly laid eggs (0-30 minutes) and late blastoderm to early germband extension stages (3-6 hours) using Roche/454 sequencing. We identified putative E. balteatus orthologues of 43% of all annotated D. melanogaster genes, including the genes of all BMP ligands and other BMP signaling components. Conclusion The diversification of several BMP signaling components in the dipteran linage of D. melanogaster preceded the origin of the amnioserosa. [Transcriptome sequence data from this study have been deposited at the NCBI Sequence Read Archive (SRP005289); individually assembled sequences have been deposited at GenBank (JN006969-JN006986).] PMID:21627820

Inkjet-based biopatterning of SDF-1β augments BMP-2-induced repair of critical size calvarial bone defects in mice

PubMed Central

Herberg, Samuel; Kondrikova, Galina; Periyasamy-Thandavan, Sudharsan; Howie, R. Nicole; Elsalanty, Mohammed E.; Weiss, Lee; Campbell, Phil; Hill, William D.; Cray, James J.

2014-01-01

Background A major problem in craniofacial surgery is non-healing bone defects. Autologous reconstruction remains the standard of care for these cases. Bone morphogenetic protein-2 (BMP-2) therapy has proven its clinical utility, although non-targeted adverse events occur due to the high milligram-level doses used. Ongoing efforts explore the use of different growth factors, cytokines, or chemokines, as well as co-therapy to augment healing. Methods Here we utilize inkjet-based biopatterning to load acellular DermaMatrix delivery matrices with nanogram-level doses of BMP-2, stromal cell-derived factor-1β (SDF-1β), transforming growth factor-β1 (TGF-β1), or co-therapies thereof. We tested the hypothesis that bioprinted SDF-1β co-delivery enhances BMP-2 and TGF-β1-driven osteogenesis both in-vitro and in-vivo using a mouse calvarial critical size defect (CSD) model. Results Our data showed that BMP-2 bioprinted in low-doses induced significant new bone formation by four weeks post-operation. TGF-β1 was less effective compared to BMP-2, and SDF-1β therapy did not enhance osteogenesis above control levels. However, co-delivery of BMP-2 + SDF-1β was shown to augment BMP-2-induced bone formation compared to BMP-2 alone. In contrast, co-delivery of TGF-β1 + SDF-1β decreased bone healing compared to TGF-β1 alone. This was further confirmed in vitro by osteogenic differentiation studies using MC3T3-E1 pre-osteoblasts. Conclusions Our data indicates that sustained release delivery of a low-dose growth factor therapy using biopatterning technology can aid in healing CSD injuries. SDF-1β augments the ability for BMP-2 to drive healing, a result confirmed in vivo and in vitro; however, because SDF-1β is detrimental to TGF-β1-driven osteogenesis, its’ effect on osteogenesis is not universal. PMID:25016095

Borrelia burgdorferi basic membrane protein A could induce chemokine production in murine microglia cell line BV2.

PubMed

Zhao, Hua; Liu, Aihua; Cui, Yuhui; Liang, Zhang; Li, Bingxue; Bao, Fukai

2017-10-01

Lyme neuroborreliosis is a nervous system infectious disease caused by Borrelia burgdorferi (B. burgdorferi). It has been demonstrated that cytokines induced by B. burgdorferi are related to Lyme neuroborreliosis. Microglia is known as a key player in the immune responses that occur within the central nervous system. In response to inflammation, it will be activated and generate cytokines and chemokines. Experiments in vitro cells have showed that B. Burgdorferi membrane protein A (BmpA), a major immunogen of B. Burgdorferi, could induce Lyme arthritis and stimulate human and murine lymphocytes to produce inflammatory cytokines. In our study, the murine microglia BV2 cell line was used as a cell model to explore the stimulating effects of recombinant BmpA (rBmpA); Chemokine chip, ELISA and QPCR technology were used to measure the production of chemokines from microglial cells stimulated by rBmpA. Compared with the negative control group, CXCL2, CCL22, and CCL5 concentrations in the cell supernatant increased significantly after the rBmpA stimulation; the concentration of these chemokines increased with rBmpA concentration increasing; the mRNA expression levels of chemokines (CXCL2, CCL22, and CCL5) in murine BV2 cells increased significantly with 10 μg/mL and 20 μg/mL rBmpA stimulation; CXCL13 was not change after the rBmpA stimulation. Our study shows that chemokines, such as CXCL2, CCL22, and CCL5 were up-regulated by the rBmpA in the BV2 cells. The production of chemokines in Lyme neuroborreliosis may be mainly from microglia cells and the rBmpA may be closely related with the development of Lyme neuroborreliosis. Copyright © 2017 Elsevier Ltd. All rights reserved.

Uncovering Molecular Bases Underlying Bone Morphogenetic Protein Receptor Inhibitor Selectivity

PubMed Central

Alsamarah, Abdelaziz; LaCuran, Alecander E.; Oelschlaeger, Peter; Hao, Jijun; Luo, Yun

2015-01-01

Abnormal alteration of bone morphogenetic protein (BMP) signaling is implicated in many types of diseases including cancer and heterotopic ossifications. Hence, small molecules targeting BMP type I receptors (BMPRI) to interrupt BMP signaling are believed to be an effective approach to treat these diseases. However, lack of understanding of the molecular determinants responsible for the binding selectivity of current BMP inhibitors has been a big hindrance to the development of BMP inhibitors for clinical use. To address this issue, we carried out in silico experiments to test whether computational methods can reproduce and explain the high selectivity of a small molecule BMP inhibitor DMH1 on BMPRI kinase ALK2 vs. the closely related TGF-β type I receptor kinase ALK5 and vascular endothelial growth factor receptor type 2 (VEGFR2) tyrosine kinase. We found that, while the rigid docking method used here gave nearly identical binding affinity scores among the three kinases; free energy perturbation coupled with Hamiltonian replica-exchange molecular dynamics (FEP/H-REMD) simulations reproduced the absolute binding free energies in excellent agreement with experimental data. Furthermore, the binding poses identified by FEP/H-REMD led to a quantitative analysis of physical/chemical determinants governing DMH1 selectivity. The current work illustrates that small changes in the binding site residue type (e.g. pre-hinge region in ALK2 vs. ALK5) or side chain orientation (e.g. Tyr219 in caALK2 vs. wtALK2), as well as a subtle structural modification on the ligand (e.g. DMH1 vs. LDN193189) will cause distinct binding profiles and selectivity among BMP inhibitors. Therefore, the current computational approach represents a new way of investigating BMP inhibitors. Our results provide critical information for designing exclusively selective BMP inhibitors for the development of effective pharmacotherapy for diseases caused by aberrant BMP signaling. PMID:26133550

Synthetic scaffold coating with adeno-associated virus encoding BMP2 to promote endogenous bone repair.

PubMed

Dupont, Kenneth M; Boerckel, Joel D; Stevens, Hazel Y; Diab, Tamim; Kolambkar, Yash M; Takahata, Masahiko; Schwarz, Edward M; Guldberg, Robert E

2012-03-01

Biomaterial scaffolds functionalized to stimulate endogenous repair mechanisms via the incorporation of osteogenic cues offer a potential alternative to bone grafting for the treatment of large bone defects. We first quantified the ability of a self-complementary adeno-associated viral vector encoding bone morphogenetic protein 2 (scAAV2.5-BMP2) to enhance human stem cell osteogenic differentiation in vitro. In two-dimensional culture, scAAV2.5-BMP2-transduced human mesenchymal stem cells (hMSCs) displayed significant increases in BMP2 production and alkaline phosphatase activity compared with controls. hMSCs and human amniotic-fluid-derived stem cells (hAFS cells) seeded on scAAV2.5-BMP2-coated three-dimensional porous polymer Poly(ε-caprolactone) (PCL) scaffolds also displayed significant increases in BMP2 production compared with controls during 12 weeks of culture, although only hMSC-seeded scaffolds displayed significantly increased mineral formation. PCL scaffolds coated with scAAV2.5-BMP2 were implanted into critically sized immunocompromised rat femoral defects, both with or without pre-seeding of hMSCs, representing ex vivo and in vivo gene therapy treatments, respectively. After 12 weeks, defects treated with acellular scAAV2.5-BMP2-coated scaffolds displayed increased bony bridging and had significantly higher bone ingrowth and mechanical properties compared with controls, whereas defects treated with scAAV2.5-BMP2 scaffolds pre-seeded with hMSCs failed to display significant differences relative to controls. When pooled, defect treatment with scAAV2.5-BMP2-coated scaffolds, both with or without inclusion of pre-seeded hMSCs, led to significant increases in defect mineral formation at all time points and increased mechanical properties compared with controls. This study thus presents a novel acellular bone-graft-free endogenous repair therapy for orthotopic tissue-engineered bone regeneration.

Experimental and Computational Investigation of the Effect of Hydrophobicity on Aggregation and Osteoinductive Potential of BMP-2-Derived Peptide in a Hydrogel Matrix

PubMed Central

Moeinzadeh, Seyedsina; Barati, Danial; Sarvestani, Samaneh K.; Karimi, Tahereh

2015-01-01

An attractive approach to reduce the undesired side effects of bone morphogenetic proteins (BMPs) in regenerative medicine is to use osteoinductive peptide sequences derived from BMPs. Although the structure and function of BMPs have been studied extensively, there is limited data on structure and activity of BMP-derived peptides immobilized in hydrogels. The objective of this work was to investigate the effect of concentration and hydrophobicity of the BMP-2 peptide, corresponding to residues 73–92 of the knuckle epitope of BMP-2 protein, on peptide aggregation and osteogenic differentiation of human mesenchymal stem cells encapsulated in a polyethylene glycol (PEG) hydrogel. The peptide hydrophobicity was varied by capping PEG chain ends with short lactide segments. The BMP-2 peptide with a positive index of hydrophobicity had a critical micelle concentration (CMC) and formed aggregates in aqueous solution. Based on simulation results, there was a slight increase in the concentration of free peptide in solution with 1000-fold increase in peptide concentration. The dose-osteogenic response curve of the BMP-2 peptide was in the 0.0005–0.005 mM range, and osteoinductive potential of the BMP-2 peptide was significantly less than that of BMP-2 protein even at 1000-fold higher concentrations, which was attributed to peptide aggregation. Further, the peptide or PEG-peptide aggregates had significantly higher interaction energy with the cell membrane compared with the free peptide, which led to a higher nonspecific interaction with the cell membrane and loss of osteoinductive potential. Conjugation of the BMP-2 peptide to PEG increased CMC and osteoinductive potential of the peptide whereas conjugation to lactide-capped PEG reduced CMC and osteoinductive potential of the peptide. Experimental and simulation results revealed that osteoinductive potential of the BMP-2 peptide is correlated with its CMC and the free peptide concentration in aqueous medium and not the total concentration. PMID:25051457

Trends Analysis of rhBMP2 Utilization in Single-Level Anterior Lumbar Interbody Fusion in the United States

PubMed Central

Lao, Lifeng; Cohen, Jeremiah R.; Buser, Zorica; Brodke, Darrel S.; Yoon, S. Tim; Youssef, Jim A.; Park, Jong-Beom; Meisel, Hans-Joerg; Wang, Jeffrey C.

2017-01-01

Study Design: Retrospective case study. Objective: To evaluate the trends and demographics of recombinant human bone morphogenetic protein 2 (rhBMP2) utilization in single-level anterior lumbar interbody fusion (ALIF) in the United States. Methods: Patients who underwent single-level ALIF from 2005 to 2011 were identified by searching ICD-9 diagnosis and procedure codes in the PearlDiver Patient Records Database (PearlDiver Technologies, Fort Wayne, IN), a national database of orthopedic insurance records. The year of procedure, age, gender, and region of the United States were analyzed for each patient. Results: A total of 921 patients were identified who underwent a single-level ALIF in this study. The average rate of single-level ALIF with rhBMP2 utilization increased (35%-48%) from 2005 to 2009, but sharply decreased to 16.7% in 2010 and 15.0% in 2011. The overall incidence of single-level ALIF without rhBMP2 (0.20 cases per 100 000 patients) was more than twice of the incidence of single-level ALIF with rhBMP2 (0.09 cases per 100 000 patients). The average rate of single-level ALIF with rhBMP2 utilization is highest in West (41.4%), followed by Midwest (33.3%), South (26.5%) and Northeast (22.2%). The highest incidence of single-level ALIF with rhBMP2 was observed in the group aged less than 65 years (compared with any other age groups, P < .001), with an incidence of 0.21 per 100 000 patients. Conclusions: The incidence of rhBMP2 utilization in single-level ALIF increased from 2006 to 2009, but decreased in 2010 and 2011. The Northeast region had the lowest incidence of rhBMP2 utilization. The group aged less than 65 years trended to have the higher incidence of single-level ALIF with rhBMP2 utilization. PMID:29662743

Trends Analysis of rhBMP2 Utilization in Single-Level Anterior Lumbar Interbody Fusion in the United States.

PubMed

Lao, Lifeng; Cohen, Jeremiah R; Buser, Zorica; Brodke, Darrel S; Yoon, S Tim; Youssef, Jim A; Park, Jong-Beom; Meisel, Hans-Joerg; Wang, Jeffrey C

2018-04-01

Retrospective case study. To evaluate the trends and demographics of recombinant human bone morphogenetic protein 2 (rhBMP2) utilization in single-level anterior lumbar interbody fusion (ALIF) in the United States. Patients who underwent single-level ALIF from 2005 to 2011 were identified by searching ICD-9 diagnosis and procedure codes in the PearlDiver Patient Records Database (PearlDiver Technologies, Fort Wayne, IN), a national database of orthopedic insurance records. The year of procedure, age, gender, and region of the United States were analyzed for each patient. A total of 921 patients were identified who underwent a single-level ALIF in this study. The average rate of single-level ALIF with rhBMP2 utilization increased (35%-48%) from 2005 to 2009, but sharply decreased to 16.7% in 2010 and 15.0% in 2011. The overall incidence of single-level ALIF without rhBMP2 (0.20 cases per 100 000 patients) was more than twice of the incidence of single-level ALIF with rhBMP2 (0.09 cases per 100 000 patients). The average rate of single-level ALIF with rhBMP2 utilization is highest in West (41.4%), followed by Midwest (33.3%), South (26.5%) and Northeast (22.2%). The highest incidence of single-level ALIF with rhBMP2 was observed in the group aged less than 65 years (compared with any other age groups, P < .001), with an incidence of 0.21 per 100 000 patients. The incidence of rhBMP2 utilization in single-level ALIF increased from 2006 to 2009, but decreased in 2010 and 2011. The Northeast region had the lowest incidence of rhBMP2 utilization. The group aged less than 65 years trended to have the higher incidence of single-level ALIF with rhBMP2 utilization.

Evaluation of rhBMP-2/collagen/TCP-HA bone graft with and without bone marrow cells in the canine femoral multi defect model.

PubMed

Luangphakdy, V; Shinohara, K; Pan, H; Boehm, C; Samaranska, A; Muschler, G F

2015-01-12

Recombinant human bone morphogenetic protein-2, when applied to an absorbable type 1 bovine collagen sponge (rhBMP-2/ACS) is an effective therapy in many bone grafting settings. Bone marrow aspirate (BMA) has also been used as a source of transplantable osteogenic connective tissue progenitors. This study was designed to characterize the performance of a scaffold comprising rhBMP-2/ACS in which the sponge wraps around tri-calcium phosphate hydroxyapatite granules (rhBMP-2/ACS/TCP-HA) and to test the hypothesis that addition of BMA will improve the performance of this construct in the Canine Femoral Multi Defect Model. In each subject, two sites were grafted with rhBMP-2/ACS/TCP-HA scaffold loaded with BMA clot and two other sites with rhBMP-2/ACS/TCP-HA scaffold loaded with wound blood (WB). After correction for unresorbed TCP-HA granules, sites grafted with rhBMP-2/ACS/TCP-HA+BMA and rhBMP-2/ACS/TCP-HA+WB were similar, with mean percent bone volumes of 10.9 %±1.2 and 11.2 %±1.2, respectively. No differences were seen in quantitative histomorphometry. While bone formation using both constructs was robust, this study did not support the hypothesis that the addition of unprocessed bone marrow aspirate clot improved bone regeneration in a site engrafted with rhBMP-2/ACS/TCP-HA+BMA. In contrast to prior studies using this model, new bone formation was greater at the center of the defect where TCP-HA was distributed. This finding suggests a potential synergy between rhBMP-2 and the centrally placed ceramic and cellular components of the graft construct. Further optimization may also require more uniform distribution of TCP-HA, alternative cell delivery strategies, and a more rigorous large animal segmental defect model.

Integration of nodal and BMP signals in the heart requires FoxH1 to create left-right differences in cell migration rates that direct cardiac asymmetry.

PubMed

Lenhart, Kari F; Holtzman, Nathalia G; Williams, Jessica R; Burdine, Rebecca D

2013-01-01

Failure to properly establish the left-right (L/R) axis is a major cause of congenital heart defects in humans, but how L/R patterning of the embryo leads to asymmetric cardiac morphogenesis is still unclear. We find that asymmetric Nodal signaling on the left and Bmp signaling act in parallel to establish zebrafish cardiac laterality by modulating cell migration velocities across the L/R axis. Moreover, we demonstrate that Nodal plays the crucial role in generating asymmetry in the heart and that Bmp signaling via Bmp4 is dispensable in the presence of asymmetric Nodal signaling. In addition, we identify a previously unappreciated role for the Nodal-transcription factor FoxH1 in mediating cell responsiveness to Bmp, further linking the control of these two pathways in the heart. The interplay between these TGFβ pathways is complex, with Nodal signaling potentially acting to limit the response to Bmp pathway activation and the dosage of Bmp signals being critical to limit migration rates. These findings have implications for understanding the complex genetic interactions that lead to congenital heart disease in humans.

Biphasic effects of FGF2 on odontoblast differentiation involve changes in the BMP and Wnt signaling pathways.

PubMed

Sagomonyants, Karen; Mina, Mina

2014-08-01

Odontoblast differentiation during physiological and reparative dentinogenesis is dependent upon multiple signaling molecules, including fibroblast growth factors (FGFs), bone morphogenetic proteins (BMPs) and Wingless/Integrated (Wnt) ligands. Recent studies in our laboratory showed that continuous exposure of primary dental pulp cultures to FGF2 exerted biphasic effects on the expression of markers of dentinogenesis. In the present study, we examined the possible involvement of the BMP and Wnt signaling pathways in mediating the effects of FGF2 on dental pulp cells. Our results showed that stimulatory effects of FGF2 on dentinogenesis during the proliferation phase of growth were associated with increased expression of the components of the BMP (Bmp2, Dlx5, Msx2, Osx) and Wnt (Wnt10a, Wisp2) pathways, and decreased expression of an inhibitor of the Wnt signaling, Nkd2. Further addition of FGF2 during the differentiation/mineralization phase of growth resulted in decreased expression of components of the BMP signaling (Bmp2, Runx2, Osx) and increased expression of inhibitors of the Wnt signaling (Nkd2, Dkk3). This suggests that both BMP and Wnt pathways may be involved in mediating the effects of FGF2 on dental pulp cells.

SIX2 and BMP4 mutations associate with anomalous kidney development.

PubMed

Weber, Stefanie; Taylor, Jaclyn C; Winyard, Paul; Baker, Kari F; Sullivan-Brown, Jessica; Schild, Raphael; Knüppel, Tanja; Zurowska, Aleksandra M; Caldas-Alfonso, Alberto; Litwin, Mieczyslaw; Emre, Sevinc; Ghiggeri, Gian Marco; Bakkaloglu, Aysin; Mehls, Otto; Antignac, Corinne; Network, Escape; Schaefer, Franz; Burdine, Rebecca D

2008-05-01

Renal hypodysplasia (RHD) is characterized by reduced kidney size and/or maldevelopment of the renal tissue following abnormal organogenesis. Mutations in renal developmental genes have been identified in a subset of affected individuals. Here, we report the first mutations in BMP4 and SIX2 identified in patients with RHD. We detected 3 BMP4 mutations in 5 RHD patients, and 3 SIX2 mutations in 5 different RHD patients. Overexpression assays in zebrafish demonstrated that these mutations affect the function of Bmp4 and Six2 in vivo. Overexpression of zebrafish six2.1 and bmp4 resulted in dorsalization and ventralization, respectively, suggesting opposing roles in mesendoderm formation. When mutant constructs containing the identified human mutations were overexpressed instead, these effects were attenuated. Morpholino knockdown of bmp4 and six2.1 affected glomerulogenesis, suggesting specific roles for these genes in the formation of the pronephros. In summary, these studies implicate conserved roles for Six2 and Bmp4 in the development of the renal system. Defects in these proteins could affect kidney development at multiple stages, leading to the congenital anomalies observed in patients with RHD.

Localized expression of a dpp/BMP2/4 ortholog in a coral embryo

PubMed Central

Hayward, David C.; Samuel, Gabrielle; Pontynen, Patricia C.; Catmull, Julian; Saint, Robert; Miller, David J.; Ball, Eldon E.

2002-01-01

As the closest outgroup to the Bilateria, the Phylum Cnidaria is likely to be critical to understanding the origins and evolution of body axes. Proteins of the decapentaplegic (DPP)/bone morphogenetic protein (BMP) 2/4 subfamily are central to the specification of the dorsoventral (D/V) axis in bilateral animals, albeit with an axis inversion between arthropods and chordates. We show that a dpp/BMP2/4 ortholog (bmp2/4-Am) is present in the reef-building scleractinian coral, Acropora millepora (Class Anthozoa) and that it is capable of causing phenotypic effects in Drosophila that mimic those of the endogenous dpp gene. We also show that, during coral embryonic development, bmp2/4-Am expression is localized in an ectodermal region adjacent to the blastopore. Thus, a representative of the DPP/BMP2/4 subfamily of ligands was present in the common ancestor of diploblastic and triploblastic animals where it was probably expressed in a localized fashion during development. A localized source of DPP/BMP2/4 may have already been used in axis formation in this ancestor, or it may have provided a means by which an axis could evolve in triploblastic animals. PMID:12048233

Towards a standardization of biomethane potential tests.

PubMed

Holliger, Christof; Alves, Madalena; Andrade, Diana; Angelidaki, Irini; Astals, Sergi; Baier, Urs; Bougrier, Claire; Buffière, Pierre; Carballa, Marta; de Wilde, Vinnie; Ebertseder, Florian; Fernández, Belén; Ficara, Elena; Fotidis, Ioannis; Frigon, Jean-Claude; de Laclos, Hélène Fruteau; Ghasimi, Dara S M; Hack, Gabrielle; Hartel, Mathias; Heerenklage, Joern; Horvath, Ilona Sarvari; Jenicek, Pavel; Koch, Konrad; Krautwald, Judith; Lizasoain, Javier; Liu, Jing; Mosberger, Lona; Nistor, Mihaela; Oechsner, Hans; Oliveira, João Vítor; Paterson, Mark; Pauss, André; Pommier, Sébastien; Porqueddu, Isabella; Raposo, Francisco; Ribeiro, Thierry; Rüsch Pfund, Florian; Strömberg, Sten; Torrijos, Michel; van Eekert, Miriam; van Lier, Jules; Wedwitschka, Harald; Wierinck, Isabella

2016-12-01

Production of biogas from different organic materials is a most interesting source of renewable energy. The biomethane potential (BMP) of these materials has to be determined to get insight in design parameters for anaerobic digesters. Although several norms and guidelines for BMP tests exist, inter-laboratory tests regularly show high variability of BMPs for the same substrate. A workshop was held in June 2015, in Leysin, Switzerland, with over 40 attendees from 30 laboratories around the world, to agree on common solutions to the conundrum of inconsistent BMP test results. This paper presents the consensus of the intense roundtable discussions and cross-comparison of methodologies used in respective laboratories. Compulsory elements for the validation of BMP results were defined. They include the minimal number of replicates, the request to carry out blank and positive control assays, a criterion for the test duration, details on BMP calculation, and last but not least criteria for rejection of the BMP tests. Finally, recommendations on items that strongly influence the outcome of BMP tests such as inoculum characteristics, substrate preparation, test setup, and data analysis are presented to increase the probability of obtaining validated and reproducible results.

Quantifying Construction Site Sediment Discharge Risk and Treatment Potential

NASA Astrophysics Data System (ADS)

Ferrell, L.; Beighley, R. E.

2006-12-01

Dealing with soil erosion and sediment transport can be a significant challenge during the construction process due to the potentially large spatial and temporal extent and conditions of bare soils. Best Management Practices (BMP) are commonly used to eliminate or reduce offsite discharge of sediment. However, few efforts have investigated the time varying risk of sediment discharge from construction sites, which often have dynamic soil conditions and the potential for less than optimal BMP installations. The goal of this research is to improve the design, implementation and effectiveness of sediment and erosion control at construction sites using site specific, temporal distributions of sediment discharge risk. Sediment risk is determined from individual factors leading to sediment expert, such as rainfall frequency, the adequacy of BMP installations, and the extent and duration of bare soil conditions. This research specifically focuses on quantifying: (a) the effectiveness of temporary sediment and control erosion control BMPs in preventing, containing, and/or treating construction site sediment discharge at varying levels of "proper" installation, and (b) sediment discharge potential from construction sites during different phases of construction, (ex., disturbed earth operations). BMPs are evaluated at selected construction sites in southern California and at the Soil Erosion Research Laboratory (SERL) in the Civil and Environmental Engineering department at San Diego State University. SERL experiments are performed on a 3-m by 10-m tilting soil bed with soil depths up to 1 meter, slopes ranging from 0 to 50 percent, and rainfall rates up to 150 mm/hr (6 in/hr). BMP performance is assessed based on experiments where BMPs are installed per manufacture specifications, potential less than optimal installations, and no treatment conditions. Soil conditions are also varied to represent site conditions during different phases of construction (i.e., loose lifts, stock piles, temporary roads, finished grade, others). Preliminary site monitoring, experimental results, and a conceptual model for estimating the time depend risk of sediment discharge over the duration of a construction project are presented.

FRAMEWORK FOR PLACEMENT OF BMP/LID IN URBAN WATERSHEDS

EPA Science Inventory

To assist stormwater management professionals in planning for BMP/LID implementation, the U.S. Environmental Protection Agency (USEPA) has funded the development of a decision support system for selection and placement of BMP/LID at strategic locations in urban watersheds. The...

Enhanced Healing of Segmental Bone Defects by Modulation of the Mechanical Environment

DTIC Science & Technology

2012-10-01

5.5 µg BMP-2, it was largely disorganized, woven bone with non-osseous soft tissue interspersed. The highest 4 dose (11 µg) of BMP-2, in contrast...various doses of BMP-2. Top row: 16x magnification Bottom row: 100x magnification N= new cortex M= marrow T=trabecular bone F= fibrous tissue ...areas of cartilagenous tissue (figure 5) it was clear that substantial areas of cartilage remained in the defects treated with 5.5 µg BMP-2. These may

Enhanced Bone Formation in Segmental Defects with BMP2 in a Biologically Relevant Molecular Context

DTIC Science & Technology

2016-10-16

gun shots . These do not heal on their own once a ‘critical size’ segment of bone is missing. One strategy to induce healing is to use bone-inducing...accelerate BMP2-induced bone formation by presenting the growth factor in a more biologically relevant context. This is based on our observation...that manganese increases the binding of BMP2 to COMP. The next steps are to validate these observations using BMP2:COMP on HA/PLG scaffolds in-vitro

Heparin Microparticle Effects on Presentation and Bioactivity of Bone Morphogenetic Protein-2

PubMed Central

Hettiaratchi, Marian H.; Miller, Tobias; Temenoff, Johnna S.; Guldberg, Robert E.; McDevitt, Todd C.

2014-01-01

Biomaterials capable of providing localized and sustained presentation of bioactive proteins are critical for effective therapeutic growth factor delivery. However, current biomaterial delivery vehicles commonly suffer from limitations that can result in low retention of growth factors at the site of interest or adversely affect growth factor bioactivity. Heparin, a highly sulfated glycosaminoglycan, is an attractive growth factor delivery vehicle due to its ability to reversibly bind positively charged proteins, provide sustained delivery, and maintain protein bioactivity. This study describes the fabrication and characterization of heparin methacrylamide (HMAm) microparticles for recombinant growth factor delivery. HMAm microparticles were shown to efficiently bind several heparin-binding growth factors (e.g. bone morphogenetic protein-2 (BMP-2), vascular endothelial growth factor (VEGF), and basic fibroblast growth factor (FGF-2)), including a wide range of BMP-2 concentrations that exceeds the maximum binding capacity of other common growth factor delivery vehicles, such as gelatin. BMP-2 bioactivity was assessed on the basis of alkaline phosphatase (ALP) activity induced in skeletal myoblasts (C2C12). Microparticles loaded with BMP-2 stimulated comparable C2C12 ALP activity to soluble BMP-2 treatment, indicating that BMP-2-loaded microparticles retain bioactivity and potently elicit a functional cell response. In summary, our results suggest that heparin microparticles stably retain large amounts of bioactive BMP-2 for prolonged periods of time, and that presentation of BMP-2 via heparin microparticles can elicit cell responses comparable to soluble BMP-2 treatment. Consequently, heparin microparticles present an effective method of delivering and spatially retaining growth factors that could be used in a variety of systems to enable directed induction of cell fates and tissue regeneration. PMID:24881028

Involvement of over-expressed BMP4 in pentylenetetrazol kindling-induced cell proliferation in the dentate gyrus of adult rats

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yin Jinbo; Ma Yuxin; Yin Qing

2007-03-30

The dentate gyrus (DG) of the hippocampus is one of a few regions in the adult mammalian brain characterized by ongoing neurogenesis. Proliferation of neural precursors in the granule cell layer of the DG has been identified in pentylenetetrazol (PTZ) kindling epilepsy model, however, little is known about the molecular mechanism. We previously reported that the expression pattern of bone morphogenetic proteins-4 (BMP4) mRNA in the hippocampus was developmentally regulated and mainly localized in the DG of the adult. To explore the role of BMP4 in epileptic activity, we detected BMP4 expression in the DG during PTZ kindling process andmore » explore its correlation with cell proliferation combined with bromodeoxyuridine (BrdU) labeling technique. We found that dynamic changes in BMP4 level and BrdU labeled cells dependent on the kindling stage of PTZ induced seizure-prone state. The number of BMP4 mRNA-positive cells and BrdU labeled cells reached the top level 1 day after PTZ kindled, then declined to base level 2 months later. Furthermore, there was a significant correlation between increased BMP4 mRNA expression and increased number of BrdU labeled cells. After effectively blocked expression of BMP4 with antisense oligodeoxynucleotides(ASODN), the BrdU labeled cells in the dentate gyrus subgranular zone(DG-SGZ) and hilus were significantly decreased 16d after First PTZ injection and 1, 3, 7, 14d after kindled respectively. These findings suggest that increased proliferation in the DG of the hippocampus resulted from kindling epilepsy elicited by PTZ maybe be modulated by BMP4 over-expression.« less

BMP delivery complements the guiding effect of scaffold architecture without altering bone microstructure in critical-sized long bone defects: A multiscale analysis.

PubMed

Cipitria, A; Wagermaier, W; Zaslansky, P; Schell, H; Reichert, J C; Fratzl, P; Hutmacher, D W; Duda, G N

2015-09-01

Scaffold architecture guides bone formation. However, in critical-sized long bone defects additional BMP-mediated osteogenic stimulation is needed to form clinically relevant volumes of new bone. The hierarchical structure of bone determines its mechanical properties. Yet, the micro- and nanostructure of BMP-mediated fast-forming bone has not been compared with slower regenerating bone without BMP. We investigated the combined effects of scaffold architecture (physical cue) and BMP stimulation (biological cue) on bone regeneration. It was hypothesized that a structured scaffold directs tissue organization through structural guidance and load transfer, while BMP stimulation accelerates bone formation without altering the microstructure at different length scales. BMP-loaded medical grade polycaprolactone-tricalcium phosphate scaffolds were implanted in 30mm tibial defects in sheep. BMP-mediated bone formation after 3 and 12 months was compared with slower bone formation with a scaffold alone after 12 months. A multiscale analysis based on microcomputed tomography, histology, polarized light microscopy, backscattered electron microscopy, small angle X-ray scattering and nanoindentation was used to characterize bone volume, collagen fiber orientation, mineral particle thickness and orientation, and local mechanical properties. Despite different observed kinetics in bone formation, similar structural properties on a microscopic and sub-micron level seem to emerge in both BMP-treated and scaffold only groups. The guiding effect of the scaffold architecture is illustrated through structural differences in bone across different regions. In the vicinity of the scaffold increased tissue organization is observed at 3 months. Loading along the long bone axis transferred through the scaffold defines bone micro- and nanostructure after 12 months. Copyright © 2015 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

BMP-2-regenerated calvarial bone: a biomechanical appraisal in a large animal model.

PubMed

Cray, James; Henderson, Sarah E; Smith, Darren M; Kinsella, Christopher R; Bykowski, Michael; Cooper, Gregory M; Almarza, Alejandro J; Losee, Joseph E

2014-11-01

Recombinant human bone morphogenetic protein-2 (rhBMP-2) is gaining popularity in craniofacial applications. Calvarial defects are, under normal circumstances, subjected to only minimal levels of the biomechanical stresses known to play an important role in osteogenesis, yet regenerated calvarial bone must be capable of withstanding traumatic forces such that the underlying neurocapsule is protected. The aim of this study is to, for the first time, assess the biomechanical properties of calvarial bone regenerated with derivations of a commercially available rhBMP-2-based system. Standardized calvarial defects were created in 23 adult male canines. These defects were treated with rhBMP-2 on one of several carriers. After 24 weeks, the biomechanical properties of the rhBMP-2-generated bone were compared to those of controls with a modified punch-out test (Bluehill 2; Instron, Norwood, Mass) and compared using a paired nonparametric analyses (SPSS, 17.0, Chicago, Ill). In a previously published report, defects across all the rhBMP-2 therapy groups were observed to have a mean rate of 99.5% radio-opacity at 24 weeks indicating nearly full bony coverage of the calvarial defect (compared to 32.7% in surgical controls). For ultimate load, ultimate energy, and first peak energy, there were significant differences (P<0.05) with the control native bone having more robust biomechanical properties than the rhBMP-2-generated bone. We conclude from these findings that rhBMP-2-generated calvarial bone is significantly less protective against trauma than native bone at 6 months. Further investigation is required to assess the efficacy of rhBMP-2 in healing calvarial defects in the longer term.

Dexamethasone Enhances Osteogenic Differentiation of Bone Marrow- and Muscle-Derived Stromal Cells and Augments Ectopic Bone Formation Induced by Bone Morphogenetic Protein-2

PubMed Central

Yuasa, Masato; Yamada, Tsuyoshi; Taniyama, Takashi; Masaoka, Tomokazu; Xuetao, Wei; Yoshii, Toshitaka; Horie, Masaki; Yasuda, Hiroaki; Uemura, Toshimasa; Okawa, Atsushi; Sotome, Shinichi

2015-01-01

We evaluated whether dexamethasone augments the osteogenic capability of bone marrow-derived stromal cells (BMSCs) and muscle tissue-derived stromal cells (MuSCs), both of which are thought to contribute to ectopic bone formation induced by bone morphogenetic protein-2 (BMP-2), and determined the underlying mechanisms. Rat BMSCs and MuSCs were cultured in growth media with or without 10-7 M dexamethasone and then differentiated under osteogenic conditions with dexamethasone and BMP-2. The effects of dexamethasone on cell proliferation and osteogenic differentiation, and also on ectopic bone formation induced by BMP-2, were analyzed. Dexamethasone affected not only the proliferation rate but also the subpopulation composition of BMSCs and MuSCs, and subsequently augmented their osteogenic capacity during osteogenic differentiation. During osteogenic induction by BMP-2, dexamethasone also markedly affected cell proliferation in both BMSCs and MuSCs. In an in vivo ectopic bone formation model, bone formation in muscle-implanted scaffolds containing dexamethasone and BMP-2 was more than two fold higher than that in scaffolds containing BMP-2 alone. Our results suggest that dexamethasone potently enhances the osteogenic capability of BMP-2 and may thus decrease the quantity of BMP-2 required for clinical application, thereby reducing the complications caused by excessive doses of BMP-2. Highlights: 1. Dexamethasone induced selective proliferation of bone marrow- and muscle-derived cells with higher differentiation potential. 2. Dexamethasone enhanced the osteogenic capability of bone marrow- and muscle-derived cells by altering the subpopulation composition. 3. Dexamethasone augmented ectopic bone formation induced by bone morphogenetic protein-2. PMID:25659106

Dura Mater Stimulates Human Adipose-Derived Stromal Cells to Undergo Bone Formation in Mouse Calvarial Defects

PubMed Central

Levi, Benjamin; Nelson, Emily R.; Li, Shuli; James, Aaron W.; Hyun, Jeong S.; Montoro, Daniel T.; Lee, Min; Glotzbach, Jason P.; Commons, George W.; Longaker, Michael T.

2015-01-01

Human adipose-derived stromal cells (hASCs) have a proven capacity to aid in osseous repair of calvarial defects. However, the bone defect microenvironment necessary for osseous healing is not fully understood. In this study, we postulated that the cell-cell interaction between engrafted ASCs and host dura mater (DM) cells is critical for the healing of calvarial defects. hASCs were engrafted into critical sized calvarial mouse defects. The DM-hASC interaction was manipulated surgically by DM removal or by insertion of a semipermeable or nonpermeable membrane between DM and hASCs. Radiographic, histologic, and gene expression analyses were performed. Next, the hASC-DM interaction is assessed by conditioned media (CM) and coculture assays. Finally, bone morphogenetic protein (BMP) signaling from DM was investigated in vivo using novel BMP-2 and anti-BMP-2/4 slow releasing scaffolds. With intact DM, osseous healing occurs both from host DM and engrafted hASCs. Interference with the DM-hASC interaction dramatically reduced calvarial healing with abrogated BMP-2–Smad-1/5 signaling. Using CM and coculture assays, mouse DM cells stimulated hASC osteogenesis via BMP signaling. Through in vivo manipulation of the BMP-2 pathway, we found that BMP-2 plays an important role in DM stimulation of hASC osteogenesis in the context of calvarial bone healing. BMP-2 supplementation to a defect with disrupted DM allowed for bone formation in a nonhealing defect. DM is an osteogenic cell type that both participates in and stimulates osseous healing in a hASC-engrafted calvarial defect. Furthermore, DM-derived BMP-2 paracrine stimulation appears to play a key role for hASC mediated repair. PMID:21656608

Differences in fat and muscle mass associated with a functional human polymorphism in a post-transcriptional BMP2 gene regulatory element.

PubMed

Devaney, Joseph M; Tosi, Laura L; Fritz, David T; Gordish-Dressman, Heather A; Jiang, Shan; Orkunoglu-Suer, Funda E; Gordon, Andrew H; Harmon, Brennan T; Thompson, Paul D; Clarkson, Priscilla M; Angelopoulos, Theodore J; Gordon, Paul M; Moyna, Niall M; Pescatello, Linda S; Visich, Paul S; Zoeller, Robert F; Brandoli, Cinzia; Hoffman, Eric P; Rogers, Melissa B

2009-08-15

A classic morphogen, bone morphogenetic protein 2 (BMP2) regulates the differentiation of pluripotent mesenchymal cells. High BMP2 levels promote osteogenesis or chondrogenesis and low levels promote adipogenesis. BMP2 inhibits myogenesis. Thus, BMP2 synthesis is tightly controlled. Several hundred nucleotides within the 3' untranslated regions of BMP2 genes are conserved from mammals to fishes indicating that the region is under stringent selective pressure. Our analyses indicate that this region controls BMP2 synthesis by post-transcriptional mechanisms. A common A to C single nucleotide polymorphism (SNP) in the BMP2 gene (rs15705, +A1123C) disrupts a putative post-transcriptional regulatory motif within the human ultra-conserved sequence. In vitro studies indicate that RNAs bearing the A or C alleles have different protein binding characteristics in extracts from mesenchymal cells. Reporter genes with the C allele of the ultra-conserved sequence were differentially expressed in mesenchymal cells. Finally, we analyzed MRI data from the upper arm of 517 healthy individuals aged 18-41 years. Individuals with the C/C genotype were associated with lower baseline subcutaneous fat volumes (P = 0.0030) and an increased gain in skeletal muscle volume (P = 0.0060) following resistance training in a cohort of young males. The rs15705 SNP explained 2-4% of inter-individual variability in the measured parameters. The rs15705 variant is one of the first genetic markers that may be exploited to facilitate early diagnosis, treatment, and/or prevention of diseases associated with poor fitness. Furthermore, understanding the mechanisms by which regulatory polymorphisms influence BMP2 synthesis will reveal novel pharmaceutical targets for these disabling conditions. (c) 2009 Wiley-Liss, Inc.

Differences in Fat and Muscle Mass Associated With a Functional Human Polymorphism in a Post-Transcriptional BMP2 Gene Regulatory Element

PubMed Central

Devaney, Joseph M.; Tosi, Laura L.; Fritz, David T.; Gordish-Dressman, Heather A.; Jiang, Shan; Orkunoglu-Suer, Funda E.; Gordon, Andrew H.; Harmon, Brennan T.; Thompson, Paul D.; Clarkson, Priscilla M.; Angelopoulos, Theodore J.; Gordon, Paul M.; Moyna, Niall M.; Pescatello, Linda S.; Visich, Paul S.; Zoeller, Robert F.; Brandoli, Cinzia; Hoffman, Eric P.; Rogers, Melissa B.

2014-01-01

A classic morphogen, bone morphogenetic protein 2 (BMP2) regulates the differentiation of pluripotent mesenchymal cells. High BMP2 levels promote osteogenesis or chondrogenesis and low levels promote adipogenesis. BMP2 inhibits myogenesis. Thus, BMP2 synthesis is tightly controlled. Several hundred nucleotides within the 3′ untranslated regions of BMP2 genes are conserved from mammals to fishes indicating that the region is under stringent selective pressure. Our analyses indicate that this region controls BMP2 synthesis by post-transcriptional mechanisms. A common A to C single nucleotide polymorphism (SNP) in the BMP2 gene (rs15705, +A1123C) disrupts a putative post-transcriptional regulatory motif within the human ultra-conserved sequence. In vitro studies indicate that RNAs bearing the A or C alleles have different protein binding characteristics in extracts from mesenchymal cells. Reporter genes with the C allele of the ultra-conserved sequence were differentially expressed in mesenchymal cells. Finally, we analyzed MRI data from the upper arm of 517 healthy individuals aged 18–41 years. Individuals with the C/C genotype were associated with lower baseline subcutaneous fat volumes (P = 0.0030) and an increased gain in skeletal muscle volume (P = 0.0060) following resistance training in a cohort of young males. The rs15705 SNP explained 2–4% of inter-individual variability in the measured parameters. The rs15705 variant is one of the first genetic markers that maybe exploited to facilitate early diagnosis, treatment, and/or prevention of diseases associated with poor fitness. Furthermore, understanding the mechanisms by which regulatory polymorphisms influence BMP2 synthesis will reveal novel pharmaceutical targets for these disabling conditions. PMID:19492344

Expression of Noggin and Gremlin1 and its implications in fine-tuning BMP activities in mouse cartilage tissues.

PubMed

Yu, Xiaodan; Kawakami, Hiroko; Tahara, Naoyuki; Olmer, Merissa; Hayashi, Shinichi; Akiyama, Ryutaro; Bagchi, Anindya; Lotz, Martin; Kawakami, Yasuhiko

2017-08-01

Increasing evidence supports the idea that bone morphogenetic proteins (BMPs) regulate cartilage maintenance in the adult skeleton. The aim of this study is to obtain insight into the regulation of BMP activities in the adult skeletal system. We analyzed expression of Noggin and Gremlin1, BMP antagonists that are known to regulate embryonic skeletal development, in the adult skeletal system by Noggin-LacZ and Gremlin1-LacZ knockin reporter mouse lines. Both reporters are expressed in the adult skeleton in a largely overlapping manner with some distinct patterns. Both are detected in the articular cartilage, pubic symphysis, facet joint in the vertebrae, and intervertebral disk, suggesting that they regulate BMP activities in these tissues. In a surgically induced knee osteoarthritis model in mice, expression of Noggin mRNA was lost from the articular cartilage, which correlated with loss of BMP2/4 and pSMAD1/5/8, an indicator of active BMP signaling. Both reporters are also expressed in the sterna and rib cartilage, suggesting an extensive role of BMP antagonism in adult cartilage tissue. Moreover, Noggin-LacZ was detected in sutures in the skull and broadly in the nasal cartilage, while Gremlin1-LacZ exhibits a weaker and more restricted expression domain in the nasal cartilage. These results suggest broad regulation of BMP activities by Noggin and Gremlin1 in cartilage tissues in the adult skeleton, and that BMP signaling and its antagonism by NOGGIN play a role in osteoarthritis development. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:1671-1682, 2017. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc.

Bone morphogenetic protein-mediated interaction of periosteum and diaphysis. Citric acid and other factors influencing the generation of parosteal bone.

PubMed

Kübler, N; Urist, M R

1990-09-01

In rabbits, after long-bone growth is complete and the cambium layer regresses, mesenchymal-type cells with embryonic potential (competence) for bone development persist in the adventitial layer of periosteum. These cells are not determined osteoprogenitor cells (stem cells) because bone tissue differentiation does not occur when adult periosteum is transplanted into a heterotopic site. In this respect, adventitial cells differ from bone marrow stroma cells. In a parosteal orthotopic site in the space between the adult periosteum and diaphysis, implants of bone morphogenetic protein (BMP) and associated noncollagenous proteins (BMP/NCP) induce adventitia and adjacent muscle connective-tissue-derived cells to switch from a fibrogenetic to a chondroosteoprogenetic pattern of bone development. The quantity of induced bone is proportional to the dose of BMP/NCP in the range from 10 to 50 mg; immature rabbits produced larger deposits than mature rabbits in response to BMP/NCP. Preoperative local intramuscular injections of citric, edetic, or hyaluronic acids in specified concentrations markedly enhanced subperiosteal BMP/NCP-induced bone formation. The quantity of bovine or human BMP/NCP-induced bone formation in rabbits is also increased by very low-dose immunosuppression but not by bone mineral, tricalcium phosphate ceramic, inorganic calcium salts, or various space-occupying, unspecific chemical irritants. Although composities of BMP/NCP and allogeneic rabbit tendon collagen increased the quantity of bone in a parosteal site, in a heterotopic site the composite failed to induce bone formation. In a parosteal site, the conditions permitting BMP/NCP-induced bone formation develop, and the end product of the morphogenetic response is a duplicate diaphysis. How BMP reactivates the morphogenetic process in postfetal mesenchymal-type adventitial cells persisting in adult periosteum (including adjacent muscle attachments) is not known.

Bone Regeneration Using N-Methyl-2-pyrrolidone as an Enhancer for Recombinant Human Bone Morphogenetic Protein-2 in a Rabbit Sinus Augmentation Model

PubMed Central

Thoma, Daniel S.

2017-01-01

The aim of this study was to determine whether N-methyl-2-pyrrolidone (NMP) can decrease the dose of recombinant human bone morphogenetic protein-2 (rhBMP-2) in sinus augmentation of rabbits. In each of 15 rabbits, 2 sinuses were randomly grafted using 1 of 3 treatment modalities: (i) biphasic calcium phosphate (BCP; control), (ii) rhBMP-2-coated BCP (BMP), or (iii) rhBMP-2-coated BCP soaked in NMP solution (BMP/NMP). The rabbits were sacrificed 2 weeks postoperatively. Histologic and histomorphometric analyses were performed. Bone formation in all groups was predominantly located close to the access window and the lateral walls. Newly formed bone within the total augmented area (NBTA) was greatest in BMP/NMP (1.94 ± 0.69 mm2), followed by BMP (1.50 ± 0.72 mm2) and BCP (1.28 ± 0.52 mm2) (P > 0.05). In the center of the augmentation (NBROI_C) and the area close to the sinus membrane (NBROI_M), BMP/NMP produced the largest area of NB (NBROI_C: 0.10 ± 0.11 mm2; NBROI_M: 0.17 ± 0.08 mm2); the corresponding NB values for BCP were 0.05 ± 0.05 mm2 and 0.08 ± 0.09 mm2, respectively (P > 0.05 for all comparisons). The effect of NMP on bone regeneration was inconsistent between the specimens. Adding NMP as an adjunct to rhBMP-2-coated BCP produced inconsistent effects on bone regeneration, resulting in no significant benefit compared to controls. PMID:28680881

Bone morphogenetic protein-7 (BMP-7) augments insulin sensitivity in mice with type II diabetes mellitus by potentiating PI3K/AKT pathway.

PubMed

Chattopadhyay, Tandrika; Singh, Rajiv Ranjan; Gupta, Sarika; Surolia, Avadhesha

2017-03-01

A direct link between development of insulin resistance and the presence of chronic inflammation, in case of obesity exists, with cytokines playing an important role in glucose metabolism. Members of TGF-β superfamily, including bone morphogenetic proteins (BMPs), have been shown to be involved in islet morphogenesis, establishment of β-cell mass and adipose cell fate determination. Here, we demonstrate a novel and direct role of BMP-4 and -7 in the regulation of glucose homeostasis and insulin resistance. An age-dependent increase in serum BMP-4 and decrease in serum BMP-7 levels was observed in animal models of type II diabetes. In this study, BMP-7 and -4 have been demonstrated to have antagonistic effects on insulin signaling and thereby on glucose homeostasis. BMP-7 augmented glucose uptake in the insulin sensitive tissues such as the adipose and muscle by increasing Glut4 translocation to the plasma membrane through phosphorylation and activation of PDK1 and Akt, and phosphorylation and translocation of FoxO1 to the cytoplasm in liver/HepG2 cells. Restoration of BMP-7 levels in serum of diabetic animals resulted in decreased blood glucose levels in contrast to age matched untreated control groups, opening up a new therapeutic avenue for diabetes. On the contrary, BMP-4 inhibited insulin signaling through activation of PKC-θ isoform, and resulted in insulin resistance through the attenuation of insulin signaling. BMP-7 therefore is an attractive candidate for tackling a multifaceted disease such as diabetes, since it not only reduces body fat, but also strengthens insulin signaling, causing improved glucose uptake and ameliorating peripheral insulin resistance. © 2017 BioFactors, 43(2):195-209, 2017. © 2017 International Union of Biochemistry and Molecular Biology.

Local treatment of cancellous bone grafts with BMP-7 and zoledronate increases both the bone formation rate and bone density

PubMed Central

2011-01-01

Background and purpose The remodeling of morselized bone grafts in revision surgery can be enhanced by an anabolic substance such as a bone morphogenetic protein (BMP). On the other hand, BMPs boost catabolism and might cause a premature resorption, both of the graft and of the new-formed bone. Bisphosphonates inactivate osteoclasts and can be used to control the resorption. We studied a combination of both drugs as a local admix to a cancellous allograft. Methods Cancellous bone allografts were harvested and freeze-dried. Either saline, BMP-7, the bisphosphonate zoledronate, or a combination of BMP-7 and zoledronate were added in solution. The grafts were placed in bone conduction chambers and implanted in the proximal tibia of 34 rats. The grafts were harvested after 6 weeks and evaluated by histomorphometry. Results Bone volume/total volume (BV/TV) was 50% in the grafts treated with the combination of BMP-7 and zoledronate and 16% in the saline controls (p < 0.001). In the zoledronate group BV/TV was 56%, and in the BMP group it was 14%. The ingrowth distance of new bone into the graft was 3.5 mm for the combination of BMP-7 and zoledronate and 2.6 mm in the saline control (p = 0.002). The net amount of retained remodeled bone was more than 4 times higher when BMP-7 and zoledronate were combined than in the controls. Interpretation An anabolic drug like BMP-7 can be combined with an anti-catabolic bisphosphonate as local bone graft adjunct, and the combination increases the amount of remaining bone after remodeling is complete. PMID:21434769

The Efficacy of Cyclic Injection of Bone Morphogenetic Protein-2 in Large-Scale Calvarial Bone Defects.

PubMed

Choi, Jin Mi; Jeong, Woo Shik; Park, Eun Jung; Choi, Jong Woo

2017-03-01

Bone morphogenetic protein-2 (BMP-2) appears to be one of the most potent growth factors thus far studied. However, recent publications on the clinical application of BMP-2 revealed that its correct control is the paramount issue in clinical practice. For improving BMP-2 delivery, the cyclic administration might be an alternative. Accordingly, the authors cyclically injected BMP-2 in a cyclic injection model of large cranial defects to maintain the proper dosage during the bone healing process. A 10-mm diameter calvarial bone defect was produced using a round drill in 8-week-old Sprague-Dawley rats. Silk-hydroxyapatite scaffolds soaked in the appropriate concentration of BMP-2 were implanted into the defect. The animals were split into 4 single-injection groups and 3 multiple-injection groups; the latter groups received weekly subcutaneous injections of BMP-2 solution (1, 5, and 10 μg/mL) for 4 weeks, whereas the former groups received a single injection of BMP-2 at these concentrations. Each rat underwent computed tomography at 8 weeks. In terms of total volumes of the new bone, the 5 μg/mL multiple-injection BMP-2 group had significantly greater increases in bone volume than the single-injection groups. In terms of bone thickness, the multiple-injection groups had better outcomes than the single-injection groups. Thus, the cyclic injection protocol restored the original thickness without overgrowth. Cyclic injection of BMP-2 permits more accurate dosage control than single injection and improves thickness and dense bone regeneration. Therefore, it may represent a promising approach for future clinical trials. Further investigation using a greater number of animals is required.

Metastatic function of BMP-2 in gastric cancer cells: The role of PI3K/AKT, MAPK, the NF-{kappa}B pathway, and MMP-9 expression

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kang, Myoung Hee; Oh, Sang Cheul; Lee, Hyun Joo

2011-07-15

Bone morphogenetic proteins (BMPs) have been implicated in tumorigenesis and metastatic progression in various types of cancer cells, but the role and cellular mechanism in the invasive phenotype of gastric cancer cells is not known. Herein, we determined the roles of phosphoinositide 3-kinase (PI3K)/AKT, extracellular signal-regulated protein kinase (ERK), nuclear factor (NF)-{kappa}B, and matrix metalloproteinase (MMP) expression in BMP-2-mediated metastatic function in gastric cancer. We found that stimulation of BMP-2 in gastric cancer cells enhanced the phosphorylation of AKT and ERK. Accompanying activation of AKT and ERK kinase, BMP-2 also enhanced phosphorylation/degradation of I{kappa}B{alpha} and the nuclear translocation/activation of NF-{kappa}B.more » Interestingly, blockade of PI3K/AKT and ERK signaling using LY294002 and PD98059, respectively, significantly inhibited BMP-2-induced motility and invasiveness in association with the activation of NF-{kappa}B. Furthermore, BMP-2-induced MMP-9 expression and enzymatic activity was also significantly blocked by treatment with PI3K/AKT, ERK, or NF-{kappa}B inhibitors. Immunohistochemistry staining of 178 gastric tumor biopsies indicated that expression of BMP-2 and MMP-9 had a significant positive correlation with lymph node metastasis and a poor prognosis. These results indicate that the BMP-2 signaling pathway enhances tumor metastasis in gastric cancer by sequential activation of the PI3K/AKT or MAPK pathway followed by the induction of NF-{kappa}B and MMP-9 activity, indicating that BMP-2 has the potential to be a therapeutic molecular target to decrease metastasis.« less

Characterisation of the biochemical methane potential (BMP) of individual material fractions in Danish source-separated organic household waste.

PubMed

Naroznova, Irina; Møller, Jacob; Scheutz, Charlotte

2016-04-01

This study is dedicated to characterising the chemical composition and biochemical methane potential (BMP) of individual material fractions in untreated Danish source-separated organic household waste (SSOHW). First, data on SSOHW in different countries, available in the literature, were evaluated and then, secondly, laboratory analyses for eight organic material fractions comprising Danish SSOHW were conducted. No data were found in the literature that fully covered the objectives of the present study. Based on laboratory analyses, all fractions were assigned according to their specific properties in relation to BMP, protein content, lipids, lignocellulose biofibres and easily degradable carbohydrates (carbohydrates other than lignocellulose biofibres). The three components in lignocellulose biofibres, i.e. lignin, cellulose and hemicellulose, were differentiated, and theoretical BMP (TBMP) and material degradability (BMP from laboratory incubation tests divided by TBMP) were expressed. Moreover, the degradability of lignocellulose biofibres (the share of volatile lignocellulose biofibre solids degraded in laboratory incubation tests) was calculated. Finally, BMP for average SSOHW composition in Denmark (untreated) was calculated, and the BMP contribution of the individual material fractions was then evaluated. Material fractions of the two general waste types, defined as "food waste" and "fibre-rich waste," were found to be anaerobically degradable with considerable BMP. Material degradability of material fractions such as vegetation waste, moulded fibres, animal straw, dirty paper and dirty cardboard, however, was constrained by lignin content. BMP for overall SSOHW (untreated) was 404 mL CH4 per g VS, which might increase if the relative content of material fractions, such as animal and vegetable food waste, kitchen tissue and dirty paper in the waste, becomes larger. Copyright © 2016 Elsevier Ltd. All rights reserved.

Smad7 mediates inhibition of Saos2 osteosarcoma cell differentiation by NF{kappa}B

DOE Office of Scientific and Technical Information (OSTI.GOV)

Eliseev, Roman A.; Schwarz, Edward M.; Zuscik, Michael J.

2006-01-01

The transcription factor NF{kappa}B is constitutively activated in various tumor cells where it promotes proliferation and represses apoptosis. The bone morphogenetic proteins (BMPs) delay cell proliferation and promote differentiation and apoptosis of bone cells through activation of Smad downstream effectors and via Smad-independent mechanisms. Thus, NF{kappa}B and BMP pathways play opposing roles in regulating osteoblastic cell fate. Here, we show that in osteosarcoma Saos2 osteoblasts, NF{kappa}B regulates the activity of the BMP/Smad signaling. Inhibition of NF{kappa}B by overexpression of mI{kappa}B leads to the induction of osteoblast differentiation. Saos2 cells overexpressing mI{kappa}B (Saos2-mI{kappa}B) exhibit higher expression of osteoblast phenotypic genes suchmore » as alkaline phosphatase, Runx2 and osteocalcin and are more responsive to BMP2 in comparison to wild-type cells (Saos2-wt) or empty vector infected controls (Saos2-EV). Furthermore, BMP-2 signaling and Smad phosphorylation are significantly increased in Saos2-mI{kappa}B cells in comparison to Saos2-EV cells. Inhibition of NF{kappa}B signaling in Saos2-mI{kappa}B cells is associated with decreased expression of the BMP signaling inhibitor Smad7. While gain of Smad7 function in Saos2-mI{kappa}B cells results in inhibition of BMP signaling, anti-sense knockdown of Smad7 in Saos2-EV cells leads to upregulation of BMP signaling. We therefore conclude that in osteosarcoma Saos2 cells, NF{kappa}B represses BMP/Smad signaling and BMP2-induced differentiation through Smad7.« less

Bone Regeneration Using N-Methyl-2-pyrrolidone as an Enhancer for Recombinant Human Bone Morphogenetic Protein-2 in a Rabbit Sinus Augmentation Model.

PubMed

Lim, Hyun-Chang; Thoma, Daniel S; Yoon, So-Ra; Cha, Jae-Kook; Lee, Jung-Seok; Jung, Ui-Won

2017-01-01

The aim of this study was to determine whether N-methyl-2-pyrrolidone (NMP) can decrease the dose of recombinant human bone morphogenetic protein-2 (rhBMP-2) in sinus augmentation of rabbits. In each of 15 rabbits, 2 sinuses were randomly grafted using 1 of 3 treatment modalities: (i) biphasic calcium phosphate (BCP; control), (ii) rhBMP-2-coated BCP (BMP), or (iii) rhBMP-2-coated BCP soaked in NMP solution (BMP/NMP). The rabbits were sacrificed 2 weeks postoperatively. Histologic and histomorphometric analyses were performed. Bone formation in all groups was predominantly located close to the access window and the lateral walls. Newly formed bone within the total augmented area (NB TA ) was greatest in BMP/NMP (1.94 ± 0.69 mm 2 ), followed by BMP (1.50 ± 0.72 mm 2 ) and BCP (1.28 ± 0.52 mm 2 ) ( P > 0.05). In the center of the augmentation (NB ROI_C ) and the area close to the sinus membrane (NB ROI_M ), BMP/NMP produced the largest area of NB (NB ROI_C : 0.10 ± 0.11 mm 2 ; NB ROI_M : 0.17 ± 0.08 mm 2 ); the corresponding NB values for BCP were 0.05 ± 0.05 mm 2 and 0.08 ± 0.09 mm 2 , respectively ( P > 0.05 for all comparisons). The effect of NMP on bone regeneration was inconsistent between the specimens. Adding NMP as an adjunct to rhBMP-2-coated BCP produced inconsistent effects on bone regeneration, resulting in no significant benefit compared to controls.

Effects of surface area and inflow on the performance of stormwater best management practices with uncertainty analysis.

PubMed

Park, Daeryong; Roesner, Larry A

2013-09-01

The performance of stormwater best management practices (BMPs) is affected by BMP geometric and hydrologic factors. The objective of this study was to investigate the effect of BMP surface area and inflow on BMP performance using the k-C* model with uncertainty analysis. Observed total suspended solids (TSS) from detention basins and retention ponds data sets in the International Stormwater BMP Database were used to build and evaluate the model. Detention basins are regarded as dry ponds because they do not always have water, whereas retention ponds have a permanent pool and are considered wet ponds. In this study, Latin hypercube sampling (LHS) was applied to consider uncertainty in both influent event mean concentration (EMC), C(in), and the areal removal constant, k. The latter was estimated from the hydraulic loading rate, q, through use of a power function relationship. Results show that effluent EMC, C(out), decreased as inflow decreased and as BMP surface area increased in both detention basins and retention ponds. However, the change in C(out), depending on inflow and BMP surface area for detention basins, differed from the change in C(out) for retention ponds. Specifically, C(in) was more dominantly associated with the performance of the k-C* model of detention basins than were BMP surface area and inflow. For retention ponds, however, results suggest that BMP surface area and inflow both influenced changes in C(out) as well as C(in). These results suggest that sensitive factors in the performance of the k-C* model are limited to C(in) for detention basins, whereas BMP surface area, inflow, and C(in) are important for retention ponds.

BMP15 suppresses progesterone production by down-regulating StAR via ALK3 in human granulosa cells.

PubMed

Chang, Hsun-Ming; Cheng, Jung-Chien; Klausen, Christian; Leung, Peter C K

2013-12-01

In addition to somatic cell-derived growth factors, oocyte-derived growth differentiation factor (GDF)9 and bone morphogenetic protein (BMP)15 play essential roles in female fertility. However, few studies have investigated their effects on human ovarian steroidogenesis, and fewer still have examined their differential effects or underlying molecular determinants. In the present study, we used immortalized human granulosa cells (SVOG) and human granulosa cell tumor cells (KGN) to compare the effects of GDF9 and BMP15 on steroidogenic enzyme expression and investigate potential mechanisms of action. In SVOG cells, neither GDF9 nor BMP15 affects the mRNA levels of P450 side-chain cleavage enzyme or 3β-hydroxysteroid dehydrogenase. However, treatment with BMP15, but not GDF9, significantly decreases steroidogenic acute regulatory protein (StAR) mRNA and protein levels as well as progesterone production. These suppressive effects, along with the induction of Sma and Mad-related protein (SMAD)1/5/8 phosphorylation, are attenuated by cotreatment with 2 different BMP type I receptor inhibitors (dorsomorphin and DMH-1). Furthermore, depletion of activin receptor-like kinase (ALK)3 using small interfering RNA reverses the effects of BMP15 on SMAD1/5/8 phosphorylation and StAR expression. Similarly, knockdown of ALK3 abolishes BMP15-induced SMAD1/5/8 phosphorylation in KGN cells. These results provide evidence that oocyte-derived BMP15 down-regulates StAR expression and decreases progesterone production in human granulosa cells, likely via ALK3-mediated SMAD1/5/8 signaling. Our findings suggest that oocyte may play a critical role in the regulation of progesterone to prevent premature luteinization during the late stage of follicle development.

Novel osteoinductive photo-cross-linkable chitosan-lactide-fibrinogen hydrogels enhance bone regeneration in critical size segmental bone defects

PubMed Central

Kim, Sungwoo; Bedigrew, Katherine; Guda, Teja; Maloney, William J.; Park, Sangwon; Wenke, Joseph C.; Yang, Yunzhi Peter

2014-01-01

The purpose of this study was to develop and characterize a novel photo-cross-linkable chitosan-lactide-fibrinogen (CLF) hydrogel and evaluate the efficacy of bone morphogenetic protein-2 (BMP-2) containing CLF hydrogel for osteogenesis in vitro and in vivo. We synthesized the CLF hydrogels and characterized their chemical structure, degradation rate, compressive modulus, and in vitro BMP-2 release kinetics. We evaluated bioactivities of the BMP-2 containing CLF hydrogels (0, 50, 100, and 500 ng/ml) in vitro using W-20-17 preosteoblast mouse bone marrow stromal cells and C2C12 mouse myoblast cells. The effect of BMP-2 containing CLF gels (0, 0.5, 1, 2, and 5μg) on bone formation was evaluated using rat critical size segmental bone defects for 4 weeks. FTIR spectra and SEM images showed chemical and structural changes by addition of fibrinogen into chitosan-lactide copolymer. Incorporation of fibrinogen molecules significantly increased compressive modulus of the hydrogels. In vitro BMP-2 release study showed initial burst releases from the CLF hydrogels followed by sustained releases, regardless of the concentration of the BMP-2 over 4 weeks. Cells in all groups were viable in the presence of the hydrogels regardless of BMP-2 doses, indicating non-cytotoxicity of hydrogels. Alkaline phosphate activity and mineralization of cells exhibited dose dependence on BMP-2 containing CLF hydrogels. Radiographs, microcomputed tomography, and histology confirmed that the BMP-2 containing CLF hydrogels prompted neo-osteogenesis and accelerated healing of the defects in a dose-dependent manner. Thus the CLF hydrogel is a promising delivery system of growth factors for bone regeneration. PMID:25174669

Paracrine and autocrine signals promoting full chondrogenic differentiation of a mesoblastic cell line.

PubMed

Locker, Morgane; Kellermann, Odile; Boucquey, Marie; Khun, Huot; Huerre, Michel; Poliard, Anne

2004-01-01

The pluripotent mesoblastic C1 cell line was used under serum-free culture conditions to investigate how paracrine and autocrine signals cooperate to drive chondrogenesis. Sequential addition of two systemic hormones, dexamethasone and triiodothyronine, permits full chondrogenic differentiation. The cell intrinsic activation of the BMP signaling pathway and Sox9 expression occurring on mesoblastic condensation is insufficient for recruitment of the progenitors. Dexamethasone-dependent Sox9 upregulation is essential for chondrogenesis. Differentiation of lineage stem cells relies on cell autonomous regulations modulated by external signals. We used the pluripotent mesoblastic C1 cell line under serum-free culture conditions to investigate how paracrine and autocrine signals cooperate to induce differentiation of a precursor clone along the chondrogenic lineage. C1 cells, cultured as aggregates, were induced toward chondrogenesis by addition of 10(-7) M dexamethasone in serum-free medium. After 30 days, dexamethasone was replaced by 10 nM triiodothyronine to promote final hypertrophic conversion. Mature and hypertrophic phenotypes were characterized by immunocytochemistry using specific antibodies against types II and X collagens, respectively. Type II collagen, bone morphogenetic proteins (BMPs), BMP receptors, Smads, and Sox9 expression were monitored by reverse transcriptase-polymerase chain reaction (RT-PCR), Northern blot, and/or Western blot analysis. Once C1 cells have formed nodules, sequential addition of two systemic hormones is sufficient to promote full chondrogenic differentiation. In response to dexamethasone, nearly 100% of the C1 precursors engage in chondrogenesis and convert within 30 days into mature chondrocytes, which triggers a typical cartilage matrix. On day 25, a switch in type II procollagen mRNA splicing acted as a limiting step in the acquisition of the mature chondrocyte phenotype. On day 30, substitution of dexamethasone with triiodothyronine triggers the final differentiation into hypertrophic chondrocytes within a further 15 days. The chondrogenic process is supported by intrinsic expression of Sox9 and BMP family genes. Similarly to the in vivo situation, activation of Sox9 expression and the BMP signaling pathway occurred on mesoblastic condensation. After induction, BMP-activated Smad nuclear translocation persisted throughout the process until the onset of hypertrophy. After dexamethasone addition, Sox9 expression was upregulated. Dexamethasone withdrawal reversed the increase in Sox9 expression and stopped differentiation. Thus, Sox9 seems to be a downstream mediator of dexamethasone action.

Assaying Oxidative Coupling Activity of CYP450 Enzymes.

PubMed

Agarwal, Vinayak

2018-01-01

Cytochrome P450 (CYP450) enzymes are ubiquitous catalysts in natural product biosynthetic schemes where they catalyze numerous different transformations using radical intermediates. In this protocol, we describe procedures to assay the activity of a marine bacterial CYP450 enzyme Bmp7 which catalyzes the oxidative radical coupling of polyhalogenated aromatic substrates. The broad substrate tolerance of Bmp7, together with rearrangements of the aryl radical intermediates leads to a large number of products to be generated by the enzymatic action of Bmp7. The complexity of the product pool generated by Bmp7 thus presents an analytical challenge for structural elucidation. To address this challenge, we describe mass spectrometry-based procedures to provide structural insights into aryl crosslinked products generated by Bmp7, which can complement subsequent spectroscopic experiments. Using the procedures described here, for the first time, we show that Bmp7 can efficiently accept polychlorinated aryl substrates, in addition to the physiological polybrominated substrates for the biosynthesis of polyhalogenated marine natural products. © 2018 Elsevier Inc. All rights reserved.

BMP signaling protects telencephalic fate by repressing eye identity and its Cxcr4-dependent morphogenesis.

PubMed

Bielen, Holger; Houart, Corinne

2012-10-16

Depletion of Wnt signaling is a major requirement for the induction of the anterior prosencephalon. However, the molecular events driving the differential regionalization of this area into eye-field and telencephalon fates are still unknown. Here we show that the BMP pathway is active in the anterior neural ectoderm during late blastula to early gastrula stage in zebrafish. Bmp2b mutants and mosaic loss-of-function experiments reveal that BMP acts as a repressor of eye-field fate through inhibition of its key transcription factor Rx3, thereby protecting the future telencephalon from acquiring eye identity. This BMP-driven mechanism initiates the establishment of the telencephalon prior to the involvement of Wnt antagonists from the anterior neural border. Furthermore, we demonstrate that Rx3 and BMP are respectively required to maintain and restrict the chemokine receptor cxcr4a, which in turn contributes to the morphogenetic separation of eye-field and telencephalic cells during early neurulation. Copyright © 2012 Elsevier Inc. All rights reserved.

Integrating patterning signals: Wnt/GSK3 regulates the duration of the BMP/Smad1 signal.

PubMed

Fuentealba, Luis C; Eivers, Edward; Ikeda, Atsushi; Hurtado, Cecilia; Kuroda, Hiroki; Pera, Edgar M; De Robertis, Edward M

2007-11-30

BMP receptors determine the intensity of BMP signals via Smad1 C-terminal phosphorylations. Here we show that a finely controlled cell biological pathway terminates this activity. The duration of the activated pSmad1(Cter) signal was regulated by sequential Smad1 linker region phosphorylations at conserved MAPK and GSK3 sites required for its polyubiquitinylation and transport to the centrosome. Proteasomal degradation of activated Smad1 and total polyubiquitinated proteins took place in the centrosome. Inhibitors of the Erk, p38, and JNK MAPKs, as well as GSK3 inhibitors, prolonged the duration of a pulse of BMP7. Wnt signaling decreased pSmad1(GSK3) antigen levels and redistributed it from the centrosome to cytoplasmic LRP6 signalosomes. In Xenopus embryos, it was found that Wnts induce epidermis and that this required an active BMP-Smad pathway. Epistatic experiments suggested that the dorsoventral (BMP) and anteroposterior (Wnt/GSK3) patterning gradients are integrated at the level of Smad1 phosphorylations during embryonic pattern formation.

Sox2 in the dermal papilla niche controls hair growth by fine-tuning Bmp signaling in differentiating hair shaft progenitors

PubMed Central

Clavel, Carlos; Grisanti, Laura; Zemla, Roland; Rezza, Amelie; Barros, Rita; Sennett, Rachel; Mazloom, Amin; Chung, Chi-Yeh; Cai, Xiaoqiang; Cai, Chen-Leng; Pevny, Larysa; Nicolis, Silvia; Ma’ayan, Avi; Rendl, Michael

2012-01-01

SUMMARY How dermal papilla (DP) niche cells regulate hair follicle progenitors to control hair growth remains unclear. Using Tbx18Cre to target embryonic DP precursors, we ablate the transcription factor Sox2 early and efficiently, resulting in diminished hair shaft outgrowth. We find that DP niche expression of Sox2 controls the migration rate of differentiating hair shaft progenitors. Transcriptional profiling of Sox2 null DPs reveals increased Bmp6 and decreased Bmp inhibitor Sostdc1, a direct Sox2 transcriptional target. Subsequently, we identify upregulated Bmp signaling in knockout hair shaft progenitors and demonstrate that Bmps inhibit cell migration, an effect that can be attenuated by Sostdc1. A shorter and Sox2-negative hair type lacks Sostdc1 in the DP and shows reduced migration and increased Bmp activity of hair shaft progenitors. Collectively, our data identify Sox2 as a key regulator of hair growth that controls progenitor migration by fine-tuning Bmp-mediated mesenchymal-epithelial crosstalk. PMID:23153495

ECSIT links TLR and BMP signaling in FOP connective tissue progenitor cells.

PubMed

Wang, Haitao; Behrens, Edward M; Pignolo, Robert J; Kaplan, Frederick S

2018-04-01

Clinical and laboratory observations strongly suggest that the innate immune system induces flare-ups in the setting of dysregulated bone morphogenetic protein (BMP) signaling in fibrodysplasia ossificans progressiva (FOP). In order to investigate the signaling substrates of this hypothesis, we examined toll-like receptor (TLR) activation and bone morphogenetic protein (BMP) signaling in connective tissue progenitor cells (CTPCs) from FOP patients and unaffected individuals. We found that inflammatory stimuli broadly activate TLR expression in FOP CTPCs and that TLR3/TLR4 signaling amplifies BMP pathway signaling through both ligand dependent and independent mechanisms. Importantly, Evolutionarily Conserved Signaling Intermediate in the Toll Pathway (ECSIT) integrates TLR injury signaling with dysregulated BMP pathway signaling in FOP CTPCs. These findings provide novel insight into the cell autonomous integration of injury signals from the innate immune system with dysregulated response signals from the BMP signaling pathway and provide new exploratory targets for therapeutic approaches to blocking the induction and amplification of FOP lesions. Copyright © 2018 Elsevier Inc. All rights reserved.

BMP signaling inhibits intestinal stem cell self-renewal through suppression of Wnt-beta-catenin signaling.

PubMed

He, Xi C; Zhang, Jiwang; Tong, Wei-Gang; Tawfik, Ossama; Ross, Jason; Scoville, David H; Tian, Qiang; Zeng, Xin; He, Xi; Wiedemann, Leanne M; Mishina, Yuji; Li, Linheng

2004-10-01

In humans, mutations in BMPR1A, SMAD4 and PTEN are responsible for juvenile polyposis syndrome, juvenile intestinal polyposis and Cowden disease, respectively. The development of polyposis is a common feature of these diseases, suggesting that there is an association between BMP and PTEN pathways. The mechanistic link between BMP and PTEN pathways and the related etiology of juvenile polyposis is unresolved. Here we show that conditional inactivation of Bmpr1a in mice disturbs homeostasis of intestinal epithelial regeneration with an expansion of the stem and progenitor cell populations, eventually leading to intestinal polyposis resembling human juvenile polyposis syndrome. We show that BMP signaling suppresses Wnt signaling to ensure a balanced control of stem cell self-renewal. Mechanistically, PTEN, through phosphatidylinosital-3 kinase-Akt, mediates the convergence of the BMP and Wnt pathways on control of beta-catenin. Thus, BMP signaling may control the duplication of intestinal stem cells, thereby preventing crypt fission and the subsequent increase in crypt number.

Enhanced BMP signaling results in supernumerary tooth formation in USAG-1 deficient mouse

DOE Office of Scientific and Technical Information (OSTI.GOV)

Murashima-Suginami, Akiko; Takahashi, Katsu; Sakata, Tomoko

2008-05-16

Uterine sensitization associated gene-1 (USAG-1) is a BMP antagonist, and also modulates Wnt signaling. We previously reported that USAG-1 deficient mice have supernumerary teeth. The supernumerary maxillary incisor appears to form as a result of the successive development of the rudimentary upper incisor. USAG-1 abrogation rescued apoptotic elimination of odontogenic mesenchymal cells. We confirmed that BMPs were expressed in both the epithelium and mesenchyme of the rudimentary incisor at E14 and E15. BMP signaling in the rudimentary maxillary incisor, assessed by expressions of Msx1 and Dlx2 and the phosphorylation of Smad protein, was significantly enhanced. Wnt signaling as demonstrated bymore » the nuclear localization of {beta}-catenin was also up-regulated. Inhibition of BMP signaling rescues supernumerary tooth formation in E15 incisor explant culture. Based upon these results, we conclude that enhanced BMP signaling results in supernumerary teeth and BMP signaling was modulated by Wnt signaling in the USAG-1 deficient mouse model.« less

rhBMP-2 (ACS and CRM formulations) overcomes pseudarthrosis in a New Zealand white rabbit posterolateral fusion model.

PubMed

Lawrence, James P; Waked, Walid; Gillon, Thomas J; White, Andrew P; Spock, Christopher R; Biswas, Debdut; Rosenberger, Patricia; Troiano, Nancy; Albert, Todd J; Grauer, Jonathan N

2007-05-15

The study design consisted of a New Zealand white rabbit model of pseudarthrosis repair. Study groups consisting of no graft, autograft, or recombinant human bone morphogenetic protein-2 (rhBMP-2) with absorbable collagen sponge (ACS) or compression resistant matrix (CRM) were evaluated. To evaluate the relative efficacy of bone graft materials (autograft, ACS, and CRM). rhBMP-2 has been shown to have a 100% fusion rate in a primary rabbit fusion model, even in the presence of nicotine, which is known to inhibit fusion. Seventy-two New Zealand white rabbits underwent posterolateral lumbar fusion with iliac crest autograft. To establish pseudarthroses, nicotine was administered to all animals. At 5 weeks, the spines were explored and all pseudarthroses were redecorticated and implanted with no graft, autograft, rhBMP-2/ACS, or rhBMP-2/CRM. At 10 weeks, fusions were assessed by manual palpation and histology. Eight rabbits (11%) were lost to complications. At 5 weeks, 66 (97%) had pseudarthroses. At 10 weeks, attempted pseudarthrosis repairs were fused in 1 of 16 of no graft rabbits (6%), 5 of 17 autograft rabbits (29%), and 31 of 31 rhBMP-2 rabbits (with ACS or CRM) (100%). Histologic analysis demonstrated more mature bone formation in the rhBMP-2 groups. The 2 rhBMP-2 formulations led to significantly higher fusion rates and histologic bone formation than no graft and autograft controls in this pseudarthrosis repair model.

Maxillary sinus floor elevation using BMP-2 and Nell-1 gene-modified bone marrow stromal cells and TCP in rabbits.

PubMed

Xia, Lunguo; Xu, Yuanjin; Chang, Qing; Sun, Xiaojuan; Zeng, Deliang; Zhang, Wenjie; Zhang, Xiuli; Zhang, Zhiyuan; Jiang, Xinquan

2011-07-01

This study evaluated the synergistic osteogenic effect of bone morphogenetic protein-2 (BMP-2) and Nel-like molecule-1 (Nell-1) genes in a rabbit maxillary sinus floor elevation model. Bone marrow stromal cells (bMSCs) were cultured and transduced with AdEGFP, AdNell-1, AdBMP-2, or AdNell-1 + AdBMP-2 overexpression virus. These gene-modified autologous bMSCs were then combined with a β-tricalcium phosphate (β-TCP) granule scaffold and used to elevate the maxillary sinus floor in rabbits. bMSCs cotransduced with AdNell-1 + AdBMP-2 demonstrated a synergistic effect on osteogenic differentiation as detected by real-time PCR analysis on markers of runt-related transcription factor-2, osteocalcin, collagen type 1, alkaline phosphatase activity, and calcium deposits in vitro. As for maxillary sinus floor elevation in a rabbit model in vivo, AdNell-1 + AdBMP-2 gene-transduced autologeous bMSCs/β-TCP complex had the largest bone area and most mature bone structure among the groups, as detected by HE staining and immunohistochemistry at weeks 2 and 8 after implantation. Our data suggested that the BMP-2 and Nell-1 genes possessed a synergistic effect on osteogenic differentiation of bMSCs, while bMSCs modified with the BMP-2 and Nell-1 genes could promote new bone formation and maturation in the rabbit maxillary sinus model.

Downregulated bone morphogenetic protein signaling in nitrofen-induced congenital diaphragmatic hernia.

PubMed

Makanga, Martine; Dewachter, Céline; Maruyama, Hidekazu; Vuckovic, Aline; Rondelet, Benoit; Naeije, Robert; Dewachter, Laurence

2013-08-01

Bone morphogenetic proteins (BMP) have been shown to play crucial roles in not only lung and heart development, but also in the pathogenesis of pulmonary vascular remodeling in pulmonary hypertension (PH). We therefore hypothesized that BMP signaling could be altered in nitrofen-induced congenital diaphragmatic hernia (CDH) and associated PH. Pregnant rats were exposed to either 100 mg nitrofen or vehicle on embryonic day (E) 9.5. On E17 and E21, fetuses were delivered by cesarean section, killed and checked for left-sided CDH. The tissue was then harvested for pathobiological evaluation. In nitrofen-induced CDH, pulmonary expressions of BMP4, BMP receptor (BMPR) type 2 and Id1 decreased on E17 and E21. On E17, pulmonary gremlin-1 expression increased, while BMP7 decreased. In the lungs, Id1 expression was correlated to BMP4 and BMPR2 and inversely correlated to gremlin-1 expression. Myocardial expressions of BMPR2, BMPR1A, BMP7 and SERCA-2A decreased, while gremlin-1 and noggin expressions increased on E17. On E21, myocardial expressions of Id1 and SERCA-2A decreased, while gremlin-1 expression increased. Moreover, BMPR2 and BMPR1A expressions were correlated to SERCA-2A expression and inversely correlated to pro-apoptotic Bax/Bcl2 ratio within the myocardium. Downregulation of BMP signaling seems to contribute to pulmonary and myocardial anomalies observed in nitrofen-induced CDH.

BmP02 Atypically Delays Kv4.2 Inactivation: Implication for a Unique Interaction between Scorpion Toxin and Potassium Channel

PubMed Central

Wu, Bin; Zhu, Yan; Shi, Jian; Tao, Jie; Ji, Yonghua

2016-01-01

BmP02, a short-chain peptide with 28 residues from the venom of Chinese scorpion Buthus martensi Karsch, has been reported to inhibit the transient outward potassium currents (Ito) in rat ventricular muscle cells. However, it remains unclear whether BmP02 modulates the Kv4.2 channel, one of the main contributors to Ito. The present study investigated the effects of BmP02 on Kv4.2 kinetics and its underlying molecular mechanism. The electrophysiological recordings showed that the inactivation of Kv4.2 expressed in HEK293T cells was significantly delayed by BmP02 in a dose-response manner with EC50 of ~850 nM while the peak current, activation and voltage-dependent inactivation of Kv4.2 were not affected. Meanwhile, the recovery from inactivation of Kv4.2 was accelerated and the deactivation was slowed after the application of BmP02. The site-directed mutagenesis combined with computational modelling identified that K347 and K353, located in the turret motif of the Kv4.2, and E4/E5, D20/D21 in BmP02 are key residues to interact with BmP02 through electrostatic force. These findings not only reveal a novel interaction between Kv4.2 channel and its peptidyl modulator, but also provide valuable information for design of highly-selective Kv4.2 modulators. PMID:27690098

BMP2-loaded hollow hydroxyapatite microspheres exhibit enhanced osteoinduction and osteogenicity in large bone defects

PubMed Central

Xiong, Long; Zeng, Jianhua; Yao, Aihua; Tu, Qiquan; Li, Jingtang; Yan, Liang; Tang, Zhiming

2015-01-01

The regeneration of large bone defects is an osteoinductive, osteoconductive, and osteogenic process that often requires a bone graft for support. Limitations associated with naturally autogenic or allogenic bone grafts have demonstrated the need for synthetic substitutes. The present study investigates the feasibility of using novel hollow hydroxyapatite microspheres as an osteoconductive matrix and a carrier for controlled local delivery of bone morphogenetic protein 2 (BMP2), a potent osteogenic inducer of bone regeneration. Hollow hydroxyapatite microspheres (100±25 μm) with a core (60±18 μm) and a mesoporous shell (180±42 m2/g surface area) were prepared by a glass conversion technique and loaded with recombinant human BMP2 (1 μg/mg). There was a gentle burst release of BMP2 from microspheres into the surrounding phosphate-buffered saline in vitro within the initial 48 hours, and continued at a low rate for over 40 days. In comparison with hollow hydroxyapatite microspheres without BMP2 or soluble BMP2 without a carrier, BMP2-loaded hollow hydroxyapatite microspheres had a significantly enhanced capacity to reconstitute radial bone defects in rabbit, as shown by increased serum alkaline phosphatase; quick and complete new bone formation within 12 weeks; and great biomechanical flexural strength. These results indicate that BMP2-loaded hollow hydroxyapatite microspheres could be a potential new option for bone graft substitutes in bone regeneration. PMID:25609957

The type I BMP receptors, Bmpr1a and Acvr1, activate multiple signaling pathways to regulate lens formation

PubMed Central

Rajagopal, Ramya; Huang, Jie; Dattilo, Lisa K.; Kaartinen, Vesa; Mishina, Yuji; Deng, Chu-Xia; Umans, Lieve; Zwijsen, An; Roberts, Anita B.; Beebe, David C.

2009-01-01

BMPs play multiple roles in development and BMP signaling is essential for lens formation. However, the mechanisms by which BMP receptors function in vertebrate development are incompletely understood. To determine the downstream effectors of BMP signaling and their functions in the ectoderm that will form the lens, we deleted the genes encoding the type I BMP receptors, Bmpr1a and Acvr1, and the canonical transducers of BMP signaling, Smad4, Smad1 and Smad5. Bmpr1a and Acvr1 regulated cell survival and proliferation, respectively. Absence of both receptors interfered with the expression of proteins involved in normal lens development and prevented lens formation, demonstrating that BMPs induce lens formation by acting directly on the prospective lens ectoderm. Remarkably, the canonical Smad signaling pathway was not needed for most of these processes. Lens formation, placode cell proliferation, the expression of FoxE3, a lens-specific transcription factor, and the lens protein, αA-crystallin were regulated by BMP receptors in a Smad-independent manner. Placode cell survival was promoted by R-Smad signaling, but in a manner that did not involve Smad4. Of the responses tested, only maintaining a high level of Sox2 protein, a transcription factor expressed early in placode formation, required the canonical Smad pathway. A key function of Smad-independent BMP receptor signaling may be reorganization of actin cytoskeleton to drive lens invagination. PMID:19733164

FAK and BMP-9 synergistically trigger osteogenic differentiation and bone formation of adipose derived stem cells through enhancing Wnt-β-catenin signaling.

PubMed

Yuan, Cheng; Gou, Xiaoli; Deng, Jiang; Dong, Zhijun; Ye, Peng; Hu, Zhenming

2018-06-14

Adipose derived stem cells (ADSCs) could undergo osteogenesis via focal adhesion kinase (FAK) and bone morphogenetic protein (BMP) 9 signals, both of which could affect Wnt-β-catenin signal, a signal pathway closely related to ADSCs osteogenesis. It's still enigma whether FAK and BMP-9 contribute to osteogenesis. Here, we examined the effect of FAK on BMP9-inducedosteogenic differentiation, unveiled the possible molecular mechanism underling this process. In the present study, ADSCs were isolated and purified, and cells of passage 3 underwent virus mediated transfection to prepare ADSCs with stable FAK shRNA expression. Cell viability and migration were detected by MTT and transwell assay, respectively. Expression of osteogenic gene, phosphorylation of FAK and GSK were detected by western blot. Osteogenic potential was evaluated by activity of alkaline phosphatase (ALP) and calcium deposition by ALP staining and Alizarin Red S staining. BMP-9 administration promoted ADSCs osteogenesis. Knocking down FAK attenuated this process, inhibited osteogenic proteins expression through Wnt-β-catenin signal. BMP-9 also triggered ADSCs proliferation and migration, and shFAK antagonized such effects too. Although Wnt signal is affected by FAK shRNA, Smad signal remains intact in ADSCs with shFAK. FAK and BMP-9 could cross talk on Wnt signal pathway and promote ADSCs osteogenesis. FAK could participate in BMP-9 induced ADSCs osteogenesis via Wnt signal pathway other than Smads signals (see in graph). Copyright © 2018. Published by Elsevier Masson SAS.

Assessment of a polyelectrolyte multilayer film coating loaded with BMP-2 on titanium and PEEK implants in the rabbit femoral condyle

PubMed Central

Guillot, R.; Pignot-Paintrand, I.; Lavaud, J.; Decambron, A.; Bourgeois, E.; Josserand, V.; Logeart-Avramoglou, D.; Viguier, E.; Picart, C.

2016-01-01

The aim of this study was to evaluate the osseointegration of titanium implants (Ti-6Al-4V, noted here TA6V) and poly(etheretherketone) PEEK implants induced by a BMP-2-delivering surface coating made of polyelectrolyte multilayer films. The in vitro bioactivity of the polyelectrolyte film-coated implants was assessed using the alkaline phosphatase assay. BMP-2-coated TA6V and PEEK implants with a total dose of 9.3 µg of BMP-2 were inserted into the femoral condyles of New Zealand white rabbits and compared to uncoated implants. Rabbits were sacrificed 4 and 8 weeks after implantation. Histomorphometric analyses on TA6V and PEEK implants and microcomputed tomography on PEEK implants revealed that the bone-to-implant contact and bone area around the implants were significantly lower for the BMP-2-coated implants than for the bare implants. This was confirmed by scanning electron microscopy imaging. This difference was more pronounced at 4 weeks in comparison to the 8-week time point. However, bone growth inside the hexagonal upper hollow cavity of the screws was higher in the case of the BMP-2 coated implants. Overall, this study shows that a high dose of BMP-2 leads to localized and temporary bone impairment, and that the dose of BMP-2 delivered at the surface of an implant needs to be carefully optimized. PMID:26965394

Bone generation in the reconstruction of a critical size calvarial defect in an experimental model.

PubMed

Por, Yong-Chen; Barceló, C Raul; Salyer, Kenneth E; Genecov, David G; Troxel, Karen; Gendler, El; Elsalanty, Mohammed E; Opperman, Lynne A

2008-03-01

This study was designed to investigate the optimal combination of known osteogenic biomaterials with shape conforming struts to achieve calvarial vault reconstruction, using a canine model. Eighteen adolescent beagles were divided equally into 6 groups. A critical-size defect of 6 x 2 cm traversed the sagittal suture. The biomaterials used for calvarial reconstruction were demineralized perforated bone matrix (DBM), recombinant human bone morphogenetic protein 2 (rhBMP2), and autogenous platelet-rich plasma (PRP). The struts used were cobalt chrome (metal) or resorbable plate. The groupings were as follows: 1) DBM + metal, 2) DBM + PRP + metal, 3) DBM + PRP + resorbable plate, 4) DBM + rhBMP2 + metal, 5) DBM + rhBMP2 + PRP + metal, and 6) DBM + rhBMP2 + resorbable plate. Animals were killed at 3 months after surgery. There was no mortality or major complications. Analysis was performed macroscopically and histologically and with computed tomography. There was complete bony regeneration in the rhBMP2 groups only. Non-rhBMP2 groups had minimal bony ingrowth from the defect edges and on the dural surface, a finding confirmed by computed tomographic scan and histology. Platelet-rich plasma did not enhance bone regeneration. Shape conformation was good with both metal and resorbable plate. rhBMP2, but not PRP, accelerated calvarial regeneration in 3 months. The DBMs in the rhBMP2 groups were substituted by new trabecular bone. Shape molding was good with both metal and resorbable plate.

BMP15 regulates AMH expression via the p38 MAPK pathway in granulosa cells from goat.

PubMed

Zhao, Zhongquan; Guo, Fangyue; Sun, Xiaowei; He, Qijie; Dai, Zinuo; Chen, Xiaochuan; Zhao, Yongju; Wang, Jian

2018-05-31

Anti-Mullerian hormone (AMH), a member of the TGF-β superfamily, is produced by granulosa cells (GCs) of preantral and small antral follicles and plays a role in regulating the recruitment of primordial follicles and the FSH-dependent development of follicles. However, the regulation of AMH expression in follicles remains poorly understood. The objectives of this study were to determine the following: 1. the association between bone morphogenetic protein 15 (BMP15) and AMH; 2. whether BMP15 can regulate the expression of AMH by inhibiting the p38 MAPK pathway; and 3. whether SRY-related HMG box 9 (SOX9), a transcription factor for AMH, is involved in the regulation of AMH expression by BMP15. In this study, an inhibitor of p38 MAPK and an siRNA specific for p38 MAPK were used to prevent the function of the p38 MAPK signaling pathway. Then, AMH mRNA expression and AMH secretion were detected in goat GCs using an RT-PCR assay and ELISA, respectively, after treatment with BMP15. The results indicated that BMP15 up-regulates the transcription of AMH and that the inhibition of p38 MAPK decreases the BMP15-induced expression of AMH and SOX9, suggesting that BMP15 up-regulates the expression of AMH via the p38 MAPK signaling pathway, and this process involves the SOX9 transcription factor. Copyright © 2018 Elsevier Inc. All rights reserved.

Continuity resection of the mandible after ameloblastoma - feasibility of oral rehabilitation with rhBMP-2 associated to bovine xenograft followed by implant installation.

PubMed

Lustosa, Romulo Maciel; Macedo, Diogo de Vasconcelos; Iwaki, Lilian Cristina Vessoni; Tolentino, Elen de Souza; Hasse, Paulo Norberto; Marson, Giordano Bruno de Oliveira; Iwaki Filho, Liogi

2015-10-01

Recombinant human morphogenetic protein (rhBMP) is a graft alternative for extensive mandibular reconstruction after tumor resections. However, the feasibility of rhBMP-2 to receive osseointegrated implants and prosthetic rehabilitation has been rarely reported. This study reports on a case of an extensive solid ameloblastoma along the mandibular body. The treatment consisted of resection followed by off-label use of rhBMP type 2 associated with bovine bone xenograft. Eleven months postoperatively, the patient was prosthetically rehabilitated with dental implants, without evidence of resorption or complications. The literature on mandibular reconstructions using rhBMP and their feasibility for future osseointegrated implant placement was also reviewed. Based on the presented case, the association between rhBMP-2 and a bovine bone xenograft could be considered a feasible option for the reconstruction and rehabilitation of large mandibular defects after tumor resection. According to the literature, the use of rhBMP as a graft material is encouraging, with good clinical outcome. However, there are no long-term studies demonstrating success and survival rates of implants placed in these grafts. Future investigations will be required to ascertain the long-term survival of implants in areas grafted with rhBMP. Also, there is a lack of information regarding the prosthetic rehabilitation of these patients. Copyright © 2015 European Association for Cranio-Maxillo-Facial Surgery. Published by Elsevier Ltd. All rights reserved.

Development of Regional Excel-Based Stormwater/Nutrient BMP Optimization Tool (Opti-Tool)

EPA Science Inventory

During 2014, EPA Region 1 contracted with Tetra Tech, Inc. to work with a regional technical Advisory Committee to develop an Excel-based stormwater/nutrient BMP optimization tool (Opti-Tool) using regional precipitation data and regionally calibrated BMP performance data from UN...

Bone marrow-derived mesenchymal stem cells assembled with low-dose BMP-2 in a three-dimensional hybrid construct enhances posterolateral spinal fusion in syngeneic rats.

PubMed

Hu, Tao; Abbah, Sunny Akogwu; Toh, Soo Yein; Wang, Ming; Lam, Raymond Wing Moon; Naidu, Mathanapriya; Bhakta, Gajadhar; Cool, Simon M; Bhakoo, Kishore; Li, Jun; Goh, James Cho-Hong; Wong, Hee-Kit

2015-12-01

The combination of potent osteoinductive growth factor, functional osteoblastic cells, and osteoconductive materials to induce bone formation is a well-established concept in bone tissue engineering. However, supraphysiological dose of growth factor, such as recombinant human bone morphogenetic protein 2 (rhBMP-2), which is necessary in contemporary clinical application, have been reported to result in severe side effects. We hypothesize that the synergistic osteoinductive capacity of low-dose bone morphogenetic protein 2 (BMP-2) combined with undifferentiated bone marrow-derived stromal cells (BMSCs) is comparable to that of osteogenically differentiated BMSCs when used in a rodent model of posterolateral spinal fusion. A prospective study using a rodent model of posterolateral spinal fusion was carried out. Thirty-six syngeneic Fischer rats comprised the patient sample. Six groups of implants were evaluated as follows (n=6): (1) 10 µg BMP-2 with undifferentiated BMSCs; (2) 10 µg BMP-2 with osteogenic-differentiated BMSCs; (3) 2.5 µg BMP-2 with undifferentiated BMSCs; (4) 2.5 µg BMP-2 with osteogenic-differentiated BMSCs; (5) 0.5 µg BMP-2 with undifferentiated BMSCs; and (6) 0.5 µg BMP-2 with osteogenic-differentiated BMSCs. Optimal in vitro osteogenic differentiation of BMSCs was determined by quantitative real-time polymerase chain reaction (qRT-PCR) gene analysis whereas in vivo bone formation capacity was evaluated by manual palpation, micro-computed tomography, and histology. Rat BMSCs cultured in fibrin matrix that was loaded into the pores of medical-grade poly epsilon caprolactone tricalcium phosphate scaffolds differentiated toward osteogenic lineage by expressing osterix, runt-related transcription factor 2, and osteocalcium mRNA when supplemented with dexamethasone, ascorbic acid, and β-glycerophosphate. Whereas qRT-PCR revealed optimal increase in osteogenic genes expression after 7 days of in vitro culture, in vivo transplantation study showed that pre-differentiation of BMSCs before transplantation failed to promote posterolateral spinal fusion when co-delivered with low-dose BMP-2 (1/6 or 17% fusion rate). In contrast, combined delivery of undifferentiated BMSCs with low-dose BMP-2 (2.5 µg) demonstrated significantly higher fusion rate (4/6 or 67%) as well as significantly increased volume of new bone formation (p<.05). In summary, this study supports the combination of undifferentiated BMSCs and low-dose rhBMP-2 for bone tissue engineering construct. Copyright © 2015 Elsevier Inc. All rights reserved.

Local rhBMP-12 on an Absorbable Collagen Sponge as an Adjuvant Therapy for Rotator Cuff Repair - A Phase 1, Randomized, Standard of Care Control, Multicenter Study: Safety and Feasibility.

PubMed

Greiner, Stefan; Ide, Junji; Van Noort, Arthur; Mochizuki, Yu; Ochi, Hiroshi; Marraffino, Shannon; Sridharan, Sudhakar; Rudicel, Sally; Itoi, Eiji

2015-08-01

Recombinant human bone morphogenetic protein-12 (rhBMP-12) has been shown to induce tendon and ligament formation in rats and to improve tendon healing; however, the safety and feasibility of implanting rhBMP-12/absorbable collagen sponge (ACS) in humans are not known. To investigate the safety and feasibility of rhBMP-12 on an ACS as an adjuvant therapy in open rotator cuff repair. Randomized controlled trial; Level of evidence, 2. This study consisted of 20 patients with full-thickness rotator cuff tears. Patients were randomized either to standard of care (SOC) treatment (open rotator cuff repair) or to receive 0.015 mg/mL rhBMP-12/ACS and SOC treatment during their open rotator cuff repair (rhBMP-12/ACS group) at a rate of 1/4 SOC/rhBMP-12/ACS. The feasibility of implanting the product and the safety of the product were evaluated during the 1-year follow-up period. The evaluation involved up to 10 postoperative visits, which included physical examinations, radiographs, computed tomography (CT) scans, magnetic resonance imaging (MRI) scans with an emphasis on heterotopic ossification (HO), pharmacokinetics, immunogenicity, laboratory evaluations, and local and systemic adverse events at specified time points. Small amounts of HO were seen on follow-up CT scans in 10 of 16 patients in the rhBMP-12/ACS group and in 2 of 3 patients in the SOC group. HO did not increase at 26 weeks and was not associated with any adverse events or unsatisfactory clinical outcomes. Pharmacokinetics demonstrated that circulating levels of rhBMP-12 were not detectable after administration. Five of 16 patients showed a postoperative immunogenic response but did not show any correlating adverse events. Complete healing of the rotator cuff was observed in 14 of 16 patients; 2 of 16 imaging results could not be analyzed because of artifacts in the rhBMP-12 group on MRI scans. In the SOC group, 1 of 4 patients showed a retear at 12 weeks after surgery. The use of rhBMP-12/ACS has been shown to be feasible and safe in a concentration of 0.015 mg/mL when used in open rotator cuff repair. Higher dose concentrations of rhBMP-12 should be evaluated in the future to evaluate their safety and potential to increase rotator cuff healing after open surgical repair. © 2015 The Author(s).

The BMP pathway is essential for re-specification and maintenance of the dorsoventral axis in regenerating and intact planarians.

PubMed

Molina, M Dolores; Saló, Emili; Cebrià, Francesc

2007-11-01

The bone morphogenetic protein (BMP) pathway has been shown to play an important role in the establishment of the dorsoventral axis during development in both vertebrate and invertebrate species. In an attempt to unravel the role of BMPs in pattern formation during planarian regeneration, we studied this signaling pathway in Schmidtea mediterranea. Here, we functionally characterize planarian homologues of two key elements of the pathway: Smed-BMP and Smed-Smad1. Whole-mount in situ hybridization showed that Smed-BMP is expressed at the planarian dorsal midline, suggesting a role in dorsoventral patterning, while Smed-Smad1 is widely expressed throughout the mesenchyme and in the central nervous system. RNA interference (RNAi) knockdowns of Smed-BMP or Smed-Smad1 led to the disappearance of dorsal markers along with the ectopic expression of ventral markers on the dorsal side of the treated animals. In almost all cases, a duplicated central nervous system differentiated dorsally after Smed-BMP or Smed-Smad1 RNAi. These defects were observed not only during regeneration but also in intact non-regenerating animals. Our results suggest that the BMP signaling pathway is conserved in planarians and that it plays a key role in the regeneration and maintenance of the dorsoventral axis.

Combined sustained release of BMP2 and MMP10 accelerates bone formation and mineralization of calvaria critical size defect in mice.

PubMed

Reyes, Ricardo; Rodríguez, Jose Antonio; Orbe, Josune; Arnau, María Rosa; Évora, Carmen; Delgado, Araceli

2018-11-01

The effect of dual delivery of bone morphogenetic protein-2 (BMP-2) and matrix metalloproteinase 10 (MMP10) on bone regeneration was investigated in a murine model of calvarial critical-size defect, hypothesizing that it would result in an enhanced bone formation. Critical-size calvarial defects (4 mm diameter) were created in mice and PLGA microspheres preloaded with either BMP-2, MMP10 or a microsphere combination of both were transplanted into defect sites at different doses. Empty microspheres were used as the negative control. Encapsulation efficiency was assessed and in vivo release kinetics of BMP-2 and MMP10 were examined over 14 days. Histological analyses were used to analyze bone formation after four and eight weeks. Combination with MMP10 (30 ng) significantly enhanced BMP-2 (600 ng)-mediated osteogenesis, as confirmed by the increase in percentage of bone fill (p < .05) at four weeks. Moreover, it also increased mineral apposition rate (p < .05), measured by double labeling with tetracycline and calceine. MMP10 accelerates bone repair by enhancing BMP-2-promoted bone healing and improving the mineralization rate. In conclusion combination of MMP10 and BMP-2 may become a promising strategy for repair and regeneration of bone defects.

Enhanced Healing of Rat Calvarial Defects with MSCs Loaded on BMP-2 Releasing Chitosan/Alginate/Hydroxyapatite Scaffolds

PubMed Central

He, Xiaoning; Liu, Yang; Yuan, Xue; Lu, Li

2014-01-01

In this study, we designed a chitosan/alginate/hydroxyapatite scaffold as a carrier for recombinant BMP-2 (CAH/B2), and evaluated the release kinetics of BMP-2. We evaluated the effect of the CAH/B2 scaffold on the viability and differentiation of bone marrow mesenchymal stem cells (MSCs) by scanning electron microscopy, MTS, ALP assay, alizarin-red staining and qRT-PCR. Moreover, MSCs were seeded on scaffolds and used in a 8 mm rat calvarial defect model. New bone formation was assessed by radiology, hematoxylin and eosin staining 12 weeks postoperatively. We found the release kinetics of BMP-2 from the CAH/B2 scaffold were delayed compared with those from collagen gel, which is widely used for BMP-2 delivery. The BMP-2 released from the scaffold increased MSC differentiation and did not show any cytotoxicity. MSCs exhibited greater ALP activity as well as stronger calcium mineral deposition, and the bone-related markers Col1α, osteopontin, and osteocalcin were upregulated. Analysis of in vivo bone formation showed that the CAH/B2 scaffold induced more bone formation than other groups. This study demonstrates that CAH/B2 scaffolds might be useful for delivering osteogenic BMP-2 protein and present a promising bone regeneration strategy. PMID:25084008

Lack of effect of bone morphogenetic protein 2 and 4 gene polymorphisms on bone density in postmenopausal Turkish women.

PubMed

Ozkan, Z S; Deveci, D; Onalan Etem, E; Yüce, H

2010-11-30

We investigated the effect of bone morphogenetic protein 2 and 4 (BMP-2 and -4) gene polymorphisms on bone density in postmenopausal Turkish women with osteoporosis. The frequency of single-nucleotide polymorphisms (SNPs) of BMP-2 and -4 genes was analyzed in 101 osteoporotic-postmenopausal women and 52 postmenopausal women with positive bone mineral density scores. We evaluated the frequency of the thymine→cytosine nucleotide variation at position 538 for BMP-4 and the transposition of adenine→thymine at codon 190 for BMP-2, with PCR. The proportions of genotypes observed for the BMP-2 SNP in the osteoporotic group were AA (47.5%), AT (39.6%), TT (12.9%), and in the non-osteoporotic group they were AA (48.1%), AT (40.4%), TT (11.5%). The corresponding frequencies for the BMP-4 SNP in the osteoporotic group were TT (30.7%), TC (45.5%), CC (23.8%), and in the non-osteoporotic group they were TT (26.9%), TC (40.4%), CC (32.7%). There were no significant differences in the frequencies of these genotypes between the patient and control groups. We conclude that genetic variations in BMP-2 and -4 do not substantially contribute to lumbar spine bone mineral density in postmenopausal Turkish women.

Follistatin-like 1 (Fstl1) is a bone morphogenetic protein (BMP) 4 signaling antagonist in controlling mouse lung development

PubMed Central

Geng, Yan; Dong, Yingying; Yu, Mingyan; Zhang, Long; Yan, Xiaohua; Sun, Jingxia; Qiao, Long; Geng, Huixia; Nakajima, Masahiro; Furuichi, Tatsuya; Ikegawa, Shiro; Gao, Xiang; Chen, Ye-Guang; Jiang, Dianhua; Ning, Wen

2011-01-01

Lung morphogenesis is a well orchestrated, tightly regulated process through several molecular pathways, including TGF-β/bone morphogenetic protein (BMP) signaling. Alteration of these signaling pathways leads to lung malformation. We investigated the role of Follistatin-like 1 (Fstl1), a secreted follistatin-module–containing glycoprotein, in lung development. Deletion of Fstl1 in mice led to postnatal lethality as a result of respiratory failure. Analysis of the mutant phenotype showed that Fstl1 is essential for tracheal cartilage formation and alveolar maturation. Deletion of the Fstl1 gene resulted in malformed tracheal rings manifested as discontinued rings and reduced ring number. Fstl1-deficient mice displayed septal hypercellularity and end-expiratory atelectasis, which were associated with impaired differentiation of distal alveolar epithelial cells and insufficient production of mature surfactant proteins. Mechanistically, Fstl1 interacted directly with BMP4, negatively regulated BMP4/Smad1/5/8 signaling, and inhibited BMP4-induced surfactant gene expression. Reducing BMP signaling activity by Noggin rescued pulmonary atelectasis of Fstl1-deficient mice. Therefore, we provide in vivo and in vitro evidence to demonstrate that Fstl1 modulates lung development and alveolar maturation, in part, through BMP4 signaling. PMID:21482757

A tissue engineering solution for segmental defect regeneration in load-bearing long bones.

PubMed

Reichert, Johannes C; Cipitria, Amaia; Epari, Devakara R; Saifzadeh, Siamak; Krishnakanth, Pushpanjali; Berner, Arne; Woodruff, Maria A; Schell, Hanna; Mehta, Manav; Schuetz, Michael A; Duda, Georg N; Hutmacher, Dietmar W

2012-07-04

The reconstruction of large defects (>10 mm) in humans usually relies on bone graft transplantation. Limiting factors include availability of graft material, comorbidity, and insufficient integration into the damaged bone. We compare the gold standard autograft with biodegradable composite scaffolds consisting of medical-grade polycaprolactone and tricalcium phosphate combined with autologous bone marrow-derived mesenchymal stem cells (MSCs) or recombinant human bone morphogenetic protein 7 (rhBMP-7). Critical-sized defects in sheep--a model closely resembling human bone formation and structure--were treated with autograft, rhBMP-7, or MSCs. Bridging was observed within 3 months for both the autograft and the rhBMP-7 treatment. After 12 months, biomechanical analysis and microcomputed tomography imaging showed significantly greater bone formation and superior strength for the biomaterial scaffolds loaded with rhBMP-7 compared to the autograft. Axial bone distribution was greater at the interfaces. With rhBMP-7, at 3 months, the radial bone distribution within the scaffolds was homogeneous. At 12 months, however, significantly more bone was found in the scaffold architecture, indicating bone remodeling. Scaffolds alone or with MSC inclusion did not induce levels of bone formation comparable to those of the autograft and rhBMP-7 groups. Applied clinically, this approach using rhBMP-7 could overcome autograft-associated limitations.

A balance of FGF and BMP signals regulates cell cycle exit and Equarin expression in lens cells

PubMed Central

Jarrin, Miguel; Pandit, Tanushree; Gunhaga, Lena

2012-01-01

In embryonic and adult lenses, a balance of cell proliferation, cell cycle exit, and differentiation is necessary to maintain physical function. The molecular mechanisms regulating the transition of proliferating lens epithelial cells to differentiated primary lens fiber cells are poorly characterized. To investigate this question, we used gain- and loss-of-function analyses to modulate fibroblast growth factor (FGF) and/or bone morphogenetic protein (BMP) signals in chick lens/retina explants. Here we show that FGF activity plays a key role for proliferation independent of BMP signals. Moreover, a balance of FGF and BMP signals regulates cell cycle exit and the expression of Ccdc80 (also called Equarin), which is expressed at sites where differentiation of lens fiber cells occurs. BMP activity promotes cell cycle exit and induces Equarin expression in an FGF-dependent manner. In contrast, FGF activity is required but not sufficient to induce cell cycle exit or Equarin expression. Furthermore, our results show that in the absence of BMP activity, lens cells have increased cell cycle length or are arrested in the cell cycle, which leads to decreased cell cycle exit. Taken together, these findings suggest that proliferation, cell cycle exit, and early differentiation of primary lens fiber cells are regulated by counterbalancing BMP and FGF signals. PMID:22718906

Regulation of Dense-Core Granule Replenishment by Autocrine BMP Signalling in Drosophila Secondary Cells.

PubMed

Redhai, Siamak; Hellberg, Josephine E E U; Wainwright, Mark; Perera, Sumeth W; Castellanos, Felix; Kroeger, Benjamin; Gandy, Carina; Leiblich, Aaron; Corrigan, Laura; Hilton, Thomas; Patel, Benjamin; Fan, Shih-Jung; Hamdy, Freddie; Goberdhan, Deborah C I; Wilson, Clive

2016-10-01

Regulated secretion by glands and neurons involves release of signalling molecules and enzymes selectively concentrated in dense-core granules (DCGs). Although we understand how many secretagogues stimulate DCG release, how DCG biogenesis is then accelerated to replenish the DCG pool remains poorly characterised. Here we demonstrate that each prostate-like secondary cell (SC) in the paired adult Drosophila melanogaster male accessory glands contains approximately ten large DCGs, which are loaded with the Bone Morphogenetic Protein (BMP) ligand Decapentaplegic (Dpp). These DCGs can be marked in living tissue by a glycophosphatidylinositol (GPI) lipid-anchored form of GFP. In virgin males, BMP signalling is sporadically activated by constitutive DCG secretion. Upon mating, approximately four DCGs are typically released immediately, increasing BMP signalling, primarily via an autocrine mechanism. Using inducible knockdown specifically in adult SCs, we show that secretion requires the Soluble NSF Attachment Protein, SNAP24. Furthermore, mating-dependent BMP signalling not only promotes cell growth, but is also necessary to accelerate biogenesis of new DCGs, restoring DCG number within 24 h. Our analysis therefore reveals an autocrine BMP-mediated feedback mechanism for matching DCG release to replenishment as secretion rates fluctuate, and might explain why in other disease-relevant systems, like pancreatic β-cells, BMP signalling is also implicated in the control of secretion.

Regulation of Dense-Core Granule Replenishment by Autocrine BMP Signalling in Drosophila Secondary Cells

PubMed Central

Redhai, Siamak; Hellberg, Josephine E. E. U.; Wainwright, Mark; Perera, Sumeth W.; Castellanos, Felix; Kroeger, Benjamin; Gandy, Carina; Leiblich, Aaron; Corrigan, Laura; Hilton, Thomas; Patel, Benjamin; Fan, Shih-Jung; Hamdy, Freddie; Goberdhan, Deborah C. I.

2016-01-01

Regulated secretion by glands and neurons involves release of signalling molecules and enzymes selectively concentrated in dense-core granules (DCGs). Although we understand how many secretagogues stimulate DCG release, how DCG biogenesis is then accelerated to replenish the DCG pool remains poorly characterised. Here we demonstrate that each prostate-like secondary cell (SC) in the paired adult Drosophila melanogaster male accessory glands contains approximately ten large DCGs, which are loaded with the Bone Morphogenetic Protein (BMP) ligand Decapentaplegic (Dpp). These DCGs can be marked in living tissue by a glycophosphatidylinositol (GPI) lipid-anchored form of GFP. In virgin males, BMP signalling is sporadically activated by constitutive DCG secretion. Upon mating, approximately four DCGs are typically released immediately, increasing BMP signalling, primarily via an autocrine mechanism. Using inducible knockdown specifically in adult SCs, we show that secretion requires the Soluble NSF Attachment Protein, SNAP24. Furthermore, mating-dependent BMP signalling not only promotes cell growth, but is also necessary to accelerate biogenesis of new DCGs, restoring DCG number within 24 h. Our analysis therefore reveals an autocrine BMP-mediated feedback mechanism for matching DCG release to replenishment as secretion rates fluctuate, and might explain why in other disease-relevant systems, like pancreatic β-cells, BMP signalling is also implicated in the control of secretion. PMID:27727275

Meat and Livestock Association Plenary Lecture 2005. Oocyte signalling molecules and their effects on reproduction in ruminants.

PubMed

McNatty, Kenneth P; Lawrence, Stephen; Groome, Nigel P; Meerasahib, Mohammed F; Hudson, Norma L; Whiting, Lynda; Heath, Derek A; Juengel, Jennifer L

2006-01-01

Sheep (Ovis aries) are a highly diverse species, with more than 900 different breeds that vary significantly in their physiological characteristics, including ovulation rate and fecundity. From examination of inherited patterns of ovulation rate, several breeds have been identified with point mutations in two growth factor genes that are expressed in oocytes. Currently, five different point mutations have been identified in the BMP15 (GDF9b) gene and one in GDF9. Animals heterozygous for the GDF9 and/or the BMP15 mutations have higher ovulation rates than their wild-type counterparts. In contrast, those homozygous for any of the aforementioned BMP15 or GDF9 mutations are sterile owing to arrested follicular development. In bovine and ovine ovaries, GDF9 was expressed exclusively in oocytes throughout follicular growth from the primordial stage of development, whereas in sheep BMP15 was expressed exclusively in oocytes from the primary stage: no data for the ontogeny of BMP15 expression are currently available for cattle. In vitro, ovine growth differentiation factor 9 (oGDF9) has no effect on (3)H-thymidine incorporation by either bovine or ovine granulosa cells, whereas ovine bone morphogenetic protein 15 (oBMP15) has modest (1.2- to 1.6-fold; P < 0.05) stimulatory effects. Ovine GDF9 or oBMP15 alone inhibited progesterone production by bovine granulosa cells, whereas in ovine cells only oGDF9 was inhibitory. The effects of oGDF9 and oBMP15 together were often cooperative and not always the same as those observed for each factor alone. Active immunisation of ewes with BMP15 and/or GDF9 peptides affected ovarian follicular development and ovulation rate. Depending on the GDF9 and/or BMP15 vaccine formulation, ovulation rate was either increased or suppressed. A primary and single booster immunisation of ewes with a BMP15 peptide in a water-based adjuvant has led to 19-40% increases in lambs born per ewe lambing. Collectively, the evidence suggests that oocyte signalling molecules have profound effects on reproduction in mammals, including rodents, humans and ruminants. Moreover, in vivo manipulation of these oocyte signalling molecules provides new opportunities for the management of the fertility of ruminants.

Vancomycin-bearing synthetic bone graft delivers rhBMP-2 and promotes healing of critical rat femoral segmental defects.

PubMed

Skelly, Jordan D; Lange, Jeffrey; Filion, Tera M; Li, Xinning; Ayers, David C; Song, Jie

2014-12-01

Bone grafts simultaneously delivering therapeutic proteins and antibiotics may be valuable in orthopaedic trauma care. Previously, we developed a poly(2-hydroxyethyl methacrylate)-nanocrystalline hydroxyapatite (pHEMA-nHA) synthetic bone graft that, when preabsorbed with 400-ng rhBMP-2/7, facilitated the functional repair of critical-size rat femoral defects. Recently, we showed that pHEMA-nHA effectively retains/releases vancomycin and rhBMP-2 in vitro. The success of such a strategy requires that the incorporation of vancomycin does not compromise the structural integrity of the graft nor its ability to promote bone healing. (1) To evaluate the ability of pHEMA-nHA-vancomycin composites in combination with 3-µg rhBMP-2 to repair 5 mm rat femoral segmental defects, and (2) To determine if the encapsulated vancomycin impairs the graft/rhBMP-2-assisted bone repair. pHEMA-nHA-vancomycin, pHEMA-nHA, or collagen sponge control with/without 3-µg rhBMP-2 were press-fit in 5 mm femoral defects in SASCO-SD male rats (289-300 g). Histology, microcomputed tomography, and torsion testing were performed on 8- and 12-week explants to evaluate the extent and quality of repair. The effect of vancomycin on the temporal absorption of endogenous BMP-2 and stromal cell-derived factor-1 was evaluated by immunohistochemistry. These factors are important for bone healing initiation and stem cell recruitment, respectively. Partial bridging of the defect with bony callus by 12 weeks was observed with pHEMA-nHA-vancomycin without rhBMP-2 while full bridging with substantially mineralized callus and partial restoration of torsional strength was achieved with 3-µg rhBMP-2. The presence of vancomycin changed the absorption patterns of endogenous proteins on the grafts, but did not appear to substantially compromise graft healing. The composite pHEMA-nHA-vancomycin preabsorbed with 3-µg rhBMP-2 promoted repair of 5 mm rat femoral segmental defects. With the sample sizes applied, vancomycin encapsulation did not appear to have a negative effect on bone healing. pHEMA-nHA-vancomycin preabsorbed with rhBMP-2 may be useful in the repair of critical-size long bone defects prone to infections.

Comparison of the osteogenesis and fusion rates between activin A/BMP-2 chimera (AB204) and rhBMP-2 in a beagle's posterolateral lumbar spine model.

PubMed

Zheng, Guang Bin; Yoon, Byung-Hak; Lee, Jae Hyup

2017-10-01

Activin A/BMP-2 chimera (AB204) could promote bone healing more effectively than recombinant bone morphogenetic protein 2 (rhBMP-2) with much lower dose in a rodent model, but there is no report about the effectiveness of AB204 in a large animal model. The purpose of this study was to compare the osteogenesis and fusion rate between AB204 and rhBMP-2 using biphasic calcium phosphate (BCP) as a carrier in a beagle's posterolateral lumbar fusion model. This is a randomized control animal study. Seventeen male beagle dogs were included. Bilateral posterolateral fusion was performed at the L1-L2 and L4-L5 levels. Biphasic calcium phosphate (2 cc), rhBMP-2 (50 µg)+BCP (2 cc), or AB204 (50 µg)+BCP (2 cc) were implanted into the intertransverse space randomly. X-ray was performed at 4 and 8 weeks. After 8 weeks, the animals were sacrificed, and new bone formation and fusion rate were evaluated by manual palpation, computed tomography (CT), and undecalcified histology. The AB204 group showed significantly higher fusion rate (90%) than the rhBMP-2 group (15%) or the Osteon group (6.3%) by manual palpation. On x-ray and CT assessment, fusion rate and the volume of newly formed bone were also significantly higher in AB204 group than other groups. In contrast, more osteolysis was found in rhBMP-2 group (40%) than in AB204 group (10%) on CT study. In histologic results, new bone formation was sufficient between transverse processes in AB204 group, and obvious trabeculation and bone remodeling were observed. But in rhBMP-2 group, new bone formation was less than AB204 group and osteolysis was observed between the intertransverse spaces. A low dose of AB204 with BCP as a carrier significantly promotes the fusion rate in a large animal model when compared with the rhBMP-2. These findings demonstrate that AB204 could be an alternative to rhBMP-2 to improve fusion rate. Copyright © 2017 Elsevier Inc. All rights reserved.

An Assessment of Mississippi's Nonindustrial Private Forest Landowners' Knowledge of Forest Best Management Practices

Treesearch

Andrew James Londo; John Benkert Auel

2004-01-01

This study examined the knowledge levels of Mississippi nonindustrial private forest (NIPF) landowners relative to best management practices (BMPs) for water quality. Data were collected through surveys of participants in BMP programs held in conjunction with County Forestry Association (CFA) meetings throughout Mississippi during 2001-02. Ten CFAs participated in this...

Binding Interactions of Keratin-Based Hair Fiber Extract to Gold, Keratin, and BMP-2

PubMed Central

de Guzman, Roche C.; Tsuda, Shanel M.; Ton, Minh-Thi N.; Zhang, Xiao; Esker, Alan R.; Van Dyke, Mark E.

2015-01-01

Hair-derived keratin biomaterials composed mostly of reduced keratin proteins (kerateines) have demonstrated their utility as carriers of biologics and drugs for tissue engineering. Electrostatic forces between negatively-charged keratins and biologic macromolecules allow for effective drug retention; attraction to positively-charged growth factors like bone morphogenetic protein 2 (BMP-2) has been used as a strategy for osteoinduction. In this study, the intermolecular surface and bulk interaction properties of kerateines were investigated. Thiol-rich kerateines were chemisorbed onto gold substrates to form an irreversible 2-nm rigid layer for surface plasmon resonance analysis. Kerateine-to-kerateine cohesion was observed in pH-neutral water with an equilibrium dissociation constant (KD) of 1.8 × 10−4 M, indicating that non-coulombic attractive forces (i.e. hydrophobic and van der Waals) were at work. The association of BMP-2 to kerateine was found to be greater (KD = 1.1 × 10−7 M), within the range of specific binding. Addition of salts (phosphate-buffered saline; PBS) shortened the Debye length or the electrostatic field influence which weakened the kerateine-BMP-2 binding (KD = 3.2 × 10−5 M). BMP-2 in bulk kerateine gels provided a limited release in PBS (~ 10% dissociation in 4 weeks), suggesting that electrostatic intermolecular attraction was significant to retain BMP-2 within the keratin matrix. Complete dissociation between kerateine and BMP-2 occurred when the PBS pH was lowered (to 4.5), below the keratin isoelectric point of 5.3. This phenomenon can be attributed to the protonation of keratin at a lower pH, leading to positive-positive repulsion. Therefore, the dynamics of kerateine-BMP-2 binding is highly dependent on pH and salt concentration, as well as on BMP-2 solubility at different pH and molarity. The study findings may contribute to our understanding of the release kinetics of drugs from keratin biomaterials and allow for the development of better, more clinically relevant BMP-2-conjugated systems for bone repair and regeneration. PMID:26317522

Topical bisphosphonate augments fixation of bone-grafted hydroxyapatite coated implants, BMP-2 causes resorption-based decrease in bone.

PubMed

Baas, Jorgen; Vestermark, Marianne; Jensen, Thomas; Bechtold, Joan; Soballe, Kjeld; Jakobsen, Thomas

2017-04-01

Bone allograft is used in total joint arthroplasties in order to enhance implant fixation. BMPs are known to stimulate new bone formation within allograft, but also known to accelerate graft resorption. Bisphosphonates are strong inhibitor of bone resorption. The aim of this study was to investigate whether the bisphosphonate zoledronate was able to counteract the accelerated graft resorption without interfering with the BMP induced bone formation. In the present study the two drugs alone and in combination were studied in our canine model of impaction bone grafting. We included 10 dogs in this study. Cancellous allograft bone grafts were soaked in either saline or zoledronate solution (0.005mg/mL) and then vehicle or BMP2 (0.15mg rhBMP2) was added. This produced four treatment groups: A) control, B) BMP2, C) zoledronate and D) BMP2+zoledronate. The allograft treated with A, B, C or D was impacted into a circumferential defect of 2.5mm around HA-coated porous Ti implants. Each dog received all four treatment groups with two implants in the distal part of each femur. The group with allograft soaked in zoledronate (C) showed better biomechanical fixation than all other groups (p<0.05). It had less allograft resorption compared to all other groups (p<0.005) without any statistically significant change in new bone formation. The addition of BMP2 to the allograft did not increase new bone formation significantly, but did accelerate allograft resorption. This was also the case where the allograft was treated with BMP2 and zoledronate in combination (D). This caused a decrease in mechanical implant fixation in both these groups compared to the control group, however only statistically significant for the BMP2 group compared to control. The study shows that topical zoledronate can be a valuable tool for augmenting bone grafts when administered optimally. The use of BMP2 in bone grafting procedures seems associated with a high risk of bone resorption and mechanical weakening. Copyright © 2017 Elsevier Inc. All rights reserved.

* Calvarial Bone Regeneration Is Enhanced by Sequential Delivery of FGF-2 and BMP-2 from Layer-by-Layer Coatings with a Biomimetic Calcium Phosphate Barrier Layer.

PubMed

Gronowicz, Gloria; Jacobs, Emily; Peng, Tao; Zhu, Li; Hurley, Marja; Kuhn, Liisa T

2017-12-01

A drug delivery coating for synthetic bone grafts has been developed to provide sequential delivery of multiple osteoinductive factors to better mimic aspects of the natural regenerative process. The coating is composed of a biomimetic calcium phosphate (bCaP) layer that is applied to a synthetic bone graft and then covered with a poly-l-Lysine/poly-l-Glutamic acid polyelectrolyte multilayer (PEM) film. Bone morphogenetic protein-2 (BMP-2) was applied before the coating process directly on the synthetic bone graft and then, bCaP-PEM was deposited followed by adsorption of fibroblast growth factor-2 (FGF-2) into the PEM layer. Cells access the FGF-2 immediately, while the bCaP-PEM temporally delays the cell access to BMP-2. In vitro studies with cells derived from mouse calvarial bones demonstrated that Sca-1 and CD-166 positive osteoblast progenitor cells proliferated in response to media dosing with FGF-2. Coated scaffolds with BMP-2 and FGF-2 were implanted in mouse calvarial bone defects and harvested at 1 and 3 weeks. After 1 week in vivo, proliferation of cells, including Sca-1+ progenitors, was observed with low dose FGF-2 and BMP-2 compared to BMP-2 alone, indicating that in vivo delivery of FGF-2 activated a similar population of cells as shown by in vitro testing. At 3 weeks, FGF-2 and BMP-2 delivery increased bone formation more than BMP-2 alone, particularly in the center of the defect, confirming that the proliferation of the Sca-1 positive osteoprogenitors by FGF-2 was associated with increased bone healing. Areas of bone mineralization were positive for double fluorochrome labeling of calcium and alkaline phosphatase staining of osteoblasts, along with increased TRAP+ osteoclasts, demonstrating active bone formation distinct from the bone-like collagen/hydroxyapatite scaffold. In conclusion, the addition of a bCaP layer to PEM delayed access to BMP-2 and allowed the FGF-2 stimulated progenitors to populate the scaffold before differentiating in response to BMP-2, leading to improved bone defect healing.

Topical Bisphosphonate Augments Fixation of Bone-grafted Hydroxyapatite coated Implants, BMP-2 causes Resorption-based decrease in Bone

PubMed Central

Baas, Jorgen; Vestermark, Marianne; Jensen, Thomas; Bechtold, Joan; Soballe, Kjeld; Jakobsen, Thomas

2017-01-01

Bone allograft is used in total joint artroplasties in order to enhance implant fixation. BMPs are known to stimulate new bone formation within allograft, but also known to accelerate graft resorption. Bisphosphonates are strong inhibitor of bone resorption. The aim of this study was to investigate whether the bisphosphonate zoledronate was able to counteract the accelerated graft resorption without interfering with the BMP induced bone formation. In the present study the two drugs alone and in combination were studied in our canine model of impaction bone grafting. We included 10 dogs in this study. Cancellous allograft bone grafts were soaked in either saline or zoledronate solution (0.005 mg/mL) and then vehicle or BMP2 (0.15 mg rhBMP2) was added. This produced four treatment groups: A) control B) BMP2 C) zoledronate and D) BMP2+ zoledronate. The allograft treated with A,B,C or D was impacted into a circumferential defect of 2.5 mm around HA-coated porous Ti implants. Each dog received all four treatment groups with two implants in the distal part of each femur. The group with allograft soaked in zoledronate (C) showed better biomechanical fixation than all other groups (p<0.05). It had less allograft resorption compared to all other groups (p<0.005) without any statistically significant change in new bone formation. The addition of BMP2 to the allograft did not increase new bone formation significantly, but did accelerate allograft resorption. This was also the case where the allograft was treated with BMP2 and zoledronate in combination (D). This caused a decrease in mechanical implant fixation in both these groups compared to the control group, however only statistically significant for the BMP2 group compared to control. The study shows that topical zoledronate can be a valuable tool for augmenting bone grafts when administered optimally. The use of BMP2 in bone grafting procedures seems associated with a high risk of bone resorption and mechanical weakening. PMID:28082076

Differential growth factor control of bone formation through osteoprogenitor differentiation.

PubMed

Chaudhary, L R; Hofmeister, A M; Hruska, K A

2004-03-01

The osteogenic factors bone morphogenetic protein (BMP-7), platelet-derived growth factor (PDGF)-BB, and fibroblast growth factor (FGF-2) regulate the recruitment of osteoprogenitor cells and their proliferation and differentiation into mature osteoblasts. However, their mechanisms of action on osteoprogenitor cell growth, differentiation, and bone mineralization remain unclear. Here, we tested the hypothesis that these osteogenic agents were capable of regulating osteoblast differentiation and bone formation in vitro. Normal human bone marrow stromal (HBMS) cells were treated with BMP-7 (40 ng ml(-1)), PDGF-BB (20 ng ml(-1)), FGF-2 (20 ng ml(-1)), or FGF-2 plus BMP-7 for 28 days in a serum-containing medium with 10 mM beta-glycerophosphate and 50 microg ml(-1) ascorbic acid. BMP-7 stimulated a morphological change to cuboidal-shaped cells, increased alkaline phosphatase (ALKP) activity, bone sialoprotein (BSP) gene expression, and alizarin red S positive nodule formation. Hydroxyapatite (HA) crystal deposition in the nodules was demonstrated by Fourier transform infrared (FTIR) spectroscopy only in BMP-7- and dexamethasone (DEX)-treated cells. DEX-treated cells appeared elongated and fibroblast-like compared to BMP-7-treated cells. FGF-2 did not stimulate ALKP, and cell morphology was dystrophic. PDGF-BB had little or no effect on ALKP activity and biomineralization. Alizarin Red S staining of cells and calcium assay indicated that BMP-7, DEX, and FGF-2 enhanced calcium mineral deposition, but FTIR spectroscopic analysis demonstrated no formation of HA similar to human bone in control, PDGF-BB-, and FGF-2-treated samples. Thus, FGF-2 stimulated amorphous octacalcium phosphate mineral deposition that failed to mature into HA. Interestingly, FGF-2 abrogated BMP-7-induced ALKP activity and HA formation. Results demonstrate that BMP-7 was competent as a sole factor in the differentiation of human bone marrow stromal cells to bone-forming osteoblasts confirmed by FTIR examination of mineralized matrix. Other growth factors, PDGF, and FGF-2 were incompetent as sole factors, and FGF-2 inhibited BMP-7-stimulated osteoblast differentiation.

Microstructured Titanium Regulates Interleukin Production by Osteoblasts, an Effect Modulated by Exogenous BMP-2

PubMed Central

Hyzy, Sharon; Olivares-Navarrete, Rene; Hutton, Daphne L.; Tan, Christian; Boyan, Barbara D.; Schwartz, Zvi

2013-01-01

Microtextured implant surfaces increase osteoblast differentiation in vitro and enhance bone-to-implant contact in vivo and clinically. These implants may be used in combination with recombinant human bone morphogenetic protein 2 (rhBMP-2) to enhance peri-implant bone formation. However, the effect of surface modifications alone or in combination with rhBMP-2 on osteoblast-produced inflammatory microenvironment is unknown. MG63 cells were cultured on tissue culture polystyrene or titanium substrates: smooth pretreated (PT, Ra=0.2μm), sandblasted/acid-etched (SLA, Ra=3.2μm), or hydrophilic-SLA (modSLA). Expression and protein production of pro-inflammatory interleukins (IL1b, IL6, IL8, IL17) and anti-inflammatory interleukins (IL10) were measured in cells with or without rhBMP-2. To determine which BMP signaling pathways were involved, cultures were incubated with BMP pathway inhibitors to blocking Smad (dorsomorphin), TAB/TAK1 ((5Z)-7-oxozeaenol), or PKA (H-8) signaling. Culture on rough SLA and modSLA surfaces decreased pro-inflammatory interleukins and increased anti-inflammatory IL10. This effect was negated in cells treated with rhBMP-2, which caused an increase in pro-inflammatory interleukins and a decrease in anti-inflammatory interleukins through TAB/TAK signaling. The results suggest that surface microtexture modulates the inflammatory process during osseointegration, an effect that may enhance healing. However, rhBMP-2 in combination with microtextured titanium implants can influence the effect of cells on these surfaces, and may adversely affect cells involved in osseointegration. PMID:23123301

BMP signaling in the human fetal ovary is developmentally regulated and promotes primordial germ cell apoptosis.

PubMed

Childs, Andrew J; Kinnell, Hazel L; Collins, Craig S; Hogg, Kirsten; Bayne, Rosemary A L; Green, Samira J; McNeilly, Alan S; Anderson, Richard A

2010-08-01

Primordial germ cells (PGCs) are the embryonic precursors of gametes in the adult organism, and their development, differentiation, and survival are regulated by a combination of growth factors collectively known as the germ cell niche. Although many candidate niche components have been identified through studies on mouse PGCs, the growth factor composition of the human PGC niche has not been studied extensively. Here we report a detailed analysis of the expression of components of the bone morphogenetic protein (BMP) signaling apparatus in the human fetal ovary, from postmigratory PGC proliferation to the onset of primordial follicle formation. We find developmentally regulated and reciprocal patterns of expression of BMP2 and BMP4 and identify germ cells to be the exclusive targets of ovarian BMP signaling. By establishing long-term cultures of human fetal ovaries in which PGCs are retained within their physiological niche, we find that BMP4 negatively regulates postmigratory PGC numbers in the human fetal ovary by promoting PGC apoptosis. Finally, we report expression of both muscle segment homeobox (MSX)1 and MSX2 in the human fetal ovary and reveal a selective upregulation of MSX2 expression in human fetal ovary in response to BMP4, suggesting this gene may act as a downstream effector of BMP-induced apoptosis in the ovary, as in other systems. These data reveal for the first time growth factor regulation of human PGC development in a physiologically relevant context and have significant implications for the development of cultures systems for the in vitro maturation of germ cells, and their derivation from pluripotent stem cells.

BMP-induced reprogramming of the neural retina into retinal pigment epithelium requires Wnt signalling

PubMed Central

Steinfeld, Jörg; Steinfeld, Ichie; Bausch, Alexander; Coronato, Nicola; Hampel, Meggi-Lee; Depner, Heike; Layer, Paul G.

2017-01-01

ABSTRACT In vertebrates, the retinal pigment epithelium (RPE) and photoreceptors of the neural retina (NR) comprise a functional unit required for vision. During vertebrate eye development, a conversion of the RPE into NR can be induced by growth factors in vivo at optic cup stages, but the reverse process, the conversion of NR tissue into RPE, has not been reported. Here, we show that bone morphogenetic protein (BMP) signalling can reprogram the NR into RPE at optic cup stages in chick. Shortly after BMP application, expression of Microphthalmia-associated transcription factor (Mitf) is induced in the NR and selective cell death on the basal side of the NR induces an RPE-like morphology. The newly induced RPE differentiates and expresses Melanosomalmatrix protein 115 (Mmp115) and RPE65. BMP-induced Wnt2b expression is observed in regions of the NR that become pigmented. Loss of function studies show that conversion of the NR into RPE requires both BMP and Wnt signalling. Simultaneous to the appearance of ectopic RPE tissue, BMP application reprogrammed the proximal RPE into multi-layered retinal tissue. The newly induced NR expresses visual segment homeobox-containing gene (Vsx2), and the ganglion and photoreceptor cell markers Brn3α and Visinin are detected. Our results show that high BMP concentrations are required to induce the conversion of NR into RPE, while low BMP concentrations can still induce transdifferentiation of the RPE into NR. This knowledge may contribute to the development of efficient standardized protocols for RPE and NR generation for cell replacement therapies. PMID:28546339

The spatial patterning of mouse cone opsin expression is regulated by BMP signaling through downstream effector COUP-TF nuclear receptors

PubMed Central

Satoh, Shinya; Tang, Ke; Iida, Atsumi; Inoue, Mariko; Kodama, Tatsuhiko; Tsai, Sophia Y.; Tsai, Ming-Jer; Furuta, Yasuhide; Watanabe, Sumiko

2009-01-01

Cone photopigments, known as opsins, are pivotal elements and the first detection module employed in color vision. In mice, cone photoreceptors are distributed throughout the retina, and S- and M-opsins have unique expression patterns in the retina with a gradient along the dorsoventral axis; however, the mechanisms regulating the spatial patterning of cone opsin expression have not been well documented. The purpose of this study was to define the mechanisms regulating the spatial patterning of cone opsin expression. By analyzing knockouts for bone morphogenetic protein (BMP) signaling, we found an essential role for BMP in forming cone opsin expression patterns in the retina; however, BMP signaling is activated only transiently in the dorsal half of the retina during early retinal development. Thus, BMP is not likely to play a direct role in opsin gene expression, which starts at a later stage of retinal development. We identified the chicken ovalbumin upstream promoter-transcription factor (COUP-TF) nuclear receptor as a link between BMP and opsin expression. BMP signaling is essential for the correct dorsoventral spatial expression of COUP-TFI and -TFII. Through gain- and loss-of-function analyses, we found that both COUP-TFI and -TFII are required to suppress S-opsin expression in the dorsal retina but that only COUP-TFI plays an essential role in suppressing M-opsin expression in the ventral retina. Based on these findings, we propose a new molecular cascade involving BMP and COUP-TFs that conveys dorsoventral information to direct the expression of cone opsins during retinal development. PMID:19812316

Abnormal Uterine Bleeding Is Associated With Increased BMP7 Expression in Human Endometrium.

PubMed

Richards, Elliott G; El-Nashar, Sherif A; Schoolmeester, John K; Keeney, Gary L; Mariani, Andrea; Hopkins, Matthew R; Dowdy, Sean C; Daftary, Gaurang S; Famuyide, Abimbola O

2017-05-01

Abnormal uterine bleeding (AUB), a common health concern of women, is a heterogeneous clinical entity that is traditionally categorized into organic and nonorganic causes. Despite varied pharmacologic treatments, few offer sustained efficacy, as most are empiric, unfocused, and do not directly address underlying dysregulated molecular mechanisms. Characterization of such molecular derangements affords the opportunity to develop and use novel, more successful treatments for AUB. Given its implication in other organ systems, we hypothesized that bone morphogenetic protein (BMP) expression is altered in patients with AUB and hence comprehensively investigated dysregulation of BMP signaling pathways by systematically screening 489 samples from 365 patients for differences in the expression of BMP2, 4, 6, and 7 ligands, BMPR1A and B receptors, and downstream SMAD4, 6, and 7 proteins. Expression analysis was correlated clinically with data abstracted from medical records, including bleeding history, age at procedure, ethnicity, body mass index, hormone treatment, and histological diagnosis of fibroids, polyps, adenomyosis, hyperplasia, and cancer. Expression of BMP7 ligand was significantly increased in patients with AUB (H-score: 18.0 vs 26.7; P < .0001). Patients reporting heavy menstrual bleeding (menorrhagia) as their specific AUB pattern demonstrated significantly higher BMP7 expression. Significantly, no differences in the expression of any other BMP ligands, receptors, or SMAD proteins were observed in this large patient cohort. However, expression of BMPR1A, BMPR1B, and SMAD4 was significantly decreased in cancer compared to benign samples. Our study demonstrates that BMP7 is a promising target for future investigation and pharmacologic treatment of AUB.

BMP2 induces PANC-1 cell invasion by MMP-2 overexpression through ROS and ERK.

PubMed

Liu, Jun; Ben, Qi-Wen; Yao, Wei-Yan; Zhang, Jian-Jun; Chen, Da-Fan; He, Xiang-Yi; Li, Lei; Yuan, Yao-Zong

2012-06-01

The emerging roles of bone morphogenetic proteins (BMPs) in the initiation and progression of multiple cancers have drawn great attention in cancer research. We hypothesized that BMP2 promotes cancer metastasis by modulating MMP-2 secretion and activity through intracellular ROS regulation and ERK activation in human pancreatic cancer. Our data show that stimulation of PANC-1 cells with BMP2 induced MMP-2 secretion and activation, associated with decreased E-cadherin expression, resulting in epithelial-to-mesenchymal transformation (EMT) and cell invasion. Blockade of ROS by the ROS scavenger, 2-MPG, abolished cell invasion, inhibited the EMT process and decreased MMP-2 expression, suggesting ROS accumulation caused an increase in MMP-2 expression in BMP2-stimulated PANC-1 cell invasion. Furthermore, treatment of PANC-1 cells with 2-MPG or ERK inhibitor PD98059 reduced the phosphorylation of ERK, resulting in attenuation of BMP2-induced cell invasion and MMP-2 activation. Taken together, these results suggest that BMP2 induces the cell invasion of PANC-1 cells by enhancing MMP-2 secretion and acting through ROS accumulation and ERK activation.

Image Size Variation Influence on Corrupted and Non-viewable BMP Image

NASA Astrophysics Data System (ADS)

Azmi, Tengku Norsuhaila T.; Azma Abdullah, Nurul; Rahman, Nurul Hidayah Ab; Hamid, Isredza Rahmi A.; Chai Wen, Chuah

2017-08-01

Image is one of the evidence component seek in digital forensics. Joint Photographic Experts Group (JPEG) format is most popular used in the Internet because JPEG files are very lossy and easy to compress that can speed up Internet transmitting processes. However, corrupted JPEG images are hard to recover due to the complexities of determining corruption point. Nowadays Bitmap (BMP) images are preferred in image processing compared to another formats because BMP image contain all the image information in a simple format. Therefore, in order to investigate the corruption point in JPEG, the file is required to be converted into BMP format. Nevertheless, there are many things that can influence the corrupting of BMP image such as the changes of image size that make the file non-viewable. In this paper, the experiment indicates that the size of BMP file influences the changes in the image itself through three conditions, deleting, replacing and insertion. From the experiment, we learnt by correcting the file size, it can able to produce a viewable file though partially. Then, it can be investigated further to identify the corruption point.

[Experimental study on repair of the defect of the pars interarticularis in rat with bone morphogenetic protein and fibrin glue].

PubMed

Nakamura, T

1992-07-01

The possibility of repairing the defect of the pars interarticularis (pars defect) with Bone Morphogenetic Protein (BMP) and fibrin glue was studied. The pars defect established in the 5th lumbar vertebra of Wistar rat was treated with surgical implantation of a composite consisting of BMP, fibrin glue and autologous cancellous bone. At 3, 6, 9 and 12 weeks after implantation, the osteoinductive activity in the pars defect was observed histologically and compared with that of other composite implants such as BMP with fibrin glue, autologous cancellous bone alone and autologous cancellous bone with fibrin glue. Although perfect bone fusion was not obtained with any of the composites employed, a significant increase in bone formation was seen in a composite of BMP, fibrin glue and autologous cancellous bone (p less than 0.01) as compared with that seen in the others. Consequently, implantation of BMP and fibrin glue combined with some biomaterials which support osteo-induction of BMP and stabilize the pars defect might be successfully applied to repair the pars defect.

Production of BMP4 by endothelial cells is crucial for endogenous thymic regeneration.

PubMed

Wertheimer, Tobias; Velardi, Enrico; Tsai, Jennifer; Cooper, Kirsten; Xiao, Shiyun; Kloss, Christopher C; Ottmüller, Katja J; Mokhtari, Zeinab; Brede, Christian; deRoos, Paul; Kinsella, Sinéad; Palikuqi, Brisa; Ginsberg, Michael; Young, Lauren F; Kreines, Fabiana; Lieberman, Sophia R; Lazrak, Amina; Guo, Peipei; Malard, Florent; Smith, Odette M; Shono, Yusuke; Jenq, Robert R; Hanash, Alan M; Nolan, Daniel J; Butler, Jason M; Beilhack, Andreas; Manley, Nancy R; Rafii, Shahin; Dudakov, Jarrod A; van den Brink, Marcel R M

2018-01-12

The thymus is not only extremely sensitive to damage but also has a remarkable ability to repair itself. However, the mechanisms underlying this endogenous regeneration remain poorly understood, and this capacity diminishes considerably with age. We show that thymic endothelial cells (ECs) comprise a critical pathway of regeneration via their production of bone morphogenetic protein 4 (BMP4) ECs increased their production of BMP4 after thymic damage, and abrogating BMP4 signaling or production by either pharmacologic or genetic inhibition impaired thymic repair. EC-derived BMP4 acted on thymic epithelial cells (TECs) to increase their expression of Foxn1 , a key transcription factor involved in TEC development, maintenance, and regeneration, and its downstream targets such as Dll4 , a key mediator of thymocyte development and regeneration. These studies demonstrate the importance of the BMP4 pathway in endogenous tissue regeneration and offer a potential clinical approach to enhance T cell immunity. Copyright © 2018 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

Lack of Association of Bone Morphogenetic Protein 2 Gene Haplotypes with Bone Mineral Density, Bone Loss, or Risk of Fractures in Men

PubMed Central

Varanasi, Satya S.; Tuck, Stephen P.; Mastana, Sarabjit S.; Dennison, Elaine; Cooper, Cyrus; Vila, Josephine; Francis, Roger M.; Datta, Harish K.

2011-01-01

Introduction. The association of bone morphogenetic protein 2 (BMP2) with BMD and risk of fracture was suggested by a recent linkage study, but subsequent studies have been contradictory. We report the results of a study of the relationship between BMP2 genotypes and BMD, annual change in BMD, and risk of fracture in male subjects. Materials and Methods. We tested three single-nucleotide polymorphisms (SNPs) across the BMP2 gene, including Ser37Ala SNP, in 342 Caucasian Englishmen, comprising 224 control and 118 osteoporotic subjects. Results. BMP2 SNP1 (Ser37Ala) genotypes were found to have similar low frequency in control subjects and men with osteoporosis. The major informative polymorphism, BMP2 SNP3 (Arg190Ser), showed no statistically significant association with weight, height, BMD, change in BMD at hip or lumbar spine, and risk of fracture. Conclusion. There were no genotypic or haplotypic effects of the BMP2 candidate gene on BMD, change in BMD, or fracture risk identified in this cohort. PMID:22013543

PDGF-AA promotes osteogenic differentiation and migration of mesenchymal stem cell by down-regulating PDGFRα and derepressing BMP-Smad1/5/8 signaling.

PubMed

Li, Anna; Xia, Xuechun; Yeh, James; Kua, Huiyi; Liu, Huijuan; Mishina, Yuji; Hao, Aijun; Li, Baojie

2014-01-01

Platelet-derived growth factors (PDGFs) play important roles in skeletal development and bone fracture healing, yet how PDGFs execute their functions remains incompletely understood. Here we show that PDGF-AA, but not -AB or -BB, could activate the BMP-Smad1/5/8 pathway in mesenchymal stem cells (MSCs), which requires BMPRIA as well as PDGFRα. PDGF-AA promotes MSC osteogenic differentiation through the BMP-Smad1/5/8-Runx2/Osx axis and MSC migration via the BMP-Smad1/5/8-Twist1/Atf4 axis. Mechanistic studies show that PDGF-AA activates BMP-Smad1/5/8 signaling by feedback down-regulating PDGFRα, which frees BMPRI and allows for BMPRI-BMPRII complex formation to activate smad1/5/8, using BMP molecules in the microenvironment. This study unravels a physical and functional interaction between PDGFRα and BMPRI, which plays an important role in MSC differentiation and migration, and establishes a link between PDGF-AA and BMPs pathways, two essential regulators of embryonic development and tissue homeostasis.

BMP regulates regional gene expression in the dorsal otocyst through canonical and non-canonical intracellular pathways

PubMed Central

2016-01-01

The inner ear consists of two otocyst-derived, structurally and functionally distinct components: the dorsal vestibular and ventral auditory compartments. BMP signaling is required to form the vestibular compartment, but how it complements other required signaling molecules and acts intracellularly is unknown. Using spatially and temporally controlled delivery of signaling pathway regulators to developing chick otocysts, we show that BMP signaling regulates the expression of Dlx5 and Hmx3, both of which encode transcription factors essential for vestibular formation. However, although BMP regulates Dlx5 through the canonical SMAD pathway, surprisingly, it regulates Hmx3 through a non-canonical pathway involving both an increase in cAMP-dependent protein kinase A activity and the GLI3R to GLI3A ratio. Thus, both canonical and non-canonical BMP signaling establish the precise spatiotemporal expression of Dlx5 and Hmx3 during dorsal vestibular development. The identification of the non-canonical pathway suggests an intersection point between BMP and SHH signaling, which is required for ventral auditory development. PMID:27151948

Flow-induced protein kinase A–CREB pathway acts via BMP signaling to promote HSC emergence

PubMed Central

Kim, Peter Geon; Nakano, Haruko; Das, Partha P.; Chen, Michael J.; Rowe, R. Grant; Chou, Stephanie S.; Ross, Samantha J.; Sakamoto, Kathleen M.; Zon, Leonard I.; Schlaeger, Thorsten M.; Orkin, Stuart H.; Nakano, Atsushi

2015-01-01

Fluid shear stress promotes the emergence of hematopoietic stem cells (HSCs) in the aorta–gonad–mesonephros (AGM) of the developing mouse embryo. We determined that the AGM is enriched for expression of targets of protein kinase A (PKA)–cAMP response element-binding protein (CREB), a pathway activated by fluid shear stress. By analyzing CREB genomic occupancy from chromatin-immunoprecipitation sequencing (ChIP-seq) data, we identified the bone morphogenetic protein (BMP) pathway as a potential regulator of CREB. By chemical modulation of the PKA–CREB and BMP pathways in isolated AGM VE-cadherin+ cells from mid-gestation embryos, we demonstrate that PKA–CREB regulates hematopoietic engraftment and clonogenicity of hematopoietic progenitors, and is dependent on secreted BMP ligands through the type I BMP receptor. Finally, we observed blunting of this signaling axis using Ncx1-null embryos, which lack a heartbeat and intravascular flow. Collectively, we have identified a novel PKA–CREB–BMP signaling pathway downstream of shear stress that regulates HSC emergence in the AGM via the endothelial-to-hematopoietic transition. PMID:25870201

Inactivation of Bmp4 from the Tbx1 Expression Domain Causes Abnormal Pharyngeal Arch Artery and Cardiac Outflow Tract Remodeling

PubMed Central

Nie, Xuguang; Brown, Christopher B.; Wang, Qin; Jiao, Kai

2011-01-01

Maldevelopment of outflow tract and aortic arch arteries is among the most common forms of human congenital heart diseases. Both Bmp4 and Tbx1 are known to play critical roles during cardiovascular development. Expression of these two genes partially overlaps in pharyngeal arch areas in mouse embryos. In this study, we applied a conditional gene inactivation approach to test the hypothesis that Bmp4 expressed from the Tbx1 expression domain plays a critical role for normal development of outflow tract and pharyngeal arch arteries. We showed that inactivation of Bmp4 from Tbx1-expressing cells leads to the spectrum of deformities resembling the cardiovascular defects observed in human DiGeorge syndrome patients. Inactivation of Bmp4 from the Tbx1 expression domain did not cause patterning defects, but affected remodeling of outflow tract and pharyngeal arch arteries. Our further examination revealed that Bmp4 is required for normal recruitment/differentiation of smooth muscle cells surrounding the PAA4 and survival of outflow tract cushion mesenchymal cells. PMID:21123999

Fine mapping of the human bone morphogenetic protein-4 gene (BMP4) to chromosome 14q22-q23 by in situ hybridization

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wijngaard, A. van den; Boersma, C.J.C.; Olijve, W.

Bone morphogenetic protein-4 (BMP-4) is a member of the transforming growth factor-{beta} (TGF-{beta}) superfamily and is involved in morphogenesis and bone cell differentiation. Recombinant BMP-4 can induce ectopic cartilage and bone formation when implanted subcutaneously or intramuscularly in rodents. This ectopic bone formation process resembles the process of bone formation during embryogenesis and fracture healing. A cosmid clone containing the complete human bone morphogenetic protein-4 gene (BMP4) was isolated (details to be published elsewhere) and used as a probe to determine the precise chromosomal localization of the human BMP4 gene. This cosmid clone was labeled with biotin-14-dATP and hybridized inmore » situ to chromosomal preparations of metaphase cells as described previously. In 20 metaphase preparations, an intense and specific fluorescence signal (FITC) was detected on the q arm of chromosome 14. The DAPI-counterstained chromosomes were computer-converted into GTG-like banding patterns, allowing the regional localization of BMP4 within 14q22-q23. 10 refs., 1 fig.« less