Reprogramming retinal neurons and standardized quantification of their differentiation in 3-dimensional retinal cultures

PubMed Central

Hiler, Daniel J.; Barabas, Marie E.; Griffiths, Lyra M.; Dyer, Michael A.

2017-01-01

Postmitotic differentiated neurons are among the most difficult cells to reprogram into induced pluripotent stem cells (iPSCs) because they have poor viability when cultured as dissociated cells. Other protocols to reprogram postmitotic neurons have required the inactivation of the p53 tumor suppressor. We describe a method that does not require p53 inactivation and induces reprogramming in cells purified from the retinae of reprogrammable mice in aggregates with wild-type retinal cells. After the first 10 days of reprogramming, the aggregates are then dispersed and plated on irradiated feeder cells to propagate and isolate individual iPSC clones. The reprogramming efficiency of different neuronal populations at any stage of development can be quantitated using this protocol. Reprogramming retinal neurons with this protocol will take 56 days, and these retina-derived iPSCs can undergo retinal differentiation to produce retinae in 34 days. In addition, we describe a quantitative assessment of retinal differentiation from these neuron-derived iPSCs called STEM-RET. The procedure quantitates eye field specification, optic cup formation, and retinal differentiation in 3-dimensional cultures using molecular, cellular and morphological criteria. An advanced level of cell culture experience is required to carry out this protocol. PMID:27658012

Psychosocial predictors of affect in adult patients undergoing orthodontic treatment.

PubMed

Peñacoba, Cecilia; González, M José; Santos, Noelia; Romero, Martín

2014-02-01

In this paper we propose to study the role of psychosocial variables in affect in adult patients undergoing orthodontic treatment, considering that affect is a key variable in treatment adherence. Seventy-four patients (average age 33,24 ± 10,56) with metal multibracket-fixed orthodontic treatment were included. Patients were assessed twice. The first stage, at the beginning of treatment, included assessment of dental impact (Psychosocial Impact of Dental Aesthetics Questionnaire), trait anxiety (State-Trait Anxiety Inventory), self-esteem (Rosenberg's self-esteem scale), and self-efficacy (General Self-efficacy Scale). In the second stage, 6 months later, positive and negative affect towards treatment was assessed using the Positive and Negative Affect Scale. Dental social impact differentiates between patients with high and low negative affect, while self-efficacy differentiates between patients with high and low positive affect. Trait anxiety and self-esteem differentiate between both types of affect (positive and negative). Trait anxiety and self-esteem (when trait anxiety weight is controlled) are significant predictor variables of affective balance. These results have important practical implications, because it seems essential to adopt a bio-psychosocial model incorporating assessment methods focusing on day-to-day changes in mood and well-being.

The relationship between right-sided tumour location, tumour microenvironment, systemic inflammation, adjuvant therapy and survival in patients undergoing surgery for colon and rectal cancer.

PubMed

Patel, Meera; McSorley, Stephen T; Park, James H; Roxburgh, Campbell S D; Edwards, Joann; Horgan, Paul G; McMillan, Donald C

2018-03-06

There has been an increasing interest in the role of tumour location in the treatment and prognosis of patients with colorectal cancer (CRC), specifically in the adjuvant setting. Together with genomic data, this has led to the proposal that right-sided and left-sided tumours should be considered as distinct biological and clinical entities. The aim of the present study was to examine the relationship between tumour location, tumour microenvironment, systemic inflammatory response (SIR), adjuvant chemotherapy and survival in patients undergoing potentially curative surgery for stage I-III colon and rectal cancer. Clinicopathological characteristics were extracted from a prospective database. MMR and BRAF status was determined using immunohistochemistry. The tumour microenvironment was assessed using routine H&E pathological sections. SIR was assessed using modified Glasgow Prognostic Score (mGPS), neutrophil:lymphocyte ratio (NLR), neutrophil:platelet score (NPS) and lymphocyte:monocyte ratio (LMR). Overall, 972 patients were included. The majority were over 65 years (68%), male (55%), TNM stage II/III (82%). In all, 40% of patients had right-sided tumours and 31% had rectal cancers. Right-sided tumour location was associated with older age (P=0.001), deficient MMR (P=0.005), higher T stage (P<0.001), poor tumour differentiation (P<0.001), venous invasion (P=0.021), and high CD3 + within cancer cell nests (P=0.048). Right-sided location was consistently associated with a high SIR, mGPS (P<0.001) and NPS (P<0.001). There was no relationship between tumour location, adjuvant chemotherapy (P=0.632) or cancer-specific survival (CSS; P=0.377). In those 275 patients who received adjuvant chemotherapy, right-sided location was not associated with the MMR status (P=0.509) but was associated with higher T stage (P=0.001), venous invasion (P=0.036), CD3 + at the invasive margin (P=0.033) and CD3 + within cancer nests (P=0.012). There was no relationship between tumour location, SIR or CSS in the adjuvant group. Right-sided tumour location was associated with an elevated tumour lymphocytic infiltrate and an elevated SIR. There was no association between tumour location and survival in the non-adjuvant or adjuvant setting in patients undergoing potentially curative surgery for stage I-III colon and rectal cancer.

An Optimized and Simplified System of Mouse Embryonic Stem Cell Cardiac Differentiation for the Assessment of Differentiation Modifiers

PubMed Central

Hartman, Matthew E.; Librande, Jason R.; Medvedev, Ivan O.; Ahmad, Rabiah N.; Moussavi-Harami, Farid; Gupta, Pritha P.; Chien, Wei-Ming; Chin, Michael T.

2014-01-01

Generating cardiomyocytes from embryonic stem cells is an important technique for understanding cardiovascular development, the origins of cardiovascular diseases and also for providing potential reagents for cardiac repair. Numerous methods have been published but often are technically challenging, complex, and are not easily adapted to assessment of specific gene contributions to cardiac myocyte differentiation. Here we report the development of an optimized protocol to induce the differentiation of mouse embryonic stem cells to cardiac myocytes that is simplified and easily adapted for genetic studies. Specifically, we made four critical findings that distinguish our protocol: 1) mouse embryonic stem cells cultured in media containing CHIR99021 and PD0325901 to maintain pluripotency will efficiently form embryoid bodies containing precardiac mesoderm when cultured in these factors at a reduced dosage, 2) low serum conditions promote cardiomyocyte differentiation and can be used in place of commercially prepared StemPro nutrient supplement, 3) the Wnt inhibitor Dkk-1 is dispensable for efficient cardiac differentiation and 4) tracking differentiation efficiency may be done with surface expression of PDGFRα alone. In addition, cardiac mesodermal precursors generated by this system can undergo lentiviral infection to manipulate the expression of specific target molecules to assess effects on cardiac myocyte differentiation and maturation. Using this approach, we assessed the effects of CHF1/Hey2 on cardiac myocyte differentiation, using both gain and loss of function. Overexpression of CHF1/Hey2 at the cardiac mesoderm stage had no apparent effect on cardiac differentiation, while knockdown of CHF1/Hey2 resulted in increased expression of atrial natriuretic factor and connexin 43, suggesting an alteration in the phenotype of the cardiomyocytes. In summary we have generated a detailed and simplified protocol for generating cardiomyocytes from mES cells that is optimized for investigating factors that affect cardiac differentiation. PMID:24667642

The non-canonical Wnt-PCP pathway shapes the mouse caudal neural plate.

PubMed

López-Escobar, Beatriz; Caro-Vega, José Manuel; Vijayraghavan, Deepthi S; Plageman, Timothy F; Sanchez-Alcazar, José A; Moreno, Roberto Carlos; Savery, Dawn; Márquez-Rivas, Javier; Davidson, Lance A; Ybot-González, Patricia

2018-05-08

The last stage of neural tube (NT) formation involves closure of the caudal neural plate (NP), an embryonic structure formed by neuromesodermal progenitors and newly differentiated cells that becomes incorporated into the NT. Here, we show in mouse that, as cell specification progresses, neuromesodermal progenitors and their progeny undergo significant changes in shape prior to their incorporation into the NT. The caudo-rostral progression towards differentiation is coupled to a gradual reliance on a unique combination of complex mechanisms that drive tissue folding, involving pulses of apical actomyosin contraction and planar polarised cell rearrangements, all of which are regulated by the Wnt-PCP pathway. Indeed, when this pathway is disrupted, either chemically or genetically, the polarisation and morphology of cells within the entire caudal NP is disturbed, producing delays in NT closure. The most severe disruptions of this pathway prevent caudal NT closure and result in spina bifida. In addition, a decrease in Vangl2 gene dosage also appears to promote more rapid progression towards a neural fate, but not the specification of more neural cells. © 2018. Published by The Company of Biologists Ltd.

Plant respirometer enables high resolution of oxygen consumption rates

NASA Technical Reports Server (NTRS)

Foster, D. L.

1966-01-01

Plant respirometer permits high resolution of relatively small changes in the rate of oxygen consumed by plant organisms undergoing oxidative metabolism in a nonphotosynthetic state. The two stage supply and monitoring system operates by a differential pressure transducer and provides a calibrated output by digital or analog signals.

Chemoablated mouse seminiferous tubular cells enriched for very small embryonic-like stem cells undergo spontaneous spermatogenesis in vitro.

PubMed

Anand, Sandhya; Patel, Hiren; Bhartiya, Deepa

2015-04-18

Extensive research is ongoing to empower cancer survivors to have biological parenthood. For this, sperm are cryopreserved prior to therapy and in younger children testicular biopsies are cryopreserved with a hope to mature the germ cells into sperm later on for assisted reproduction. In addition, lot of hope was bestowed on pluripotent embryonic and induced pluripotent stem cells to differentiate into sperm and oocytes. However, obtaining functional gametes from pluripotent stem cells still remains a distant dream and major bottle-neck appears to be their inefficient differentiation into primordial germ cells (PGCs). There exists yet another population of pluripotent stem cells termed very small embryonic-like stem cells (VSELs) in adult body organs including gonads. We have earlier reported that busulphan (25 mg/Kg) treatment to 4 weeks old mice destroys actively dividing cells and sperm but VSELs survive and differentiate into sperm when a healthy niche is provided in vivo. Mouse testicular VSELs that survived busulphan treatment were cultured for 3 weeks. A mix of surviving cells in seminiferous tubules (VSELs, possibly few spermatogonial stem cells and Sertoli cells) were cultured using Sertoli cells conditioned medium containing fetal bovine serum, follicle stimulating hormone and with no additional growth factors. Stem cells underwent proliferation and clonal expansion in culture and spontaneously differentiated into sperm whereas Sertoli cells attached and provided a somatic support. Transcripts specific for various stages of spermatogenesis were up-regulated by qRT-PCR studies on day 7 suggesting VSELs (Sca1) and SSCs (Gfra) proliferate (Pcna), undergo spermatogenesis (spermatocyte specific marker prohibitin), meiosis (Scp3) and differentiate into sperm (post-meiotic marker protamine). Process of spermatogenesis and spermiogenesis was replicated in vitro starting with testicular cells that survived busulphan treatment. We have earlier reported similar ability of ovarian VSELs enriched in the ovary surface epithelial cells to form oocyte-like structures in vitro. This striking potential of spontaneous differentiation of primitive testicular cells including VSELs that survive chemotherapy is being described for the first time in the present study.

Adult-specific insulin-producing neurons in Drosophila melanogaster.

PubMed

Ohhara, Yuya; Kobayashi, Satoru; Yamakawa-Kobayashi, Kimiko; Yamanaka, Naoki

2018-06-01

Holometabolous insects undergo metamorphosis to reorganize their behavioral and morphological features into adult-specific ones. In the central nervous system (CNS), some larval neurons undergo programmed cell death, whereas others go through remodeling of axonal and dendritic arbors to support functions of re-established adult organs. Although there are multiple neuropeptides that have stage-specific roles in holometabolous insects, the reorganization pattern of the entire neuropeptidergic system through metamorphosis still remains largely unclear. In this study, we conducted a mapping and lineage tracing of peptidergic neurons in the larval and adult CNS by using Drosophila genetic tools. We found that Diuretic hormone 44-producing median neurosecretory cells start expressing Insulin-like peptide 2 in the pharate adult stage. This neuronal cluster projects to the corpora cardiaca and dorsal vessel in both larval and adult stages, and also innervates an adult-specific structure in the digestive tract, the crop. We propose that the adult-specific insulin-producing cells may regulate functions of the digestive system in a stage-specific manner. Our study provides a neuroanatomical basis for understanding remodeling of the neuropeptidergic system during insect development and evolution. © 2018 Wiley Periodicals, Inc.

Adherence to treatment, emotional state and quality of life in patients with end-stage renal disease undergoing dialysis.

PubMed

García-Llana, Helena; Remor, Eduardo; Selgas, Rafael

2013-02-01

A low rate of adherence to treatment is a widespread problem of great clinical relevance among dialysis patients. The objective of the present study is to determine the relationship between adherence, emotional state (depression, anxiety, and perceived stress), and health-related quality of life (HRQOL) in renal patients undergoing dialysis. Two patient groups (30 in hemodialysis and 31 in peritoneal dialysis) participated in this study. We evaluated aspects of adherence, depression, anxiety, perceived stress, and HRQOL with self-report and standardized instruments. Peritoneal dialysis patients reported significantly higher levels of adherence to treatment and better HRQOL in Physical Function and Bodily Pain domains. Depression level is associated with HRQOL indicators. We did not find any differences regarding specific adherence to antihypertensive and phosphate binder drugs or in psychological variables depending on the modality of dialysis. Patients with adherence to antihypertensive drugs show better physical HRQOL. The predictors of HRQOL in dialysis patients were: work, gender and depression. Our results suggest that the modality of dialysis does not differentially affect the emotional state or specific adherence to drugs, but it is nevertheless related to their overall adherence to treatment and to their HRQOL.

In vivo differentiation of human periodontal ligament cells leads to formation of dental hard tissue.

PubMed

Wolf, M; Lossdörfer, S; Abuduwali, N; Meyer, R; Kebir, S; Götz, W; Jäger, A

2013-11-01

Following trauma, periodontal disease, or orthodontic tooth movement, residual periodontal ligament (PDL) cells at the defect site are considered mandatory for successful regeneration of the injured structures. Recent developments in tissue engineering focus, as one pillar, on the transplantation of PDL cells to support periodontal regeneration processes. Here, we examined the ability of osteogenically predifferentiated PDL cells to undergo further osteoblastic or cementoblastic differentiation and to mineralize their extracellular matrix when transplanted in an in vivo microenvironment. Using collagen sponges as carriers, osteogenically predifferentiated human PDL cells were transplanted subcutaneously into six immunocompromised CD-1® nude mice. Following explantation after 28 days, osteogenic and cementogenic marker protein expression was visualized immunohistochemically. After 28 days, transplanted PDL cells revealed both cellular, cytoplasmatic and extracellular immunoreactivity for the chosen markers alkaline phosphatase, osteopontin, PTH-receptor 1, and osteocalcin. Specific osteogenic and cementoblastic differentiation was demonstrated by RUNX2 and CEMP1 immunoreactivity. Early stages of mineralization were demonstrated by calcium and phosphate staining. Our results reinforce the previously published reports of PDL cell mineralization in vivo and further demonstrate the successful induction of specific osteogenic and cementogenic differentiation of transplanted human PDL cells in vivo. These findings reveal promising possibilities for supporting periodontal remodeling and regeneration processes with PDL cells being potential target cells with which to influence the process of orthodontically induced root resorption.

Expression of GFP under the control of the RNA helicase VASA permits fluorescence-activated cell sorting isolation of human primordial germ cells.

PubMed

Tilgner, Katarzyna; Atkinson, Stuart P; Yung, Sun; Golebiewska, Anna; Stojkovic, Miodrag; Moreno, Ruben; Lako, Majlinda; Armstrong, Lyle

2010-01-01

The isolation of significant numbers of human primordial germ cells at several developmental stages is important for investigations of the mechanisms by which they are able to undergo epigenetic reprogramming. Only small numbers of these cells can be obtained from embryos of appropriate developmental stages, so the differentiation of human embryonic stem cells is essential to obtain sufficient numbers of primordial germ cells to permit epigenetic examination. Despite progress in the enrichment of human primordial germ cells using fluorescence-activated cell sorting (FACS), there is still no definitive marker of the germ cell phenotype. Expression of the widely conserved RNA helicase VASA is restricted to germline cells, but in contrast to species such as Mus musculus in which reporter constructs expressing green fluorescent protein (GFP) under the control of a Vasa promoter have been developed, such reporter systems are lacking in human in vitro models. We report here the generation and characterization of human embryonic stem cell lines stably carrying a VASA-pEGFP-1 reporter construct that expresses GFP in a population of differentiating human embryonic stem cells that show expression of characteristic markers of primordial germ cells. This population shows a different pattern of chromatin modifications to those obtained by FACS enrichment of Stage Specific Antigen one expressing cells in our previous publication.

Whole-genome fingerprint of the DNA methylome during human B cell differentiation.

PubMed

Kulis, Marta; Merkel, Angelika; Heath, Simon; Queirós, Ana C; Schuyler, Ronald P; Castellano, Giancarlo; Beekman, Renée; Raineri, Emanuele; Esteve, Anna; Clot, Guillem; Verdaguer-Dot, Néria; Duran-Ferrer, Martí; Russiñol, Nuria; Vilarrasa-Blasi, Roser; Ecker, Simone; Pancaldi, Vera; Rico, Daniel; Agueda, Lidia; Blanc, Julie; Richardson, David; Clarke, Laura; Datta, Avik; Pascual, Marien; Agirre, Xabier; Prosper, Felipe; Alignani, Diego; Paiva, Bruno; Caron, Gersende; Fest, Thierry; Muench, Marcus O; Fomin, Marina E; Lee, Seung-Tae; Wiemels, Joseph L; Valencia, Alfonso; Gut, Marta; Flicek, Paul; Stunnenberg, Hendrik G; Siebert, Reiner; Küppers, Ralf; Gut, Ivo G; Campo, Elías; Martín-Subero, José I

2015-07-01

We analyzed the DNA methylome of ten subpopulations spanning the entire B cell differentiation program by whole-genome bisulfite sequencing and high-density microarrays. We observed that non-CpG methylation disappeared upon B cell commitment, whereas CpG methylation changed extensively during B cell maturation, showing an accumulative pattern and affecting around 30% of all measured CpG sites. Early differentiation stages mainly displayed enhancer demethylation, which was associated with upregulation of key B cell transcription factors and affected multiple genes involved in B cell biology. Late differentiation stages, in contrast, showed extensive demethylation of heterochromatin and methylation gain at Polycomb-repressed areas, and genes with apparent functional impact in B cells were not affected. This signature, which has previously been linked to aging and cancer, was particularly widespread in mature cells with an extended lifespan. Comparing B cell neoplasms with their normal counterparts, we determined that they frequently acquire methylation changes in regions already undergoing dynamic methylation during normal B cell differentiation.

Quality of Life and Survivorship Care in Patients Undergoing Hyperthermic Intraperitoneal Chemotherapy (HIPEC)

ClinicalTrials.gov

2017-05-25

Advanced Malignant Mesothelioma; Carcinoma of the Appendix; Ovarian Sarcoma; Ovarian Stromal Cancer; Pseudomyxoma Peritonei; Recurrent Colon Cancer; Recurrent Malignant Mesothelioma; Recurrent Ovarian Epithelial Cancer; Recurrent Ovarian Germ Cell Tumor; Stage III Colon Cancer; Stage III Ovarian Epithelial Cancer; Stage III Ovarian Germ Cell Tumor; Stage IV Colon Cancer; Stage IV Ovarian Epithelial Cancer; Stage IV Ovarian Germ Cell Tumor; Unspecified Childhood Solid Tumor, Protocol Specific

Dynamics of γ-tubulin cytoskeleton in HL-60 leukemia cells undergoing differentiation and apoptosis by all-trans retinoic acid.

PubMed

Shariftabrizi, Ahmad; Ahmadian, Shahin; Pazhang, Yaghub

2012-02-01

Microtubules are important components of the cell cytoskeleton, participating in protein localization and cell signaling. The capacity of leukemia cells to re-organize their microtubules is considered an integral part of differentiation in these cells in order to become mature granulocytes through treatment with all-trans retinoic acid (ATRA), an established drug for treating acute promyelocytic leukemia. In this study we examined γ-, α- and acetylated-α-tubulin content, their patterns of distribution in the cytoplasm, and the potency of centrosomes in re-organizing microtubules in different stages of ATRA-induced differentiation and apoptosis of the HL-60 cell line. The γ-tubulin content was dramatically increased following differentiation of HL-60 cells, and was then decreased after apoptosis. We also found that γ-tubulin had a diffuse, cytoplasmic pattern following apoptosis compared to the focal, centrosomal accumulation of γ-tubulin in differentiated cells. Differentiated cells had the ability to re-organize their microtubule network following nocodazole challenge testing, whereas undifferentiated cells did not show a similar ability. α-tubulin was more regularly organized in differentiated cells, and did not reveal any specific pattern of polymerization in apoptotic cells. Acetylated-α-tubulin generally followed the same organization patterns after differentiation, as that which occurred for α-tubulin. Our data is suggestive of a centrosomal and organized nucleation pattern of microtubules in HL-60 cells following differentiation, possibly mediated through up-regulation of γ-tubulin.

Genome-Level Determination of Plasmodium falciparum Blood-Stage Targets of Malarial Clinical Immunity in the Peruvian Amazon

PubMed Central

Torres, Katherine J.; Castrillon, Carlos E.; Moss, Eli L.; Saito, Mayuko; Tenorio, Roy; Molina, Douglas M.; Davies, Huw; Neafsey, Daniel E.; Felgner, Philip; Vinetz, Joseph M.; Gamboa, Dionicia

2015-01-01

Background. Persons with blood-stage Plasmodium falciparum parasitemia in the absence of symptoms are considered to be clinically immune. We hypothesized that asymptomatic subjects with P. falciparum parasitemia would differentially recognize a subset of P. falciparum proteins on a genomic scale. Methods and Findings. Compared with symptomatic subjects, sera from clinically immune, asymptomatically infected individuals differentially recognized 51 P. falciparum proteins, including the established vaccine candidate PfMSP1. Novel, hitherto unstudied hypothetical proteins and other proteins not previously recognized as potential vaccine candidates were also differentially recognized. Genes encoding the proteins differentially recognized by the Peruvian clinically immune individuals exhibited a significant enrichment of nonsynonymous nucleotide variation, an observation consistent with these genes undergoing immune selection. Conclusions. A limited set of P. falciparum protein antigens was associated with the development of naturally acquired clinical immunity in the low-transmission setting of the Peruvian Amazon. These results imply that, even in a low-transmission setting, an asexual blood-stage vaccine designed to reduce clinical malaria symptoms will likely need to contain large numbers of often-polymorphic proteins, a finding at odds with many current efforts in the design of vaccines against asexual blood-stage P. falciparum. PMID:25381370

Primary Metabolism during Biosynthesis of Secondary Wall Polymers of Protoxylem Vessel Elements1[OPEN

PubMed Central

Morisaki, Keiko; Sawada, Yuji; Sano, Ryosuke; Yamamoto, Atsushi; Kurata, Tetsuya; Suzuki, Shiro; Matsuda, Mami; Hasunuma, Tomohisa; Hirai, Masami Yokota

2016-01-01

Xylem vessels, the water-conducting cells in vascular plants, undergo characteristic secondary wall deposition and programmed cell death. These processes are regulated by the VASCULAR-RELATED NAC-DOMAIN (VND) transcription factors. Here, to identify changes in metabolism that occur during protoxylem vessel element differentiation, we subjected tobacco (Nicotiana tabacum) BY-2 suspension culture cells carrying an inducible VND7 system to liquid chromatography-mass spectrometry-based wide-target metabolome analysis and transcriptome analysis. Time-course data for 128 metabolites showed dynamic changes in metabolites related to amino acid biosynthesis. The concentration of glyceraldehyde 3-phosphate, an important intermediate of the glycolysis pathway, immediately decreased in the initial stages of cell differentiation. As cell differentiation progressed, specific amino acids accumulated, including the shikimate-related amino acids and the translocatable nitrogen-rich amino acid arginine. Transcriptome data indicated that cell differentiation involved the active up-regulation of genes encoding the enzymes catalyzing fructose 6-phosphate biosynthesis from glyceraldehyde 3-phosphate, phosphoenolpyruvate biosynthesis from oxaloacetate, and phenylalanine biosynthesis, which includes shikimate pathway enzymes. Concomitantly, active changes in the amount of fructose 6-phosphate and phosphoenolpyruvate were detected during cell differentiation. Taken together, our results show that protoxylem vessel element differentiation is associated with changes in primary metabolism, which could facilitate the production of polysaccharides and lignin monomers and, thus, promote the formation of the secondary cell wall. Also, these metabolic shifts correlate with the active transcriptional regulation of specific enzyme genes. Therefore, our observations indicate that primary metabolism is actively regulated during protoxylem vessel element differentiation to alter the cell’s metabolic activity for the biosynthesis of secondary wall polymers. PMID:27600813

Functional high-resolution time-course expression analysis of human embryonic stem cells undergoing cardiac induction.

PubMed

Piccini, Ilaria; Araúzo-Bravo, Marcos; Seebohm, Guiscard; Greber, Boris

2016-12-01

Cardiac induction of human embryonic stem cells (hESCs) is a process bearing increasing medical relevance, yet it is poorly understood from a developmental biology perspective. Anticipated technological progress in deriving stably expandable cardiac precursor cells or in advancing cardiac subtype specification protocols will likely require deeper insights into this fascinating system. Recent improvements in controlling hESC differentiation now enable a near-homogeneous induction of the cardiac lineage. This is based on an optimized initial stimulation of mesoderm-inducing signaling pathways such as Activin and/or FGF, BMP, and WNT, followed by WNT inhibition as a secondary requirement. Here, we describe a comprehensive data set based on varying hESC differentiation conditions in a systematic manner and recording high-resolution differentiation time-courses analyzed by genome-wide expression profiling (GEO accession number GSE67154). As a baseline, hESCs were differentiated into cardiomyocytes under optimal conditions. Moreover, in additional time-series, individual signaling factors were withdrawn from the initial stimulation cocktail to reveal their specific roles via comparison to the standard condition. Hence, this data set presents a rich resource for hypothesis generation in studying human cardiac induction, as we reveal numbers of known as well as uncharacterized genes prominently marking distinct intermediate stages in the process. These data will also be useful for identifying putative cardiac master regulators in the human system as well as for characterizing expandable cardiac stem cells.

EUS for the staging of gastric cancer: a meta-analysis.

PubMed

Mocellin, Simone; Marchet, Alberto; Nitti, Donato

2011-06-01

The role of EUS in the locoregional staging of gastric carcinoma is undefined. We aimed to comprehensively review and quantitatively summarize the available evidence on the staging performance of EUS. We systematically searched the MEDLINE, Cochrane, CANCERLIT, and EMBASE databases for relevant studies published until July 2010. Formal meta-analysis of diagnostic accuracy parameters was performed by using a bivariate random-effects model. Fifty-four studies enrolling 5601 patients with gastric cancer undergoing disease staging with EUS were eligible for the meta-analysis. EUS staging accuracy across eligible studies was measured by computing overall sensitivity, specificity, positive likelihood ratio (PLR), negative likelihood ratio (NLR), and diagnostic odds ratio (DOR). EUS can differentiate T1-2 from T3-4 gastric cancer with high accuracy, with overall sensitivity, specificity, PLR, NLR, and DOR of 0.86 (95% CI, 0.81-0.90), 0.91 (95% CI, 0.89-0.93), 9.8 (95% CI, 7.5-12.8), 0.15 (95% CI, 0.11-0.21), and 65 (95% CI, 41-105), respectively. In contrast, the diagnostic performance of EUS for lymph node status is less reliable, with overall sensitivity, specificity, PLR, NLR, and DOR of 0.69 (95% CI, 0.63-0.74), 0.84 (95% CI, 0.81-0.88), 4.4 (95% CI, 3.6-5.4), 0.37 (95% CI, 0.32-0.44), and 12 (95% CI, 9-16), respectively. Results regarding single T categories (including T1 substages) and Bayesian nomograms to calculate posttest probabilities for any target condition prevalence are also provided. Statistical heterogeneity was generally high; unfortunately, subgroup analysis did not identify a consistent source of the heterogeneity. Our results support the use of EUS for the locoregional staging of gastric cancer, which can affect the therapeutic management of these patients. However, clinicians must be aware of the performance limits of this staging tool. Copyright © 2011 American Society for Gastrointestinal Endoscopy. Published by Mosby, Inc. All rights reserved.

Brominated and organophosphate flame retardants target different neurodevelopmental stages, characterized with embryonic neural stem cells and neuronotypic PC12 cells.

PubMed

Slotkin, Theodore A; Skavicus, Samantha; Stapleton, Heather M; Seidler, Frederic J

2017-09-01

In addition to their activity as endocrine disruptors, brominated and organophosphate flame retardants are suspected to be developmental neurotoxicants, although identifying their specific mechanisms for that activity has been elusive. In the current study, we evaluated the effects of several flame retardants on neurodifferentiation using two in vitro models that assess distinct "decision nodes" in neural cell development: embryonic rat neural stem cells (NSCs), which evaluate the origination of neurons and glia from precursors, and rat neuronotypic PC12 cells, which characterize a later stage where cells committed to a neuronal phenotype undergo neurite outgrowth and neurotransmitter specification. In NSCs, both brominated and organophosphate flame retardants diverted the phenotype in favor of glia and away from formation of neurons, leading to an increased glia/neuron ratio, a common hallmark of the in vivo effects of neurotoxicants. For this early decision node, the brominated flame retardants were far more potent than the organophosphates. In PC12 cells, the brominated flame retardants were far less effective, whereas tris (1,3-dichloro-2-propyl) phosphate, an organophosphate, was more effective. Thus, the two classes of flame retardants differentially impact the two distinct vulnerable periods of neurodifferentiation. Furthermore, the effects on neurodifferentiation were separable from outright cytotoxicity, an important requirement in establishing a specific effect of these agents on neural cell development. These results reinforce the likelihood that flame retardants act as developmental neurotoxicants via direct effects on neural cell differentiation, over and above other activities that can impact nervous system development, such as endocrine disruption. Copyright © 2017 Elsevier B.V. All rights reserved.

Prolyl Isomerase Pin1 Regulates Neuronal Differentiation via β-Catenin

PubMed Central

Nakamura, Kazuhiro; Kosugi, Isao; Lee, Daniel Y.; Hafner, Angela; Sinclair, David A.

2012-01-01

The Wnt/β-catenin pathway promotes proliferation of neural progenitor cells (NPCs) at early stages and induces neuronal differentiation from NPCs at late stages, but the molecular mechanisms that control this stage-specific response are unclear. Pin1 is a prolyl isomerase that regulates cell signaling uniquely by controlling protein conformation after phosphorylation, but its role in neuronal differentiation is not known. Here we found that whereas Pin1 depletion suppresses neuronal differentiation, Pin1 overexpression enhances it, without any effects on gliogenesis from NPCs in vitro. Consequently, Pin1-null mice have significantly fewer upper layer neurons in the motor cortex and severely impaired motor activity during the neonatal stage. A proteomic approach identified β-catenin as a major substrate for Pin1 in NPCs, in which Pin1 stabilizes β-catenin. As a result, Pin1 knockout leads to reduced β-catenin during differentiation but not proliferation of NPCs in developing brains. Importantly, defective neuronal differentiation in Pin1 knockout NPCs is fully rescued in vitro by overexpression of β-catenin but not a β-catenin mutant that fails to act as a Pin1 substrate. These results show that Pin1 is a novel regulator of NPC differentiation by acting on β-catenin and provides a new postphosphorylation signaling mechanism to regulate developmental stage-specific functioning of β-catenin signaling in neuronal differentiation. PMID:22645310

Detection of a ventricular-specific myosin heavy chain in adult and developing chicken heart

PubMed Central

1986-01-01

In the present study, a monoclonal antibody (McAb), ALD19, generated against myosin of slow tonic muscle, was shown to react with the heavy chain of ventricular myosin in the adult chicken heart. With this antibody, it was possible to detect a ventricular-specific myosin during myocardial differentiation and to show that the epitope recognized by ALD19 was present from the earliest stages of ventricular differentiation and maintained throughout development only in the ventricle. A second McAb, specific for atrial myosin heavy chain (MHC) (Gonzalez-Sanchez, A., and D. Bader, 1984, Dev. Biol., 103:151-158), was used as a control to detect an atrial-specific myosin in the caudal portion of the developing heart at Hamburger-Hamilton stage 15. It was found that the appearance of ventricular MHC predated the expression of atrial MHC by approximately 1 d in ovo and that specific MHCs were always differentially distributed. While a common primordial MHC may be present in the early heart, this study showed the tissue-specific expression of a ventricular MHC during the initial stages of heart development and its differential accumulation throughout development. PMID:3514633

Genome-level determination of Plasmodium falciparum blood-stage targets of malarial clinical immunity in the Peruvian Amazon.

PubMed

Torres, Katherine J; Castrillon, Carlos E; Moss, Eli L; Saito, Mayuko; Tenorio, Roy; Molina, Douglas M; Davies, Huw; Neafsey, Daniel E; Felgner, Philip; Vinetz, Joseph M; Gamboa, Dionicia

2015-04-15

Persons with blood-stage Plasmodium falciparum parasitemia in the absence of symptoms are considered to be clinically immune. We hypothesized that asymptomatic subjects with P. falciparum parasitemia would differentially recognize a subset of P. falciparum proteins on a genomic scale. Compared with symptomatic subjects, sera from clinically immune, asymptomatically infected individuals differentially recognized 51 P. falciparum proteins, including the established vaccine candidate PfMSP1. Novel, hitherto unstudied hypothetical proteins and other proteins not previously recognized as potential vaccine candidates were also differentially recognized. Genes encoding the proteins differentially recognized by the Peruvian clinically immune individuals exhibited a significant enrichment of nonsynonymous nucleotide variation, an observation consistent with these genes undergoing immune selection. A limited set of P. falciparum protein antigens was associated with the development of naturally acquired clinical immunity in the low-transmission setting of the Peruvian Amazon. These results imply that, even in a low-transmission setting, an asexual blood-stage vaccine designed to reduce clinical malaria symptoms will likely need to contain large numbers of often-polymorphic proteins, a finding at odds with many current efforts in the design of vaccines against asexual blood-stage P. falciparum. © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

The Safety of Ovarian Preservation in Stage I Endometrial Endometrioid Adenocarcinoma Based on Propensity Score Matching.

PubMed

Hou, Ting; Sun, Yidi; Li, Junyi; Liu, Chenglin; Wang, Zhen; Li, Yixue; Lu, Yuan

2017-01-01

Most patients with early stage endometrial endometrioid adenocarcinoma (EEAC) are treated with hysterectomy and bilateral oophorectomy. But this surgical menopause leads to long-term sequelae for premenopausal women, especially for young women of childbearing age. This population-based study was to evaluate the safety of ovarian preservation in young women with stage I EEAC. Patients of age 50 or younger with stage I EEAC were explored from the Surveillance, Epidemiology and End Results program database during 2004 to 2013. Propensity score matching was used to randomize the data set and reduce the selection biases of doctors. Univariate analysis and multivariate cox proportional hazards model were utilized to estimate the safety of ovarian preservation. A total of 7183 patients were identified, and ovarian preservation was performed in 863 (12 %) patients. Compared with women treated with oophorectomy, patients with ovarian preservation significantly tend to be younger at diagnosis (P-value < 0.001) and more likely diagnosed as stage IA EEAC, to have better differentiated tumor tissues and smaller tumors, as well as less likely to undergo radiation and lymphadenectomy. 863 patients treated with oophorectomy were selected by propensity score matching. After propensity score matching, the differences of all characteristics between ovarian preservation and oophorectomy were not significant and potential confounders in the two groups decreased. In univariate analysis of matched population, ovarian preservation had no effect on overall (P-value=0.928) and cancer-specific (P-value=0.390) mortality. In propensityadjusted multivariate analysis, ovarian preservation was not significantly associated with overall (HR=0.69, 95%CI=0.41-1.68, P-value=0.611) and cancer-specific (HR=1.65, 95%CI=0.54-5.06, Pvalue= 0.379) survival. Ovarian preservation is safe for young women with stage I EEAC, which is not significantly associated with overall and cancer-specific mortality. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Changes in inflorescence protein during advanced stages of floret development in Buchloe dactyloides (Poaceae).

PubMed

Zhou, Y-J; Xue, J-G; Wang, X-G; Zhang, X-Q

2012-11-12

Buffalograss, Buchloe dactyloides, is a dioecious species native to the Great Plains of North America. The florets at the early stages of development possess both gynoecium and androecium organ primordia but later become unisexual. Very little is known about the proteomic changes that occur when the florets change from hermaphroditism to unisexuality. We compared the protein composition of florets at the hermaphroditic stage with that at the unisexual stage. The development stage of the floret was determined by stereomicroscopic observation. Two-dimensional gel electrophoresis was used to separate the proteins extracted from female and male inflorescences. Stage- specific protein maps, with an average of about 400 spots per map, were analyzed with the protein analysis software. Eighteen spots were found to be differentially expressed between the hermaphrodite and unisexual stages. Of these, 12 were present at both stages but with a different expression value. Four specific spots appeared at the hermaphrodite stage and disappeared at the unisexual stage. Two specific protein spots were associated with female and male floret differentiation. One appears to be associated with contabescence in the female floret and the final protein appears to lead to the abortion of gynoecium in the male floret. The MALDI TOF/TOF technique was used for peptide mass fingerprinting of the differentially expressed proteins and the MASCOT software was used to search the protein database. However, only two protein spots were identified from the database. These were aldolase1 and Os05g0574400 (similar to malate dehydrogenase). This type of proteomic study can help to identify novel protein products and determine the mechanisms involved in the floral sex differentiation process in buffalo grass.

Differential effects of insulin-like growth factor I and growth hormone on developmental stages of rat growth plate chondrocytes in vivo.

PubMed Central

Hunziker, E B; Wagner, J; Zapf, J

1994-01-01

Skeletal growth depends upon enchondral ossification in growth plate cartilage, within which chondrocytes undergo well defined stages of maturation. We infused IGF-I or growth hormone (GH), two key regulators of skeletal growth, into hypophysectomized rats and compared their effects on growth plate chondrocyte differentiation using qualitative and quantitative autoradiography, stereology, and incident light fluorescence microscopy. Stem cell cycle time was shortened from 50 to 15 and 8 d after treatment with IGF-I and GH, respectively. Proliferating cell cycle time decreased from 11 to 4.5 and 3 d, and duration of the hypertrophic phase decreased from 6 to 4 and 2.8 d. Average matrix volume per cell at each differentiation stage was similar for normal, hormone-treated, and untreated hypophysectomized groups. Mean cell volume and cell height were significantly reduced by hypophysectomy at the proliferative and hypertrophic stages, but were restored to physiological values by IGF-I and GH. In contrast, cell productivity, i.e., increases in cell volume, height, and matrix production per unit of time, did not reach normal values with either IGF-I or GH, and this parameter was inversely proportional to cell cycle time or phase duration. IGF-I and GH are thus capable of stimulating growth plate chondrocytes at all stages of differentiation, albeit to variable degrees with respect to individual cell activities. Although it is generally accepted that GH acts at both the stem and proliferating phases of chondrocyte differentiation, our data represent the first evidence in vivo that IGF-I is also capable of stimulating stem cells. Images PMID:8132746

Large-scale identification of differentially expressed genes during pupa development reveals solute carrier gene is essential for pupal pigmentation in Chilo suppressalis.

PubMed

Sun, Yang; Huang, Shuijin; Wang, Shuping; Guo, Dianhao; Ge, Chang; Xiao, Huamei; Jie, Wencai; Yang, Qiupu; Teng, Xiaolu; Li, Fei

2017-04-01

Insects undergo metamorphosis, involving an abrupt change in body structure through cell growth and differentiation. Rice stem stripped borer (SSB), Chilo suppressalis, is one of the most destructive rice pests. However, little is known about the regulation mechanism of metamorphosis development in this notorious insect pest. Here, we studied the expression of 22,197 SSB genes at seven time points during pupa development with a customized microarray, identifying 622 differentially expressed genes (DEG) during pupa development. Gene ontology (GO) analysis of these DEGs indicated that the genes related to substance metabolism were highly expressed in the early pupa, which participate in the physiological processes of larval tissue disintegration at these stages. In comparison, highly expressed genes in the late pupal stages were mainly associated with substance biosynthesis, consistent with adult organ formation at these stages. There were 27 solute carrier (SLC) genes that were highly expressed during pupa development. We knocked down SLC22A3 at the prepupal stage, demonstrating that silencing SLC22A3 induced a deficiency in pupa stiffness and pigmentation. The RNAi-treated individuals had white and soft pupa, suggesting that this gene has an essential role in pupal development. Copyright © 2016 Elsevier Ltd. All rights reserved.

Utility of Inflammatory Marker- and Nutritional Status-based Prognostic Factors for Predicting the Prognosis of Stage IV Gastric Cancer Patients Undergoing Non-curative Surgery.

PubMed

Mimatsu, Kenji; Fukino, Nobutada; Ogasawara, Yasuo; Saino, Yoko; Oida, Takatsugu

2017-08-01

The present study aimed to compare the utility of various inflammatory marker- and nutritional status-based prognostic factors, including many previous established prognostic factors, for predicting the prognosis of stage IV gastric cancer patients undergoing non-curative surgery. A total of 33 patients with stage IV gastric cancer who had undergone palliative gastrectomy and gastrojejunostomy were included in the study. Univariate and multivariate analyses were performed to evaluate the relationships between the mGPS, PNI, NLR, PLR, the CONUT, various clinicopathological factors and cancer-specific survival (CS). Among patients who received non-curative surgery, univariate analysis of CS identified the following significant risk factors: chemotherapy, mGPS and NLR, and multivariate analysis revealed that the mGPS was independently associated with CS. The mGPS was a more useful prognostic factor than the PNI, NLR, PLR and CONUT in patients undergoing non-curative surgery for stage IV gastric cancer. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

Cytosolic labile zinc: a marker for apoptosis in the developing rat brain.

PubMed

Lee, Joo-Yong; Hwang, Jung Jin; Park, Mi-Ha; Koh, Jae-Young

2006-01-01

Cytosolic zinc accumulation was thought to occur specifically in neuronal death (necrosis) following acute injury. However, a recent study demonstrated that zinc accumulation also occurs in adult rat neurons undergoing apoptosis following target ablation, and in vitro experiments have shown that zinc accumulation may play a causal role in various forms of apoptosis. Here, we examined whether intraneuronal zinc accumulation occurs in central neurons undergoing apoptosis during development. Embryonic and newborn Sprague-Dawley rat brains were double-stained for terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labelling (TUNEL) detection of apoptosis and immunohistochemical detection of stage-specific neuronal markers, such as nestin, proliferating cell nuclear antigen (PCNA), TuJ1 and neuronal nuclear specific protein (NeuN). The results revealed that apoptotic cell death occurred in neurons of diverse stages (neural stem cells, and dividing, young and adult neurons) throughout the brain during the embryonic and early postnatal periods. Further staining of brain sections with acid fuchsin or zinc-specific fluorescent dyes showed that all of the apoptotic neurons were acidophilic and contained labile zinc in their cell bodies. Cytosolic zinc accumulation was also observed in cultured cortical neurons undergoing staurosporine- or sodium nitroprusside (SNP)-induced apoptosis. In contrast, zinc chelation with CaEDTA or N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine (TPEN) reduced SNP-induced apoptosis but not staurosporine-induced apoptosis, indicating that cytosolic zinc accumulation does not play a causal role in all forms of apoptosis. Finally, the specific cytosolic zinc accumulation may have a practical application as a relatively simple marker for neurons undergoing developmental apoptosis.

Pair-wise comparison analysis of differential expression of mRNAs in early and advanced stage primary colorectal adenocarcinomas

PubMed Central

Lau, Tze Pheng; Roslani, April Camilla; Lian, Lay Hoong; Chai, Hwa Chia; Lee, Ping Chin; Hilmi, Ida; Goh, Khean Lee; Chua, Kek Heng

2014-01-01

Objectives To characterise the mRNA expression patterns of early and advanced stage colorectal adenocarcinomas of Malaysian patients. Design Comparative expression analysis. Setting and participants We performed a combination of annealing control primer (ACP)-based PCR and reverse transcription-quantitative real-time PCR for the identification of differentially expressed genes (DEGs) associated with early and advanced stage primary colorectal tumours. We recruited four paired samples from patients with colorectal cancer (CRC) of Dukes’ A and B for the preliminary differential expression study, and a total of 27 paired samples, ranging from CRC stages I to IV, for subsequent confirmatory test. The tumouric samples were obtained from the patients with CRC undergoing curative surgical resection without preoperative chemoradiotherapy. The recruited patients with CRC were newly diagnosed with CRC, and were not associated with any hereditary syndromes, previously diagnosed cancer or positive family history of CRC. The paired non-cancerous tissue specimens were excised from macroscopically normal colonic mucosa distally located from the colorectal tumours. Primary and secondary outcome measures The differential mRNA expression patterns of early and advanced stage colorectal adenocarcinomas compared with macroscopically normal colonic mucosa were characterised by ACP-based PCR and reverse transcription-quantitative real-time PCR. Results The RPL35, RPS23 and TIMP1 genes were found to be overexpressed in both early and advanced stage colorectal adenocarcinomas (p<0.05). However, the ARPC2 gene was significantly underexpressed in early colorectal adenocarcinomas, while the advanced stage primary colorectal tumours exhibited an additional overexpression of the C6orf173 gene (p<0.05). Conclusions We characterised two distinctive gene expression patterns to aid in the stratification of primary colorectal neoplasms among Malaysian patients with CRC. Further work can be done to assess and compare the mRNA expression levels of these identified DEGs between each CRC stage group, stages I–IV. PMID:25107436

The development and plasticity of alveolar type 1 cells

PubMed Central

Yang, Jun; Hernandez, Belinda J.; Martinez Alanis, Denise; Narvaez del Pilar, Odemaris; Vila-Ellis, Lisandra; Akiyama, Haruhiko; Evans, Scott E.; Ostrin, Edwin J.; Chen, Jichao

2016-01-01

Alveolar type 1 (AT1) cells cover >95% of the gas exchange surface and are extremely thin to facilitate passive gas diffusion. The development of these highly specialized cells and its coordination with the formation of the honeycomb-like alveolar structure are poorly understood. Using new marker-based stereology and single-cell imaging methods, we show that AT1 cells in the mouse lung form expansive thin cellular extensions via a non-proliferative two-step process while retaining cellular plasticity. In the flattening step, AT1 cells undergo molecular specification and remodel cell junctions while remaining connected to their epithelial neighbors. In the folding step, AT1 cells increase in size by more than 10-fold and undergo cellular morphogenesis that matches capillary and secondary septa formation, resulting in a single AT1 cell spanning multiple alveoli. Furthermore, AT1 cells are an unexpected source of VEGFA and their normal development is required for alveolar angiogenesis. Notably, a majority of AT1 cells proliferate upon ectopic SOX2 expression and undergo stage-dependent cell fate reprogramming. These results provide evidence that AT1 cells have both structural and signaling roles in alveolar maturation and can exit their terminally differentiated non-proliferative state. Our findings suggest that AT1 cells might be a new target in the pathogenesis and treatment of lung diseases associated with premature birth. PMID:26586225

Cell population structure prior to bifurcation predicts efficiency of directed differentiation in human induced pluripotent cells

PubMed Central

Bargaje, Rhishikesh; Trachana, Kalliopi; Shelton, Martin N.; McGinnis, Christopher S.; Zhou, Joseph X.; Chadick, Cora; Cook, Savannah; Cavanaugh, Christopher; Huang, Sui; Hood, Leroy

2017-01-01

Steering the differentiation of induced pluripotent stem cells (iPSCs) toward specific cell types is crucial for patient-specific disease modeling and drug testing. This effort requires the capacity to predict and control when and how multipotent progenitor cells commit to the desired cell fate. Cell fate commitment represents a critical state transition or “tipping point” at which complex systems undergo a sudden qualitative shift. To characterize such transitions during iPSC to cardiomyocyte differentiation, we analyzed the gene expression patterns of 96 developmental genes at single-cell resolution. We identified a bifurcation event early in the trajectory when a primitive streak-like cell population segregated into the mesodermal and endodermal lineages. Before this branching point, we could detect the signature of an imminent critical transition: increase in cell heterogeneity and coordination of gene expression. Correlation analysis of gene expression profiles at the tipping point indicates transcription factors that drive the state transition toward each alternative cell fate and their relationships with specific phenotypic readouts. The latter helps us to facilitate small molecule screening for differentiation efficiency. To this end, we set up an analysis of cell population structure at the tipping point after systematic variation of the protocol to bias the differentiation toward mesodermal or endodermal cell lineage. We were able to predict the proportion of cardiomyocytes many days before cells manifest the differentiated phenotype. The analysis of cell populations undergoing a critical state transition thus affords a tool to forecast cell fate outcomes and can be used to optimize differentiation protocols to obtain desired cell populations. PMID:28167799

Cell population structure prior to bifurcation predicts efficiency of directed differentiation in human induced pluripotent cells.

PubMed

Bargaje, Rhishikesh; Trachana, Kalliopi; Shelton, Martin N; McGinnis, Christopher S; Zhou, Joseph X; Chadick, Cora; Cook, Savannah; Cavanaugh, Christopher; Huang, Sui; Hood, Leroy

2017-02-28

Steering the differentiation of induced pluripotent stem cells (iPSCs) toward specific cell types is crucial for patient-specific disease modeling and drug testing. This effort requires the capacity to predict and control when and how multipotent progenitor cells commit to the desired cell fate. Cell fate commitment represents a critical state transition or "tipping point" at which complex systems undergo a sudden qualitative shift. To characterize such transitions during iPSC to cardiomyocyte differentiation, we analyzed the gene expression patterns of 96 developmental genes at single-cell resolution. We identified a bifurcation event early in the trajectory when a primitive streak-like cell population segregated into the mesodermal and endodermal lineages. Before this branching point, we could detect the signature of an imminent critical transition: increase in cell heterogeneity and coordination of gene expression. Correlation analysis of gene expression profiles at the tipping point indicates transcription factors that drive the state transition toward each alternative cell fate and their relationships with specific phenotypic readouts. The latter helps us to facilitate small molecule screening for differentiation efficiency. To this end, we set up an analysis of cell population structure at the tipping point after systematic variation of the protocol to bias the differentiation toward mesodermal or endodermal cell lineage. We were able to predict the proportion of cardiomyocytes many days before cells manifest the differentiated phenotype. The analysis of cell populations undergoing a critical state transition thus affords a tool to forecast cell fate outcomes and can be used to optimize differentiation protocols to obtain desired cell populations.

Uncovering the pathways underlying whole body regeneration in a chordate model, Botrylloides leachi using de novo transcriptome analysis.

PubMed

Zondag, Lisa E; Rutherford, Kim; Gemmell, Neil J; Wilson, Megan J

2016-02-16

Regenerative capacity differs greatly between animals. In vertebrates regenerative abilities are highly limited and tissue or organ specific. However the closest related chordate to the vertebrate clade, Botrylloides leachi, can undergo whole body regeneration (WBR). Therefore, research on WBR in B. leachi has focused on pathways known to be important for regeneration in vertebrates. To obtain a comprehensive vision of this unique process we have carried out the first de novo transcriptome sequencing for multiple stages of WBR occurring in B. leachi. The identified changes in gene expression during B. leachi WBR offer novel insights into this remarkable ability to regenerate. The transcriptome of B. leachi tissue undergoing WBR were analysed using differential gene expression, gene ontology and pathway analyses. We observed up-regulation in the expression of genes involved in wound healing and known developmental pathways including WNT, TGF-β and Notch, during the earliest stages of WBR. Later in WBR, the expression patterns in several pathways required for protein synthesis, biogenesis and the organisation of cellular components were up-regulated. While the genes expressed early on are characteristic of a necessary wound healing response to an otherwise lethal injury, the subsequent vast increase in protein synthesis conceivably sustains the reestablishment of the tissue complexity and body axis polarity within the regenerating zooid. We have, for the first time, provided a global overview of the genes and their corresponding pathways that are modulated during WBR in B. leachi.

Diagnostic value of sleep stage dissociation as visualized on a 2-dimensional sleep state space in human narcolepsy.

PubMed

Olsen, Anders Vinther; Stephansen, Jens; Leary, Eileen; Peppard, Paul E; Sheungshul, Hong; Jennum, Poul Jørgen; Sorensen, Helge; Mignot, Emmanuel

2017-04-15

Type 1 narcolepsy (NT1) is characterized by symptoms believed to represent Rapid Eye Movement (REM) sleep stage dissociations, occurrences where features of wake and REM sleep are intermingled, resulting in a mixed state. We hypothesized that sleep stage dissociations can be objectively detected through the analysis of nocturnal Polysomnography (PSG) data, and that those affecting REM sleep can be used as a diagnostic feature for narcolepsy. A Linear Discriminant Analysis (LDA) model using 38 features extracted from EOG, EMG and EEG was used in control subjects to select features differentiating wake, stage N1, N2, N3 and REM sleep. Sleep stage differentiation was next represented in a 2D projection. Features characteristic of sleep stage differences were estimated from the residual sleep stage probability in the 2D space. Using this model we evaluated PSG data from NT1 and non-narcoleptic subjects. An LDA classifier was used to determine the best separation plane. This method replicates the specificity/sensitivity from the training set to the validation set better than many other methods. Eight prominent features could differentiate narcolepsy and controls in the validation dataset. Using a composite measure and a specificity cut off 95% in the training dataset, sensitivity was 43%. Specificity/sensitivity was 94%/38% in the validation set. Using hypersomnia subjects, specificity/sensitivity was 84%/15%. Analyzing treated narcoleptics the specificity/sensitivity was 94%/10%. Sleep stage dissociation can be used for the diagnosis of narcolepsy. However the use of some medications and presence of undiagnosed hypersomnolence patients impacts the result. Copyright © 2017 Elsevier B.V. All rights reserved.

METACASPASE9 modulates autophagy to confine cell death to the target cells during Arabidopsis vascular xylem differentiation

PubMed Central

Escamez, Sacha; André, Domenique; Zhang, Bo; Bollhöner, Benjamin; Pesquet, Edouard; Tuominen, Hannele

2016-01-01

ABSTRACT We uncovered that the level of autophagy in plant cells undergoing programmed cell death determines the fate of the surrounding cells. Our approach consisted of using Arabidopsis thaliana cell cultures capable of differentiating into two different cell types: vascular tracheary elements (TEs) that undergo programmed cell death (PCD) and protoplast autolysis, and parenchymatic non-TEs that remain alive. The TE cell type displayed higher levels of autophagy when expression of the TE-specific METACASPASE9 (MC9) was reduced using RNAi (MC9-RNAi). Misregulation of autophagy in the MC9-RNAi TEs coincided with ectopic death of the non-TEs, implying the existence of an autophagy-dependent intercellular signalling from within the TEs towards the non-TEs. Viability of the non-TEs was restored when AUTOPHAGY2 (ATG2) was downregulated specifically in MC9-RNAi TEs, demonstrating the importance of autophagy in the spatial confinement of cell death. Our results suggest that other eukaryotic cells undergoing PCD might also need to tightly regulate their level of autophagy to avoid detrimental consequences for the surrounding cells. PMID:26740571

Regulated expression and role of c-Myb in the cardiovascular-directed differentiation of mouse embryonic stem cells.

PubMed

Ishida, Masayoshi; El-Mounayri, Omar; Kattman, Steven; Zandstra, Peter; Sakamoto, Hiroshi; Ogawa, Minetaro; Keller, Gordon; Husain, Mansoor

2012-01-20

c-myb null (knockout) embryonic stem cells (ESC) can differentiate into cardiomyocytes but not contractile smooth muscle cells (SMC) in embryoid bodies (EB). To define the role of c-Myb in SMC differentiation from ESC. In wild-type (WT) EB, high c-Myb levels on days 0-2 of differentiation undergo ubiquitin-mediated proteosomal degradation on days 2.5-3, resurging on days 4-6, without changing c-myb mRNA levels. Activin-A and bone morphogenetic protein 4-induced cardiovascular progenitors were isolated by FACS for expression of vascular endothelial growth factor receptor (VEGFR)2 and platelet-derived growth factor receptor (PDGFR)α. By day 3.75, hematopoesis-capable VEGFR2+ cells were fewer, whereas cardiomyocyte-directed VEGFR2+/PDGFRα+ cells did not differ in abundance in knockout versus WT EB. Importantly, highest and lowest levels of c-Myb were observed in VEGFR2+ and VEGFR2+/PDGFRα+ cells, respectively. Proteosome inhibitor MG132 and lentiviruses enabling inducible expression or knockdown of c-myb were used to regulate c-Myb in WT and knockout EB. These experiments showed that c-Myb promotes expression of VEGFR2 over PDGFRα, with chromatin immunopreciptation and promoter-reporter assays defining specific c-Myb-responsive binding sites in the VEGFR2 promoter. Next, FACS-sorted VEGFR2+ cells expressed highest and lowest levels of SMC- and fibroblast-specific markers, respectively, at days 7-14 after retinoic acid (RA) as compared with VEGFR2+/PDGFRα+ cells. By contrast, VEGFR2+/PDGFRα+ cells cultured without RA beat spontaneously, like cardiomyocytes between days 7 and 14, and expressed cardiac troponin. Notably, RA was required to more fully differentiate SMC from VEGFR2+ cells and completely blocked differentiation of cardiomyocytes from VEGFR2+/PDGFRα+ cells. c-Myb is tightly regulated by proteosomal degradation during cardiovascular-directed differentiation of ESC, expanding early-stage VEGFR2+ progenitors capable of RA-responsive SMC formation.

Breast Cancer after Augmentation: Oncologic and Reconstructive Considerations among Women Undergoing Mastectomy.

PubMed

Cho, Eugenia H; Shammas, Ronnie L; Phillips, Brett T; Greenup, Rachel A; Hwang, E Shelley; Hollenbeck, Scott T

2017-06-01

Breast augmentation with subglandular versus subpectoral implants may differentially impact the early detection of breast cancer and treatment recommendations. The authors assessed the impact of prior augmentation on the diagnosis and management of breast cancer in women undergoing mastectomy. Breast cancer diagnosis and management were retrospectively analyzed in all women with prior augmentation undergoing therapeutic mastectomy at the authors' institution from 1993 to 2014. Comparison was made to all women with no prior augmentation undergoing mastectomy in 2010. Subanalyses were performed according to prior implant placement. A total of 260 women with (n = 89) and without (n = 171) prior augmentation underwent mastectomy for 95 and 179 breast cancers, respectively. Prior implant placement was subglandular (n = 27) or subpectoral (n = 63) (For five breasts, the placement was unknown). Breast cancer stage at diagnosis (p = 0.19) and detection method (p = 0.48) did not differ for women with and without prior augmentation. Compared to subpectoral augmentation, subglandular augmentation was associated with the diagnosis of invasive breast cancer rather than ductal carcinoma in situ (p = 0.01) and detection by self-palpation rather than screening mammography (p = 0.03). Immediate two-stage implant reconstruction was the preferred reconstructive method in women with augmentation (p < 0.01). Breast cancer stage at diagnosis was similar for women with and without prior augmentation. Among women with augmentation, however, subglandular implants were associated with more advanced breast tumors commonly detected on palpation rather than mammography. Increased vigilance in breast cancer screening is recommended among women with subglandular augmentation. Therapeutic, III.

Differentiation of embryonic stem cells into hepatocytes that coexpress coagulation factors VIII and IX.

PubMed

Cao, Jun; Shang, Chang-zhen; Lü, Li-hong; Qiu, De-chuan; Ren, Meng; Chen, Ya-jin; Min, Jun

2010-11-01

To establish an efficient culture system to support embryonic stem (ES) cell differentiation into hepatocytes that coexpress F-VIII and F-IX. Mouse E14 ES cells were cultured in differentiation medium containing sodium butyrate (SB), basic fibroblast growth factor (bFGF), and/or bone morphogenetic protein 4 (BMP4) to induce the differentiation of endoderm cells and hepatic progenitor cells. Hepatocyte growth factor, oncostatin M, and dexamethasone were then used to induce the maturation of ES cell-derived hepatocytes. The mRNA expression levels of endoderm-specific genes and hepatocyte-specific genes, including the levels of F-VIII and F-IX, were detected by RT-PCR and real-time PCR during various stages of differentiation. Protein expression was examined by immunofluorescence and Western blot. At the final stage of differentiation, flow cytometry was performed to determine the percentage of cells coexpressing F-VIII and F-IX, and ELISA was used to detect the levels of F-VIII and F-IX protein secreted into the culture medium. The expression of endoderm-specific and hepatocyte-specific markers was upregulated to highest level in response to the combination of SB, bFGF, and BMP4. Treatment with the three inducers during hepatic progenitor differentiation significantly enhanced the mRNA and protein levels of F-VIII and F-IX in ES cell-derived hepatocytes. More importantly, F-VIII and F-IX were coexpressed with high efficiency at the final stage of differentiation, and they were also secreted into the culture medium. We have established a novel in vitro differentiation protocol for ES-derived hepatocytes that coexpress F-VIII and F-IX that may provide a foundation for stem cell replacement therapy for hemophilia.

Parental contributions to early embryo development: influences of urinary phthalate and phthalate alternatives among couples undergoing IVF treatment.

PubMed

Wu, Haotian; Ashcraft, Lisa; Whitcomb, Brian W; Rahil, Tayyab; Tougias, Ellen; Sites, Cynthia K; Pilsner, J Richard

2017-01-01

Are preconception urinary concentrations of phthalates and phthalate alternatives associated with diminished early stage embryo quality in couples undergoing IVF? Male, but not female, urinary concentrations of select metabolites of phthalates and phthalate alternatives are associated with diminished blastocyst quality. Although phthalates are endocrine disrupting compounds associated with adverse reproductive health, they are in widespread use across the world. Male and female preconception exposures to select phthalates have been previously associated with adverse reproductive outcomes in both the general population and in those undergoing IVF. This prospective cohort included 50 subfertile couples undergoing IVF in western Massachusetts. This study includes the first 50 couples recruited from the Baystate Medical Center's Fertility Center in Springfield, MA, as part of the Sperm Environmental Epigenetics and Development Study (SEEDS). Relevant data from both partners, including embryo quality at the cleavage (Day 3) and blastocyst (Day 5) stages, were collected by clinic personnel during the normal course of an IVF cycle. A spot urine sample was collected from both male and female partners on the same day as semen sample procurement and oocyte retrieval. Concentrations of 17 urinary metabolite were quantified by liquid chromatography mass spectrometry and normalized via specific gravity. Generalized estimating equations were used to estimate odds ratios (OR) and 95% CI, with urinary phthalates and phthalate alternatives fitted as continuous variables and embryo quality as a binary variable. The 50 couples contributed 761 oocytes, of which 423 progressed to the cleavage stage, 261 were high-quality cleavage stage embryos, 137 were transferrable quality blastocysts and 47 were high-quality blastocysts. At the cleavage stage, male urinary monoethyl phthalate concentrations were positively associated with high-quality cleavage stage embryos (OR = 1.20, 95% CI 1.01-1.43, P = 0.04); no other significant associations were observed at this stage. At the blastocyst stage, male urinary concentrations of monobenzyl phthalate (OR = 0.55, 95% CI 0.36-0.84, P = 0.01), mono-3-hydroxybutyl phthalate (OR = 0.37, 95% CI 0.18-0.76, P = 0.01), mono-n-butyl phthalate (OR = 0.55, 95% CI 0.42-0.73, P < 0.01) and monomethyl phthalate (OR = 0.39, 95% CI 0.26-0.60, P < 0.01) were inversely associated with high-quality blastocysts. A borderline statistically significant relationship was observed for male concentrations of mono(2-ethylhexyl) phthalate (OR = 0.52, 95% CI 0.27-1.00, P = 0.05) and cyclohexane-1,2-dicarboxylic acid-monocarboxy isooctyl ester (OR = 0.21, 95% CI 0.04-1.03, P = 0.05) at the blastocyst stage. Similar inverse associations were observed between male urinary phthalate metabolite concentrations and likelihood of transferrable quality blastocysts. For female partners, select metabolites were positively associated with odds of high or transferrable blastocyst quality, but the observed associations were not consistent across blastocyst quality measures or between sex-specific and couples-level models. All models were adjusted for age of both partners, urinary metabolite concentrations of female partners and male infertility status, while models of blastocysts were additionally adjusted for embryo quality at cleavage stage. Our modest sample included only 50 couples contributing one cycle each. In addition, non-differential misclassification of exposure remains a concern given the single-spot urine collection and the short half-life of phthalates. Our results suggest an inverse association between male preconception concentrations of select phthalate metabolites and blastocyst quality, likely occurring after genomic activation. If corroborated with other studies, such findings will have public health and clinical significance for both the general population and those undergoing IVF. This work was generously supported by grant K22-ES023085 from the National Institute of Environmental Health Sciences. The authors declare no competing interests. N/A. © The Author 2016. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

The C. elegans tailless/Tlx homolog nhr-67 regulates a stage-specific program of linker cell migration in male gonadogenesis.

PubMed

Kato, Mihoko; Sternberg, Paul W

2009-12-01

Cell migration is a common event during organogenesis, yet little is known about how migration is temporally coordinated with organ development. We are investigating stage-specific programs of cell migration using the linker cell (LC), a migratory cell crucial for male gonadogenesis of C. elegans. During the L3 and L4 larval stages of wild-type males, the LC undergoes changes in its position along the migratory route, in transcriptional regulation of the unc-5 netrin receptor and zmp-1 zinc matrix metalloprotease, and in cell morphology. We have identified the tailless homolog nhr-67 as a cell-autonomous, stage-specific regulator of timing in LC migration programs. In nhr-67-deficient animals, each of the L3 and L4 stage changes is either severely delayed or never occurs, yet LC development before the early L3 stage or after the mid-L4 stage occurs with normal timing. We propose that there is a basal migration program utilized throughout LC migration that is modified by stage-specific regulators such as nhr-67.

Novel histopathologic feature identified through image analysis augments stage II colorectal cancer clinical reporting

PubMed Central

Caie, Peter D.; Zhou, Ying; Turnbull, Arran K.; Oniscu, Anca; Harrison, David J.

2016-01-01

A number of candidate histopathologic factors show promise in identifying stage II colorectal cancer (CRC) patients at a high risk of disease-specific death, however they can suffer from low reproducibility and none have replaced classical pathologic staging. We developed an image analysis algorithm which standardized the quantification of specific histopathologic features and exported a multi-parametric feature-set captured without bias. The image analysis algorithm was executed across a training set (n = 50) and the resultant big data was distilled through decision tree modelling to identify the most informative parameters to sub-categorize stage II CRC patients. The most significant, and novel, parameter identified was the ‘sum area of poorly differentiated clusters’ (AreaPDC). This feature was validated across a second cohort of stage II CRC patients (n = 134) (HR = 4; 95% CI, 1.5– 11). Finally, the AreaPDC was integrated with the significant features within the clinical pathology report, pT stage and differentiation, into a novel prognostic index (HR = 7.5; 95% CI, 3–18.5) which improved upon current clinical staging (HR = 4.26; 95% CI, 1.7– 10.3). The identification of poorly differentiated clusters as being highly significant in disease progression presents evidence to suggest that these features could be the source of novel targets to decrease the risk of disease specific death. PMID:27322148

Identification of Cells at Early and Late Stages of Polarization During Odontoblast Differentiation Using pOBCol3.6GFP and pOBCol2.3GFP Transgenic Mice

PubMed Central

Balic, Anamaria; Aguila, H. Leonardo; Mina, Mina

2010-01-01

Transgenic mouse lines in which GFP expression is under the control of tissue-and stage specific promoters have provided powerful experimental tools for identification and isolation of cells at specific stage of differentiation along a lineage. In the present study we used primary cell cultures derived from the dental pulp from pOBCol3.6GFP and pOBCol2.3GFP transgenic mice as a model to develop markers for early stages of odontoblast differentiation from progenitor cells. We analyzed the temporal and spatial expression of 2.3-GFP and 3.6-GFP during in vitro mineralization. Using FACS to separate cells based on GFP expression, we obtained relatively homogenous sub-populations of cells and analyzed their dentinogenic potentials and their progression into odontoblasts. Our observations showed that these transgenes were activated before the onset of matrix deposition and in cells at different stages of polarization. The 3.6-GFP transgene was activated in cells in early stages of polarization whereas the 2.3-GFP transgene was activated at a later stage of polarization just before or at the time of formation of secretory odontoblast. PMID:20728593

Physiological and biological patterns of a highland and a coastal population of the European cherry fruit fly during diapause.

PubMed

Papanastasiou, Stella A; Nestel, David; Diamantidis, Alexandros D; Nakas, Christos T; Papadopoulos, Nikos T

2011-01-01

Adult emergence of univoltine temperate insect species and its synchronization with specific host phenological stages is mainly regulated by obligatory pupal diapause. Although a few studies have investigated the factors affecting diapause intensity, little attention has been paid to the physiological alterations and metabolic regulation that take place during diapause. Here, we describe differences in diapause between a highland and a coastal Greek population of the European cherry fruit fly Rhagoletis cerasi, a major pest of sweet and sour cherries in many European countries. Pupae of both populations were exposed to the environmental conditions prevailing in the two areas and diapause termination was observed under laboratory conditions. The regulation of energetic metabolites during the long pupae stage was examined under both field and laboratory conditions. Differences in diapause intensity revealed that the two populations have adapted to the local geographical and climatic conditions and have different requirements for low temperatures to terminate diapause. The coastal population undergoes a shorter diapause and adults emerge more rapidly, especially in the highland area. The highland population failed to terminate diapause (<40% adult emergence) in the coastal area. Both populations draw on their major energetic reserves (lipids and protein) similarly during diapause. Nevertheless, regulation of carbohydrate and glycogen reserves seems to vary between the populations: major peaks of these stored nutrients occur on different dates in the two populations, suggesting a differential regulation. Differences in diapause intensity imply a genetic differentiation between the two populations. The importance of our findings in understanding the physiological patterns during obligatory diapause of a univoltine insect species, as well as the practical implications for the development of specific phenological models for the European cherry fruit fly are discussed. 2010 Elsevier Ltd. All rights reserved.

Genetic Profile of Adenoid Cystic Carcinomas (ACC) with High-Grade Transformation versus Solid Type

PubMed Central

Costa, Ana Flávia; Altemani, Albina; Vékony, Hedy; Bloemena, Elisabeth; Fresno, Florentino; Suárez, Carlos; Llorente, José Luis; Hermsen, Mario

2010-01-01

Background: ACC can occasionally undergo dedifferentiation also referred to as high-grade transformation (ACC-HGT). However, ACC-HGT can also undergo transformation to adenocarcinomas which are not poorly differentiated. ACC-HGT is generally considered to be an aggressive variant of ACC, even more than solid ACC. This study was aimed to describe the genetic changes of ACC-HGT in relation to clinico-pathological features and to compare results to solid ACC. Methods: Genome-wide DNA copy number changes were analyzed by microarray CGH in ACC-HGT, 4 with transformation into moderately differentiated adenocarcinoma (MDA) and two into poorly differentiated carcinoma (PDC), 5 solid ACC. In addition, Ki-67 index and p53 immunopositivity was assessed. Results: ACC-HGT carried fewer copy number changes compared to solid ACC. Two ACC-HGT cases harboured a breakpoint at 6q23, near the cMYB oncogene. The complexity of the genomic profile concurred with the clinical course of the patient. Among the ACC-HGT, p53 positivity significantly increased from the conventional to the transformed (both MDA and PDC) component. Conclusion: ACC-HGT may not necessarily reflect a more advanced stage of tumor progression, but rather a transformation to another histological form in which the poorly differentiated forms (PDC) presents a genetic complexity similar to the solid ACC. PMID:20978318

Genetic profile of adenoid cystic carcinomas (ACC) with high-grade transformation versus solid type.

PubMed

Costa, Ana Flávia; Altemani, Albina; Vékony, Hedy; Bloemena, Elisabeth; Fresno, Florentino; Suárez, Carlos; Llorente, José Luis; Hermsen, Mario

2010-01-01

ACC can occasionally undergo dedifferentiation also referred to as high-grade transformation (ACC-HGT). However, ACC-HGT can also undergo transformation to adenocarcinomas which are not poorly differentiated. ACC-HGT is generally considered to be an aggressive variant of ACC, even more than solid ACC. This study was aimed to describe the genetic changes of ACC-HGT in relation to clinico-pathological features and to compare results to solid ACC. genome-wide DNA copy number changes were analyzed by microarray CGH in ACC-HGT, 4 with transformation into moderately differentiated adenocarcinoma (MDA) and two into poorly differentiated carcinoma (PDC), 5 solid ACC. In addition, Ki-67 index and p53 immunopositivity was assessed. ACC-HGT carried fewer copy number changes compared to solid ACC. Two ACC-HGT cases harboured a breakpoint at 6q23, near the cMYB oncogene. The complexity of the genomic profile concurred with the clinical course of the patient. Among the ACC-HGT, p53 positivity significantly increased from the conventional to the transformed (both MDA and PDC) component. ACC-HGT may not necessarily reflect a more advanced stage of tumor progression, but rather a transformation to another histological form in which the poorly differentiated forms (PDC) presents a genetic complexity similar to the solid ACC.

Genetic profile of adenoid cystic carcinomas (ACC) with high-grade transformation versus solid type.

PubMed

Costa, Ana Flávia; Altemani, Albina; Vékony, Hedy; Bloemena, Elisabeth; Fresno, Florentino; Suárez, Carlos; Llorente, José Luis; Hermsen, Mario

2011-08-01

ACC can occasionally undergo dedifferentiation also referred to as high-grade transformation (ACC-HGT). However, ACC-HGT can also undergo transformation to adenocarcinomas which are not poorly differentiated. ACC-HGT is generally considered to be an aggressive variant of ACC, even more than solid ACC. This study was aimed to describe the genetic changes of ACC-HGT in relation to clinico-pathological features, and to compare results to solid ACC. Genome wide DNA copy number changes were analyzed by microarray CGH in ACC-HGT, four with transformation into moderately differentiated adenocarcinoma (MDA) and two into poorly differentiated carcinoma (PDC), and five solid ACC. In addition, Ki67 index and p53 immunopositivity was assessed. ACC-HGT carried fewer copy number changes compared to solid ACC. Two ACC-HGT cases harboured a breakpoint at 6q23, near the cMYB oncogene. The complexity of the genomic profile concurred with the clinical course of the patient. Among the ACC-HGT, p53 positivity significantly increased from the conventional to the transformed (both MDA and PDC) component. ACC-HGT may not necessarily reflect a more advanced stage of tumor progression, but rather a transformation to another histological form in which the poorly differentiated forms (PDC) presents a genetic complexity similar to the solid ACC.

Metamorphosis in Teleosts

PubMed Central

McMenamin, Sarah K.; Parichy, David M.

2017-01-01

Teleosts are the largest and most diverse group of vertebrates, and many species undergo morphological, physiological, and behavioral transitions, “metamorphoses,” as they progress between morphologically divergent life stages. The larval metamorphosis that generally occurs as teleosts mature from larva to juvenile involves the loss of embryo-specific features, the development of new adult features, major remodeling of different organ systems, and changes in physical proportions and overall phenotype. Yet, in contrast to anuran amphibians, for example, teleost metamorphosis can entail morphological change that is either sudden and profound, or relatively gradual and subtle. Here, we review the definition of metamorphosis in teleosts, the diversity of teleost metamorphic strategies and the transitions they involve, and what is known of their underlying endocrine and genetic bases. We suggest that teleost metamorphosis offers an outstanding opportunity for integrating our understanding of endocrine mechanisms, cellular processes of morphogenesis and differentiation, and the evolution of diverse morphologies and life histories. PMID:23347518

Development of extracellular matrix in chick paravertebral sympathetic ganglia.

PubMed

Luckenbill-Edds, L

1986-08-01

Alcian blue staining coupled with enzyme digestion or critical electrolyte staining revealed differences in the development of extracellular matrix (ECM) within sympathetic ganglia compared with the surrounding capsule. On day 5 of chick development (Hamburger-Hamilton stage 26) only hyaluronic acid (HA) could be detected in the ECM surrounding condensing primary ganglia. By day 7 (st 30) the ganglionic capsule contained HA, as well as sulfated glycosaminoglycans (GAGs), and this pattern continued into the adult stage. During the later stages of embryonic life (st 41-45) satellite cells appear, showing fine structural characteristics that point to their role in the secretion of intraganglionic ECM. Only during these stages could ECM be detected histochemically within ganglia, the same stages (days 15-19) when routine electron microscopic methods reveal collagen fibrils embedded in a granular ground substance. Thus, the intraganglionic environment appears as a separate compartment free of detectable amounts of GAG until late embryonic stages when ECM is secreted around satellite cells. This developmental pattern could represent a role of ECM in the histological stabilization of ganglia during the late stages of differentiation, since the appearance of intraganglionic ECM is correlated with the appearance of small dense-cored vesicles characteristic of adult neurons. The developmental pattern of ECM in differentiating sympathetic ganglia is compared with that of other tissues that undergo condensation and morphogenesis.

Expression profile of senescence-associated beta-galactosidase and activation of telomerase in human ovarian surface epithelial cells undergoing immortalization.

PubMed

Litaker, J R; Pan, J; Cheung, Y; Zhang, D K; Liu, Y; Wong, S C; Wan, T S; Tsao, S W

1998-11-01

Senescence is a specific physiological stage of cells characterized by long population doubling time. It accounts for the inability of normal somatic cells to undergo indefinite cell division. As the number of population doublings increase, cell cycle regulatory mechanisms come into play and signal cells to exit the cell cycle and become senescent. Senescence has been implicated in the aging process and may function as a tumor suppressor mechanism in human cells. The ability to measure the degree of cellular senescence is important in understanding the biological processes regulating cell aging and immortalization. Senescent cells exhibit an enzyme termed senescence-associated histochemical staining. Cells immortalized by viral oncogenes often enter a stage of crisis at the early phase of immortalization. The cells at crisis have a long population doubling time. Cells at the crisis stage resemble senescent cells and the expression of SA- beta-Gal may be used to monitor the process of immortalization. In this study the expression profile of SA-beta-Gal was examined in human ovarian surface epithelial cells (HOSE 6-3) undergoing immortalization by the human papilloma viral oncogene E6 and E7 (HPV E6 and E7). Our results showed a low percentage (12.0%) of HOSE 6-3 cells expressing SA-beta-Gal activity at the pre-crisis stage. The percentage of HOSE 6-3 cells expressing SA-beta-Gal activity was highest (39.2%) at the crisis stage. When HOSE 6-3 cells achieved immortalized status there was a sharp decrease in cells (1. 3%) expressing SA-beta-Gal activity. In addition, an inverse relationship between the expression of SA-beta-Gal activity and telomerase activity was noted in cells undergoing immortalization. The results confirm that the SA-beta-Gal enzyme is a good marker for monitoring the population of cells undergoing senescence at different stages of immortalization and that telomerase activation is a characteristic feature of post-crisis cells.

Differentially co-expressed interacting protein pairs discriminate samples under distinct stages of HIV type 1 infection.

PubMed

Yoon, Dukyong; Kim, Hyosil; Suh-Kim, Haeyoung; Park, Rae Woong; Lee, KiYoung

2011-01-01

Microarray analyses based on differentially expressed genes (DEGs) have been widely used to distinguish samples across different cellular conditions. However, studies based on DEGs have not been able to clearly determine significant differences between samples of pathophysiologically similar HIV-1 stages, e.g., between acute and chronic progressive (or AIDS) or between uninfected and clinically latent stages. We here suggest a novel approach to allow such discrimination based on stage-specific genetic features of HIV-1 infection. Our approach is based on co-expression changes of genes known to interact. The method can identify a genetic signature for a single sample as contrasted with existing protein-protein-based analyses with correlational designs. Our approach distinguishes each sample using differentially co-expressed interacting protein pairs (DEPs) based on co-expression scores of individual interacting pairs within a sample. The co-expression score has positive value if two genes in a sample are simultaneously up-regulated or down-regulated. And the score has higher absolute value if expression-changing ratios are similar between the two genes. We compared characteristics of DEPs with that of DEGs by evaluating their usefulness in separation of HIV-1 stage. And we identified DEP-based network-modules and their gene-ontology enrichment to find out the HIV-1 stage-specific gene signature. Based on the DEP approach, we observed clear separation among samples from distinct HIV-1 stages using clustering and principal component analyses. Moreover, the discrimination power of DEPs on the samples (70-100% accuracy) was much higher than that of DEGs (35-45%) using several well-known classifiers. DEP-based network analysis also revealed the HIV-1 stage-specific network modules; the main biological processes were related to "translation," "RNA splicing," "mRNA, RNA, and nucleic acid transport," and "DNA metabolism." Through the HIV-1 stage-related modules, changing stage-specific patterns of protein interactions could be observed. DEP-based method discriminated the HIV-1 infection stages clearly, and revealed a HIV-1 stage-specific gene signature. The proposed DEP-based method might complement existing DEG-based approaches in various microarray expression analyses.

Ovule development: identification of stage-specific and tissue-specific cDNAs.

PubMed Central

Nadeau, J A; Zhang, X S; Li, J; O'Neill, S D

1996-01-01

A differential screening approach was used to identify seven ovule-specific cDNAs representing genes that are expressed in a stage-specific manner during ovule development. The Phalaenopsis orchid takes 80 days to complete the sequence of ovule developmental events, making it a good system to isolate stage-specific ovule genes. We constructed cDNA libraries from orchid ovule tissue during archesporial cell differentiation, megasporocyte formation, and the transition to meiosis, as well as during the final mitotic divisions of female gametophyte development. RNA gel blot hybridization analysis revealed that four clones were stage specific and expressed solely in ovule tissue, whereas one clone was specific to pollen tubes. Two other clones were not ovule specific. Sequence analysis and in situ hybridization revealed the identities and domain of expression of several of the cDNAs. O39 encodes a putative homeobox transcription factor that is expressed early in the differentiation of the ovule primordium; O40 encodes a cytochrome P450 monooxygenase (CYP78A2) that is pollen tube specific. O108 encodes a protein of unknown function that is expressed exclusively in the outer layer of the outer integument and in the female gametophyte of mature ovules. O126 encodes a glycine-rich protein that is expressed in mature ovules, and O141 encodes a cysteine proteinase that is expressed in the outer integument of ovules during seed formation. Sequences homologous to these ovule clones can now be isolated from other organisms, and this should facilitate their functional characterization. PMID:8742709

Differential requirements for Runx proteins in CD4 repression and epigenetic silencing during T lymphocyte development.

PubMed

Taniuchi, Ichiro; Osato, Motomi; Egawa, Takeshi; Sunshine, Mary Jean; Bae, Suk Chul; Komori, Toshihisa; Ito, Yoshiaki; Littman, Dan R

2002-11-27

T lymphocytes differentiate in discrete stages within the thymus. Immature thymocytes lacking CD4 and CD8 coreceptors differentiate into double-positive cells (CD4(+)CD8(+)), which are selected to become either CD4(+)CD8(-)helper cells or CD4(-)CD8(+) cytotoxic cells. A stage-specific transcriptional silencer regulates expression of CD4 in both immature and CD4(-)CD8(+) thymocytes. We show here that binding sites for Runt domain transcription factors are essential for CD4 silencer function at both stages, and that different Runx family members are required to fulfill unique functions at each stage. Runx1 is required for active repression in CD4(-)CD8(-) thymocytes whereas Runx3 is required for establishing epigenetic silencing in cytotoxic lineage thymocytes. Runx3-deficient cytotoxic T cells, but not helper cells, have defective responses to antigen, suggesting that Runx proteins have critical functions in lineage specification and homeostasis of CD8-lineage T lymphocytes.

Advanced stages of embryonic development and cotylocidial morphogenesis in the intrauterine eggs of Aspidogaster limacoides Diesing, 1835 (Aspidogastrea), with comments on their phylogenetic implications.

PubMed

Świderski, Zdzisław; Poddubnaya, Larisa G; Gibson, David I; Młocicki, Daniel

2012-06-01

Ultrastructural aspects of the advanced embryonic development and cotylocidial morphogenesis of the aspidogastrean Aspidogaster limacoides are described. The posterior or distal regions of the uterus are filled with eggs containing larvae at advanced stages of morphogenesis and fully-formed cotylocidia. Various stages and organs of this larva are described in detail, including the aspects of the developing and fully-differentiated cotylocidium, the body wall (tegument and musculature), glandular regions and the protonephridial excretory system. Blastomere multiplication by means of mitotic divisions takes place simultaneously with the degeneration or apoptosis of some micromeres; this frequently observed characteristic is compared and discussed in relation to corresponding reports for other neodermatans. During the advanced stages of the embryonic development of A. limacoides, the vitelline syncytium disappears and the size of the embryo increases rapidly. Evident polarization of the differentiating larva was observed; towards one pole of the egg, cytodifferentiation of the mouth, surrounded by the oral sucker and cephalic glands, takes place, whereas, towards the opposite pole, differentiation of the posterior sucker (incipient ventral disc) occurs. The oral and posterior suckers are formed from numerous embryonic cells which have differentiated into myocytes. The central part of the oral sucker undergoes invagination and forms the future pharynx and intestine. Fully-developed cotylocidia of A. limacoides have a neodermatan type of tegument, flame cells and two types of glandular structures. These results suggest a sister relationship between the Aspidogastrea and the Digenea, although the systematic position of aspidogastreans in relation to other platyhelminth taxa remains somewhat equivocal.

Characterization and differential expression patterns of conserved microRNAs and mRNAs in three genders of the rice field eel (Monopterus albus).

PubMed

Gao, Yu; Guo, Wei; Hu, Qing; Zou, Ming; Tang, Rong; Chi, Wei; Li, Dapeng

2014-01-01

MicroRNAs (miRNAs) are endogenous small RNAs that can regulate target mRNAs by binding to their sequences in the 3' untranslated region. The expression of miRNAs and their biogenetic pathway are involved in sexual differentiation and in the regulation of the development of germ cells and gonadal somatic cells. The rice field eel (Monopterus albus) undergoes a natural sexual transformation from female to male via an intersex stage during its life cycle. To investigate the molecular mechanisms of this sexual transformation, miRNAs present in the different sexual stages of the rice field eel were identified by high-throughput sequencing technology. A significantly differential expression among the 3 genders (p < 0.001) was observed for 48 unique miRNAs and 3 miRNAs*. Only 9 unique miRNAs showed a more than 8-fold change in their expression among the 3 genders, including mal-miR-430a and mal-miR-430c which were higher in females than in males. However, mal-miR-430b was only detected in males. Several potential miRNA target genes (cyp19a, cyp19b, nr5a1b, foxl2 amh, and vasa) were also investigated. Real-time RT-PCR demonstrated highly specific expression patterns of these genes in the 3 genders of the rice field eel. Many of these genes are targets of mal-miR-430b according to the TargetScan and miRTarBase. These results suggest that the miR-430 family may be involved in the sexual transformation of the rice field eel. © 2014 S. Karger AG, Basel.

Temporal dynamics of the Saccharopolyspora erythraea phosphoproteome.

PubMed

Licona-Cassani, Cuauhtemoc; Lim, Sooa; Marcellin, Esteban; Nielsen, Lars K

2014-05-01

Actinomycetes undergo a dramatic reorganization of metabolic and cellular machinery during a brief period of growth arrest ("metabolic switch") preceding mycelia differentiation and the onset of secondary metabolite biosynthesis. This study explores the role of phosphorylation in coordinating the metabolic switch in the industrial actinomycete Saccharopolyspora erythraea. A total of 109 phosphopeptides from 88 proteins were detected across a 150-h fermentation using open-profile two-dimensional LC-MS proteomics and TiO(2) enrichment. Quantitative analysis of the phosphopeptides and their unphosphorylated cognates was possible for 20 pairs that also displayed constant total protein expression. Enzymes from central carbon metabolism such as putative acetyl-coenzyme A carboxylase, isocitrate lyase, and 2-oxoglutarate dehydrogenase changed dramatically in the degree of phosphorylation during the stationary phase, suggesting metabolic rearrangement for the reutilization of substrates and the production of polyketide precursors. In addition, an enzyme involved in cellular response to environmental stress, trypsin-like serine protease (SACE_6340/NC_009142_6216), decreased in phosphorylation during the growth arrest stage. More important, enzymes related to the regulation of protein synthesis underwent rapid phosphorylation changes during this stage. Whereas the degree of phosphorylation of ribonuclease Rne/Rng (SACE_1406/NC_009142_1388) increased during the metabolic switch, that of two ribosomal proteins, S6 (SACE_7351/NC_009142_7233) and S32 (SACE_6101/NC_009142_5981), dramatically decreased during this stage of the fermentation, supporting the hypothesis that ribosome subpopulations differentially regulate translation before and after the metabolic switch. Overall, we show the great potential of phosphoproteomic studies to explain microbial physiology and specifically provide evidence of dynamic protein phosphorylation events across the developmental cycle of actinomycetes.

Temporal Dynamics of the Saccharopolyspora erythraea Phosphoproteome*

PubMed Central

Licona-Cassani, Cuauhtemoc; Lim, SooA; Marcellin, Esteban; Nielsen, Lars K.

2014-01-01

Actinomycetes undergo a dramatic reorganization of metabolic and cellular machinery during a brief period of growth arrest (“metabolic switch”) preceding mycelia differentiation and the onset of secondary metabolite biosynthesis. This study explores the role of phosphorylation in coordinating the metabolic switch in the industrial actinomycete Saccharopolyspora erythraea. A total of 109 phosphopeptides from 88 proteins were detected across a 150-h fermentation using open-profile two-dimensional LC-MS proteomics and TiO2 enrichment. Quantitative analysis of the phosphopeptides and their unphosphorylated cognates was possible for 20 pairs that also displayed constant total protein expression. Enzymes from central carbon metabolism such as putative acetyl-coenzyme A carboxylase, isocitrate lyase, and 2-oxoglutarate dehydrogenase changed dramatically in the degree of phosphorylation during the stationary phase, suggesting metabolic rearrangement for the reutilization of substrates and the production of polyketide precursors. In addition, an enzyme involved in cellular response to environmental stress, trypsin-like serine protease (SACE_6340/NC_009142_6216), decreased in phosphorylation during the growth arrest stage. More important, enzymes related to the regulation of protein synthesis underwent rapid phosphorylation changes during this stage. Whereas the degree of phosphorylation of ribonuclease Rne/Rng (SACE_1406/NC_009142_1388) increased during the metabolic switch, that of two ribosomal proteins, S6 (SACE_7351/NC_009142_7233) and S32 (SACE_6101/NC_009142_5981), dramatically decreased during this stage of the fermentation, supporting the hypothesis that ribosome subpopulations differentially regulate translation before and after the metabolic switch. Overall, we show the great potential of phosphoproteomic studies to explain microbial physiology and specifically provide evidence of dynamic protein phosphorylation events across the developmental cycle of actinomycetes. PMID:24615062

Regulation of proliferation in developing human tooth germs by MSX homeodomain proteins and cyclin-dependent kinase inhibitor p19INK4d.

PubMed

Kero, Darko; Vukojevic, Katarina; Stazic, Petra; Sundov, Danijela; Mardesic Brakus, Snjezana; Saraga-Babic, Mirna

2017-10-02

Before the secretion of hard dental tissues, tooth germs undergo several distinctive stages of development (dental lamina, bud, cap and bell). Every stage is characterized by specific proliferation patterns, which is regulated by various morphogens, growth factors and homeodomain proteins. The role of MSX homeodomain proteins in odontogenesis is rather complex. Expression domains of genes encoding for murine Msx1/2 during development are observed in tissues containing highly proliferative progenitor cells. Arrest of tooth development in Msx knockout mice can be attributed to impaired proliferation of progenitor cells. In Msx1 knockout mice, these progenitor cells start to differentiate prematurely as they strongly express cyclin-dependent kinase inhibitor p19 INK4d . p19 INK4d induces terminal differentiation of cells by blocking the cell cycle in mitogen-responsive G1 phase. Direct suppression of p19 INK4d by Msx1 protein is, therefore, important for maintaining proliferation of progenitor cells at levels required for the normal progression of tooth development. In this study, we examined the expression patterns of MSX1, MSX2 and p19 INK4d in human incisor tooth germs during the bud, cap and early bell stages of development. The distribution of expression domains of p19 INK4d throughout the investigated period indicates that p19 INK4d plays active role during human tooth development. Furthermore, comparison of expression domains of p19 INK4d with those of MSX1, MSX2 and proliferation markers Ki67, Cyclin A2 and pRb, indicates that MSX-mediated regulation of proliferation in human tooth germs might not be executed by the mechanism similar to one described in developing tooth germs of wild-type mouse.

Distinct functional and temporal requirements for zebrafish Hdac1 during neural crest-derived craniofacial and peripheral neuron development.

PubMed

Ignatius, Myron S; Unal Eroglu, Arife; Malireddy, Smitha; Gallagher, Glen; Nambiar, Roopa M; Henion, Paul D

2013-01-01

The regulation of gene expression is accomplished by both genetic and epigenetic means and is required for the precise control of the development of the neural crest. In hdac1(b382) mutants, craniofacial cartilage development is defective in two distinct ways. First, fewer hoxb3a, dlx2 and dlx3-expressing posterior branchial arch precursors are specified and many of those that are consequently undergo apoptosis. Second, in contrast, normal numbers of progenitors are present in the anterior mandibular and hyoid arches, but chondrocyte precursors fail to terminally differentiate. In the peripheral nervous system, there is a disruption of enteric, DRG and sympathetic neuron differentiation in hdac1(b382) mutants compared to wildtype embryos. Specifically, enteric and DRG-precursors differentiate into neurons in the anterior gut and trunk respectively, while enteric and DRG neurons are rarely present in the posterior gut and tail. Sympathetic neuron precursors are specified in hdac1(b382) mutants and they undergo generic neuronal differentiation but fail to undergo noradrenergic differentiation. Using the HDAC inhibitor TSA, we isolated enzyme activity and temporal requirements for HDAC function that reproduce hdac1(b382) defects in craniofacial and sympathetic neuron development. Our study reveals distinct functional and temporal requirements for zebrafish hdac1 during neural crest-derived craniofacial and peripheral neuron development.

Changes in the Golgi Apparatus of Neocortical and Hippocampal Neurons in the Hibernating Hamster.

PubMed

Antón-Fernández, Alejandro; León-Espinosa, Gonzalo; DeFelipe, Javier; Muñoz, Alberto

2015-01-01

Hibernating animals have been used as models to study several aspects of the plastic changes that occur in the metabolism and physiology of neurons. These models are also of interest in the study of Alzheimer's disease because the microtubule-associated protein tau is hyperphosphorylated during the hibernation state known as torpor, similar to the pretangle stage of Alzheimer's disease. Hibernating animals undergo torpor periods with drops in body temperature and metabolic rate, and a virtual cessation of neural activity. These processes are accompanied by morphological and neurochemical changes in neurons, which reverse a few hours after coming out of the torpor state. Since tau has been implicated in the structural regulation of the neuronal Golgi apparatus (GA) we have used Western Blot and immunocytochemistry to analyze whether the GA is modified in cortical neurons of the Syrian hamster at different hibernation stages. The results show that, during the hibernation cycle, the GA undergo important structural changes along with differential modifications in expression levels and distribution patterns of Golgi structural proteins. These changes were accompanied by significant transitory reductions in the volume and surface area of the GA elements during torpor and arousal stages as compared with euthermic animals.

Changes in the Golgi Apparatus of Neocortical and Hippocampal Neurons in the Hibernating Hamster

PubMed Central

Antón-Fernández, Alejandro; León-Espinosa, Gonzalo; DeFelipe, Javier; Muñoz, Alberto

2015-01-01

Hibernating animals have been used as models to study several aspects of the plastic changes that occur in the metabolism and physiology of neurons. These models are also of interest in the study of Alzheimer's disease because the microtubule-associated protein tau is hyperphosphorylated during the hibernation state known as torpor, similar to the pretangle stage of Alzheimer's disease. Hibernating animals undergo torpor periods with drops in body temperature and metabolic rate, and a virtual cessation of neural activity. These processes are accompanied by morphological and neurochemical changes in neurons, which reverse a few hours after coming out of the torpor state. Since tau has been implicated in the structural regulation of the neuronal Golgi apparatus (GA) we have used Western Blot and immunocytochemistry to analyze whether the GA is modified in cortical neurons of the Syrian hamster at different hibernation stages. The results show that, during the hibernation cycle, the GA undergo important structural changes along with differential modifications in expression levels and distribution patterns of Golgi structural proteins. These changes were accompanied by significant transitory reductions in the volume and surface area of the GA elements during torpor and arousal stages as compared with euthermic animals. PMID:26696838

Development of Planning Abilities in Normal Aging: Differential Effects of Specific Cognitive Demands

ERIC Educational Resources Information Center

Köstering, Lena; Stahl, Christoph; Leonhart, Rainer; Weiller, Cornelius; Kaller, Christoph P.

2014-01-01

In line with the frontal hypothesis of aging, the ability to plan ahead undergoes substantial change during normal aging. Although impairments on the Tower of London planning task were reported earlier, associations between age-related declines and specific cognitive demands on planning have not been studied. Here we investigated the impact of…

Tissue Specificity and Dynamics of Sex-Biased Gene Expression in a Common Frog Population with Differentiated, Yet Homomorphic, Sex Chromosomes.

PubMed

Ma, Wen-Juan; Veltsos, Paris; Toups, Melissa A; Rodrigues, Nicolas; Sermier, Roberto; Jeffries, Daniel L; Perrin, Nicolas

2018-06-12

Sex-biased genes are central to the study of sexual selection, sexual antagonism, and sex chromosome evolution. We describe a comprehensive de novo assembled transcriptome in the common frog Rana temporaria based on five developmental stages and three adult tissues from both sexes, obtained from a population with karyotypically homomorphic but genetically differentiated sex chromosomes. This allows the study of sex-biased gene expression throughout development, and its effect on the rate of gene evolution while accounting for pleiotropic expression, which is known to negatively correlate with the evolutionary rate. Overall, sex-biased genes had little overlap among developmental stages and adult tissues. Late developmental stages and gonad tissues had the highest numbers of stage- or tissue-specific genes. We find that pleiotropic gene expression is a better predictor than sex bias for the evolutionary rate of genes, though it often interacts with sex bias. Although genetically differentiated, the sex chromosomes were not enriched in sex-biased genes, possibly due to a very recent arrest of XY recombination. These results extend our understanding of the developmental dynamics, tissue specificity, and genomic localization of sex-biased genes.

Upright Imaging of Drosophila Egg Chambers

PubMed Central

Manning, Lathiena; Starz-Gaiano, Michelle

2015-01-01

Drosophila melanogaster oogenesis provides an ideal context for studying varied developmental processes since the ovary is relatively simple in architecture, is well-characterized, and is amenable to genetic analysis. Each egg chamber consists of germ-line cells surrounded by a single epithelial layer of somatic follicle cells. Subsets of follicle cells undergo differentiation during specific stages to become several different cell types. Standard techniques primarily allow for a lateral view of egg chambers, and therefore a limited view of follicle cell organization and identity. The upright imaging protocol describes a mounting technique that enables a novel, vertical view of egg chambers with a standard confocal microscope. Samples are first mounted between two layers of glycerin jelly in a lateral (horizontal) position on a glass microscope slide. The jelly with encased egg chambers is then cut into blocks, transferred to a coverslip, and flipped to position egg chambers upright. Mounted egg chambers can be imaged on either an upright or an inverted confocal microscope. This technique enables the study of follicle cell specification, organization, molecular markers, and egg development with new detail and from a new perspective. PMID:25867882

Comparative Digital Gene Expression Analysis of the Arabidopsis Response to Volatiles Emitted by Bacillus amyloliquefaciens

PubMed Central

Hao, Hai-Ting; Zhao, Xia; Shang, Qian-Han; Wang, Yun; Guo, Zhi-Hong; Zhang, Yu-Bao; Xie, Zhong-Kui; Wang, Ruo-Yu

2016-01-01

Some plant growth-promoting rhizobacteria (PGPR) regulated plant growth and elicited plant basal immunity by volatiles. The response mechanism to the Bacillus amyloliquefaciens volatiles in plant has not been well studied. We conducted global gene expression profiling in Arabidopsis after treatment with Bacillus amyloliquefaciens FZB42 volatiles by Illumina Digital Gene Expression (DGE) profiling of different growth stages (seedling and mature) and tissues (leaves and roots). Compared with the control, 1,507 and 820 differentially expressed genes (DEGs) were identified in leaves and roots at the seedling stage, respectively, while 1,512 and 367 DEGs were identified in leaves and roots at the mature stage. Seventeen genes with different regulatory patterns were validated using quantitative RT-PCR. Numerous DEGs were enriched for plant hormones, cell wall modifications, and protection against stress situations, which suggests that volatiles have effects on plant growth and immunity. Moreover, analyzes of transcriptome difference in tissues and growth stage using DGE profiling showed that the plant response might be tissue-specific and/or growth stage-specific. Thus, genes encoding flavonoid biosynthesis were downregulated in leaves and upregulated in roots, thereby indicating tissue-specific responses to volatiles. Genes related to photosynthesis were downregulated at the seedling stage and upregulated at the mature stage, respectively, thereby suggesting growth period-specific responses. In addition, the emission of bacterial volatiles significantly induced killing of cells of other organism pathway with up-regulated genes in leaves and the other three pathways (defense response to nematode, cell morphogenesis involved in differentiation and trichoblast differentiation) with up-regulated genes were significantly enriched in roots. Interestingly, some important alterations in the expression of growth-related genes, metabolic pathways, defense response to biotic stress and hormone-related genes were firstly founded response to FZB42 volatiles. PMID:27513952

Comparative anatomy and histology of developmental and parasitic stages in the life cycle of the lined sea anemone Edwardsiella lineata.

PubMed

Reitzel, Adam M; Daly, Marymegan; Sullivan, James C; Finnerty, John R

2009-02-01

The evolution of parasitism is often accompanied by profound changes to the developmental program. However, relatively few studies have directly examined the developmental evolution of parasitic species from free-living ancestors. The lined sea anemone Edwardsiella lineata is a relatively recently evolved parasite for which closely related free-living outgroups are known, including the starlet sea anemone Nematostella vectensis. The larva of E. lineata parasitizes the ctenophore Mnemiopsis leidyi, and, once embedded in its host, the anemone assumes a novel vermiform body plan. That we might begin to understand how the developmental program of this species has been transformed during the evolution of parasitism, we characterized the gross anatomy, histology, and cnidom of the parasitic stage, post-parasitic larval stage, and adult stage of the E. lineata life cycle. The distinct parasitic stage of the life cycle differs from the post-parasitic larva with respect to overall shape, external ciliation, cnida frequency, and tissue architecture. The parasitic stage and planula both contain holotrichs, a type of cnida not previously reported in Edwardsiidae. The internal morphology of the post-parasitic planula is extremely similar to the adult morphology, with a complete set of mesenterial tissue and musculature despite this stage having little external differentiation. Finally, we observed 2 previously undocumented aspects of asexual reproduction in E. lineata: (1) the parasitic stage undergoes transverse fission via physal pinching, the first report of asexual reproduction in a pre-adult stage in the Edwardsiidae; and (2) the juvenile polyp undergoes transverse fission via polarity reversal, the first time this form of fission has been reported in E. lineata.

Chitosan-Intercalated Montmorillonite/Poly(vinyl alcohol) Nanofibers as a Platform to Guide Neuronlike Differentiation of Human Dental Pulp Stem Cells.

PubMed

Ghasemi Hamidabadi, Hatef; Rezvani, Zahra; Nazm Bojnordi, Maryam; Shirinzadeh, Haji; Seifalian, Alexander M; Joghataei, Mohammad Taghi; Razaghpour, Mojgan; Alibakhshi, Abbas; Yazdanpanah, Abolfazl; Salimi, Maryam; Mozafari, Masoud; Urbanska, Aleksandra M; Reis, Rui L; Kundu, Subhas C; Gholipourmalekabadi, Mazaher

2017-04-05

In this study, we present a novel chitosan-intercalated montmorillonite/poly(vinyl alcohol) (OMMT/PVA) nanofibrous mesh as a microenvironment for guiding differentiation of human dental pulp stem cells (hDPSCs) toward neuronlike cells. The OMMT was prepared through ion exchange reaction between the montmorillonite (MMT) and chitosan. The PVA solutions containing various concentrations of OMMT were electrospun to form 3D OMMT-PVA nanofibrous meshes. The biomechanical and biological characteristics of the nanofibrous meshes were evaluated by ATR-FTIR, XRD, SEM, MTT, and LDH specific activity, contact angle, and DAPI staining. They were carried out for mechanical properties, overall viability, and toxicity of the cells. The hDPSCs were seeded on the prepared scaffolds and induced with neuronal specific differentiation media at two differentiation stages (2 days at preinduction stage and 6 days at induction stage). The neural differentiation of the cells cultured on the meshes was evaluated by determining the expression of Oct-4, Nestin, NF-M, NF-H, MAP2, and βIII-tubulin in the cells after preinduction, at induction stages by real-time PCR (RT-PCR) and immunostaining. All the synthesized nanofibers exhibited a homogeneous morphology with a favorable mechanical behavior. The population of the cells differentiated into neuronlike cells in all the experimental groups was significantly higher than that in control group. The expression level of the neuronal specific markers in the cells cultured on 5% OMMT/PVA meshes was significantly higher than the other groups. This study demonstrates the feasibility of the OMMT/PVA artificial nerve graft cultured with hDPSCs for regeneration of damaged neural tissues. These fabricated matrices may have a potential in neural tissue engineering applications.

Specific TRPC6 Channel Activation, a Novel Approach to Stimulate Keratinocyte Differentiation*S⃞

PubMed Central

Müller, Margarethe; Essin, Kirill; Hill, Kerstin; Beschmann, Heike; Rubant, Simone; Schempp, Christoph M.; Gollasch, Maik; Boehncke, W. Henning; Harteneck, Christian; Müller, Walter E.; Leuner, Kristina

2008-01-01

The protective epithelial barrier in our skin undergoes constant regulation, whereby the balance between differentiation and proliferation of keratinocytes plays a major role. Impaired keratinocyte differentiation and proliferation are key elements in the pathophysiology of several important dermatological diseases, including atopic dermatitis and psoriasis. Ca2+ influx plays an essential role in this process presumably mediated by different transient receptor potential (TRP) channels. However, investigating their individual role was hampered by the lack of specific stimulators or inhibitors. Because we have recently identified hyperforin as a specific TRPC6 activator, we investigated the contribution of TRPC6 to keratinocyte differentiation and proliferation. Like the endogenous differentiation stimulus high extracellular Ca2+ concentration ([Ca2+]o), hyperforin triggers differentiation in HaCaT cells and in primary cultures of human keratinocytes by inducing Ca2+ influx via TRPC6 channels and additional inhibition of proliferation. Knocking down TRPC6 channels prevents the induction of Ca2+- and hyperforin-induced differentiation. Importantly, TRPC6 activation is sufficient to induce keratinocyte differentiation similar to the physiological stimulus [Ca2+]o. Therefore, TRPC6 activation by hyperforin may represent a new innovative therapeutic strategy in skin disorders characterized by altered keratinocyte differentiation. PMID:18818211

Differentiating Mouse Embryonic Stem Cells into Embryoid Bodies by Hanging-Drop Cultures.

PubMed

Behringer, Richard; Gertsenstein, Marina; Nagy, Kristina Vintersten; Nagy, Andras

2016-12-01

Embryonic stem (ES) cells can develop into many types of differentiated tissues if they are placed into a differentiating environment. This can occur in vivo when the ES cells are injected into or aggregated with an embryo, or in vitro if their culture conditions are modified to induce differentiation. There are an increasing number of differentiating culture conditions that can bias the differentiation of ES cells into desired cell types. Determining the mechanisms that control ES cell differentiation into therapeutically important cell types is a quickly growing area of research. Knowledge gained from these studies may eventually lead to the use of stem cells to repair specific damaged tissues. Many times ES cell differentiation proceeds through an intermediate stage called the embryoid body (EB). EBs are round structures composed of ES cells that have undergone some of the initial stages of differentiation. EBs can then be manipulated further to generate more specific cell types. This protocol describes a method to differentiate ES cells into EBs. It produces EBs of comparable size. This aspect is important because the differentiation processes taking place inside an EB are influenced by its size. © 2016 Cold Spring Harbor Laboratory Press.

Is knowledge translation adequate? A quality assurance study of staging investigations in early stage breast cancer patients.

PubMed

Han, Dolly; Hogeveen, Sophie; Sweet Goldstein, Miriam; George, Ralph; Brezden-Masley, Christine; Hoch, Jeffrey; Haq, Rashida; Simmons, Christine E

2012-02-01

After primary surgery, patients diagnosed with early stage breast cancer undergo radiological investigations based on pathologic stage of disease to rule out distant metastases. Published guidelines can aid clinicians in determining which tests are appropriate based on stage of disease. We wished to assess the consistency of radiological staging in an academic community oncology setting with standard guidelines and to determine the overall impact of non-adherence to these guidelines. A retrospective cohort study was conducted for new breast cancer patients seen at a single institution between January 2009 and April 2010. Patients were included if initial diagnosis and primary surgery was at this institution. Pathologic stage and radiological tests completed were recorded. A literature review was performed and the results were compared with those from this study to determine overall adherence rates. Subsequently, a cost analysis was performed to determine the financial impact at this centre. 231 patients met eligibility criteria for inclusion in this study. A large proportion of patients were over-staged with 129 patients (55%) undergoing unnecessary investigations according to guidelines. Specifically, 59% of stage I patients and 58% of stage II patients were over-investigated. Distant metastases at the time of diagnosis were found in three patients, all of whom had stage III disease (1.3%). The literature reviewed revealed similar non-adherence rates in other centres. The estimated cost of such non-adherence is in the range of $78 (CDN) per new early stage breast cancer patient seen at this centre. This oncology centre has a low adherence to practice guidelines for staging investigations in breast cancer patients, with 55% of patients undergoing unnecessary tests. Very few patients had metastases at diagnosis, and all had pathological stage III disease. Efforts may need to focus on improving knowledge translation across clinical oncology settings to increase guideline adherence.

Temporal Expression of a Master Regulator Drives Synchronous Sporulation in Budding Yeast.

PubMed

Chia, Minghao; van Werven, Folkert J

2016-09-07

Yeast cells enter and undergo gametogenesis relatively asynchronously, making it technically challenging to perform stage-specific genomic and biochemical analyses. Cell-to-cell variation in the expression of the master regulator of entry into sporulation IME1, has been implicated to be the underlying cause of asynchronous sporulation. Here we find that timing of IME1 expression is of critical importance for inducing cells to undergo sporulation synchronously. When we force expression of IME1 from an inducible promoter in cells incubated in sporulation medium for two hours, the vast majority of cells exhibit synchrony during pre-meiotic DNA replication and meiotic divisions. Inducing IME1 expression too early or too late affects the synchrony of sporulation. Surprisingly, our approach for synchronous sporulation does not require growth in acetate containing medium, but can be achieved in cells grown in rich medium until saturation. Our system solely requires IME1 because the expression of the N6-methyladenosine methyltransferase IME4, another key regulator of early sporulation, is controlled by IME1 itself. The approach described here can be easily combined with other stage specific synchronization methods, and thereby applied to study specific stages of sporulation or the complete sporulation program. Copyright © 2016 Author et al.

High oxygen condition facilitates the differentiation of mouse and human pluripotent stem cells into pancreatic progenitors and insulin-producing cells.

PubMed

Hakim, Farzana; Kaitsuka, Taku; Raeed, Jamiruddin Mohd; Wei, Fan-Yan; Shiraki, Nobuaki; Akagi, Tadayuki; Yokota, Takashi; Kume, Shoen; Tomizawa, Kazuhito

2014-04-04

Pluripotent stem cells have potential applications in regenerative medicine for diabetes. Differentiation of stem cells into insulin-producing cells has been achieved using various protocols. However, both the efficiency of the method and potency of differentiated cells are insufficient. Oxygen tension, the partial pressure of oxygen, has been shown to regulate the embryonic development of several organs, including pancreatic β-cells. In this study, we tried to establish an effective method for the differentiation of induced pluripotent stem cells (iPSCs) into insulin-producing cells by culturing under high oxygen (O2) conditions. Treatment with a high O2 condition in the early stage of differentiation increased insulin-positive cells at the terminus of differentiation. We found that a high O2 condition repressed Notch-dependent gene Hes1 expression and increased Ngn3 expression at the stage of pancreatic progenitors. This effect was caused by inhibition of hypoxia-inducible factor-1α protein level. Moreover, a high O2 condition activated Wnt signaling. Optimal stage-specific treatment with a high O2 condition resulted in a significant increase in insulin production in both mouse embryonic stem cells and human iPSCs and yielded populations containing up to 10% C-peptide-positive cells in human iPSCs. These results suggest that culturing in a high O2 condition at a specific stage is useful for the efficient generation of insulin-producing cells.

Adipogenesis in thyroid eye disease.

PubMed

Crisp, M; Starkey, K J; Lane, C; Ham, J; Ludgate, M

2000-10-01

Adipogenesis contributes to the pathogenesis of thyroid eye disease (TED). Thyrotropin receptor (TSHR) transcripts are present in orbital fat. This study was conducted to determine whether they are expressed as functional protein, and if so, whether this is restricted to TED orbits or to a particular stage in adipocyte differentiation. Samples of fat were obtained from 18 TED-affected orbits and 4 normal orbits, and 9 were obtained from nonorbital locations. Frozen sections were examined by immunocytochemistry using monoclonal antibodies specific for the human TSHR. Samples were disaggregated and the preadipocytes separated from the mature by differential centrifugation and cultured in serum-free or DM and examined for morphologic changes, oil red O and TSHR staining, and TSH-induced cyclic adenosine monophosphate (cAMP) production. Marked immunoreactivity was observed in frozen sections from all three TED samples and faint staining in both normal orbital fat samples. In vitro, 1% to 5% of preadipocytes displayed TSHR immunoreactivity in five of six TED and two of three normal orbital samples and in three of five nonorbital samples. Differentiation, was induced in all 14 orbital samples. Three of four nonorbital samples contained occasional differentiated cells. Fifty percent to 70% of differentiating cells demonstrated receptor immunoreactivity. Two of three TED and four of four nonorbital preadipocytes in DM and/or mature adipocytes displayed a TSH-mediated increase in cAMP. The results indicate that orbital fat TSHR transcripts are expressed as protein, which can be functional. This is not aberrant in TED orbits, although expression may be upregulated. The majority of preadipocytes undergoing differentiation express the receptor, indicating a key role for this population in one mechanism for increasing orbital volume.

The predictive value of preoperative carcinoembryonic antigen level in the prognosis of colon cancer.

PubMed

Kirat, Hasan T; Ozturk, Ersin; Lavery, Ian C; Kiran, Ravi P

2012-10-01

We evaluated factors associated with an increased preoperative carcinoembryonic antigen (CEA) level for colon cancer patients undergoing elective curative surgery and assessed whether this was associated with prognosis when accounting for other potential confounders. Prospectively accrued data of patients with stage I, II, and III colon cancer undergoing surgery (1980-2008) were retrieved retrospectively. Patients with a preoperative CEA level greater than 5 ng/mL (group B) were compared with those with a CEA level of 5 ng/mL or less (group A). There were 651 patients (379 men) with a median age of 67 years (range, 21-94 y) and a median follow-up period of 5.9 years. Groups A (n = 451) and B (n = 200) had similar ages and tumor locations. Group B had larger tumors; more patients with T3 and N1/N2; and more patients with stage II/III tumors, and hence greater use of chemotherapy (P = .04). On multivariate analysis, patient age, tumor stage, and differentiation were associated with oncologic outcomes. A CEA level greater than 5 ng/mL was not associated independently with recurrence, recurrence-free survival (P = .47), or overall survival (P = .3). An increased preoperative CEA level is a marker for a more advanced tumor stage. For adequately staged patients, a high preoperative CEA level is not associated independently with oncologic outcomes. Copyright © 2012 Elsevier Inc. All rights reserved.

Evaluation and validation of the diagnostic value of the apparent diffusion coefficient for differentiating early-stage endometrial carcinomas from benign mimickers at 3T MRI.

PubMed

Wang, Xue; Zhao, Yu; Hu, Yumin; Zhou, Yongjin; Ye, Xinjian; Liu, Kun; Bai, Guanghui; Guo, Anna; Du, Meimei; Jiang, Lezhen; Wang, Jinhong; Yan, Zhihan

2017-07-11

Previous researchers obtained various apparent diffusion coefficient (ADC) cutoff values to differentiate endometrial carcinoma from benign mimickers with 1.5T magnetic resonance imaging (MRI). Few studies have used 3T MRI or validated the effectiveness of these cutoff ADC values prospectively. This study was designed in two stages to obtain a cutoff ADC value at 3T MRI and to validate prospectively the role of the ADC value. First, we conducted a retrospective study of 60 patients to evaluate the diagnostic value of ADC by obtain a theoretical cutoff ADC value for differentiating between benign and malignant endometrial lesions. Student's t test revealed that ADC values for stage I endometrial carcinomas were significantly lower than those for benign lesions. The area under the curve value of the receiver operating characteristic curve was 0.993, and the cutoff ADC value was 0.98 × 10-3 mm2/s. The sensitivity, specificity, and overall accuracy of diagnosing stage I endometrial carcinoma were 100%, 97.1%, and 98.3%, respectively. Second, we conducted a prospective study of 26 patients to validate the use of the cutoff ADC value obtained in the study's first stage. The sensitivity, specificity, and overall accuracy for differentiating malignant from benign endometrial lesions based on the cutoff ADC value obtained earlier were as follows: radiologist 1 attained 86.67%, 100.0%, and 92.31%, respectively; radiologist 2 attained 86.67%, 91.0%, and 88.5%, respectively. Our results suggest that ADC values could be a potential biomarker for use as a quantitative and qualitative tool for differentiating between early-stage endometrial carcinomas and benign mimickers.

Evaluation and validation of the diagnostic value of the apparent diffusion coefficient for differentiating early-stage endometrial carcinomas from benign mimickers at 3T MRI

PubMed Central

Hu, Yumin; Zhou, Yongjin; Ye, Xinjian; Liu, Kun; Bai, Guanghui; Guo, Anna; Du, Meimei; Jiang, Lezhen

2017-01-01

Previous researchers obtained various apparent diffusion coefficient (ADC) cutoff values to differentiate endometrial carcinoma from benign mimickers with 1.5T magnetic resonance imaging (MRI). Few studies have used 3T MRI or validated the effectiveness of these cutoff ADC values prospectively. This study was designed in two stages to obtain a cutoff ADC value at 3T MRI and to validate prospectively the role of the ADC value. First, we conducted a retrospective study of 60 patients to evaluate the diagnostic value of ADC by obtain a theoretical cutoff ADC value for differentiating between benign and malignant endometrial lesions. Student's t test revealed that ADC values for stage I endometrial carcinomas were significantly lower than those for benign lesions. The area under the curve value of the receiver operating characteristic curve was 0.993, and the cutoff ADC value was 0.98 × 10−3 mm2/s. The sensitivity, specificity, and overall accuracy of diagnosing stage I endometrial carcinoma were 100%, 97.1%, and 98.3%, respectively. Second, we conducted a prospective study of 26 patients to validate the use of the cutoff ADC value obtained in the study's first stage. The sensitivity, specificity, and overall accuracy for differentiating malignant from benign endometrial lesions based on the cutoff ADC value obtained earlier were as follows: radiologist 1 attained 86.67%, 100.0%, and 92.31%, respectively; radiologist 2 attained 86.67%, 91.0%, and 88.5%, respectively. Our results suggest that ADC values could be a potential biomarker for use as a quantitative and qualitative tool for differentiating between early-stage endometrial carcinomas and benign mimickers. PMID:28634318

Differentiation and Characterization of Myeloid Cells

PubMed Central

Gupta, Dipti; Shah, Hetavi Parag; Malu, Krishnakumar; Berliner, Nancy; Gaines, Peter

2015-01-01

Recent molecular studies of myeloid differentiation have utilized several in vitro models of myelopoiesis, generated from either ex vivo differentiated bone marrow progenitors or induced immortalized myeloid cell lines. Ex vivo differentiation begins with an enriched population of bone marrow-derived hematopoietic stem cells generated by lineage depletion and/or positive selection for CD34+ antigen (human) or Sca-1+ (mouse) cells, which are then expanded and subsequently induced in vitro in a process that recapitulates normal myeloid development. Myeloid cell lines include two human leukemic cell lines, NB-4 and HL-60, which have been demonstrated to undergo retinoic acid–induced myeloid development, however, both cell lines exhibit defects in the upregulation of late-expressed neutrophil-specific genes. Multiple murine factor–dependent cell models of myelopoiesis are also available that express the full range of neutrophil maturation markers, including: 32Dcl3 cells, which undergo G-CSF-induced myeloid maturation, EML/EPRO cells, which develop into mature neutrophils in response to cytokines and retinoic acid, and ER-Hoxb8 cells, which undergo myeloid maturation upon removal of estradial in the maintenance medium. In this unit, the induction of myeloid maturation in each of these model systems is described, including their differentiation to either neutrophils or macrophages, if applicable. Commonly used techniques to test for myeloid characteristics of developing cells are also described, including flow cytometry and real time RT-PCR. Together, these assays provide a solid foundation for in vitro investigations of myeloid development with either human or mouse models. PMID:24510620

XBtg2 is required for notochord differentiation during early Xenopus development.

PubMed

Sugimoto, Kaoru; Hayata, Tadayoshi; Asashima, Makoto

2005-09-01

The notochord is essential for normal vertebrate development, serving as both a structural support for the embryo and a signaling source for the patterning of adjacent tissues. Previous studies on the notochord have mostly focused on its formation and function in early organogenesis but gene regulation in the differentiation of notochord cells itself remains poorly defined. In the course of screening for genes expressed in developing notochord, we have isolated Xenopus homolog of Btg2 (XBtg2). The mammalian Btg2 genes, Btg2/PC3/TIS21, have been reported to have multiple functions in the regulation of cell proliferation and differentiation but their roles in early development are still unclear. Here we characterized XBtg2 in early Xenopus laevis embryogenesis with focus on notochord development. Translational inhibition of XBtg2 resulted in a shortened and bent axis phenotype and the abnormal structures in the notochord tissue, which did not undergo vacuolation. The XBtg2-depleted notochord cells expressed early notochord markers such as chordin and Xnot at the early tailbud stage, but failed to express differentiation markers of notochord such as Tor70 and 5-D-4 antigens in the later stages. These results suggest that XBtg2 is required for the differentiation of notochord cells such as the process of vacuolar formation after determination of notochord cell fate.

EOG and EMG: two important switches in automatic sleep stage classification.

PubMed

Estrada, E; Nazeran, H; Barragan, J; Burk, J R; Lucas, E A; Behbehani, K

2006-01-01

Sleep is a natural periodic state of rest for the body, in which the eyes are usually closed and consciousness is completely or partially lost. In this investigation we used the EOG and EMG signals acquired from 10 patients undergoing overnight polysomnography with their sleep stages determined by expert sleep specialists based on RK rules. Differentiation between Stage 1, Awake and REM stages challenged a well trained neural network classifier to distinguish between classes when only EEG-derived signal features were used. To meet this challenge and improve the classification rate, extra features extracted from EOG and EMG signals were fed to the classifier. In this study, two simple feature extraction algorithms were applied to EOG and EMG signals. The statistics of the results were calculated and displayed in an easy to visualize fashion to observe tendencies for each sleep stage. Inclusion of these features show a great promise to improve the classification rate towards the target rate of 100%

Possible pathways used to predict different stages of lung adenocarcinoma.

PubMed

Chen, Xiaodong; Duan, Qiongyu; Xuan, Ying; Sun, Yunan; Wu, Rong

2017-04-01

We aimed to find some specific pathways that can be used to predict the stage of lung adenocarcinoma.RNA-Seq expression profile data and clinical data of lung adenocarcinoma (stage I [37], stage II 161], stage III [75], and stage IV [45]) were obtained from the TCGA dataset. The differentially expressed genes were merged, correlation coefficient matrix between genes was constructed with correlation analysis, and unsupervised clustering was carried out with hierarchical clustering method. The specific coexpression network in every stage was constructed with cytoscape software. Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis was performed with KOBAS database and Fisher exact test. Euclidean distance algorithm was used to calculate total deviation score. The diagnostic model was constructed with SVM algorithm.Eighteen specific genes were obtained by getting intersection of 4 group differentially expressed genes. Ten significantly enriched pathways were obtained. In the distribution map of 10 pathways score in different groups, degrees that sample groups deviated from the normal level were as follows: stage I < stage II < stage III < stage IV. The pathway score of 4 stages exhibited linear change in some pathways, and the score of 1 or 2 stages were significantly different from the rest stages in some pathways. There was significant difference between dead and alive for these pathways except thyroid hormone signaling pathway.Those 10 pathways are associated with the development of lung adenocarcinoma and may be able to predict different stages of it. Furthermore, these pathways except thyroid hormone signaling pathway may be able to predict the prognosis.

Effector Regulatory T Cell Differentiation and Immune Homeostasis Depend on the Transcription Factor Myb.

PubMed

Dias, Sheila; D'Amico, Angela; Cretney, Erika; Liao, Yang; Tellier, Julie; Bruggeman, Christine; Almeida, Francisca F; Leahy, Jamie; Belz, Gabrielle T; Smyth, Gordon K; Shi, Wei; Nutt, Stephen L

2017-01-17

FoxP3-expressing regulatory T (Treg) cells are essential for maintaining immune homeostasis. Activated Treg cells undergo further differentiation into an effector state that highly expresses genes critical for Treg cell function, although how this process is coordinated on a transcriptional level is poorly understood. Here, we demonstrate that mice lacking the transcription factor Myb in Treg cells succumbed to a multi-organ inflammatory disease. Myb was specifically expressed in, and required for the differentiation of, thymus-derived effector Treg cells. The combination of transcriptome and genomic footprint analyses revealed that Myb directly regulated a large proportion of the gene expression specific to effector Treg cells, identifying Myb as a critical component of the gene regulatory network controlling effector Treg cell differentiation and function. Copyright © 2017 Elsevier Inc. All rights reserved.

Expressed Sequence Reference Standards for Evaluating Stage-specific Gene Expression in Southern Green Lacewings, Chrysoperla rufilabris

USDA-ARS?s Scientific Manuscript database

Five developmental stages of Chrysoperla rufilabris were tested using nine primer pairs. Three sequences were highly expressed at all life stages and six were differentially expressed. These primer pairs may be used as standards to quantitate functional gene expression associated with physiological ...

Genetic Variation in Dopamine Pathways Differentially Associated with Smoking Progression in Adolescence

ERIC Educational Resources Information Center

Laucht, Manfred; Becker, Katja; Frank, Josef; Schmidt, Martin H.; Esser, Gunter; Treutlein, Jens; Skowronek, Markus H.; Schumann, Gunter

2008-01-01

A study examines whether genetic variation in dopamine pathways differentially associate with smoking progression in adolescence. Results indicate the influence of specific dopamine genes in different stages of smoking progression in adolescents.

Spermatogonial stem cells in the testis of an endangered bovid: Indian black buck (Antilope cervicapra L.).

PubMed

Goel, Sandeep; Reddy, Niranjan; Mahla, Ranjeet Singh; Suman, Sanjay Kumar; Pawar, Rahul Mohanchandra

2011-07-01

Numerous wild bovids are facing threat of extinction owing to the loss of habitat and various other reasons. Spermatogonial stem cells (SSCs) represent the only germline stem cells in adult body that are capable of self-renewal and that can undergo differentiation to produce haploid germ cells. SSCs can, therefore, serve as a useful resource for preservation of germplasm of threatened and endangered mammals. The Indian black buck (Antilope cervicapra L.) is a small Indian antelope that is listed as endangered by the Indian Wildlife Protection Act, 1972. Immunohistochemical analysis of testes tissues of black buck revealed the presence of spermatogonia that were specifically stained by lectin-Dolichos biflorus agglutinin (DBA). The expression of pluripotent cell-specific markers, NANOG and stage-specific embryonic antigen-1 (SSEA-1), was detected in spermatogonia. Interestingly, the expression of POU5F1 (OCT3/4) was absent from spermatogonia, however, it was detected in differentiating cells such as spermatocytes and round spermatids but not in elongated spermatids. The expression of NANOG protein was also present in spermatocytes but absent in round and elongated spermatids. Using the testis transplantation assay, stem cell potential of black buck spermatogonia was confirmed as indicated by the presence of colonized DBA-stained cells in the basal membrane of seminiferous tubules of xenotransplanted mice testis. The findings from this study suggest the presence of SSCs in the testis of an endangered bovid for the first time and open new possibility to explore the use of SSCs in conservation. Copyright © 2011 Elsevier B.V. All rights reserved.

Deciphering the Developmental Dynamics of the Mouse Liver Transcriptome

PubMed Central

Gunewardena, Sumedha S.; Yoo, Byunggil; Peng, Lai; Lu, Hong; Zhong, Xiaobo; Klaassen, Curtis D.; Cui, Julia Yue

2015-01-01

During development, liver undergoes a rapid transition from a hematopoietic organ to a major organ for drug metabolism and nutrient homeostasis. However, little is known on a transcriptome level of the genes and RNA-splicing variants that are differentially regulated with age, and which up-stream regulators orchestrate age-specific biological functions in liver. We used RNA-Seq to interrogate the developmental dynamics of the liver transcriptome in mice at 12 ages from late embryonic stage (2-days before birth) to maturity (60-days after birth). Among 21,889 unique NCBI RefSeq-annotated genes, 9,641 were significantly expressed in at least one age, 7,289 were differently regulated with age, and 859 had multiple (> = 2) RNA splicing-variants. Factor analysis showed that the dynamics of hepatic genes fall into six distinct groups based on their temporal expression. The average expression of cytokines, ion channels, kinases, phosphatases, transcription regulators and translation regulators decreased with age, whereas the average expression of peptidases, enzymes and transmembrane receptors increased with age. The average expression of growth factors peak between Day-3 and Day-10, and decrease thereafter. We identified critical biological functions, upstream regulators, and putative transcription modules that seem to govern age-specific gene expression. We also observed differential ontogenic expression of known splicing variants of certain genes, and 1,455 novel splicing isoform candidates. In conclusion, the hepatic ontogeny of the transcriptome ontogeny has unveiled critical networks and up-stream regulators that orchestrate age-specific biological functions in liver, and suggest that age contributes to the complexity of the alternative splicing landscape of the hepatic transcriptome. PMID:26496202

Deciphering the Developmental Dynamics of the Mouse Liver Transcriptome.

PubMed

Gunewardena, Sumedha S; Yoo, Byunggil; Peng, Lai; Lu, Hong; Zhong, Xiaobo; Klaassen, Curtis D; Cui, Julia Yue

2015-01-01

During development, liver undergoes a rapid transition from a hematopoietic organ to a major organ for drug metabolism and nutrient homeostasis. However, little is known on a transcriptome level of the genes and RNA-splicing variants that are differentially regulated with age, and which up-stream regulators orchestrate age-specific biological functions in liver. We used RNA-Seq to interrogate the developmental dynamics of the liver transcriptome in mice at 12 ages from late embryonic stage (2-days before birth) to maturity (60-days after birth). Among 21,889 unique NCBI RefSeq-annotated genes, 9,641 were significantly expressed in at least one age, 7,289 were differently regulated with age, and 859 had multiple (> = 2) RNA splicing-variants. Factor analysis showed that the dynamics of hepatic genes fall into six distinct groups based on their temporal expression. The average expression of cytokines, ion channels, kinases, phosphatases, transcription regulators and translation regulators decreased with age, whereas the average expression of peptidases, enzymes and transmembrane receptors increased with age. The average expression of growth factors peak between Day-3 and Day-10, and decrease thereafter. We identified critical biological functions, upstream regulators, and putative transcription modules that seem to govern age-specific gene expression. We also observed differential ontogenic expression of known splicing variants of certain genes, and 1,455 novel splicing isoform candidates. In conclusion, the hepatic ontogeny of the transcriptome ontogeny has unveiled critical networks and up-stream regulators that orchestrate age-specific biological functions in liver, and suggest that age contributes to the complexity of the alternative splicing landscape of the hepatic transcriptome.

Stage-specific effects of Notch activation during skeletal myogenesis

PubMed Central

Bi, Pengpeng; Yue, Feng; Sato, Yusuke; Wirbisky, Sara; Liu, Weiyi; Shan, Tizhong; Wen, Yefei; Zhou, Daoguo; Freeman, Jennifer; Kuang, Shihuan

2016-01-01

Skeletal myogenesis involves sequential activation, proliferation, self-renewal/differentiation and fusion of myogenic stem cells (satellite cells). Notch signaling is known to be essential for the maintenance of satellite cells, but its function in late-stage myogenesis, i.e. post-differentiation myocytes and post-fusion myotubes, is unknown. Using stage-specific Cre alleles, we uncovered distinct roles of Notch1 in mononucleated myocytes and multinucleated myotubes. Specifically, constitutive Notch1 activation dedifferentiates myocytes into Pax7 quiescent satellite cells, leading to severe defects in muscle growth and regeneration, and postnatal lethality. By contrast, myotube-specific Notch1 activation improves the regeneration and exercise performance of aged and dystrophic muscles. Mechanistically, Notch1 activation in myotubes upregulates the expression of Notch ligands, which modulate Notch signaling in the adjacent satellite cells to enhance their regenerative capacity. These results highlight context-dependent effects of Notch activation during myogenesis, and demonstrate that Notch1 activity improves myotube’s function as a stem cell niche. DOI: http://dx.doi.org/10.7554/eLife.17355.001 PMID:27644105

[Effect of Supportan on nutritional status and immune function of late-staged gastric cancer patients undergoing chemotherapy].

PubMed

Zhong, Hai-jun; Ying, Jie-er; Ma, Sheng-lin

2006-09-01

To evaluate the effect of Supportan, an enteral nutrition (EN) specific for tumor patients, on the nutritional status and immune function of late-staged gastric cancer patients undergoing chemotherapy. Sixty-six late-staged gastric cancer patients undergoing chemotherapy were randomly divided into EN group (n=33) and control group (n=33). During chemotherapy, the patients in EN group received Supportan and the patients in the control group received basic diet. On the 14th day before chemotherapy and after chemotherapy, nutritional status and cell immune indicators were evaluated. As for nutrition indicators, there were no significant differences in EN group before and after chemotherapy (P > 0.05). Total protein, hemoglobin, prealbumin and transferrin significantly decreased after chemotherapy compared with those before chemotherapy in the control group (P< 0.01). The levels of CD4(+), CD8(+) T cells and CD4/CD8 were significantly increased, and NK cells, serum levels of IL-1, IL-6 were significantly decreased after chemotherapy in EN group (P< 0.01). The levels of IL-6 and TNF-alpha were significantly higher after chemotherapy than those before chemotherapy in the control group(P< 0.01). Curative effects of immune nutrition in EN group were superior to that in the control group, however, the differences were not statistically significant. The incidences of nausea, vomiting and marrow inhibition in Supportan group was lower compared with those in the control group, but with no significant difference. Supportan can prevent malnutrition of the late-staged gastric cancer patients undergoing chemotherapy, and improve immune function and alleviate adverse effects of chemotherapy.

Microarray Detection Call Methodology as a Means to Identify and Compare Transcripts Expressed within Syncytial Cells from Soybean (Glycine max) Roots Undergoing Resistant and Susceptible Reactions to the Soybean Cyst Nematode (Heterodera glycines)

PubMed Central

Klink, Vincent P.; Overall, Christopher C.; Alkharouf, Nadim W.; MacDonald, Margaret H.; Matthews, Benjamin F.

2010-01-01

Background. A comparative microarray investigation was done using detection call methodology (DCM) and differential expression analyses. The goal was to identify genes found in specific cell populations that were eliminated by differential expression analysis due to the nature of differential expression methods. Laser capture microdissection (LCM) was used to isolate nearly homogeneous populations of plant root cells. Results. The analyses identified the presence of 13,291 transcripts between the 4 different sample types. The transcripts filtered down into a total of 6,267 that were detected as being present in one or more sample types. A comparative analysis of DCM and differential expression methods showed a group of genes that were not differentially expressed, but were expressed at detectable amounts within specific cell types. Conclusion. The DCM has identified patterns of gene expression not shown by differential expression analyses. DCM has identified genes that are possibly cell-type specific and/or involved in important aspects of plant nematode interactions during the resistance response, revealing the uniqueness of a particular cell population at a particular point during its differentiation process. PMID:20508855

Fluorescence microscopy study on the cytoskeletal displacements during sperm differentiation in the bush-cricket Tylopsis liliifolia (Fabricius) (Orthoptera: Tettigoniidae).

PubMed

Viscuso, Renata; Federico, Concetta; Saccone, Salvatore; Bonaccorsi, Bianca; Vitale, Danilo G M

2016-02-01

A study by fluorescence microscopy has been carried out on male gametes from testicular follicles, seminal vesicles, spermatophores, and seminal receptacles of the bush-cricket Tylopsis liliifolia, focusing the attention on localization and movements of F-actin and α-tubulin during sperm differentiation, since data in this respect are lacking in the Orthoptera. F-actin and α-tubulin positivity was detected in the testicular follicles, in particular at the bridges connecting spermatids of a same clone and around their nucleus, during the first differentiation stages. During the following differentiation stages in the testes, F-actin was found at one of the spermatid poles and then, during nucleus elongation, at the whole acrosomal region. A peculiar F-actin-positivity was found at the flagellum, more markedly immediately posterior to the nucleus, at the basal body region of the gametes from the testicular follicles and from the other examined districts. Other interesting data from our investigations concerns the α-tubulin displacements during the differentiation stages of the spermatid and a constant absence of α-tubulin-positivity where the centrioles are located. No positivity was also found for both α-tubulin and nuclear markers at the anterior region of the gamete, where the acrosomal wings are localized. Our results, compared with what is so far known in literature for the insects, lead us to assert that microfilaments and microtubules undergo gradual displacements, markedly in the testicular follicles, during the morphogenesis of the male gamete of T. liliifolia aimed to its organization and motility and probably also to its interaction with the female gamete. © 2015 Wiley Periodicals, Inc.

Troglitazone induces differentiation in Trypanosoma brucei

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

Denninger, Viola; Figarella, Katherine; Schoenfeld, Caroline

2007-05-15

Trypanosoma brucei, a protozoan parasite causing sleeping sickness, is transmitted by the tsetse fly and undergoes a complex lifecycle including several defined stages within the insect vector and its mammalian host. In the latter, differentiation from the long slender to the short stumpy form is induced by a yet unknown factor of trypanosomal origin. Here we describe that some thiazolidinediones are also able to induce differentiation. In higher eukaryotes, thiazolidinediones are involved in metabolism and differentiation processes mainly by binding to the intracellular receptor peroxisome proliferator activated receptor {gamma}. Our studies focus on the effects of troglitazone on bloodstream formmore » trypanosomes. Differentiation was monitored using mitochondrial markers (membrane potential, succinate dehydrogenase activity, inhibition of oxygen uptake by KCN, amount of cytochrome transcripts), morphological changes (Transmission EM and light microscopy), and transformation experiments (loss of the Variant Surface Glycoprotein coat and increase of dihydroliponamide dehydrogenase activity). To further investigate the mechanisms responsible for these changes, microarray analyses were performed, showing an upregulation of expression site associated gene 8 (ESAG8), a potential differentiation regulator.« less

NGF-conjugated iron oxide nanoparticles promote differentiation and outgrowth of PC12 cells

NASA Astrophysics Data System (ADS)

Marcus, M.; Skaat, H.; Alon, N.; Margel, S.; Shefi, O.

2014-12-01

The search for regenerative agents that promote neuronal differentiation and repair is of great importance. Nerve growth factor (NGF) which is an essential contributor to neuronal differentiation has shown high pharmacological potential for the treatment of central neurodegenerative diseases such as Alzheimer's and Parkinson's. However, growth factors undergo rapid degradation, leading to a short biological half-life. In our study, we describe a new nano-based approach to enhance the NGF activity resulting in promoted neuronal differentiation. We covalently conjugated NGF to iron oxide nanoparticles (NGF-NPs) and studied the effect of the novel complex on the differentiation of PC12 cells. We found that the NGF-NP treatment, at the same concentration as free NGF, significantly promoted neurite outgrowth and increased the complexity of the neuronal branching trees. Examination of neuronal differentiation gene markers demonstrated higher levels of expression in PC12 cells treated with the conjugated factor. By manipulating the NGF specific receptor, TrkA, we have demonstrated that NGF-NPs induce cell differentiation via the regular pathway. Importantly, we have shown that NGF-NPs undergo slower degradation than free NGF, extending their half-life and increasing NGF availability. Even a low concentration of conjugated NGF treatment has led to an effective response. We propose the use of the NGF-NP complex which has magnetic characteristics, also as a useful method to enhance NGF efficiency and activity, thus, paving the way for substantial neuronal repair therapeutics.The search for regenerative agents that promote neuronal differentiation and repair is of great importance. Nerve growth factor (NGF) which is an essential contributor to neuronal differentiation has shown high pharmacological potential for the treatment of central neurodegenerative diseases such as Alzheimer's and Parkinson's. However, growth factors undergo rapid degradation, leading to a short biological half-life. In our study, we describe a new nano-based approach to enhance the NGF activity resulting in promoted neuronal differentiation. We covalently conjugated NGF to iron oxide nanoparticles (NGF-NPs) and studied the effect of the novel complex on the differentiation of PC12 cells. We found that the NGF-NP treatment, at the same concentration as free NGF, significantly promoted neurite outgrowth and increased the complexity of the neuronal branching trees. Examination of neuronal differentiation gene markers demonstrated higher levels of expression in PC12 cells treated with the conjugated factor. By manipulating the NGF specific receptor, TrkA, we have demonstrated that NGF-NPs induce cell differentiation via the regular pathway. Importantly, we have shown that NGF-NPs undergo slower degradation than free NGF, extending their half-life and increasing NGF availability. Even a low concentration of conjugated NGF treatment has led to an effective response. We propose the use of the NGF-NP complex which has magnetic characteristics, also as a useful method to enhance NGF efficiency and activity, thus, paving the way for substantial neuronal repair therapeutics. Electronic supplementary information (ESI) available: Conjugation ratio determination and supplementary figures. See DOI: 10.1039/c4nr05193a

LIF inhibits osteoblast differentiation at least in part by regulation of HAS2 and its product hyaluronan.

PubMed

Falconi, Dominic; Aubin, Jane E

2007-08-01

LIF arrests osteogenesis in fetal rat calvaria cells in a differentiation stage-specific manner. Differential display identified HAS2 as a LIF-induced gene and its product, HA, modulated osteoblast differentiation similarly to LIF. Our data suggest that LIF arrests osteoblast differentiation by altering HA content of the extracellular matrix. Leukemia inhibitory factor (LIF) elicits both anabolic and catabolic effects on bone. We previously showed in the fetal rat calvaria (RC) cell system that LIF inhibits osteoblast differentiation at the late osteoprogenitor/early osteoblast stage. To uncover potential molecular mediators of this inhibitory activity, we used a positive-negative genome-wide differential display screen to identify LIF-induced changes in the developing osteoblast transcriptome. Although LIF signaling is active throughout the RC cell proliferation-differentiation sequence, only a relatively small number of genes, in several different functional clusters, are modulated by LIF specifically during the LIF-sensitive inhibitory time window. Based on their known and predicted functions, most of the LIF-regulated genes identified are plausible candidates to be involved in the LIF-induced arrest of osteoprogenitor differentiation. To test this hypothesis, we further analyzed the function of one of the genes identified, hyaluronan synthase 2 (HAS2), in the LIF-induced inhibition. Synthesis of hyaluronan (HA), the product of HAS enzymatic activity, was stimulated by LIF and mimicked the HAS2 expression profile, with highest expression in early/proliferative and late/maturing cultures and lowest levels in intermediate/late osteoprogenitor-early osteoblast cultures. Exogenously added high molecular weight HA, the product of HAS2, dose-dependently inhibited osteoblast differentiation, with pulse-treatment effective in the same differentiation stage-specific inhibitory window as seen with LIF. In addition, however, pulse treatment with HA in early cultures slightly increased bone nodule formation. Treatment with hyaluronidase, on the other hand, stimulated bone nodule formation in early cultures but caused a small dose-dependent inhibition of osteoblast differentiation in the LIF- and HA-sensitive late time window. Together the data suggest that osteoblast differentiation is acutely sensitive to HA levels and that LIF inhibits osteoblast development at least in part by stimulating high molecular weight HA synthesis through HAS2.

Identification of Secretory Odontoblasts Using DMP1-GFP Transgenic Mice

PubMed Central

Balic, Anamaria; Mina, Mina

2011-01-01

Terminal differentiation of odontoblasts from dental papilla is a long process involving several intermediate steps and changes in the transcriptional profile and expression of proteins secreted by cells in the odontoblast lineage. Transgenic mouse lines in which GFP expression is under the control of tissue-and stage specific promoters have provided powerful experimental tools for identification and isolation of cells at specific stages of differentiation along a lineage. Our previous studies showed utilization of pOBCol3.6GFP and pOBCol2.3GFP animals for identification of odontoblasts at early and late stages of polarization respectively. In the present study we used the DMP1-GFP transgenic animal as an experimental model to examine its expression during the differentiation of odontoblasts from progenitor cells in vivo and in vitro. Our observations showed that DMP1-GFP transgene is first activated in secretory/functional odontoblasts engaged in secretion of predentin and then transiently expressed at high levels in newly differentiated odontoblasts. Expression of DMP1-GFP was down-regulated in highly differentiated odontoblasts. The temporal and spatial pattern of expression of DMP1-GFP transgene closely mimics the expression of endogenous DMP1. This transgenic animal will facilitate studies of gene expression and biological functions in secretory/functional odontoblasts. PMID:21172466

[The development of gender identity beyond rigid dichotomy].

PubMed

Quindeau, Ilka

2014-01-01

The conflicts individuals with ambiguous sexual characteristics suffer from are not the result of genetic features but of the rigid and dichotomous gender order, which is currently undergoing a renaissance. This also applies to individuals with an uncertain gender identity. In the best interests of the child a concept of gender seems necessary, that goes beyond a binary separation and allows gender-specific intermediary stages in the personal development of identity. Such a gender concept can be developed following psychoanalytic theories. The present discourse contains a scale of connecting factors for a differentiated and less normative conceptualization of gender development. Starting from Freud's concept of constitutional bisexuality, Robert Stoller's theory, which has been firmly rooted in the mainstream of psychoanalysis for more than 40 years, will be critically reviewed. By involving Reimut Reiche's and Jean Laplanche's arguments, a continuative psychological gender theory will be drafted, which does not normatively and reductively claim the demarcation of gender, but rather opens up a space for gender diversity.

Myeloblastic leukemia cells conditionally blocked by myc-estrogen receptor chimeric transgenes for terminal differentiation coupled to growth arrest and apoptosis.

PubMed

Selvakumaran, M; Liebermann, D; Hoffman-Liebermann, B

1993-05-01

Conditional mutants of the myeloblastic leukemic M1 cell line, expressing the chimeric mycer transgene, have been established. It is shown that M1 mycer cells, like M1, undergo terminal differentiation coupled to growth arrest and programmed cell death (apoptosis) after treatment with the physiologic differentiation inducer interleukin-6. However, when beta-estradiol is included in the culture medium, M1 mycer cells respond to differentiation inducers like M1 myc cell lines, where the differentiation program is blocked at an intermediate stage. By manipulating the function of the mycer transgene product, it is shown that there is a 10-hour window during myeloid differentiation, from 30 to 40 hours after the addition of the differentiation inducer, when the terminal differentiation program switches from being dependent on c-myc suppression to becoming c-myc suppression independent, where activation of c-myc has no apparent effect on mature macrophages. M1 mycer cell lines provide a powerful tool to increase our understanding of the role of c-myc in normal myelopoiesis and in leukemogenesis, also providing a strategy to clone c-myc target genes.

Thermal characterization of Titan's tholins by simultaneous TG-MS, DTA, DSC analysis

NASA Astrophysics Data System (ADS)

Nna-Mvondo, Delphine; de la Fuente, José L.; Ruiz-Bermejo, Marta; Khare, Bishun; McKay, Christopher P.

2013-09-01

Three samples of Titan's tholins synthesized in laboratory under simulated Titan's conditions and presenting different degrees of exposure to ambient atmosphere have been used to study in detail their thermal behavior using thermogravimetry coupled with a mass spectrometer (TG-MS), differential thermal analysis (DTA) and differential scanning calorimetry (DSC). The degradation of Titan's tholins under inert atmosphere follows a three-step consecutive decomposition: a drying stage (>150 °C) where moisture is desorbed, this stage indicated the high hydrophilicity of the tholins; a second stage, the main pyrolysis stage (150-575 °C) where endothermic decomposition begins releasing mainly ammonia, HCN, acetonitrile, and methane over a broad temperature range. Few other hydrocarbon fragments such as ethylene and propane are released but no cyclic molecules, aliphatic or aromatic, are observed. The last stage (>575 °C) is the carbonization of the material leading to a non-crystalline graphitic residue. The thermal degradation under oxygen atmosphere shows the same stages as in argon, with a shift of the thermogravimetric peaks toward lower temperatures indicating a lower thermal stability. The last stage in this case is an oxidative combustion of the char residue. This research concludes that even if Titan tholins, subjected to air contamination for few minutes to several years (varying with the storage conditions) transform to produce different C/N and C/O ratios and thermal stabilities, they undergo the same thermal degradation phases and products. This suggests that the studied three tholins have a similar main chemical structure which does not alter by the air exposure. We discuss on the possible nature of this structure.

Notch/Delta signaling constrains reengineering of pro-T cells by PU.1

PubMed Central

Franco, Christopher B.; Scripture-Adams, Deirdre D.; Proekt, Irina; Taghon, Tom; Weiss, Angela H.; Yui, Mary A.; Adams, Stephanie L.; Diamond, Rochelle A.; Rothenberg, Ellen V.

2006-01-01

PU.1 is essential for early stages of mouse T cell development but antagonizes it if expressed constitutively. Two separable mechanisms are involved: attenuation and diversion. Dysregulated PU.1 expression inhibits pro-T cell survival, proliferation, and passage through β-selection by blocking essential T cell transcription factors, signaling molecules, and Rag gene expression, which expression of a rearranged T cell antigen receptor transgene cannot rescue. However, Bcl2 transgenic cells are protected from this attenuation and may even undergo β-selection, as shown by PU.1 transduction of defined subsets of Bcl2 transgenic fetal thymocytes with differentiation in OP9-DL1 and OP9 control cultures. The outcome of PU.1 expression in these cells depends on Notch/Delta signaling. PU.1 can efficiently divert thymocytes toward a myeloid-like state with multigene regulatory changes, but Notch/Delta signaling vetoes diversion. Gene expression analysis distinguishes sets of critical T lineage regulatory genes with different combinatorial responses to PU.1 and Notch/Delta signals, suggesting particular importance for inhibition of E proteins, Myb, and/or Gfi1 (growth factor independence 1) in diversion. However, Notch signaling only protects against diversion of cells that have undergone T lineage specification after Thy-1 and CD25 up-regulation. The results imply that in T cell precursors, Notch/Delta signaling normally acts to modulate and channel PU.1 transcriptional activities during the stages from T lineage specification until commitment. PMID:16880393

Cell wall proteomics of the green alga Haematococcus pluvialis (Chlorophyceae).

PubMed

Wang, Sheng-Bing; Hu, Qiang; Sommerfeld, Milton; Chen, Feng

2004-03-01

The green microalga Haematococcus pluvialis can synthesize and accumulate large amounts of the ketocarotenoid astaxanthin, and undergo profound changes in cell wall composition and architecture during the cell cycle and in response to environmental stresses. In this study, cell wall proteins (CWPs) of H. pluvialis were systematically analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) coupled with peptide mass fingerprinting (PMF) and sequence-database analysis. In total, 163 protein bands were analyzed, which resulted in positive identification of 81 protein orthologues. The highly complex and dynamic composition of CWPs is manifested by the fact that the majority of identified CWPs are differentially expressed at specific stages of the cell cycle along with a number of common wall-associated 'housekeeping' proteins. The detection of cellulose synthase orthologue in the vegetative cells suggested that the biosynthesis of cellulose occurred during primary wall formation, in contrast to earlier observations that cellulose was exclusively present in the secondary wall of the organism. A transient accumulation of a putative cytokinin oxidase at the early stage of encystment pointed to a possible role in cytokinin degradation while facilitating secondary wall formation and/or assisting in cell expansion. This work represents the first attempt to use a proteomic approach to investigate CWPs of microalgae. The reference protein map constructed and the specific protein markers obtained from this study provide a framework for future characterization of the expression and physiological functions of the proteins involved in the biogenesis and modifications in the cell wall of Haematococcus and related organisms.

Differential receipt of sentinel lymph node biopsy within practice-based research networks

PubMed Central

Meyer, Anne-Marie; Reeder-Hayes, Katherine E.; Liu, Huan; Wheeler, Stephanie B.; Penn, Dolly; Weiner, Bryan J.; Carpenter, William R.

2013-01-01

Background Provider-based research networks (PBRNs) are promising for accelerating not only research, but also dissemination of research-based evidence into broader community practice. Sentinel lymph node biopsy (SLNB) is an innovation in breast cancer care associated with equivalent survival and lower morbidity, as compared to standard axillary lymph node dissection. We examined the diffusion of SLNB into practice and whether affiliation with the Community Clinical Oncology Program (CCOP), a cancer-focused PBRN, was associated with more rapid uptake of SLNB. Research Design Surveillance Epidemiology and End Results(SEER)-Medicare data were used to study women diagnosed with stage I or II breast cancer in the years 2000 to 2005 and undergoing breast conserving surgery with axillary staging (n=6,226). The primary outcome was undergoing SLNB. CCOP affiliation of the surgical physician was ascertained from NCI records. Multivariable generalized linear modeling with generalized estimating equations was used to measure association between CCOP exposure and undergoing SLNB, controlling for potential confounders. Results Women treated by a CCOP physician had significantly higher odds of receiving SLNB compared to women treated by a non-CCOP physician (OR 2.68; 95% CI 1.35, 5.34). The magnitude of this association was larger than that observed among patients treated by physicians operating in medical school-affiliated hospitals (OR 1.76; 95% CI 1.30–2.39). Conclusion Women treated by CCOP-affiliated physicians were more likely to undergo SLNB irrespective of the hospital’s medical school affiliation, suggesting that the CCOP PBRN may play a role in the rapid adoption of research-based innovation in community practice. PMID:23942221

Assessment of cerebral microbleeds by susceptibility-weighted imaging at 3T in patients with end-stage organ failure.

PubMed

Sparacia, Gianvincenzo; Cannella, Roberto; Lo Re, Vincenzina; Gambino, Angelo; Mamone, Giuseppe; Miraglia, Roberto

2018-02-17

Cerebral microbleeds (CMBs) are small rounded lesions representing cerebral hemosiderin deposits surrounded by macrophages that results from previous microhemorrhages. The aim of this study was to review the distribution of cerebral microbleeds in patients with end-stage organ failure and their association with specific end-stage organ failure risk factors. Between August 2015 and June 2017, we evaluated 15 patients, 9 males, and 6 females, (mean age 65.5 years). Patients population was subdivided into three groups according to the organ failure: (a) chronic kidney failure (n = 8), (b) restrictive cardiomyopathy undergoing heart transplantation (n = 1), and (c) end-stage liver failure undergoing liver transplantation (n = 6). The MR exams were performed on a 3T MR unit and the SWI sequence was used for the detection of CMBs. CMBs were subdivided in supratentorial lobar distributed, supratentorial non-lobar distributed, and infratentorial distributed. A total of 91 microbleeds were observed in 15 patients. Fifty-nine CMBs lesions (64.8%) had supratentorial lobar distribution, 17 CMBs lesions (18.8%) had supratentorial non-lobar distribution and the remaining 15 CMBs lesions (16.4%) were infratentorial distributed. An overall predominance of supratentorial multiple lobar localizations was found in all types of end-stage organ failure. The presence of CMBs was significantly correlated with age, hypertension, and specific end-stage organ failure risk factors (p < 0.001). CMBs are mostly founded in supratentorial lobar localization in end-stage organ failure. The improved detection of CMBs with SWI sequences may contribute to a more accurate identification of patients with cerebral risk factors to prevent complications during or after the organ transplantation.

When Genomes Collide: Aberrant Seed Development Following Maize Interploidy Crosses

PubMed Central

Pennington, Paul D.; Costa, Liliana M.; Gutierrez-Marcos, Jose F.; Greenland, Andy J.; Dickinson, Hugh G.

2008-01-01

Background and Aims The results of wide- or interploidy crosses in angiosperms are unpredictable and often lead to seed abortion. The consequences of reciprocal interploidy crosses have been explored in maize in detail, focusing on alterations to tissue domains in the maize endosperm, and changes in endosperm-specific gene expression. Methods Following reciprocal interploidy crosses between diploid and tetraploid maize lines, development of endosperm domains was studied using GUS reporter lines, and gene expression in resulting kernels was investigated using semi-quantitative RT-PCR on endosperms isolated at different stages of development. Key Results Reciprocal interploidy crosses result in very small, largely infertile seeds with defective endosperms. Seeds with maternal genomic excess are smaller than those with paternal genomic excess, their endosperms cellularize earlier and they accumulate significant quantities of starch. Endosperms from the reciprocal cross undergo an extended period of cell proliferation, and accumulate little starch. Analysis of reporter lines and gene expression studies confirm that functional domains of the endosperm are severely disrupted, and are modified differently according to the direction of the interploidy cross. Conclusions Interploidy crosses affect factors which regulate the balance between cell proliferation and cell differentiation within the endosperm. In particular, unbalanced crosses in maize affect transfer cell differentiation, and lead to the temporal deregulation of the ontogenic programme of endosperm development. PMID:18276791

Trends in Relative Mortality Between Hispanic and Non-Hispanic Whites Initiating Dialysis: A Retrospective Study of the US Renal Data System

PubMed Central

Arce, Cristina M.; Goldstein, Benjamin A.; Mitani, Aya A.; Winkelmayer, Wolfgang C.

2014-01-01

Background Hispanic patients undergoing long-term dialysis experience better survival compared with non-Hispanic whites. It is unknown whether this association differs by age, has changed over time, or is due to differential access to kidney transplantation. Study Design National retrospective cohort study. Setting & Participants Using the US Renal Data System, we identified 615,618 white patients 18 years or older who initiated dialysis therapy between January 1, 1995, and December 31, 2007. Predictors Hispanic ethnicity (vs non-Hispanic whites), year of end-stage renal disease incidence, age (as potential effect modifier). Outcomes All-cause and cause-specific mortality. Results We found that Hispanics initiating dialysis therapy experienced lower mortality, but age modified this association (P < 0.001). Compared with non-Hispanic whites, mortality in Hispanics was 33% lower at ages 18–39 years (adjusted cause-specific HR [HRcs], 0.67; 95% CI, 0.64–0.71) and 40–59 years (HRcs, 0.67; 95% CI, 0.66–0.68), 19% lower at ages 60–79 years (HRcs, 0.81; 95% CI, 0.80–0.82), and 6% lower at 80 years or older (HRcs, 0.94; 95% CI, 0.91–0.97). Accounting for the differential rates of kidney transplantation, the associations were attenuated markedly in the younger age strata; the survival benefit for Hispanics was reduced from 33% to 10% at ages 18–39 years (adjusted subdistribution-specific HR [HRsd], 0.90; 95% CI, 0.85–0.94) and from 33% to 19% among those aged 40–59 years (HRsd, 0.81; 95% CI, 0.80–0.83). Limitations Inability to analyze Hispanic subgroups that may experience heterogeneous mortality outcomes. Conclusions Overall, Hispanics experienced lower mortality, but differential access to kidney transplantation was responsible for much of the apparent survival benefit noted in younger Hispanics. Am J Kidney Dis. 62(2):312–321. PMID:23647836

Glycogen serves as an energy source that maintains astrocyte cell proliferation in the neonatal telencephalon.

PubMed

Gotoh, Hitoshi; Nomura, Tadashi; Ono, Katsuhiko

2017-06-01

Large amounts of energy are required when cells undergo cell proliferation and differentiation for mammalian neuronal development. Early neonatal mice face transient starvation and use stored energy for survival or to support development. Glycogen is a branched polysaccharide that is formed by glucose, and serves as an astrocytic energy store for rapid energy requirements. Although it is present in radial glial cells and astrocytes, the role of glycogen during development remains unclear. In the present study, we demonstrated that glycogen accumulated in glutamate aspartate transporter (GLAST)+ astrocytes in the subventricular zone and rostral migratory stream. Glycogen levels markedly decreased after birth due to the increase of glycogen phosphorylase, an essential enzyme for glycogen metabolism. In primary cultures and in vivo, the inhibition of glycogen phosphorylase decreased the proliferation of astrocytic cells. The number of cells in the G1 phase increased in combination with the up-regulation of cyclin-dependent kinase inhibitors or down-regulation of the phosphorylation of retinoblastoma protein (pRB), a determinant for cell cycle progression. These results suggest that glycogen accumulates in astrocytes located in specific areas during the prenatal stage and is used as an energy source to maintain normal development in the early postnatal stage.

Egr-5 is a post-mitotic regulator of planarian epidermal differentiation

PubMed Central

Tu, Kimberly C; Cheng, Li-Chun; TK Vu, Hanh; Lange, Jeffrey J; McKinney, Sean A; Seidel, Chris W; Sánchez Alvarado, Alejandro

2015-01-01

Neoblasts are an abundant, heterogeneous population of adult stem cells (ASCs) that facilitate the maintenance of planarian tissues and organs, providing a powerful system to study ASC self-renewal and differentiation dynamics. It is unknown how the collective output of neoblasts transit through differentiation pathways to produce specific cell types. The planarian epidermis is a simple tissue that undergoes rapid turnover. We found that as epidermal progeny differentiate, they progress through multiple spatiotemporal transition states with distinct gene expression profiles. We also identified a conserved early growth response family transcription factor, egr-5, that is essential for epidermal differentiation. Disruption of epidermal integrity by egr-5 RNAi triggers a global stress response that induces the proliferation of neoblasts and the concomitant expansion of not only epidermal, but also multiple progenitor cell populations. Our results further establish the planarian epidermis as a novel paradigm to uncover the molecular mechanisms regulating ASC specification in vivo. DOI: http://dx.doi.org/10.7554/eLife.10501.001 PMID:26457503

Work-Family Conflict: An Exploration of the Differential Effects of a Dependent Childs Age on Working Parents

ERIC Educational Resources Information Center

Darcy, Colette; McCarthy, Alma

2007-01-01

Purpose--The purpose of this article is to explore the impact of life cycle stage, specifically parenting stage, on work-family conflict among working parents to determine whether discernible differences are evident among those individuals at the early stage of their parenting cycle compared with those with older children.…

Clinical presentation and initial management of Black men and White men with prostate cancer in the United Kingdom: the PROCESS cohort study

PubMed Central

Evans, S; Metcalfe, C; Patel, B; Ibrahim, F; Anson, K; Chinegwundoh, F; Corbishley, C; Gillatt, D; Kirby, R; Muir, G; Nargund, V; Popert, R; Wilson, P; Persad, R; Ben-Shlomo, Y

2009-01-01

Background: In the United States, Black men have a higher risk of prostate cancer and worse survival than do White men, but it is unclear whether this is because of differences in diagnosis and management. We re-examined these differences in the United Kingdom, where health care is free and unlikely to vary by socioeconomic status. Methods: This study is a population-based retrospective cohort study of men diagnosed with prostate cancer with data on ethnicity, prognostic factors, and clinical care. A Delphi panel considered the appropriateness of investigations and treatments received. Results: At diagnosis, Black men had similar clinical stage and Gleason scores but higher age-adjusted prostate-specific antigen levels (geometric mean ratio 1.41, 95% confidence interval (95% CI): 1.15–1.73). Black men underwent more investigations and were more likely to undergo radical treatment, although this was largely explained by their younger age. Even after age adjustment, Black men were more likely to undergo a bone scan (odds ratio 1.37, 95% CI: 1.05–1.80). The Delphi analysis did not suggest differential management by ethnicity. Conclusions: This UK-based study comparing Black men with White men found no evidence of differences in disease characteristics at the time of prostate cancer diagnosis, nor of under-investigation or under-treatment in Black men. PMID:19935788

Disparities in staging prostate magnetic resonance imaging utilization for nonmetastatic prostate cancer patients undergoing definitive radiation therapy.

PubMed

Ajayi, Ayobami; Hwang, Wei-Ting; Vapiwala, Neha; Rosen, Mark; Chapman, Christina H; Both, Stefan; Shah, Meera; Wang, Xingmei; Agawu, Atu; Gabriel, Peter; Christodouleas, John; Tochner, Zelig; Deville, Curtiland

2016-01-01

There is growing evidence supporting incorporating multiparametric (mp) magnetic resonance imaging (MRI) scans into risk stratification, active surveillance, and treatment paradigms for prostate cancer. The purpose of our study was to determine whether demographic disparities exist in staging MRI utilization for prostate cancer patients. An institutional database of 705 nonmetastatic prostate cancer patients treated with radiation therapy from 2005 through 2013 was used to identify patients undergoing versus not undergoing pretreatment diagnostic prostate mpMRI. Uni- and multivariable logistic regression evaluated the relationship of clinical and demographic characteristics with MRI utilization. All demographic variables assessed, except the other race category, were significantly associated with MRI utilization (all P < .05), including age (odds ratio [OR], 0.92), black race (OR, 0.51), poverty (OR, 0.53), closer distance to radiation facility (OR, 1.79), and nonprivate primary insurance (OR, 0.57) on univariable analysis, while clinical stage T3 (OR, 3.37) was the only clinical characteristic. On multivariable analysis stratified by D'Amico risk group, age remained significant across all risk groups, whereas the black versus white racial (OR, 0.21; 95% confidence interval, 0.08-0.55) and nonprivate versus private insurance type (OR, 0.37; 95% confidence interval, 0.16-0.86) disparities persisted in the low-risk group. Clinical stage T3 remained associated in the high-risk group. For race specifically, the percentages of whites, blacks, and others undergoing MRI in the overall cohort and by risk group were, respectively: overall, 80% (343/427), 68% (156/231), and 85% (40/47); low risk, 86%, 56%, and 63%; intermediate risk, 79%, 72%, and 95%; and high risk, 72%, 72%, and 100%. In this urban, academic center cohort, older patients across all risk groups and black or nonprivate insurance patients in the low risk group were less likely to undergo staging prostate MRI scans. Further research should investigate these differences to ensure equitable utilization across all demographic groups considering the burden of prostate cancer disparities.

Ucma--A novel secreted factor represents a highly specific marker for distal chondrocytes.

PubMed

Tagariello, Andreas; Luther, Julia; Streiter, Melanie; Didt-Koziel, Lydia; Wuelling, Manuela; Surmann-Schmitt, Cordula; Stock, Michael; Adam, Nadia; Vortkamp, Andrea; Winterpacht, Andreas

2008-01-01

Growth and development of most parts of the vertebrate skeleton takes place by endochondral ossification, a process during which chondrocytes undergo distinct stages of differentiation resulting in a successive replacement of the cartilage anlagen by bone. In the context of an EST project we isolated a novel transcript from a human fetal growth plate cartilage cDNA library. The transcript which we called Ucma (unique cartilage matrix-associated protein) encodes a short protein of 138 amino acids. The protein sequence is evolutionary conserved throughout vertebrates and comprises a signal peptide, a coiled-coil domain, and a putative dibasic cleavage site for proprotein convertases. Using RNA in situ hybridization and immunohistochemistry with a polyclonal anti-Ucma antibody we found high expression of Ucma uniquely in distal (resting) chondrocytes in developing long bones of wildtype mice. This restricted expression could also be observed in Ihh(-/-), Ihh(-/-); Gli3(-/-), Gli3(-/-) mice, and in mice that overexpress Ihh under the control of the Col2a1 promoter indicating that expression of Ucma is regulated independent of hedgehog signaling. During insulin-induced differentiation of ATDC5 cells we found gradual increase of Ucma expression at day 21 with a maximum at day 24 and a decrease correlating with a simultaneous increase in the expression of cartilage link protein (Crtl1), a protein with maximum expression in column-forming proliferating chondrocytes. The present data strongly suggest an important function of Ucma in the early phase of chondrocyte differentiation.

Differential sensitivity of Glioma stem cells to Aurora kinase A inhibitors: implications for stem cell mitosis and centrosome dynamics.

PubMed

Mannino, Mariella; Gomez-Roman, Natividad; Hochegger, Helfrid; Chalmers, Anthony J

2014-07-01

Glioma stem-cell-like cells are considered to be responsible for treatment resistance and tumour recurrence following chemo-radiation in glioblastoma patients, but specific targets by which to kill the cancer stem cell population remain elusive. A characteristic feature of stem cells is their ability to undergo both symmetric and asymmetric cell divisions. In this study we have analysed specific features of glioma stem cell mitosis. We found that glioma stem cells appear to be highly prone to undergo aberrant cell division and polyploidization. Moreover, we discovered a pronounced change in the dynamic of mitotic centrosome maturation in these cells. Accordingly, glioma stem cell survival appeared to be strongly dependent on Aurora A activity. Unlike differentiated cells, glioma stem cells responded to moderate Aurora A inhibition with spindle defects, polyploidization and a dramatic increase in cellular senescence, and were selectively sensitive to Aurora A and Plk1 inhibitor treatment. Our study proposes inhibition of centrosomal kinases as a novel strategy to selectively target glioma stem cells. Copyright © 2014 The Authors. Published by Elsevier B.V. All rights reserved.

Stage- and subunit-specific functions of polycomb repressive complex 2 in bladder urothelial formation and regeneration

PubMed Central

Guo, Chunming; Balsara, Zarine R.

2017-01-01

ABSTRACT Urothelium is the protective lining of the urinary tract. The mechanisms underlying urothelial formation and maintenance are largely unknown. Here, we report the stage-specific roles of PRC2 epigenetic regulators in embryonic and adult urothelial progenitors. Without Eed, the obligatory subunit of PRC2, embryonic urothelial progenitors demonstrate reduced proliferation with concomitant dysregulation of genes including Cdkn2a (p16), Cdkn2b (p15) and Shh. These mutants display premature differentiation of keratin 5-positive (Krt5+) basal cells and ectopic expression of squamous-like differentiation markers. Deletion of Ezh2, the major enzymatic component of PRC2, causes upregulation of Upk3a+ superficial cells. Unexpectedly, Eed and Eed/Ezh2 double mutants exhibit delayed superficial cell differentiation. Furthermore, Eed regulates the proliferative and regenerative capacity of adult urothelial progenitors and prevents precocious differentiation. Collectively, these findings uncover the epigenetic mechanism by which PRC2 controls urothelial progenitor cell fate and the timing of differentiation, and further suggest an epigenetic basis of urothelial maintenance and regeneration. PMID:28049658

The neoteny-helper hypothesis: When to expect and when not to expect endocrine mechanisms to regulate allo-parental care?

PubMed

Schradin, Carsten; Vuarin, Pauline; Rimbach, Rebecca

2018-05-02

Family groups with helpers occur in several species of fish, birds and mammals. In such cooperatively breeding species all group members help with raising the offspring, i.e. parents and offspring from previous litters. While the ecological reasons and ultimate consequences of allo-parental care have been studied in detail, we know little about its physiological regulation. We propose three alternative hypotheses for the endocrine regulation of allo-parental care. 1. The neoteny-helper hypothesis predicts that helpers that did not undergo adolescence yet show helping behavior without any endocrine mechanisms activating it, as helping is the default response towards infant stimuli. The endocrine changes during adolescence would then deactivate helping behavior. 2. The parent-helper hypothesis predicts that helpers undergo the same endocrine changes as parents (increased prolactin and corticosterone levels; decreased testosterone in males but increased estrogen in females). We predict that this hypothesis is especially important in post-adolescent helpers. 3. The helper-specific hypothesis predicts that there are specific endocrine mechanisms that only exist in helpers but not in breeders. We review evidence for these three hypotheses in 23 species of fish, birds, and mammals. We found no evidence for the helper-specific hypothesis but for both other hypotheses. As predicted, this depended on whether helpers were pre- or post-adolescent, but information on whether or not helpers underwent adolescence was often missing. Thus, future studies should investigate whether or not helpers have reached sexual maturity, differentiate between pre- and post-adolescent helpers, and study behavioral changes in helping behavior during adolescence. We conclude that the neurobiological circuits in the brain necessary for allo-parental care might often be the default stage in helpers from cooperative breeding species, which might be deactivated by specific endocrine mechanisms during adolescence, and then would need reactivation for allo-parental and parental care. Copyright © 2017 Elsevier Inc. All rights reserved.

Diagnostic accuracy of apparent diffusion coefficient and 123I-metaiodobenzylguanidine for differentiation of multiple system atrophy and Parkinson's disease.

PubMed

Umemura, Atsushi; Oeda, Tomoko; Hayashi, Ryutaro; Tomita, Satoshi; Kohsaka, Masayuki; Yamamoto, Kenji; Sawada, Hideyuki

2013-01-01

It is often hard to differentiate Parkinson's disease (PD) and parkinsonian variant of multiple system atrophy (MSA-P), especially in the early stages. Cardiac sympathetic denervation and putaminal rarefaction are specific findings for PD and MSA-P, respectively. We investigated diagnostic accuracy of putaminal apparent diffusion coefficient (ADC) test for MSA-P and (123)I-metaiodobenzylguanidine (MIBG) scintigram for PD, especially in early-stage patients. The referral standard diagnosis of PD and MSA-P were the diagnostic criteria of the United Kingdom Parkinson's Disease Society Brain Bank Criteria and the second consensus criteria, respectively. Based on the referral standard criteria, diagnostic accuracy [area under the receiver-operator characteristic curve (AUC), sensitivity and specificity] of the ADC and MIBG tests was estimated retrospectively. Diagnostic accuracy of these tests performed within 3 years of symptom onset was also investigated. ADC and MIBG tests were performed on 138 patients (20 MSA and 118 PD). AUC was 0.95 and 0.83 for the ADC and MIBG tests, respectively. Sensitivity and specificity were 85.0% and 89.0% for MSA-P diagnosis by ADC test and 67.0% and 80.0% for PD diagnosis by MIBG test. When these tests were restricted to patients with disease duration ≤ 3 years, the sensitivity and specificity were 75.0% and 91.4% for the ADC test (MSA-P diagnosis) and 47.7% and 92.3% for the MIBG test (PD diagnosis). Both tests were useful in differentiating between PD and MSA-P, even in the early stages. In early-stage patients, elevated putaminal ADC was a diagnostic marker for MSA-P. Despite high specificity of the MIBG test, careful neurological history and examinations were required for PD diagnosis because of possible false-negative results.

Thoroughness of Mediastinal Staging in Stage IIIA Non-Small Cell Lung Cancer

PubMed Central

Vest, Michael T.; Tanoue, Lynn; Soulos, Pamela R.; Kim, Anthony W.; Detterbeck, Frank; Morgensztern, Daniel; Gross, Cary P.

2011-01-01

Introduction Guidelines recommend that patients with clinical stage IIIA non-small cell lung cancer (NSCLC) undergo histologic confirmation of pathologic lymph nodes. Studies have suggested that invasive mediastinal staging is underutilized, though practice patterns have not been rigorously evaluated. Methods We used the Surveillance, Epidemiology, and End Results-Medicare database to identify patients with stage IIIA NSCLC diagnosed from 1998 through 2005. Invasive staging and use of positron emission tomography (PET) scanning were assessed using Medicare claims. Multivariable logistic regression was used to identify patient characteristics associated with use of invasive staging. Results Of 7583 stage IIIA NSCLC patients, 1678 (22%) underwent invasive staging. Patients who received curative intent cancer treatment were more likely to undergo invasive staging than patients who did not receive cancer specific therapy (30% vs. 9.8%, adjusted odds ratio [OR} 3.31, 95% CI 2.78–3.95). The oldest patients (age 85–94) were less likely to receive invasive staging than the youngest ((age 67–69) (27.6 % vs. 11.9%, OR 0.46, 95% CI 0.34–0.61)). Sex, marital status, income and race were not associated with the use of the invasive staging. The use of invasive staging was stable throughout the study period, despite an increase in the use of PET scanning from less than 10% of patients prior to 2000 to almost 70% in 2005. Conclusion Nearly 80% of Medicare beneficiaries with stage IIIA NSCLC do not receive guideline adherent mediastinal staging; this failure cannot be entirely explained by patient factors or a reliance on PET imaging. Incentives to encourage use of invasive staging may improve care. PMID:22134069

Minimally Invasive Surgical Staging for Ovarian Carcinoma: A Propensity-Matched Comparison With Traditional Open Surgery.

PubMed

Ditto, Antonino; Bogani, Giorgio; Martinelli, Fabio; Signorelli, Mauro; Chiappa, Valentina; Scaffa, Cono; Indini, Alice; Leone Roberti Maggiore, Umberto; Lorusso, Domenica; Raspagliesi, Francesco

2017-01-01

Growing evidence supports the safety of a laparoscopic approach for patients affected by apparent early-stage ovarian cancer. However, no well-designed studies comparing laparoscopic and open surgical staging are available. In the present investigation we aimed to provide a balanced long-term comparison between these 2 approaches. Retrospective study (Canadian Task Force classification II-2). Tertiary center. Data of consecutive patients affected by early-stage ovarian cancer who had laparoscopic staging were matched 1:1 with a cohort of patients undergoing open surgical staging. The matching was conducted by a propensity-score comparison. Laparoscopic and open surgical staging. Fifty patient pairs (100 patients: 50 undergoing laparoscopic staging vs 50 undergoing open surgical staging) were included. Demographic and baseline oncologic characteristics were balanced between groups (p > .2). We observed that patients undergoing laparoscopic staging experienced longer operative time (207.2 [71.6] minutes vs 180.7 [47.0] minutes; p = .04), lower blood loss (150 [52.7] mL vs 339.8 [225.9] mL; p < .001), and shorter length of hospital stay (4.0 [2.6] days vs 6.1 [1.6] days; p < .001) compared with patients undergoing open surgical staging. No conversion to open surgery occurred. Complication rate was similar between groups. No difference in survival outcomes were observed, after a mean (SD) follow-up of 49.5 (64) and 52.6 (31.7) months after laparoscopic and open surgical staging, respectively. Our findings suggest that the implementation of minimally invasive staging does not influence survival outcomes of patients affected by early-stage ovarian cancer. Laparoscopic staging improved patient outcomes, reducing length of hospital stay. Further large prospective studies are warranted. Copyright © 2016 AAGL. Published by Elsevier Inc. All rights reserved.

Proteomic analysis of maize grain development using iTRAQ reveals temporal programs of diverse metabolic processes.

PubMed

Yu, Tao; Li, Geng; Dong, Shuting; Liu, Peng; Zhang, Jiwang; Zhao, Bin

2016-11-04

Grain development in maize is an essential process in the plant's life cycle and is vital for use of the plant as a crop for animals and humans. However, little is known regarding the protein regulatory networks that control grain development. Here, isobaric tag for relative and absolute quantification (iTRAQ) technology was used to analyze temporal changes in protein expression during maize grain development. Maize grain proteins and changes in protein expression at eight developmental stages from 3 to 50 d after pollination (DAP) were performed using iTRAQ-based proteomics. Overall, 4751 proteins were identified; 2639 of these were quantified and 1235 showed at least 1.5-fold changes in expression levels at different developmental stages and were identified as differentially expressed proteins (DEPs). The DEPs were involved in different cellular and metabolic processes with a preferential distribution to protein synthesis/destination and metabolism categories. A K-means clustering analysis revealed coordinated protein expression associated with different functional categories/subcategories at different development stages. Our results revealed developing maize grain display different proteomic characteristics at distinct stages, such as numerous DEPs for cell growth/division were highly expressed during early stages, whereas those for starch biosynthesis and defense/stress accumulated in middle and late stages, respectively. We also observed coordinated expression of multiple proteins of the antioxidant system, which are essential for the maintenance of reactive oxygen species (ROS) homeostasis during grain development. Particularly, some DEPs, such as zinc metallothionein class II, pyruvate orthophosphate dikinase (PPDK) and 14-3-3 proteins, undergo major changes in expression at specific developmental stages, suggesting their roles in maize grain development. These results provide a valuable resource for analyzing protein function on a global scale and also provide new insights into the potential protein regulatory networks that control grain yield and quality.

Different tissue phagocytes sample apoptotic cells to direct distinct homeostasis programs.

PubMed

Cummings, Ryan J; Barbet, Gaetan; Bongers, Gerold; Hartmann, Boris M; Gettler, Kyle; Muniz, Luciana; Furtado, Glaucia C; Cho, Judy; Lira, Sergio A; Blander, J Magarian

2016-11-24

Recognition and removal of apoptotic cells by professional phagocytes, including dendritic cells and macrophages, preserves immune self-tolerance and prevents chronic inflammation and autoimmune pathologies. The diverse array of phagocytes that reside within different tissues, combined with the necessarily prompt nature of apoptotic cell clearance, makes it difficult to study this process in situ. The full spectrum of functions executed by tissue-resident phagocytes in response to homeostatic apoptosis, therefore, remains unclear. Here we show that mouse apoptotic intestinal epithelial cells (IECs), which undergo continuous renewal to maintain optimal barrier and absorptive functions, are not merely extruded to maintain homeostatic cell numbers, but are also sampled by a single subset of dendritic cells and two macrophage subsets within a well-characterized network of phagocytes in the small intestinal lamina propria. Characterization of the transcriptome within each subset before and after in situ sampling of apoptotic IECs revealed gene expression signatures unique to each phagocyte, including macrophage-specific lipid metabolism and amino acid catabolism, and a dendritic-cell-specific program of regulatory CD4 + T-cell activation. A common 'suppression of inflammation' signature was noted, although the specific genes and pathways involved varied amongst dendritic cells and macrophages, reflecting specialized functions. Apoptotic IECs were trafficked to mesenteric lymph nodes exclusively by the dendritic cell subset and served as critical determinants for the induction of tolerogenic regulatory CD4 + T-cell differentiation. Several of the genes that were differentially expressed by phagocytes bearing apoptotic IECs overlapped with susceptibility genes for inflammatory bowel disease. Collectively, these findings provide new insights into the consequences of apoptotic cell sampling, advance our understanding of how homeostasis is maintained within the mucosa and set the stage for development of novel therapeutics to alleviate chronic inflammatory diseases such as inflammatory bowel disease.

miR-203 modulates epithelial differentiation of human embryonic stem cells towards epidermal stratification.

PubMed

Nissan, Xavier; Denis, Jérôme Alexandre; Saidani, Manoubia; Lemaitre, Gilles; Peschanski, Marc; Baldeschi, Christine

2011-08-15

The molecular mechanisms controlling the differentiation of human basal keratinocyte stem cells towards the epidermis are well characterized, whereas the earliest process leading to the specification of embryonic stem cells into keratinocytes is still not well understood. MicroRNAs are regulators of many cellular events, but evidence for microRNA acting on the differentiation of human embryonic stem cells into a specific lineage has been elusive. By using our recent protocol for obtaining functional keratinocytes from hESC, we attempted to analyze the role of microRNAs in the early stages of epidermal differentiation. Thus, we identified a set of 5 microRNAs, namely miR-200a, miR-200b, miR-203, miR-205 and miR-429, that are specifically overexpressed during the early stages of the differentiation process. Interestingly, our functional analyses revealed an instrumental role of miR-203, which had been previously shown to play a key role during the formation of the pluristratified epidermis by basal keratinocyte stem cells, in the early keratinocyte commitment. These results highlight the determinant and unique role of miR-203 during the entire process of epidermal development by extending its spectrum of action from the early commitment of embryonic stem cells to ultimate differentiation of the organ. Copyright © 2011 Elsevier Inc. All rights reserved.

Peptide-specific recognition of human cytomegalovirus strains controls adaptive natural killer cells.

PubMed

Hammer, Quirin; Rückert, Timo; Borst, Eva Maria; Dunst, Josefine; Haubner, André; Durek, Pawel; Heinrich, Frederik; Gasparoni, Gilles; Babic, Marina; Tomic, Adriana; Pietra, Gabriella; Nienen, Mikalai; Blau, Igor Wolfgang; Hofmann, Jörg; Na, Il-Kang; Prinz, Immo; Koenecke, Christian; Hemmati, Philipp; Babel, Nina; Arnold, Renate; Walter, Jörn; Thurley, Kevin; Mashreghi, Mir-Farzin; Messerle, Martin; Romagnani, Chiara

2018-05-01

Natural killer (NK) cells are innate lymphocytes that lack antigen-specific rearranged receptors, a hallmark of adaptive lymphocytes. In some people infected with human cytomegalovirus (HCMV), an NK cell subset expressing the activating receptor NKG2C undergoes clonal-like expansion that partially resembles anti-viral adaptive responses. However, the viral ligand that drives the activation and differentiation of adaptive NKG2C + NK cells has remained unclear. Here we found that adaptive NKG2C + NK cells differentially recognized distinct HCMV strains encoding variable UL40 peptides that, in combination with pro-inflammatory signals, controlled the population expansion and differentiation of adaptive NKG2C + NK cells. Thus, we propose that polymorphic HCMV peptides contribute to shaping of the heterogeneity of adaptive NKG2C + NK cell populations among HCMV-seropositive people.

Transcriptional diversity during lineage commitment of human blood progenitors.

PubMed

Chen, Lu; Kostadima, Myrto; Martens, Joost H A; Canu, Giovanni; Garcia, Sara P; Turro, Ernest; Downes, Kate; Macaulay, Iain C; Bielczyk-Maczynska, Ewa; Coe, Sophia; Farrow, Samantha; Poudel, Pawan; Burden, Frances; Jansen, Sjoert B G; Astle, William J; Attwood, Antony; Bariana, Tadbir; de Bono, Bernard; Breschi, Alessandra; Chambers, John C; Consortium, Bridge; Choudry, Fizzah A; Clarke, Laura; Coupland, Paul; van der Ent, Martijn; Erber, Wendy N; Jansen, Joop H; Favier, Rémi; Fenech, Matthew E; Foad, Nicola; Freson, Kathleen; van Geet, Chris; Gomez, Keith; Guigo, Roderic; Hampshire, Daniel; Kelly, Anne M; Kerstens, Hindrik H D; Kooner, Jaspal S; Laffan, Michael; Lentaigne, Claire; Labalette, Charlotte; Martin, Tiphaine; Meacham, Stuart; Mumford, Andrew; Nürnberg, Sylvia; Palumbo, Emilio; van der Reijden, Bert A; Richardson, David; Sammut, Stephen J; Slodkowicz, Greg; Tamuri, Asif U; Vasquez, Louella; Voss, Katrin; Watt, Stephen; Westbury, Sarah; Flicek, Paul; Loos, Remco; Goldman, Nick; Bertone, Paul; Read, Randy J; Richardson, Sylvia; Cvejic, Ana; Soranzo, Nicole; Ouwehand, Willem H; Stunnenberg, Hendrik G; Frontini, Mattia; Rendon, Augusto

2014-09-26

Blood cells derive from hematopoietic stem cells through stepwise fating events. To characterize gene expression programs driving lineage choice, we sequenced RNA from eight primary human hematopoietic progenitor populations representing the major myeloid commitment stages and the main lymphoid stage. We identified extensive cell type-specific expression changes: 6711 genes and 10,724 transcripts, enriched in non-protein-coding elements at early stages of differentiation. In addition, we found 7881 novel splice junctions and 2301 differentially used alternative splicing events, enriched in genes involved in regulatory processes. We demonstrated experimentally cell-specific isoform usage, identifying nuclear factor I/B (NFIB) as a regulator of megakaryocyte maturation-the platelet precursor. Our data highlight the complexity of fating events in closely related progenitor populations, the understanding of which is essential for the advancement of transplantation and regenerative medicine. Copyright © 2014, American Association for the Advancement of Science.

Ultrastructural aspects of pollen ontogeny in an endangered plant species, Pancratium maritimum L. (Amaryllidaceae).

PubMed

Tütüncü Konyar, Sevil

2017-03-01

Pollen ontogeny in Pancratium maritimum L. was studied from the sporogenous cell to mature pollen grain stages using transmission electron, scanning electron, and light microscopy to determine whether the pollen development in P. maritimum follows the basic scheme in angiosperms or not. In the course of microsporogenesis and microgametogenesis, special attention was given to the considerable ultrastructural changes that are observed in the cytoplasm of microsporocytes, microspores, and mature pollen grains throughout the successive stages of pollen development. Microsporocyte differentiation concerning number and ultrastructure of organelles facilitates the transition of microsporocytes from the sporophytic phase to the gametophytic phase. However, cytoplasmic differentiation of generative and vegetative cells supports their functional distinctness and pollen maturation. Although microsporogenesis and microgametogenesis in P. maritimum generally follow the usual angiosperm pattern, abnormalities such as formation of unreduced gametes were observed. During normal microsporogenesis, meiocytes undergo meiosis and successive cytokinesis, resulting in the formation of isobilateral, decussate, and linear tetrads. Subsequent to the development of free and vacuolated microspores, the first mitotic division occurs and bicellular monosulcate pollen grains are produced. Pollen grains are shed from the anther at binucleate stage. During pollen ontogeny, three periods of vacuolization were observed: in meiocytes, in mononucleate free microspores, and in the generative cell.

Continued Expression of GATA3 Is Necessary for Cochlear Neurosensory Development

PubMed Central

Duncan, Jeremy S.; Fritzsch, Bernd

2013-01-01

Hair cells of the developing mammalian inner ear are progressively defined through cell fate restriction. This process culminates in the expression of the bHLH transcription factor Atoh1, which is necessary for differentiation of hair cells, but not for their specification. Loss of several genes will disrupt ear morphogenesis or arrest of neurosensory epithelia development. We previously showed in null mutants that the loss of the transcription factor, Gata3, results specifically in the loss of all cochlear neurosensory development. Temporal expression of Gata3 is broad from the otic placode stage through the postnatal ear. It therefore remains unclear at which stage in development Gata3 exerts its effect. To better understand the stage specific effects of Gata3, we investigated the role of Gata3 in cochlear neurosensory specification and differentiation utilizing a LoxP targeted Gata3 line and two Cre lines. Foxg1Cre∶Gata3f/f mice show recombination of Gata3 around E8.5 but continue to develop a cochlear duct without differentiated hair cells and spiral ganglion neurons. qRT-PCR data show that Atoh1 was down-regulated but not absent in the duct whereas other hair cell specific genes such as Pou4f3 were completely absent. In addition, while Sox2 levels were lower in the Foxg1Cre:Gata3f/f cochlea, Eya1 levels remained normal. We conclude that Eya1 is unable to fully upregulate Atoh1 or Pou4f3, and drive differentiation of hair cells without Gata3. Pax2-Cre∶Gata3f/f mice show a delayed recombination of Gata3 in the ear relative to Foxg1Cre:Gata3f/f. These mice exhibited a cochlear duct containing patches of partially differentiated hair cells and developed only few and incorrectly projecting spiral ganglion neurons. Our conditional deletion studies reveal a major role of Gata3 in the signaling of prosensory genes and in the differentiation of cochlear neurosenory cells. We suggest that Gata3 may act in combination with Eya1, Six1, and Sox2 in cochlear prosensory gene signaling. PMID:23614009

Desmocollin 2 is a new immunohistochemical marker indicative of squamous differentiation in urothelial carcinoma.

PubMed

Hayashi, Tetsutaro; Sentani, Kazuhiro; Oue, Naohide; Anami, Katsuhiro; Sakamoto, Naoya; Ohara, Shinya; Teishima, Jun; Noguchi, Tsuyoshi; Nakayama, Hirofumi; Taniyama, Kiyomi; Matsubara, Akio; Yasui, Wataru

2011-10-01

Urothelial carcinoma (UC) with squamous differentiation tends to present at higher stages than pure UC. To distinguish UC with squamous differentiation from pure UC, a sensitive and specific marker is needed. Desmocollin 2 (DSC2) is a protein localized in desmosomal junctions of stratified epithelium, but little is known about its biological significance in bladder cancer. We examined the utility of DSC2 as a diagnostic marker. We analysed the immunohistochemical characteristics of DSC2, and studied the relationship of DSC2 expression with the expression of the known markers uroplakin III (UPIII), cytokeratin (CK)7, CK20, epidermal growth factor receptor (EGFR), and p53. DSC2 staining was detected in 24 of 25 (96%) cases of UC with squamous differentiation, but in none of 85 (0%) cases of pure UC. DSC2 staining was detected only in areas of squamous differentiation. DSC2 expression was mutually exclusive of UPIII expression, and was correlated with EGFR expression. Furthermore, DSC2 expression was correlated with higher stage (P = 0.0314) and poor prognosis (P = 0.0477). DSC2 staining offers high sensitivity (96%) and high specificity (100%) for the detection of squamous differentiation in UC. DSC2 is a useful immunohistochemical marker for separation of UC with squamous differentiation from pure UC. 2011 Blackwell Publishing Limited.

Lipidomic profiling of patient-specific iPSC-derived hepatocyte-like cells

PubMed Central

Viiri, Leena E.; Vihervaara, Terhi; Koistinen, Kaisa M.; Hilvo, Mika; Ekroos, Kim; Käkelä, Reijo; Aalto-Setälä, Katriina

2017-01-01

ABSTRACT Hepatocyte-like cells (HLCs) differentiated from human induced pluripotent stem cells (iPSCs) offer an alternative model to primary human hepatocytes to study lipid aberrations. However, the detailed lipid profile of HLCs is yet unknown. In the current study, functional HLCs were differentiated from iPSCs generated from dermal fibroblasts of three individuals by a three-step protocol through the definitive endoderm (DE) stage. In parallel, detailed lipidomic analyses as well as gene expression profiling of a set of lipid-metabolism-related genes were performed during the entire differentiation process from iPSCs to HLCs. Additionally, fatty acid (FA) composition of the cell culture media at different stages was determined. Our results show that major alterations in the molecular species of lipids occurring during DE and early hepatic differentiation stages mainly mirror the quality and quantity of the FAs supplied in culture medium at each stage. Polyunsaturated phospholipids and sphingolipids with a very long FA were produced in the cells at a later stage of differentiation. This work uncovers the previously unknown lipid composition of iPSC-HLCs and its alterations during the differentiation in conjunction with the expression of key lipid-associated genes. Together with biochemical, functional and gene expression measurements, the lipidomic analyses allowed us to improve our understanding of the concerted influence of the exogenous metabolite supply and cellular biosynthesis essential for iPSC-HLC differentiation and function. Importantly, the study describes in detail a cell model that can be applied in exploring, for example, the lipid metabolism involved in the development of fatty liver disease or atherosclerosis. PMID:28733363

Fibroblast growth factor receptors in in vitro and in vivo chondrogenesis: relating tissue engineering using adult mesenchymal stem cells to embryonic development.

PubMed

Hellingman, Catharine A; Koevoet, Wendy; Kops, Nicole; Farrell, Eric; Jahr, Holger; Liu, Wei; Baatenburg de Jong, Robert J; Frenz, Dorothy A; van Osch, Gerjo J V M

2010-02-01

Adult mesenchymal stem cells (MSCs) are considered promising candidate cells for therapeutic cartilage and bone regeneration. Because tissue regeneration and embryonic development may involve similar pathways, understanding common pathways may lead to advances in regenerative medicine. In embryonic limb development, fibroblast growth factor receptors (FGFRs) play a role in chondrogenic differentiation. The aim of this study was to investigate and compare FGFR expression in in vivo embryonic limb development and in vitro chondrogenesis of MSCs. Our study showed that in in vitro chondrogenesis of MSCs three sequential stages can be found, as in embryonic limb development. A mesenchymal condensation (indicated by N-cadherin) is followed by chondrogenic differentiation (indicated by collagen II), and hypertrophy (indicated by collagen X). FGFR1-3 are expressed in a stage-dependent pattern during in vitro differentiation and in vivo embryonic limb development. In both models FGFR2 is clearly expressed by cells in the condensation phase. No FGFR expression was observed in differentiating and mature hyaline chondrocytes, whereas hypertrophic chondrocytes stained strongly for all FGFRs. To evaluate whether stage-specific modulation of chondrogenic differentiation in MSCs is possible with different subtypes of FGF, FGF2 and FGF9 were added to the chondrogenic medium during different stages in the culture process (early or late). FGF2 and FGF9 differentially affected the amount of cartilage formed by MSCs depending on the stage in which they were added. These results will help us understand the role of FGF signaling in chondrogenesis and find new tools to monitor and control chondrogenic differentiation.

Impact of different variables on the outcome of patients with clinically confined prostate carcinoma: prediction of pathologic stage and biochemical failure using an artificial neural network.

PubMed

Ziada, A M; Lisle, T C; Snow, P B; Levine, R F; Miller, G; Crawford, E D

2001-04-15

The advent of advanced computing techniques has provided the opportunity to analyze clinical data using artificial intelligence techniques. This study was designed to determine whether a neural network could be developed using preoperative prognostic indicators to predict the pathologic stage and time of biochemical failure for patients who undergo radical prostatectomy. The preoperative information included TNM stage, prostate size, prostate specific antigen (PSA) level, biopsy results (Gleason score and percentage of positive biopsy), as well as patient age. All 309 patients underwent radical prostatectomy at the University of Colorado Health Sciences Center. The data from all patients were used to train a multilayer perceptron artificial neural network. The failure rate was defined as a rise in the PSA level > 0.2 ng/mL. The biochemical failure rate in the data base used was 14.2%. Univariate and multivariate analyses were performed to validate the results. The neural network statistics for the validation set showed a sensitivity and specificity of 79% and 81%, respectively, for the prediction of pathologic stage with an overall accuracy of 80% compared with an overall accuracy of 67% using the multivariate regression analysis. The sensitivity and specificity for the prediction of failure were 67% and 85%, respectively, demonstrating a high confidence in predicting failure. The overall accuracy rates for the artificial neural network and the multivariate analysis were similar. Neural networks can offer a convenient vehicle for clinicians to assess the preoperative risk of disease progression for patients who are about to undergo radical prostatectomy. Continued investigation of this approach with larger data sets seems warranted. Copyright 2001 American Cancer Society.

Preferential Hyperacuity Perimeter (PreView PHP) for detecting choroidal neovascularization study.

PubMed

Alster, Yair; Bressler, Neil M; Bressler, Susan B; Brimacombe, Judith A; Crompton, R Michael; Duh, Yi-Jing; Gabel, Veit-Peter; Heier, Jeffrey S; Ip, Michael S; Loewenstein, Anat; Packo, Kirk H; Stur, Michael; Toaff, Techiya

2005-10-01

To assess the ability of the Preferential Hyperacuity Perimeter (PreView PHP; Carl Zeiss Meditec, Dublin, CA) to detect recent-onset choroidal neovascularization (CNV) resulting from age-related macular degeneration (AMD) and to differentiate it from an intermediate stage of AMD. Prospective, comparative, concurrent, nonrandomized, multicenter study. Eligible participants' study eyes had a corrected visual acuity of 20/160 or better and either untreated CNV from AMD diagnosed within the last 60 days or an intermediate stage of AMD. After obtaining consent, visual acuity with habitual correction, masked PHP testing, stereoscopic color fundus photography, and fluorescein angiography were performed. Photographs and angiograms were evaluated by graders masked to diagnosis and PHP results. The reading center's diagnosis determined if the patient was categorized as having intermediate AMD or neovascular AMD. A successful study outcome was defined a priori as a sensitivity of at least 80% and a specificity of at least 80%. Of 185 patients who gave consent to be enrolled, 11 (6%) had PHP results judged to be unreliable. An additional 52 were not included because they did not meet all eligibility criteria. Of the remaining 122 patients, 57 had an intermediate stage of AMD and 65 had neovascular AMD. The sensitivity to detect newly diagnosed CNV using PHP testing was 82% (95% confidence interval [CI], 70%-90%). The specificity to differentiate newly diagnosed CNV from the intermediate stage of AMD using PHP testing was 88% (95% CI, 76%-95%). Preferential Hyperacuity Perimeter testing can detect recent-onset CNV resulting from AMD and can differentiate it from an intermediate stage of AMD with high sensitivity and specificity. These data suggest that monitoring with PHP should detect most cases of CNV of recent onset with few false-positive results at a stage when treatment usually would be beneficial. Thus, this monitoring should be considered in the management of the intermediate stage of AMD.

Sex-specific gene expression in early life stage fathead minnows (Pimephales promelas) throughout development and after exposure to synthetic hormones

EPA Science Inventory

There is evidence that exposure to endocrine disrupting chemicals (EDCs) during early life stages can alter sex differentiation in fishes. Fathead minnows (Pimephales promelas) are commonly used as a model fish species in endocrine disruption studies. However, limited knowledge...

Embryonic mouse pre-metatarsal development in organ culture

NASA Technical Reports Server (NTRS)

Klement, B. J.; Spooner, B. S.

1993-01-01

Embryonic mouse pre-metatarsals were removed from embryos at 13 days of gestation and cultured in a defined, serum-free medium for up to 15 days. By histological analysis, we observe that the cultured pre-metatarsal tissue undergoes a similar developmental profile as pre-metatarsals growing normally in vivo. The initial mesenchyme condensation regions undergo differentiation and morphogenesis to form distinct rods made up of cartilage tissue. A marker of this differentiation step is the synthesis of type II collagen. Metabolic labelling, pepsin digestion, SDS-PAGE, and autoradiography were used to demonstrate this protein when cartilage tissue is present in the cultures. After additional culture time, terminal chondrocyte differentiation and morphogenesis take place in specific regions of the cartilage rods to form bands of hypertrophied chondrocytes. One marker of this differentiation step is the synthesis of the enzyme alkaline phosphatase. We have measured the activity of this enzyme throughout the culture period and see a substantial increase at the time of terminal chondrocyte differentiation. Another feature of hypertrophied chondrocytes is that the matrix around the cells becomes calcified. Calcified matrix in our cultured pre-metatarsals was visualized by staining with alizarin red. By supplementing the defined culture medium with ITS, we observed that terminal chondrocyte differentiation took place in a shorter culture time. Supplementation of the medium with serum results in a similar acceleration of terminal differentiation, and, with additional culture time, an osteoid-like matrix forms around the central region of the rods.

MicroRNA miR-124 Controls the Choice between Neuronal and Astrocyte Differentiation by Fine-tuning Ezh2 Expression*

PubMed Central

Neo, Wen Hao; Yap, Karen; Lee, Suet Hoay; Looi, Liang Sheng; Khandelia, Piyush; Neo, Sheng Xiong; Makeyev, Eugene V.; Su, I-hsin

2014-01-01

Polycomb group protein Ezh2 is a histone H3 Lys-27 histone methyltransferase orchestrating an extensive epigenetic regulatory program. Several nervous system-specific genes are known to be repressed by Ezh2 in stem cells and derepressed during neuronal differentiation. However, the molecular mechanisms underlying this regulation remain poorly understood. Here we show that Ezh2 levels are dampened during neuronal differentiation by brain-enriched microRNA miR-124. Expression of miR-124 in a neuroblastoma cells line was sufficient to up-regulate a significant fraction of nervous system-specific Ezh2 target genes. On the other hand, naturally elevated expression of miR-124 in embryonic carcinoma cells undergoing neuronal differentiation correlated with down-regulation of Ezh2 levels. Importantly, overexpression of Ezh2 mRNA with a 3′-untranslated region (3′-UTR) lacking a functional miR-124 binding site, but not with the wild-type Ezh2 3′-UTR, hampered neuronal and promoted astrocyte-specific differentiation in P19 and embryonic mouse neural stem cells. Overall, our results uncover a molecular mechanism that allows miR-124 to balance the choice between alternative differentiation possibilities through fine-tuning the expression of a critical epigenetic regulator. PMID:24878960

MicroRNA miR-124 controls the choice between neuronal and astrocyte differentiation by fine-tuning Ezh2 expression.

PubMed

Neo, Wen Hao; Yap, Karen; Lee, Suet Hoay; Looi, Liang Sheng; Khandelia, Piyush; Neo, Sheng Xiong; Makeyev, Eugene V; Su, I-hsin

2014-07-25

Polycomb group protein Ezh2 is a histone H3 Lys-27 histone methyltransferase orchestrating an extensive epigenetic regulatory program. Several nervous system-specific genes are known to be repressed by Ezh2 in stem cells and derepressed during neuronal differentiation. However, the molecular mechanisms underlying this regulation remain poorly understood. Here we show that Ezh2 levels are dampened during neuronal differentiation by brain-enriched microRNA miR-124. Expression of miR-124 in a neuroblastoma cells line was sufficient to up-regulate a significant fraction of nervous system-specific Ezh2 target genes. On the other hand, naturally elevated expression of miR-124 in embryonic carcinoma cells undergoing neuronal differentiation correlated with down-regulation of Ezh2 levels. Importantly, overexpression of Ezh2 mRNA with a 3'-untranslated region (3'-UTR) lacking a functional miR-124 binding site, but not with the wild-type Ezh2 3'-UTR, hampered neuronal and promoted astrocyte-specific differentiation in P19 and embryonic mouse neural stem cells. Overall, our results uncover a molecular mechanism that allows miR-124 to balance the choice between alternative differentiation possibilities through fine-tuning the expression of a critical epigenetic regulator.

Fine regulation of RhoA and Rock is required for skeletal muscle differentiation.

PubMed

Castellani, Loriana; Salvati, Erica; Alemà, Stefano; Falcone, Germana

2006-06-02

The RhoA GTPase controls a variety of cell functions such as cell motility, cell growth, and gene expression. Previous studies suggested that RhoA mediates signaling inputs that promote skeletal myogenic differentiation. We show here that levels and activity of RhoA protein are down-regulated in both primary avian myoblasts and mouse satellite cells undergoing differentiation, suggesting that a fine regulation of this GTPase is required. In addition, ectopic expression of activated RhoA in primary quail myocytes, but not in mouse myocytes, inhibits accumulation of muscle-specific proteins and cell fusion. By disrupting RhoA signaling with specific inhibitors, we have shown that this GTPase, although required for cell identity in proliferating myoblasts, is not essential for commitment to terminal differentiation and muscle gene expression. Ectopic expression of an activated form of its downstream effector, Rock, impairs differentiation of both avian and mouse myoblasts. Conversely, Rock inhibition with specific inhibitors and small interfering RNA-mediated gene silencing leads to accelerated progression in the lineage and enhanced cell fusion, underscoring a negative regulatory function of Rock in myogenesis. Finally, we have reported that Rock acts independently from RhoA in preventing myoblast exit from the cell cycle and commitment to differentiation and may receive signaling inputs from Raf-1 kinase.

Lace plant ethylene receptors, AmERS1a and AmERS1c, regulate ethylene-induced programmed cell death during leaf morphogenesis.

PubMed

Rantong, Gaolathe; Evans, Rodger; Gunawardena, Arunika H L A N

2015-10-01

The lace plant, Aponogeton madagascariensis, is an aquatic monocot that forms perforations in its leaves as part of normal leaf development. Perforation formation occurs through developmentally regulated programmed cell death (PCD). The molecular basis of PCD regulation in the lace plant is unknown, however ethylene has been shown to play a significant role. In this study, we examined the role of ethylene receptors during perforation formation. We isolated three lace plant ethylene receptors AmERS1a, AmERS1b and AmERS1c. Using quantitative PCR, we examined their transcript levels at seven stages of leaf development. Through laser-capture microscopy, transcript levels were also determined in cells undergoing PCD and cells not undergoing PCD (NPCD cells). AmERS1a transcript levels were significantly lower in window stage leaves (in which perforation formation and PCD are occurring) as compared to all other leaf developmental stages. AmERS1a and AmERS1c (the most abundant among the three receptors) had the highest transcript levels in mature stage leaves, where PCD is not occurring. Their transcript levels decreased significantly during senescence-associated PCD. AmERS1c had significantly higher transcript levels in NPCD compared to PCD cells. Despite being significantly low in window stage leaves, AmERS1a transcripts were not differentially expressed between PCD and NPCD cells. The results suggested that ethylene receptors negatively regulate ethylene-controlled PCD in the lace plant. A combination of ethylene and receptor levels determines cell fate during perforation formation and leaf senescence. A new model for ethylene emission and receptor expression during lace plant perforation formation and senescence is proposed.

SVEP1 is a novel marker of activated pre-determined skeletal muscle satellite cells.

PubMed

Shefer, Gabi; Benayahu, Dafna

2010-03-01

In this study we explored the expression pattern of SVEP1, a novel cell adhesion molecule (CAM), in bona fide satellite cells and their immediate progeny. We show that SVEP1 is expressed in activated satellite cells prior to their determination to the myogenic lineage. SVEP1 was also expressed during early phases of myogenic differentiation through the initial stage of myoblast fusion and myotube formation. The expression of SVEP1 was shown by immunostaining two cell culture systems: freshly isolated myofibers and primary myoblasts. Pax7 was used to pinpoint satellite cells situated in their niche on myofibers, and activated satellite cells were determined based on BrdU incorporation (Pax7(+)/BrdU(+)cells). MyoD marked satellite cells fated to undergo myogenesis as well as proliferating and differentiating myoblasts. Differentiating myoblasts and myotubes were identified based on their sarcomeric myosin expression. We showed that SVEP1 was specifically expressed in pre-determined activated satellite cells (Pax7(+)/ BrdU(+) /MyoD(-)) accounting for about 24% of total satellite cells. On the other hand, SVEP1 expression was absent in quiescent satellite cells (Pax7(+)/BrdU(-)/MyoD(-)). Moreover, based on MyoD/sarcomeric myosin co-expression SVEP1 was shown to be expressed throughout the early phases of myogenesis up until myoblast fusion and myotube formation. A decline in SVEP1 expression occurred upon myotube maturation. We suggest SVEP1 as a potential biomarker for pre-fated satellite cells. The impact of this finding is that it may allow scrutinizing signals that affect differentiation commitment. Thus, holds a therapeutic potential for maladies that involve deregulated stem cell fate-decision.

Embryonic development of Python sebae - II: Craniofacial microscopic anatomy, cell proliferation and apoptosis.

PubMed

Buchtová, Marcela; Boughner, Julia C; Fu, Katherine; Diewert, Virginia M; Richman, Joy M

2007-01-01

This study explores the microscopic craniofacial morphogenesis of the oviparous African rock python (Python sebae) spanning the first two-thirds of the post-oviposition period. At the time of laying, the python embryo consists of largely undifferentiated mesenchyme and epithelium with the exception of the cranial base and trabeculae cranii, which are undergoing chondrogenesis. The facial prominences are well defined and are at a late stage, close to the time when lip fusion begins. Later (11-12d), specializations in the epithelia begin to differentiate (vomeronasal and olfactory epithelia, teeth). Dental development in snakes is different from that of mammals in several aspects including an extended dental lamina with the capacity to form 4 sets of generational teeth. In addition, the ophidian olfactory system is very different from the mammalian. There is a large vomeronasal organ, a nasal cavity proper and an extraconchal space. All of these areas are lined with a greatly expanded olfactory epithelium. Intramembranous bone differentiation is taking place at stage 3 with some bones already ossifying whereas most are only represented as mesenchymal condensations. In addition to routine histological staining, PCNA immunohistochemistry reveals relatively higher levels of proliferation in the extending dental laminae, in osseous mesenchymal condensations and in the olfactory epithelia. Areas undergoing apoptosis were noted in the enamel organs of the teeth and osseous mesenchymal condensations. We propose that localized apoptosis helps to divide a single condensation into multiple ossification centres and this is a mechanism whereby novel morphology can be selected in response to evolutionary pressures. Several additional differences in head morphology between snakes and other amniotes were noted including a palatal groove separating the inner and outer row of teeth in the upper jaw, a tracheal opening within the tongue and a pharyngeal adhesion that closes off the pharynx from the oral cavity between stages 1 and 4. Our studies on these and other differences in the python will provide valuable insights into in developmental, molecular and evolutionary mechanisms of patterning.

Neural Differentiation of Embryonic Stem Cells In Vitro: A Road Map to Neurogenesis in the Embryo

PubMed Central

Abranches, Elsa; Silva, Margarida; Pradier, Laurent; Schulz, Herbert; Hummel, Oliver; Henrique, Domingos; Bekman, Evguenia

2009-01-01

Background The in vitro generation of neurons from embryonic stem (ES) cells is a promising approach to produce cells suitable for neural tissue repair and cell-based replacement therapies of the nervous system. Available methods to promote ES cell differentiation towards neural lineages attempt to replicate, in different ways, the multistep process of embryonic neural development. However, to achieve this aim in an efficient and reproducible way, a better knowledge of the cellular and molecular events that are involved in the process, from the initial specification of neuroepithelial progenitors to their terminal differentiation into neurons and glial cells, is required. Methodology/Principal Findings In this work, we characterize the main stages and transitions that occur when ES cells are driven into a neural fate, using an adherent monolayer culture system. We established improved conditions to routinely produce highly homogeneous cultures of neuroepithelial progenitors, which organize into neural tube-like rosettes when they acquire competence for neuronal production. Within rosettes, neuroepithelial progenitors display morphological and functional characteristics of their embryonic counterparts, namely, apico-basal polarity, active Notch signalling, and proper timing of production of neurons and glia. In order to characterize the global gene activity correlated with each particular stage of neural development, the full transcriptome of different cell populations that arise during the in vitro differentiation protocol was determined by microarray analysis. By using embryo-oriented criteria to cluster the differentially expressed genes, we define five gene expression signatures that correlate with successive stages in the path from ES cells to neurons. These include a gene signature for a primitive ectoderm-like stage that appears after ES cells enter differentiation, and three gene signatures for subsequent stages of neural progenitor development, from an early stage that follows neural induction to a final stage preceding terminal differentiation. Conclusions/Significance Overall, our work confirms and extends the cellular and molecular parallels between monolayer ES cell neural differentiation and embryonic neural development, revealing in addition novel aspects of the genetic network underlying the multistep process that leads from uncommitted cells to differentiated neurons. PMID:19621087

Infectivity of Plasmodium falciparum sporozoites determines emerging parasitemia in infected volunteers.

PubMed

McCall, Matthew B B; Wammes, Linda J; Langenberg, Marijke C C; van Gemert, Geert-Jan; Walk, Jona; Hermsen, Cornelus C; Graumans, Wouter; Koelewijn, Rob; Franetich, Jean-François; Chishimba, Sandra; Gerdsen, Max; Lorthiois, Audrey; van de Vegte, Marga; Mazier, Dominique; Bijker, Else M; van Hellemond, Jaap J; van Genderen, Perry J J; Sauerwein, Robert W

2017-06-21

Malaria sporozoites must first undergo intrahepatic development before a pathogenic blood-stage infection is established. The success of infection depends on host and parasite factors. In healthy human volunteers undergoing controlled human malaria infection (CHMI), we directly compared three clinical Plasmodium falciparum isolates for their ability to infect primary human hepatocytes in vitro and to drive the production of blood-stage parasites in vivo. Our data show a correlation between the efficiency of strain-specific sporozoite invasion of human hepatocytes and the dynamics of patent parasitemia in study subjects, highlighting intrinsic differences in infectivity among P. falciparum isolates from distinct geographical locales. The observed heterogeneity in infectivity among strains underscores the value of assessing the protective efficacy of candidate malaria vaccines against heterologous strains in the CHMI model. Copyright © 2017 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

Blood-stage malaria of Plasmodium chabaudi induces differential Tlr expression in the liver of susceptible and vaccination-protected Balb/c mice.

PubMed

Al-Quraishy, Saleh; Dkhil, Mohamed A; Alomar, Suliman; Abdel-Baki, Abdel Azeem S; Delic, Denis; Wunderlich, Frank; Araúzo-Bravo, Marcos J

2016-05-01

Protective vaccination induces self-healing of otherwise lethal blood-stage infections of Plasmodium chabaudi malaria. Here, we investigate mRNA expression patterns of all 12 members of the Toll-like receptor (Tlr) gene family in the liver, a major effector organ against blood-stage malaria, during lethal and vaccination-induced self-healing infections of P. chabaudi in female Balb/c mice. Gene expression microarrays reveal that all 12 Tlr genes are constitutively expressed, though at varying levels, and specifically respond to infection. Protective vaccination does not affect constitutive expression of any of the 12 Tlr genes but leads to differential expression (p < 0.05) of seven Tlrs (1, 2, 4, 7, 8, 12, and 13) in response to malaria. Quantitative PCR substantiates differential expression at p < 0.01. There is an increased expression of Tlr2 by approximately five-fold on day 1 post-infection (p.i.) and Tlr1 by approximately threefold on day 4 p.i.. At peak parasitemia on day 8 p.i., none of the 12 Tlrs display any differential expression. After peak parasitemia, towards the end of the crisis phase on day 11 p.i., expression of Tlrs 1, 4, and 12 is increased by approximately four-, two-, and three-fold, respectively, and that of Tlr7 is decreased by approximately two-fold. Collectively, our data suggest that though all 12 members of the Tlr gene family are specifically responsive to malaria in the liver, not only Tlr2 at the early stage of infection but also the Tlrs 1, 4, 7, and 12 towards the end of crisis phase are critical for vaccination-induced resolution and survival of otherwise lethal blood-stage malaria.

Robotic assisted kidney transplantation

PubMed Central

Modi, Pranjal; Pal, Bipinchandra; Modi, Jayesh; Kumar, Suresh; Sood, Akshay; Menon, Mani

2014-01-01

Kidney transplantation is the standard of care for patients with end stage renal disease. While open surgery remains the gold standard, minimally invasive surgery has recently been introduced for the recipient undergoing kidney transplantation. We review the evolution of techniques of minimally invasive surgery for kidney transplantation with specific emphasis on technical aspects of robotic assisted kidney transplantation. PMID:25097315

Regulation of H3K4me3 at Transcriptional Enhancers Characterizes Acquisition of Virus-Specific CD8+ T Cell-Lineage-Specific Function.

PubMed

Russ, Brendan E; Olshansky, Moshe; Li, Jasmine; Nguyen, Michelle L T; Gearing, Linden J; Nguyen, Thi H O; Olson, Matthew R; McQuilton, Hayley A; Nüssing, Simone; Khoury, Georges; Purcell, Damian F J; Hertzog, Paul J; Rao, Sudha; Turner, Stephen J

2017-12-19

Infection triggers large-scale changes in the phenotype and function of T cells that are critical for immune clearance, yet the gene regulatory mechanisms that control these changes are largely unknown. Using ChIP-seq for specific histone post-translational modifications (PTMs), we mapped the dynamics of ∼25,000 putative CD8 + T cell transcriptional enhancers (TEs) differentially utilized during virus-specific T cell differentiation. Interestingly, we identified a subset of dynamically regulated TEs that exhibited acquisition of a non-canonical (H3K4me3 + ) chromatin signature upon differentiation. This unique TE subset exhibited characteristics of poised enhancers in the naive CD8 + T cell subset and demonstrated enrichment for transcription factor binding motifs known to be important for virus-specific CD8 + T cell differentiation. These data provide insights into the establishment and maintenance of the gene transcription profiles that define each stage of virus-specific T cell differentiation. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

The development of a cancer genetic-specific measure of coping: the GRACE.

PubMed

Phelps, C; Bennett, P; Jones, H; Hood, K; Brain, K; Murray, A

2010-08-01

Generic measures of coping fail to capture the process of undergoing specific health processes such as cancer genetic risk assessment. The Genetic Risk Assessment Coping Evaluation (GRACE) has been developed to provide greater specificity of measurement. Based upon previous research findings, the GRACE measures the degree of stress associated with 11 recognised sources of stress for individuals undergoing the early stages of cancer genetic risk assessment, and the use of up to eight coping strategies they may elicit. This paper reports preliminary data from the piloting of the GRACE within a randomised trial of a coping intervention. Of the 265 participants who completed and returned their baseline questionnaire (prior to being informed of their level of genetic risk), 257 completed the GRACE. The most highly endorsed sources of stress involved concerns relating to family members, endorsed by over 60% of respondents, and concerns about how the participants would cope if found to be at increased risk (59%). Participants made use of multiple coping strategies across different sources of stress. The most frequently reported coping strategies were emotion-focused, which may reflect the stage of the assessment process. The completion rates for the matrix and specificity of responses provided suggest that the GRACE may be an acceptable measurement tool. Further data collection and validation is ongoing. (c) 2009 John Wiley & Sons, Ltd.

Physico-electrochemical Characterization of Pluripotent Stem Cells during Self-Renewal or Differentiation by a Multi-modal Monitoring System.

PubMed

Low, Karen; Wong, Lauren Y; Maldonado, Maricela; Manjunath, Chetas; Horner, Christopher B; Perez, Mark; Myung, Nosang V; Nam, Jin

2017-05-09

Monitoring pluripotent stem cell behaviors (self-renewal and differentiation to specific lineages/phenotypes) is critical for a fundamental understanding of stem cell biology and their translational applications. In this study, a multi-modal stem cell monitoring system was developed to quantitatively characterize physico-electrochemical changes of the cells in real time, in relation to cellular activities during self-renewal or lineage-specific differentiation, in a non-destructive, label-free manner. The system was validated by measuring physical (mass) and electrochemical (impedance) changes in human induced pluripotent stem cells undergoing self-renewal, or subjected to mesendodermal or ectodermal differentiation, and correlating them to morphological (size, shape) and biochemical changes (gene/protein expression). An equivalent circuit model was used to further dissect the electrochemical (resistive and capacitive) contributions of distinctive cellular features. Overall, the combination of the physico-electrochemical measurements and electrical circuit modeling collectively offers a means to longitudinally quantify the states of stem cell self-renewal and differentiation. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Biomineralization, life-time of odontogenic cells and differential expression of the two homeobox genes MSX-1 and DLX-2 in transgenic mice.

PubMed

Lézot, F; Thomas, B; Hotton, D; Forest, N; Orestes-Cardoso, S; Robert, B; Sharpe, P; Berdal, A

2000-03-01

Msx and Dlx homeobox genes encode for transcription factors that control early morphogenesis. More specifically, Msx-1, Msx-2, and Dlx-2 homeobox genes contribute to the initial patterning of the dentition. The present study is devoted to the potential role of those homeobox genes during the late formation of mineralized tissues, using the rodent incisor as an experimental system. The continuously erupting mandibular incisor allows (1) the coinvestigation of the whole sequences of amelogenesis and dentinogenesis, aligned along the main dental axis in a single sample in situ and (2) the differential characterization of transcripts generated by epithelial and ectomesenchymal odontogenic cells. Northern blot experiments on microdissected cells showed the continuing expression of Msx-2 and Dlx-2 in the later stages of dental biomineralization, differentially in epithelial and ectomesenchymal compartments. Transgenic mice produced with LacZ reporter constructs for Dlx-2 and Msx-1 were used to detect different components of the gene expression patterns with the sensitive beta-galactosidase histoenzymology. The results show a prominent epithelial involvement of Dlx-2, with stage-specific variations in the cells involved in enamel formation. Quantitative analyses identified specific modulations of Dlx-2 expression in ameloblasts depending on the anatomical sites of the incisor, showing more specifically an inverse linear relationship between the Dlx-2 promoter activity level and enamel thickness. This investigation extends the role of homeoproteins to postmitotic stages, which would control secretory cell activity, in a site-specific manner as shown here for Dlx-2.

Reconstructing human pancreatic differentiation by mapping specific cell populations during development.

PubMed

Ramond, Cyrille; Glaser, Nicolas; Berthault, Claire; Ameri, Jacqueline; Kirkegaard, Jeannette Schlichting; Hansson, Mattias; Honoré, Christian; Semb, Henrik; Scharfmann, Raphaël

2017-07-21

Information remains scarce on human development compared to animal models. Here, we reconstructed human fetal pancreatic differentiation using cell surface markers. We demonstrate that at 7weeks of development, the glycoprotein 2 (GP2) marks a multipotent cell population that will differentiate into the acinar, ductal or endocrine lineages. Development towards the acinar lineage is paralleled by an increase in GP2 expression. Conversely, a subset of the GP2 + population undergoes endocrine differentiation by down-regulating GP2 and CD142 and turning on NEUROG3 , a marker of endocrine differentiation. Endocrine maturation progresses by up-regulating SUSD2 and lowering ECAD levels. Finally, in vitro differentiation of pancreatic endocrine cells derived from human pluripotent stem cells mimics key in vivo events. Our work paves the way to extend our understanding of the origin of mature human pancreatic cell types and how such lineage decisions are regulated.

Nuclear magnetic resonance studies of ancient buried wood-II. Observations on the origin of coal from lignite to bituminous coal

USGS Publications Warehouse

Hatcher, P.G.; Breger, I.A.; Szeverenyi, N.; Maciel, G.E.

1982-01-01

Coalified logs ranging in age from Late Pennsylvania to Miocene and in rank from lignite B to bituminous coal were analyzed by 13C nuclear magnetic resonance (NMR) utilizing the cross-polarization, magic-angle spinning technique, as well as by infrared spectroscopy. The results of this study indicate that at least three major stages of coalification can be observed as wood gradually undergoes transformation to bituminous coal. The first stage involves hydrolysis and loss of cellulose from wood with retention and differential concentration of the resistant lignin. The second stage involves conversion of the lignin residues directly to coalified wood of lignitic rank, during which the oxygen content of intermediate diagenetic products remains constant as the hydrogen content and the carbon content increases. These changes are thought to involve loss of methoxyl groups, water, and C3 side chains from the lignin. In the third major stage of coalification, the coalified wood increases in rank to subbituminous and bituminous coal; during this stage the oxygen content decreases, hydrogen remains constant, and the carbon content increases. These changes are thought to result from loss of soluble humic acids that are rich in oxygen and that are mobilized during compaction and dewatering. Relatively resistant resinous substances are differentially concentrated in the coal during this stage. The hypothesis that humic acids are formed as mobile by-products of the coalification of lignin and function only as vehicles for removal of oxygen represents a dramatic departure from commonly accepted views that they are relatively low-molecular-weight intermediates formed during the degradation of lignin that then condense to form high-molecular-weight coal structures. ?? 1982.

The development of human mast cells. An historical reappraisal

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

Ribatti, Domenico, E-mail: domenico.ribatti@uniba.it

2016-03-15

The understanding of mast cell (MC) differentiation is derived mainly from in vitro studies of different stages of stem and progenitor cells. The hematopoietic lineage development of human MCs is unique compared to other myeloid-derived cells. Human MCs originate from CD34{sup +}/CD117{sup +}/CD13{sup +}multipotent hematopoietic progenitors, which undergo transendothelial recruitment into peripheral tissues, where they complete differentiation. Stem cell factor (SCF) is a major chemotactic factor for MCs and their progenitors. SCF also elicits cell-cell and cell-substratum adhesion, facilitates the proliferation, and sustains the survival, differentiation, and maturation, of MCs. Because MC maturation is influenced by local microenvironmental factors, differentmore » MC phenotypes can develop in different tissues and organs. - Highlights: • Human mast cells originate from CD34/CD117/CD13 positive multipotent hematopoietic progenitors. • Stem cell factor is a major chemotactic factor for mast cells and their progenitors. • Different mast cell phenotypes can develop in different tissues and organs.« less

Differential Diagnosis of Selective Mutism in Bilingual Children

PubMed Central

TOPPELBERG, CLAUDIO O.; TABORS, PATTON; COGGINS, ALISSA; LUM, KIRK; BURGER, CLAUDIA

2012-01-01

Early diagnosis of selective mutism (SM) is an important concern. SM prevalence is higher than initially thought and at least three times higher in immigrant language minority children. Although the DSM-IV precludes diagnosing SM in immigrant children with limited language proficiency (as children acquiring a second language may normally undergo a “silent period”), specific diagnostic boundaries are not clear. The specific focus of this article is, therefore, the exact circumstances in which a language minority child should be diagnosed with SM. PMID:15908842

Development of the arterial pattern in the upper limb of staged human embryos: normal development and anatomic variations

PubMed Central

RODRÍGUEZ-NIEDENFÜHR, M.; BURTON, G. J.; DEU, J.; SAÑUDO, J. R.

2001-01-01

A total of 112 human embryos (224 upper limbs) between stages 12 and 23 of development were examined. It was observed that formation of the arterial system in the upper limb takes place as a dual process. An initial capillary plexus appears from the dorsal aorta during stage 12 and develops at the same rate as the limb. At stage 13, the capillary plexus begins a maturation process involving the enlargement and differentiation of selected parts. This remodelling process starts in the aorta and continues in a proximal to distal sequence. By stage 15 the differentiation has reached the subclavian and axillary arteries, by stage 17 it has reached the brachial artery as far as the elbow, by stage 18 it has reached the forearm arteries except for the distal part of the radial, and finally by stage 21 the whole arterial pattern is present in its definitive morphology. This differentiation process parallels the development of the skeletal system chronologically. A number of arterial variations were observed, and classified as follows: superficial brachial (7.7%), accessory brachial (0.6%), brachioradial (14%), superficial brachioulnar (4.7%), superficial brachioulnoradial (0.7%), palmar pattern of the median (18.7%) and superficial brachiomedian (0.7%) arteries. They were observed in embryos belonging to stages 17–23 and were not related to a specific stage of development. Statistical comparison with the rates of variations reported in adults did not show significant differences. It is suggested that the variations arise through the persistence, enlargement and differentiation of parts of the initial network which would normally remain as capillaries or even regress. PMID:11693301

Incorporating Exercise Into the Cancer Treatment Paradigm.

PubMed

Haas, Barbara K; Hermanns, Melinda; Kimmel, Gary

2016-12-01

The benefits of exercise for patients with cancer are well documented. However, exercise is still not a standard of care for this population. Several factors contribute to the lack of exercise prescriptions for patients with cancer, including challenges posed by treatment-related side effects, lack of knowledge among healthcare providers and the laypeople, and inadequate resources. This article reviews the benefits of exercise in general and specifically to patients with cancer, discusses the specific challenges and considerations required in recommending exercise to this population, and provides specific recommendations for healthcare providers to incorporate exercise into treatment plans. Using a case study exemplar, this article discusses the benefits and challenges to exercise while undergoing treatment for cancer and proposes specific solutions and recommendations. Oncology practitioners can provide the opportunity for patients to safely engage in exercise with the appropriate resources and trained personnel using a successful model of delivering exercise to patients undergoing treatment for cancer. Exercise improves quality of life in all patients, including those with advanced-stage cancers and those actively receiving treatment.

Developmental expression and function analysis of protein tyrosine phosphatase receptor type D in oligodendrocyte myelination

PubMed Central

Zhu, Qiang; Tan, Zhou; Zhao, Shufang; Huang, Hao; Zhao, Xiaofeng; Hu, Xuemei; Zhang, Yiping; Shields, Christopher B; Uetani, Noriko; Qiu, Mengsheng

2015-01-01

Receptor protein tyrosine phosphatases (RPTPs) are extensively expressed in the central nervous system (CNS), and have distinct spatial and temporal patterns in different cell types during development. Previous studies have demonstrated possible roles for RPTPs in axon outgrowth, guidance, and synaptogenesis. In the present study, our results revealed that protein tyrosine phosphatase, receptor type D (PTPRD) was initially expressed in mature neurons in embryonic CNS, and later in oligodendroglial cells at postnatal stages when oligodendrocyte undergo active axonal myelination process. In PTPRD mutants, oligodendrocyte differentiation was normal and a transient myelination delay occurred at early postnatal stages, indicating the contribution of PTPRD to the initiation of axonal myelination. Our results also showed that the remyelination process was not affected in the absence of PTPRD function after a cuprizone-induced demyelination in adult animals. PMID:26341907

Delineation of Matriptase Protein Expression by Enzymatic Gene Trapping Suggests Diverging Roles in Barrier Function, Hair Formation, and Squamous Cell Carcinogenesis

PubMed Central

List, Karin; Szabo, Roman; Molinolo, Alfredo; Nielsen, Boye Schnack; Bugge, Thomas H.

2006-01-01

The membrane serine protease matriptase is required for epidermal barrier function, hair formation, and thymocyte development in mice, and dysregulated matriptase expression causes epidermal squamous cell carcinoma. To elucidate the specific functions of matriptase in normal and aberrant epidermal differentiation, we used enzymatic gene trapping combined with immunohistochemical, ultrastructural, and barrier function assays to delineate the spatio-temporal expression and function of matriptase in mouse keratinized tissue development, homeostasis, and malignant transformation. In the interfollicular epidermis, matriptase expression was restricted to postmitotic transitional layer keratinocytes undergoing terminal differentiation. Matriptase was also expressed in keratinizing oral epithelium, where it was required for oral barrier function, and in thymic epithelium. In all three tissues, matriptase colocalized with profilaggrin. In staged embryos, the onset of epidermal matriptase expression coincided with that of profilaggrin expression and acquisition of the epidermal barrier. In marked contrast to stratifying keritinized epithelium, matripase expression commenced already in undifferentiated and rapidly proliferating profilaggrin-negative matrix cells and displayed hair growth cycle-dependent expression. Exposure of the epidermis to carcinogens led to the gradual appearance of matriptase in a keratin-5-positive proliferative cell compartment during malignant progression. Combined with previous studies, these data suggest that matriptase has diverging functions in the genesis of stratified keratinized epithelium, hair follicles, and squamous cell carcinoma. PMID:16651618

Delineation of matriptase protein expression by enzymatic gene trapping suggests diverging roles in barrier function, hair formation, and squamous cell carcinogenesis.

PubMed

List, Karin; Szabo, Roman; Molinolo, Alfredo; Nielsen, Boye Schnack; Bugge, Thomas H

2006-05-01

The membrane serine protease matriptase is required for epidermal barrier function, hair formation, and thymocyte development in mice, and dysregulated matriptase expression causes epidermal squamous cell carcinoma. To elucidate the specific functions of matriptase in normal and aberrant epidermal differentiation, we used enzymatic gene trapping combined with immunohistochemical, ultrastructural, and barrier function assays to delineate the spatio-temporal expression and function of matriptase in mouse keratinized tissue development, homeostasis, and malignant transformation. In the interfollicular epidermis, matriptase expression was restricted to postmitotic transitional layer keratinocytes undergoing terminal differentiation. Matriptase was also expressed in keratinizing oral epithelium, where it was required for oral barrier function, and in thymic epithelium. In all three tissues, matriptase colocalized with profilaggrin. In staged embryos, the onset of epidermal matriptase expression coincided with that of profilaggrin expression and acquisition of the epidermal barrier. In marked contrast to stratifying keritinized epithelium, matripase expression commenced already in undifferentiated and rapidly proliferating profilaggrin-negative matrix cells and displayed hair growth cycle-dependent expression. Exposure of the epidermis to carcinogens led to the gradual appearance of matriptase in a keratin-5-positive proliferative cell compartment during malignant progression. Combined with previous studies, these data suggest that matriptase has diverging functions in the genesis of stratified keratinized epithelium, hair follicles, and squamous cell carcinoma.

Hyperspectral imaging based on compressive sensing to determine cancer margins in human pancreatic tissue ex vivo

NASA Astrophysics Data System (ADS)

Peller, Joseph; Thompson, Kyle J.; Siddiqui, Imran; Martinie, John; Iannitti, David A.; Trammell, Susan R.

2017-02-01

Pancreatic cancer is the fourth leading cause of cancer death in the US. Currently, surgery is the only treatment that offers a chance of cure, however, accurately identifying tumor margins in real-time is difficult. Research has demonstrated that optical spectroscopy can be used to distinguish between healthy and diseased tissue. The design of a single-pixel imaging system for cancer detection is discussed. The system differentiates between healthy and diseased tissue based on differences in the optical reflectance spectra of these regions. In this study, pancreatic tissue samples from 6 patients undergoing Whipple procedures are imaged with the system (total number of tissue sample imaged was N=11). Regions of healthy and unhealthy tissue are determined based on SAM analysis of these spectral images. Hyperspectral imaging results are then compared to white light imaging and histological analysis. Cancerous regions were clearly visible in the hyperspectral images. Margins determined via spectral imaging were in good agreement with margins identified by histology, indicating that hyperspectral imaging system can differentiate between healthy and diseased tissue. After imaging the system was able to detect cancerous regions with a sensitivity of 74.50±5.89% and a specificity of 75.53±10.81%. Possible applications of this imaging system include determination of tumor margins during surgery/biopsy and assistance with cancer diagnosis and staging.

Metabolic reprogramming during neuronal differentiation.

PubMed

Agostini, M; Romeo, F; Inoue, S; Niklison-Chirou, M V; Elia, A J; Dinsdale, D; Morone, N; Knight, R A; Mak, T W; Melino, G

2016-09-01

Newly generated neurons pass through a series of well-defined developmental stages, which allow them to integrate into existing neuronal circuits. After exit from the cell cycle, postmitotic neurons undergo neuronal migration, axonal elongation, axon pruning, dendrite morphogenesis and synaptic maturation and plasticity. Lack of a global metabolic analysis during early cortical neuronal development led us to explore the role of cellular metabolism and mitochondrial biology during ex vivo differentiation of primary cortical neurons. Unexpectedly, we observed a huge increase in mitochondrial biogenesis. Changes in mitochondrial mass, morphology and function were correlated with the upregulation of the master regulators of mitochondrial biogenesis, TFAM and PGC-1α. Concomitant with mitochondrial biogenesis, we observed an increase in glucose metabolism during neuronal differentiation, which was linked to an increase in glucose uptake and enhanced GLUT3 mRNA expression and platelet isoform of phosphofructokinase 1 (PFKp) protein expression. In addition, glutamate-glutamine metabolism was also increased during the differentiation of cortical neurons. We identified PI3K-Akt-mTOR signalling as a critical regulator role of energy metabolism in neurons. Selective pharmacological inhibition of these metabolic pathways indicate existence of metabolic checkpoint that need to be satisfied in order to allow neuronal differentiation.

Metabolic reprogramming during neuronal differentiation

PubMed Central

Agostini, M; Romeo, F; Inoue, S; Niklison-Chirou, M V; Elia, A J; Dinsdale, D; Morone, N; Knight, R A; Mak, T W; Melino, G

2016-01-01

Newly generated neurons pass through a series of well-defined developmental stages, which allow them to integrate into existing neuronal circuits. After exit from the cell cycle, postmitotic neurons undergo neuronal migration, axonal elongation, axon pruning, dendrite morphogenesis and synaptic maturation and plasticity. Lack of a global metabolic analysis during early cortical neuronal development led us to explore the role of cellular metabolism and mitochondrial biology during ex vivo differentiation of primary cortical neurons. Unexpectedly, we observed a huge increase in mitochondrial biogenesis. Changes in mitochondrial mass, morphology and function were correlated with the upregulation of the master regulators of mitochondrial biogenesis, TFAM and PGC-1α. Concomitant with mitochondrial biogenesis, we observed an increase in glucose metabolism during neuronal differentiation, which was linked to an increase in glucose uptake and enhanced GLUT3 mRNA expression and platelet isoform of phosphofructokinase 1 (PFKp) protein expression. In addition, glutamate–glutamine metabolism was also increased during the differentiation of cortical neurons. We identified PI3K–Akt–mTOR signalling as a critical regulator role of energy metabolism in neurons. Selective pharmacological inhibition of these metabolic pathways indicate existence of metabolic checkpoint that need to be satisfied in order to allow neuronal differentiation. PMID:27058317

Congenital heart disease protein 5 associates with CASZ1 to maintain myocardial tissue integrity.

PubMed

Sojka, Stephen; Amin, Nirav M; Gibbs, Devin; Christine, Kathleen S; Charpentier, Marta S; Conlon, Frank L

2014-08-01

The identification and characterization of the cellular and molecular pathways involved in the differentiation and morphogenesis of specific cell types of the developing heart are crucial to understanding the process of cardiac development and the pathology associated with human congenital heart disease. Here, we show that the cardiac transcription factor CASTOR (CASZ1) directly interacts with congenital heart disease 5 protein (CHD5), which is also known as tryptophan-rich basic protein (WRB), a gene located on chromosome 21 in the proposed region responsible for congenital heart disease in individuals with Down's syndrome. We demonstrate that loss of CHD5 in Xenopus leads to compromised myocardial integrity, improper deposition of basement membrane, and a resultant failure of hearts to undergo cell movements associated with cardiac formation. We further report that CHD5 is essential for CASZ1 function and that the CHD5-CASZ1 interaction is necessary for cardiac morphogenesis. Collectively, these results establish a role for CHD5 and CASZ1 in the early stages of vertebrate cardiac development. © 2014. Published by The Company of Biologists Ltd.

Implications of inaccurate clinical nodal staging in pancreatic adenocarcinoma.

PubMed

Swords, Douglas S; Firpo, Matthew A; Johnson, Kirsten M; Boucher, Kenneth M; Scaife, Courtney L; Mulvihill, Sean J

2017-07-01

Many patients with stage I-II pancreatic adenocarcinoma do not undergo resection. We hypothesized that (1) clinical staging underestimates nodal involvement, causing stage IIB to have a greater percent of resected patients and (2) this stage-shift causes discrepancies in observed survival. The Surveillance, Epidemiology, and End Results (SEER) research database was used to evaluate cause-specific survival in patients with pancreatic adenocarcinoma from 2004-2012. Survival was compared using the log-rank test. Single-center data on 105 patients who underwent resection of pancreatic adenocarcinoma without neoadjuvant treatment were used to compare clinical and pathologic nodal staging. In SEER data, medium-term survival in stage IIB was superior to IB and IIA, with median cause-specific survival of 14, 9, and 11 months, respectively (P < .001). Seventy-two percent of stage IIB patients underwent resection vs 28% in IB and 36% in IIA (P < .001). In our institutional data, 12.4% of patients had clinical evidence of nodal involvement vs 69.5% by pathologic staging (P < .001). Among clinical stage IA-IIA patients, 71.6% had nodal involvement by pathologic staging. Both SEER and institutional data support substantial underestimation of nodal involvement by clinical staging. This finding has implications in decisions regarding neoadjuvant therapy and analysis of outcomes in the absence of pathologic staging. Copyright © 2017 Elsevier Inc. All rights reserved.

Glycoconjugates reveal diversity of human neural stem cells (hNSCs) derived from human induced pluripotent stem cells (hiPSCs).

PubMed

Kandasamy, Majury; Roll, Lars; Langenstroth, Daniel; Brüstle, Oliver; Faissner, Andreas

2017-06-01

Neural stem cells (NSCs) have the ability to self-renew and to differentiate into various cell types of the central nervous system. This potential can be recapitulated by human induced pluripotent stem cells (hiPSCs) in vitro. The differentiation capacity of hiPSCs is characterized by several stages with distinct morphologies and the expression of various marker molecules. We used the monoclonal antibodies (mAbs) 487 LeX , 5750 LeX and 473HD to analyze the expression pattern of particular carbohydrate motifs as potential markers at six differentiation stages of hiPSCs. Mouse ESCs were used as a comparison. At the pluripotent stage, 487 LeX -, 5750 LeX - and 473HD-related glycans were differently expressed. Later, cells of the three germ layers in embryoid bodies (hEBs) and, even after neuralization of hEBs, subpopulations of cells were labeled with these surface antibodies. At the human rosette-stage of NSCs (hR-NSC), LeX- and 473HD-related epitopes showed antibody-specific expression patterns. We also found evidence that these surface antibodies could be used to distinguish the hR-NSCs from the hSR-NSCs stages. Characterization of hNSCs FGF-2/EGF derived from hSR-NSCs revealed that both LeX antibodies and the 473HD antibody labeled subpopulations of hNSCs FGF-2/EGF . Finally, we identified potential LeX carrier molecules that were spatiotemporally regulated in early and late stages of differentiation. Our study provides new insights into the regulation of glycoconjugates during early human stem cell development. The mAbs 487 LeX , 5750 LeX and 473HD are promising tools for identifying distinct stages during neural differentiation.

Nervous systems in 3D: a comparison of Caridean, anomuran, and brachyuran zoea-I (Decapoda).

PubMed

Geiselbrecht, Hannes; Melzer, Roland R

2013-12-01

Using serial semi-thin sections and digital 3D-reconstructions we studied the nervous systems of zoea-I larvae in three decapod species, Hippolyte inermis (Leach, 1815), Porcellana platycheles (Pennant, 1777), and Pachygrapsus marmoratus (Fabricius, 1787). These taxa represent three decapod lineages, that is, Caridea, Anomura, and Brachyura, each characterized by specific zoea-I morphology. Special attention was paid to development of ganglia, neuropil composition, and segmental nerves. In all zoeae studied, the overall elements, for example, the segmental ganglia, their neuropils and most of the nerves of the adult decapod nervous system are present. Ongoing differentiation processes are observable as well, most obvious in segments with well-developed limbs the ganglia are in a more advanced stage of differentiation and more voluminous compared to segments with only limb buds or without externally visible limb anlagen. Intra- and interspecific comparisons indicate that neuromere differentiation thus deviates from a simple anterior-posterior gradient as, for example, posterior thoracic neuromeres are less developed than those of the pleon. In addition, the differences in the progress of the development of ganglia between the studied taxa can best be attributed to heterochronic mechanisms. Taxon and stage-specific morphologies indicate that neuronal architecture reflects both, morphogenesis to the adult stage and specific larval adaptions, and provides sets of characters relevant to understanding the corresponding phylogeny. © 2013 Wiley Periodicals, Inc.

Dynamics of genomic H3K27me3 domains and role of EZH2 during pancreatic endocrine specification

PubMed Central

Xu, Cheng-Ran; Li, Lin-Chen; Donahue, Greg; Ying, Lei; Zhang, Yu-Wei; Gadue, Paul; Zaret, Kenneth S

2014-01-01

Endoderm cells undergo sequential fate choices to generate insulin-secreting beta cells. Ezh2 of the PRC2 complex, which generates H3K27me3, modulates the transition from endoderm to pancreas progenitors, but the role of Ezh2 and H3K27me3 in the next transition to endocrine progenitors is unknown. We isolated endoderm cells, pancreas progenitors, and endocrine progenitors from different staged mouse embryos and analyzed H3K27me3 genome-wide. Unlike the decline in H3K27me3 domains reported during embryonic stem cell differentiation in vitro, we find that H3K27me3 domains increase in number during endocrine progenitor development in vivo. Genes that lose the H3K27me3 mark typically encode transcriptional regulators, including those for pro-endocrine fates, whereas genes that acquire the mark typically are involved in cell biology and morphogenesis. Deletion of Ezh2 at the pancreas progenitor stage enhanced the production of endocrine progenitors and beta cells. Inhibition of EZH2 in embryonic pancreas explants and in human embryonic stem cell cultures increased endocrine progenitors in vitro. Our studies reveal distinct dynamics in H3K27me3 targets in vivo and a means to modulate beta cell development from stem cells. PMID:25107471

Mucositis and salivary antioxidants in patients undergoing bone marrow transplantation (BMT)

PubMed Central

Mazzeo, Marcelo A.; López, María M.; Linares, Jorge A.; Jarchum, Gustavo; Wietz, Fernando M.; Finkelberg, Ana B.

2014-01-01

Objectives: High doses of chemotherapy generate DNA damage in patients undergoing bone marrow transplantation (BMT), due to the production of reactive oxygen species (ROS). In order to evaluate the local defensive effectiveness of the patient undergoing BMT, the concentrations of the antioxidants superoxide dismutase (SOD) and uric acid (UA) were measured in saliva. Study Design: Basal saliva samples were collected from 20 patients undergoing BMT at the Oncology Department, Sanatorio Allende (Córdoba), in the stages: initial, prior to conditioning therapy (I); middle: 7 to 10 days after BMT (M) and final stage, 30 days after discharge from isolation (F). SOD levels were determined using a RANDOX kit (RANSOD superoxide dismutase manual), and for uric acid enzymatic UOD / PAP spectrophotometric method, ( Trinder Color Kit , Wiener Lab) was used. Results: 85% of the patients developed oral mucositis. SOD concentration in the M stage was significantly higher (p<0.01) compared with stage I, and it reversed in stage F. UA concentration was significantly lower (p<0.001) in stage M compared with stage I, and in stage F it recovered the initial values. Conclusions: SOD increase in stage M coincided with the appearance of mucositis, which could be interpreted as a defensive mechanism of saliva against oxidative stress produced by chemotherapy. UA decrease in stage M would favour the development of higher degrees of mucositis. Key words:Bone marrow transplantation, mucositis, superoxide dismutase, uric acid. PMID:24608218

Gangliosides in the Nervous System: Biosynthesis and Degradation

NASA Astrophysics Data System (ADS)

Yu, Robert K.; Ariga, Toshio; Yanagisawa, Makoto; Zeng, Guichao

Gangliosides, abundant in the nervous system, are known to play crucial modulatory roles in cellular recognition, interaction, adhesion, and signal transduction, particularly during early developmental stages. The expression of gangliosides in the nervous system is developmentally regulated and is closely related to the differentiation state of the cell. Ganglioside biosynthesis occurs in intracellular organelles, from which gangliosides are transported to the plasma membrane. During brain development, the ganglioside composition of the nervous system undergoes remarkable changes and is strictly regulated by the activities of glycosyltransferases, which can occur at different levels of control, including glycosyltransferase gene transcription and posttranslational modification. Genes for glycosyltransferase involved in ganglioside biosynthesis have been cloned and classified into families of glycosyltransferases based on their amino acid sequence similarities. The donor and acceptor substrate specificities are determined by enzymatic analysis of the glycosyltransferase gene products. Cell-type specific regulation of these genes has also been studied. Gangliosides are degraded by lysosomal exoglycosidases. The action of these enzymes occurs frequently in cooperation with activator proteins. Several human diseases are caused by defects of degradative enzymes, resulting in massive accumulation of certain glycolipids, including gangliosides in the lysosomal compartment and other organelles in the brain and visceral organs. Some of the representative lysosomal storage diseases (LSDs) caused by the accumulation of lipids in late endosomes and lysosomes will be discussed.

Expression profiles of genes involved in jasmonic acid biosynthesis and signaling during growth and development of carrot.

PubMed

Wang, Guanglong; Huang, Wei; Li, Mengyao; Xu, Zhisheng; Wang, Feng; Xiong, Aisheng

2016-09-01

Jasmonates (JAs) are recognized as essential regulators in response to environmental stimuli and plant development. Carrot is an Apiaceae vegetable with great value and undergoes significant size changes over the course of plant growth. However, JA accumulation and its potential roles in carrot growth remain unclear. Here, methyl JA (MeJA) levels and expression profiles of JA-related genes were analyzed in carrot roots and leaves at five developmental stages. MeJA levels in the roots and leaves were the highest at the first stage and decreased as carrot growth proceeded. Transcript levels of several JA-related genes (Dc13-LOX1, Dc13-LOX2, DcAOS, DcAOC, DcOPR2, DcOPR3, DcOPCL1, DcJAR1, DcJMT, DcCOI1, DcJAZ1, DcJAZ2, DcMYC2, DcCHIB/PR3, DcLEC, and DcVSP2) were not well correlated with MeJA accumulation during carrot root and leaf development. In addition, some JA-related genes (DcJAR1, DcJMT, DcCOI1, DcMYC2, and DcVSP2) showed differential expression between roots and leaves. These results suggest that JAs may regulate carrot plant growth in stage-dependent and organ-specific manners. Our work provides novel insights into JA accumulation and its potential roles during carrot growth and development. © The Author 2016. Published by Oxford University Press on behalf of the Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Regulation of Plasmodium yoelii oocyst development by strain- and stage-specific small-subunit rRNA.

PubMed

Qi, Yanwei; Zhu, Feng; Eastman, Richard T; Fu, Young; Zilversmit, Martine; Pattaradilokrat, Sittiporn; Hong, Lingxian; Liu, Shengfa; McCutchan, Thomas F; Pan, Weiqing; Xu, Wenyue; Li, Jian; Huang, Fusheng; Su, Xin-zhuan

2015-03-10

One unique feature of malaria parasites is the differential transcription of structurally distinct rRNA (rRNA) genes at different developmental stages: the A-type genes are transcribed mainly in asexual stages, whereas the S-type genes are expressed mostly in sexual or mosquito stages. Conclusive functional evidence of different rRNAs in regulating stage-specific parasite development, however, is still absent. Here we performed genetic crosses of Plasmodium yoelii parasites with one parent having an oocyst development defect (ODD) phenotype and another producing normal oocysts to identify the gene(s) contributing to the ODD. The parent with ODD--characterized as having small oocysts and lacking infective sporozoites--was obtained after introduction of a plasmid with a green fluorescent protein gene into the parasite genome and subsequent passages in mice. Quantitative trait locus analysis of genome-wide microsatellite genotypes of 48 progeny from the crosses linked an ~200-kb segment on chromosome 6 containing one of the S-type genes (D-type small subunit rRNA gene [D-ssu]) to the ODD. Fine mapping of the plasmid integration site, gene expression pattern, and gene knockout experiments demonstrated that disruption of the D-ssu gene caused the ODD phenotype. Interestingly, introduction of the D-ssu gene into the same parasite strain (self), but not into a different subspecies, significantly affected or completely ablated oocyst development, suggesting a stage- and subspecies (strain)-specific regulation of oocyst development by D-ssu. This study demonstrates that P. yoelii D-ssu is essential for normal oocyst and sporozoite development and that variation in the D-ssu sequence can have dramatic effects on parasite development. Malaria parasites are the only known organisms that express structurally distinct rRNA genes at different developmental stages. The differential expression of these genes suggests that they play unique roles during the complex life cycle of the parasites. Conclusive functional proof of different rRNAs in regulating parasite development, however, is still absent or controversial. Here we functionally demonstrate for the first time that a stage-specifically expressed D-type small-subunit rRNA gene (D-ssu) is essential for oocyst development of the malaria parasite Plasmodium yoelii in the mosquito. This study also shows that variations in D-ssu sequence and/or the timing of transcription may have profound effects on parasite oocyst development. The results show that in addition to protein translation, rRNAs of malaria parasites also regulate parasite development and differentiation in a strain-specific manner, which can be explored for controlling parasite transmission. Copyright © 2015 Qi et al.

Identification of a myometrial molecular profile for dystocic labor

PubMed Central

2011-01-01

Background The most common indication for cesarean section (CS) in nulliparous women is dystocia secondary to ineffective myometrial contractility. The aim of this study was to identify a molecular profile in myometrium associated with dystocic labor. Methods Myometrial biopsies were obtained from the upper incisional margins of nulliparous women undergoing lower segment CS for dystocia (n = 4) and control women undergoing CS in the second stage who had demonstrated efficient uterine action during the first stage of labor (n = 4). All patients were in spontaneous (non-induced) labor and had received intrapartum oxytocin to accelerate labor. RNA was extracted from biopsies and hybridized to Affymetrix HuGene U133A Plus 2 microarrays. Internal validation was performed using quantitative SYBR Green Real-Time PCR. Results Seventy genes were differentially expressed between the two groups. 58 genes were down-regulated in the dystocia group. Gene ontology analysis revealed 12 of the 58 down-regulated genes were involved in the immune response. These included (ERAP2, (8.67 fold change (FC)) HLA-DQB1 (7.88 FC) CD28 (2.60 FC), LILRA3 (2.87 FC) and TGFBR3 (2.1 FC)) Hierarchical clustering demonstrated a difference in global gene expression patterns between the samples from dystocic and non-dystocic labours. RT-PCR validation was performed on 4 genes ERAP2, CD28, LILRA3 and TGFBR3 Conclusion These findings suggest an underlying molecular basis for dystocia in nulliparous women in spontaneous labor. Differentially expressed genes suggest an important role for the immune response in dystocic labor and may provide important indicators for new diagnostic assays and potential intrapartum therapeutic targets. PMID:21999197

Toxicity of silver nanoparticles in mouse embryonic stem cells and chemical based reprogramming of somatic cells to sphere cells

NASA Astrophysics Data System (ADS)

Rajanahalli Krishnamurthy, Pavan

Abstract 1: Silver nanoparticles (Ag Np's) have an interesting surface chemistry and unique plasmonic properties. They are used in a wide variety of applications ranging from consumer products like socks, medical dressing, computer chips and it is also shown to have antimicrobial, anti bacterial activity and wound healing. Ag Np toxicity studies have been limited to date which needs to be critically addressed due to its wide applications. Mouse embryonic stem (MES) cells represent a unique cell population with the ability to undergo both self renewal and differentiation. They exhibit very stringent and tightly regulated mechanisms to circumvent DNA damage and stress response. We used 10 nm coated (polysaccharide) and uncoated Ag Np's to test its toxic effects on MES cells. MES cells and embryoid bodies (EB's) were treated with two concentrations of Ag Np's: 5 microg/ml and 50 ug/ml and exposed for 24, 48 and 72 hours. Increased cell death, ROS production and loss of mitochondrial membrane potential and alkaline phosphatase (AP) occur in a time and a concentration dependant manner. Due to increased cell death, there is a progressive increase in Annexin V (apoptosis) and Propidium Iodide (PI) staining (necrosis). Oct4 and Nanog undergo ubiquitination and dephosphorylation post-translational modifications in MES cells thereby altering gene expression of pluripotency factors and differentiation of EB's into all the three embryonic germ layers with specific growth factors were also inhibited after Ag Np exposure. Flow cytometry analysis revealed Ag Np's treated cells had altered cell cycle phases correlating with altered self renewal capacity. Our results suggest that Ag Np's effect MES cell self renewal, pluripotency and differentiation and serves as a perfect model system for studying toxicity induced by engineered Ag Np's. Abstract 2: The reprogramming of fibroblasts to pluripotent stem cells and the direct conversion of fibroblasts to functional neurons has been successfully manipulated by ectopic expression of defined factors. We demonstrate that mouse fibroblasts can be converted into sphere cells by detaching fibroblast cells by proteases and then using AlbuMAX I-containing culture medium without genetic alteration. AlbuMAX I is a lipid-rich albumin. Albumin-associated lipids arachidonic acid (AA) and pluronic F-68 were responsible for this effect. The converted colonies were positive for both alkaline phosphatase and stage specific embryonic antigen-1 (SSEA-1) staining. Global gene expression analysis indicated that the sphere cells were in an intermediate state compared with MES cells and MEF cells. The sphere cells were able to differentiate into tissues representing all three embryonic germ layers following retinoic acid treatment, and also differentiated into smooth muscle cells following treatment with vascular endothelial growth factor (VEGF). The study presented a potential novel approach to transdifferentiate mouse fibroblast cells into other cell lineages mediated by AlbuMAX I-containing culture medium.

Comprehensive Cell-specific Protein Analysis in Early and Late Pollen Development from Diploid Microsporocytes to Pollen Tube Growth*

PubMed Central

Ischebeck, Till; Valledor, Luis; Lyon, David; Gingl, Stephanie; Nagler, Matthias; Meijón, Mónica; Egelhofer, Volker; Weckwerth, Wolfram

2014-01-01

Pollen development in angiosperms is one of the most important processes controlling plant reproduction and thus productivity. At the same time, pollen development is highly sensitive to environmental fluctuations, including temperature, drought, and nutrition. Therefore, pollen biology is a major focus in applied studies and breeding approaches for improving plant productivity in a globally changing climate. The most accessible developmental stages of pollen are the mature pollen and the pollen tubes, and these are thus most frequently analyzed. To reveal a complete quantitative proteome map, we additionally addressed the very early stages, analyzing eight stages of tobacco pollen development: diploid microsporocytes, meiosis, tetrads, microspores, polarized microspores, bipolar pollen, desiccated pollen, and pollen tubes. A protocol for the isolation of the early stages was established. Proteins were extracted and analyzed by means of a new gel LC-MS fractionation protocol. In total, 3817 protein groups were identified. Quantitative analysis was performed based on peptide count. Exceedingly stage-specific differential protein regulation was observed during the conversion from the sporophytic to the gametophytic proteome. A map of highly specialized functionality for the different stages could be revealed from the metabolic activity and pronounced differentiation of proteasomal and ribosomal protein complex composition up to protective mechanisms such as high levels of heat shock proteins in the very early stages of development. PMID:24078888

Comprehensive cell-specific protein analysis in early and late pollen development from diploid microsporocytes to pollen tube growth.

PubMed

Ischebeck, Till; Valledor, Luis; Lyon, David; Gingl, Stephanie; Nagler, Matthias; Meijón, Mónica; Egelhofer, Volker; Weckwerth, Wolfram

2014-01-01

Pollen development in angiosperms is one of the most important processes controlling plant reproduction and thus productivity. At the same time, pollen development is highly sensitive to environmental fluctuations, including temperature, drought, and nutrition. Therefore, pollen biology is a major focus in applied studies and breeding approaches for improving plant productivity in a globally changing climate. The most accessible developmental stages of pollen are the mature pollen and the pollen tubes, and these are thus most frequently analyzed. To reveal a complete quantitative proteome map, we additionally addressed the very early stages, analyzing eight stages of tobacco pollen development: diploid microsporocytes, meiosis, tetrads, microspores, polarized microspores, bipolar pollen, desiccated pollen, and pollen tubes. A protocol for the isolation of the early stages was established. Proteins were extracted and analyzed by means of a new gel LC-MS fractionation protocol. In total, 3817 protein groups were identified. Quantitative analysis was performed based on peptide count. Exceedingly stage-specific differential protein regulation was observed during the conversion from the sporophytic to the gametophytic proteome. A map of highly specialized functionality for the different stages could be revealed from the metabolic activity and pronounced differentiation of proteasomal and ribosomal protein complex composition up to protective mechanisms such as high levels of heat shock proteins in the very early stages of development.

[Monoclonal antibodies in diagnosis of acute leukemias].

PubMed

Krawczyńska, A; Robak, T

1996-01-01

Immunophenotyping has become an essential component for the study of acute myeloblastic (AML) and lymphoblastic (ALL) leukaemias. The recent development of highly specific monoclonal antibodies (Mc Ab) to differentiation antigens (CD) of haematopoetic cells have made it readily available to clinical laboratories in most major hospitals. Immunophenotyping complements standard morphology by providing information on lineage, stage of differentiation and clonality. In addition some of the flow cytometry findings have independent prognostic significance. Monoclonal antibodies useful in defining lineage (B-cell versus T-cell) and stages of differentiation of ALL. It can be also used in identifying characteristic feature of AML and aiding in lineage determination in acute leukaemias that are morphologically undifferentiated. Surface immunophenotyping is especially helpful for recognizing mixed lineage acute leukaemia and diagnosing certain rare entities such as erythroleukaemia (M6), acute megakaryocytic leukaemia (M7) and minimally differentiation acute myeloid leukaemia.

Host-secreted antimicrobial peptide enforces symbiotic selectivity in Medicago truncatula.

PubMed

Wang, Qi; Yang, Shengming; Liu, Jinge; Terecskei, Kata; Ábrahám, Edit; Gombár, Anikó; Domonkos, Ágota; Szűcs, Attila; Körmöczi, Péter; Wang, Ting; Fodor, Lili; Mao, Linyong; Fei, Zhangjun; Kondorosi, Éva; Kaló, Péter; Kereszt, Attila; Zhu, Hongyan

2017-06-27

Legumes engage in root nodule symbioses with nitrogen-fixing soil bacteria known as rhizobia. In nodule cells, bacteria are enclosed in membrane-bound vesicles called symbiosomes and differentiate into bacteroids that are capable of converting atmospheric nitrogen into ammonia. Bacteroid differentiation and prolonged intracellular survival are essential for development of functional nodules. However, in the Medicago truncatula - Sinorhizobium meliloti symbiosis, incompatibility between symbiotic partners frequently occurs, leading to the formation of infected nodules defective in nitrogen fixation (Fix - ). Here, we report the identification and cloning of the M. truncatula NFS2 gene that regulates this type of specificity pertaining to S. meliloti strain Rm41. We demonstrate that NFS2 encodes a nodule-specific cysteine-rich (NCR) peptide that acts to promote bacterial lysis after differentiation. The negative role of NFS2 in symbiosis is contingent on host genetic background and can be counteracted by other genes encoded by the host. This work extends the paradigm of NCR function to include the negative regulation of symbiotic persistence in host-strain interactions. Our data suggest that NCR peptides are host determinants of symbiotic specificity in M. truncatula and possibly in closely related legumes that form indeterminate nodules in which bacterial symbionts undergo terminal differentiation.

Quantitative expression patterns of GDF9 and BMP15 genes in sheep ovarian follicles grown in vivo or cultured in vitro.

PubMed

Kona, S S R; Praveen Chakravarthi, V; Siva Kumar, A V N; Srividya, D; Padmaja, K; Rao, V H

2016-01-15

Quantitative patterns of expression of the growth differentiation factor 9 (GDF9) and bone morphogenic protein 15 (BMP15) genes in different development stages of in vivo and in vitro grown ovarian follicles in sheep were studied for the first time. Both GDF9 and BMP15 were expressed in the cumulus cells and oocytes at all the development stages of in vivo and in vitro grown ovarian follicles. Growth differentiation factor 9 and bone morphogenic protein 15 exhibited stage-specific undulations in the expression in the cumulus cells and oocytes isolated from in vivo grown ovarian follicles. These undulations could be related to discrete development events during the ovarian follicle development. The expression of GDF9 and BMP15 was highest (3.38 ± 0.02 and 2.69 ± 0.06, respectively; P ≤ 0.05) in the primordial follicles compared with preantral, early antral, antral, and large antral stages. Similarly, GDF9 and BMP15 expression in the cumulus cells (0 ± 0.16 and 0 ± 0.07) and oocytes (1.47 ± 0.07 and 1.32 ± 0.03) was lowest (P ≤ 0.05) in the in vivo grown antral follicles. In the cultured follicles, the stage-specific undulations observed in the expression of GDF9 and BMP15 in the in vivo grown follicles were either different or abolished. For example, in the oocytes from in vitro grown follicles, the expression of BMP15 did not change as the development progressed all the way from preantral to large antral follicle stage although in the oocytes from in vivo grown follicles BMP15 expression exhibited stage-specific variations. It is concluded that GDF9 and BMP15 follow a stage-specific pattern of expression during the in vivo development of ovarian follicles in sheep, and in vitro culture altered the stage-specific changes in the expression of these two genes. Copyright © 2016 Elsevier Inc. All rights reserved.

Origin and specification of type II neuroblasts in the Drosophila embryo.

PubMed

Álvarez, José-Andrés; Díaz-Benjumea, Fernando J

2018-04-05

In Drosophila , neural stem cells or neuroblasts (NBs) acquire different identities according to their site of origin in the embryonic neuroectoderm. Their identity determines the number of times they will divide and the types of daughter cells they will generate. All NBs divide asymmetrically, with type I NBs undergoing self-renewal and generating another cell that will divide only once more. By contrast, a small set of NBs in the larval brain, type II NBs, divides differently, undergoing self-renewal and generating an intermediate neural progenitor (INP) that continues to divide asymmetrically several more times, generating larger lineages. In this study, we have analysed the origin of type II NBs and how they are specified. Our results indicate that these cells originate in three distinct clusters in the dorsal protocerebrum during stage 12 of embryonic development. Moreover, it appears that their specification requires the combined action of EGFR signalling and the activity of the related genes buttonhead and Drosophila Sp1 In addition, we also show that the INPs generated in the embryo enter quiescence at the end of embryogenesis, resuming proliferation during the larval stage. © 2018. Published by The Company of Biologists Ltd.

Development of endosperm transfer cells in barley.

PubMed

Thiel, Johannes

2014-01-01

Endosperm transfer cells (ETCs) are positioned at the intersection of maternal and filial tissues in seeds of cereals and represent a bottleneck for apoplasmic transport of assimilates into the endosperm. Endosperm cellularization starts at the maternal-filial boundary and generates the highly specialized ETCs. During differentiation barley ETCs develop characteristic flange-like wall ingrowths to facilitate effective nutrient transfer. A comprehensive morphological analysis depicted distinct developmental time points in establishment of transfer cell (TC) morphology and revealed intracellular changes possibly associated with cell wall metabolism. Embedded inside the grain, ETCs are barely accessible by manual preparation. To get tissue-specific information about ETC specification and differentiation, laser microdissection (LM)-based methods were used for transcript and metabolite profiling. Transcriptome analysis of ETCs at different developmental stages by microarrays indicated activated gene expression programs related to control of cell proliferation and cell shape, cell wall and carbohydrate metabolism reflecting the morphological changes during early ETC development. Transporter genes reveal distinct expression patterns suggesting a switch from active to passive modes of nutrient uptake with the onset of grain filling. Tissue-specific RNA-seq of the differentiating ETC region from the syncytial stage until functionality in nutrient transfer identified a high number of novel transcripts putatively involved in ETC differentiation. An essential role for two-component signaling (TCS) pathways in ETC development of barley emerged from this analysis. Correlative data provide evidence for abscisic acid and ethylene influences on ETC differentiation and hint at a crosstalk between hormone signal transduction and TCS phosphorelays. Collectively, the data expose a comprehensive view on ETC development, associated pathways and identified candidate genes for ETC specification.

Development of endosperm transfer cells in barley

PubMed Central

Thiel, Johannes

2014-01-01

Endosperm transfer cells (ETCs) are positioned at the intersection of maternal and filial tissues in seeds of cereals and represent a bottleneck for apoplasmic transport of assimilates into the endosperm. Endosperm cellularization starts at the maternal-filial boundary and generates the highly specialized ETCs. During differentiation barley ETCs develop characteristic flange-like wall ingrowths to facilitate effective nutrient transfer. A comprehensive morphological analysis depicted distinct developmental time points in establishment of transfer cell (TC) morphology and revealed intracellular changes possibly associated with cell wall metabolism. Embedded inside the grain, ETCs are barely accessible by manual preparation. To get tissue-specific information about ETC specification and differentiation, laser microdissection (LM)-based methods were used for transcript and metabolite profiling. Transcriptome analysis of ETCs at different developmental stages by microarrays indicated activated gene expression programs related to control of cell proliferation and cell shape, cell wall and carbohydrate metabolism reflecting the morphological changes during early ETC development. Transporter genes reveal distinct expression patterns suggesting a switch from active to passive modes of nutrient uptake with the onset of grain filling. Tissue-specific RNA-seq of the differentiating ETC region from the syncytial stage until functionality in nutrient transfer identified a high number of novel transcripts putatively involved in ETC differentiation. An essential role for two-component signaling (TCS) pathways in ETC development of barley emerged from this analysis. Correlative data provide evidence for abscisic acid and ethylene influences on ETC differentiation and hint at a crosstalk between hormone signal transduction and TCS phosphorelays. Collectively, the data expose a comprehensive view on ETC development, associated pathways and identified candidate genes for ETC specification. PMID:24723929

Transcriptome Analysis of the Arabidopsis Megaspore Mother Cell Uncovers the Importance of RNA Helicases for Plant Germline Development

PubMed Central

Schmidt, Anja; Wuest, Samuel E.; Vijverberg, Kitty; Baroux, Célia; Kleen, Daniela; Grossniklaus, Ueli

2011-01-01

Germ line specification is a crucial step in the life cycle of all organisms. For sexual plant reproduction, the megaspore mother cell (MMC) is of crucial importance: it marks the first cell of the plant “germline” lineage that gets committed to undergo meiosis. One of the meiotic products, the functional megaspore, subsequently gives rise to the haploid, multicellular female gametophyte that harbours the female gametes. The MMC is formed by selection and differentiation of a single somatic, sub-epidermal cell in the ovule. The transcriptional network underlying MMC specification and differentiation is largely unknown. We provide the first transcriptome analysis of an MMC using the model plant Arabidopsis thaliana with a combination of laser-assisted microdissection and microarray hybridizations. Statistical analyses identified an over-representation of translational regulation control pathways and a significant enrichment of DEAD/DEAH-box helicases in the MMC transcriptome, paralleling important features of the animal germline. Analysis of two independent T-DNA insertion lines suggests an important role of an enriched helicase, MNEME (MEM), in MMC differentiation and the restriction of the germline fate to only one cell per ovule primordium. In heterozygous mem mutants, additional enlarged MMC-like cells, which sometimes initiate female gametophyte development, were observed at higher frequencies than in the wild type. This closely resembles the phenotype of mutants affected in the small RNA and DNA-methylation pathways important for epigenetic regulation. Importantly, the mem phenotype shows features of apospory, as female gametophytes initiate from two non-sister cells in these mutants. Moreover, in mem gametophytic nuclei, both higher order chromatin structure and the distribution of LIKE HETEROCHROMATIN PROTEIN1 were affected, indicating epigenetic perturbations. In summary, the MMC transcriptome sets the stage for future functional characterization as illustrated by the identification of MEM, a novel gene involved in the restriction of germline fate. PMID:21949639

Long range epigenetic silencing is a trans-species mechanism that results in cancer specific deregulation by overriding the chromatin domains of normal cells.

PubMed

Forn, Marta; Muñoz, Mar; Tauriello, Daniele V F; Merlos-Suárez, Anna; Rodilla, Verónica; Bigas, Anna; Batlle, Eduard; Jordà, Mireia; Peinado, Miguel A

2013-12-01

DNA methylation and chromatin remodeling are frequently implicated in the silencing of genes involved in carcinogenesis. Long Range Epigenetic Silencing (LRES) is a mechanism of gene inactivation that affects multiple contiguous CpG islands and has been described in different human cancer types. However, it is unknown whether there is a coordinated regulation of the genes embedded in these regions in normal cells and in early stages of tumor progression. To better characterize the molecular events associated with the regulation and remodeling of these regions we analyzed two regions undergoing LRES in human colon cancer in the mouse model. We demonstrate that LRES also occurs in murine cancer in vivo and mimics the molecular features of the human phenomenon, namely, downregulation of gene expression, acquisition of inactive histone marks, and DNA hypermethylation of specific CpG islands. The genes embedded in these regions showed a dynamic and autonomous regulation during mouse intestinal cell differentiation, indicating that, in the framework considered here, the coordinated regulation in LRES is restricted to cancer. Unexpectedly, benign adenomas in Apc(Min/+) mice showed overexpression of most of the genes affected by LRES in cancer, which suggests that the repressive remodeling of the region is a late event. Chromatin immunoprecipitation analysis of the transcriptional insulator CTCF in mouse colon cancer cells revealed disrupted chromatin domain boundaries as compared with normal cells. Malignant regression of cancer cells by in vitro differentiation resulted in partial reversion of LRES and gain of CTCF binding. We conclude that genes in LRES regions are plastically regulated in cell differentiation and hyperproliferation, but are constrained to a coordinated repression by abolishing boundaries and the autonomous regulation of chromatin domains in cancer cells. Copyright © 2013 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

[Identification of genes that are specifically/preferentially expressed in developing cotton fibers by mRNA fluorescence differential display (FDD)].

PubMed

Sun, Jie; Li, Yuan-Li; Wang, Ruo-Hai; Xia, Gui-Xian

2004-01-01

Fluorescence differential display (FDD) technique was used to identify genes that are specifically or preferentially expressed in different developmental stages of cotton fiber cells. One hundred and nine differentially displayed cDNA fragments were isolated using 9, 21 and 27 DPA (days postanthesis) fibers as experimental materials. By a combination of two rounds of reverse Northern hybridization and Northern blot analyses, a number of such cDNA fragments were proved to represent fiber-specific/preferential genes. Sequencing determination and database searching indicated that most of these genes are novel. This work is an important step towards cloning the full-length cDNAs and characterizing the cellular functions of aforementioned genes in fiber development.

Systematically labeling developmental stage-specific genes for the study of pancreatic β-cell differentiation from human embryonic stem cells.

PubMed

Liu, Haisong; Yang, Huan; Zhu, Dicong; Sui, Xin; Li, Juan; Liang, Zhen; Xu, Lei; Chen, Zeyu; Yao, Anzhi; Zhang, Long; Zhang, Xi; Yi, Xing; Liu, Meng; Xu, Shiqing; Zhang, Wenjian; Lin, Hua; Xie, Lan; Lou, Jinning; Zhang, Yong; Xi, Jianzhong; Deng, Hongkui

2014-10-01

The applications of human pluripotent stem cell (hPSC)-derived cells in regenerative medicine has encountered a long-standing challenge: how can we efficiently obtain mature cell types from hPSCs? Attempts to address this problem are hindered by the complexity of controlling cell fate commitment and the lack of sufficient developmental knowledge for guiding hPSC differentiation. Here, we developed a systematic strategy to study hPSC differentiation by labeling sequential developmental genes to encompass the major developmental stages, using the directed differentiation of pancreatic β cells from hPSCs as a model. We therefore generated a large panel of pancreas-specific mono- and dual-reporter cell lines. With this unique platform, we visualized the kinetics of the entire differentiation process in real time for the first time by monitoring the expression dynamics of the reporter genes, identified desired cell populations at each differentiation stage and demonstrated the ability to isolate these cell populations for further characterization. We further revealed the expression profiles of isolated NGN3-eGFP(+) cells by RNA sequencing and identified sushi domain-containing 2 (SUSD2) as a novel surface protein that enriches for pancreatic endocrine progenitors and early endocrine cells both in human embryonic stem cells (hESC)-derived pancreatic cells and in the developing human pancreas. Moreover, we captured a series of cell fate transition events in real time, identified multiple cell subpopulations and unveiled their distinct gene expression profiles, among heterogeneous progenitors for the first time using our dual reporter hESC lines. The exploration of this platform and our new findings will pave the way to obtain mature β cells in vitro.

Analysis of global gene expression profiles to identify differentially expressed genes critical for embryo development in Brassica rapa.

PubMed

Zhang, Yu; Peng, Lifang; Wu, Ya; Shen, Yanyue; Wu, Xiaoming; Wang, Jianbo

2014-11-01

Embryo development represents a crucial developmental period in the life cycle of flowering plants. To gain insights into the genetic programs that control embryo development in Brassica rapa L., RNA sequencing technology was used to perform transcriptome profiling analysis of B. rapa developing embryos. The results generated 42,906,229 sequence reads aligned with 32,941 genes. In total, 27,760, 28,871, 28,384, and 25,653 genes were identified from embryos at globular, heart, early cotyledon, and mature developmental stages, respectively, and analysis between stages revealed a subset of stage-specific genes. We next investigated 9,884 differentially expressed genes with more than fivefold changes in expression and false discovery rate ≤ 0.001 from three adjacent-stage comparisons; 1,514, 3,831, and 6,633 genes were detected between globular and heart stage embryo libraries, heart stage and early cotyledon stage, and early cotyledon and mature stage, respectively. Large numbers of genes related to cellular process, metabolism process, response to stimulus, and biological process were expressed during the early and middle stages of embryo development. Fatty acid biosynthesis, biosynthesis of secondary metabolites, and photosynthesis-related genes were expressed predominantly in embryos at the middle stage. Genes for lipid metabolism and storage proteins were highly expressed in the middle and late stages of embryo development. We also identified 911 transcription factor genes that show differential expression across embryo developmental stages. These results increase our understanding of the complex molecular and cellular events during embryo development in B. rapa and provide a foundation for future studies on other oilseed crops.

Biomechanics of the Chick Embryonic Heart Outflow Tract at HH18 Using 4D Optical Coherence Tomography Imaging and Computational Modeling

PubMed Central

Liu, Aiping; Yin, Xin; Shi, Liang; Li, Peng; Thornburg, Kent L.; Wang, Ruikang; Rugonyi, Sandra

2012-01-01

During developmental stages, biomechanical stimuli on cardiac cells modulate genetic programs, and deviations from normal stimuli can lead to cardiac defects. Therefore, it is important to characterize normal cardiac biomechanical stimuli during early developmental stages. Using the chicken embryo model of cardiac development, we focused on characterizing biomechanical stimuli on the Hamburger–Hamilton (HH) 18 chick cardiac outflow tract (OFT), the distal portion of the heart from which a large portion of defects observed in humans originate. To characterize biomechanical stimuli in the OFT, we used a combination of in vivo optical coherence tomography (OCT) imaging, physiological measurements and computational fluid dynamics (CFD) modeling. We found that, at HH18, the proximal portion of the OFT wall undergoes larger circumferential strains than its distal portion, while the distal portion of the OFT wall undergoes larger wall stresses. Maximal wall shear stresses were generally found on the surface of endocardial cushions, which are protrusions of extracellular matrix onto the OFT lumen that later during development give rise to cardiac septa and valves. The non-uniform spatial and temporal distributions of stresses and strains in the OFT walls provide biomechanical cues to cardiac cells that likely aid in the extensive differential growth and remodeling patterns observed during normal development. PMID:22844414

Bioactive treatment promotes osteoblast differentiation on titanium materials fabricated by selective laser melting technology.

PubMed

Tsukanaka, Masako; Fujibayashi, Shunsuke; Takemoto, Mitsuru; Matsushita, Tomiharu; Kokubo, Tadashi; Nakamura, Takashi; Sasaki, Kiyoyuki; Matsuda, Shuichi

2016-01-01

Selective laser melting (SLM) technology is useful for the fabrication of porous titanium implants with complex shapes and structures. The materials fabricated by SLM characteristically have a very rough surface (average surface roughness, Ra=24.58 µm). In this study, we evaluated morphologically and biochemically the specific effects of this very rough surface and the additional effects of a bioactive treatment on osteoblast proliferation and differentiation. Flat-rolled titanium materials (Ra=1.02 µm) were used as the controls. On the treated materials fabricated by SLM, we observed enhanced osteoblast differentiation compared with the flat-rolled materials and the untreated materials fabricated by SLM. No significant differences were observed between the flat-rolled materials and the untreated materials fabricated by SLM in their effects on osteoblast differentiation. We concluded that the very rough surface fabricated by SLM had to undergo a bioactive treatment to obtain a positive effect on osteoblast differentiation.

A sequential EMT-MET mechanism drives the differentiation of human embryonic stem cells towards hepatocytes.

PubMed

Li, Qiuhong; Hutchins, Andrew P; Chen, Yong; Li, Shengbiao; Shan, Yongli; Liao, Baojian; Zheng, Dejin; Shi, Xi; Li, Yinxiong; Chan, Wai-Yee; Pan, Guangjin; Wei, Shicheng; Shu, Xiaodong; Pei, Duanqing

2017-05-03

Reprogramming has been shown to involve EMT-MET; however, its role in cell differentiation is unclear. We report here that in vitro differentiation of hESCs to hepatic lineage undergoes a sequential EMT-MET with an obligatory intermediate mesenchymal phase. Gene expression analysis reveals that Activin A-induced formation of definitive endoderm (DE) accompanies a synchronous EMT mediated by autocrine TGFβ signalling followed by a MET process. Pharmacological inhibition of TGFβ signalling blocks the EMT as well as DE formation. We then identify SNAI1 as the key EMT transcriptional factor required for the specification of DE. Genetic ablation of SNAI1 in hESCs does not affect the maintenance of pluripotency or neural differentiation, but completely disrupts the formation of DE. These results reveal a critical mesenchymal phase during the acquisition of DE, highlighting a role for sequential EMT-METs in both differentiation and reprogramming.

Antagonism between the transcription factors NANOG and OTX2 specifies rostral or caudal cell fate during neural patterning transition.

PubMed

Su, Zhenghui; Zhang, Yanqi; Liao, Baojian; Zhong, Xiaofen; Chen, Xin; Wang, Haitao; Guo, Yiping; Shan, Yongli; Wang, Lihui; Pan, Guangjin

2018-03-23

During neurogenesis, neural patterning is a critical step during which neural progenitor cells differentiate into neurons with distinct functions. However, the molecular determinants that regulate neural patterning remain poorly understood. Here we optimized the "dual SMAD inhibition" method to specifically promote differentiation of human pluripotent stem cells (hPSCs) into forebrain and hindbrain neural progenitor cells along the rostral-caudal axis. We report that neural patterning determination occurs at the very early stage in this differentiation. Undifferentiated hPSCs expressed basal levels of the transcription factor orthodenticle homeobox 2 (OTX2) that dominantly drove hPSCs into the "default" rostral fate at the beginning of differentiation. Inhibition of glycogen synthase kinase 3β (GSK3β) through CHIR99021 application sustained transient expression of the transcription factor NANOG at early differentiation stages through Wnt signaling. Wnt signaling and NANOG antagonized OTX2 and, in the later stages of differentiation, switched the default rostral cell fate to the caudal one. Our findings have uncovered a mutual antagonism between NANOG and OTX2 underlying cell fate decisions during neural patterning, critical for the regulation of early neural development in humans. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

Differential gene expression patterns in the autogamous plant Hordeum euclaston (Poaceae).

PubMed

Georg-Kraemer, J E; Ferreira, C A S; Cavalli, S S

2011-02-22

Sib-seedlings of 95 strains of the strictly autogamous grass Hordeum euclaston were analyzed by horizontal polyacrylamide gel electrophoresis for four isoenzyme systems at a specific ontogenetic stage. We found differences in the activity of some genes among individuals of this species. Hence, an ontogenetic analysis was carried out to investigate 12 strains at five ontogenetic stages, to determine the patterns of expression of these genes during development. The differences in the presence versus absence of certain isoenzyme bands may be due to differential regulatory activation in response to environmental differences, as all plants showed the same structural genes, although these genes were active in different tissues and/or times of development. These results indicate the importance of differential gene activation in the metabolic phenotype variability of this strictly autogamous, highly homozygous species. The same structural alleles for isoenzymes showed the active form of the enzymes (phenotypic expression) to be present in different tissues and/or stages of development. Differential isoenzyme gene activation was shown to be directly responsible for the enzymatic variability (metabolic phenotype) presented by the plants, which seem to possess almost no heterozygosis.

Rapamycin and CHIR99021 Coordinate Robust Cardiomyocyte Differentiation From Human Pluripotent Stem Cells Via Reducing p53-Dependent Apoptosis.

PubMed

Qiu, Xiao-Xu; Liu, Yang; Zhang, Yi-Fan; Guan, Ya-Na; Jia, Qian-Qian; Wang, Chen; Liang, He; Li, Yong-Qin; Yang, Huang-Tian; Qin, Yong-Wen; Huang, Shuang; Zhao, Xian-Xian; Jing, Qing

2017-10-02

Cardiomyocytes differentiated from human pluripotent stem cells can serve as an unexhausted source for a cellular cardiac disease model. Although small molecule-mediated cardiomyocyte differentiation methods have been established, the differentiation efficiency is relatively unsatisfactory in multiple lines due to line-to-line variation. Additionally, hurdles including line-specific low expression of endogenous growth factors and the high apoptotic tendency of human pluripotent stem cells also need to be overcome to establish robust and efficient cardiomyocyte differentiation. We used the H9-human cardiac troponin T-eGFP reporter cell line to screen for small molecules that promote cardiac differentiation in a monolayer-based and growth factor-free differentiation model. We found that collaterally treating human pluripotent stem cells with rapamycin and CHIR99021 during the initial stage was essential for efficient and reliable cardiomyocyte differentiation. Moreover, this method maintained consistency in efficiency across different human embryonic stem cell and human induced pluripotent stem cell lines without specifically optimizing multiple parameters (the efficiency in H7, H9, and UQ1 human induced pluripotent stem cells is 98.3%, 93.3%, and 90.6%, respectively). This combination also increased the yield of cardiomyocytes (1:24) and at the same time reduced medium consumption by about 50% when compared with the previous protocols. Further analysis indicated that inhibition of the mammalian target of rapamycin allows efficient cardiomyocyte differentiation through overcoming p53-dependent apoptosis of human pluripotent stem cells during high-density monolayer culture via blunting p53 translation and mitochondrial reactive oxygen species production. We have demonstrated that mammalian target of rapamycin exerts a stage-specific and multifaceted regulation over cardiac differentiation and provides an optimized approach for generating large numbers of functional cardiomyocytes for disease modeling and in vitro drug screening. © 2017 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley.

Efficient stage-specific differentiation of human pluripotent stem cells toward retinal photoreceptor cells.

PubMed

Mellough, Carla B; Sernagor, Evelyne; Moreno-Gimeno, Inmaculada; Steel, David H W; Lako, Majlinda

2012-04-01

Recent successes in the stem cell field have identified some of the key chemical and biological cues which drive photoreceptor derivation from human embryonic stem cells (hESC) and human induced pluripotent stem cells (hiPSC); however, the efficiency of this process is variable. We have designed a three-step photoreceptor differentiation protocol combining previously published methods that direct the differentiation of hESC and hiPSC toward a retinal lineage, which we further modified with additional supplements selected on the basis of reports from the eye field and retinal development. We report that hESC and hiPSC differentiating under our regimen over a 60 day period sequentially acquire markers associated with neural, retinal field, retinal pigmented epithelium and photoreceptor cells, including mature photoreceptor markers OPN1SW and RHODOPSIN with a higher efficiency than previously reported. In addition, we report the ability of hESC and hiPSC cultures to generate neural and retinal phenotypes under minimal culture conditions, which may be linked to their ability to endogenously upregulate the expression of a range of factors important for retinal cell type specification. However, cultures that were differentiated with full supplementation under our photoreceptor-induction regimen achieve this within a significantly shorter time frame and show a substantial increase in the expression of photoreceptor-specific markers in comparison to cultures differentiated under minimal conditions. Interestingly, cultures supplemented only with B27 and/or N2 displayed comparable differentiation efficiency to those under full supplementation, indicating a key role for B27 and N2 during the differentiation process. Furthermore, our data highlight an important role for Dkk1 and Noggin in enhancing the differentiation of hESC and hiPSC toward retinal progenitor cells and photoreceptor precursors during the early stages of differentiation, while suggesting that further maturation of these cells into photoreceptors may not require additional factors and can ensue under minimal culture conditions. Copyright © 2012 AlphaMed Press.

Root hydrotropism is controlled via a cortex-specific growth mechanism.

PubMed

Dietrich, Daniela; Pang, Lei; Kobayashi, Akie; Fozard, John A; Boudolf, Véronique; Bhosale, Rahul; Antoni, Regina; Nguyen, Tuan; Hiratsuka, Sotaro; Fujii, Nobuharu; Miyazawa, Yutaka; Bae, Tae-Woong; Wells, Darren M; Owen, Markus R; Band, Leah R; Dyson, Rosemary J; Jensen, Oliver E; King, John R; Tracy, Saoirse R; Sturrock, Craig J; Mooney, Sacha J; Roberts, Jeremy A; Bhalerao, Rishikesh P; Dinneny, José R; Rodriguez, Pedro L; Nagatani, Akira; Hosokawa, Yoichiroh; Baskin, Tobias I; Pridmore, Tony P; De Veylder, Lieven; Takahashi, Hideyuki; Bennett, Malcolm J

2017-05-08

Plants can acclimate by using tropisms to link the direction of growth to environmental conditions. Hydrotropism allows roots to forage for water, a process known to depend on abscisic acid (ABA) but whose molecular and cellular basis remains unclear. Here we show that hydrotropism still occurs in roots after laser ablation removed the meristem and root cap. Additionally, targeted expression studies reveal that hydrotropism depends on the ABA signalling kinase SnRK2.2 and the hydrotropism-specific MIZ1, both acting specifically in elongation zone cortical cells. Conversely, hydrotropism, but not gravitropism, is inhibited by preventing differential cell-length increases in the cortex, but not in other cell types. We conclude that root tropic responses to gravity and water are driven by distinct tissue-based mechanisms. In addition, unlike its role in root gravitropism, the elongation zone performs a dual function during a hydrotropic response, both sensing a water potential gradient and subsequently undergoing differential growth.

Developmental Progression in the Coral Acropora digitifera Is Controlled by Differential Expression of Distinct Regulatory Gene Networks

PubMed Central

Reyes-Bermudez, Alejandro; Villar-Briones, Alejandro; Ramirez-Portilla, Catalina; Hidaka, Michio; Mikheyev, Alexander S.

2016-01-01

Corals belong to the most basal class of the Phylum Cnidaria, which is considered the sister group of bilaterian animals, and thus have become an emerging model to study the evolution of developmental mechanisms. Although cell renewal, differentiation, and maintenance of pluripotency are cellular events shared by multicellular animals, the cellular basis of these fundamental biological processes are still poorly understood. To understand how changes in gene expression regulate morphogenetic transitions at the base of the eumetazoa, we performed quantitative RNA-seq analysis during Acropora digitifera’s development. We collected embryonic, larval, and adult samples to characterize stage-specific transcription profiles, as well as broad expression patterns. Transcription profiles reconstructed development revealing two main expression clusters. The first cluster grouped blastula and gastrula and the second grouped subsequent developmental time points. Consistently, we observed clear differences in gene expression between early and late developmental transitions, with higher numbers of differentially expressed genes and fold changes around gastrulation. Furthermore, we identified three coexpression clusters that represented discrete gene expression patterns. During early transitions, transcriptional networks seemed to regulate cellular fate and morphogenesis of the larval body. In late transitions, these networks seemed to play important roles preparing planulae for switch in lifestyle and regulation of adult processes. Although developmental progression in A. digitifera is regulated to some extent by differential coexpression of well-defined gene networks, stage-specific transcription profiles appear to be independent entities. While negative regulation of transcription is predominant in early development, cell differentiation was upregulated in larval and adult stages. PMID:26941230

The protein expression landscape of mitosis and meiosis in diploid budding yeast.

PubMed

Becker, Emmanuelle; Com, Emmanuelle; Lavigne, Régis; Guilleux, Marie-Hélène; Evrard, Bertrand; Pineau, Charles; Primig, Michael

2017-03-06

Saccharomyces cerevisiae is an established model organism for the molecular analysis of fundamental biological processes. The genomes of numerous strains have been sequenced, and the transcriptome and proteome ofmajor phases during the haploid and diploid yeast life cycle have been determined. However, much less is known about dynamic changes of the proteome when cells switch from mitotic growth to meiotic development. We report a quantitative protein profiling analysis of yeast cell division and differentiation based on mass spectrometry. Information about protein levels was integrated with strand-specific tiling array expression data. We identified a total of 2366 proteins in at least one condition, including 175 proteins showing a statistically significant>5-fold change across the sample set, and 136 proteins detectable in sporulating but not respiring cells. We correlate protein expression patterns with biological processes and molecular function by Gene Ontology term enrichment, chemoprofiling, transcription interference and the formation of double stranded RNAs by overlapping sense/antisense transcripts. Our work provides initial quantitative insight into protein expression in diploid respiring and differentiating yeast cells. Critically, it associates developmentally regulated induction of antisense long noncoding RNAs and double stranded RNAs with fluctuating protein concentrations during growth and development. This integrated genomics analysis helps better understand how the transcriptome and the proteome correlate in diploid yeast cells undergoing mitotic growth in the presence of acetate (respiration) versus meiotic differentiation (Meiosis I and II). The study (i) provides quantitative expression data for 2366 proteins and their cognate mRNAs in at least one sample, (ii) shows strongly fluctuating protein levels during growth and differentiation for 175 cases, and (iii) identifies 136 proteins absent in mitotic but present in meiotic yeast cells. We have integrated protein profiling data using mass spectrometry with tiling array RNA profiling data and information on double-stranded RNAs (dsRNAs) by overlapping sense/antisense transcripts from an RNA-Sequencing experiment. This work therefore provides quantitative insight into protein expression during cell division and development and associates changing protein levels with developmental stage specific induction of antisense transcripts and the formation of dsRNAs. Copyright © 2017 Elsevier B.V. All rights reserved.

Single-Cell RNA-Sequencing Reveals a Continuous Spectrum of Differentiation in Hematopoietic Cells.

PubMed

Macaulay, Iain C; Svensson, Valentine; Labalette, Charlotte; Ferreira, Lauren; Hamey, Fiona; Voet, Thierry; Teichmann, Sarah A; Cvejic, Ana

2016-02-02

The transcriptional programs that govern hematopoiesis have been investigated primarily by population-level analysis of hematopoietic stem and progenitor cells, which cannot reveal the continuous nature of the differentiation process. Here we applied single-cell RNA-sequencing to a population of hematopoietic cells in zebrafish as they undergo thrombocyte lineage commitment. By reconstructing their developmental chronology computationally, we were able to place each cell along a continuum from stem cell to mature cell, refining the traditional lineage tree. The progression of cells along this continuum is characterized by a highly coordinated transcriptional program, displaying simultaneous suppression of genes involved in cell proliferation and ribosomal biogenesis as the expression of lineage specific genes increases. Within this program, there is substantial heterogeneity in the expression of the key lineage regulators. Overall, the total number of genes expressed, as well as the total mRNA content of the cell, decreases as the cells undergo lineage commitment. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Viral Superantigen Drives Extrafollicular and Follicular B Cell Differentiation Leading to Virus-specific Antibody Production

PubMed Central

Luther, Sanjiv A.; Gulbranson-Judge, Adam; Acha-Orbea, Hans; MacLennan, Ian C.M.

1997-01-01

Mouse mammary tumor virus (MMTV[SW]) encodes a superantigen expressed by infected B cells. It evokes an antibody response specific for viral envelope protein, indicating selective activation of antigen-specific B cells. The response to MMTV(SW) in draining lymph nodes was compared with the response to haptenated chicken gamma globulin (NP-CGG) using flow cytometry and immunohistology. T cell priming occurs in both responses, with T cells proliferating in association with interdigitating dendritic cells in the T zone. T cell proliferation continues in the presence of B cells in the outer T zone, and B blasts then undergo exponential growth and differentiation into plasma cells in the medullary cords. Germinal centers develop in both responses, but those induced by MMTV(SW) appear later and are smaller. Most T cells activated in the T zone and germinal centers in the MMTV(SW) response are superantigen specific and these persist for weeks in lymph nodes draining the site MMTV(SW) injection; this contrasts with the selective loss of superantigen-specific T cells from other secondary lymphoid tissues. The results indicate that this viral superantigen, when expressed by professional antigen-presenting cells, drives extrafollicular and follicular B cell differentiation leading to virus-specific antibody production. PMID:9053455

The CpG island methylator phenotype may confer a survival benefit in patients with stage II or III colorectal carcinomas receiving fluoropyrimidine-based adjuvant chemotherapy

PubMed Central

2011-01-01

Background Colorectal carcinoma (CRC) with CpG island methylator phenotype (CIMP) is recognized as a distinct subgroup of CRC, and CIMP status affects prognosis and response to chemotherapy. Identification of CIMP status in CRC is important for proper patient management. In Eastern countries, however, the clinicopathologic and molecular characteristics and prognosis of CRCs with CIMP are still unclear. Methods A total of 245 patients who underwent their first surgical resection for sporadic CRC were enrolled and CIMP status of the CRCs was determined using the quantitative MethyLight assay. The clinicopathologic and molecular characteristics were reviewed and compared according to CIMP status. In addition, the three-year recurrence-free survival (RFS) of 124 patients with stage II or stage III CRC was analyzed in order to assess the effectiveness of fluoropyrimidine-based adjuvant chemotherapy with respect to CIMP status. Results CIMP-high CRCs were identified in 34 cases (13.9%), and were significantly associated with proximal tumor location, poorly differentiated carcinoma, mucinous histology, and high frequencies of BRAF mutation, MGMT methylation, and MSI-high compared to CIMP-low/negative carcinomas. For patients with stage II or III CIMP-low/negative CRCs, no significant difference was found in RFS between those undergoing surgery alone and those receiving surgery with fluoropyrimidine-based adjuvant chemotherapy. However, for patients with CIMP-high CRCs, patients undergoing surgery with fluoropyrimidine-based adjuvant chemotherapy (n = 17; three-year RFS: 100%) showed significantly better RFS than patients treated with surgery alone (n = 7; three-year RFS: 71.4%) (P = 0.022). Conclusions Our results suggest that selected patients with CIMP-high CRC may benefit from fluoropyrimidine-based adjuvant chemotherapy with longer RFS. Further large scale-studies are required to confirm our results. PMID:21827707

Analysis of phosphorylated proteins and inhibition of kinase activity during Giardia intestinalis excystation.

PubMed

Alvarado, Magda E; Wasserman, Moisés

2010-03-01

The parasite Giardia intestinalis undergoes a differentiation process that allows it to infect its mammal host. That process is excystation. We examined the importance of protein phosphorylation during the passage from cyst to trophozoite. Cysts obtained from patients with giardiasis were excysted in vitro and the soluble cytoplasmic proteins were analyzed during the three phases of the process, using a specific staining for phosphoproteins. We found two phosphorylated proteins and identified them with MALDI-TOF as 14-3-3 and Hsp70. Modifications were detected in both proteins, which could indicate a role in differentiation of the parasite. In addition, the inhibition of serine-threonine kinases during excystation specifically affected the cytokinesis of the excyzoite, thus inhibiting the completion of trophozoite formation. Copyright 2009 Elsevier Ireland Ltd. All rights reserved.

Developmental stage related patterns of codon usage and genomic GC content: searching for evolutionary fingerprints with models of stem cell differentiation

PubMed Central

2007-01-01

Background The usage of synonymous codons shows considerable variation among mammalian genes. How and why this usage is non-random are fundamental biological questions and remain controversial. It is also important to explore whether mammalian genes that are selectively expressed at different developmental stages bear different molecular features. Results In two models of mouse stem cell differentiation, we established correlations between codon usage and the patterns of gene expression. We found that the optimal codons exhibited variation (AT- or GC-ending codons) in different cell types within the developmental hierarchy. We also found that genes that were enriched (developmental-pivotal genes) or specifically expressed (developmental-specific genes) at different developmental stages had different patterns of codon usage and local genomic GC (GCg) content. Moreover, at the same developmental stage, developmental-specific genes generally used more GC-ending codons and had higher GCg content compared with developmental-pivotal genes. Further analyses suggest that the model of translational selection might be consistent with the developmental stage-related patterns of codon usage, especially for the AT-ending optimal codons. In addition, our data show that after human-mouse divergence, the influence of selective constraints is still detectable. Conclusion Our findings suggest that developmental stage-related patterns of gene expression are correlated with codon usage (GC3) and GCg content in stem cell hierarchies. Moreover, this paper provides evidence for the influence of natural selection at synonymous sites in the mouse genome and novel clues for linking the molecular features of genes to their patterns of expression during mammalian ontogenesis. PMID:17349061

Metastatic pancreatic cancer presenting as linitis plastica of the stomach.

PubMed

Garg, Shivani; Mulki, Ramzi; Sher, Daniel

2016-03-08

Metastatic disease from pancreatic carcinoma involving the stomach is an unusual event, and the pattern of spread in the form of linitis plastica, to our knowledge, has not been reported previously. Local recurrence after curative resection for pancreatic cancer is the most common pattern of disease. We report a case of metastatic pancreatic adenocarcinoma presenting as linitis plastica of the stomach 4 years after curative resection. A 52-year-old man presented with epigastric pain and melaena 4 years after undergoing a Whipple's procedure for a poorly-differentiated pancreatic adenocarcinoma, stage IB; T2N0M0. CT imaging of the abdomen revealed thickening of the gastric wall, and subsequent oesophagogastroduodenoscopy (OGD) revealed diffuse friable erythaematous tissue. The biopsy specimen obtained during the OGD revealed a poorly differentiated adenocarcinoma, with similar appearance to the prior specimen obtained from the pancreas. 2016 BMJ Publishing Group Ltd.

Differentially regulated gene expression associated with hepatitis C virus clearance.

PubMed

Grimes, Carolyn Z; Hwang, Lu-Yu; Wei, Peng; Shah, Dimpy P; Volcik, Kelly A; Brown, Eric L

2013-03-01

Human chronic hepatitis C virus (HCV) infections pose a significant public health threat, necessitating the development of novel treatments and vaccines. HCV infections range from spontaneous resolution to end-stage liver disease. Approximately 10-30% of HCV infections undergo spontaneous resolution independent of treatment by yet-to-be-defined mechanisms. These individuals test positive for anti-HCV antibodies in the absence of detectable viral serum RNA. To identify genes associated with HCV clearance, this study compared gene expression profiles between current drug users chronically infected with HCV and drug users who cleared their HCV infection. This analysis identified 91 differentially regulated (up- or downregulated by twofold or more) genes potentially associated with HCV clearance. The majority of genes identified were associated with immune function, with the remaining genes categorized either as cancer related or 'other'. Identification of factors and pathways that may influence virus clearance will be essential to the development of novel treatment strategies.

Comparison of transcript profiles in different life stages of the nematode Globodera pallida under different host potato genotypes.

PubMed

Palomares-Rius, Juan E; Hedley, Pete E; Cock, Peter J A; Morris, Jenny A; Jones, John T; Vovlas, Nikos; Blok, Vivian

2012-12-01

The potato cyst nematodes (PCNs) Globodera pallida and Globodera rostochiensis are important parasites of potato. PCNs undergo complex biotrophic interactions with their hosts that involve gene expression changes in both the nematode and the host plant. The aim of this study was to determine key genes that are differentially expressed in Globodera pallida life cycle stages and during the initiation of the feeding site in susceptible and partially resistant potato genotypes. For this purpose, two microarray experiments were designed: (i) a comparison of eggs, infective second-stage juveniles (J2s) and sedentary parasitic-stage J2s (SJ2); (ii) a comparison of SJ2s at 8 days after inoculation (DAI) in the susceptible cultivar (Desirée) and two partially resistant lines. The results showed differential expression of G. pallida genes during the stages studied, including previously characterized effectors. In addition, a large number of genes changed their expression between SJ2s in the susceptible cultivar and those infecting partially resistant lines; the number of genes with modified expression was lower when the two partially resistant lines were compared. Moreover, a histopathological study was performed at several time points (7, 14 and 30 DAI) and showed the similarities between both partially resistant lines with a delay and degeneration in the formation of the syncytia in comparison with the susceptible cultivar. Females at 30 DAI in partially resistant lines showed a delay in their development in comparison with those in the susceptible cultivar. © 2012 THE AUTHORS. MOLECULAR PLANT PATHOLOGY © 2012 BSPP AND BLACKWELL PUBLISHING LTD.

Evaluation of the 8th AJCC staging system for pathologically versus clinically staged pancreatic adenocarcinoma: A time to revisit a dogma?

PubMed

Abdel-Rahman, Omar

2018-02-01

The 8th edition of the American Joint Committee on Cancer (AJCC) staging system for pancreatic exocrine adenocarcinoma has been released. The current study seeks to assess the 7th and 8th editions among patients registered within the surveillance, epidemiology and end results (SEER) database. SEER database (2010-2013) has been accessed through SEER*Stat program and AJCC 8th edition stages were reconstructed utilizing the collaborative stage descriptions. Kaplan-Meier analysis of overall survival and pancreatic cancer-specific survival analyses (according to both 7th and 8th editions and according to whether pathological or clinical staging were conducted) has been performed. Multivariate analysis of factors affecting pancreatic cancer-specific survival was also conducted through a Cox proportional hazard model. A total of 18  948 patients with pancreatic adenocarcinoma were identified in the period from 2010-2013. Pancreatic cancer-specific survival among pathologically staged patients and according to the 8th edition showed significant differences for all pair wise comparisons among different stages (P < 0.0001) except for the comparison between stage IA and stage IB (P = 0.307) and the comparison between stage IB and stage IIA (P = 0.116). Moreover, P value for stage IA vs IIA was 0.014; while pancreatic cancer-specific survival according to the 7th edition among pathologically staged patients showed significant differences for all pair wise comparisons among different stages (P < 0.0001) except for the comparison between IA and IB (P = 0.072), the comparison between stage IIA and stage IIB (P = 0.065), the comparison between stage IIA and stage III (P = 0.059) and the comparison between IIB and III (P = 0.595). Among clinically staged patients (i.e. those who did not undergo initial radical surgery), the prognostic performance of both 7th and 8th stages for both overall survival and pancreatic cancer-specific survival was limited. There is clearly a need to have two staging systems for pancreatic adenocarcinoma: pathological and clinical staging systems. Copyright © 2018 First Affiliated Hospital, Zhejiang University School of Medicine in China. Published by Elsevier B.V. All rights reserved.

Gene trapping in differentiating cell lines: regulation of the lysosomal protease cathepsin B in skeletal myoblast growth and fusion.

PubMed

Gogos, J A; Thompson, R; Lowry, W; Sloane, B F; Weintraub, H; Horwitz, M

1996-08-01

To identify genes regulated during skeletal muscle differentiation, we have infected mouse C2C12 myoblasts with retroviral gene trap vectors, containing a promoterless marker gene with a 5' splice acceptor signal. Integration of the vector adjacent to an actively transcribed gene places the marker under the transcriptional control of the endogenous gene, while the adjacent vector sequences facilitate cloning. The vector insertionally mutates the trapped locus and may also form fusion proteins with the endogenous gene product. We have screened several hundred clones, each containing a trapping vector integrated into a different endogenous gene. In agreement with previous estimates based on hybridization kinetics, we find that a large proportion of all genes expressed in myoblasts are regulated during differentiation. Many of these genes undergo unique temporal patterns of activation or repression during cell growth and myotube formation, and some show specific patterns of subcellular localization. The first gene we have identified with this strategy is the lysosomal cysteine protease cathepsin B. Expression from the trapped allele is upregulated during early myoblast fusion and downregulated in myotubes. A direct role for cathepsin B in myoblast growth and fusion is suggested by the observation that the trapped cells deficient in cathepsin B activity have an unusual morphology and reduced survival in low-serum media and undergo differentiation with impaired cellular fusion. The phenotype is reproduced by antisense cathepsin B expression in parental C2C12 myoblasts. The cellular phenotype is similar to that observed in cultured myoblasts from patients with I cell disease, in which there is diminished accumulation of lysosomal enzymes. This suggests that a specific deficiency of cathepsin B could contribute to the myopathic component of this illness.

Species-specific differentiation of variola, monkeypox, and varicella-zoster viruses by multiplex real-time PCR assay.

PubMed

Maksyutov, Rinat A; Gavrilova, Elena V; Shchelkunov, Sergei N

2016-10-01

A method of one-stage rapid detection and differentiation of epidemiologically important variola virus (VARV), monkeypox virus (MPXV), and varicella-zoster virus (VZV) utilizing multiplex real-time TaqMan PCR assay was developed. Four hybridization probes with various fluorescent dyes and the corresponding fluorescence quenchers were simultaneously used for the assay. The hybridization probes specific for the VARV sequence contained FAM/BHQ1 as a dye/quencher pair; MPXV-specific, JOE/BHQ1; VZV-specific, TAMRA/BHQ2; and internal control-specific, Cy5/BHQ3. The specificity and sensitivity of the developed method were assessed by analyzing DNA of 32 strains belonging to orthopoxvirus and herpesvirus species. Copyright © 2016 Elsevier B.V. All rights reserved.

MAPK signaling pathways and HDAC3 activity are disrupted during differentiation of emerin-null myogenic progenitor cells

PubMed Central

Collins, Carol M.; Ellis, Joseph A.

2017-01-01

ABSTRACT Mutations in the gene encoding emerin cause Emery–Dreifuss muscular dystrophy (EDMD). Emerin is an integral inner nuclear membrane protein and a component of the nuclear lamina. EDMD is characterized by skeletal muscle wasting, cardiac conduction defects and tendon contractures. The failure to regenerate skeletal muscle is predicted to contribute to the skeletal muscle pathology of EDMD. We hypothesize that muscle regeneration defects are caused by impaired muscle stem cell differentiation. Myogenic progenitors derived from emerin-null mice were used to confirm their impaired differentiation and analyze selected myogenic molecular pathways. Emerin-null progenitors were delayed in their cell cycle exit, had decreased myosin heavy chain (MyHC) expression and formed fewer myotubes. Emerin binds to and activates histone deacetylase 3 (HDAC3). Here, we show that theophylline, an HDAC3-specific activator, improved myotube formation in emerin-null cells. Addition of the HDAC3-specific inhibitor RGFP966 blocked myotube formation and MyHC expression in wild-type and emerin-null myogenic progenitors, but did not affect cell cycle exit. Downregulation of emerin was previously shown to affect the p38 MAPK and ERK/MAPK pathways in C2C12 myoblast differentiation. Using a pure population of myogenic progenitors completely lacking emerin expression, we show that these pathways are also disrupted. ERK inhibition improved MyHC expression in emerin-null cells, but failed to rescue myotube formation or cell cycle exit. Inhibition of p38 MAPK prevented differentiation in both wild-type and emerin-null progenitors. These results show that each of these molecular pathways specifically regulates a particular stage of myogenic differentiation in an emerin-dependent manner. Thus, pharmacological targeting of multiple pathways acting at specific differentiation stages may be a better therapeutic approach in the future to rescue muscle regeneration in vivo. PMID:28188262

Dynamic changes in copper homeostasis and post-transcriptional regulation of Atp7a during myogenic differentiation.

PubMed

Vest, Katherine E; Paskavitz, Amanda L; Lee, Joseph B; Padilla-Benavides, Teresita

2018-02-21

Copper (Cu) is an essential metal required for activity of a number of redox active enzymes that participate in critical cellular pathways such as metabolism and cell signaling. Because it is also a toxic metal, Cu must be tightly controlled by a series of transporters and chaperone proteins that regulate Cu homeostasis. The critical nature of Cu is highlighted by the fact that mutations in Cu homeostasis genes cause pathologic conditions such as Menkes and Wilson diseases. While Cu homeostasis in highly affected tissues like the liver and brain is well understood, no study has probed the role of Cu in development of skeletal muscle, another tissue that often shows pathology in these conditions. Here, we found an increase in whole cell Cu content during differentiation of cultured immortalized or primary myoblasts derived from mouse satellite cells. We demonstrate that Cu is required for both proliferation and differentiation of primary myoblasts. We also show that a key Cu homeostasis gene, Atp7a, undergoes dynamic changes in expression during myogenic differentiation. Alternative polyadenylation and stability of Atp7a mRNA fluctuates with differentiation stage of the myoblasts, indicating post-transcriptional regulation of Atp7a that depends on the differentiation state. This is the first report of a requirement for Cu during myogenic differentiation and provides the basis for understanding the network of Cu transport associated with myogenesis.

"Even if I Don't Remember, I Feel Better". A Qualitative Study of Patients with Early-Stage Non-Small Cell Lung Cancer Undergoing Stereotactic Body Radiotherapy or Surgery.

PubMed

Golden, Sara E; Thomas, Charles R; Deffebach, Mark E; Sukumar, Mithran S; Schipper, Paul H; Tieu, Brandon H; Kee, Andrew Y; Tsen, Andrew C; Slatore, Christopher G

2016-08-01

While surgical resection is recommended for most patients with early stage lung cancer according to the National Comprehensive Cancer Network guidelines, stereotactic body radiotherapy is increasingly being used. Provider-patient communication regarding the risks and benefits of each approach may be a modifiable factor leading to improved patient-centered outcomes. To qualitatively describe the experiences of patients undergoing either surgery or stereotactic body radiotherapy for early stage non-small cell lung cancer. We qualitatively evaluated and used content analysis to describe the experiences of 13 patients with early clinical stage non-small cell lung cancer before undergoing treatment in three health care systems in the Pacific Northwest, with a focus on knowledge obtained, communication, and feelings of distress. Although most participants reported rarely having been told about other options for treatment and could not readily recall many details about specific risks of recommended treatment, they were satisfied with their care. The patients paradoxically described clinicians as displaying caring and empathy despite not explicitly addressing their concerns and worries. We found that the communication domains that underlie shared decision making occurred infrequently, but that participants were still pleased with their role in the decision-making process. We did not find substantially different themes based on where the participant received care or the treatment selected. Patients were satisfied with all aspects of their care, despite reporting little knowledge about risks or other treatment options, no direct elicitation of worries from providers, and a lack of shared decision making. While the development of effective communication strategies to address these gaps is warranted, their effect on patient-centered outcomes, such as distress and decisional conflict, is unclear.

Gene expression profiling during asexual development of the late blight pathogen Phytophthora infestans reveals a highly dynamic transcriptome.

PubMed

Judelson, Howard S; Ah-Fong, Audrey M V; Aux, George; Avrova, Anna O; Bruce, Catherine; Cakir, Cahid; da Cunha, Luis; Grenville-Briggs, Laura; Latijnhouwers, Maita; Ligterink, Wilco; Meijer, Harold J G; Roberts, Samuel; Thurber, Carrie S; Whisson, Stephen C; Birch, Paul R J; Govers, Francine; Kamoun, Sophien; van West, Pieter; Windass, John

2008-04-01

Much of the pathogenic success of Phytophthora infestans, the potato and tomato late blight agent, relies on its ability to generate from mycelia large amounts of sporangia, which release zoospores that encyst and form infection structures. To better understand these stages, Affymetrix GeneChips based on 15,650 unigenes were designed and used to profile the life cycle. Approximately half of P. infestans genes were found to exhibit significant differential expression between developmental transitions, with approximately (1)/(10) being stage-specific and most changes occurring during zoosporogenesis. Quantitative reverse-transcription polymerase chain reaction assays confirmed the robustness of the array results and showed that similar patterns of differential expression were obtained regardless of whether hyphae were from laboratory media or infected tomato. Differentially expressed genes encode potential cellular regulators, especially protein kinases; metabolic enzymes such as those involved in glycolysis, gluconeogenesis, or the biosynthesis of amino acids or lipids; regulators of DNA synthesis; structural proteins, including predicted flagellar proteins; and pathogenicity factors, including cell-wall-degrading enzymes, RXLR effector proteins, and enzymes protecting against plant defense responses. Curiously, some stage-specific transcripts do not appear to encode functional proteins. These findings reveal many new aspects of oomycete biology, as well as potential targets for crop protection chemicals.

In vivo Diagnosis of Cervical Intraepithelial Neoplasia Using 337-nm- Excited Laser-Induced Fluorescence

NASA Astrophysics Data System (ADS)

Ramanujam, N.; Mitchell, M. F.; Mahadevan, A.; Warren, S.; Thomsen, S.; Silva, E.; Richards-Kortum, R.

1994-10-01

Laser-induced fluorescence at 337-nm excitation was used in vivo to differentiate neoplastic [cervical intraepithelial neoplasia (CIN)], nonneoplastic abnormal (inflammation and human papilloma viral infection), and normal cervical tissues. A colposcope (low-magnification microscope used to view the cervix with reflected light) was used to identify 66 normal and 49 abnormal (5 inflammation, 21 human papilloma virus infection, and 23 CIN) sites on the cervix in 28 patients. These sites were then interrogated spectroscopically. A two-stage algorithm was developed to diagnose CIN. The first stage differentiated histologically abnormal tissues from colposcopically normal tissues with a sensitivity, specificity, and positive predictive value of 92%, 90%, and 88%, respectively. The second stage differentiated preneoplastic and neoplastic tissues from nonneoplastic abnormal tissues with a sensitivity, specificity, and positive predictive value of 87%, 73%, and 74%, respectively. Spectroscopic differences were consistent with a decrease in the absolute contribution of collagen fluorescence, an increase in the absolute contribution of oxyhemoglobin attenuation, and an increase in the relative contribution of reduced nicotinamide dinucleotide phosphate [NAD(P)H] fluorescence as tissue progresses from normal to abnormal in the same patient. These results suggest that in vivo fluorescence spectroscopy of the cervix can be used to diagnose CIN at colposcopy.

FMRFamide-like peptides expand the behavioral repertoire of a densely connected nervous system

PubMed Central

Schaedel, Oren N.; Rogers, Alicia K.

2017-01-01

Animals, including humans, can adapt to environmental stress through phenotypic plasticity. The free-living nematode Caenorhabditis elegans can adapt to harsh environments by undergoing a whole-animal change, involving exiting reproductive development and entering the stress-resistant dauer larval stage. The dauer is a dispersal stage with dauer-specific behaviors for finding and stowing onto carrier animals, but how dauers acquire these behaviors, despite having a physically limited nervous system of 302 neurons, is poorly understood. We compared dauer and reproductive development using whole-animal RNA sequencing at fine time points and at sufficient depth to measure transcriptional changes within single cells. We detected 8,042 genes differentially expressed during dauer and reproductive development and observed striking up-regulation of neuropeptide genes during dauer entry. We knocked down neuropeptide processing using sbt-1 mutants and demonstrate that neuropeptide signaling promotes the decision to enter dauer rather than reproductive development. We also demonstrate that during dauer neuropeptides modulate the dauer-specific nictation behavior (carrier animal-hitchhiking) and are necessary for switching from repulsion to CO2 (a carrier animal cue) in nondauers to CO2 attraction in dauers. We tested individual neuropeptides using CRISPR knockouts and existing strains and demonstrate that the combined effects of flp-10 and flp-17 mimic the effects of sbt-1 on nictation and CO2 attraction. Through meta-analysis, we discovered similar up-regulation of neuropeptides in the dauer-like infective juveniles of diverse parasitic nematodes, suggesting the antiparasitic target potential of SBT-1. Our findings reveal that, under stress, increased neuropeptide signaling in C. elegans enhances their decision-making accuracy and expands their behavioral repertoire. PMID:29167374

Pancreatic Endoderm-Derived From Diabetic Patient-Specific Induced Pluripotent Stem Cell Generates Glucose-Responsive Insulin-Secreting Cells.

PubMed

Rajaei, Bahareh; Shamsara, Mehdi; Amirabad, Leila Mohammadi; Massumi, Mohammad; Sanati, Mohammad Hossein

2017-10-01

Human-induced pluripotent stem cells (hiPSCs) can potentially serve as an invaluable source for cell replacement therapy and allow the creation of patient- and disease-specific stem cells without the controversial use of embryos and avoids any immunological incompatibility. The generation of insulin-producing pancreatic β-cells from pluripotent stem cells in vitro provides an unprecedented cell source for personal drug discovery and cell transplantation therapy in diabetes. A new five-step protocol was introduced in this study, effectively induced hiPSCs to differentiate into glucose-responsive insulin-producing cells. This process mimics in vivo pancreatic organogenesis by directing cells through stages resembling definitive endoderm, primitive gut-tube endoderm, posterior foregut, pancreatic endoderm, and endocrine precursor. Each stage of differentiation were characterized by stage-specific markers. The produced cells exhibited many properties of functional β-cells, including expression of critical β-cells transcription factors, the potency to secrete C-peptide in response to high levels of glucose and the presence of mature endocrine secretory granules. This high efficient differentiation protocol, established in this study, yielded 79.18% insulin-secreting cells which were responsive to glucose five times higher than the basal level. These hiPSCs-derived glucose-responsive insulin-secreting cells might provide a promising approach for the treatment of type I diabetes mellitus. J. Cell. Physiol. 232: 2616-2625, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Genome-wide analysis of miRNA and mRNA transcriptomes during amelogenesis.

PubMed

Yin, Kaifeng; Hacia, Joseph G; Zhong, Zhe; Paine, Michael L

2014-11-19

In the rodent incisor during amelogenesis, as ameloblast cells transition from secretory stage to maturation stage, their morphology and transcriptome profiles change dramatically. Prior whole genome transcriptome analysis has given a broad picture of the molecular activities dominating both stages of amelogenesis, but this type of analysis has not included miRNA transcript profiling. In this study, we set out to document which miRNAs and corresponding target genes change significantly as ameloblasts transition from secretory- to maturation-stage amelogenesis. Total RNA samples from both secretory- and maturation-stage rat enamel organs were subjected to genome-wide miRNA and mRNA transcript profiling. We identified 59 miRNAs that were differentially expressed at the maturation stage relative to the secretory stage of enamel development (False Discovery Rate (FDR)<0.05, fold change (FC)≥1.8). In parallel, transcriptome profiling experiments identified 1,729 mRNA transcripts that were differentially expressed in the maturation stage compared to the secretory stage (FDR<0.05, FC≥1.8). Based on bioinformatics analyses, 5.8% (629 total) of these differentially expressed genes (DEGS) were highlighted as being the potential targets of 59 miRNAs that were differentially expressed in the opposite direction, in the same tissue samples. Although the number of predicted target DEGs was not higher than baseline expectations generated by examination of stably expressed miRNAs, Gene Ontology (GO) analysis showed that these 629 DEGS were enriched for ion transport, pH regulation, calcium handling, endocytotic, and apoptotic activities. Seven differentially expressed miRNAs (miR-21, miR-31, miR-488, miR-153, miR-135b, miR-135a and miR298) in secretory- and/or maturation-stage enamel organs were confirmed by in situ hybridization. Further, we used luciferase reporter assays to provide evidence that two of these differentially expressed miRNAs, miR-153 and miR-31, are potential regulators for their predicated target mRNAs, Lamp1 (miR-153) and Tfrc (miR-31). In conclusion, these data indicate that miRNAs exhibit a dynamic expression pattern during the transition from secretory-stage to maturation-stage tooth enamel formation. Although they represent only one of numerous mechanisms influencing gene activities, miRNAs specific to the maturation stage could be involved in regulating several key processes of enamel maturation by influencing mRNA stability and translation.

The value of histological grading of biopsy and resection specimens in early stage oral squamous cell carcinomas.

PubMed

Dik, Eric A; Ipenburg, Norbertus A; Kessler, Peter A; van Es, Robert J J; Willems, Stefan M

2018-04-05

In oral squamous cell carcinoma (OSCC) the differentiation grade of the tumor is determined on the biopsy and the resection specimen. The relation between tumor grade, nodal metastasis and survival is debatable. The aims of this study were to determine the correlation between differentiation grade of the biopsy and the resection specimen. Furthermore, we wanted to correlate tumor differentiation grade with nodal stage and survival. One-hundred and forty-five patients with OSCC staged as T1-2, N0 of the tongue, floor of mouth or cheek with primary resection of the tumor were examined. Biopsy and resection specimen were histologically re-assessed with regard to differentiation grade, as well as infiltrative, peri-neural and vascular invasive growth. This study showed a poor correlation between differentiation grade in the incisional biopsy and the resection specimen of the same tumor. No significant relation between differentiation grade of the resection specimen and nodal involvement, as well as overall and disease-specific survival was found. In early OSCC the differentiation grade determined by biopsy is of little predictive value for the grading of the resection specimen. Poor differentiation grade could not be related to the presence of nodal metastasis or survival and seems not to have any prognostic value concerning outcome. Treatment planning must be related to these findings. Copyright © 2018 European Association for Cranio-Maxillo-Facial Surgery. Published by Elsevier Ltd. All rights reserved.

TGF-β control of stem cell differentiation genes.

PubMed

Massagué, Joan; Xi, Qiaoran

2012-07-04

The canonical TGF-β/Smad signaling pathway was delineated in the mid 90s and enriched over the past decade with many findings about its specificity, regulation, networking, and malfunctions in disease. However, a growing understanding of the chromatin status of a critical class of TGF-β target genes - the genes controlling differentiation of embryonic stem cells - recently prompted a reexamination of this pathway and its critical role in the regulation of stem cell differentiation. The new findings reveal master regulators of the pluripotent state set the stage for Smad-mediated activation of master regulators of the next differentiation stage. Furthermore, a novel branch of the TGF-β/Smad pathway has been identified in which a chromatin-reading Smad complex makes the master differentiation genes accessible to canonical Smad complexes for transcriptional activation. These findings provide exciting new insights into the global role of TGF-β signaling in the regulators of stem cell fate. Copyright © 2012 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Stage-specific effects of FGF2 on the differentiation of dental pulp cells

PubMed Central

Sagomonyants, Karen; Mina, Mina

2015-01-01

Dentinogenesis is a complex and multistep process, which is regulated by various growth factors, including members of the Fibroblast Growth Factor (FGF) family. Both positive and negative effects of FGFs on dentinogenesis have been reported but the underlying mechanisms of these conflicting results are still unclear. To gain better insight into the role of FGF2 in dentinogenesis, we used dental pulp cells from various transgenic mice, in which fluorescent protein expression identifies cells at different stages of odontoblast differentiation. Our results showed that continuous exposure of pulp cells to FGF2 inhibited mineralization and revealed both stimulatory and inhibitory effects of FGF2 on expression of markers of dentinogenesis and various transgenes. During the proliferation phase of in vitro growth FGF2 increased expression of markers of dentinogenesis and the percentages of DMP1-GFP+ functional odontoblasts and DSPP-Cerulean+ odontoblasts. Additional exposure to FGF2 during the differentiation/mineralization phase of in vitro growth decreased the extent of mineralization, expression of markers of dentinogenesis, and expression of DMP1-GFP and DSPP-Cerulean transgenes. Recovery experiments showed that the inhibitory effects of FGF2 on dentinogenesis were related to the blocking of differentiation of cells into mature odontoblasts. These observations together showed stage-specific effects of FGF2 on dentinogenesis by dental pulp cells and provide critical information for the development of improved treatments for vital pulp therapy and dentin regeneration. PMID:25823776

Early embryonic expression of FGF4/6/9 gene and its role in the induction of mesenchyme and notochord in Ciona savignyi embryos.

PubMed

Imai, Kaoru S; Satoh, Nori; Satou, Yutaka

2002-04-01

In early Ciona savignyi embryos, nuclear localization of beta-catenin is the first step of endodermal cell specification, and triggers the activation of various target genes. A cDNA for Cs-FGF4/6/9, a gene activated downstream of beta-catenin signaling, was isolated and shown to encode an FGF protein with features of both FGF4/6 and FGF9/20. The early embryonic expression of Cs-FGF4/6/9 was transient and the transcript was seen in endodermal cells at the 16- and 32-cell stages, in notochord and muscle cells at the 64-cell stage, and in nerve cord and muscle cells at the 110-cell stage; the gene was then expressed again in cells of the nervous system after neurulation. When the gene function was suppressed with a specific antisense morpholino oligo, the differentiation of mesenchyme cells was completely blocked, and the fate of presumptive mesenchyme cells appeared to change into that of muscle cells. The inhibition of mesenchyme differentiation was abrogated by coinjection of the morpholino oligo and synthetic Cs-FGF4/6/9 mRNA. Downregulation of beta-catenin nuclear localization resulted in the absence of mesenchyme cell differentiation due to failure of the formation of signal-producing endodermal cells. Injection of synthetic Cs-FGF4/6/9 mRNA in beta-catenin-downregulated embryos evoked mesenchyme cell differentiation. These results strongly suggest that Cs-FGF4/6/9 produced by endodermal cells acts an inductive signal for the differentiation of mesenchyme cells. On the other hand, the role of Cs-FGF4/6/9 in the induction of notochord cells is partial; the initial process of the induction was inhibited by Cs-FGF4/6/9 morpholino oligo, but notochord-specific genes were expressed later to form a partial notochord.

Irx1 regulates dental outer enamel epithelial and lung alveolar type II epithelial differentiation

PubMed Central

Yu, Wenjie; Li, Xiao; Eliason, Steven; Romero-Bustillos, Miguel; Ries, Ryan J.; Cao, Huojun; Amendt, Brad A.

2017-01-01

The Iroquois genes (Irx) appear to regulate fundamental processes that lead to cell proliferation, differentiation, and maturation during development. In this report, the Iroquois homeobox 1 (Irx1) transcription factor was functionally disrupted using a LacZ insert and LacZ expression demonstrated stage-specific expression during embryogenesis. Irx1 is highly expressed in the brain, lung, digits, kidney, testis and developing teeth. Irx1 null mice are neonatal lethal and this lethality it due to pulmonary immaturity. Irx1−/− mice show delayed lung maturation characterized by defective surfactant protein secretion and Irx1 marks a population of SP-C expressing alveolar type II cells. Irx1 is specifically expressed in the outer enamel epithelium (OEE), stellate reticulum (SR) and stratum intermedium (SI) layers of the developing tooth. Irx1 mediates dental epithelial cell differentiation in the lower incisors resulting in delayed growth of the lower incisors. Irx1 is specifically and temporally expressed during developmental stages and we have focused on lung and dental development in this report. Irx1+ cells are unique to the development of the incisor outer enamel epithelium, patterning of Lef-1+ and Sox2+ cells as well as a new marker for lung alveolar type II cells. Mechanistically, Irx1 regulates Foxj1 and Sox9 to control cell differentiation during development. PMID:28746823

Irx1 regulates dental outer enamel epithelial and lung alveolar type II epithelial differentiation.

PubMed

Yu, Wenjie; Li, Xiao; Eliason, Steven; Romero-Bustillos, Miguel; Ries, Ryan J; Cao, Huojun; Amendt, Brad A

2017-09-01

The Iroquois genes (Irx) appear to regulate fundamental processes that lead to cell proliferation, differentiation, and maturation during development. In this report, the Iroquois homeobox 1 (Irx1) transcription factor was functionally disrupted using a LacZ insert and LacZ expression demonstrated stage-specific expression during embryogenesis. Irx1 is highly expressed in the brain, lung, digits, kidney, testis and developing teeth. Irx1 null mice are neonatal lethal and this lethality it due to pulmonary immaturity. Irx1 -/- mice show delayed lung maturation characterized by defective surfactant protein secretion and Irx1 marks a population of SP-C expressing alveolar type II cells. Irx1 is specifically expressed in the outer enamel epithelium (OEE), stellate reticulum (SR) and stratum intermedium (SI) layers of the developing tooth. Irx1 mediates dental epithelial cell differentiation in the lower incisors resulting in delayed growth of the lower incisors. Irx1 is specifically and temporally expressed during developmental stages and we have focused on lung and dental development in this report. Irx1+ cells are unique to the development of the incisor outer enamel epithelium, patterning of Lef-1+ and Sox2+ cells as well as a new marker for lung alveolar type II cells. Mechanistically, Irx1 regulates Foxj1 and Sox9 to control cell differentiation during development. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

Characterizing and differentiating task-based and resting state fMRI signals via two-stage sparse representations.

PubMed

Zhang, Shu; Li, Xiang; Lv, Jinglei; Jiang, Xi; Guo, Lei; Liu, Tianming

2016-03-01

A relatively underexplored question in fMRI is whether there are intrinsic differences in terms of signal composition patterns that can effectively characterize and differentiate task-based or resting state fMRI (tfMRI or rsfMRI) signals. In this paper, we propose a novel two-stage sparse representation framework to examine the fundamental difference between tfMRI and rsfMRI signals. Specifically, in the first stage, the whole-brain tfMRI or rsfMRI signals of each subject were composed into a big data matrix, which was then factorized into a subject-specific dictionary matrix and a weight coefficient matrix for sparse representation. In the second stage, all of the dictionary matrices from both tfMRI/rsfMRI data across multiple subjects were composed into another big data-matrix, which was further sparsely represented by a cross-subjects common dictionary and a weight matrix. This framework has been applied on the recently publicly released Human Connectome Project (HCP) fMRI data and experimental results revealed that there are distinctive and descriptive atoms in the cross-subjects common dictionary that can effectively characterize and differentiate tfMRI and rsfMRI signals, achieving 100% classification accuracy. Moreover, our methods and results can be meaningfully interpreted, e.g., the well-known default mode network (DMN) activities can be recovered from the very noisy and heterogeneous aggregated big-data of tfMRI and rsfMRI signals across all subjects in HCP Q1 release.

Reconstructing human pancreatic differentiation by mapping specific cell populations during development

PubMed Central

Ramond, Cyrille; Glaser, Nicolas; Berthault, Claire; Ameri, Jacqueline; Kirkegaard, Jeannette Schlichting; Hansson, Mattias; Honoré, Christian; Semb, Henrik; Scharfmann, Raphaël

2017-01-01

Information remains scarce on human development compared to animal models. Here, we reconstructed human fetal pancreatic differentiation using cell surface markers. We demonstrate that at 7weeks of development, the glycoprotein 2 (GP2) marks a multipotent cell population that will differentiate into the acinar, ductal or endocrine lineages. Development towards the acinar lineage is paralleled by an increase in GP2 expression. Conversely, a subset of the GP2+ population undergoes endocrine differentiation by down-regulating GP2 and CD142 and turning on NEUROG3, a marker of endocrine differentiation. Endocrine maturation progresses by up-regulating SUSD2 and lowering ECAD levels. Finally, in vitro differentiation of pancreatic endocrine cells derived from human pluripotent stem cells mimics key in vivo events. Our work paves the way to extend our understanding of the origin of mature human pancreatic cell types and how such lineage decisions are regulated. DOI: http://dx.doi.org/10.7554/eLife.27564.001 PMID:28731406

Hematopoietic stem cells can differentiate into restricted myeloid progenitors before cell division in mice.

PubMed

Grinenko, Tatyana; Eugster, Anne; Thielecke, Lars; Ramasz, Beáta; Krüger, Anja; Dietz, Sevina; Glauche, Ingmar; Gerbaulet, Alexander; von Bonin, Malte; Basak, Onur; Clevers, Hans; Chavakis, Triantafyllos; Wielockx, Ben

2018-05-15

Hematopoietic stem cells (HSCs) continuously replenish all blood cell types through a series of differentiation steps and repeated cell divisions that involve the generation of lineage-committed progenitors. However, whether cell division in HSCs precedes differentiation is unclear. To this end, we used an HSC cell-tracing approach and Ki67 RFP knock-in mice, in a non-conditioned transplantation model, to assess divisional history, cell cycle progression, and differentiation of adult HSCs. Our results reveal that HSCs are able to differentiate into restricted progenitors, especially common myeloid, megakaryocyte-erythroid and pre-megakaryocyte progenitors, without undergoing cell division and even before entering the S phase of the cell cycle. Additionally, the phenotype of the undivided but differentiated progenitors correlated with the expression of lineage-specific genes and loss of multipotency. Thus HSC fate decisions can be uncoupled from physical cell division. These results facilitate a better understanding of the mechanisms that control fate decisions in hematopoietic cells.

Gene expression profiling of mesenteric lymph nodes from sheep with natural scrapie

PubMed Central

2014-01-01

Background Prion diseases are characterized by the accumulation of the pathogenic PrPSc protein, mainly in the brain and the lymphoreticular system. Although prions multiply/accumulate in the lymph nodes without any detectable pathology, transcriptional changes in this tissue may reflect biological processes that contribute to the molecular pathogenesis of prion diseases. Little is known about the molecular processes that occur in the lymphoreticular system in early and late stages of prion disease. We performed a microarray-based study to identify genes that are differentially expressed at different disease stages in the mesenteric lymph node of sheep naturally infected with scrapie. Oligo DNA microarrays were used to identify gene-expression profiles in the early/middle (preclinical) and late (clinical) stages of the disease. Results In the clinical stage of the disease, we detected 105 genes that were differentially expressed (≥2-fold change in expression). Of these, 43 were upregulated and 62 downregulated as compared with age-matched negative controls. Fewer genes (50) were differentially expressed in the preclinical stage of the disease. Gene Ontology enrichment analysis revealed that the differentially expressed genes were largely associated with the following terms: glycoprotein, extracellular region, disulfide bond, cell cycle and extracellular matrix. Moreover, some of the annotated genes could be grouped into 3 specific signaling pathways: focal adhesion, PPAR signaling and ECM-receptor interaction. We discuss the relationship between the observed gene expression profiles and PrPSc deposition and the potential involvement in the pathogenesis of scrapie of 7 specific differentially expressed genes whose expression levels were confirmed by real time-PCR. Conclusions The present findings identify new genes that may be involved in the pathogenesis of natural scrapie infection in the lymphoreticular system, and confirm previous reports describing scrapie-induced alterations in the expression of genes involved in protein misfolding, angiogenesis and the oxidative stress response. Further studies will be necessary to determine the role of these genes in prion replication, dissemination and in the response of the organism to this disease. PMID:24450868

Impact of Marital Status on Tumor Stage at Diagnosis and on Survival in Male Breast Cancer.

PubMed

Adekolujo, Orimisan Samuel; Tadisina, Shourya; Koduru, Ujwala; Gernand, Jill; Smith, Susan Jane; Kakarala, Radhika Ramani

2017-07-01

The effect of marital status (MS) on survival varies according to cancer type and gender. There has been no report on the impact of MS on survival in male breast cancer (MBC). This study aims to determine the influence of MS on tumor stage at diagnosis and survival in MBC. Men with MBC ≥18 years of age in the SEER database from 1990 to 2011 were included in the study. MS was classified as married and unmarried (including single, divorced, separated, widowed). Kaplan-Meier method was used to estimate the 5-year cancer-specific survival. Multivariate regression analyses were done to determine the effect of MS on presence of Stage IV disease at diagnosis and on cancer-specific mortality. The study included 3,761 men; 2,647 (70.4%) were married. Unmarried men were more often diagnosed with Stage IV MBC compared with married (10.7% vs. 5.5%, p < .001). Unmarried men (compared with married) were significantly less likely to undergo surgery (92.4% vs. 96.7%, p < .001). Overall unmarried males with Stages II, III, and IV MBC have significantly worse 5-year cancer-specific survival compared with married. On multivariate analysis, being unmarried was associated with increased hazard of death (HR = 1.43, p < .001) and increased likelihood of Stage IV disease at diagnosis ( OR = 1.96, p < .001). Unmarried males with breast cancer are at greater risk for Stage IV disease at diagnosis and poorer outcomes compared with married males.

The transcription factors Thpok and LRF are necessary and partly redundant for T helper cell differentiation

PubMed Central

Carpenter, Andrea C.; Grainger, John R.; Xiong, Yumei; Kanno, Yuka; Chu, H. Hamlet; Wang, Lie; Naik, Shruti; dos Santos, Liliane; Wei, Lai; Jenkins, Marc K.; O’Shea, John J.; Belkaid, Yasmine; Bosselut, Rémy

2014-01-01

Summary T helper (Th) cells are critical for defenses against infection and recognize peptides bound to Class II Major Histocompatibility Complex (MHC-II) molecules. Although transcription factors have been identified that direct helper cells into specific effector fates, whether a ‘master’ regulator controls the developmental program common to all Th cells remains unclear. Here we showed that the two transcription factors Thpok and LRF share this function. Although disruption of both factors did not prevent the generation of MHC II-specific T cells, these cells failed to express Th cell genes or undergo Th cell differentiation in vivo. In contrast, T cells lacking Thpok only displayed LRF-dependent functions and contributed to multiple effector responses, both in vitro and in vivo, with the notable exception of Th2 cell responses that control extra-cellular parasites. These findings identify the Thpok-LRF pair as a core node of Th cell differentiation and function. PMID:23041065

Proteins involved in neuronal differentiation of neuroblastoma cell line N1E-115.

PubMed

Oh, Ji-Eun; Freilinger, Angelika; Gelpi, Ellen; Pollak, Arnold; Hengstschläger, Markus; Lubec, Gert

2007-06-01

Neuronal differentiation (ND) represents a well-defined phenomenon in biological terms but proteins involved have not been studied systematically. We therefore aimed to study ND by retinoic acid (RA) in a widely used neuroblastoma cell line by comparative proteomics. The ND was induced in the N1E-115 cell line by serum deprivation and RA treatment. Undifferentiated cells and cells undergoing serum deprivation served as controls. Protein extracts were run on 2-DE followed by MALDI-TOF or MALDI-TOF-TOF analysis. Quantification was carried out using specific software and stringent statistical analysis was performed. Tubulin beta 5, cat eye syndrome critical region protein 5 homolog, putative GTP-binding protein PTD004 homolog, and the metabolic proteins glyceraldehyde-3-phosphate dehydrogenase and transketolase were differentially regulated. Differential protein levels of cytoskeleton proteins including tubulins and metabolic proteins have been reported to be regulated by ND. Herein, specific signaling differences as reflected by putative GTP-binding protein PTD004 changes in differentiated cells are shown and a possible role for the Cat eye syndrome critical region protein 5 homolog is proposed. The protein disulfide isomerase associated 3 protein fits the already proposed findings of chaperon regulation by ND. The study forms the molecular basis for further evaluation of the functional roles of the differentially expressed proteins in ND.

Vertebrate homologues of Frodo are dynamically expressed during embryonic development in tissues undergoing extensive morphogenetic movements.

PubMed

Hunter, Nina L; Hikasa, Hiroki; Dymecki, Susan M; Sokol, Sergei Y

2006-01-01

Frodo has been identified as a protein interacting with Dishevelled, an essential mediator of the Wnt signaling pathway, critical for the determination of cell fate and polarity in embryonic development. In this study, we use specific gene probes to characterize stage- and tissue-specific expression patterns of the mouse Frodo homologue and compare them with Frodo expression patterns in Xenopus embryos. In situ hybridization analysis of mouse Frodo transcripts demonstrates that, similar to Xenopus Frodo, mouse Frodo is expressed in primitive streak mesoderm, neuroectoderm, neural crest, presomitic mesoderm, and somites. In many cases, Frodo expression is confined to tissues undergoing extensive morphogenesis, suggesting that Frodo may be involved in the regulation of cell shape and motility. Highly conserved dynamic expression patterns of Frodo homologues indicate a similar function for these proteins in different vertebrates. 2005 Wiley-Liss, Inc.

Characterization of Mediterranean hail-bearing storms using an operational polarimetric X-band radar

NASA Astrophysics Data System (ADS)

Vulpiani, G.; Baldini, L.; Roberto, N.

2015-07-01

This work documents the fruitul use of X-band radar observations for the monitoring of severe storms in an operational framework. More specifically, a couple of severe hail-bearing Mediterranean storms occurred in 2013 in southern Italy, flooding two important cities of Sicily, are described in terms of their polarimetric radar signatures and retrieved rainfall fields. It is used the X-band dual-polarization radar operating inside the Catania airport (Sicily, Italy), managed by the Italian Department of Civil Protection. A suitable processing is applied to X-band radar measurements. The crucial procedural step relies on the differential phase processing based on an iterative approach that uses a very short-length (1 km) moving window allowing to properly catch the observed high radial gradients of the differential phase. The parameterization of the attenuation correction algorithm, which use the reconstructed differential phase shift, is derived from electromagnetic simulations based on 3 years of DSD observations collected in Rome (Italy). A Fuzzy Logic hydrometeor classification algorithm was also adopted to support the analysis of the storm characteristics. The precipitation fields amount were reconstructed using a combined polarimetric rainfall algorithm based on reflectivity and specific differential phase. The first considered storm was observed on the 21 February, when a winter convective system, originated in the Tyrrhenian sea, hit only marginally the central-eastern coastline of Sicily causing the flash-flood of Catania. Due to the optimal radar location (the system is located at just few kilometers from the city center), it was possible to well retrieve the storm characteristics, including the amount of rainfall field at ground. Extemporaneous signal extinction, caused by close-range hail core causing significant differential phase shift in very short range path, is documented. The second storm, occurred on 21 August 2013, is a summer mesoscale convective system originated by the temperature gradient between sea and land surface, lasted a few hours and eventually flooded the city of Siracusa. The undergoing physical process, including the storm dynamics, is inferred by analysing the vertical sections of the polarimetric radar measurements. The high registered precipitation amount was fairly well reconstructed even though with a trend to underestimation at increasing distances. Several episodes of signal extinction clearly manifested during the mature stage of the observed supercell.

Racial and ethnic disparities over time in the treatment and mortality of women with gynecological malignancies.

PubMed

Rauh-Hain, J Alejandro; Melamed, Alexander; Schaps, Diego; Bregar, Amy J; Spencer, Ryan; Schorge, John O; Rice, Laurel W; Del Carmen, Marcela G

2018-04-01

To examine temporal trends in treatment and survival among black, Asian, Hispanic, and white women diagnosed with endometrial, ovarian, cervical, and vulvar cancer. Using the National Cancer Database (2004-2014), we identified women diagnosed with endometrial, ovarian, cervical, and vulvar cancer. For each disease site, we analyzed race/ethnicity-specific trends in receipt of evidence-based practices. Professional societies' recommendations were used to define these practices. Using data from the Surveillance, Epidemiology, and End Results Program (2000-2009) we analyzed trends in 5-year survival. Throughout the study period black (64.8%) and Hispanic (68.3%) women were less likely to undergo lymphadenectomy for stage I ovarian cancer compared to Asian (79.5%) and white patients (74.6%). Black women were the least likely group to undergo lymphadenectomy in all periods. Among patients with stage II-IV ovarian cancer, 76.6% of white and Asian women received both surgery and chemotherapy, compared to 70.8% of black and 73.9% Hispanic women. Hispanic women with deeply invasive or high-grade stage I endometrial cancer underwent lymphadenectomy less frequently (74.5%) than all other groups (80.7%). Black women were less likely to have chemo-radiotherapy for stage IIB-IVA cervical cancer (75.6% versus 80.4% of all others). Black women were also less likely to have a surgical lymph node evaluation for vulvar cancer (58.8% versus 63.5% of all others). Among women diagnosed with ovarian, endometrial, and cervical cancer, black women had lower five-year survival than other groups. Significant racial disparities persist in the delivery of evidence-based care. Black women with ovarian, endometrial, and cervical cancer continue to experience higher cancer-specific mortality than other groups. Copyright © 2017 Elsevier Inc. All rights reserved.

Minimal clinically important difference and the effect of clinical variables on the ankle osteoarthritis scale in surgically treated end-stage ankle arthritis.

PubMed

Coe, Marcus P; Sutherland, Jason M; Penner, Murray J; Younger, Alastair; Wing, Kevin J

2015-05-20

There is much debate regarding the best outcome tool for use in foot and ankle surgery, specifically in patients with ankle arthritis. The Ankle Osteoarthritis Scale (AOS) is a validated, disease-specific score. The goals of this study were to investigate the clinical performance of the AOS and to determine a minimal clinically important difference (MCID) for it, using a large cohort of 238 patients undergoing surgery for end-stage ankle arthritis. Patients treated with total ankle arthroplasty or ankle arthrodesis were prospectively followed for a minimum of two years at a single site. Data on demographics, comorbidities, AOS score, Short Form-36 results, and the relationship between expectations and satisfaction were collected at baseline (preoperatively), at six and twelve months, and then yearly thereafter. A linear regression analysis examined the variables affecting the change in AOS scores between baseline and the two-year follow-up. An MCID in the AOS change score was then determined by employing an anchor question, which asked patients to rate their relief from symptoms after surgery. Surgical treatment of end-stage ankle arthritis resulted in a mean improvement (and standard deviation) of 31.2 ± 22.7 points in the AOS score two years after surgery. The MCID of the AOS change score was a mean of 28.0 ± 17.9 points. The change in AOS score was significantly affected by the preoperative AOS score, smoking, back pain, and age. Patients undergoing arthroplasty or arthrodesis for end-stage ankle arthritis experienced a mean improvement in AOS score that was greater than the estimated MCID (31.2 versus 28.0 points). Copyright © 2015 by The Journal of Bone and Joint Surgery, Incorporated.

Origin, differentiation and functional ultrastructure of egg envelopes in the cestode Echinococcus multilocularis Leuckart, 1863 (Cyclophyllidea: Taeniidae).

PubMed

Świderski, Zdzisław; Miquel, Jordi; Azzouz-Maache, Samira; Pétavy, Anne-Françoise

2017-07-01

The origin, differentiation and functional ultrastructure of oncospheral or egg envelopes in Echinococcus multilocularis Leuckart, 1863 were studied by transmission electron microscopy (TEM) and cytochemistry. The purpose of our study is to describe the formation of the four primary embryonic envelopes, namely vitelline capsule, outer envelope, inner envelope and oncospheral membrane, and their transformation into the oncospheral or egg envelopes surrounding the mature hexacanth. This transformation takes place in the preoncospheral phase of embryonic development. The vitelline capsule and oncospheral membrane are thin membranes, while the outer and inner envelopes are thick cytoplasmic layers formed by two specific types of blastomeres: the outer envelope by cytoplasmic fusion of two macromeres and the inner envelope by cytoplasmic fusion of three mesomeres. Both outer and inner envelopes are therefore cellular in origin and syncytial in nature. During the advanced phase of embryonic development, the outer and inner envelopes undergo great modifications. The outer envelope remains as a metabolically active layer involved in the storage of glycogen and lipids for the final stages of egg development and survival. The inner envelope is the most important protective layer because of its thick layer of embryophoric blocks that assures oncospheral protection and survival. This embryophore is the principal layer of mature eggs, affording physical and physiological protection for the differentiated embryo or oncosphere, since the outer envelope is stripped from the egg before it is liberated. The embryophore is very thick and impermeable, consisting of polygonal blocks of an inert keratin-like protein held together by a cementing substance. The embryophore therefore assures extreme resistance of eggs, enabling them to withstand a wide range of environmental temperatures and physicochemical conditions.

Complete life cycle of a pennellid Peniculus minuticaudae Shiino, 1956 (Copepoda: Siphonostomatoida) infecting cultured threadsail filefish, Stephanolepis cirrhifer

PubMed Central

Ismail, Norshida; Ohtsuka, Susumu; Venmathi Maran, Balu Alagar; Tasumi, Satoshi; Zaleha, Kassim; Yamashita, Hirofumi

2013-01-01

The complete life cycle of a pennellid copepod Peniculus minuticaudae Shiino, 1956 is proposed based on the discovery of all post-embryonic stages together with the post-metamorphic adult females infecting the fins of threadsail filefish Stephanolepis cirrhifer (Monacanthidae) cultured in a fish farm at Ehime Prefecture, Japan. The hatching stage was the infective copepodid. The life cycle of P. minuticaudae consists of six stages separated by moults: the copepodid, four chalimi and adult. In this study, the adult males were observed frequently in precopulatory amplexus with various stages of females however, copulation occurs only between adults. Fertilized pre-metamorphic adult females carrying spermatophores may detach from the host and settle again before undergoing massive differential growth into the post-metamorphic adult female. Comparison of the life cycle of P. minuticaudae has been made with three known pennellids: Lernaeocera branchialis (Linnaeus, 1767), Cardiodectes medusaeus (Wilson, 1908) and Lernaeenicus sprattae (Sowerby, 1806). Among the compared species, P. minuticaudae is the first ectoparasitic pennellid to be discovered to complete its life cycle on a single host without any change in infection site preferences between infective copepodid and fertilized pre-metamorphic female. PMID:24165196

Early cell lineage specification in a marsupial: a case for diverse mechanisms among mammals.

PubMed

Frankenberg, Stephen; Shaw, Geoff; Freyer, Claudia; Pask, Andrew J; Renfree, Marilyn B

2013-03-01

Early cell lineage specification in eutherian mammals results in the formation of a pluripotent inner cell mass (ICM) and trophoblast. By contrast, marsupials have no ICM. Here, we present the first molecular analysis of mechanisms of early cell lineage specification in a marsupial, the tammar wallaby. There was no overt differential localisation of key lineage-specific transcription factors in cleavage and early unilaminar blastocyst stages. Pluriblast cells (equivalent to the ICM) became distinguishable from trophoblast cells by differential expression of POU5F1 and, to a greater extent, POU2, a paralogue of POU5F1. Unlike in the mouse, pluriblast-trophoblast differentiation coincided with a global nuclear-to-cytoplasmic transition of CDX2 localisation. Also unlike in the mouse, Hippo pathway factors YAP and WWTR1 showed mutually distinct localisation patterns that suggest non-redundant roles. NANOG and GATA6 were conserved as markers of epiblast and hypoblast, respectively, but some differences to the mouse were found in their mode of differentiation. Our results suggest that there is considerable evolutionary plasticity in the mechanisms regulating early lineage specification in mammals.

Molecular hierarchy in neurons differentiated from mouse ES cells containing a single human chromosome 21.

PubMed

Wang, Chi Chiu; Kadota, Mitsutaka; Nishigaki, Ryuichi; Kazuki, Yasuhiro; Shirayoshi, Yasuaki; Rogers, Michael Scott; Gojobori, Takashi; Ikeo, Kazuho; Oshimura, Mitsuo

2004-02-06

Defects in neurogenesis and neuronal differentiation in the fetal brain of Down syndrome (DS) patients lead to the apparent neuropathological abnormalities and contribute to the phenotypic characters of mental retardation, and premature development of Alzheimer's disease, those being the most common phenotype in DS. In order to understand the molecular mechanism underlying the cause of phenotypic abnormalities in the DS brain, we have utilized an in vitro model of TT2F mouse embryonic stem cells containing a single human chromosome 21 (hChr21) to study neuron development and neuronal differentiation by microarray containing 15K developmentally expressed cDNAs. Defective neuronal differentiation in the presence of extra hChr21 manifested primarily the post-transcriptional and translational modification, such as Mrpl10, SNAPC3, Srprb, SF3a60 in the early neuronal stem cell stage, and Mrps18a, Eef1g, and Ubce8 in the late differentiated stage. Hierarchical clustering patterned specific expression of hChr21 gene dosage effects on neuron outgrowth, migration, and differentiation, such as Syngr2, Dncic2, Eif3sf, and Peg3.

A comparison of the prognostic value of preoperative inflammation-based scores and TNM stage in patients with gastric cancer.

PubMed

Pan, Qun-Xiong; Su, Zi-Jian; Zhang, Jian-Hua; Wang, Chong-Ren; Ke, Shao-Ying

2015-01-01

People's Republic of China is one of the countries with the highest incidence of gastric cancer, accounting for 45% of all new gastric cancer cases in the world. Therefore, strong prognostic markers are critical for the diagnosis and survival of Chinese patients suffering from gastric cancer. Recent studies have begun to unravel the mechanisms linking the host inflammatory response to tumor growth, invasion and metastasis in gastric cancers. Based on this relationship between inflammation and cancer progression, several inflammation-based scores have been demonstrated to have prognostic value in many types of malignant solid tumors. To compare the prognostic value of inflammation-based prognostic scores and tumor node metastasis (TNM) stage in patients undergoing gastric cancer resection. The inflammation-based prognostic scores were calculated for 207 patients with gastric cancer who underwent surgery. Glasgow prognostic score (GPS), neutrophil lymphocyte ratio (NLR), platelet lymphocyte ratio (PLR), prognostic nutritional index (PNI), and prognostic index (PI) were analyzed. Linear trend chi-square test, likelihood ratio chi-square test, and receiver operating characteristic were performed to compare the prognostic value of the selected scores and TNM stage. In univariate analysis, preoperative serum C-reactive protein (P<0.001), serum albumin (P<0.001), GPS (P<0.001), PLR (P=0.002), NLR (P<0.001), PI (P<0.001), PNI (P<0.001), and TNM stage (P<0.001) were significantly associated with both overall survival and disease-free survival of patients with gastric cancer. In multivariate analysis, GPS (P=0.024), NLR (P=0.012), PI (P=0.001), TNM stage (P<0.001), and degree of differentiation (P=0.002) were independent predictors of gastric cancer survival. GPS and TNM stage had a comparable prognostic value and higher linear trend chi-square value, likelihood ratio chi-square value, and larger area under the receiver operating characteristic curve as compared to other inflammation-based prognostic scores. The present study indicates that preoperative GPS and TNM stage are robust predictors of gastric cancer survival as compared to NLR, PLR, PI, and PNI in patients undergoing tumor resection.

A comparison of the prognostic value of preoperative inflammation-based scores and TNM stage in patients with gastric cancer

PubMed Central

Pan, Qun-Xiong; Su, Zi-Jian; Zhang, Jian-Hua; Wang, Chong-Ren; Ke, Shao-Ying

2015-01-01

Background People’s Republic of China is one of the countries with the highest incidence of gastric cancer, accounting for 45% of all new gastric cancer cases in the world. Therefore, strong prognostic markers are critical for the diagnosis and survival of Chinese patients suffering from gastric cancer. Recent studies have begun to unravel the mechanisms linking the host inflammatory response to tumor growth, invasion and metastasis in gastric cancers. Based on this relationship between inflammation and cancer progression, several inflammation-based scores have been demonstrated to have prognostic value in many types of malignant solid tumors. Objective To compare the prognostic value of inflammation-based prognostic scores and tumor node metastasis (TNM) stage in patients undergoing gastric cancer resection. Methods The inflammation-based prognostic scores were calculated for 207 patients with gastric cancer who underwent surgery. Glasgow prognostic score (GPS), neutrophil lymphocyte ratio (NLR), platelet lymphocyte ratio (PLR), prognostic nutritional index (PNI), and prognostic index (PI) were analyzed. Linear trend chi-square test, likelihood ratio chi-square test, and receiver operating characteristic were performed to compare the prognostic value of the selected scores and TNM stage. Results In univariate analysis, preoperative serum C-reactive protein (P<0.001), serum albumin (P<0.001), GPS (P<0.001), PLR (P=0.002), NLR (P<0.001), PI (P<0.001), PNI (P<0.001), and TNM stage (P<0.001) were significantly associated with both overall survival and disease-free survival of patients with gastric cancer. In multivariate analysis, GPS (P=0.024), NLR (P=0.012), PI (P=0.001), TNM stage (P<0.001), and degree of differentiation (P=0.002) were independent predictors of gastric cancer survival. GPS and TNM stage had a comparable prognostic value and higher linear trend chi-square value, likelihood ratio chi-square value, and larger area under the receiver operating characteristic curve as compared to other inflammation-based prognostic scores. Conclusion The present study indicates that preoperative GPS and TNM stage are robust predictors of gastric cancer survival as compared to NLR, PLR, PI, and PNI in patients undergoing tumor resection. PMID:26124667

2-(trimethylammonium)ethyl (R)-3-methoxy-3-oxo-2-stearamidopropyl phosphate promotes megakaryocytic differentiation of myeloid leukaemia cells and primary human CD34⁺ haematopoietic stem cells.

PubMed

Limb, Jin-Kyung; Song, Doona; Jeon, Mijeong; Han, So-Yeop; Han, Gyoonhee; Jhon, Gil-Ja; Bae, Yun Soo; Kim, Jaesang

2015-04-01

In this study we showed that 2-(trimethylammonium)ethyl (R)-3-methoxy-3-oxo-2-stearamidopropyl phosphate [(R)-TEMOSPho], a derivative of an organic chemical identified from a natural product library, promotes highly efficient differentiation of megakaryocytes. Specifically, (R)-TEMOSPho induces cell cycle arrest, cell size increase and polyploidization from K562 and HEL cells, which are used extensively to model megakaryocytic differentiation. In addition, megakaryocyte-specific cell surface markers showed a dramatic increase in expression in response to (R)-TEMOSPho treatment. Importantly, we demonstrated that such megakaryocytic differentiation can also be induced from primary human CD34(+) haematopoietic stem cells. Activation of the PI3K-AKT pathway and, to a lesser extent, the MEK-ERK pathway appears to be required for this process, as blocking with specific inhibitors interferes with the differentiation of K562 cells. A subset of (R)-TEMOSPho-treated K562 cells undergoes spontaneous apoptosis and produces platelets that are apparently functional, as they bind to fibrinogen, express P-selectin and aggregate in response to SFLLRN and AYPGFK, the activating peptides for the PAR1 and PAR4 receptors, respectively. Taken together, these results indicate that (R)-TEMOSPho will be useful for dissecting the molecular mechanisms of megakaryocytic differentiation, and that this class of compounds represents potential therapeutic reagents for thrombocytopenia. Copyright © 2012 John Wiley & Sons, Ltd.

Different tissue phagocytes sample apoptotic cells to direct distinct homeostasis programs

PubMed Central

Cummings, Ryan J.; Barbet, Gaetan; Bongers, Gerold; Hartmann, Boris M.; Gettler, Kyle; Muniz, Luciana; Furtado, Glaucia C.; Cho, Judy; Lira, Sergio A.; Blander, J. Magarian

2017-01-01

Recognition and removal of apoptotic cells by professional phagocytes, including dendritic cells and macrophages, preserves immune self-tolerance and prevents chronic inflammation and autoimmune pathologies1,2. The diverse array of phagocytes that reside within different tissues, combined with the necessarily prompt nature of apoptotic cell clearance, makes it difficult to study this process in situ. The full spectrum of functions executed by tissue-resident phagocytes in response to homeostatic apoptosis, therefore, remains unclear. Here we show that mouse apoptotic intestinal epithelial cells (IECs), which undergo continuous renewal to maintain optimal barrier and absorptive functions3, are not merely extruded to maintain homeostatic cell numbers4, but are also sampled by a single subset of dendritic cells and two macrophage subsets within a well-characterized network of phagocytes in the small intestinal lamina propria5,6. Characterization of the transcriptome within each subset before and after in situ sampling of apoptotic IECs revealed gene expression signatures unique to each phagocyte, including macrophage-specific lipid metabolism and amino acid catabolism, and a dendritic-cell-specific program of regulatory CD4+ T-cell activation. A common ‘suppression of inflammation’ signature was noted, although the specific genes and pathways involved varied amongst dendritic cells and macrophages, reflecting specialized functions. Apoptotic IECs were trafficked to mesenteric lymph nodes exclusively by the dendritic cell subset and served as critical determinants for the induction of tolerogenic regulatory CD4+ T-cell differentiation. Several of the genes that were differentially expressed by phagocytes bearing apoptotic IECs overlapped with susceptibility genes for inflammatory bowel disease7. Collectively, these findings provide new insights into the consequences of apoptotic cell sampling, advance our understanding of how homeostasis is maintained within the mucosa and set the stage for development of novel therapeutics to alleviate chronic inflammatory diseases such as inflammatory bowel disease. PMID:27828940

Conditional induction of Math1 specifies embryonic stem cells to cerebellar granule neuron lineage and promotes differentiation into mature granule neurons.

PubMed

Srivastava, Rupali; Kumar, Manoj; Peineau, Stéphane; Csaba, Zsolt; Mani, Shyamala; Gressens, Pierre; El Ghouzzi, Vincent

2013-04-01

Directing differentiation of embryonic stem cells (ESCs) to specific neuronal subtype is critical for modeling disease pathology in vitro. An attractive means of action would be to combine regulatory differentiation factors and extrinsic inductive signals added to the culture medium. In this study, we have generated mature cerebellar granule neurons by combining a temporally controlled transient expression of Math1, a master gene in granule neuron differentiation, with inductive extrinsic factors involved in cerebellar development. Using a Tetracyclin-On transactivation system, we overexpressed Math1 at various stages of ESCs differentiation and found that the yield of progenitors was considerably increased when Math1 was induced during embryonic body stage. Math1 triggered expression of Mbh1 and Mbh2, two target genes directly involved in granule neuron precursor formation and strong expression of early cerebellar territory markers En1 and NeuroD1. Three weeks after induction, we observed a decrease in the number of glial cells and an increase in that of neurons albeit still immature. Combining Math1 induction with extrinsic factors specifically increased the number of neurons that expressed Pde1c, Zic1, and GABAα6R characteristic of mature granule neurons, formed "T-shaped" axons typical of granule neurons, and generated synaptic contacts and action potentials in vitro. Finally, in vivo implantation of Math1-induced progenitors into young adult mice resulted in cell migration and settling of newly generated neurons in the cerebellum. These results show that conditional induction of Math1 drives ESCs toward the cerebellar fate and indicate that acting on both intrinsic and extrinsic factors is a powerful means to modulate ESCs differentiation and maturation into a specific neuronal lineage. Copyright © 2012 AlphaMed Press.

Arid3a is essential to execution of the first cell fate decision via direct embryonic and extraembryonic transcriptional regulation

PubMed Central

Rhee, Catherine; Lee, Bum-Kyu; Beck, Samuel; Anjum, Azeen; Cook, Kendra R.; Popowski, Melissa

2014-01-01

Despite their origin from the inner cell mass, embryonic stem (ES) cells undergo differentiation to the trophectoderm (TE) lineage by repression of the ES cell master regulator Oct4 or activation of the TE master regulator Caudal-type homeobox 2 (Cdx2). In contrast to the in-depth studies of ES cell self-renewal and pluripotency, few TE-specific regulators have been identified, thereby limiting our understanding of mechanisms underlying the first cell fate decision. Here we show that up-regulation and nuclear entry of AT-rich interactive domain 3a (Arid3a) drives TE-like transcriptional programs in ES cells, maintains trophoblast stem (TS) cell self-renewal, and promotes further trophoblastic differentiation both upstream and independent of Cdx2. Accordingly, Arid3a−/− mouse post-implantation placental development is severely impaired, resulting in early embryonic death. We provide evidence that Arid3a directly activates TE-specific and trophoblast lineage-specific genes while directly repressing pluripotency genes via differential regulation of epigenetic acetylation or deacetylation. Our results identify Arid3a as a critical regulator of TE and placental development through execution of the commitment and differentiation phases of the first cell fate decision. PMID:25319825

Anti-citrullinated protein antibody and rheumatoid factor in patients with end-stage renal disease.

PubMed

Romic, Zeljko; Unic, Adriana; Derek, Lovorka; Zivkovic, Marcela; Marijancevic, Domagoj; Kes, Petar; Pehar, Mario

2009-01-01

Patients with end-stage renal disease (ESRD) and on hemodialysis (HD) are at increased risk for developing rheumatoid arthritis (RA), as a result of defective immunity. Our aim was to examine if ESRD and the length of HD treatment impact the clinical utility of antibodies to cyclic citrullinated peptides (anti-CCP) and rheumatoid factor (RF) as diagnostic tools for RA. We included 94 subjects in our study: 37 healthy volunteers and 57 patients with ESRD who had been undergoing HD for 1-12 years, and without confirmed RA. In order to test our hypothesis, we measured and correlated anti-CCP and RF as laboratory markers of RA. Our study showed that there is no significant difference between values for anti-CCP (p=0.11) and RF (p=0.98) in control subjects as well as in patients undergoing HD, regardless of the length of time that patients had been undergoing HD treatment. Our study indicates that HD does not impair the specificity of anti-CCP and RF for RA in patients where the disease has not yet developed. Future prospective studies may show whether there is any use in determinating RF, and especially anti-CCP, as early predictors of RA in patients with ESRD who are at greater risk of developing this condition.

Adult human pancreas-derived cells expressing stage-specific embryonic antigen 4 differentiate into Sox9-expressing and Ngn3-expressing pancreatic ducts in vivo.

PubMed

Lee, Song; Lee, Chan Mi; Kim, Song Cheol

2016-11-11

Tissue-specific stem/progenitor cells are found in various adult tissues and may have the capacity for lineage-specific differentiation, facilitating applications in autologous transplantation. Stage-specific embryonic antigen 4 (SSEA-4), an early embryonic glycolipid antigen, is expressed in cells derived from adult human pancreas exocrine tissue. Here, we examined the characteristics and lineage-specific differentiation capacity of SSEA-4 + cells. Human adult partial pancreas tissues were obtained from different donors and cultured in vitro. SSEA-4 + and CA19-9 + cells were isolated from adult human pancreas exocrine cells using magnetic-activated cell sorting, and gene expression was validated by quantitative polymerase chain reaction. To confirm in-vivo differentiation, SSEA-4 + and CA19-9 + cells were transplanted into the dorsal subcutaneous region of mice. Finally, morphological features of differentiated areas were confirmed by immunostaining and morphometric analysis. SSEA-4-expressing cells were detected in isolated pancreas exocrine cells from adult humans. These SSEA-4 + cells exhibited coexpression of CA19-9, a marker of pancreatic duct cells, but not amylase expression, as shown by immunostaining and flow cytometry. SSEA-4 + cells exhibited higher relative expression of Oct4, Nanog, Klf4, Sox2, and c-Myc mRNAs than CA19-9 + cells. Pancreatic intralobular ducts (PIDs) were generated from SSEA-4 + or CA19-9 + cells in vivo at 5 weeks after transplantation. However, newly formed PIDs from CA19-9 + cells were less abundant and showed an incomplete PID morphology. In contrast, newly formed PIDs from SSEA-4 + cells were abundant in the transplanted area and showed a crowded morphology, typical of PIDs. Sox9 and Ngn3, key transcription factors associated with pancreatic development and regeneration, were expressed in PIDs from SSEA-4 + cells. SSEA-4-expressing cells in the adult human pancreas may have the potential for regeneration of the pancreas and may be used as a source of stem/progenitor cells for pancreatic cell lineage-specific differentiation.

Proteomic analysis of the nuclear matrix in the early stages of rat liver carcinogenesis: Identification of differentially expressed and MAR-binding proteins

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

Barboro, Paola; D'Arrigo, Cristina; Repaci, Erica

Tumor progression is characterized by definite changes in the protein composition of the nuclear matrix (NM). The interactions of chromatin with the NM occur via specific DNA sequences called MARs (matrix attachment regions). In the present study, we applied a proteomic approach along with a Southwestern assay to detect both differentially expressed and MAR-binding NM proteins, in persistent hepatocyte nodules (PHN) in respect with normal hepatocytes (NH). In PHN, the NM undergoes changes both in morphology and in protein composition. We detected over 500 protein spots in each two dimensional map and 44 spots were identified. Twenty-three proteins were differentiallymore » expressed; among these, 15 spots were under-expressed and 8 spots were over-expressed in PHN compared to NH. These changes were synchronous with several modifications in both NM morphology and the ability of NM proteins to bind nuclear RNA and/or DNA containing MARs sequences. In PHN, we observed a general decrease in the expression of the basic proteins that bound nuclear RNA and the over-expression of two species of Mw 135 kDa and 81 kDa and pI 6.7-7.0 and 6.2-7.4, respectively, which exclusively bind to MARs. These results suggest that the deregulated expression of these species might be related to large-scale chromatin reorganization observed in the process of carcinogenesis by modulating the interaction between MARs and the scaffold structure.« less

Combined Transcriptome and Proteome Analysis Identifies Pathways and Markers Associated with the Establishment of Rapeseed Microspore-Derived Embryo Development1[W

PubMed Central

Joosen, Ronny; Cordewener, Jan; Supena, Ence Darmo Jaya; Vorst, Oscar; Lammers, Michiel; Maliepaard, Chris; Zeilmaker, Tieme; Miki, Brian; America, Twan; Custers, Jan; Boutilier, Kim

2007-01-01

Microspore-derived embryo (MDE) cultures are used as a model system to study plant cell totipotency and as an in vitro system to study embryo development. We characterized and compared the transcriptome and proteome of rapeseed (Brassica napus) MDEs from the few-celled stage to the globular/heart stage using two MDE culture systems: conventional cultures in which MDEs initially develop as unorganized clusters that usually lack a suspensor, and a novel suspensor-bearing embryo culture system in which the embryo proper originates from the distal cell of a suspensor-like structure and undergoes the same ordered cell divisions as the zygotic embryo. Improved histodifferentiation of suspensor-bearing MDEs suggests a new role for the suspensor in driving embryo cell identity and patterning. An MDE culture cDNA array and two-dimensional gel electrophoresis and protein sequencing were used to compile global and specific expression profiles for the two types of MDE cultures. Analysis of the identities of 220 candidate embryo markers, as well as the identities of 32 sequenced embryo up-regulated protein spots, indicate general roles for protein synthesis, glycolysis, and ascorbate metabolism in the establishment of MDE development. A collection of 135 robust markers for the transition to MDE development was identified, a number of which may be coregulated at the gene and protein expression level. Comparison of the expression profiles of preglobular-stage conventional MDEs and suspensor-bearing MDEs identified genes whose differential expression may reflect improved histodifferentiation of suspensor-bearing embryos. This collection of early embryo-expressed genes and proteins serves as a starting point for future marker development and gene function studies aimed at understanding the molecular regulation of cell totipotency and early embryo development in plants. PMID:17384159

Transient chondrogenic phase in the intramembranous pathway during normal skeletal development.

PubMed

Nah, H D; Pacifici, M; Gerstenfeld, L C; Adams, S L; Kirsch, T

2000-03-01

Calvarial and facial bones form by intramembranous ossification, in which bone cells arise directly from mesenchyme without an intermediate cartilage anlage. However, a number of studies have reported the emergence of chondrocytes from in vitro calvarial cell or organ cultures and the expression of type II collagen, a cartilage-characteristic marker, in developing calvarial bones. Based on these findings we hypothesized that a covert chondrogenic phase may be an integral part of the normal intramembranous pathway. To test this hypothesis, we analyzed the temporal and spatial expression patterns of cartilage characteristic genes in normal membranous bones from chick embryos at various developmental stages (days 12, 15 and 19). Northern and RNAse protection analyses revealed that embryonic frontal bones expressed not only the type I collagen gene but also a subset of cartilage characteristic genes, types IIA and XI collagen and aggrecan, thus resembling a phenotype of prechondrogenic-condensing mesenchyme. The expression of cartilage-characteristic genes decreased with the progression of bone maturation. Immunohistochemical analyses of developing embryonic chick heads indicated that type II collagen and aggrecan were produced by alkaline phosphatase activity positive cells engaged in early stages of osteogenic differentiation, such as cells in preosteogenic-condensing mesenchyme, the cambium layer of periosteum, the advancing osteogenic front, and osteoid bone. Type IIB and X collagen messenger RNAs (mRNA), markers for mature chondrocytes, were also detected at low levels in calvarial bone but not until late embryonic stages (day 19), indicating that some calvarial cells may undergo overt chondrogenesis. On the basis of our findings, we propose that the normal intramembranous pathway in chicks includes a previously unrecognized transient chondrogenic phase similar to prechondrogenic mesenchyme, and that the cells in this phase retain chondrogenic potential that can be expressed in specific in vitro and in vivo microenvironments.

Differential expression of syndecan isoforms during mouse incisor amelogenesis.

PubMed

Muto, Taro; Miyoshi, Keiko; Munesue, Seiichi; Nakada, Hiroshi; Okayama, Minoru; Matsuo, Takashi; Noma, Takafumi

2007-08-01

Syndecans are transmembranous heparan sulfate proteoglycans (HSPGs) with covalently attached glycosaminoglycan side-chains located on the cell surface. The mammalian syndecan family is composed of four types of syndecans (syndecan-1 to -4). Syndecans interact with the intracellular cytoskeleton through the cytoplasmic domains of their core proteins and membrane proteins, extracellular enzymes, growth factors, and matrix components, through their heparan-sulfate chains, to regulate developmental processes.Here, as a first step to assess the possible roles of syndecan proteins in amelogenesis, we examined the expression patterns of all syndecan isoforms in continuously growing mouse incisors, in which we can overview major differentiation stages of amelogenesis at a glance. Understanding the expression domain of each syndecan isoform during specific developmental stages seems useful for investigating their physiological roles in amelogenesis. Immunohistochemical analysis of syndecan core proteins in the lower incisors from postnatal day 1 mice revealed spatially and temporally specific expression patterns, with syndecan-1 expressed in undifferentiated epithelial and mesenchymal cells, and syndecan-2, -3, and -4 in more differentiated cells. These findings suggest that each syndecan isoform functions distinctly during the amelogenesis of the incisors of mice.

In vitro development of Strongylus edentatus to the fourth larval stage with notes on Strongylus vulgaris and Strongylus equinus.

PubMed

Farrar, R G; Klei, T R

1985-08-01

Strongylus edentatus was successfully cultured in vitro to the fourth larval stage (L4). Some growth continued for periods of 40-50 days at which time reductions in viability were observed in some of the culture systems tested. Various combinations of media, sera, buffers and organ explant cultures were tested. All cultures were incubated at 37 C in an atmosphere of 95% air and 5% CO2. Larvae underwent growth and differentiation to the L4 in all medium-serum combinations with and without organ explant cultures. Development and growth did occur but viability was reduced to insignificant levels in media without serum or cells. Optimal growth, differentiation, and longevity were observed in bicarbonate buffered RPMI-1640 containing 10% fetal calf serum and gerbil (Meriones unguiculatus) cecum explant cultures. Observations indicated that Strongylus vulgaris and Strongylus equinus also developed to the L4 stage using similar techniques. However, viability of S. vulgaris L4 was markedly limited. Specific morphological changes marked phases of development of S. edentatus, categorized as early, middle and late third stage, third molt and early fourth stage. Strongylus equinus appeared to follow the same developmental pattern in vitro as S. edentatus. Distinct differences in morphological features during differentiation were observed between S. edentatus and S. vulgaris.

Plasticity of brain wave network interactions and evolution across physiologic states

PubMed Central

Liu, Kang K. L.; Bartsch, Ronny P.; Lin, Aijing; Mantegna, Rosario N.; Ivanov, Plamen Ch.

2015-01-01

Neural plasticity transcends a range of spatio-temporal scales and serves as the basis of various brain activities and physiologic functions. At the microscopic level, it enables the emergence of brain waves with complex temporal dynamics. At the macroscopic level, presence and dominance of specific brain waves is associated with important brain functions. The role of neural plasticity at different levels in generating distinct brain rhythms and how brain rhythms communicate with each other across brain areas to generate physiologic states and functions remains not understood. Here we perform an empirical exploration of neural plasticity at the level of brain wave network interactions representing dynamical communications within and between different brain areas in the frequency domain. We introduce the concept of time delay stability (TDS) to quantify coordinated bursts in the activity of brain waves, and we employ a system-wide Network Physiology integrative approach to probe the network of coordinated brain wave activations and its evolution across physiologic states. We find an association between network structure and physiologic states. We uncover a hierarchical reorganization in the brain wave networks in response to changes in physiologic state, indicating new aspects of neural plasticity at the integrated level. Globally, we find that the entire brain network undergoes a pronounced transition from low connectivity in Deep Sleep and REM to high connectivity in Light Sleep and Wake. In contrast, we find that locally, different brain areas exhibit different network dynamics of brain wave interactions to achieve differentiation in function during different sleep stages. Moreover, our analyses indicate that plasticity also emerges in frequency-specific networks, which represent interactions across brain locations mediated through a specific frequency band. Comparing frequency-specific networks within the same physiologic state we find very different degree of network connectivity and link strength, while at the same time each frequency-specific network is characterized by a different signature pattern of sleep-stage stratification, reflecting a remarkable flexibility in response to change in physiologic state. These new aspects of neural plasticity demonstrate that in addition to dominant brain waves, the network of brain wave interactions is a previously unrecognized hallmark of physiologic state and function. PMID:26578891

1α,25(OH)2-Vitamin D3 Inhibits C2C12 Cell Differentiation by Activating c-Src and ERK1/2.

PubMed

Wang, Zhonghua; Jiang, Aijun; Mei, Jingwei; Zhang, Xinyan

2018-05-01

The steroid hormone 1α,25(OH)2-vitamin D3 (1,25-D3) induced some biological responses through activation of MAPK cascades in various cell types. It seems that 1,25-D3 plays different roles at different stages of proliferating, differentiating, and differentiated C2C12 cells. We wanted to detect the effect of 1,25-D3 on myogenic differentiation and the role of ERK1/2 in differentiating stage induced by 2% horse serum with 1,25-D3. In this study, cells were induced to differentiate with 2% horse serum until the 7th day (with addition of 1,25-D3 every two days). The protein level of MHC (myosin heavy chain) and phosphorylation level of Src and ERK1/2 were determined with western blot. U0126 (MEK inhibitor) and PP2 (Src specific inhibitor) were used to confirm the relationship between 1,25-D3, MHC, Src, and ERK1/2. 1,25-D3 inhibited differentiation of C2C12 cells and fusion of myotubes by phosphorylating and activating Src and ERK1/2. Phosphorylation of ERK1/2 was inhibited, not only by U0126 but also by PP2 (a Src specific inhibitor) which led to the promotion of differentiation of C2C12 cells; however, U0126 did not inhibit Src phosphorylation. These results suggested that 1,25-D3 possibly inhibited C2C12 differentiation through Src and ERK1/2, and Src played an upstream role in this signaling pathway.

Site-specific tumor grading system in colorectal cancer: multicenter pathologic review of the value of quantifying poorly differentiated clusters.

PubMed

Ueno, Hideki; Hase, Kazuo; Hashiguchi, Yojiro; Shimazaki, Hideyuki; Tanaka, Masafumi; Miyake, Ohki; Masaki, Tadahiko; Shimada, Yoshifumi; Kinugasa, Yusuke; Mori, Yoshiyuki; Kishimoto, Mitsuo; Kameoka, Shingo; Sato, Yu; Matsuda, Keiji; Nakadoi, Koichi; Shinto, Eiji; Nakamura, Takahiro; Sugihara, Kenichi

2014-02-01

The study aimed to determine the value of a novel site-specific grading system based on quantifying poorly differentiated clusters (PDC; Grade(PDC)) in colorectal cancer (CRC). A multicenter pathologic review involving 12 institutions was performed on 3243 CRC cases (stage I, 583; II, 1331; III, 1329). Cancer clusters of ≥5 cancer cells and lacking a gland-like structure (PDCs) were counted under a ×20 objective lens in a field containing the maximum clusters. Tumors with <5, 5 to 9, and ≥10 PDCs were classified as grades G1, G2, and G3, respectively. According to Grade(PDC), 1594, 1005, and 644 tumors were classified as G1, G2, and G3 and had 5-year recurrence-free survival rates of 91.6%, 75.4%, and 59.6%, respectively (P<0.0001). Multivariate analysis showed that Grade exerted an influence on prognostic outcome independently of TNM staging; approximately 20% and 46% of stage I and II patients, respectively, were selected by Grade(PDC) as a population whose survival estimate was comparable to or even worse than that of stage III patients. Grade(PDC) surpassed TNM staging in the ability to stratify patients by recurrence-free survival (Akaike information criterion, 2915.6 vs. 2994.0) and had a higher prognostic value than American Joint Committee on Cancer (AJCC) grading (Grade(AJCC)) at all stages. Regarding judgment reproducibility of grading tumors, weighted κ among the 12 institutions was 0.40 for Grade(AJCC) and 0.52 for Grade(PDC). Grade(PDC) has a robust prognostic power and promises to be of sufficient clinical value to merit implementation as a site-specific grading system in CRC.

Determination of 17OHPreg and DHEAS by LC-MS/MS: Impact of Age, Sex, Pubertal Stage, and BMI on the Δ5 Steroid Pathway.

PubMed

Kulle, Alexandra E; Reinehr, Thomas; Simic-Schleicher, Gunter; Hornig, Nadine C; Holterhus, Paul-Martin

2017-01-01

Dehydroepiandrosterone sulfate (DHEAS) and 17-hydroxypregnenolone (17OHPreg) are important for understanding the Δ5 pathway (e.g., in adrenarche and obesity). Although mass spectrometry has become the state-of-the-art method for quantifying steroids, there are few comprehensive age-, sex-, and pubertal stage-specific reference ranges for children. To develop a sensitive and reliable ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method for simultaneous quantification of DHEAS and 17OHPreg and to establish entire age-, sex- and pubertal stage-specific reference ranges in children. A total of 684 children, 453 (243 female, 210 male) with normal body mass index (BMI; <90th) and 231 (132 female, 99 male) obese subjects (>97th), were categorized into 11 age groups, and age- and Tanner stage (PH)-specific reference ranges were determined. The limit of detection was 0.05 nmol/L for 17OHPreg and 0.5 nmol/L for DHEAS. Levels of both steroids declined after the neonatal period. Comparisons with RIA assays (Siemens, Munich, Germany) (DHEAS) and an in-house kit (17OHPreg) revealed 0.95 and 0.93, respectively, as coefficients of determination. Although DHEAS-generally higher in boys-increased continuously starting at 3 to 6 years, 17OHPreg remained largely constant. In obese patients, both were significantly elevated, also in part after alignment to Tanner stages (PH). UPLC-MS/MS is sensitive and reliable for quantifying DHEAS and 17OHPreg. Our data support differential maturation of CYP17 during adrenarche with successively increasing 17,20-lyase activity but largely constant 17α-hydroxylation activity. Endocrine interpretation of 17OHPreg and DHEAS must consider differential patterns for age, sex, pubertal stage, and BMI. Copyright © 2017 by the Endocrine Society

Specific Biomimetic Hydroxyapatite Nanotopographies Enhance Osteoblastic Differentiation and Bone Graft Osteointegration

PubMed Central

Loiselle, Alayna E.; Wei, Lai; Faryad, Muhammad; Paul, Emmanuel M.; Lewis, Gregory S.; Gao, Jun; Lakhtakia, Akhlesh

2013-01-01

Impaired healing of cortical bone grafts represents a significant clinical problem. Cadaveric bone grafts undergo extensive chemical processing to decrease the risk of disease transmission; however, these processing techniques alter the bone surface and decrease the osteogenic potential of cells at the healing site. Extensive work has been done to optimize the surface of bone grafts, and hydroxyapatite (HAP) and nanotopography both increase osteoblastic differentiation. HAP is the main mineral component of bone and can enhance osteoblastic differentiation and bone implant healing in vivo, while nanotopography can enhance osteoblastic differentiation, adhesion, and proliferation. This is the first study to test the combined effects of HAP and nanotopographies on bone graft healing. With the goal of identifying the optimized surface features to improve bone graft healing, we tested the hypothesis that HAP-based nanotopographic resurfacing of bone grafts improves integration of cortical bone grafts by enhancing osteoblastic differentiation. Here we show that osteoblastic cells cultured on processed bones coated with specific-scale (50–60 nm) HAP nanotopographies display increased osteoblastic differentiation compared to cells on uncoated bone, bones coated with poly-l-lactic acid nanotopographies, or other HAP nanotopographies. Further, bone grafts coated with 50–60-nm HAP exhibited increased formation of new bone and improved healing, with mechanical properties equivalent to live autografts. These data indicate the potential for specific HAP nanotopographies to not only increase osteoblastic differentiation but also improve bone graft incorporation, which could significantly increase patient quality of life after traumatic bone injuries or resection of an osteosarcoma. PMID:23510012

Development of thalamocortical connections between the mediodorsal thalamus and the prefrontal cortex and its implication in cognition

PubMed Central

Ferguson, Brielle R.; Gao, Wen-Jun

2015-01-01

The mediodorsal thalamus (MD) represents a fundamental subcortical relay to the prefrontal cortex (PFC), and is thought to be highly implicated in modulation of cognitive performance. Additionally, it undergoes highly conserved developmental stages, which, when dysregulated, can have detrimental consequences. Embryonically, the MD experiences a tremendous surge in neurogenesis and differentiation, and disruption of this process may underlie the pathology in certain neurodevelopmental disorders. However, during the postnatal period, a vast amount of cell loss in the MD occurs. These together may represent an extended critical period for postnatal development, in which disturbances in the normal growth or reduction of the MD afferents to the PFC, can result in PFC-dependent cognitive, affective, or psychotic abnormalities. In this review, we explore the current knowledge supporting this hypothesis of a protracted critical period, and propose how developmental changes in the MD contribute to successful prefrontal cortical development and function. Specifically, we elaborate on the unique properties of MD-PFC connections compared with other thalamocortical afferents in sensory cortices, examine how MD-PFC innervation modulates synaptic transmission in the local prefrontal circuitry, and speculate on what occurs during postnatal development, particularly within the early neonatal stage, as well as juvenile and adolescent periods. Finally, we discuss the questions that remain and propose future experiments in order to provide perspective and novel insights into the cause of neuropsychiatric disorders associated with MD-PFC development. PMID:25620923

Quantifying Ciliary Dynamics during Assembly Reveals Step-wise Waveform Maturation in Airway Cells.

PubMed

Oltean, Alina; Schaffer, Andrew J; Bayly, Philip V; Brody, Steven L

2018-05-31

Motile cilia are essential for clearance of particulates and pathogens from airways. For effective transport, ciliary motor proteins and axonemal structures interact to generate the rhythmic, propulsive bending, but the mechanisms that produce a dynamic waveform remain incompletely understood. Biomechanical measures of human cilia motion and their relationships to cilia assembly are needed to illuminate the biophysics of normal cilia function, and to quantify dysfunction in ciliopathies. To these ends, we analyzed cilia motion from high-speed video microscopy of ciliated cells sampled from human lung airways compared to primary-culture cells that undergo ciliogenesis in vitro. Quantitative assessment of waveform parameters showed variations in waveform shape between individual cilia; however, general trends in waveform parameters emerged, associated with progression of cilia length and stage of differentiation. When cilia emerged from cultured cells, beat frequency was initially elevated, then fell and remained stable as cilia lengthened. In contrast, the average bending amplitude and the ability to generate force gradually increased and eventually approached values observed in ex vivo samples. Dynein arm motor proteins DNAH5, DNAH9, DNAH11, and DNAH6 were localized within specific regions of the axoneme in the ex vivo cells; however distinct stages of in vitro waveform development identified by biomechanical features were associated with the progressive movement of dyneins to the appropriate proximal or distal sections of the cilium. These observations suggest that the step-wise variation in waveform development during ciliogenesis is dependent on cilia length and potentially outer dynein arm assembly.

Laparoscopic optical coherence tomographic imaging of human ovarian cancer

NASA Astrophysics Data System (ADS)

Hariri, Lida P.; Bonnema, Garret T.; Schmidt, Kathy; Hatch, Kenneth; Brewer, Molly; Barton, Jennifer K.

2008-02-01

Ovarian cancer is the fourth leading cause of cancer-related death among women in the United States. If diagnosed at an early stage, the 5-year survival rate is 94%, but drops to 68% for regional disease and 29% for distant metastasis; only 19% of all cases are diagnosed at the early, localized stage. Optical coherence tomography is a recently emerging non-destructive imaging technology, achieving high axial resolutions (10-20 microns) at imaging depths up to 2 mm. Previously, we studied OCT imaging in normal and diseased human ovary ex vivo to determine the features OCT is capable of resolving. Changes in collagen were suggested with several of the images that correlated with changes in collagen seen in malignancy. Areas of necrosis and blood vessels were also visualized using OCT, indicative of an underlying tissue abnormality. We recently developed a custom side-firing laparoscopic OCT (LOCT) probe fabricated specifically for in vivo laparoscopic imaging. The LOCT probe consists of a 38 mm diameter handpiece terminated in an 280 mm long, 4.6 mm diameter tip for insertion into the laparoscopic trocar and is capable of obtaining up to 9.5 mm image lengths at 10 micron axial resolution. In this study, we utilize the LOCT probe to image one or both ovaries of 20 patients undergoing laparotomy or transabdominal endoscopy and oophorectomy to determine if OCT is capable of identifying and/or differentiating normal and neoplastic ovary. To date, we have laparoscopically imaged the ovaries of ten patients successfully with no known complications.

Vascular and Cognitive Assessments in Patients With Breast Cancer Undergoing Chemotherapy After Surgery

ClinicalTrials.gov

2017-05-25

Cognitive/Functional Effects; Recurrent Breast Cancer; Stage IA Breast Cancer; Stage IB Breast Cancer; Stage II Breast Cancer; Stage IIIA Breast Cancer; Stage IIIB Breast Cancer; Stage IIIC Breast Cancer

Rosuvastatin in Treating Women With Cardiovascular Complications Who Are Undergoing Chemotherapy For Breast Cancer

ClinicalTrials.gov

2017-05-25

Cardiovascular Complications; Recurrent Breast Cancer; Stage I Breast Cancer; Stage II Breast Cancer; Stage IIIA Breast Cancer; Stage IIIB Breast Cancer; Stage IIIC Breast Cancer; Stage IV Breast Cancer

Impairment of organ-specific T cell negative selection by diabetes susceptibility genes: genomic analysis by mRNA profiling.

PubMed

Liston, Adrian; Hardy, Kristine; Pittelkow, Yvonne; Wilson, Susan R; Makaroff, Lydia E; Fahrer, Aude M; Goodnow, Christopher C

2007-01-01

T cells in the thymus undergo opposing positive and negative selection processes so that the only T cells entering circulation are those bearing a T cell receptor (TCR) with a low affinity for self. The mechanism differentiating negative from positive selection is poorly understood, despite the fact that inherited defects in negative selection underlie organ-specific autoimmune disease in AIRE-deficient people and the non-obese diabetic (NOD) mouse strain Here we use homogeneous populations of T cells undergoing either positive or negative selection in vivo together with genome-wide transcription profiling on microarrays to identify the gene expression differences underlying negative selection to an Aire-dependent organ-specific antigen, including the upregulation of a genomic cluster in the cytogenetic band 2F. Analysis of defective negative selection in the autoimmune-prone NOD strain demonstrates a global impairment in the induction of the negative selection response gene set, but little difference in positive selection response genes. Combining expression differences with genetic linkage data, we identify differentially expressed candidate genes, including Bim, Bnip3, Smox, Pdrg1, Id1, Pdcd1, Ly6c, Pdia3, Trim30 and Trim12. The data provide a molecular map of the negative selection response in vivo and, by analysis of deviations from this pathway in the autoimmune susceptible NOD strain, suggest that susceptibility arises from small expression differences in genes acting at multiple points in the pathway between the TCR and cell death.

Impairment of organ-specific T cell negative selection by diabetes susceptibility genes: genomic analysis by mRNA profiling

PubMed Central

Liston, Adrian; Hardy, Kristine; Pittelkow, Yvonne; Wilson, Susan R; Makaroff, Lydia E; Fahrer, Aude M; Goodnow, Christopher C

2007-01-01

Background T cells in the thymus undergo opposing positive and negative selection processes so that the only T cells entering circulation are those bearing a T cell receptor (TCR) with a low affinity for self. The mechanism differentiating negative from positive selection is poorly understood, despite the fact that inherited defects in negative selection underlie organ-specific autoimmune disease in AIRE-deficient people and the non-obese diabetic (NOD) mouse strain Results Here we use homogeneous populations of T cells undergoing either positive or negative selection in vivo together with genome-wide transcription profiling on microarrays to identify the gene expression differences underlying negative selection to an Aire-dependent organ-specific antigen, including the upregulation of a genomic cluster in the cytogenetic band 2F. Analysis of defective negative selection in the autoimmune-prone NOD strain demonstrates a global impairment in the induction of the negative selection response gene set, but little difference in positive selection response genes. Combining expression differences with genetic linkage data, we identify differentially expressed candidate genes, including Bim, Bnip3, Smox, Pdrg1, Id1, Pdcd1, Ly6c, Pdia3, Trim30 and Trim12. Conclusion The data provide a molecular map of the negative selection response in vivo and, by analysis of deviations from this pathway in the autoimmune susceptible NOD strain, suggest that susceptibility arises from small expression differences in genes acting at multiple points in the pathway between the TCR and cell death. PMID:17239257

Heavy Metal Exposure in Predicting Peripheral Neuropathy in Patients With Stage I-III Breast Cancer Undergoing Chemotherapy

ClinicalTrials.gov

2017-06-14

Male Breast Cancer; Neurotoxicity; Peripheral Neuropathy; Stage IA Breast Cancer; Stage IB Breast Cancer; Stage II Breast Cancer; Stage IIIA Breast Cancer; Stage IIIB Breast Cancer; Stage IIIC Breast Cancer

Comparative proteomics of two life cycle stages of stable isotope-labeled Trypanosoma brucei reveals novel components of the parasite's host adaptation machinery.

PubMed

Butter, Falk; Bucerius, Ferdinand; Michel, Margaux; Cicova, Zdenka; Mann, Matthias; Janzen, Christian J

2013-01-01

Trypanosoma brucei developed a sophisticated life cycle to adapt to different host environments. Although developmental differentiation of T. brucei has been the topic of intensive research for decades, the mechanisms responsible for adaptation to different host environments are not well understood. We developed stable isotope labeling by amino acids in cell culture in trypanosomes to compare the proteomes of two different life cycle stages. Quantitative comparison of 4364 protein groups identified many proteins previously not known to be stage-specifically expressed. The identification of stage-specific proteins helps to understand how parasites adapt to different hosts and provides new insights into differences in metabolism, gene regulation, and cell architecture. A DEAD-box RNA helicase, which is highly up-regulated in the bloodstream form of this parasite and which is essential for viability and proper cell cycle progression in this stage is described as an example.

A novel gene, RSD-3/HSD-3.1, encodes a meiotic-related protein expressed in rat and human testis.

PubMed

Zhang, Xiaodong; Liu, Huixian; Zhang, Yan; Qiao, Yuan; Miao, Shiying; Wang, Linfang; Zhang, Jianchao; Zong, Shudong; Koide, S S

2003-06-01

The expression of stage-specific genes during spermatogenesis was determined by isolating two segments of rat seminiferous tubule at different stages of the germinal epithelium cycle delineated by transillumination-delineated microdissection, combined with differential display polymerase chain reaction to identify the differential transcripts formed. A total of 22 cDNAs were identified and accepted by GenBank as new expressed sequence tags. One of the expressed sequence tags was radiolabeled and used as a probe to screen a rat testis cDNA library. A novel full-length cDNA composed of 2228 bp, designated as RSD-3 (rat sperm DNA no.3, GenBank accession no. AF094609) was isolated and characterized. The reading frame encodes a polypeptide consisting of 526 amino acid residues, containing a number of DNA binding motifs and phosphorylation sites for PKC, CK-II, and p34cdc2. Northern blot of mRNA prepared from various tissues of adult rats showed that RSD-3 is expressed only in the testis. The initial expression of the RSD-3 gene was detected in the testis on the 30th postnatal day and attained adult level on the 60th postnatal day. Immunolocalization of RSD-3 in germ cells of rat testis showed that its expression is restricted to primary spermatocytes, undergoing meiosis division I. A human testis homologue of RSD-3 cDNA, designated as HSD-3.1 (GenBank accession no. AF144487) was isolated by screening the Human Testis Rapid-Screen arrayed cDNA library panels by RT-PCR. The exon-intron boundaries of HSD-3.1 gene were determined by aligning the cDNA sequence with the corresponding genome sequence. The cDNA consisted of 12 exons that span approximately 52.8 kb of the genome sequence and was mapped to chromosome 14q31.3.

Salt-sensitivity of σ(H) and Spo0A prevents sporulation of Bacillus subtilis at high osmolarity avoiding death during cellular differentiation.

PubMed

Widderich, Nils; Rodrigues, Christopher D A; Commichau, Fabian M; Fischer, Kathleen E; Ramirez-Guadiana, Fernando H; Rudner, David Z; Bremer, Erhard

2016-04-01

The spore-forming bacterium Bacillus subtilis frequently experiences high osmolarity as a result of desiccation in the soil. The formation of a highly desiccation-resistant endospore might serve as a logical osmostress escape route when vegetative growth is no longer possible. However, sporulation efficiency drastically decreases concomitant with an increase in the external salinity. Fluorescence microscopy of sporulation-specific promoter fusions to gfp revealed that high salinity blocks entry into the sporulation pathway at a very early stage. Specifically, we show that both Spo0A- and SigH-dependent transcription are impaired. Furthermore, we demonstrate that the association of SigH with core RNA polymerase is reduced under these conditions. Suppressors that modestly increase sporulation efficiency at high salinity map to the coding region of sigH and in the regulatory region of kinA, encoding one the sensor kinases that activates Spo0A. These findings led us to discover that B. subtilis cells that overproduce KinA can bypass the salt-imposed block in sporulation. Importantly, these cells are impaired in the morphological process of engulfment and late forespore gene expression and frequently undergo lysis. Altogether our data indicate that B. subtilis blocks entry into sporulation in high-salinity environments preventing commitment to a developmental program that it cannot complete. © 2015 John Wiley & Sons Ltd.

Salt-sensitivity of σH and Spo0A prevents sporulation of Bacillus subtilis at high osmolarity avoiding death during cellular differentiation

PubMed Central

Widderich, Nils; Rodrigues, Christopher D.A.; Commichau, Fabian M.; Fischer, Kathleen E.; Ramirez-Guadiana, Fernando H.; Rudner, David Z.; Bremer, Erhard

2016-01-01

Summary The spore-forming bacterium Bacillus subtilis frequently experiences high osmolarity as a result of desiccation in the soil. The formation of a highly desiccation-resistant endospore might serve as a logical osmostress escape route when vegetative growth is no longer possible. However, sporulation efficiency drastically decreases concomitant with an increase in the external salinity. Fluorescence microscopy of sporulation-specific promoter fusions to gfp revealed that high salinity blocks entry into the sporulation pathway at a very early stage. Specifically, we show that both Spo0A- and SigH-dependent transcription are impaired. Furthermore, we demonstrate that the association of SigH with core RNA polymerase is reduced under these conditions. Suppressors that modestly increase sporulation efficiency at high salinity map to the coding region of sigH and in the regulatory region of kinA, encoding one the sensor kinases that activates Spo0A. These findings led us to discover that B. subtilis cells that overproduce KinA can bypass the salt-imposed block in sporulation. Importantly, these cells are impaired in the morphological process of engulfment and late forespore gene expression and frequently undergo lysis. Altogether our data indicate that B. subtilis blocks entry into sporulation in high-salinity environments preventing commitment to a developmental program that it cannot complete. PMID:26712348

Expression of Ulex europaeus agglutinin I lectin-binding sites in squamous cell carcinomas and their absence in basal cell carcinomas. Indicator of tumor type and differentiation.

PubMed

Heng, M C; Fallon-Friedlander, S; Bennett, R

1992-06-01

Lectins bind tightly to carbohydrate moieties on cell surfaces. Alterations in lectin binding have been reported to accompany epidermal cell differentiation, marking alterations in membrane sugars during this process. The presence of UEA I (Ulex europaeus agglutinin I) L-fucose-specific lectin-binding sites has been used as a marker for terminally differentiated (committed) keratinocytes. In this article, we report the presence of UEA-I-binding sites on squamous keratinocytes of well-differentiated squamous cell carcinomas, with patchy loss of UEA I positivity on poorly differentiated cells of squamous cell carcinomas, suggesting a possible use for this technique in the rapid assessment of less differentiated areas within the squamous cell tumor. The absence of UEA-I-binding sites on basal cell carcinomas may be related to an inability of cells comprising this tumor to convert the L-D-pyranosyl moiety on basal cells to the L-fucose moiety, resulting in an inability of basal cell carcinoma cell to undergo terminal differentiation into a committed keratinocyte.

Lenvatinib and Pembrolizumab in DTC

ClinicalTrials.gov

2018-05-21

Columnar Cell Variant Thyroid Gland Papillary Carcinoma; Follicular Variant Thyroid Gland Papillary Carcinoma; Poorly Differentiated Thyroid Gland Carcinoma; Recurrent Thyroid Gland Carcinoma; Stage III Differentiated Thyroid Gland Carcinoma AJCC v7; Stage III Thyroid Gland Follicular Carcinoma AJCC v7; Stage III Thyroid Gland Papillary Carcinoma AJCC v7; Stage IV Thyroid Gland Follicular Carcinoma AJCC v7; Stage IV Thyroid Gland Papillary Carcinoma AJCC v7; Stage IVA Differentiated Thyroid Gland Carcinoma AJCC v7; Stage IVA Thyroid Gland Follicular Carcinoma AJCC v7; Stage IVA Thyroid Gland Papillary Carcinoma AJCC v7; Stage IVB Differentiated Thyroid Gland Carcinoma AJCC v7; Stage IVB Thyroid Gland Follicular Carcinoma AJCC v7; Stage IVB Thyroid Gland Papillary Carcinoma AJCC v7; Stage IVC Differentiated Thyroid Gland Carcinoma AJCC v7; Stage IVC Thyroid Gland Follicular Carcinoma AJCC v7; Stage IVC Thyroid Gland Papillary Carcinoma AJCC v7; Tall Cell Variant Thyroid Gland Papillary Carcinoma; Thyroid Gland Oncocytic Follicular Carcinoma

Differentiation of Spermatogonia Stem Cells into Functional Mature Neurons Characterized with Differential Gene Expression.

PubMed

Bojnordi, Maryam Nazm; Azizi, Hossein; Skutella, Thomas; Movahedin, Mansoureh; Pourabdolhossein, Fereshteh; Shojaei, Amir; Hamidabadi, Hatef Ghasemi

2017-09-01

Transplantation of embryonic stem cells (ESCs) is a promising therapeutic approach for the treatment of neurodegenerative diseases. However, ESCs are not usable clinically due to immunological and ethical limitations. The identification of an alternative safe cell source opens novel options via autologous transplantation in neuro-regeneration circumventing these problems. Here, we examined the neurogenic capacity of embryonic stem-like cells (ES-like cells) derived from the testis using neural growth factor inducers and utilized them to generate functional mature neurons. The neuronal differentiation of ES-like cells is induced in three stages. Stage 1 is related to embryoid body (EB) formation. To induce neuroprogenitor cells, EBs were cultured in the presence of retinoic acid, N 2 supplement and fibroblast growth factor followed by culturing in a neurobasal medium containing B 27 , N 2 supplements for additional 10 days, to allow the maturation and development of neuronal progenitor cells. The neurogenic differentiation was confirmed by immunostaining for markers of mature neurons. The differentiated neurons were positive for Tuj1 and Tau1. Real-time PCR dates indicated the expression of Nestin and Neuro D (neuroprogenitor markers) in induced cells at the second stage of the differentiation protocol. The differentiated mature neurons exhibited the specific neuron markers Map2 and β-tubulin. The functional maturity of neurons was confirmed by an electrophysiological analysis of passive and active neural membrane properties. These findings indicated a differentiation capacity of ES-like cells derived from the testis to functionally mature neurons, which proposes them as a novel cell source for neuroregenerative medicine.

Development of an occupational advice intervention for patients undergoing lower limb arthroplasty (the OPAL study).

PubMed

Baker, Paul; Coole, Carol; Drummond, Avril; McDaid, Catriona; Khan, Sayeed; Thomson, Louise; Hewitt, Catherine; McNamara, Iain; McDonald, David; Fitch, Judith; Rangan, Amar

2018-06-28

There are an increasing number of patients of working age undergoing hip and knee replacements. Currently there is variation in the advice and support given about sickness absence, recovery to usual activities and return to work after these procedures. Earlier, sustainable, return to work improves the health of patients and benefits their employers and society. An intervention that encourages and supports early recovery to usual activities, including work, has the potential to reduce the health and socioeconomic burden of hip and knee replacements. A two-phase research programme delivered over 27 months will be used to develop and subsequently test the feasibility of an occupational advice intervention to facilitate return to work and usual activities in patients undergoing lower limb arthroplasty. The 2 phases will incorporate a six-stage intervention mapping process: Phase 1: Intervention mapping stages 1-3: 1 Needs assessment (including rapid evidence synthesis, prospective cohort analysis and structured stakeholder interviews) 2 Identification of intended outcomes and performance objectives 3 Selection of theory-based methods and practical strategies Phase 2: Intervention mapping stages 4-6: 4 Development of components and materials for the occupational advice intervention using a modified Delphi process 5 Adoption and implementation of the intervention 6 Evaluation and feasibility testing The study will be undertaken in four National Health Service (NHS) hospitals in the United Kingdom and two Higher Education Institutions. OPAL (Occupational advice for Patients undergoing Arthroplasty of the Lower limb) aims to develop an occupational advice intervention to support early recovery to usual activities including work, which is tailored to the requirements of patients undergoing hip and knee replacements. The developed intervention will then be assessed with a specific focus on evaluating its feasibility as a potential trial intervention to improve speed of recovery to usual activities including work. The study was registered retrospectively with the International Standard Randomised Controlled Trials Number (ISRCTN): 27426982 (Date 20/12/2016) and the International prospective register of systematic reviews (PROSPERO): CRD42016045235 (Date 04/08/2016).

Two-Stage, In Silico Deconvolution of the Lymphocyte Compartment of the Peripheral Whole Blood Transcriptome in the Context of Acute Kidney Allograft Rejection

PubMed Central

Shannon, Casey P.; Balshaw, Robert; Ng, Raymond T.; Wilson-McManus, Janet E.; Keown, Paul; McMaster, Robert; McManus, Bruce M.; Landsberg, David; Isbel, Nicole M.; Knoll, Greg; Tebbutt, Scott J.

2014-01-01

Acute rejection is a major complication of solid organ transplantation that prevents the long-term assimilation of the allograft. Various populations of lymphocytes are principal mediators of this process, infiltrating graft tissues and driving cell-mediated cytotoxicity. Understanding the lymphocyte-specific biology associated with rejection is therefore critical. Measuring genome-wide changes in transcript abundance in peripheral whole blood cells can deliver a comprehensive view of the status of the immune system. The heterogeneous nature of the tissue significantly affects the sensitivity and interpretability of traditional analyses, however. Experimental separation of cell types is an obvious solution, but is often impractical and, more worrying, may affect expression, leading to spurious results. Statistical deconvolution of the cell type-specific signal is an attractive alternative, but existing approaches still present some challenges, particularly in a clinical research setting. Obtaining time-matched sample composition to biologically interesting, phenotypically homogeneous cell sub-populations is costly and adds significant complexity to study design. We used a two-stage, in silico deconvolution approach that first predicts sample composition to biologically meaningful and homogeneous leukocyte sub-populations, and then performs cell type-specific differential expression analysis in these same sub-populations, from peripheral whole blood expression data. We applied this approach to a peripheral whole blood expression study of kidney allograft rejection. The patterns of differential composition uncovered are consistent with previous studies carried out using flow cytometry and provide a relevant biological context when interpreting cell type-specific differential expression results. We identified cell type-specific differential expression in a variety of leukocyte sub-populations at the time of rejection. The tissue-specificity of these differentially expressed probe-set lists is consistent with the originating tissue and their functional enrichment consistent with allograft rejection. Finally, we demonstrate that the strategy described here can be used to derive useful hypotheses by validating a cell type-specific ratio in an independent cohort using the nanoString nCounter assay. PMID:24733377

Adolescent Experiences with the Vaginal Ring

PubMed Central

Epstein, Laura B.; Sokal-Gutierrez, Karen; Ivey, Susan L.; Raine, Tina; Auerswald, Colette

2011-01-01

Purpose To understand racial/ethnic minority adolescent females’ experiences with the vaginal ring. Methods We conducted in-depth interviews with a clinic-based sample of 32 young women aged 15–24 years who had used the vaginal ring. Results Qualitative analysis using grounded theory revealed that adolescents undergo a multi-stage process when trying the ring and adopting ring use. These stages include hearing about the ring, initial reactions, first experiences with insertion and removal, and first sexual experiences. Adolescents subsequently enter an assessment and adjustment stage in which they decide whether to adopt or discontinue ring use. Ultimately they share their experiences with friends. Conclusions The model developed provides a context within which providers may advise adolescents as they begin use of the ring. Some specific recommendations are offered. PMID:18565439

Switching of the core structures of glycosphingolipids from globo- and lacto- to ganglio-series upon human embryonic stem cell differentiation.

PubMed

Liang, Yuh-Jin; Kuo, Huan-Hsien; Lin, Chi-Hung; Chen, Yen-Ying; Yang, Bei-Chia; Cheng, Yuan-Yuan; Yu, Alice L; Khoo, Kay-Hooi; Yu, John

2010-12-28

A systematic survey of expression profiles of glycosphingolipids (GSLs) in two hESC lines and their differentiated embryoid body (EB) outgrowth with three germ layers was carried out using immunofluorescence, flow cytometry, and MALDI-MS and MS/MS analyses. In addition to the well-known hESC-specific markers stage-specific embryonic antigen 3 (SSEA-3) and SSEA-4, we identified several globosides and lacto-series GSLs, previously unrevealed in hESCs, including Gb(4)Cer, Lc(4)Cer, fucosyl Lc(4)Cer, Globo H, and disialyl Gb(5)Cer. During hESC differentiation into EBs, MS analysis revealed a clear-cut switch in the core structures of GSLs from globo- and lacto- to ganglio-series, which was not as evident by immunostaining with antibodies against SSEA-3 and SSEA-4, owing to their cross-reactivities with various glycosphingolipids. Such a switch was attributable to altered expression of key glycosyltransferases (GTs) in the biosynthetic pathways by the up-regulation of ganglio-series-related GTs with simultaneous down-regulation of globo- and lacto-series-related GTs. Thus, these results provide insights into the unique stage-specific transition and mechanism for alterations of GSL core structures during hESC differentiation. In addition, unique glycan structures uncovered by MS analyses may serve as surface markers for further delineation of hESCs and help identify of their functional roles not only in hESCs but also in cancers.

Donor‐Dependent and Other Nondefined Factors Have Greater Influence on the Hepatic Phenotype Than the Starting Cell Type in Induced Pluripotent Stem Cell Derived Hepatocyte‐Like Cells

PubMed Central

Heslop, James A.; Kia, Richard; Pridgeon, Christopher S.; Sison‐Young, Rowena L.; Liloglou, Triantafillos; Elmasry, Mohamed; Fenwick, Stephen W.; Mills, John S.; Kitteringham, Neil R.; Park, Bong K.

2017-01-01

Abstract Drug‐induced liver injury is the greatest cause of post‐marketing drug withdrawal; therefore, substantial resources are directed toward triaging potentially dangerous new compounds at all stages of drug development. One of the major factors preventing effective screening of new compounds is the lack of a predictive in vitro model of hepatotoxicity. Primary human hepatocytes offer a metabolically relevant model for which the molecular initiating events of hepatotoxicity can be examined; however, these cells vary greatly between donors and dedifferentiate rapidly in culture. Induced pluripotent stem cell (iPSC)‐derived hepatocyte‐like cells (HLCs) offer a reproducible, physiologically relevant and genotypically normal model cell; however, current differentiation protocols produce HLCs with a relatively immature phenotype. During the reprogramming of somatic cells, the epigenome undergoes dramatic changes; however, this “resetting” is a gradual process, resulting in an altered differentiation propensity, skewed toward the lineage of origin, particularly in early passage cultures. We, therefore, performed a comparison of human hepatocyte‐ and dermal fibroblast‐derived iPSCs, assessing the impact of epigenetic memory at all stages of HLC differentiation. These results provide the first isogenic assessment of the starting cell type in human iPSC‐derived HLCs. Despite a trend toward improvement in hepatic phenotype in albumin secretion and gene expression, few significant differences in hepatic differentiation capacity were found between hepatocyte and fibroblast‐derived iPSCs. We conclude that the donor and inter‐clonal differences have a greater influence on the hepatocyte phenotypic maturity than the starting cell type. Therefore, it is not necessary to use human hepatocytes for generating iPSC‐derived HLCs. Stem Cells Translational Medicine 2017;6:1321–1331 PMID:28456008

Endocrine disrupting chemicals affect the adipogenic differentiation of mesenchymal stem cells in distinct ontogenetic windows

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

Biemann, Ronald, E-mail: ronald.biemann@medizin.uni-halle.de; Navarrete Santos, Anne; Navarrete Santos, Alexander

Highlights: Black-Right-Pointing-Pointer Endocrine disrupting chemicals affect adipogenesis in mesenchymal stem cells (MSC). Black-Right-Pointing-Pointer The adipogenic impact depends strongly on the window of exposure. Black-Right-Pointing-Pointer Bisphenol A reduces the potential of MSC to differentiate into adipocytes. Black-Right-Pointing-Pointer DEHP and TBT trigger the adipogenic differentiation of mesenchymal stem cells. Black-Right-Pointing-Pointer BPA, DEHP and TBT did not affect adipogenesis in embryonic stem cells. -- Abstract: Endocrine disrupting chemicals (EDC) like bisphenol A (BPA), bis(2-ethylhexyl)phthalate (DEHP) and tributyltin (TBT) are ubiquitously present in the environment and in human tissues. They bind to nuclear hormone receptors and affect cellular and developmental processes. In this study,more » we show that BPA, DEHP and TBT affect the adipogenic differentiation of murine mesenchymal stem cells (MSC, C3H/10T1/2) in a concentration-, stage- and compound-specific manner. C3H/10T1/2 cells and embryonic stem cells (CGR8) were exposed to BPA, DEHP or TBT at different stages of cell determination and differentiation (undifferentiated growth, adipogenic induction and terminal adipogenic differentiation). The final amount of differentiated adipocytes, cellular triglyceride content and mRNA expression of adipogenic marker genes (adiponectin, FABP4, PPAR{gamma}2, LPL) were quantified and compared with corresponding unexposed cells. BPA (10 {mu}M) decreased subsequent adipogenic differentiation of MSC, when cells were exposed during undifferentiated growth. In contrast, DEHP (100 {mu}M) during the hormonal induction period, and TBT (100 nM) in all investigated stages, enhanced adipogenesis. Importantly, exposure of undifferentiated murine embryonic stem cells did not show any effect of the investigated EDC on subsequent adipogenic differentiation.« less

Relationship Between Clinicopathological Characteristics and PET/CT Uptake in Esophageal Squamous Cell Carcinoma: [18F]Alfatide versus [18F]FDG.

PubMed

Dong, Yinjun; Wei, Yuchun; Chen, Guanxuan; Huang, Yong; Song, Pingping; Liu, Shuguang; Zheng, Jinsong; Cheng, Monica; Yuan, Shuanghu

2018-06-04

To assess a novel radiotracer aluminum [ 18 F]fluoride-1,4,7-triazacyclononane-triacetic acid-pegylated dimeric RGD ([ 18 F]ALF-NOTA-PRGD 2 , denoted as [ 18 F]Alfatide) for positron emission tomography (PET)/X-ray computed tomography (CT) and explore the relationships between clinicopathological characteristics and maximum standard uptake values in primary (SUV P ) and metastatic lymph nodes (SUV LN ) of patients with esophageal squamous cell carcinoma (ESCC), as verified by pathologic examination and compared with those obtained with 2-deoxy-2-[ 18 F]fluoro-D-glucose ([ 18 F]DG) PET. We prospectively enrolled patients with newly diagnosed ESCC who agreed to undergo [ 18 F]Alfatide PET/CT or [ 18 F]FDG PET/CT scans before surgery at Shandong Cancer Hospital from May 2011 to July 2017. SUVs and the pathological tumor-node-metastasis (pTNM) stages of primary tumors and metastatic lymph nodes (LNs) were measured and confirmed pathologically. Immunohistochemical (IHC) staining for integrin αvβ3 was performed on tumor samples (both primary tumors and metastatic LNs) collected from nine patients. Of 61 patients who underwent PET/CT scans, 46 then underwent curative surgery and were included in our analysis (n = 21 for [ 18 F]Alfatide PET/CT and n = 25 for [ 18 F]FDG PET/CT). No significant differences in the SUV P on [ 18 F]Alfatide PET/CT or [ 18 F]FDG PET/CT were observed among the cohorts according to gender, pathological stage, T stage, status of LNs, and differentiation (all P > 0.05). The SUV LN differed significantly between the pathological stages and status of LNs both on [ 18 F]Alfatide PET/CT (P = 0.03, 0.003) and [ 18 F]FDG PET/CT (P = 0.001. < 0.001), but not according to gender (P = 0.128, 0.129), T stage (P = 0.791, 0.727), or tumor differentiation (P = 0.049, 0.053). Significant positive correlations were observed between the SUV LN on [ 18 F]Alfatide PET/CT and [ 18 F]FDG PET/CT, and pathological stage (r = 0.52, P = 0.016; r = 0.503, P = 0.01), LN status (r = 0.73, P < 0.001; r = 0.649, P < 0.001), and differentiation (r = 0.509, P < 0.019; r = 0.459, P = 0.021) were observed. No significant differences were found between the relationships of SUV P with SUV LN , length, age, gender, pathological stage, T stage, status of LN, or differentiation, or of SUV LN with length, age, gender, or T stage both on [ 18 F]Alfatide PET/CT and [ 18 F]FDG PET/CT (all P > 0.05). The quantitated expression levels of αvβ3 in primary tumors and metastatic LNs were 1.67 ± 1.12 and 3.42 ± 2.93, respectively (P = 0.031). Our results suggest that SUV LN is influenced by pathological stage, LN status, and differentiation. SUV LN may therefore serve as a new parameter for risk stratification of with ESCC patients. Moreover, [ 18 F]Alfatide PET can provide complementary molecular information about ESCC metastasis.

HoxBlinc RNA Recruits Set1/MLL Complexes to Activate Hox Gene Expression Patterns and Mesoderm Lineage Development.

PubMed

Deng, Changwang; Li, Ying; Zhou, Lei; Cho, Joonseok; Patel, Bhavita; Terada, Naohiro; Li, Yangqiu; Bungert, Jörg; Qiu, Yi; Huang, Suming

2016-01-05

Trithorax proteins and long-intergenic noncoding RNAs are critical regulators of embryonic stem cell pluripotency; however, how they cooperatively regulate germ layer mesoderm specification remains elusive. We report here that HoxBlinc RNA first specifies Flk1(+) mesoderm and then promotes hematopoietic differentiation through regulation of hoxb pathways. HoxBlinc binds to the hoxb genes, recruits Setd1a/MLL1 complexes, and mediates long-range chromatin interactions to activate transcription of the hoxb genes. Depletion of HoxBlinc by shRNA-mediated knockdown or CRISPR-Cas9-mediated genetic deletion inhibits expression of hoxb genes and other factors regulating cardiac/hematopoietic differentiation. Reduced hoxb expression is accompanied by decreased recruitment of Set1/MLL1 and H3K4me3 modification, as well as by reduced chromatin loop formation. Re-expression of hoxb2-b4 genes in HoxBlinc-depleted embryoid bodies rescues Flk1(+) precursors that undergo hematopoietic differentiation. Thus, HoxBlinc plays an important role in controlling hoxb transcription networks that mediate specification of mesoderm-derived Flk1(+) precursors and differentiation of Flk1(+) cells into hematopoietic lineages. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

HoxBlinc RNA recruits Set1/MLL complexes to activate Hox gene expression patterns and mesoderm lineage development

PubMed Central

Deng, Changwang; Li, Ying; Zhou, Lei; Cho, Joonseok; Patel, Bhavita; Terada, Nao; Li, Yangqiu; Bungert, Jörg; Qiu, Yi; Huang, Suming

2015-01-01

Summary Trithorax proteins and long-intergenic noncoding RNAs are critical regulators of embryonic stem cell pluripotency; however, how they cooperatively regulate germ layer mesoderm specification remains elusive. We report here that HoxBlinc RNA first specifies Flk1+ mesoderm and then promotes hematopoietic differentiation through regulating hoxb gene pathways. HoxBlinc binds to the hoxb genes, recruits Setd1a/MLL1 complexes, and mediates long-range chromatin interactions to activate transcription of the hoxb genes. Depletion of HoxBlinc by shRNA-mediated KD or CRISPR-Cas9-mediated genetic deletion inhibits expression of hoxb genes and other factors regulating cardiac/hematopoietic differentiation. Reduced hoxb gene expression is accompanied by decreased recruitment of Set1/MLL1 and H3K4me3 modification, as well as by reduced chromatin loop formation. Re-expression of hoxb2-b4 genes in HoxBlinc-depleted embryoid bodies rescues Flk1+ precursors that undergo hematopoietic differentiation. Thus, HoxBlinc plays an important role in controlling hoxb transcription networks that mediate specification of mesoderm-derived Flk1+ precursors and differentiation of Flk1+ cells into hematopoietic lineages. PMID:26725110

Lysophosphatidylcholine Regulates Sexual Stage Differentiation in the Human Malaria Parasite Plasmodium falciparum.

PubMed

Brancucci, Nicolas M B; Gerdt, Joseph P; Wang, ChengQi; De Niz, Mariana; Philip, Nisha; Adapa, Swamy R; Zhang, Min; Hitz, Eva; Niederwieser, Igor; Boltryk, Sylwia D; Laffitte, Marie-Claude; Clark, Martha A; Grüring, Christof; Ravel, Deepali; Blancke Soares, Alexandra; Demas, Allison; Bopp, Selina; Rubio-Ruiz, Belén; Conejo-Garcia, Ana; Wirth, Dyann F; Gendaszewska-Darmach, Edyta; Duraisingh, Manoj T; Adams, John H; Voss, Till S; Waters, Andrew P; Jiang, Rays H Y; Clardy, Jon; Marti, Matthias

2017-12-14

Transmission represents a population bottleneck in the Plasmodium life cycle and a key intervention target of ongoing efforts to eradicate malaria. Sexual differentiation is essential for this process, as only sexual parasites, called gametocytes, are infective to the mosquito vector. Gametocyte production rates vary depending on environmental conditions, but external stimuli remain obscure. Here, we show that the host-derived lipid lysophosphatidylcholine (LysoPC) controls P. falciparum cell fate by repressing parasite sexual differentiation. We demonstrate that exogenous LysoPC drives biosynthesis of the essential membrane component phosphatidylcholine. LysoPC restriction induces a compensatory response, linking parasite metabolism to the activation of sexual-stage-specific transcription and gametocyte formation. Our results reveal that malaria parasites can sense and process host-derived physiological signals to regulate differentiation. These data close a critical knowledge gap in parasite biology and introduce a major component of the sexual differentiation pathway in Plasmodium that may provide new approaches for blocking malaria transmission. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

Dynamic and differential expression of the gonadal aromatase during the process of sexual differentiation in a novel transgenic cyp19a1a-eGFP zebrafish line.

PubMed

Hinfray, Nathalie; Sohm, Frédéric; Caulier, Morgane; Chadili, Edith; Piccini, Benjamin; Torchy, Camille; Porcher, Jean-Marc; Guiguen, Yann; Brion, François

2018-05-15

In zebrafish, there exists a clear need for new tools to study sex differentiation dynamic and its perturbation by endocrine disrupting chemicals. In this context, we developed and characterized a novel transgenic zebrafish line expressing green fluorescent protein (GFP) under the control of the zebrafish cyp19a1a (gonadal aromatase) promoter. In most gonochoristic fish species including zebrafish, cyp19a1a, the enzyme responsible for the synthesis of estrogens, has been shown to play a critical role in the processes of reproduction and sexual differentiation. This novel cyp19a1a-eGFP transgenic line allowed a deeper characterization of expression and localization of cyp19a1a gene in zebrafish gonads both at the adult stage and during development. At the adult stage, GFP expression was higher in ovaries than in testis. We showed a perfect co-expression of GFP and endogenous Cyp19a1a protein in gonads that was mainly localized in the cytoplasm of peri-follicular cells in the ovary and of Leydig and germ cells in the testis. During development, GFP was expressed in all immature gonads of 20 dpf-old zebrafish. Then, GFP expression increased in early differentiated female at 30 and 35dpf to reach a high GFP intensity in well-differentiated ovaries at 40dpf. On the contrary, males consistently displayed low GFP expression as compared to female whatever their stage of development, resulting in a clear dimorphic expression between both sexes. Interestingly, fish that undergoes ovary-to-testis transition (35 and 40dpf) presented GFP levels similar to males or intermediate between females and males. In this transgenic line our results confirm that cyp19a1a is expressed early during development, before the histological differentiation of the gonads, and that the down-regulation of cyp19a1a expression is likely responsible for the testicular differentiation. Moreover, we show that although cyp19a1a expression exhibits a clear dimorphic expression pattern in gonads during sexual differentiation, its expression persists whatever the sex suggesting that estradiol synthesis is important for gonadal development of both sexes. Monitoring the expression of GFP in control and exposed-fish will help determine the sensitivity of this transgenic line to EDCs and to refine mechanistic based-assays for the study of EDCs. In fine, this transgenic zebrafish line will be a useful tool to study physiological processes such as reproduction and sexual differentiation, and their perturbations by EDCs. Copyright © 2017 Elsevier Inc. All rights reserved.

Pazopanib Hydrochloride in Treating Patients With Advanced Thyroid Cancer

ClinicalTrials.gov

2018-05-08

Recurrent Thyroid Gland Carcinoma; Stage III Differentiated Thyroid Gland Carcinoma AJCC v7; Stage III Thyroid Gland Medullary Carcinoma AJCC v7; Stage IVA Differentiated Thyroid Gland Carcinoma AJCC v7; Stage IVA Thyroid Gland Medullary Carcinoma AJCC v7; Stage IVA Thyroid Gland Undifferentiated (Anaplastic) Carcinoma AJCC v7; Stage IVB Differentiated Thyroid Gland Carcinoma AJCC v7; Stage IVB Thyroid Gland Medullary Carcinoma AJCC v7; Stage IVB Thyroid Gland Undifferentiated (Anaplastic) Carcinoma AJCC v7; Stage IVC Differentiated Thyroid Gland Carcinoma AJCC v7; Stage IVC Thyroid Gland Medullary Carcinoma AJCC v7; Stage IVC Thyroid Gland Undifferentiated (Anaplastic) Carcinoma AJCC v7; Thyroglobulin Antibody Negative; Thyroid Gland Undifferentiated (Anaplastic) Carcinoma

Unicuspid and bicuspid tooth crown formation in squamates.

PubMed

Handrigan, Gregory R; Richman, Joy M

2011-12-15

The molecular and developmental factors that regulate tooth morphogenesis in nonmammalian species, such as snakes and lizards, have received relatively little attention compared to mammals. Here we describe the development of unicuspid and bicuspid teeth in squamate species. The simple, cone-shaped tooth crown of the bearded dragon and ball python is established at cap stage and fixed in shape by the differentiation of cells and the secretion of dental matrices. Enamel production, as demonstrated by amelogenin expression, occurs relatively earlier in squamate teeth than in mouse molars. We suggest that the early differentiation in squamate unicuspid teeth at cap stage correlates with a more rudimentary tooth crown shape. The leopard gecko can form a bicuspid tooth crown despite the early onset of differentiation. Cusp formation in the gecko does not occur by the folding of the inner enamel epithelium, as in the mouse molar, but by the differential secretion of enamel. Ameloblasts forming the enamel epithelial bulge, a central swelling of cells in the inner enamel epithelium, secrete amelogenin at cap stage, but cease to do so by bell stage. Meanwhile, other ameloblasts in the inner enamel epithelium continue to secrete enamel, forming cusp tips on either side of the bulge. Bulge cells specifically express the gene Bmp2, which we suggest serves as a pro-differentiation signal for cells of the gecko enamel organ. In this regard, the enamel epithelial bulge of the gecko may be more functionally analogous to the secondary enamel knot of mammals than the primary enamel knot. Copyright © 2011 Wiley Periodicals, Inc., A Wiley Company.

Stage-Specific Human Induced Pluripotent Stem Cells Map the Progression of Myeloid Transformation to Transplantable Leukemia.

PubMed

Kotini, Andriana G; Chang, Chan-Jung; Chow, Arthur; Yuan, Han; Ho, Tzu-Chieh; Wang, Tiansu; Vora, Shailee; Solovyov, Alexander; Husser, Chrystel; Olszewska, Malgorzata; Teruya-Feldstein, Julie; Perumal, Deepak; Klimek, Virginia M; Spyridonidis, Alexandros; Rampal, Raajit K; Silverman, Lewis; Reddy, E Premkumar; Papaemmanuil, Elli; Parekh, Samir; Greenbaum, Benjamin D; Leslie, Christina S; Kharas, Michael G; Papapetrou, Eirini P

2017-03-02

Myeloid malignancy is increasingly viewed as a disease spectrum, comprising hematopoietic disorders that extend across a phenotypic continuum ranging from clonal hematopoiesis to myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML). In this study, we derived a collection of induced pluripotent stem cell (iPSC) lines capturing a range of disease stages encompassing preleukemia, low-risk MDS, high-risk MDS, and secondary AML. Upon their differentiation, we found hematopoietic phenotypes of graded severity and/or stage specificity that together delineate a phenotypic roadmap of disease progression culminating in serially transplantable leukemia. We also show that disease stage transitions, both reversal and progression, can be modeled in this system using genetic correction or introduction of mutations via CRISPR/Cas9 and that this iPSC-based approach can be used to uncover disease-stage-specific responses to drugs. Our study therefore provides insight into the cellular events demarcating the initiation and progression of myeloid transformation and a new platform for testing genetic and pharmacological interventions. Copyright © 2017 Elsevier Inc. All rights reserved.

Interactive Gentle Yoga in Improving Quality of Life in Patients With Stage I-III Breast Cancer Undergoing Radiation Therapy

ClinicalTrials.gov

2017-07-28

Anxiety Disorder; Depression; Ductal Breast Carcinoma in Situ; Fatigue; Stage IA Breast Cancer; Stage IB Breast Cancer; Stage II Breast Cancer; Stage IIIA Breast Cancer; Stage IIIB Breast Cancer; Stage IIIC Breast Cancer

Chitosan-coated amyloid fibrils increase adipogenesis of mesenchymal stem cells.

PubMed

Gilbert, Jay; Reynolds, Nicholas P; Russell, Sarah M; Haylock, David; McArthur, Sally; Charnley, Mirren; Jones, Owen G

2017-10-01

Mesenchymal stem cells (MSCs) have the potential to revolutionize medicine due to their ability to differentiate into specific lineages for targeted tissue repair. Development of materials and cell culture platforms that improve differentiation of either autologous or allogenic stem cell sources into specific lineages would enhance clinical utilization of MCSs. In this study, nanoscale amyloid fibrils were evaluated as substrate materials to encourage viability, proliferation, multipotency, and differentiation of MSCs. Fibrils assembled from the proteins lysozyme or β-lactoglobulin, with and without chitosan coatings, were deposited on planar mica surfaces. MSCs were cultured and differentiated on fibril-covered surfaces, as well as on unstructured controls and tissue culture plastic. Expression of CD44 and CD90 proteins indicated that multipotency was maintained for all fibrils, and osteogenic differentiation was similarly comparable among all tested materials. MSCs grown for 7days on fibril-covered surfaces favored multicellular spheroid formation and demonstrated a >75% increase in adipogenesis compared to tissue culture plastic controls, although this benefit could only be achieved if MSCs were transferred to TCP for the final differentiation step. The largest spheroids and greatest tendency to undergo adipogenesis was evidenced among MSCs grown on fibrils coated with the positively-charged polysaccharide chitosan, suggesting that spheroid formation is prompted by both topography and cell-surface interactivity and that there is a connection between multicellular spheroid formation and adipogenesis. Copyright © 2017 Elsevier B.V. All rights reserved.

ISSUES IN ENDOCRINE DISRUPTION: COMPARING CRITICAL PERIODS OF HORMONE SENSITIVITY

EPA Science Inventory

Japanese medaka (Oryzias latipes) have been developed as a model species to compare the effects of endocrine active chemicals at critical life-stage periods of hormonal sensitivity, specifically as reproductively active adults, during the developmental period of differentiation, ...

ASSESSMENT OF THE REPRODUCTIVE AXIS: COMPARING CRITICAL PERIODS OF HORMONE SENSITIVITY.

EPA Science Inventory

Japanese medaka (Oryzias latipes) have been developed as a model species to compare the effects of endocrine active chemicals at critical life-stage periods of hormonal sensitivity, specifically as reproductively active adults, during the developmental period of differentiation, ...

In vitro haematopoiesis of a novel dendritic-like cell present in murine spleen.

PubMed

Tan, Jonathan K H; O'Neill, Helen C

2010-12-01

Dendritic cells (DC) are important antigen presenting cells (APC) which induce and control the adaptive immune response. In spleen alone, multiple DC subsets can be distinguished by cell surface marker phenotype. Most of these have been shown to develop from progenitors in bone marrow and to seed lymphoid and tissue sites during development. This study advances in vitro methodology for haematopoiesis of dendritic-like cells from progenitors in spleen. Since spleen progenitors undergo differentiation in vitro to produce these cells, the possibility exists that spleen represents a specific niche for differentiation of this subset. The fact that an equivalent cell subset has been shown to exist in spleen also supports that hypothesis. Studies have been directed at investigating the specific functional role of this novel subset as an APC accessible to blood-borne antigen, as well as the conditions under which haematopoiesis is initiated in spleen, and the type of progenitor involved.

Liver-enriched transcription factors are critical for the expression of hepatocyte marker genes in mES-derived hepatocyte-lineage cells.

PubMed

Kheolamai, Pakpoom; Dickson, Alan J

2009-04-23

Induction of stem cell differentiation toward functional hepatocytes is hampered by lack of knowledge of the hepatocyte differentiation processes. The overall objective of this project is to characterize key stages in the hepatocyte differentiation process. We established a mouse embryonic stem (mES) cell culture system which exhibited changes in gene expression profiles similar to those observed in the development of endodermal and hepatocyte-lineage cells previously described in the normal mouse embryo. Transgenic mES cells were established that permitted isolation of enriched hepatocyte-lineage populations. This approach has isolated mES-derived hepatocyte-lineage cells that express several markers of mature hepatocytes including albumin, glucose-6-phosphatase, tyrosine aminotransferase, cytochrome P450-3a, phosphoenolpyruvate carboxykinase and tryptophan 2,3-dioxygenase. In addition, our results show that the up-regulation of the expression levels of hepatocyte nuclear factor-3alpha, -4alpha, -6, and CCAAT-enhancer binding protein-beta might be critical for passage into late-stage differentiation towards functional hepatocytes. These data present important steps for definition of regulatory phenomena that direct specific cell fate determination. The mES cell culture system generated in this study provides a model for studying transition between stages of the hepatocyte development and has significant potential value for studying the molecular basis of hepatocyte differentiation in vitro.

Sex-specific inhibition and stimulation of worker-reproductive transition in a termite.

PubMed

Sun, Qian; Haynes, Kenneth F; Hampton, Jordan D; Zhou, Xuguo

2017-09-06

In social insects, the postembryonic development of individuals exhibits strong phenotypic plasticity in response to the environment, thus generating the caste system. Different from eusocial Hymenoptera, in which queens dominate reproduction and inhibit worker fertility, the primary reproductive caste in termites (kings and queens) can be replaced by neotenic reproductives derived from functionally sterile individuals. Feedback regulation of nestmate differentiation into reproductives has been suggested, but the sex specificity remains inconclusive. In the eastern subterranean termite, Reticulitermes flavipes, we tested the hypothesis that neotenic reproductives regulate worker-reproductive transition in a sex-specific manner. With this R. flavipes system, we demonstrate a sex-specific regulatory mechanism with both inhibitory and stimulatory functions. Neotenics inhibit workers of the same sex from differentiating into additional reproductives but stimulate workers of the opposite sex to undergo this transition. Furthermore, this process is not affected by the presence of soldiers. Our results highlight the reproductive plasticity of termites in response to social cues and provide insights into the regulation of reproductive division of labor in a hemimetabolous social insect.

Sex-specific inhibition and stimulation of worker-reproductive transition in a termite

NASA Astrophysics Data System (ADS)

Sun, Qian; Haynes, Kenneth F.; Hampton, Jordan D.; Zhou, Xuguo

2017-10-01

In social insects, the postembryonic development of individuals exhibits strong phenotypic plasticity in response to the environment, thus generating the caste system. Different from eusocial Hymenoptera, in which queens dominate reproduction and inhibit worker fertility, the primary reproductive caste in termites (kings and queens) can be replaced by neotenic reproductives derived from functionally sterile individuals. Feedback regulation of nestmate differentiation into reproductives has been suggested, but the sex specificity remains inconclusive. In the eastern subterranean termite, Reticulitermes flavipes, we tested the hypothesis that neotenic reproductives regulate worker-reproductive transition in a sex-specific manner. With this R. flavipes system, we demonstrate a sex-specific regulatory mechanism with both inhibitory and stimulatory functions. Neotenics inhibit workers of the same sex from differentiating into additional reproductives but stimulate workers of the opposite sex to undergo this transition. Furthermore, this process is not affected by the presence of soldiers. Our results highlight the reproductive plasticity of termites in response to social cues and provide insights into the regulation of reproductive division of labor in a hemimetabolous social insect.

Ovarian Conservation and Overall Survival in Young Women With Early-Stage Cervical Cancer.

PubMed

Matsuo, Koji; Machida, Hiroko; Shoupe, Donna; Melamed, Alexander; Muderspach, Laila I; Roman, Lynda D; Wright, Jason D

2017-01-01

To identify predictors of ovarian conservation at hysterectomy and to examine the association of ovarian conservation and survival of young women with early-stage cervical cancer. This is a retrospective cohort study using the Surveillance, Epidemiology, and End Results Program to identify hysterectomy-based surgically treated patients with stage I cervical cancer diagnosed between 1983 and 2012 (N=16,511). Multivariable models were used to identify independent factors associated with ovarian conservation. Among the subgroup of 9,419 women younger than 50 years of age with stage I disease, survival outcomes and causes of death were examined for 3,908 (41.5%) women who underwent ovarian conservation at hysterectomy without radiotherapy. On multivariable analysis, age younger than 50 years, stage IA disease, and squamous histology were independent factors associated with ovarian conservation (all, P<.001). Among 5,526 women younger than 50 years of age with stage IA disease who underwent hysterectomy without radiotherapy, overall survival was significantly higher in patients undergoing ovarian conservation than in those undergoing oophorectomy (20-year rate, 93.5% compared with 86.8%, P<.001); cervical cancer-specific survival was similar between the patients who underwent ovarian conservation and those who underwent oophorectomy (98.8% compared with 97.8%, P=.12). On multivariable analysis, ovarian conservation remained an independent prognostic factor for improved overall survival (adjusted hazard ratio 0.63, 95% confidence interval [CI] 0.49-0.82, P=.001) and was independently associated with lower cumulative risks of death resulting from cardiovascular disease (20-year cumulative rate, 1.2% compared with 3.3%, adjusted hazard ratio 0.47, 95% CI 0.26-0.86, P=.014) and other chronic disease (0.5% compared with 1.4%, adjusted hazard ratio 0.24, 95% CI 0.09-0.65, P=.005) compared with oophorectomy. Both cervical cancer-specific survival (20-year rate, 93.1% compared with 92.0%, P=.37) and overall survival (86.7% compared with 84.6%, P=.12) were similar between ovarian conservation and oophorectomy among 3,893 women younger than 50 years of age with stage IB disease who underwent hysterectomy without radiotherapy. Among young women with stage IA cervical cancer, ovarian conservation at hysterectomy is associated with decreased all-cause mortality including death resulting from cardiovascular disease and other chronic diseases.

Is Duodenal Invasion a Relevant Prognosticator in Patients Undergoing Adjuvant Chemoradiotherapy for Distal Common Bile Duct Cancer?

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

Kim, Kyubo; Chie, Eui Kyu, E-mail: ekchie93@snu.ac.k; Jang, Jin-Young

2010-07-15

Purpose: To analyze the outcome of adjuvant chemoradiotherapy for patients with distal common bile duct (CBD) cancer who underwent curative surgery, and to identify the prognostic factors for these patients. Methods and Materials: Between January 1991 and December 2002, 38 patients with adenocarcinoma of the distal CBD underwent curative resection followed by adjuvant chemoradiotherapy. There were 27 men and 11 women, and the median age was 60 years (range, 34-73). Adjuvant radiotherapy was delivered to the tumor bed and regional lymph nodes up to 40 Gy at 2 Gy/fraction with a 2-week planned rest. Intravenous 5-fluorouracil (500mg/m{sup 2}/day) was givenmore » on day 1 to day 3 of each split course. The median follow-up period was 39 months. Results: The 5-year overall survival rate of all patients was 49.1%. On univariate analysis, only histologic differentiation (p = 0.0005) was associated with overall survival. Tumor size ({<=}2cm vs. >2cm) had a marginally significant impact on the treatment outcome (p = 0.0624). However, there was no difference in overall survival rates between T3 and T4 tumors (p = 0.6189), for which the main determinants were pancreatic and duodenal invasion, respectively. On multivariate analysis, histologic differentiation (p = 0.0092) and tumor size (p = 0.0046) were independent risk factors for overall survival. Conclusions: Long-term survival can be expected in patients with distal CBD cancer undergoing curative surgery and adjuvant chemoradiotherapy. Histologic differentiation and tumor size were significant prognostic factors predicting overall survival, whereas duodenal invasion was not. This finding suggests the need for further refinement in tumor staging.« less

Effects of Swallowing Exercises on Patients Undergoing Radiation Treatment for Head and Neck Cancer

ClinicalTrials.gov

2017-05-25

Head and Neck Cancer; Stage I Hypopharyngeal Cancer; Stage I Laryngeal Cancer; Stage I Oropharyngeal Cancer; Stage II Hypopharyngeal Cancer; Stage II Laryngeal Cancer; Stage II Oropharyngeal Cancer; Stage III Hypopharyngeal Cancer; Stage III Laryngeal Cancer; Stage III Oropharyngeal Cancer; Stage IV Hypopharyngeal Cancer; Stage IV Laryngeal Cancer; Stage IV Oropharyngeal Cancer

A rat RNA-Seq transcriptomic BodyMap across 11 organs and 4 developmental stages

PubMed Central

Yu, Ying; Fuscoe, James C.; Zhao, Chen; Guo, Chao; Jia, Meiwen; Qing, Tao; Bannon, Desmond I.; Lancashire, Lee; Bao, Wenjun; Du, Tingting; Luo, Heng; Su, Zhenqiang; Jones, Wendell D.; Moland, Carrie L.; Branham, William S.; Qian, Feng; Ning, Baitang; Li, Yan; Hong, Huixiao; Guo, Lei; Mei, Nan; Shi, Tieliu; Wang, Kevin Y.; Wolfinger, Russell D.; Nikolsky, Yuri; Walker, Stephen J.; Duerksen-Hughes, Penelope; Mason, Christopher E.; Tong, Weida; Thierry-Mieg, Jean; Thierry-Mieg, Danielle; Shi, Leming; Wang, Charles

2014-01-01

The rat has been used extensively as a model for evaluating chemical toxicities and for understanding drug mechanisms. However, its transcriptome across multiple organs, or developmental stages, has not yet been reported. Here we show, as part of the SEQC consortium efforts, a comprehensive rat transcriptomic BodyMap created by performing RNA-Seq on 320 samples from 11 organs of both sexes of juvenile, adolescent, adult and aged Fischer 344 rats. We catalogue the expression profiles of 40,064 genes, 65,167 transcripts, 31,909 alternatively spliced transcript variants and 2,367 non-coding genes/non-coding RNAs (ncRNAs) annotated in AceView. We find that organ-enriched, differentially expressed genes reflect the known organ-specific biological activities. A large number of transcripts show organ-specific, age-dependent or sex-specific differential expression patterns. We create a web-based, open-access rat BodyMap database of expression profiles with crosslinks to other widely used databases, anticipating that it will serve as a primary resource for biomedical research using the rat model. PMID:24510058

Micropattern differentiation of mouse pluripotent stem cells recapitulates embryo regionalized cell fate patterning

PubMed Central

Morgani, Sophie M; Metzger, Jakob J; Nichols, Jennifer

2018-01-01

During gastrulation epiblast cells exit pluripotency as they specify and spatially arrange the three germ layers of the embryo. Similarly, human pluripotent stem cells (PSCs) undergo spatially organized fate specification on micropatterned surfaces. Since in vivo validation is not possible for the human, we developed a mouse PSC micropattern system and, with direct comparisons to mouse embryos, reveal the robust specification of distinct regional identities. BMP, WNT, ACTIVIN and FGF directed mouse epiblast-like cells to undergo an epithelial-to-mesenchymal transition and radially pattern posterior mesoderm fates. Conversely, WNT, ACTIVIN and FGF patterned anterior identities, including definitive endoderm. By contrast, epiblast stem cells, a developmentally advanced state, only specified anterior identities, but without patterning. The mouse micropattern system offers a robust scalable method to generate regionalized cell types present in vivo, resolve how signals promote distinct identities and generate patterns, and compare mechanisms operating in vivo and in vitro and across species. PMID:29412136

Characterization of keratinocyte differentiation induced by ascorbic acid: protein kinase C involvement and vitamin C homeostasis.

PubMed

Savini, Isabella; Catani, Maria Valeria; Rossi, Antonello; Duranti, Guglielmo; Melino, Gerry; Avigliano, Luciana

2002-02-01

Epidermal keratinocytes undergo differentiation in response to several stimuli to form the cornified envelope, a structure that contributes to the barrier function of skin. Although differentiation has been extensively analyzed, the precise role of vitamin C during this process is still not defined. Ascorbic acid, besides acting as a radical scavenger, has been shown to promote mesenchymal differentiation. In this study, we found that keratinocytes grown in ascorbate-supplemented medium developed a differentiated phenotype, as demonstrated by enhanced expression of marker genes and increase in cornified envelope content. The pro-differentiating effects of ascorbate were mediated by the protein-kinase-C-dependent induction of activating protein 1 DNA binding activity; indeed, down-modulation of protein kinase C activity abolished differentiation triggered by ascorbic acid. Although vitamin C appeared to regulate the same signaling pathway modulated by calcium, a classical in vitro inducer of epidermal differentiation, nonetheless terminally differentiated keratinocytes exhibited different ascorbate homeostasis and cellular antioxidant status. Indeed, we found that, unlike calcium, differentiation promoted by ascorbate was accompanied by (i) an enhanced ascorbate transport, due to overexpression of specific transporters, (ii) a great efficiency of dehydroascorbate uptake, and (iii) an increase in glutathione content with respect to proliferating cells. Ascorbic acid may be useful to promote epidermal differentiation, avoiding depletion of hydrophilic antioxidant stores.

Temporal Impact of Substrate Mechanics on Differentiation of Human Embryonic Stem Cells to Cardiomyocytes

PubMed Central

Hazeltine, Laurie B.; Badur, Mehmet G.; Lian, Xiaojun; Das, Amritava; Han, Wenqing; Palecek, Sean P.

2014-01-01

A significant clinical need exists to differentiate human pluripotent stem cells (hPSCs) into cardiomyocytes, enabling tissue modeling for in vitro discovery of new drugs or cell-based therapies for heart repair in vivo. Chemical and mechanical microenvironmental factors are known to impact efficiency of stem cell differentiation, but cardiac differentiation protocols in hPSCs are typically performed on rigid tissue culture polystyrene (TCPS) surfaces which do not present a physiological mechanical setting. To investigate the temporal effects of mechanics on cardiac differentiation, we cultured human embryonic stem cells (hESCs) and their derivatives on polyacrylamide hydrogel substrates with a physiologically relevant range of stiffnesses. In directed differentiation and embryoid body culture systems, differentiation of hESCs to cardiac Troponin T-expressing (cTnT+) cardiomyocytes peaked on hydrogels of intermediate stiffness. Brachyury expression also peaked on intermediate stiffness hydrogels at day 1 of directed differentiation, suggesting that stiffness impacted the initial differentiation trajectory of hESCs to mesendoderm. To investigate the impact of substrate mechanics during cardiac specification of mesodermal progenitors, we initiated directed cardiomyocyte differentiation on TCPS and transferred cells to hydrogels at the Nkx2.5/Isl1+ cardiac progenitor cell stage. No differences in cardiomyocyte purity with stiffness were observed on day 15. These experiments indicate that differentiation of hESCs is sensitive to substrate mechanics at early stages of mesodermal induction, and proper application of substrate mechanics can increase the propensity of hESCs to differentiate to cardiomyocytes. PMID:24200714

Flower development and sex specification in wild grapevine.

PubMed

Ramos, Miguel Jesus Nunes; Coito, João Lucas; Silva, Helena Gomes; Cunha, Jorge; Costa, Maria Manuela Ribeiro; Rocheta, Margarida

2014-12-12

Wild plants of Vitis closely related to the cultivated grapevine (V. v. vinifera) are believed to have been first domesticated 10,000 years BC around the Caspian Sea. V. v. vinifera is hermaphrodite whereas V. v. sylvestris is a dioecious species. Male flowers show a reduced pistil without style or stigma and female flowers present reflexed stamens with infertile pollen. V. vinifera produce perfect flowers with all functional structures. The mechanism for flower sex determination and specification in grapevine is still unknown. To understand which genes are involved during the establishment of male, female and complete flowers, we analysed and compared the transcription profiles of four developmental stages of the three genders. We showed that sex determination is a late event during flower development and that the expression of genes from the ABCDE model is not directly correlated with the establishment of sexual dimorphism. We propose a temporal comprehensive model in which two mutations in two linked genes could be players in sex determination and indirectly establish the Vitis domestication process. Additionally, we also found clusters of genes differentially expressed between genders and between developmental stages that suggest a role involved in sex differentiation. Also, the detection of differentially transcribed regions that extended existing gene models (intergenic regions) between sexes suggests that they may account for some of the variation between the subspecies. There is no evidence of differences of expression levels in genes from the ABCDE model that could explain the shift from hermaphroditism to dioecy. We propose that sex specification occurs after floral organ identity has been established and therefore, sex determination genes might be having an effect downstream of the ABCDE model genes.For the first time a full transcriptomic analysis was performed in different flower developmental stages in the same individual. Our experimental approach enabled us to create a comprehensive catalogue of transcribed genes across developmental stages and genders that will contribute for future work in sex determination in seed plants.

Semi-Quantitative Evaluation of Secondary Carbonates via Portable X-ray Fluorescence Spectrometry

NASA Astrophysics Data System (ADS)

Chakraborty, Somsubhra; Weindorf, David; Weindorf, Camille; Duda, Bogdan; Pennington, Sarah; Ortiz, Rebekah

2017-04-01

Secondary calcium carbonate commonly occurs in subsoils of semi-arid soils worldwide. In US Soil Taxonomy, such horizons are frequently described as Bk, Bkk, Bkm, Bkkm, or Ck horizons at variable stages of development. Specifically, the Soil Survey Staff uses a qualitative scale of one through six to indicate differential developmental stages. However, considerable disagreement exists even among experienced soil scientists. Evaluating 75 soil samples from across four US states, a portable X-ray fluorescence (PXRF) spectrometer was used to quantify the total soil Ca content and compare it to average developmental stage scores as determined by a panel of Soil Survey Staff personnel. Samples were evaluated both as intact aggregates as well as ground (<2 mm), homogenized powders. PXRF readings of total soil Ca concentration steadily increased under both conditions as developmental stage progressed. However, minimal difference was observed between stage five and six carbonate accumulation. Stage three showed the widest variability in total soil Ca. Given than PXRF cannot distinguish between primary and secondary CaCO3 in soils, interpretation by the analyst remains essential. Nonetheless, PXRF provides an important tool for assessing carbonate laden subsoils providing elemental differentiation beyond that perceived by the human eye.

BARHL2 differentially regulates the development of retinal amacrine and ganglion neurons

PubMed Central

Ding, Qian; Chen, Hui; Xie, Xiaoling; Libby, Richard T.; Tian, Ning; Gan, Lin

2009-01-01

Summary Through transcriptional regulations the BarH family of homeodomain proteins play essential roles in cell fate specification, cell differentiation, migration and survival. Barhl2, a member of the Barh gene family, is expressed in retinal ganglion cells (RGCs), amacrine cells (ACs) and horizontal cells. Here, to investigate the role of Barhl2 in retinal development, Barhl2 deficient mice were generated. Analysis of AC subtypes in Barhl2 deficient retinas suggests that Barhl2 plays a critical role in AC subtype determination. A significant reduction of glycinergic and GABAergic ACs with a substantial increase in the number of cholinergic ACs was observed in Barhl2-null retinas. Barhl2 is also critical for the development of a normal complement of RGCs. Barhl2 deficiency resulted in a 35% increase in RGCs undergoing apoptosis during development. Genetic analysis revealed that Barhl2 functions downstream of the Atoh7-Pou4f3 regulatory pathway and regulates the maturation and/or survival of RGCs. Thus, BARHL2 appears to have numerous roles in retinal development, including regulating neuronal subtype specification, differentiation, and survival. PMID:19339595

Mesenchymal stem cell-derived exosomes have altered microRNA profiles and induce osteogenic differentiation depending on the stage of differentiation

PubMed Central

Wang, Xiaoqin; Omar, Omar; Vazirisani, Forugh; Thomsen, Peter

2018-01-01

Human mesenchymal stem cell (hMSC)-derived exosomes have shown regenerative effects, but their role in osteogenesis and the underlying mechanism are yet to be determined. In this study, we examined the time-course secretion of exosomes by hMSCs during the entire process of osteogenic differentiation. Exosomes derived from hMSCs in various stages of osteogenic differentiation committed homotypic cells to differentiate towards osteogenic lineage, but only exosomes from late stages of osteogenic differentiation induced extracellular matrix mineralisation. Exosomes from expansion and early and late stages of osteogenic differentiation were internalised by a subpopulation of hMSCs. MicroRNA profiling revealed a set of differentially expressed exosomal microRNAs from the late stage of osteogenic differentiation, which were osteogenesis related. Target prediction demonstrated that these microRNAs enriched pathways involved in regulation of osteogenic differentiation and general mechanisms how exosomes exert their functions, such as “Wnt signalling pathway” and “endocytosis”. Taken together, the results show that MSCs secrete exosomes with different biological properties depending on differentiation stage of their parent cells. The exosomal cargo transferred from MSCs in the late stage of differentiation induces osteogenic differentiation and mineralisation. Moreover, it is suggested that the regulatory effect on osteogenesis by exosomes is at least partly exerted by exosomal microRNA. PMID:29447276

The landscape of sex-differential transcriptome and its consequent selection in human adults.

PubMed

Gershoni, Moran; Pietrokovski, Shmuel

2017-02-07

The prevalence of several human morbid phenotypes is sometimes much higher than intuitively expected. This can directly arise from the presence of two sexes, male and female, in one species. Men and women have almost identical genomes but are distinctly dimorphic, with dissimilar disease susceptibilities. Sexually dimorphic traits mainly result from differential expression of genes present in both sexes. Such genes can be subject to different, and even opposing, selection constraints in the two sexes. This can impact human evolution by differential selection on mutations with dissimilar effects on the two sexes. We comprehensively mapped human sex-differential genetic architecture across 53 tissues. Analyzing available RNA-sequencing data from 544 adults revealed thousands of genes differentially expressed in the reproductive tracts and tissues common to both sexes. Sex-differential genes are related to various biological systems, and suggest new insights into the pathophysiology of diverse human diseases. We also identified a significant association between sex-specific gene transcription and reduced selection efficiency and accumulation of deleterious mutations, which might affect the prevalence of different traits and diseases. Interestingly, many of the sex-specific genes that also undergo reduced selection efficiency are essential for successful reproduction in men or women. This seeming paradox might partially explain the high incidence of human infertility. This work provides a comprehensive overview of the sex-differential transcriptome and its importance to human evolution and human physiology in health and in disease.

Biogenesis of protein bodies during legumin accumulation in developing olive (Olea europaea L.) seed.

PubMed

Jimenez-Lopez, Jose C; Zienkiewicz, Agnieszka; Zienkiewicz, Krzysztof; Alché, Juan D; Rodríguez-García, Maria I

2016-03-01

Much of our current knowledge about seed development and differentiation regarding reserves synthesis and accumulation come from monocot (cereals) plants. Studies in dicotyledonous seeds differentiation are limited to a few species and in oleaginous species are even scarcer despite their agronomic and economic importance. We examined the changes accompanying the differentiation of olive endosperm and cotyledon with a focus on protein bodies (PBs) biogenesis during legumin protein synthesis and accumulation, with the aim of getting insights and a better understanding of the PBs' formation process. Cotyledon and endosperm undergo differentiation during seed development, where an asynchronous time-course of protein synthesis, accumulation, and differential PB formation patterns was found in both tissues. At the end of seed maturation, a broad population of PBs, particularly in cotyledon cells, was distinguishable in terms of number per cell and morphometric and cytochemical features. Olive seed development is a tissue-dependent process characterized by differential rates of legumin accumulation and PB formation in the main tissues integrating seed. One of the main features of the impressive differentiation process is the specific formation of a broad group of PBs, particularly in cotyledon cells, which might depend on selective accumulation and packaging of proteins and specific polypeptides into PBs. The nature and availability of the major components detected in the PBs of olive seed are key parameters in order to consider the potential use of this material as a suitable source of carbon and nitrogen for animal or even human use.

Retinoic Acid Therapy Resistance Progresses from Unilineage to Bilineage in HL-60 Leukemic Blasts

PubMed Central

Jensen, Holly A.; Bunaciu, Rodica P.; Ibabao, Christopher N.; Myers, Rebecca; Varner, Jeffrey D.; Yen, Andrew

2014-01-01

Emergent resistance can be progressive and driven by global signaling aberrations. All-trans retinoic acid (RA) is the standard therapeutic agent for acute promyelocytic leukemia, but 10–20% of patients are not responsive, and initially responsive patients relapse and develop retinoic acid resistance. The patient-derived, lineage-bipotent acute myeloblastic leukemia (FAB M2) HL-60 cell line is a potent tool for characterizing differentiation-induction therapy responsiveness and resistance in t(15;17)-negative cells. Wild-type (WT) HL-60 cells undergo RA-induced granulocytic differentiation, or monocytic differentiation in response to 1,25-dihydroxyvitamin D3 (D3). Two sequentially emergent RA-resistant HL-60 cell lines, R38+ and R38-, distinguishable by RA-inducible CD38 expression, do not arrest in G1/G0 and fail to upregulate CD11b and the myeloid-associated signaling factors Vav1, c-Cbl, Lyn, Fgr, and c-Raf after RA treatment. Here, we show that the R38+ and R38- HL-60 cell lines display a progressive reduced response to D3-induced differentiation therapy. Exploiting the biphasic dynamic of induced HL-60 differentiation, we examined if resistance-related defects occurred during the first 24 h (the early or “precommitment” phase) or subsequently (the late or “lineage-commitment” phase). HL-60 were treated with RA or D3 for 24 h, washed and retreated with either the same, different, or no differentiation agent. Using flow cytometry, D3 was able to induce CD38, CD11b and CD14 expression, and G1/G0 arrest when present during the lineage-commitment stage in R38+ cells, and to a lesser degree in R38- cells. Clustering analysis of cytometry and quantified Western blot data indicated that WT, R38+ and R38- HL-60 cells exhibited decreasing correlation between phenotypic markers and signaling factor expression. Thus differentiation induction therapy resistance can develop in stages, with initial partial RA resistance and moderate vitamin D3 responsiveness (unilineage maturation block), followed by bilineage maturation block and progressive signaling defects, notably the reduced expression of Vav1, Fgr, and c-Raf. PMID:24922062

Gene function in early mouse embryonic stem cell differentiation

PubMed Central

Sene, Kagnew Hailesellasse; Porter, Christopher J; Palidwor, Gareth; Perez-Iratxeta, Carolina; Muro, Enrique M; Campbell, Pearl A; Rudnicki, Michael A; Andrade-Navarro, Miguel A

2007-01-01

Background Little is known about the genes that drive embryonic stem cell differentiation. However, such knowledge is necessary if we are to exploit the therapeutic potential of stem cells. To uncover the genetic determinants of mouse embryonic stem cell (mESC) differentiation, we have generated and analyzed 11-point time-series of DNA microarray data for three biologically equivalent but genetically distinct mESC lines (R1, J1, and V6.5) undergoing undirected differentiation into embryoid bodies (EBs) over a period of two weeks. Results We identified the initial 12 hour period as reflecting the early stages of mESC differentiation and studied probe sets showing consistent changes of gene expression in that period. Gene function analysis indicated significant up-regulation of genes related to regulation of transcription and mRNA splicing, and down-regulation of genes related to intracellular signaling. Phylogenetic analysis indicated that the genes showing the largest expression changes were more likely to have originated in metazoans. The probe sets with the most consistent gene changes in the three cell lines represented 24 down-regulated and 12 up-regulated genes, all with closely related human homologues. Whereas some of these genes are known to be involved in embryonic developmental processes (e.g. Klf4, Otx2, Smn1, Socs3, Tagln, Tdgf1), our analysis points to others (such as transcription factor Phf21a, extracellular matrix related Lama1 and Cyr61, or endoplasmic reticulum related Sc4mol and Scd2) that have not been previously related to mESC function. The majority of identified functions were related to transcriptional regulation, intracellular signaling, and cytoskeleton. Genes involved in other cellular functions important in ESC differentiation such as chromatin remodeling and transmembrane receptors were not observed in this set. Conclusion Our analysis profiles for the first time gene expression at a very early stage of mESC differentiation, and identifies a functional and phylogenetic signature for the genes involved. The data generated constitute a valuable resource for further studies. All DNA microarray data used in this study are available in the StemBase database of stem cell gene expression data [1] and in the NCBI's GEO database. PMID:17394647

Transcriptome Profiling of Wheat Inflorescence Development from Spikelet Initiation to Floral Patterning Identified Stage-Specific Regulatory Genes1[OPEN

PubMed Central

Feng, Nan; Song, Gaoyuan; Guan, Jiantao; Chen, Kai; Jia, Meiling; Huang, Dehua; Wu, Jiajie; Zhang, Lichao; Kong, Xiuying; Geng, Shuaifeng

2017-01-01

Early reproductive development in cereals is crucial for final grain number per spike and hence the yield potential of the crop. To date, however, no systematic analyses of gene expression profiles during this important process have been conducted for common wheat (Triticum aestivum). Here, we studied the transcriptome profiles at four stages of early wheat reproductive development, from spikelet initiation to floral organ differentiation. K-means clustering and stage-specific transcript identification detected dynamically expressed homeologs of important transcription regulators in spikelet and floral meristems that may be involved in spikelet initiation, floret meristem specification, and floral organ patterning, as inferred from their homologs in model plants. Small RNA transcriptome sequencing discovered key microRNAs that were differentially expressed during wheat inflorescence development alongside their target genes, suggesting that miRNA-mediated regulatory mechanisms for floral development may be conserved in cereals and Arabidopsis. Our analysis was further substantiated by the functional characterization of the ARGONAUTE1d (AGO1d) gene, which was initially expressed in stamen primordia and later in the tapetum during anther maturation. In agreement with its stage-specific expression pattern, the loss of function of the predominantly expressed B homeolog of AGO1d in a tetraploid durum wheat mutant resulted in smaller anthers with more infertile pollens than the wild type and a reduced grain number per spike. Together, our work provides a first glimpse of the gene regulatory networks in wheat inflorescence development that may be pivotal for floral and grain development, highlighting potential targets for genetic manipulation to improve future wheat yields. PMID:28515146

De Novo Assembly and Comparative Transcriptome Analysis Provide Insight into Lysine Biosynthesis in Toona sinensis Roem.

PubMed

Zhang, Xia; Song, Zhenqiao; Liu, Tian; Guo, Linlin; Li, Xingfeng

2016-01-01

Toona sinensis Roem is a popular leafy vegetable in Chinese cuisine and is also used as a traditional Chinese medicine. In this study, leaf samples were collected from the same plant on two development stages and then used for high-throughput Illumina RNA-sequencing (RNA-Seq). 125,884 transcripts and 54,628 unigenes were obtained through de novo assembly. A total of 25,570 could be annotated with known biological functions, which indicated that the T. sinensis leaves and shoots were undergoing multiple developmental processes especially for active metabolic processes. Analysis of differentially expressed unigenes between the two libraries showed that the lysine biosynthesis was an enriched KEGG pathway, and candidate genes involved in the lysine biosynthesis pathway in T. sinensis leaves and shoots were identified. Our results provide a primary analysis of the gene expression files of T. sinensis leaf and shoot on different development stages and afford a valuable resource for genetic and genomic research on plant lysine biosynthesis.

Construction of a Lotus japonicus late nodulin expressed sequence tag library and identification of novel nodule-specific genes.

PubMed Central

Szczyglowski, K; Hamburger, D; Kapranov, P; de Bruijn, F J

1997-01-01

A range of novel expressed sequence tags (ESTs) associated with late developmental events during nodule organogenesis in the legume Lotus japonicus were identified using mRNA differential display; 110 differentially displayed polymerase chain reaction products were cloned and analyzed. Of 88 unique cDNAs obtained, 22 shared significant homology to DNA/protein sequences in the respective databases. This group comprises, among others, a nodule-specific homolog of protein phosphatase 2C, a peptide transporter protein, and a nodule-specific form of cytochrome P450. RNA gel-blot analysis of 16 differentially displayed ESTs confirmed their nodule-specific expression pattern. The kinetics of mRNA accumulation of the majority of the ESTs analyzed were found to resemble the expression pattern observed for the L. japonicus leghemoglobin gene. These results indicate that the newly isolated molecular markers correspond to genes induced during late developmental stages of L. japonicus nodule organogenesis and provide important, novel tools for the study of nodulation. PMID:9276951

Transcriptome Analysis of the Signalling Networks in Coronatine-Induced Secondary Laticifer Differentiation from Vascular Cambia in Rubber Trees

PubMed Central

Wu, Shaohua; Zhang, Shixin; Chao, Jinquan; Deng, Xiaomin; Chen, Yueyi; Shi, Minjing; Tian, Wei-Min

2016-01-01

The secondary laticifer in rubber tree (Hevea brasiliensis Muell. Arg.) is a specific tissue within the secondary phloem. This tissue differentiates from the vascular cambia, and its function is natural rubber biosynthesis and storage. Given that jasmonates play a pivotal role in secondary laticifer differentiation, we established an experimental system with jasmonate (JA) mimic coronatine (COR) for studying the secondary laticifer differentiation: in this system, differentiation occurs within five days of the treatment of epicormic shoots with COR. In the present study, the experimental system was used to perform transcriptome sequencing and gene expression analysis. A total of 67,873 unigenes were assembled, and 50,548 unigenes were mapped at least in one public database. Of these being annotated unigenes, 15,780 unigenes were differentially expressed early after COR treatment, and 19,824 unigenes were differentially expressed late after COR treatment. At the early stage, 8,646 unigenes were up-regulated, while 7,134 unigenes were down-regulated. At the late stage, the numbers of up- and down-regulated unigenes were 7,711 and 12,113, respectively. The annotation data and gene expression analysis of the differentially expressed unigenes suggest that JA-mediated signalling, Ca2+ signal transduction and the CLAVATA-MAPK-WOX signalling pathway may be involved in regulating secondary laticifer differentiation in rubber trees. PMID:27808245

Quantitative Proteomics Uncovers Novel Factors Involved in Developmental Differentiation of Trypanosoma brucei

PubMed Central

Dejung, Mario; Subota, Ines; Bucerius, Ferdinand; Dindar, Gülcin; Freiwald, Anja; Engstler, Markus; Boshart, Michael; Butter, Falk; Janzen, Christian J.

2016-01-01

Developmental differentiation is a universal biological process that allows cells to adapt to different environments to perform specific functions. African trypanosomes progress through a tightly regulated life cycle in order to survive in different host environments when they shuttle between an insect vector and a vertebrate host. Transcriptomics has been useful to gain insight into RNA changes during stage transitions; however, RNA levels are only a moderate proxy for protein abundance in trypanosomes. We quantified 4270 protein groups during stage differentiation from the mammalian-infective to the insect form and provide classification for their expression profiles during development. Our label-free quantitative proteomics study revealed previously unknown components of the differentiation machinery that are involved in essential biological processes such as signaling, posttranslational protein modifications, trafficking and nuclear transport. Furthermore, guided by our proteomic survey, we identified the cause of the previously observed differentiation impairment in the histone methyltransferase DOT1B knock-out strain as it is required for accurate karyokinesis in the first cell division during differentiation. This epigenetic regulator is likely involved in essential chromatin restructuring during developmental differentiation, which might also be important for differentiation in higher eukaryotic cells. Our proteome dataset will serve as a resource for detailed investigations of cell differentiation to shed more light on the molecular mechanisms of this process in trypanosomes and other eukaryotes. PMID:26910529

Evaluation of the AJCC 8th Edition Staging System for Pathologically Versus Clinically Staged Intrahepatic Cholangiocarcinoma (iCCA): a Time to Revisit a Dogma? A Surveillance, Epidemiology, and End Results (SEER) Analysis.

PubMed

Kamarajah, Sivesh K

2018-03-07

Recently, the AJCC has released its 8th edition changes to the staging system for intrahepatic cholangiocarcinoma (iCCA). This study sought to validate the proposed changes to the 8th edition of AJCC system for T and N classification of iCCA using a population-based data set. Using the Surveillance, Epidemiology, and End Results (SEER) database (1998-2013), patients undergoing resection or non-surgical management for non-metastatic iCCA were identified. Overall survival was estimated using the Kaplan-Meier method and compared using log-rank tests. Concordance indices (c-indices) calculated from Cox proportional hazards models were calculated to evaluate discriminatory power. The study included 2630 patients resected (37%) or non-surgically managed (63%) for iCCA. Nodal staging was performed in 56%, of whom 31% had positive nodes. For all patients with iCCA, the median 5-year survival by AJCC T classification for T1a, T1b, T2, T3, and T4 was 32, 21, 14, 10, and 10 months, respectively (p < 0.001). The concordance index for the staging system was 0.57 for all patients, 0.62 for those who underwent resection, and 0.54 for patients who did not undergo resection. In summary, the new AJCC 8th edition staging system is comparable to the 7th edition and valid in stratifying patients with iCCA. However, the performance of the staging system is better in patients undergoing surgical resection than those undergoing non-surgical management. These findings further highlight the need for improved accuracy of radiological imaging in clinically staging patients to guide prognosis.

Chromatin plasticity as a differentiation index during muscle differentiation of C2C12 myoblasts

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

Watanabe, Tomonobu M.; World Premier Initiative, iFREC, Osaka University, Osaka 565-0871; Higuchi, Sayaka

Highlights: Black-Right-Pointing-Pointer Change in the epigenetic landscape during myogenesis was optically investigated. Black-Right-Pointing-Pointer Mobility of nuclear proteins was used to state the epigenetic status of the cell. Black-Right-Pointing-Pointer Mobility of nuclear proteins decreased as myogenesis progressed in C2C12. Black-Right-Pointing-Pointer Differentiation state diagram was developed using parameters obtained. -- Abstract: Skeletal muscle undergoes complicated differentiation steps that include cell-cycle arrest, cell fusion, and maturation, which are controlled through sequential expression of transcription factors. During muscle differentiation, remodeling of the epigenetic landscape is also known to take place on a large scale, determining cell fate. In an attempt to determine the extentmore » of epigenetic remodeling during muscle differentiation, we characterized the plasticity of the chromatin structure using C2C12 myoblasts. Differentiation of C2C12 cells was induced by lowering the serum concentration after they had reached full confluence, resulting in the formation of multi-nucleated myotubes. Upon induction of differentiation, the nucleus size decreased whereas the aspect ratio increased, indicating the presence of force on the nucleus during differentiation. Movement of the nucleus was also suppressed when differentiation was induced, indicating that the plasticity of chromatin changed upon differentiation. To evaluate the histone dynamics during differentiation, FRAP experiment was performed, which showed an increase in the immobile fraction of histone proteins when differentiation was induced. To further evaluate the change in the histone dynamics during differentiation, FCS was performed, which showed a decrease in histone mobility on differentiation. We here show that the plasticity of chromatin decreases upon differentiation, which takes place in a stepwise manner, and that it can be used as an index for the differentiation stage during myogenesis using the state diagram developed with the parameters obtained in this study.« less

Transdermal or Oral Telapristone Acetate in Treating Patients Undergoing Mastectomy

ClinicalTrials.gov

2018-01-22

BRCA1 Mutation Carrier; BRCA2 Mutation Carrier; Ductal Breast Carcinoma In Situ; Lobular Breast Carcinoma In Situ; Stage 0 Breast Cancer; Stage IA Breast Cancer; Stage IB Breast Cancer; Stage IIA Breast Cancer; Stage IIB Breast Cancer

Location and cellular stages of NK cell development

PubMed Central

Yu, Jianhua; Freud, Aharon G.; Caligiuri, Michael A

2013-01-01

The identification of distinct tissue-specific natural killer (NK) cell populations that apparently mature from local precursor populations has brought new insight into the diversity and developmental regulation of this important lymphoid subset. NK cells provide a necessary link between the early (innate) and late (adaptive) immune responses to infection. Gaining a better understanding of the processes that govern NK cell development should allow us to better harness NK cell functions in multiple clinical settings as well as to gain further insight into how these cells undergo malignant transformation. In this review, we summarize recent advances in understanding sites and cellular stages of NK cell development in humans and mice. PMID:24055329

Sex determination in mammalian germ cells

PubMed Central

Spiller, Cassy M; Bowles, Josephine

2015-01-01

Germ cells are the precursors of the sperm and oocytes and hence are critical for survival of the species. In mammals, they are specified during fetal life, migrate to the developing gonads and then undergo a critical period during which they are instructed, by the soma, to adopt the appropriate sexual fate. In a fetal ovary, germ cells enter meiosis and commit to oogenesis, whereas in a fetal testis, they avoid entry into meiosis and instead undergo mitotic arrest and mature toward spermatogenesis. Here, we discuss what we know so far about the regulation of sex-specific differentiation of germ cells, considering extrinsic molecular cues produced by somatic cells, as well as critical intrinsic changes within the germ cells. This review focuses almost exclusively on our understanding of these events in the mouse model. PMID:25791730

Complete life cycle of a pennellid Peniculus minuticaudae Shiino, 1956 (Copepoda: Siphonostomatoida) infecting cultured threadsail filefish, Stephanolepis cirrhifer.

PubMed

Ismail, Norshida; Ohtsuka, Susumu; Maran, Balu Alagar Venmathi; Tasumi, Satoshi; Zaleha, Kassim; Yamashita, Hirofumi

2013-01-01

The complete life cycle of a pennellid copepod Peniculus minuticaudae Shiino, 1956 is proposed based on the discovery of all post-embryonic stages together with the post-metamorphic adult females infecting the fins of threadsail filefish Stephanolepis cirrhifer (Monacanthidae) cultured in a fish farm at Ehime Prefecture, Japan. The hatching stage was the infective copepodid. The life cycle of P. minuticaudae consists of six stages separated by moults: the copepodid, four chalimi and adult. In this study, the adult males were observed frequently in precopulatory amplexus with various stages of females however, copulation occurs only between adults. Fertilized pre-metamorphic adult females carrying spermatophores may detach from the host and settle again before undergoing massive differential growth into the post-metamorphic adult female. Comparison of the life cycle of P. minuticaudae has been made with three known pennellids: Lernaeocera branchialis (Linnaeus, 1767), Cardiodectes medusaeus (Wilson, 1908) and Lernaeenicus sprattae (Sowerby, 1806). Among the compared species, P. minuticaudae is the first ectoparasitic pennellid to be discovered to complete its life cycle on a single host without any change in infection site preferences between infective copepodid and fertilized pre-metamorphic female. © N. Ismail et al., published by EDP Sciences, 2013.

Moral motivation based on multiple developmental structures: an exploration of cognitive and emotional dynamics.

PubMed

Kaplan, Ulas; Tivnan, Terrence

2014-01-01

Intrapersonal variability and multiplicity in the complexity of moral motivation were examined from Dynamic Systems and Self-Determination Theory perspectives. L. Kohlberg's (1969) stages of moral development are reconceptualized as soft-assembled and dynamically transformable process structures of motivation that may operate simultaneously within person in different degrees. Moral motivation is conceptualized as the real-time process of self-organization of cognitive and emotional dynamics out of which moral judgment and action emerge. A detailed inquiry into intrapersonal variation in moral motivation is carried out based on the differential operation of multiple motivational structures. A total of 74 high school students and 97 college students participated in the study by completing a new questionnaire, involving 3 different hypothetical moral judgments. As hypothesized, findings revealed significant multiplicity in the within-person operation of developmental stage structures, and intrapersonal variability in the degrees to which stages were used. Developmental patterns were found in terms of different distributions of multiple stages between high school and college samples, as well as the association between age and overall motivation scores. Differential relations of specific emotions to moral motivation revealed and confirmed the value of differentiating multiple emotions. Implications of the present theoretical perspective and the findings for understanding the complexity of moral judgment and motivation are discussed.

Limiting parental feedback disrupts vocal development in marmoset monkeys

PubMed Central

Gultekin, Yasemin B.; Hage, Steffen R.

2017-01-01

Vocalizations of human infants undergo dramatic changes across the first year by becoming increasingly mature and speech-like. Human vocal development is partially dependent on learning by imitation through social feedback between infants and caregivers. Recent studies revealed similar developmental processes being influenced by parental feedback in marmoset monkeys for apparently innate vocalizations. Marmosets produce infant-specific vocalizations that disappear after the first postnatal months. However, it is yet unclear whether parental feedback is an obligate requirement for proper vocal development. Using quantitative measures to compare call parameters and vocal sequence structure we show that, in contrast to normally raised marmosets, marmosets that were separated from parents after the third postnatal month still produced infant-specific vocal behaviour at subadult stages. These findings suggest a significant role of social feedback on primate vocal development until the subadult stages and further show that marmoset monkeys are a compelling model system for early human vocal development. PMID:28090084

Oncological Outcomes in Japanese Men Undergoing Orchiectomy for Stage I Testicular Germ Cell Tumor

PubMed Central

Harada, Ken-ichi; Miyake, Hideaki; Ogawa, Takayoshi; Inoue, Taka-aki; Fujisawa, Masato

2015-01-01

Background The objective of this study was to retrospectively review oncological outcomes in patients with stage I testicular germ cell tumor (GCT). Patients and Methods This study included 265 consecutive Japanese men undergoing orchiectomy for stage I testicular GCT, and a retrospective review of their records was performed. Results Of these 265 patients, 192 and 73 were pathologically classified with seminoma and nonseminoma, respectively. Prophylactic radiation and chemotherapy were performed in 62 patients with seminoma and 6 with nonseminoma, respectively. Disease recurrence occurred in 12 seminoma patients, of whom 11 had not received prophylactic radiation therapy; however, all 12 achieved a complete response to bleomycin, etoposide and cisplatin therapy. Of the nonseminoma patients, 19 experienced disease recurrence and were then treated with bleomycin, etoposide and cisplatin followed additionally by the surgical resection of residual tumors and salvage chemotherapy in 7 and 4, respectively. There was no cancer-specific death in the 265 patients, and 5-year recurrence-free survival rates in patients with seminoma and nonseminoma were 92.6 and 72.8%, respectively. Furthermore, following factors appeared to be significantly associated with recurrence-free survival in these patients: age, T classification, microvascular invasion and adjuvant therapy for those with seminoma, and microvascular invasion for those with nonseminoma. Conclusions Despite a generally favorable prognosis in Japanese men with stage I testicular GCT, intensive follow-up or prophylactic therapy should be considered for men with possible risk factors of disease recurrence. PMID:26889123

Perturbed thymopoiesis in vitro in the absence of Suppressor of Cytokine Signalling 1 and 3

PubMed Central

Croom, Hayley A.; Izon, David J.; Chong, Mark M.; Curtis, David J.; Roberts, Andrew W.; Kay, Thomas W.H.; Hilton, Douglas J.; Alexander, Warren S.; Starr, Robyn

2014-01-01

Cytokine signals are central to the differentiation of thymocytes and their stepwise progression through defined developmental stages. The intensity and duration of cytokine signals are regulated by the suppressor of cytokine signalling (SOCS) proteins. A clear role for SOCS1 during the later stages of thymopoiesis has been established, but little is known about its role during early thymopoiesis, nor the function of its closest relative, SOCS3. Here, we find that both SOCS1 and SOCS3 are expressed during early thymopoiesis, with expression coincident during the double negative (DN)2 and DN3 stages. We examined thymocyte differentiation in vitro by co-culture of SOCS-deficient bone marrow cells with OP9 cells expressing the Notch ligand Delta-like1 (OP9-DL1). Cells lacking SOCS1 were retarded at the DN3:DN4 transition and appeared unable to differentiate into double positive (DP) thymocytes. Cells lacking both SOCS1 and SOCS3 were more severely affected, and displayed an earlier block in T cell differentiation at DN2, the stage at which expression of SOCS1 and SOCS3 coincides. This indicates that, in addition to their specific roles, SOCS1 and SOCS3 share overlapping roles during thymopoiesis. This is the first demonstration of functional redundancy within the SOCS family, and has uncovered a vital role for SOCS1 and SOCS3 during two important checkpoints in early T cell development. PMID:18321577

Transcriptome analysis of functional differentiation between haploid and diploid cells of Emiliania huxleyi, a globally significant photosynthetic calcifying cell.

PubMed

von Dassow, Peter; Ogata, Hiroyuki; Probert, Ian; Wincker, Patrick; Da Silva, Corinne; Audic, Stéphane; Claverie, Jean-Michel; de Vargas, Colomban

2009-01-01

Eukaryotes are classified as either haplontic, diplontic, or haplo-diplontic, depending on which ploidy levels undergo mitotic cell division in the life cycle. Emiliania huxleyi is one of the most abundant phytoplankton species in the ocean, playing an important role in global carbon fluxes, and represents haptophytes, an enigmatic group of unicellular organisms that diverged early in eukaryotic evolution. This species is haplo-diplontic. Little is known about the haploid cells, but they have been hypothesized to allow persistence of the species between the yearly blooms of diploid cells. We sequenced over 38,000 expressed sequence tags from haploid and diploid E. huxleyi normalized cDNA libraries to identify genes involved in important processes specific to each life phase (2N calcification or 1N motility), and to better understand the haploid phase of this prominent haplo-diplontic organism. The haploid and diploid transcriptomes showed a dramatic differentiation, with approximately 20% greater transcriptome richness in diploid cells than in haploid cells and only

Transcriptome analysis of functional differentiation between haploid and diploid cells of Emiliania huxleyi, a globally significant photosynthetic calcifying cell

PubMed Central

2009-01-01

Background Eukaryotes are classified as either haplontic, diplontic, or haplo-diplontic, depending on which ploidy levels undergo mitotic cell division in the life cycle. Emiliania huxleyi is one of the most abundant phytoplankton species in the ocean, playing an important role in global carbon fluxes, and represents haptophytes, an enigmatic group of unicellular organisms that diverged early in eukaryotic evolution. This species is haplo-diplontic. Little is known about the haploid cells, but they have been hypothesized to allow persistence of the species between the yearly blooms of diploid cells. We sequenced over 38,000 expressed sequence tags from haploid and diploid E. huxleyi normalized cDNA libraries to identify genes involved in important processes specific to each life phase (2N calcification or 1N motility), and to better understand the haploid phase of this prominent haplo-diplontic organism. Results The haploid and diploid transcriptomes showed a dramatic differentiation, with approximately 20% greater transcriptome richness in diploid cells than in haploid cells and only ≤ 50% of transcripts estimated to be common between the two phases. The major functional category of transcripts differentiating haploids included signal transduction and motility genes. Diploid-specific transcripts included Ca2+, H+, and HCO3- pumps. Potential factors differentiating the transcriptomes included haploid-specific Myb transcription factor homologs and an unusual diploid-specific histone H4 homolog. Conclusions This study permitted the identification of genes likely involved in diploid-specific biomineralization, haploid-specific motility, and transcriptional control. Greater transcriptome richness in diploid cells suggests they may be more versatile for exploiting a diversity of rich environments whereas haploid cells are intrinsically more streamlined. PMID:19832986

Differential expression of myogenic regulatory genes and Msx-1 during dedifferentiation and redifferentiation of regenerating amphibian limbs.

PubMed

Simon, H G; Nelson, C; Goff, D; Laufer, E; Morgan, B A; Tabin, C

1995-01-01

An amputated limb of an adult urodele amphibian is capable of undergoing regeneration. The new structures form from an undifferentiated mass of cells called the regenerative blastema. The cells of the blastema are believed to derive from differentiated tissues of the adult limb. However, the exact source of these cells and the process by which they undergo dedifferentiation are poorly understood. In order to elucidate the molecular and cellular basis for dedifferentiation we isolated a number of genes which are potential regulators of the process. These include Msx-1, which is believed to support the undifferentiated and proliferative state of cells in the embryonic limb bud; and two members of the myogenic regulatory gene family, MRF-4 and Myf-5, which are expressed in differentiated muscle and regulate muscle-specific gene activity. As anticipated, we find that Msx-1 is strongly up-regulated during the initiation of regeneration. It remains expressed throughout regeneration but is not found in the fully regenerated limb. The myogenic gene MRF-4 has the reverse expression pattern. It is expressed in adult limb muscle, is rapidly shut off in early regenerative blastemas, and is only reexpressed at the completion of regeneration. These kinetics are paralleled by those of a muscle-specific Myosin gene. In contrast Myf-5, a second member of the myogenic gene family, continues to be expressed throughout the regenerative process. Thus, MRF-4 and Myf-5 are likely to play distinct roles during regeneration. MRF-4 may directly regulate muscle phenotype and as such its repression may be a key event in dedifferentiation.(ABSTRACT TRUNCATED AT 250 WORDS)

PET Imaging of Tau Pathology and Relationship to Amyloid, Longitudinal MRI, and Cognitive Change in Down Syndrome: Results from the Down Syndrome Biomarker Initiative (DSBI).

PubMed

Rafii, Michael S; Lukic, Ana S; Andrews, Randolph D; Brewer, James; Rissman, Robert A; Strother, Stephen C; Wernick, Miles N; Pennington, Craig; Mobley, William C; Ness, Seth; Matthews, Dawn C

2017-01-01

Adults with Down syndrome (DS) represent an enriched population for the development of Alzheimer's disease (AD), which could aid the study of therapeutic interventions, and in turn, could benefit from discoveries made in other AD populations. 1) Understand the relationship between tau pathology and age, amyloid deposition, neurodegeneration (MRI and FDG PET), and cognitive and functional performance; 2) detect and differentiate AD-specific changes from DS-specific brain changes in longitudinal MRI. Twelve non-demented adults, ages 30 to 60, with DS were enrolled in the Down Syndrome Biomarker Initiative (DSBI), a 3-year, observational, cohort study to demonstrate the feasibility of conducting AD intervention/prevention trials in adults with DS. We collected imaging data with 18F-AV-1451 tau PET, AV-45 amyloid PET, FDG PET, and volumetric MRI, as well as cognitive and functional measures and additional laboratory measures. All amyloid negative subjects imaged were tau-negative. Among the amyloid positive subjects, three had tau in regions associated with Braak stage VI, two at stage V, and one at stage II. Amyloid and tau burden correlated with age. The MRI analysis produced two distinct volumetric patterns. The first differentiated DS from normal (NL) and AD, did not correlate with age or amyloid, and was longitudinally stable. The second pattern reflected AD-like atrophy and differentiated NL from AD. Tau PET and MRI atrophy correlated with several cognitive and functional measures. Tau accumulation is associated with amyloid positivity and age, as well as with progressive neurodegeneration measurable using FDG and MRI. Tau correlates with cognitive decline, as do AD-specific hypometabolism and atrophy.

Postoperative Stiffness Requiring Manipulation Under Anesthesia Is Significantly Reduced After Simultaneous Versus Staged Bilateral Total Knee Arthroplasty.

PubMed

Meehan, John P; Monazzam, Shafagh; Miles, Troy; Danielsen, Beate; White, Richard H

2017-12-20

For patients with symptomatic bilateral knee arthritis, it is unknown whether the risk of developing stiffness requiring manipulation under anesthesia postoperatively is higher or lower for those undergoing simultaneous bilateral total knee arthroplasty (TKA) compared with those having staged bilateral TKA. Therefore, we undertook this study to evaluate the risk of requiring manipulation under anesthesia in staged versus simultaneous bilateral TKA as well as patients undergoing unilateral TKA. We utilized the California Patient Discharge Database, which is linked with the California Emergency Department, Ambulatory Surgery, and master death file databases. Using a literature-based estimate of the number of patients who failed to undergo the second stage of a staged bilateral TKA, replacement cases were randomly selected from patients who had unilateral TKA and were matched on 8 clinical characteristics of the patients who had staged bilateral TKA. Hierarchical multivariate logistic regression was performed to determine the risk-adjusted odds of manipulation in patients undergoing unilateral TKA, staged bilateral TKA, and simultaneous bilateral TKA using yearly hospital TKA volume as a random effect. Adjustment was made to allow fair comparison of the outcome at 90 and 180 days of follow-up after staged compared with simultaneous bilateral TKA. During the time period from 2005 through 2013, the cumulative incidence of manipulation within 90 days was 2.14% for unilateral TKA (4,398 events per 205,744 patients), 2.11% for staged bilateral TKA (724 events per 34,352 patients), and 1.62% for simultaneous bilateral TKA (195 events per 12,013 patients). At 180 days of complete follow-up, the cumulative incidence of manipulation was 3.07% after unilateral TKA (6,313 events per 205,649 patients), 2.89% after staged bilateral TKAs (957 events per 33,169 patients), and 2.29% after simultaneous bilateral TKA (267 events per 11,653 patients). With multivariate analyses used to adjust for relevant risk factors, the 90-day odds ratio (OR) of undergoing manipulation after simultaneous bilateral TKA was significantly lower than that for unilateral TKA (OR = 0.70; 95% confidence interval [CI], 0.57 to 0.86) and staged bilateral TKA (OR = 0.71; 95% CI, 0.57 to 0.90). Similarly, at 180 days, the odds of undergoing manipulation were significantly lower after simultaneous bilateral TKA than after both unilateral TKA (OR = 0.71; 95% CI, 0.59 to 0.84) and staged bilateral TKA (OR = 0.76; 95% CI, 0.63 to 0.93). The frequency of manipulation was significantly associated with younger age, fewer comorbidities, black race, and the absence of obesity. Although the ORs were small (close to 1), simultaneous bilateral TKA had a significantly decreased rate of stiffness requiring manipulation under anesthesia at 90 days and 180 days after knee replacement compared with that after staged bilateral TKA and unilateral TKA. Therapeutic Level III. See Instructions for Authors for a complete description of levels of evidence.

Derivation, propagation and differentiation of human embryonic stem cells.

PubMed

Conley, Brock J; Young, Julia C; Trounson, Alan O; Mollard, Richard

2004-04-01

Embryonic stem (ES) cells are in vitro cultivated pluripotent cells derived from the inner cell mass (ICM) of the embryonic blastocyst. Attesting to their pluripotency, ES cells can be differentiated into representative derivatives of all three embryonic germ layers (endoderm, ectoderm and mesoderm) both in vitro and in vivo. Although mouse ES cells have been studied for many years, human ES cells have only more recently been derived and successfully propagated. Many biochemical differences and culture requirements between mouse and human ES cells have been described, yet despite these differences the study of murine ES cells has provided important insights into methodologies aimed at generating a greater and more in depth understanding of human ES cell biology. One common feature of both mouse and human ES cells is their capacity to undergo controlled differentiation into spheroid structures termed embryoid bodies (EBs). EBs recapitulate several aspects of early development, displaying regional-specific differentiation programs into derivatives of all three embryonic germ layers. For this reason, EB formation has been utilised as an initial step in a wide range of studies aimed at differentiating both mouse and human ES cells into a specific and desired cell type. Recent reports utilising specific growth factor combinations and cell-cell induction systems have provided alternative strategies for the directed differentiation of cells into a desired lineage. According to each one of these strategies, however, a relatively high cell lineage heterogeneity remains, necessitating subsequent purification steps including mechanical dissection, selective media or fluorescent or magnetic activated cell sorting (FACS and MACS, respectively). In the future, the ability to specifically direct differentiation of human ES cells at 100% efficiency into a desired lineage will allow us to fully explore the potential of these cells in the analysis of early human development, drug discovery, drug testing and repair of damaged or diseased tissues via transplantation.

Identification of miRNAs during mouse postnatal ovarian development and superovulation.

PubMed

Khan, Hamid Ali; Zhao, Yi; Wang, Li; Li, Qian; Du, Yu-Ai; Dan, Yi; Huo, Li-Jun

2015-07-08

MicroRNAs are small noncoding RNAs that play critical roles in regulation of gene expression in wide array of tissues including the ovary through sequence complementarity at post-transcriptional level. Tight regulation of multitude of genes involved in ovarian development and folliculogenesis could be regulated at transcription level by these miRNAs. Therefore, tissue specific miRNAs identification is considered a key step towards understanding the role of miRNAs in biological processes. To investigate the role of microRNAs during ovarian development and folliculogenesis we sequenced eight different libraries using Illumina deep sequencing technology. Different developmental stages were selected to explore miRNAs expression pattern at different stages of gonadal maturation with/without treatment of PMSG/hCG for superovulation. From massive sequencing reads, clean reads of 16-26 bp were selected for further analysis of differential expression analysis and novel microRNA annotation. Expression analysis of all miRNAs at different developmental stages showed that some miRNAs were present ubiquitously while others were differentially expressed at different stages. Among differentially expressed miRNAs we reported 61 miRNAs with a fold change of more than 2 at different developmental stages among all libraries. Among the up-regulated miRNAs, mmu-mir-1298 had the highest fold change with 4.025 while mmu-mir-150 was down-regulated more than 3 fold. Furthermore, we found 2659 target genes for 20 differentially expressed microRNAs using seven different target predictions programs (DIANA-mT, miRanda, miRDB, miRWalk, RNAhybrid, PICTAR5, TargetScan). Analysis of the predicted targets showed certain ovary specific genes targeted by single or multiple microRNAs. Furthermore, pathway annotation and Gene ontology showed involvement of these microRNAs in basic cellular process. These results suggest the presence of different miRNAs at different stages of ovarian development and superovulation. Potential role of these microRNAs was elucidated using bioinformatics tools in regulation of different pathways, biological functions and cellular components underlying ovarian development and superovulation. These results provide a framework for extended analysis of miRNAs and their roles during ovarian development and superovulation. Furthermore, this study provides a base for characterization of individual miRNAs to discover their role in ovarian development and female fertility.

Epithelial–mesenchymal transition-associated secretory phenotype predicts survival in lung cancer patients

PubMed Central

Reka, Ajaya Kumar; Chen, Guoan; Keshamouni, Venkateshwar G.

2014-01-01

In cancer cells, the process of epithelial–mesenchymal transition (EMT) confers migratory and invasive capacity, resistance to apoptosis, drug resistance, evasion of host immune surveillance and tumor stem cell traits. Cells undergoing EMT may represent tumor cells with metastatic potential. Characterizing the EMT secretome may identify biomarkers to monitor EMT in tumor progression and provide a prognostic signature to predict patient survival. Utilizing a transforming growth factor-β-induced cell culture model of EMT, we quantitatively profiled differentially secreted proteins, by GeLC-tandem mass spectrometry. Integrating with the corresponding transcriptome, we derived an EMT-associated secretory phenotype (EASP) comprising of proteins that were differentially upregulated both at protein and mRNA levels. Four independent primary tumor-derived gene expression data sets of lung cancers were used for survival analysis by the random survival forests (RSF) method. Analysis of 97-gene EASP expression in human lung adenocarcinoma tumors revealed strong positive correlations with lymph node metastasis, advanced tumor stage and histological grade. RSF analysis built on a training set (n = 442), including age, sex and stage as variables, stratified three independent lung cancer data sets into low-, medium- and high-risk groups with significant differences in overall survival. We further refined EASP to a 20 gene signature (rEASP) based on variable importance scores from RSF analysis. Similar to EASP, rEASP predicted survival of both adenocarcinoma and squamous carcinoma patients. More importantly, it predicted survival in the early-stage cancers. These results demonstrate that integrative analysis of the critical biological process of EMT provides mechanism-based and clinically relevant biomarkers with significant prognostic value. PMID:24510113

Shape-Shifted Red Blood Cells: A Novel Red Blood Cell Stage?

PubMed Central

Chico, Verónica; Puente-Marin, Sara; Ciordia, Sergio; Mena, María Carmen; Carracedo, Begoña; Mercado, Luis; Coll, Julio

2018-01-01

Primitive nucleated erythroid cells in the bloodstream have long been suggested to be more similar to nucleated red cells of fish, amphibians, and birds than the red cells of fetal and adult mammals. Rainbow trout Ficoll-purified red blood cells (RBCs) cultured in vitro undergo morphological changes, especially when exposed to stress, and enter a new cell stage that we have coined shape-shifted RBCs (shRBCs). We have characterized these shRBCs using transmission electron microscopy (TEM) micrographs, Wright–Giemsa staining, cell marker immunostaining, and transcriptomic and proteomic evaluation. shRBCs showed reduced density of the cytoplasm, hemoglobin loss, decondensed chromatin in the nucleus, and striking expression of the B lymphocyte molecular marker IgM. In addition, shRBCs shared some features of mammalian primitive pyrenocytes (extruded nucleus surrounded by a thin rim of cytoplasm and phosphatidylserine (PS) exposure on cell surface). These shRBCs were transiently observed in heat-stressed rainbow trout bloodstream for three days. Functional network analysis of combined transcriptomic and proteomic studies resulted in the identification of proteins involved in pathways related to the regulation of cell morphogenesis involved in differentiation, cellular response to stress, and immune system process. In addition, shRBCs increased interleukin 8 (IL8), interleukin 1 β (IL1β), interferon ɣ (IFNɣ), and natural killer enhancing factor (NKEF) protein production in response to viral hemorrhagic septicemia virus (VHSV). In conclusion, shRBCs may represent a novel cell stage that participates in roles related to immune response mediation, homeostasis, and the differentiation and development of blood cells. PMID:29671811

Shape-Shifted Red Blood Cells: A Novel Red Blood Cell Stage?

PubMed

Chico, Verónica; Puente-Marin, Sara; Nombela, Iván; Ciordia, Sergio; Mena, María Carmen; Carracedo, Begoña; Villena, Alberto; Mercado, Luis; Coll, Julio; Ortega-Villaizan, María Del Mar

2018-04-19

Primitive nucleated erythroid cells in the bloodstream have long been suggested to be more similar to nucleated red cells of fish, amphibians, and birds than the red cells of fetal and adult mammals. Rainbow trout Ficoll-purified red blood cells (RBCs) cultured in vitro undergo morphological changes, especially when exposed to stress, and enter a new cell stage that we have coined shape-shifted RBCs (shRBCs). We have characterized these shRBCs using transmission electron microscopy (TEM) micrographs, Wright⁻Giemsa staining, cell marker immunostaining, and transcriptomic and proteomic evaluation. shRBCs showed reduced density of the cytoplasm, hemoglobin loss, decondensed chromatin in the nucleus, and striking expression of the B lymphocyte molecular marker IgM. In addition, shRBCs shared some features of mammalian primitive pyrenocytes (extruded nucleus surrounded by a thin rim of cytoplasm and phosphatidylserine (PS) exposure on cell surface). These shRBCs were transiently observed in heat-stressed rainbow trout bloodstream for three days. Functional network analysis of combined transcriptomic and proteomic studies resulted in the identification of proteins involved in pathways related to the regulation of cell morphogenesis involved in differentiation, cellular response to stress, and immune system process. In addition, shRBCs increased interleukin 8 (IL8), interleukin 1 β (IL1β), interferon ɣ (IFNɣ), and natural killer enhancing factor (NKEF) protein production in response to viral hemorrhagic septicemia virus (VHSV). In conclusion, shRBCs may represent a novel cell stage that participates in roles related to immune response mediation, homeostasis, and the differentiation and development of blood cells.

Three primary synchronous malignancies of the uterus, cervix, and fallopian tube: A case report.

PubMed

Song, Liang; Li, Qingli; Yang, Kaixuan; Yin, Rutie; Wang, Danqing

2018-06-01

Multiple primary malignancies can occur in the same organ or in multiple organs or systems. Likewise, they can occur simultaneously or successively. Based on the timing of the diagnosis, they are classified as multiple synchronous (i.e., concurrent) or metachronous (i.e., successive) primary malignancies. The vast majority of patients have multiple metachronous malignant tumors; multiple synchronous tumors are rare. A 63-year-old woman presented with the chief complaint of vaginal fluid discharge for 3 months and abdominal pain for 1 month. The patient was diagnosed with multiple synchronous primary malignancies: 1) endometrial poorly differentiated serous adenocarcinoma, stage IV; 2) poorly differentiated squamous cell carcinoma of the cervix, stage IB1; and 3) left-sided fallopian tube carcinoma in situ. After total abdominal hysterectomy, bilateral salpingo-oophorectomy, and comprehensive staging and debulking, the patient was administered eight courses of adjuvant chemotherapy (taxane carboplatin/taxane cisplatin). After chemotherapy completion, the patient has been undergoing regular follow-up examinations; no recurrence has been noted at 18 months. It is important to distinguish between multiple synchronous primary malignancies and metastasis of a primary tumor to select the appropriate treatment regimen and to adequately assess the patient's prognosis. When a cancer patient shows clinical manifestations of another tumor, not only metastasis but also the possibility of multiple synchronous primary malignant tumors should be considered. The duration of follow-up in patients with malignant tumors should be extended as much as possible, as the timely detection and treatment of other primary malignant tumors can prolong survival and improve the quality of life.

STAT signaling in mammary gland differentiation, cell survival and tumorigenesis

PubMed Central

Haricharan, S; Li, Y

2013-01-01

The mammary gland is a unique organ that undergoes extensive and profound changes during puberty, menstruation, pregnancy, lactation and involution. The changes that take place during puberty involve large-scale proliferation and invasion of the fat-pad. During pregnancy and lactation, the mammary cells are exposed to signaling pathways that inhibit apoptosis, induce proliferation and invoke terminal differentiation. Finally, during involution the mammary gland is exposed to milk stasis, programed cell death and stromal reorganization to clear the differentiated milk-producing cells. Not surprisingly, the signaling pathways responsible for bringing about these changes in breast cells are often subverted during the process of tumorigenesis. The STAT family of proteins is involved in every stage of mammary gland development, and is also frequently implicated in breast tumorigenesis. While the roles of STAT3 and STAT5 during mammary gland development and tumorigenesis are well studied, others members, e.g. STAT1 and STAT6, have only recently been observed to play a role in mammary gland biology. Continued investigation into the STAT protein network in the mammary gland will likely yield new biomarkers and risk factors for breast cancer, and may also lead to novel prophylactic or therapeutic strategies against breast cancer. PMID:23541951

Single-cell mechanical phenotype is an intrinsic marker of reprogramming and differentiation along the mouse neural lineage.

PubMed

Urbanska, Marta; Winzi, Maria; Neumann, Katrin; Abuhattum, Shada; Rosendahl, Philipp; Müller, Paul; Taubenberger, Anna; Anastassiadis, Konstantinos; Guck, Jochen

2017-12-01

Cellular reprogramming is a dedifferentiation process during which cells continuously undergo phenotypical remodeling. Although the genetic and biochemical details of this remodeling are fairly well understood, little is known about the change in cell mechanical properties during the process. In this study, we investigated changes in the mechanical phenotype of murine fetal neural progenitor cells (fNPCs) during reprogramming to induced pluripotent stem cells (iPSCs). We find that fNPCs become progressively stiffer en route to pluripotency, and that this stiffening is mirrored by iPSCs becoming more compliant during differentiation towards the neural lineage. Furthermore, we show that the mechanical phenotype of iPSCs is comparable with that of embryonic stem cells. These results suggest that mechanical properties of cells are inherent to their developmental stage. They also reveal that pluripotent cells can differentiate towards a more compliant phenotype, which challenges the view that pluripotent stem cells are less stiff than any cells more advanced developmentally. Finally, our study indicates that the cell mechanical phenotype might be utilized as an inherent biophysical marker of pluripotent stem cells. © 2017. Published by The Company of Biologists Ltd.

Knock-in strategy at 3'-end of Crx gene by CRISPR/Cas9 system shows the gene expression profiles during human photoreceptor differentiation.

PubMed

Homma, Kohei; Usui, Sumiko; Kaneda, Makoto

2017-03-01

Fluorescent reporter gene knock-in induced pluripotent stem cell (iPSC) lines have been used to evaluate the efficiency of differentiation into specific cell lineages. Here, we report a knock-in strategy for the generation of human iPSC reporter lines in which a 2A peptide sequence and a red fluorescent protein (E2-Crimson) gene were inserted at the termination codon of the cone-rod homeobox (Crx) gene, a photoreceptor-specific transcriptional factor gene. The knock-in iPSC lines were differentiated into fluorescence-expressing cells in 3D retinal differentiation culture, and the fluorescent cells also expressed Crx specifically in the nucleus. We found that the fluorescence intensity was positively correlated with the expression levels of Crx mRNA and that fluorescent cells expressed rod photoreceptor-specific genes in the later stage of differentiation. Finally, we treated the fluorescent cells with DAPT, a Notch inhibitor, and found that DAPT-enhanced retinal differentiation was associated with up-regulation of Crx, Otx2 and NeuroD1, and down-regulation of Hes5 and Ngn2. These suggest that this knock-in strategy at the 3'-end of the target gene, combined with the 2A peptide linked to fluorescent proteins, offers a useful tool for labeling specific cell lineages or monitoring expression of any marker genes without affecting the function of the target gene. © 2017 Molecular Biology Society of Japan and John Wiley & Sons Australia, Ltd.

Was Miyakejima undergoing subsidence before the 2000 caldera collapse? JERS1 InSAR results: 1992-1998

NASA Astrophysics Data System (ADS)

Furuya, M.

2003-12-01

Miyakejima volcano is a basaltic strato volcano island on the eastern edge of the Philippine Sea Plate, and was undergoing a number of eruption activities over the past centuries. In July-August 2000, the Miyakejima volcano underwent a caldera collapse, prompting many modern geodetic and geophysical measurements (e.g., Geshi et al. 2002; Furuya et al. 2003). The observation results on the pre-caldera-collapse stages are, however, limitted. Were there any precursory secular subsidence before the collapse? Though Miyazaki (1990) reported a secular subsidence at the Miyakejima, using leveling technique, there are no documented reports, to my knowledge, which employed radar interferometry to examine the ground displacements at Miyakejima. Here I will report on the results derived from the radar interferometry at Miyakejima volcano. I chose JERS-1 data (L-band HH) for the analysis, so that I could get rid of the loss of coherence; most of the Miyakejima is covered with vegetation. To remove the topographic fringes as well as to re-estimate the spatial baseline data (Rosen et al. 1996), I employed 10-meter resolution digital elevation map derived by Geographical Survey Institute, Japan. I could generate 24 differential interferograms at the time of writing this text. However, I do not yet recognize any significant "signals" that can be discriminated with the atmospheric "noise". There appears to be no specific subsidence pattern, which are detected in a number of other volcanos in the world (e.g., Lu et al. 2002; Yarai et al. 2002; Okuyama et al. 2002). I am going to show a stacked interferogram like that in Fujiwara et al. (1998) and to examine the existence of volcanic signals.

Testicular involution prior to sex change in gilthead seabream is characterized by a decrease in DMRT1 gene expression and by massive leukocyte infiltration

PubMed Central

Liarte, Sergio; Chaves-Pozo, Elena; García-Alcazar, Alicia; Mulero, Victoriano; Meseguer, José; García-Ayala, Alfonsa

2007-01-01

Background Leukocytes are found within the testis of most, if not all, mammals and are involved in immunological surveillance, physiological regulation and tissue remodelling. The testis of seasonal breeding fish undergoes a regression process. In the present study, the second reproductive cycle (RC) of the protandrous seasonal teleost fish, gilthead seabream, was investigated and the presence of leukocytes analysed. Special attention has been paid to the testicular degenerative process which is particularly active in the last stage of the second RC probably due to the immediacy of the sex change process. Methods Sexually mature specimens (n = 10–18 fish/month) were sampled during the second RC. Some specimens were intraperitoneally injected with bromodeoxyuridin (BrdU) before sampling. Light and electron microscopy was used to determine the different stages of gonadal development and the presence of leukocytes and PCR was used to analyse the gene expression of a testis-differentiating gene and of specific markers for macrophages and B and T lymphocytes. Immunocytochemistry and flow cytometry were performed using a specific antibody against acidophilic granulocytes from the gilthead seabream. Cell proliferation was detected by immunocytochemistry using an anti-BrdU antibody and apoptotic cells by in situ detection of DNA fragmentation. Results The fish in the western Mediterranean area developed as males during the first two RCs. The testis of all the specimens during the second RC underwent a degenerative process, which started at post-spawning and was enhanced during the testicular involution stage, when vitellogenic oocytes appeared in the ovary accompanied by a progressive increase in the ovarian index. However, only 40% of specimens were females in the third RC. Leukocytes (acidophilic granulocytes, macrophages and lymphocytes) were present in the gonad and acidophilic granulocyte infiltration occurred during the last two stages. At the same time DMRT1 gene expression decreased. Conclusions The results demonstrate that innate and adaptive immune cells are present in the gonads of gilthead seabream. Moreover, the whole fish population underwent a testicular degenerative process prior to sex change, characterized by high rates of apoptosis and necrosis and accompanied by an infiltration of acidophilic granulocytes and a decrease in DMRT1 levels. PMID:17547755

Functional cartilage repair capacity of de-differentiated, chondrocyte- and mesenchymal stem cell-laden hydrogels in vitro

PubMed Central

Rackwitz, Lars; Djouad, Farida; Janjanin, Sasa; Nöth, Ulrich; Tuan, Rocky S.

2017-01-01

Objective The long-term performance of cell seeded matrix based cartilage constructs depends on (1) the development of sufficient biomechanical properties, and (2) lateral integration with host tissues, both of which require cartilage specific matrix deposition within the scaffold. In this study, we have examined the potential of tissue-engineered cartilage analogs developed using different cell types, i.e., MSCs versus chondrocytes and de-differentiated chondrocytes, in an established “construct in cartilage ring” model. Design Cell-laden constructs of differentiated chondrocytes, de-differentiated chondrocytes after 2, 5 or 8 population doublings, and MSCs were either implanted into a native cartilage ring immediately after fabrication (immature group) or pretreated for 21 days in a transforming growth factor-β3 (TGF-β3) containing medium prior to implantation. After additional culture for 28 days in a serum-free, chemically defined medium, the extent of lateral integration, and biochemical and biomechanical characteristics of the implants as hybrid constructs were assessed. Results The quality of integration, the amount of accumulated cartilage-specific matrix components and associated biomechanical properties were found to be highest when using differentiated chondrocytes. De-differentiation of chondrocytes negatively impacted the properties of the implants, as even two population doublings of the chondrocytes in culture significantly lowered cartilage repair capacity. In contrast, MSCs showed chondrogenic differentiation with TGF-β3 pre-treatment and superior integrational behavior. Conclusions Chondrocyte expansion and de-differentiation impaired the cell response, resulting in inferior cartilage repair in vitro. With TGF-β3 pre-treatment, MSCs were able to undergo sustained chondrogenic differentiation and exhibited superior matrix deposition and integration compared to de-differentiated chondrocytes. PMID:24887551

Karyopherin alpha 1 regulates satellite cell proliferation and survival by modulating nuclear import

PubMed Central

Choo, Hyo-Jung; Cutler, Alicia; Rother, Franziska; Bader, Michael; Pavlath, Grace K.

2016-01-01

Satellite cells are stem cells with an essential role in skeletal muscle repair. Precise regulation of gene expression is critical for proper satellite cell quiescence, proliferation, differentiation and self -renewal. Nuclear proteins required for gene expression are dependent on the nucleocytoplasmic transport machinery to access to nucleus, however little is known about regulation of nuclear transport in satellite cells. The best characterized nuclear import pathway is classical nuclear import which depends on a classical nuclear localization signal (cNLS) in a cargo protein and the heterodimeric import receptors, karyopherin alpha (KPNA) and beta (KPNB). Multiple KPNA1 paralogs exist and can differ in importing specific cNLS proteins required for cell differentiation and function. We show that transcripts for six Kpna paralogs underwent distinct changes in mouse satellite cells during muscle regeneration accompanied by changes in cNLS proteins in nuclei. Depletion of KPNA1, the most dramatically altered KPNA, caused satellite cells in uninjured muscle to prematurely activate, proliferate and undergo apoptosis leading to satellite cell exhaustion with age. Increased proliferation of satellite cells led to enhanced muscle regeneration at early stages of regeneration. In addition, we observed impaired nuclear localization of two key KPNA1 cargo proteins: p27, a cyclin-dependent kinase inhibitor associated with cell cycle control and lymphoid enhancer factor 1, a critical co-transcription factor for β-catenin. These results indicate that regulated nuclear import of proteins by KPNA1 is critical for satellite cell proliferation and survival and establish classical nuclear import as a novel regulatory mechanism for controlling satellite cell fate. PMID:27434733

Fibroblast growth factor signaling is required for early somatic gonad development in zebrafish.

PubMed

Leerberg, Dena M; Sano, Kaori; Draper, Bruce W

2017-09-01

The vertebrate ovary and testis develop from a sexually indifferent gonad. During early development of the organism, primordial germ cells (the gamete lineage) and somatic gonad cells coalesce and begin to undergo growth and morphogenesis to form this bipotential gonad. Although this aspect of development is requisite for a fertile adult, little is known about the genetic regulation of early gonadogenesis in any vertebrate. Here, we provide evidence that fibroblast growth factor (Fgf) signaling is required for the early growth phase of a vertebrate bipotential gonad. Based on mutational analysis in zebrafish, we show that the Fgf ligand 24 (Fgf24) is required for proliferation, differentiation, and morphogenesis of the early somatic gonad, and as a result, most fgf24 mutants are sterile as adults. Additionally, we describe the ultrastructural elements of the early zebrafish gonad and show that distinct somatic cell populations can be identified soon after the gonad forms. Specifically, we show that fgf24 is expressed in an epithelial population of early somatic gonad cells that surrounds an inner population of mesenchymal somatic gonad cells that are in direct contact with the germ cells, and that fgf24 is required for stratification of the somatic tissue. Furthermore, based on gene expression analysis, we find that differentiation of the inner mesenchymal somatic gonad cells into functional cell types in the larval and early juvenile-stage gonad is dependent on Fgf24 signaling. Finally, we argue that the role of Fgf24 in zebrafish is functionally analogous to the role of tetrapod FGF9 in early gonad development.

Pax-3 expression in segmental mesoderm marks early stages in myogenic cell specification.

PubMed

Williams, B A; Ordahl, C P

1994-04-01

Specification of the myogenic lineage begins prior to gastrulation and culminates in the emergence of determined myogenic precursor cells from the somites. The myoD family (MDF) of transcriptional activators controls late step(s) in myogenic specification that are closely followed by terminal muscle differentiation. Genes expressed in myogenic specification at stages earlier than MDFs are unknown. The Pax-3 gene is expressed in all the cells of the caudal segmental plate, the early mesoderm compartment that contains the precursors of skeletal muscle. As somites form from the segmental plate and mature, Pax-3 expression is progressively modulated. Beginning at the time of segmentation, Pax-3 becomes repressed in the ventral half of the somite, leaving Pax-3 expression only in the dermomyotome. Subsequently, differential modulation of Pax-3 expression levels delineates the medial and lateral halves of the dermomyotome, which contain precursors of axial (back) muscle and limb muscle, respectively. Pax-3 expression is then repressed as dermomyotome-derived cells activate MDFs. Quail-chick chimera and ablation experiments confirmed that the migratory precursors of limb muscle continue to express Pax-3 during migration. Since limb muscle precursors do not activate MDFs until 2 days after they leave the somite, Pax-3 represents the first molecular marker for this migratory cell population. A null mutation of the mouse Pax-3 gene, Splotch, produces major disruptions in early limb muscle development (Franz, T., Kothary, R., Surani, M. A. H., Halata, Z. and Grim, M. (1993) Anat. Embryol. 187, 153-160; Goulding, M., Lumsden, A. and Paquette, A. (1994) Development 120, 957-971). We conclude, therefore, that Pax-3 gene expression in the paraxial mesoderm marks earlier stages in myogenic specification than MDFs and plays a crucial role in the specification and/or migration of limb myogenic precursors.

Antithrombin III is associated with acute liver failure in patients with end-stage heart failure undergoing mechanical circulatory support.

PubMed

Hoefer, Judith; Ulmer, Hanno; Kilo, Juliane; Margreiter, Raimund; Grimm, Michael; Mair, Peter; Ruttmann, Elfriede

2017-06-01

There are few data on the role of liver dysfunction in patients with end-stage heart failure supported by mechanical circulatory support. The aim of our study was to investigate predictors for acute liver failure in patients with end-stage heart failure undergoing mechanical circulatory support. A consecutive 164 patients with heart failure with New York Heart Association class IV undergoing mechanical circulatory support were investigated for acute liver failure using the King's College criteria. Clinical characteristics of heart failure together with hemodynamic and laboratory values were analyzed by logistic regression. A total of 45 patients (27.4%) with heart failure developed subsequent acute liver failure with a hospital mortality of 88.9%. Duration of heart failure, cause, cardiopulmonary resuscitation, use of vasopressors, central venous pressure, pulmonary capillary wedge pressure, pulmonary pulsatility index, cardiac index, and transaminases were not significantly associated with acute liver failure. Repeated decompensation, atrial fibrillation (P < .001) and the use of inotropes (P = .007), mean arterial (P = .005) and pulmonary pressures (P = .042), cholinesterase, international normalized ratio, bilirubin, lactate, and pH (P < .001) were predictive of acute liver failure in univariate analysis only. In multivariable analysis, decreased antithrombin III was the strongest single measurement indicating acute liver failure (relative risk per %, 0.84; 95% confidence interval, 0.77-0.93; P = .001) and remained an independent predictor when adjustment for the Model for End-Stage Liver Disease score was performed (relative risk per %, 0.89; 95% confidence interval, 0.80-0.99; P = .031). Antithrombin III less than 59.5% was identified as a cutoff value to predict acute liver failure with a corresponding sensitivity of 81% and specificity of 87%. In addition to the Model for End-Stage Liver Disease score, decreased antithrombin III activity tends to be superior in predicting acute liver failure compared with traditionally thought predictors. Antithrombin III measurement may help to identify patients more precisely who are developing acute liver failure during mechanical circulatory support. Copyright © 2017 The American Association for Thoracic Surgery. Published by Elsevier Inc. All rights reserved.

Silver nanoparticles induce developmental stage-specific embryonic phenotypes in zebrafish

NASA Astrophysics Data System (ADS)

Lee, Kerry J.; Browning, Lauren M.; Nallathamby, Prakash D.; Osgood, Christopher J.; Xu, Xiao-Hong Nancy

2013-11-01

Much is anticipated from the development and deployment of nanomaterials in biological organisms, but concerns remain regarding their biocompatibility and target specificity. Here we report our study of the transport, biocompatibility and toxicity of purified and stable silver nanoparticles (Ag NPs, 13.1 +/- 2.5 nm in diameter) upon the specific developmental stages of zebrafish embryos using single NP plasmonic spectroscopy. We find that single Ag NPs passively diffuse into five different developmental stages of embryos (cleavage, early-gastrula, early-segmentation, late-segmentation, and hatching stages), showing stage-independent diffusion modes and diffusion coefficients. Notably, the Ag NPs induce distinctive stage and dose-dependent phenotypes and nanotoxicity, upon their acute exposure to the Ag NPs (0-0.7 nM) for only 2 h. The late-segmentation embryos are most sensitive to the NPs with the lowest critical concentration (CNP,c << 0.02 nM) and highest percentages of cardiac abnormalities, followed by early-segmentation embryos (CNP,c < 0.02 nM), suggesting that disruption of cell differentiation by the NPs causes the most toxic effects on embryonic development. The cleavage-stage embryos treated with the NPs develop into a wide variety of phenotypes (abnormal finfold, tail/spinal cord flexure, cardiac malformation/edema, yolk sac edema, and acephaly). These organ structures are not yet developed in cleavage-stage embryos, suggesting that the earliest determinative events to create these structures are ongoing, and disrupted by NPs, which leads to the downstream effects. In contrast, the hatching embryos are most resistant to the Ag NPs, and majority of embryos (94%) develop normally, and none of them develop abnormally. Interestingly, early-gastrula embryos are less sensitive to the NPs than cleavage and segmentation stage embryos, and do not develop abnormally. These important findings suggest that the Ag NPs are not simple poisons, and they can target specific pathways in development, and potentially enable target specific study and therapy for early embryonic development.Much is anticipated from the development and deployment of nanomaterials in biological organisms, but concerns remain regarding their biocompatibility and target specificity. Here we report our study of the transport, biocompatibility and toxicity of purified and stable silver nanoparticles (Ag NPs, 13.1 +/- 2.5 nm in diameter) upon the specific developmental stages of zebrafish embryos using single NP plasmonic spectroscopy. We find that single Ag NPs passively diffuse into five different developmental stages of embryos (cleavage, early-gastrula, early-segmentation, late-segmentation, and hatching stages), showing stage-independent diffusion modes and diffusion coefficients. Notably, the Ag NPs induce distinctive stage and dose-dependent phenotypes and nanotoxicity, upon their acute exposure to the Ag NPs (0-0.7 nM) for only 2 h. The late-segmentation embryos are most sensitive to the NPs with the lowest critical concentration (CNP,c << 0.02 nM) and highest percentages of cardiac abnormalities, followed by early-segmentation embryos (CNP,c < 0.02 nM), suggesting that disruption of cell differentiation by the NPs causes the most toxic effects on embryonic development. The cleavage-stage embryos treated with the NPs develop into a wide variety of phenotypes (abnormal finfold, tail/spinal cord flexure, cardiac malformation/edema, yolk sac edema, and acephaly). These organ structures are not yet developed in cleavage-stage embryos, suggesting that the earliest determinative events to create these structures are ongoing, and disrupted by NPs, which leads to the downstream effects. In contrast, the hatching embryos are most resistant to the Ag NPs, and majority of embryos (94%) develop normally, and none of them develop abnormally. Interestingly, early-gastrula embryos are less sensitive to the NPs than cleavage and segmentation stage embryos, and do not develop abnormally. These important findings suggest that the Ag NPs are not simple poisons, and they can target specific pathways in development, and potentially enable target specific study and therapy for early embryonic development. Electronic supplementary information (ESI) available. See DOI: 10.1039/c3nr03210h

Defining a Model for Mitochondrial Function in mESC Differentiation

EPA Science Inventory

Defining a Model for Mitochondrial Function in mESC DifferentiationDefining a Model for Mitochondrial Function in mESC Differentiation Differentiating embryonic stem cells (ESCs) undergo mitochondrial maturation leading to a switch from a system dependent upon glycolysis to a re...

Deep analysis of wild Vitis flower transcriptome reveals unexplored genome regions associated with sex specification.

PubMed

Ramos, Miguel Jesus Nunes; Coito, João Lucas; Fino, Joana; Cunha, Jorge; Silva, Helena; de Almeida, Patrícia Gomes; Costa, Maria Manuela Ribeiro; Amâncio, Sara; Paulo, Octávio S; Rocheta, Margarida

2017-01-01

RNA-seq of Vitis during early stages of bud development, in male, female and hermaphrodite flowers, identified new loci outside of annotated gene models, suggesting their involvement in sex establishment. The molecular mechanisms responsible for flower sex specification remain unclear for most plant species. In the case of V. vinifera ssp. vinifera, it is not fully understood what determines hermaphroditism in the domesticated subspecies and male or female flowers in wild dioecious relatives (Vitis vinifera ssp. sylvestris). Here, we describe a de novo assembly of the transcriptome of three flower developmental stages from the three Vitis vinifera flower types. The validation of de novo assembly showed a correlation of 0.825. The main goals of this work were the identification of V. v. sylvestris exclusive transcripts and the characterization of differential gene expression during flower development. RNA from several flower developmental stages was used previously to generate Illumina sequence reads. Through a sequential de novo assembly strategy one comprehensive transcriptome comprising 95,516 non-redundant transcripts was assembled. From this dataset 81,064 transcripts were annotated to V. v. vinifera reference transcriptome and 11,084 were annotated against V. v. vinifera reference genome. Moreover, we found 3368 transcripts that could not be mapped to Vitis reference genome. From all the non-redundant transcripts that were assembled, bioinformatics analysis identified 133 specific of V. v. sylvestris and 516 transcripts differentially expressed among the three flower types. The detection of transcription from areas of the genome not currently annotated suggests active transcription of previously unannotated genomic loci during early stages of bud development.

Genetic Mutations in Blood and Tissue Samples in Predicting Response to Treatment in Patients With Locally Advanced Rectal Cancer Undergoing Chemoradiation

ClinicalTrials.gov

2017-09-08

Mucinous Adenocarcinoma of the Rectum; Recurrent Rectal Cancer; Signet Ring Adenocarcinoma of the Rectum; Stage IIA Rectal Cancer; Stage IIB Rectal Cancer; Stage IIC Rectal Cancer; Stage IIIA Rectal Cancer; Stage IIIB Rectal Cancer; Stage IIIC Rectal Cancer

Genome-wide association study in East Asians suggests UHMK1 as a novel bone mineral density susceptibility gene.

PubMed

Choi, Hyung Jin; Park, Hyojung; Zhang, Lei; Kim, Jung Hee; Kim, Ye An; Yang, Jae-Yeon; Pei, Yu-Fang; Tian, Qing; Shen, Hui; Hwang, Joo-Yeon; Deng, Hong-Wen; Cho, Nam H; Shin, Chan Soo

2016-10-01

To identify genetic variants that influence bone mineral density (BMD) in East Asians, we performed a quantitative trait analysis of lumbar spine, total hip and femoral neck BMD in a Korean population-based cohort (N=2729) and follow-up replication analysis in a Chinese Han population and two Caucasian populations (N=1547, 2250 and 987, respectively). From the meta-analysis of the stage 1 discovery analysis and stage 2 replication analysis, we identified four BMD loci that reached near-genome-wide significance level (P<5×10(-7)). One locus on 1q23 (UHMK1, rs16863247, P=4.1×10(-7) for femoral neck BMD and P=3.2×10(-6) for total hip BMD) was a novel BMD signal. Interestingly, rs16863247 was very rare in Caucasians (minor allele frequency<0.01), indicating that this association could be specific to East Asians. In gender specific analysis, rs1160574 on 1q32 (KCNH1) was associated with femoral neck BMD (P=2.1×10(-7)) in female subjects. rs9371538 in the known BMD region on 6q25 ESR1 was associated with lumbar spine BMD (P=5.6×10(-9)). rs7776725 in the known BMD region on 7q31 WTN16 was associated with total hip BMD (P=8.6×10(-9)). In osteoblasts, endogenous UHMK1 expression was increased during differentiation and UHMK1 knockdown decreased its differentiation, while UHMK1 overexpression increased its differentiation. In osteoclasts, endogenous UHMK1 expression was decreased during differentiation and UHMK1 knockdown increased its differentiation, while UHMK1 overexpression decreased its differentiation. In conclusion, our genome-wide association study identified the UHMK1 gene as a novel BMD locus specific to East Asians. Functional studies suggest a role of UHMK1 on regulation of osteoblasts and osteoclasts. Copyright © 2016 Elsevier Inc. All rights reserved.

Osteograft, plastic material for regenerative medicine

NASA Astrophysics Data System (ADS)

Zaidman, A. M.; Korel, A. V.; Shevchenko, A. I.; Shchelkunova, E. I.; Sherman, K. M.; Predein, Yu. A.; Kosareva, O. S.

2016-08-01

Creating tissue-engineering constructs based on the mechanism of cartilage-bone evolution is promising for traumatology and orthopedics. Such a graft was obtained from a chondrograft by transdifferentiation. The hondrograft placed in osteogenic medium is undergoing osteogenic differentiation for 14-30 days. Tissue specificity of the osteograft was studied by morphology, immunohistochemistry, electron microscopy, and the expression of the corresponding genes was estimated. The expression of osteonectin, fibronectin, collagen of type I, izolektin and CD 44 is determined. Alkaline phosphatase and matrix vesicles are determined in osteoblasts. Calcificates are observed in the matrix. Chondrogenic proteins expression is absent. These findings evidence the tissue specificity of the developed osteograft.

Lead exposure delays the differentiation of oligodendroglial progenitors in vitro.

PubMed

Deng, W; McKinnon, R D; Poretz, R D

2001-08-01

Lead (Pb) is an environmental neurotoxicant that can cause hypo- and demyelination. Oligodendrocytes (OLs), the myelin-forming cells in the central nervous system, may be a possible target for Pb toxicity. The present study describes the effect of Pb on the maturation of rat OL progenitor (OP) cells and the developmental expression of myelin-specific galactolipids. Dose-response studies showed that OP cultures were more sensitive to Pb than mature OLs. Pb delayed the differentiation of OL progenitors, as demonstrated by cell morphology and immunostaining with a panel of stage-specific differentiation markers. Pb given prior to and during differentiation caused a decrease in the biosynthesis of galactolipids in both undifferentiated and differentiated OLs, as detected by metabolic radiolabeling with 3H-D-galactose. While the ratios of galacto/gluco-cerebrosides, hydroxy fatty acid/nonhydroxy fatty acid galactolipids, and galactocerebrosides/sulfatides increased in control cultures during cell differentiation, Pb treatment prevented these changes. The results suggest that chronic Pb exposure may impact brain development by interfering with the timely developmental maturation of OL progenitors. Copyright 2001 Academic Press.

Effect of activated autologous platelet-rich plasma on proliferation and osteogenic differentiation of human adipose-derived stem cells in vitro

PubMed Central

Xu, Fang-Tian; Li, Hong-Mian; Yin, Qing-Shui; Liang, Zhi-Jie; Huang, Min-Hong; Chi, Guang-Yi; Huang, Lu; Liu, Da-Lie; Nan, Hua

2015-01-01

To investigate whether activated autologous platelet-rich plasma (PRP) can promote proliferation and osteogenic differentiation of human adipose-derived stem cells (hASCs) in vitro. hASCs were isolated from lipo-aspirates, and characterized by specific cell markers and multilineage differentiation capacity after culturing to the 3rd passage. PRP was collected and activated from human peripheral blood of the same patient. Cultured hASCs were treated with normal osteogenic inductive media alone (group A, control) or osteogenic inductive media plus 5%, 10%, 20%, 40%PRP (group B, C, D, E, respectively). Cell proliferation was assessed by CCK-8 assay. mRNA expression of osteogenic marker genes including alkaline phosphatase (ALP), osteopontin (OPN), osteocalcin (OCN) and core binding factor alpha 1 (Cbfa1) were determined by Real-Time Quantitative PCR Analysis (qPCR). Data revealed that different concentrations of activated autologous PRP significantly promoted hASCs growth in the proliferation phase compared to the without PRP group and resulted in a dose-response relationship. At 7-d and 14-d time point of the osteogenic induced stage, ALP activity in PRP groups gradually increased with the increasing of concentrations of PRP and showed that dose-response relationship. At 21-d time point of the osteogenic induced stage, PRP groups make much more mineralization and mRNA relative expression of ALP, OPN, OCN and Cbfa1 than that without PRP groups and show that dose-response relationship. This study indicated that different concentrations of activated autologous PRP can promote cell proliferation at earlier stage and promote osteogenic differentiation at later stage of hASCs in vitro. Moreover, it displayed a dose-dependent effect of activated autologous PRP on cell proliferation and osteogenic differentiation of hASCs in vitro. PMID:25901195

Plasmodium P-Type Cyclin CYC3 Modulates Endomitotic Growth during Oocyst Development in Mosquitoes

PubMed Central

Ferguson, David J. P.; Kaindama, Mbinda L.; Brusini, Lorenzo; Joshi, Nimitray; Rchiad, Zineb; Brady, Declan; Guttery, David S.; Wheatley, Sally P.; Yamano, Hiroyuki; Holder, Anthony A.; Pain, Arnab; Wickstead, Bill; Tewari, Rita

2015-01-01

Cell-cycle progression and cell division in eukaryotes are governed in part by the cyclin family and their regulation of cyclin-dependent kinases (CDKs). Cyclins are very well characterised in model systems such as yeast and human cells, but surprisingly little is known about their number and role in Plasmodium, the unicellular protozoan parasite that causes malaria. Malaria parasite cell division and proliferation differs from that of many eukaryotes. During its life cycle it undergoes two types of mitosis: endomitosis in asexual stages and an extremely rapid mitotic process during male gametogenesis. Both schizogony (producing merozoites) in host liver and red blood cells, and sporogony (producing sporozoites) in the mosquito vector, are endomitotic with repeated nuclear replication, without chromosome condensation, before cell division. The role of specific cyclins during Plasmodium cell proliferation was unknown. We show here that the Plasmodium genome contains only three cyclin genes, representing an unusual repertoire of cyclin classes. Expression and reverse genetic analyses of the single Plant (P)-type cyclin, CYC3, in the rodent malaria parasite, Plasmodium berghei, revealed a cytoplasmic and nuclear location of the GFP-tagged protein throughout the lifecycle. Deletion of cyc3 resulted in defects in size, number and growth of oocysts, with abnormalities in budding and sporozoite formation. Furthermore, global transcript analysis of the cyc3-deleted and wild type parasites at gametocyte and ookinete stages identified differentially expressed genes required for signalling, invasion and oocyst development. Collectively these data suggest that cyc3 modulates oocyst endomitotic development in Plasmodium berghei. PMID:26565797

Impact of Xist RNA on chromatin modifications and transcriptional silencing maintenance at different stages of imprinted X chromosome inactivation in vole Microtus levis.

PubMed

Shevchenko, Alexander I; Grigor'eva, Elena V; Medvedev, Sergey P; Zakharova, Irina S; Dementyeva, Elena V; Elisaphenko, Eugeny A; Malakhova, Anastasia A; Pavlova, Sophia V; Zakian, Suren M

2018-03-01

In vole Microtus levis, cells of preimplantation embryo and extraembryonic tissues undergo imprinted X chromosome inactivation (iXCI) which is triggered by a long non-coding nuclear RNA, Xist. At early stages of iXCI, chromatin of vole inactive X chromosome is enriched with the HP1 heterochromatin-specific protein, trimethylated H3K9 and H4K20 attributable to constitutive heterochromatin. In the study, using vole trophoblast stem (TS) cells as a model of iXCI, we further investigated chromatin of the inactive X chromosome of M. levis and tried to find out the role of Xist RNA. We demonstrated that chromatin of the inactive X chromosome in vole TS cells also contained the SETDB1 histone methyltransferase and KAP1 protein. In addition, we observed that Xist RNA did not contribute significantly to maintenance of X chromosome inactive state during iXCI in vole TS cells. Xist repression affected neither transcriptional silencing caused by iXCI nor maintenance of trimethylated H3K9 and H4K20 as well as HP1, KAP1, and SETDB1 on the inactive X chromosome. Moreover, the unique repertoire of chromatin modifications on the inactive X chromosome in vole TS cells could be disrupted by a chemical compound, DZNep, and then restored even in the absence of Xist RNA. However, Xist transcript was necessary for recruitment of an additional repressive histone modification, trimethylated H3K27, to the inactive X chromosome during vole TS cell differentiation.

Identifying States along the Hematopoietic Stem Cell Differentiation Hierarchy with Single Cell Specificity via Raman Spectroscopy.

PubMed

Ilin, Yelena; Choi, Ji Sun; Harley, Brendan A C; Kraft, Mary L

2015-11-17

A major challenge for expanding specific types of hematopoietic cells ex vivo for the treatment of blood cell pathologies is identifying the combinations of cellular and matrix cues that direct hematopoietic stem cells (HSC) to self-renew or differentiate into cell populations ex vivo. Microscale screening platforms enable minimizing the number of rare HSCs required to screen the effects of numerous cues on HSC fate decisions. These platforms create a strong demand for label-free methods that accurately identify the fate decisions of individual hematopoietic cells at specific locations on the platform. We demonstrate the capacity to identify discrete cells along the HSC differentiation hierarchy via multivariate analysis of Raman spectra. Notably, cell state identification is accurate for individual cells and independent of the biophysical properties of the functionalized polyacrylamide gels upon which these cells are cultured. We report partial least-squares discriminant analysis (PLS-DA) models of single cell Raman spectra enable identifying four dissimilar hematopoietic cell populations across the HSC lineage specification. Successful discrimination was obtained for a population enriched for long-term repopulating HSCs (LT-HSCs) versus their more differentiated progeny, including closely related short-term repopulating HSCs (ST-HSCs) and fully differentiated lymphoid (B cells) and myeloid (granulocytes) cells. The lineage-specific differentiation states of cells from these four subpopulations were accurately identified independent of the stiffness of the underlying biomaterial substrate, indicating subtle spectral variations that discriminated these populations were not masked by features from the culture substrate. This approach enables identifying the lineage-specific differentiation stages of hematopoietic cells on biomaterial substrates of differing composition and may facilitate correlating hematopoietic cell fate decisions with the extrinsic cues that elicited them.

Diagnostic accuracy of transabdominal high-resolution US for staging gallbladder cancer and differential diagnosis of neoplastic polyps compared with EUS.

PubMed

Lee, Jeong Sub; Kim, Jung Hoon; Kim, Yong Jae; Ryu, Ji Kon; Kim, Yong-Tae; Lee, Jae Young; Han, Joon Koo

2017-07-01

To compare the diagnostic accuracy of transabdominal high-resolution ultrasound (HRUS) for staging gallbladder cancer and differential diagnosis of neoplastic polyps compared with endoscopic ultrasound (EUS) and pathology. Among 125 patients who underwent both HRUS and EUS, we included 29 pathologically proven cancers (T1 = 7, T2 = 19, T3 = 3) including 15 polypoid cancers and 50 surgically proven polyps (neoplastic = 30, non-neoplastic = 20). We reviewed formal reports and assessed the accuracy of HRUS and EUS for diagnosing cancer as well as the differential diagnosis of neoplastic polyps. Statistical analyses were performed using chi-square tests. The sensitivity, specificity, PPV, and NPV for gallbladder cancer were 82.7 %, 44.4 %, 82.7 %, and 44 % using HRUS and 86.2 %, 22.2 %, 78.1 %, and 33.3 % using EUS. HRUS and EUS correctly diagnosed the stage in 13 and 12 patients. The sensitivity, specificity, PPV, and NPV for neoplastic polyps were 80 %, 80 %, 86 %, and 73 % using HRUS and 73 %, 85 %, 88 %, and 69 % using EUS. Single polyps (8/20 vs. 21/30), larger (1.0 ± 0.28 cm vs. 1.9 ± 0.85 cm) polyps, and older age (52.5 ± 13.2 vs. 66.1 ± 10.3 years) were common in neoplastic polyps (p < 0.05). Transabdominal HRUS showed comparable accuracy for diagnosing gallbladder cancer and differentiating neoplastic polyps compared with EUS. HRUS is also easy to use during our routine ultrasound examinations. • HRUS showed comparable diagnostic accuracy for GB cancer compared with EUS. • HRUS and EUS showed similar diagnostic accuracy for differentiating neoplastic polyps. • Single, larger polyps and older age were common in neoplastic polyps. • HRUS is less invasive compared with EUS.

Fulvestrant and/or Anastrozole in Treating Postmenopausal Patients With Stage II-III Breast Cancer Undergoing Surgery

ClinicalTrials.gov

2018-06-08

Estrogen Receptor-positive Breast Cancer; HER2-negative Breast Cancer; Invasive Ductal Breast Carcinoma; Invasive Lobular Breast Carcinoma; Recurrent Breast Cancer; Stage II Breast Cancer; Stage IIIA Breast Cancer; Stage IIIB Breast Cancer; Stage IIIC Breast Cancer

Applying NGS Data to Find Evolutionary Network Biomarkers from the Early and Late Stages of Hepatocellular Carcinoma

PubMed Central

Wu, Chia-Chou; Lin, Chih-Lung; Chen, Ting-Shou

2015-01-01

Hepatocellular carcinoma (HCC) is a major liver tumor (~80%), besides hepatoblastomas, angiosarcomas, and cholangiocarcinomas. In this study, we used a systems biology approach to construct protein-protein interaction networks (PPINs) for early-stage and late-stage liver cancer. By comparing the networks of these two stages, we found that the two networks showed some common mechanisms and some significantly different mechanisms. To obtain differential network structures between cancer and noncancer PPINs, we constructed cancer PPIN and noncancer PPIN network structures for the two stages of liver cancer by systems biology method using NGS data from cancer cells and adjacent noncancer cells. Using carcinogenesis relevance values (CRVs), we identified 43 and 80 significant proteins and their PPINs (network markers) for early-stage and late-stage liver cancer. To investigate the evolution of network biomarkers in the carcinogenesis process, a primary pathway analysis showed that common pathways of the early and late stages were those related to ordinary cancer mechanisms. A pathway specific to the early stage was the mismatch repair pathway, while pathways specific to the late stage were the spliceosome pathway, lysine degradation pathway, and progesterone-mediated oocyte maturation pathway. This study provides a new direction for cancer-targeted therapies at different stages. PMID:26366411

Genome-wide expression profiling of in vivo-derived bloodstream parasite stages and dynamic analysis of mRNA alterations during synchronous differentiation in Trypanosoma brucei

PubMed Central

Kabani, Sarah; Fenn, Katelyn; Ross, Alan; Ivens, Al; Smith, Terry K; Ghazal, Peter; Matthews, Keith

2009-01-01

Background Trypanosomes undergo extensive developmental changes during their complex life cycle. Crucial among these is the transition between slender and stumpy bloodstream forms and, thereafter, the differentiation from stumpy to tsetse-midgut procyclic forms. These developmental events are highly regulated, temporally reproducible and accompanied by expression changes mediated almost exclusively at the post-transcriptional level. Results In this study we have examined, by whole-genome microarray analysis, the mRNA abundance of genes in slender and stumpy forms of T.brucei AnTat1.1 cells, and also during their synchronous differentiation to procyclic forms. In total, five biological replicates representing the differentiation of matched parasite populations derived from five individual mouse infections were assayed, with RNAs being derived at key biological time points during the time course of their synchronous differentiation to procyclic forms. Importantly, the biological context of these mRNA profiles was established by assaying the coincident cellular events in each population (surface antigen exchange, morphological restructuring, cell cycle re-entry), thereby linking the observed gene expression changes to the well-established framework of trypanosome differentiation. Conclusion Using stringent statistical analysis and validation of the derived profiles against experimentally-predicted gene expression and phenotypic changes, we have established the profile of regulated gene expression during these important life-cycle transitions. The highly synchronous nature of differentiation between stumpy and procyclic forms also means that these studies of mRNA profiles are directly relevant to the changes in mRNA abundance within individual cells during this well-characterised developmental transition. PMID:19747379

Characterization and potential role of microRNA in the Chinese dominant malaria mosquito Anopheles sinensis (Diptera: Culicidae) throughout four different life stages.

PubMed

Feng, Xinyu; Wu, Jiatong; Zhou, Shuisen; Wang, Jingwen; Hu, Wei

2018-01-01

microRNAs (miRNAs) are one kind of small non-coding RNAs widely distributed in insects. Many studies have shown that miRNAs play critical roles in development, differentiation, apoptosis, and innate immunity. However, there are a few reports describing miRNAs in Anopheles sinensis , the most common, and one of the dominant malaria mosquito in China. Here, we investigated the global miRNA expression profile across four different developmental stages including embryo, larval, pupal, and adult stages using Illumina Hiseq 2500 sequencing. In total, 164 miRNAs were obtained out of 107.46 million raw sequencing reads. 99 of them identified as known miRNAs, and the remaining 65 miRNAs were considered as novel. By analyzing the read counts of miRNAs in all developmental stages, 95 miRNAs showed stage-specific expression (q < 0.01 and |log2 (fold change)| > 1) in consecutive stages, indicating that these miRNAs may be involved in critical physiological activity during development. Sixteen miRNAs were identified to be commonly dysregulated throughout four developmental stages. Many miRNAs showed stage-specific expression, such as asi-miR-2943 was exclusively expressed in the embryo stage, and asi-miR-1891 could not be detected in larval stage. The expression of six selected differentially expressed miRNAs identified by qRT-PCR were consistent with our sequencing results. Furthermore, 5296 and 1902 target genes were identified for the dysregulated 68 known and 27 novel miRNAs respectively by combining miRanda and RNAhybrid prediction. GO annotation and KEGG pathway analysis for the predicted genes of dysregulated miRNAs revealed that they might be involved in a broad range of biological processes related with the development, such as membrane, organic substance transport and several key pathways including protein processing in endoplasmic reticulum, propanoate metabolism and folate biosynthesis. Thirty-two key miRNAs were identified by microRNA-gene network analysis. The present study represents the first global characterization of An. sinensis miRNAs in its four developmental stages. The presence and differential expression of An. sinensis miRNAs imply that such miRNAs may play critical roles in An. sinensis life cycle. A better understanding of the functions of these miRNAs will have great implication for the effective control of vector population and therefore interrupting malaria transmission.

Proteomic Analysis of Tendon Extracellular Matrix Reveals Disease Stage-specific Fragmentation and Differential Cleavage of COMP (Cartilage Oligomeric Matrix Protein)*

PubMed Central

Dakin, Stephanie Georgina; Smith, Roger Kenneth Whealands; Heinegård, Dick; Önnerfjord, Patrik; Khabut, Areej; Dudhia, Jayesh

2014-01-01

During inflammatory processes the extracellular matrix (ECM) is extensively remodeled, and many of the constituent components are released as proteolytically cleaved fragments. These degradative processes are better documented for inflammatory joint diseases than tendinopathy even though the pathogenesis has many similarities. The aims of this study were to investigate the proteomic composition of injured tendons during early and late disease stages to identify disease-specific cleavage patterns of the ECM protein cartilage oligomeric matrix protein (COMP). In addition to characterizing fragments released in naturally occurring disease, we hypothesized that stimulation of tendon explants with proinflammatory mediators in vitro would induce fragments of COMP analogous to natural disease. Therefore, normal tendon explants were stimulated with IL-1β and prostaglandin E2, and their effects on the release of COMP and its cleavage patterns were characterized. Analyses of injured tendons identified an altered proteomic composition of the ECM at all stages post injury, showing protein fragments that were specific to disease stage. IL-1β enhanced the proteolytic cleavage and release of COMP from tendon explants, whereas PGE2 had no catabolic effect. Of the cleavage fragments identified in early stage tendon disease, two fragments were generated by an IL-1-mediated mechanism. These fragments provide a platform for the development of neo-epitope assays specific to injury stage for tendon disease. PMID:24398684

Impact of prostate weight on probability of positive surgical margins in patients with low-risk prostate cancer after robotic-assisted laparoscopic radical prostatectomy.

PubMed

Marchetti, Pablo E; Shikanov, Sergey; Razmaria, Aria A; Zagaja, Gregory P; Shalhav, Arieh L

2011-03-01

To evaluate the impact of prostate weight (PW) on probability of positive surgical margin (PSM) in patients undergoing robotic-assisted radical prostatectomy (RARP) for low-risk prostate cancer. The cohort consisted of 690 men with low-risk prostate cancer (clinical stage T1c, prostate-specific antigen <10 ng/mL, biopsy Gleason score ≤6) who underwent RARP with bilateral nerve-sparing at our institution by 1 of 2 surgeons from 2003 to 2009. PW was obtained from the pathologic specimen. The association between probability of PSM and PW was assessed with univariate and multivariate logistic regression analysis. A PSM was identified in 105 patients (15.2%). Patients with PSM had significant higher prostate-specific antigen (P = .04), smaller prostates (P = .0001), higher Gleason score (P = .004), and higher pathologic stage (P < .0001). After logistic regression, we found a significant inverse relation between PSM and PW (OR 0.97%; 95% confidence interval [CI] 0.96, 0.99; P = .0003) in univariate analysis. This remained significant in the multivariate model (OR 0.98%; 95% CI 0.96, 0.99; P = .006) adjusting for age, body mass index, surgeon experience, pathologic Gleason score, and pathologic stage. In this multivariate model, the predicted probability of PSM for 25-, 50-, 100-, and 150-g prostates were 22% (95% CI 16%, 30%), 13% (95% CI 11%, 16%), 5% (95% CI 1%, 8%), and 1% (95% CI 0%, 3%), respectively. Lower PW is independently associated with higher probability of PSM in low-risk patients undergoing RARP with bilateral nerve-sparing. Copyright © 2011 Elsevier Inc. All rights reserved.

Generation of monoclonal antibodies specific for cell surface molecules expressed on early mouse endoderm.

PubMed

Gadue, Paul; Gouon-Evans, Valerie; Cheng, Xin; Wandzioch, Ewa; Zaret, Kenneth S; Grompe, Markus; Streeter, Philip R; Keller, Gordon M

2009-09-01

The development of functional cell populations such as hepatocytes and pancreatic beta cells from embryonic stem cell (ESC) is dependent on the efficient induction of definitive endoderm early in the differentiation process. To monitor definitive endoderm formation in mouse ESC differentiation cultures in a quantitative fashion, we generated a reporter cell line that expresses human CD25 from the Foxa3 locus and human CD4 from the Foxa2 locus. Induction of these reporter ESCs with high concentrations of activin A led to the development of a CD25-Foxa3+CD4-Foxa2+ population within 4-5 days of culture. Isolation and characterization of this population showed that it consists predominantly of definitive endoderm that is able to undergo hepatic specification under the appropriate conditions. To develop reagents that can be used for studies on endoderm development from unmanipulated ESCs, from induced pluripotent stem cells, and from the mouse embryo, we generated monoclonal antibodies against the CD25-Foxa3+CD4-Foxa2+ population. With this approach, we identified two antibodies that react specifically with endoderm from ESC cultures and from the early embryo. The specificity of these antibodies enables one to quantitatively monitor endoderm development in ESC differentiation cultures, to study endoderm formation in the embryo, and to isolate pure populations of culture- or embryo-derived endodermal cells.

Expression of different functional isoforms in haematopoiesis.

PubMed

Grech, Godfrey; Pollacco, Joel; Portelli, Mark; Sacco, Keith; Baldacchino, Shawn; Grixti, Justine; Saliba, Christian

2014-01-01

Haematopoiesis is a complex process regulated at various levels facilitating rapid responses to external factors including stress, modulation of lineage commitment and terminal differentiation of progenitors. Although the transcription program determines the RNA pool of a cell, various mRNA strands can be obtained from the same template, giving rise to multiple protein isoforms. The majority of variants and isoforms co-occur in normal haematopoietic cells or are differentially expressed at various maturity stages of progenitor maturation and cellular differentiation within the same lineage or across lineages. Genetic aberrations or specific cellular states result in the predominant expression of abnormal isoforms leading to deregulation and disease. The presence of upstream open reading frames (uORF) in 5' untranslated regions (UTRs) of a transcript, couples the utilization of start codons with the cellular status and availability of translation initiation factors (eIFs). In addition, tissue-specific and cell lineage-specific alternative promoter use, regulates several transcription factors producing transcript variants with variable 5' exons. In this review, we propose to give a detailed account of the differential isoform formation, causing haematological malignancies.

Developmental effects of tobacco smoke exposure during human embryonic stem cell differentiation are mediated through the transforming growth factor-β superfamily member, Nodal

PubMed Central

Liszewski, Walter; Ritner, Carissa; Aurigui, Julian; Wong, Sharon S. Y.; Hussain, Naveed; Krueger, Winfried; Oncken, Cheryl; Bernstein, Harold S.

2012-01-01

While the pathologies associated with in utero smoke exposure are well established, their underlying molecular mechanisms are incompletely understood. We differentiated human embryonic stem cells in the presence of physiological concentrations of tobacco smoke and nicotine. Using post hoc microarray analysis, quantitative PCR, and immunoblot analysis, we demonstrated that tobacco smoke has lineage- and stage-specific effects on human embryonic stem cell differentiation, through both nicotine-dependent and -independent pathways. We show that three major stem cell pluripotency/differentiation pathways, Notch, canonical Wnt, and transforming growth factor-β, are affected by smoke exposure, and that Nodal signaling through SMAD2 is specifically impacted by effects on Lefty1, Nodal, and FoxH1. These events are associated with upregulation of microRNA-302a, a post-transcriptional silencer of Lefty1. The described studies provide insight into the mechanisms by which tobacco smoke influences fetal development at the cellular level, and identify specific transcriptional, post-transcriptional, and signaling pathways by which this likely occurs. PMID:22381624

Telomere Length in Predicting Toxicity in Older Patients With Stage III-IV Colorectal Cancer Undergoing Chemotherapy

ClinicalTrials.gov

2018-02-06

Mucinous Adenocarcinoma of the Colon; Mucinous Adenocarcinoma of the Rectum; Signet Ring Adenocarcinoma of the Colon; Signet Ring Adenocarcinoma of the Rectum; Stage IIIA Colon Cancer; Stage IIIA Rectal Cancer; Stage IIIB Colon Cancer; Stage IIIB Rectal Cancer; Stage IIIC Colon Cancer; Stage IIIC Rectal Cancer; Stage IV Colon Cancer; Stage IV Rectal Cancer

Computational annotation of genes differentially expressed along olive fruit development

PubMed Central

Galla, Giulio; Barcaccia, Gianni; Ramina, Angelo; Collani, Silvio; Alagna, Fiammetta; Baldoni, Luciana; Cultrera, Nicolò GM; Martinelli, Federico; Sebastiani, Luca; Tonutti, Pietro

2009-01-01

Background Olea europaea L. is a traditional tree crop of the Mediterranean basin with a worldwide economical high impact. Differently from other fruit tree species, little is known about the physiological and molecular basis of the olive fruit development and a few sequences of genes and gene products are available for olive in public databases. This study deals with the identification of large sets of differentially expressed genes in developing olive fruits and the subsequent computational annotation by means of different software. Results mRNA from fruits of the cv. Leccino sampled at three different stages [i.e., initial fruit set (stage 1), completed pit hardening (stage 2) and veraison (stage 3)] was used for the identification of differentially expressed genes putatively involved in main processes along fruit development. Four subtractive hybridization libraries were constructed: forward and reverse between stage 1 and 2 (libraries A and B), and 2 and 3 (libraries C and D). All sequenced clones (1,132 in total) were analyzed through BlastX against non-redundant NCBI databases and about 60% of them showed similarity to known proteins. A total of 89 out of 642 differentially expressed unique sequences was further investigated by Real-Time PCR, showing a validation of the SSH results as high as 69%. Library-specific cDNA repertories were annotated according to the three main vocabularies of the gene ontology (GO): cellular component, biological process and molecular function. BlastX analysis, GO terms mapping and annotation analysis were performed using the Blast2GO software, a research tool designed with the main purpose of enabling GO based data mining on sequence sets for which no GO annotation is yet available. Bioinformatic analysis pointed out a significantly different distribution of the annotated sequences for each GO category, when comparing the three fruit developmental stages. The olive fruit-specific transcriptome dataset was used to query all known KEGG (Kyoto Encyclopaedia of Genes and Genomes) metabolic pathways for characterizing and positioning retrieved EST records. The integration of the olive sequence datasets within the MapMan platform for microarray analysis allowed the identification of specific biosynthetic pathways useful for the definition of key functional categories in time course analyses for gene groups. Conclusion The bioinformatic annotation of all gene sequences was useful to shed light on metabolic pathways and transcriptional aspects related to carbohydrates, fatty acids, secondary metabolites, transcription factors and hormones as well as response to biotic and abiotic stresses throughout olive drupe development. These results represent a first step toward both functional genomics and systems biology research for understanding the gene functions and regulatory networks in olive fruit growth and ripening. PMID:19852839

Temporal impact of substrate mechanics on differentiation of human embryonic stem cells to cardiomyocytes.

PubMed

Hazeltine, Laurie B; Badur, Mehmet G; Lian, Xiaojun; Das, Amritava; Han, Wenqing; Palecek, Sean P

2014-02-01

A significant clinical need exists to differentiate human pluripotent stem cells (hPSCs) into cardiomyocytes, enabling tissue modeling for in vitro discovery of new drugs or cell-based therapies for heart repair in vivo. Chemical and mechanical microenvironmental factors are known to impact the efficiency of stem cell differentiation, but cardiac differentiation protocols in hPSCs are typically performed on rigid tissue culture polystyrene (TCPS) surfaces, which do not present a physiological mechanical setting. To investigate the temporal effects of mechanics on cardiac differentiation, we cultured human embryonic stem cells (hESCs) and their derivatives on polyacrylamide hydrogel substrates with a physiologically relevant range of stiffnesses. In directed differentiation and embryoid body culture systems, differentiation of hESCs to cardiac troponin T-expressing (cTnT+) cardiomyocytes peaked on hydrogels of intermediate stiffness. Brachyury expression also peaked on intermediate stiffness hydrogels at day 1 of directed differentiation, suggesting that stiffness impacted the initial differentiation trajectory of hESCs to mesendoderm. To investigate the impact of substrate mechanics during cardiac specification of mesodermal progenitors, we initiated directed cardiomyocyte differentiation on TCPS and transferred cells to hydrogels at the Nkx2.5/Isl1+ cardiac progenitor cell stage. No differences in cardiomyocyte purity with stiffness were observed on day 15. These experiments indicate that differentiation of hESCs is sensitive to substrate mechanics at early stages of mesodermal induction, and proper application of substrate mechanics can increase the propensity of hESCs to differentiate to cardiomyocytes. Copyright © 2013 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Baicalin maintains late-stage functional cardiomyocytes in embryoid bodies derived from murine embryonic stem cells.

PubMed

Tang, Meilin; Yin, Mengmeng; Tang, Ming; Liang, Huamin; Yu, Chong; Hu, Xinwu; Luo, Hongyan; Baudis, Birte; Haustein, Moritz; Khalil, Markus; Sarić, Tomo; Hescheler, Jürgen; Xi, Jiaoya

2013-01-01

Low efficiency of cardiomyocyte (CM) differentiation from embryonic stem (ES) cells limits their therapeutic use. The objective of this study was to investigate the effect of baicalin, a natural flavonoid compound, on the in vitro cardiac differentiation of murine ES cells. The induction of ES cells into cardiac-like cells was performed by embryoid body (EB)-based differentiation method. The electrophysiological properties of the ES cell-derived CMs (ES-CMs) were measured by patch-clamp. The biomarkers of ES-CMs were determined by quantitative RT-PCR and immunofluorescence. Continuous baicalin treatment decreased the size of EBs, and increased the proportion of α-actinin-positive CMs and transcript level of cardiac specific markers in beating EBs by inducing cell death of non-CMs. Baicalin increased the percentage of working ES-CMs which had typical responses to β-adrenergic and muscarinic stimulations. Baicalin maintains the late-stage functional CMs in EBs derived from murine ES cells. This study describes a new insight into the various biological effects of baicalin on cardiac differentiation of pluripotent stem cells. Copyright © 2013 S. Karger AG, Basel.

Blood based cell biopsy for early detection of cancer

NASA Astrophysics Data System (ADS)

Tang, Cha-Mei; Adams, Daniel; Adams, Diane; Alpaugh, R. Katherine; Cristofanilli, Massimo; Martin, Stuart; Chumsri, Saranya; Marks, Jeffrey

Early detection (ED) of cancer holds the promise for less aggressive treatments and better outcome. However, there are few accepted methods for ED. We report on a previously unknown blood cell found specifically in the peripheral blood of many solid tumors. They are defined as Cancer Associated Macrophage-Like cells (CAMLs) and are characterized by large size (25-300 μm) and expression of cancer markers. CAMLs were isolated on precision filters during blood filtration. We conducted prospective studies in breast cancer (BC) to ascertain CAML prevalence, specificity and sensitivity in relation to disease status at clinical presentation. We report on two related but separate studies: 1) the isolation of CAMLs from patients with known invasive BC, compared to healthy volunteers and, 2) a double blind study conducted on women undergoing core needle biopsy to evaluate suspicious breast masses. The studies show that CAMLs are found in all stages of BC and suggest that detection of CAMLs can differentiate patients with BC from those with benign breast conditions and healthy individuals. This non-invasive blood test can be potentially used for ED of BC and other malignancies after validation studies with the advantage of a minimally invasive procedure and longitudinal monitoring. This work was supported by Grants from Maryland TEDCO MTTCF, R01-CA154624 from NIH, KG100240 from Susan G. Komen Foundation, Era of Hope Scholar award from DoD (BC100675), and U01-CA084955 from NCI EDRN.

Quality of Life, Perceptions, and Health Satisfaction of Older Adults with End-Stage Renal Disease: A Systematic Review.

PubMed

Balogun, Seki A; Balogun, Rasheed; Philbrick, John; Abdel-Rahman, Emaad

2017-04-01

To explore the quality of life (QOL), perceptions, and health satisfaction of older adults with end-stage renal disease (ESRD) undergoing renal replacement therapy (RRT). Systematic review of literature. Individuals with ESRD undergoing RRT aged 65 and older. Articles identified from PubMed database search from January 1994 to December 2014. The methodological quality of each of the selected articles was assessed using eight standards adapted from well-established research quality review criteria. Of the initial 1,401 articles identified, 23 met the inclusion criteria. The age range of study participants examined was 65 to 90. Seventy-eight percent of the studies met six or more of the methodological standards; 47% found overall health-related and mental component summary QOL scores in elderly adults with ESRD to be similar to or higher than those of age-matched controls or younger individuals, although the physical component summary QOL scores tended to be lower in older adults. Only six studies addressed health satisfaction and perceptions of elderly adults with ESRD, with widely variable findings. Few studies specifically examine QOL in elderly adults with ESRD undergoing RRT and even fewer address issues of perceptions and health satisfaction. However, the limited data from the QOL studies looks promising with a significant proportion showing similar or higher overall health-related and mental component summary QOL scores in elderly adults with ESRD. The very limited data on perceptions and health satisfaction of elderly adults with ESRD undergoing RRT makes it difficult to make any generalizable conclusions. Overall, more research is needed to examine these factors in elderly adults with ESRD. © 2016, Copyright the Authors Journal compilation © 2016, The American Geriatrics Society.

Contrast media use in patients with chronic kidney disease undergoing coronary angiography: A systematic review and meta-analysis of randomized trials.

PubMed

Pandya, Bhavi; Chalhoub, Jean M; Parikh, Valay; Gaddam, Sainath; Spagnola, Jonathan; El-Sayegh, Suzanne; Bogin, Marc; Kandov, Ruben; Lafferty, James; Bangalore, Sripal

2017-02-01

Patients with chronic kidney disease (CKD) undergoing coronary angiography (CA), adequate hydration and minimizing volume of contrast media (CM) are class 1b recommendations for preventing contrast induced nephropathy (CIN). Current data are insufficient to justify specific recommendations about isoosmolar vs. low-osmolar contrast media by the ACCF/AHA/SCAI guidelines. Randomized trials comparing IOCM to LOCM in CKD stage 3 and above patients undergoing CA, and reporting incidence of CIN (defined by a rise in creatinine of 25% from baseline) were included in the analysis. The secondary outcome of the study was the incidence of serum creatinine increase by >1mg/dl. A total of 2839 patients were included in 10 trials, in which 1430 patients received IOCM and 1393 received LOCM. When compared to LOCM, IOCM was not associated with significant benefit in preventing CIN (OR=0.72, [CI: 0.50-1.04], P=0.08, I2=59%). Subgroup analysis revealed non-significant difference in incidence of CIN based on baseline use of N-acetylcystine (NAC), diabetes status, ejection fraction, and whether percutaneous coronary intervention vs coronary angiography alone was performed. The difference between IOCM and LOCM was further attenuated when restricted to studies with larger sample size (>250 patients) (OR=0.93; [CI: 0.66-1.30]) or when compared with non-ionic LOCM (OR=0.79, [CI: 0.52-1.21]). In patients with CKD stage 3 and above undergoing coronary angiography, use of IOCM showed overall non-significant difference in incidence of CIN compared to LOCM. The difference was further attenuated when IOCM was compared with non-ionic LOCM. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Regulation of Aicda expression and AID activity

PubMed Central

ZAN, HONG; CASALI, PAOLO

2013-01-01

Activation-induced cytidine deaminase (AID) is expressed in a B cell differentiation stage-specific fashion and is essential for immunoglobulin (Ig) gene class switch DNA recombination (CSR) and somatic hypermutation (SHM). CSR and SHM play a central role in the maturation of antibody and autoantibody responses. AID displays a mutagenic activity by catalyzing targeted deamination of deoxycytidine (dC) residues in DNA resulting in dU:dG mismatches, which are processed into point-mutations in SHM or double-strand breaks (DSBs) in CSR. Although AID specifically targets the Ig gene loci (IgH, Igκ and Igλ), it can also home into a wide array of non-Ig genes in B- and non-B-cell backgrounds. Aberrant expression of AID is associated with multiple diseases such as allergy, inflammation, autoimmunity and cancer. In autoimmune systemic lupus erythematosus, dysregulated AID expression underpins increased CSR, SHM and autoantibody production. As a potent mutator, AID is under stringent transcriptional, post-transcriptional and post-translational regulation. AID is also regulated in its targeting and enzymatic function. In resting naïve or memory B cells, AID transcripts and protein are undetectable. These, however, are readily and significantly upregulated in B cells induced to undergo CSR and/or SHM. Transcription factors, such as HoxC4 and NF-κB, which are upregulated in a B cell lineage- and/or differentiation stage-specific manner, regulate the induction of AID. HoxC4 induces AID expression by directly binding to the AID gene promoter through an evolutionarily conserved 5’-ATTT-3’ motif. HoxC4 is induced by the same stimuli that induce AID and CSR. It is further upregulated by estrogen through three estrogen responsive elements in its promoter region. The targeting of AID to switch (S) regions is mediated by 14-3-3 adaptor proteins, which specifically bind to 5′-AGCT-3′ repeats that are exist at high frequency in S region cores. Like HoxC4, 14-3-3 adaptors are induced by the same stimuli that induce AID. These include “primary” inducing stimuli, that is, those that play a major role in inducing AID, i.e., engagement of CD40 by CD154, engagement of Toll-like receptors (TLRs) by microbial-associated molecular patterns (MAMPs) and cross-linking of the BCR, as synergized by “secondary” inducing stimuli, that is, those that synergize for AID induction and specify CSR to different isotypes, i.e., switch-directing cytokines IL-4, TGF-β or IFN-γ. In this review, we focus on the multi-levels regulation of AID expression and activity. We also discuss the dysregulation or misexpression of AID in autoimmunity and tumorigenesis. PMID:23181381

SAGE analysis of early oogenesis in the silkworm, Bombyx mori.

PubMed

Funaguma, Shunsuke; Hashimoto, Shin-ichi; Suzuki, Yutaka; Omuro, Naoko; Sugano, Sumio; Mita, Kazuei; Katsuma, Susumu; Shimada, Toru

2007-02-01

To identify genes involved in the differentiation of Bombyx cystoblast, we constructed two 3' long serial analysis of gene expression (Long SAGE) libraries from stage 1-3 or stage 2-3 egg chambers and compared their gene expression profiles. In both libraries, the most frequent tags were derived from the same novel transcript. The transcript does not have any open reading frame capable of encoding a protein with over 100 amino acids in length. RNA blot analysis revealed that this transcript is specifically and abundantly expressed in the Bombyx ovary, mainly the germ line cells in the ovarioles. These results suggest that Bombyx oogenesis may be regulated by a previously unidentified non-coding RNA. Comparison of the gene expression profiles between the stage 1-3 and stage 2-3 egg chamber libraries revealed that 272 tags were significantly more abundant in stage 1-3 egg chambers (p<0.05 and at least two-fold change) than in library 2. Among the differentially expressed transcripts were the sequences that correspond to ATP synthase subunit d (3.1-fold enriched) and ATP synthase coupling factor 6 (9.1-fold enriched), suggesting that they are involved in regulation of cell cycle of cystocytes.

Cell wall invertase as a regulator in determining sequential development of endosperm and embryo through glucose signaling early in seed development.

PubMed

Wang, Lu; Liao, Shengjin; Ruan, Yong-Ling

2013-01-01

Seed development depends on coordination among embryo, endosperm and seed coat. Endosperm undergoes nuclear division soon after fertilization, whereas embryo remains quiescent for a while. Such a developmental sequence is of great importance for proper seed development. However, the underlying mechanism remains unclear. Recent results on the cellular domain- and stage-specific expression of invertase genes in cotton and Arabidopsis revealed that cell wall invertase may positively and specifically regulate nuclear division of endosperm after fertilization, thereby playing a role in determining the sequential development of endosperm and embryo, probably through glucose signaling.

Accuracy of magnetic resonance imaging in the pre-operative staging of rectal adenocarcinoma: Experience from a regional Australian cancer center.

PubMed

White, Rohen; Ung, Kim Ann; Mathlum, Maitham

2013-12-01

Selection of the optimal treatment pathway in patients with rectal adenocarcinoma relies on accurate locoregional staging. This study aims to assess the accuracy of staging with magnetic resonance imaging (MRI) and in particular, its accuracy in differentiating patients with early stage disease from those with more advanced disease who benefit from a different treatment approach. Patients who were staged with MRI and received surgery as the first line of treatment for biopsy-proven adenocarcinoma of the rectum were identified. Comparison was made between the clinical stage on MRI and the pathological stage of the surgical specimen. The sensitivity, specificity and overall accuracy of MRI was assessed. In all, 58 eligible patients were identified. In 31% of patients, the extent of disease was underrepresented on preoperative MRI. Sensitivity, specificity and overall accuracy of anorectal MRI in detecting stage II/III disease status in this cohort was 59, 71 and 62%, respectively. MRI underestimated the pathological stage in many patients in this series who may have benefited from the addition of neoadjuvant radiotherapy to their management. This study supports further refinement of preoperative staging and demonstrates that impressive results from highly controlled settings may be difficult to reproduce in community practice. © 2012 Wiley Publishing Asia Pty Ltd.

Glycoprotein and Glycan in Tissue and Blood Samples of Patients With Stage IB-IVA Cervical Cancer Undergoing Surgery to Remove Pelvic and Abdominal Lymph Nodes

ClinicalTrials.gov

2017-08-23

Cervical Adenocarcinoma; Cervical Adenosquamous Carcinoma; Cervical Small Cell Carcinoma; Cervical Squamous Cell Carcinoma, Not Otherwise Specified; Stage IB Cervical Cancer; Stage IIA Cervical Cancer; Stage IIB Cervical Cancer; Stage III Cervical Cancer; Stage IVA Cervical Cancer

Floral reversion mechanism in longan (Dimocarpus longan Lour.) revealed by proteomic and anatomic analyses.

PubMed

You, Xiangrong; Wang, Lingxia; Liang, Wenyu; Gai, Yonghong; Wang, Xiaoyan; Chen, Wei

2012-02-02

Two-dimensional gel electrophoresis (2-DE) was used to analyze the proteins related to floral reversion in Dimocarpus longan Lour. Proteins were extracted from buds undergoing the normal process of flowering and from those undergoing floral reversion in three developing stages in D. longan. Differentially expressed proteins were identified from the gels after 2-DE analysis, which were confirmed using matrix-assisted laser desorption/ionization-time of flying-mass spectroscopy and protein database search. A total of 39 proteins, including 18 up-regulated and 21 down-regulated proteins, were classified into different categories, such as energy and substance metabolism, protein translation, secondary metabolism, phytohormone, cytoskeleton structure, regulation, and stress tolerance. Among these, the largest functional class was associated with primary metabolism. Down-regulated proteins were involved in photosynthesis, transcription, and translation, whereas up-regulated proteins were involved in respiration. Decreased flavonoid synthesis and up-regulated GA20ox might be involved in the floral reversion process. Up-regulated 14-3-3 proteins played a role in the regulation of floral reversion in D. longan by responding to abiotic stress. Observations via transmission electron microscopy revealed the ultrastructure changes in shedding buds undergoing floral reversion. Overall, the results provided insights into the molecular basis for the floral reversion mechanism in D. longan. Copyright © 2011 Elsevier B.V. All rights reserved.

Free and cued memory in relation to biomarker-defined abnormalities in clinically normal older adults and those at risk for Alzheimer's disease.

PubMed

Papp, Kathryn V; Amariglio, Rebecca E; Mormino, Elizabeth C; Hedden, Trey; Dekhytar, Maria; Johnson, Keith A; Sperling, Reisa A; Rentz, Dorene M

2015-07-01

Furthering our understanding of the relationship between amyloidosis (Aβ), neurodegeneration (ND), and cognition is imperative for early identification and early intervention of Alzheimer's disease (AD). However, the subtle cognitive decline differentially associated with each biomarker-defined stage of preclinical AD has yet to be fully characterized. Recent work indicates that different components of memory performance (free and cued recall) may be differentially specific to memory decline in prodromal AD. We sought to examine the relationship between free and cued recall paradigms, in addition to global composites of memory, executive functioning, and processing speed in relation to stages of preclinical AD. A total of 260 clinically normal (CN) older adults (CDR=0) from the Harvard Aging Brain study were grouped according to preclinical AD stages including Stage 0 (Aβ-/ND-), Stage 1 (Aβ+/ND-), Stage 2 (Aβ+/ND+), and suspected non-Alzheimer's associated pathology (SNAP; Aβ-/ND+). General linear models controlling for age, sex, and education were used to assess for stage-based performance differences on cognitive composites of executive functioning, processing speed, and memory in addition to free and cued delayed recall on the Selective Reminding Test (SRT) and Memory Capacity Test (MCT). Global memory performance differed between preclinical stages with Stage 2 performing worse compared with Stage 0. When examining free and cued paradigms by memory test, only the MCT (and not the SRT) revealed group differences. More specifically, Stage 1 was associated with decrements in free recall compared with Stage 0 while Stage 2 was associated with decrements in both free and cued recall. There was a trend for the SNAP group to perform worse on free recall compared with Stage 0. Finally, there was no association between preclinical stage and global composites of executive functioning or processing speed. Clinically normal older adults with underlying evidence of amyloidosis and neurodegeneration exhibit subtle, yet measurable differences in memory performance, but only on a challenging associative test. The sensitivity of free vs. cued memory paradigms may be dependent on preclinical stage such that reduced free recall is associated with amyloidosis alone (Stage 1) while a decline in cued recall may represent progression to amyloidosis and neurodegeneration (Stage 2). These findings may have practical applications for clinical assessment and clinical trial design. Copyright © 2015. Published by Elsevier Ltd.

Free and Cued Memory in relation to Biomarker-Defined Abnormalities in Clinically Normal Older Adults and Those at Risk for Alzheimer’s Disease

PubMed Central

Papp, Kathryn V.; Amariglio, Rebecca E.; Mormino, Elizabeth; Hedden, Trey; Dekhytar, Maria; Johnson, Keith A.; Sperling, Reisa A.; Rentz, Dorene M.

2015-01-01

Objectives Furthering our understanding of the relationship between amyloidosis (Aβ), neurodegeneration (ND), and cognition is imperative for early identification and early intervention of Alzheimer’s disease (AD). However, the subtle cognitive decline differentially associated with each biomarker-defined stage of preclinical AD has yet to be fully characterized. Recent work indicates that different components of memory performance (free and cued recall) may be differentially specific to memory decline in prodromal AD. We sought to examine the relationship between free and cued recall paradigms, in addition to global composites of memory, executive functioning, and processing speed in relation to stages of preclinical AD. Methods A total of 260 clinically normal (CN) older adults (CDR=0) from the Harvard Aging Brain study were grouped according to preclinical AD stages including Stage 0 (Aβ−/ND−), Stage 1 (Aβ+/ND−), Stage 2 (Aβ+/ND+), and suspected non-Alzheimer’s associated pathology (SNAP; Aβ−/ND+). General linear models controlling for age, sex, and education were used to assess for stage-based performance differences on cognitive composites of executive functioning, processing speed, and memory in addition to free and cued delayed recall on the Selective Reminding Test (SRT) and Memory Capacity Test (MCT). Results Global memory performance differed between preclinical stages with Stage 2 performing worse compared with Stage 0. When examining free and cued paradigms by memory test, only the MCT (and not the SRT) revealed group differences. More specifically, Stage 1 was associated with decrements in free recall compared with Stage 0 while Stage 2 was associated with decrements in both free and cued recall. There was a trend for the SNAP group to perform worse on free recall compared with Stage 0. Finally, there was no association between preclinical stage and global composites of executive functioning or processing speed. Conclusions Clinically normal older adults with underlying evidence of amyloidosis and neurodegeneration exhibit subtle, yet measurable differences in memory performance, but only on a challenging associative test. The sensitivity of free vs. cued memory paradigms may be dependent on preclinical stage such that reduced free recall is associated with amyloidosis alone (Stage 1) while a decline in cued recall may represent progression to amyloidosis and neurodegeneration (Stage 2). These findings may have practical applications for clinical assessment and clinical trial design. PMID:26002757

Health-related quality of life with long-term retention of the PROSthesis of Antibiotic Loaded Acrylic Cement system following infection resolution in low demand patients.

PubMed

Beaupre, Lauren A; Stampe, Kyle; Masson, Edward; O'Connor, Gregory; Clark, Marcia; Joffe, A Mark; Boychuk, Lesia R; Lavoie, Guy

2017-01-01

The study purpose was to (1) evaluate health-related quality of life (HRQL) with the PROSthesis of Antibiotic Loaded Acrylic Cement (PROSTALAC) in situ for infected total hip arthroplasty (THA), (2) determine infection resolution, and (3) compare subjects who underwent second stage surgery with those who retained the PROSTALAC on a longer term basis. Demographics, physical demand level, and comorbidities were recorded prospectively in 29 subjects followed to at least 24 months after initial PROSTALAC insertion. HRQL was evaluated using the Western Ontario McMaster Osteoarthritis Index (WOMAC) and RAND 36-Item Health Survey. Infection resolution was determined using a pre-specified clinical definition. Twenty-five of 29 (86%) subjects' infections resolved. Three subjects died, of whom two had resolved infections. For survivors, 22/26 (85%) completed HRQL evaluations. After PROSTALAC insertion, pain and function improved within 3-6 months and was retained at 24 months. Of those followed to 24 months, 7/22 (32%) subjects underwent second stage surgery. They were higher physical demand subjects ( p = 0.03) than those not undergoing second stage surgery. We found no difference in WOMAC scores at 24 months between those who underwent second stage surgery and those who retained the PROSTALAC ( p > 0.32). The PROSTALAC system for THA appears to allow acceptable HRQL while in situ for at least 2 years in low physical demand patients. Subjects with higher physical demand levels are more likely to undergo second stage surgery. Further evaluation is required to determine whether longer term PROSTALAC retention may be appropriate for specific patient groups.

Transcriptome profiling of the whitefly Bemisia tabaci reveals stage-specific gene expression signatures for thiamethoxam resistance

PubMed Central

Yang, N; Xie, W; Jones, CM; Bass, C; Jiao, X; Yang, X; Liu, B; Li, R; Zhang, Y

2013-01-01

Bemisia tabaci has developed high levels of resistance to many insecticides including the neonicotinoids and there is strong evidence that for some compounds resistance is stage-specific. To investigate the molecular basis of B. tabaci resistance to the neonicotinoid thiamethoxam we used a custom whitefly microarray to compare gene expression in the egg, nymph and adult stages of a thiamethoxam-resistant strain (TH-R) with a susceptible strain (TH-S). Gene ontology and bioinformatic analyses revealed that in all life stages many of the differentially expressed transcripts encoded enzymes involved in metabolic processes and/or metabolism of xenobiotics. Several of these are candidate resistance genes and include the cytochrome P450 CYP6CM1, which has been shown to confer resistance to several neonicotinoids previously, a P450 belonging to the Cytochrome P450s 4 family and a glutathione S-transferase (GST) belonging to the sigma class. Finally several ATP-binding cassette transporters of the ABCG subfamily were highly over-expressed in the adult stage of the TH-R strain and may play a role in resistance by active efflux. Here, we evaluated both common and stage-specific gene expression signatures and identified several candidate resistance genes that may underlie B. tabaci resistance to thiamethoxam. PMID:23889345

Focal adhesion kinase is a regulator of F-actin dynamics

PubMed Central

Li, Stephen YT; Mruk, Dolores D; Cheng, C Yan

2013-01-01

During spermatogenesis, spermatogonia (2n, diploid) undergo a series of mitotic divisions as well as differentiation to become spermatocytes, which enter meiosis I to be followed by meiosis II to form round spermatids (1n, haploid), and then differentiate into spermatozoa (1n, haploid) via spermiogenesis. These events take place in the epithelium of the seminiferous tubule, involving extensive junction restructuring at the Sertoli-Sertoli and Sertoli-germ cell interface to allow the transport of developing germ cells across the epithelium. Although structural aspects of these cell-cell junctions have been studied, the underlying mechanism(s) that governs these events has yet to be explored. Earlier studies have shown that a non-receptor protein tyrosine kinase known as focal adhesion kinase (FAK) is a likely regulator of these events due to the stage-specific and spatiotemporal expression of its various phosphorylated/activated forms at the testis-specific anchoring junctions in the testis, as well as its association with actin regulatory proteins. Recent studies have shown that FAK, in particular its two activated phosphorylated forms p-FAK-Tyr407 and p-FAK-Tyr397, are crucial regulators in modulating junction restructuring at the Sertoli cell-cell interface at the blood-testis barrier (BTB) known as the basal ectoplasmic specialization (basal ES), as well as at the Sertoli-spermatid interface called apical ES during spermiogenesis via its effects on the filamentous (F)-actin organization at the ES. We herein summarize and critically evaluate the current knowledge regarding the physiological significance of FAK in regulating BTB and apical ES dynamics by governing the conversion of actin filaments at the ES from a “bundled” to a “de-bundled/branched” configuration and vice versa. We also provide a molecular model on the role of FAK in regulating these events based on the latest findings in the field. PMID:24381802

Utilization of nuclear structural proteins for targeted therapy and detection of proliferative and differentiation disorders

DOEpatents

Lelievre, Sophie; Bissell, Mina

2001-01-01

The localization of nuclear apparatus proteins (NUMA) is used to identify tumor cells and different stages in the tumor progression and differentiation processes. There is a characteristic organization of NuMA in tumor cells and in phenotypically normal cells. NuMA distribution patterns are significantly less diffuse in proliferating non-malignant cells compared to malignant cells. The technique encompasses cell immunostaining using a NuMA specific antibody, and microscopic analysis of NuMA distribution within each nucleus.

Predicting the nodal status in gastric cancers: The role of apparent diffusion coefficient histogram characteristic analysis.

PubMed

Liu, Song; Zhang, Yujuan; Xia, Jie; Chen, Ling; Guan, Wenxian; Guan, Yue; Ge, Yun; He, Jian; Zhou, Zhengyang

2017-10-01

To explore the application of histogram analysis in preoperative T and N staging of gastric cancers, with a focus on characteristic parameters of apparent diffusion coefficient (ADC) maps. Eighty-seven patients with gastric cancers underwent diffusion weighted magnetic resonance imaging (b=0, 1000s/mm 2 ), which generated ADC maps. Whole-volume histogram analysis was performed on ADC maps and 7 characteristic parameters were obtained. All those patients underwent surgery and postoperative pathologic T and N stages were determined. Four parameters, including skew, kurtosis, s-sD av and sample number, showed significant differences among gastric cancers at different T and N stages. Most parameters correlated with T and N stages significantly and worked in differentiating gastric cancers at different T or N stages. Especially skew yielded a sensitivity of 0.758, a specificity of 0.810, and an area under the curve (AUC) of 0.802 for differentiating gastric cancers with and without lymph node metastasis (P<0.001). All the parameters, except AUC low , showed good or excellent inter-observer agreement with intra-class correlation coefficients ranging from 0.710 to 0.991. Characteristic parameters derived from whole-volume ADC histogram analysis could help assessing preoperative T and N stages of gastric cancers. Copyright © 2017. Published by Elsevier Inc.

Effective myotube formation in human adipose tissue-derived stem cells expressing dystrophin and myosin heavy chain by cellular fusion with mouse C2C12 myoblasts

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

Eom, Young Woo; Biomedical Research Institute, Lifeliver Co., Ltd., Suwon; Lee, Jong Eun

2011-04-29

Highlights: {yields} hASCs were differentiated into skeletal muscle cells by treatment with 5-azacytidine, FGF-2, and the supernatant of cultured hASCs. {yields} Dystrophin and MyHC were expressed in late differentiation step by treatment with the supernatant of cultured hASCs. {yields} hASCs expressing dystrophin and MyHC contributed to myotube formation during co-culture with mouse myoblast C2C12 cells. -- Abstract: Stem cell therapy for muscular dystrophies requires stem cells that are able to participate in the formation of new muscle fibers. However, the differentiation steps that are the most critical for this process are not clear. We investigated the myogenic phases of humanmore » adipose tissue-derived stem cells (hASCs) step by step and the capability of myotube formation according to the differentiation phase by cellular fusion with mouse myoblast C2C12 cells. In hASCs treated with 5-azacytidine and fibroblast growth factor-2 (FGF-2) for 1 day, the early differentiation step to express MyoD and myogenin was induced by FGF-2 treatment for 6 days. Dystrophin and myosin heavy chain (MyHC) expression was induced by hASC conditioned medium in the late differentiation step. Myotubes were observed only in hASCs undergoing the late differentiation step by cellular fusion with C2C12 cells. In contrast, hASCs that were normal or in the early stage were not involved in myotube formation. Our results indicate that stem cells expressing dystrophin and MyHC are more suitable for myotube formation by co-culture with myoblasts than normal or early differentiated stem cells expressing MyoD and myogenin.« less

Differentiation induction of mouse embryonic stem cells into sinus node-like cells by suramin

PubMed Central

Wiese, Cornelia; Nikolova, Teodora; Zahanich, Ihor; Sulzbacher, Sabine; Fuchs, Joerg; Yamanaka, Satoshi; Graf, Eva; Ravens, Ursula; Boheler, Kenneth R.; Wobus, Anna M.

2015-01-01

Background Embryonic stem (ES) cells differentiate into cardiac phenotypes representing early pacemaker-, atrial-, ventricular-, and sinus node-like cells, however, ES-derived specification into sinus nodal cells is not yet known. By using the naphthylamine derivative of urea, suramin, we were able to follow the process of cardiac specialization into sinus node-like cells. Methods Differentiating mouse ES cells were treated with suramin (500 μM) from day 5 to 7 of embryoid body formation, and cells were analysed for their differentiation potential via morphological analysis, flow cytometry, RT-PCR, immunohistochemistry and patch clamp analysis. Results Application of suramin resulted in an increased number of cardiac cells, but inhibition of neuronal, skeletal muscle and definitive endoderm differentiation. Immediately after suramin treatment, a decreased mesendoderm differentiation was found. Brachyury, FGF10, Wnt8 and Wnt3a transcript levels were significantly down-regulated, followed by a decrease in mesoderm- and cardiac progenitor-specific markers BMP2, GATA4/5, Wnt11, Isl1, Nkx2.5 and Tbx5 immediately after removal of the substance. With continued differentiation, a significant up-regulation of Brachyury, FGF10 and GATA5 transcript levels was observed, whereas Nkx2.5, Isl1, Tbx5, BMP2 and Wnt11 levels were normalized to control levels. At advanced differentiation stages, sinus node-specific HCN4, Tbx2 and Tbx3 transcript levels were significantly up-regulated. Immunofluorescence and patch-clamp analysis confirmed the increased number of sinus node-like cells, and electrophysiological analysis revealed a lower number of atrial- and ventricular-like cardiomyocytes following suramin treatment. Conclusion We conclude that the interference of suramin with the cardiac differentiation process modified mesoderm- and cardiac-specific gene expression resulting in enhanced formation of sinus node-like cells. PMID:19775764

Stem Cell Differentiation Stage Factors and Their Role in Triggering Symmetry Breaking Processes during Cancer Development: A Quantum Field Theory Model for Reprogramming Cancer Cells to Healthy Phenotypes.

PubMed

Biava, Pier Mario; Burigana, Fabio; Germano, Roberto; Kurian, Philip; Verzegnassi, Claudio; Vitiello, Giuseppe

2017-09-20

A long history of research has pursued the use of embryonic factors isolated during cell differentiation processes for the express purpose of transforming cancer cells back to healthy phenotypes. Recent results have clarified that the substances present at different stages of cell differentiation-which we call stem cell differentiation stage factors (SCDSFs)-are proteins with low molecular weight and nucleic acids that regulate genomic expression. The present review summarizes how these substances, taken at different stages of cellular maturation, are able to retard proliferation of many human tumor cell lines and thereby reprogram cancer cells to healthy phenotypes. The model presented here is a quantum field theory (QFT) model in which SCDSFs are able to trigger symmetry breaking processes during cancer development. These symmetry breaking processes, which lie at the root of many phenomena in elementary particle physics and condensed matter physics, govern the phase transitions of totipotent cells to higher degrees of diversity and order, resulting in cell differentiation. In cancers, which share many genomic and metabolic similarities with embryonic stem cells, stimulated re-differentiation often signifies the phenotypic reversion back to health and non-proliferation. In addition to acting on key components of the cellular cycle, SCDSFs are able to reprogram cancer cells by delicately influencing the cancer microenvironment, modulating the electrochemistry and thus the collective electrodynamic behaviors between dipole networks in biomacromolecules and the interstitial water field. Coherent effects in biological water, which are derived from a dissipative QFT framework, may offer new diagnostic and therapeutic targets at a systemic level, before tumor instantiation occurs in specific tissues or organs. Thus, by including the environment as an essential component of our model, we may push the prevailing paradigm of mutation-driven oncogenesis toward a closer description of reality. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Parvovirus B19 Replication and Expression in Differentiating Erythroid Progenitor Cells

PubMed Central

Bua, Gloria; Manaresi, Elisabetta; Bonvicini, Francesca; Gallinella, Giorgio

2016-01-01

The pathogenic Parvovirus B19 (B19V) is characterized by a strict adaptation to erythroid progenitor cells (EPCs), a heterogeneous population of differentiating cells with diverse phenotypic and functional properties. In our work, we studied the dynamics of B19V infection in EPCs in dependence on the cell differentiation stage, in terms of distribution of infected cells, synthesis of viral nucleic acids and production of infectious virus. EPCs at early differentiation stage led to an abortive infection, without viral genome replication and a very low transcriptional activity. EPCs at later stages were permissive, with highest levels of viral replicative activity at day 9 (+3.0 Log from 2 to 48 hpi) and lower levels at day 18 (+1.5 Log from 2 to 48 hpi). B19V DNA increment was in accordance with the percentage of cells positive to flow-FISH assay (41.4% at day 9, 1.1% at day 18). Quantitation of total RNA indicated a close association of genome replication and transcription with viral RNA accumulation within infected cells related to viral DNA increase during the course of infection. Analysis of the different classes of mRNAs revealed two distinct pattern of genome expression profile with a fine regulation in the frequency utilization of RNA processing signals: an early phase, when cleavage at the proximal site leading to a higher relative production of mRNA for NS protein, and a late phase, when cleavage at the distal site was more frequent leading to higher relative abundance of mRNA for VP and 11 kDA proteins. Infectious virus was released from cells at day 6–15, but not at day 18. Our results, providing a detailed description of B19V replication and expression profile in differentiating EPCs, highlight the very tight adaptation of B19V to a specific cellular target defined both by its erythroid lineage and its differentiation stage. PMID:26845771

Surface Antigens Common to Mouse Cleavage Embryos and Primitive Teratocarcinoma Cells in Culture

PubMed Central

Artzt, Karen; Dubois, Philippe; Bennett, Dorothea; Condamine, Hubert; Babinet, Charles; Jacob, François

1973-01-01

Syngeneic antisera have been produced in mouse strain 129/Sv-CP males against the primitive cells of teratocarcinoma. These sera react specifically with the primitive cells and are negative on various types of differentiated teratoma cells derived from the same original tumor. They are negative on all other mouse cells tested, with the exception of male germ cells and cleavage-stage embryos. Thus, teratoma cells possess cell-surface antigens in common with normal cleavage-stage embryos. Images PMID:4355379

Parallel processing of general and specific threat during early stages of perception

PubMed Central

2016-01-01

Differential processing of threat can consummate as early as 100 ms post-stimulus. Moreover, early perception not only differentiates threat from non-threat stimuli but also distinguishes among discrete threat subtypes (e.g. fear, disgust and anger). Combining spatial-frequency-filtered images of fear, disgust and neutral scenes with high-density event-related potentials and intracranial source estimation, we investigated the neural underpinnings of general and specific threat processing in early stages of perception. Conveyed in low spatial frequencies, fear and disgust images evoked convergent visual responses with similarly enhanced N1 potentials and dorsal visual (middle temporal gyrus) cortical activity (relative to neutral cues; peaking at 156 ms). Nevertheless, conveyed in high spatial frequencies, fear and disgust elicited divergent visual responses, with fear enhancing and disgust suppressing P1 potentials and ventral visual (occipital fusiform) cortical activity (peaking at 121 ms). Therefore, general and specific threat processing operates in parallel in early perception, with the ventral visual pathway engaged in specific processing of discrete threats and the dorsal visual pathway in general threat processing. Furthermore, selectively tuned to distinctive spatial-frequency channels and visual pathways, these parallel processes underpin dimensional and categorical threat characterization, promoting efficient threat response. These findings thus lend support to hybrid models of emotion. PMID:26412811

Plasticity of Human THP-1 Cell Phagocytic Activity during Macrophagic Differentiation.

PubMed

Kurynina, A V; Erokhina, M V; Makarevich, O A; Sysoeva, V Yu; Lepekha, L N; Kuznetsov, S A; Onishchenko, G E

2018-03-01

Studies of the role of macrophages in phagocytosis are of great theoretical and practical importance for understanding how these cells are involved in the organism's defense response and in the development of various pathologies. Here we investigated phagocytic plasticity of THP-1 (acute monocytic human leukemia) cells at different stages (days 1, 3, and 7) of phorbol ester (PMA)-induced macrophage differentiation. Analysis of cytokine profiles showed that PMA at a concentration of 100 nM induced development of the proinflammatory macrophage population. The functional activity of macrophages was assessed on days 3 and 7 of differentiation using unlabeled latex beads and latex beads conjugated with ligands (gelatin, mannan, and IgG Fc fragment) that bind to the corresponding specific receptors. The general phagocytic activity increased significantly (1.5-2.0-fold) in the course of differentiation; phagocytosis occurred mostly through the Fc receptors, as shown previously for M1 macrophages. On day 7, the levels of phagocytosis of gelatin- and Fc-covered beads were high; however, the intensity of ingestion of mannan-conjugated beads via mannose receptors increased 2.5-3.0-fold as well, which indicated formation of cells with an alternative phenotype similar to that of M2 macrophages. Thus, the type and the plasticity of phagocytic activity at certain stages of macrophage differentiation can be associated with the formation of functionally mature morphological phenotype. This allows macrophages to exhibit their phagocytic potential in response to specific ligands. These data are of fundamental importance and can be used to develop therapeutic methods for correcting the M1/M2 macrophage ratio in an organism.

Transcriptomic insights into the genetic basis of mammalian limb diversity.

PubMed

Maier, Jennifer A; Rivas-Astroza, Marcelo; Deng, Jenny; Dowling, Anna; Oboikovitz, Paige; Cao, Xiaoyi; Behringer, Richard R; Cretekos, Chris J; Rasweiler, John J; Zhong, Sheng; Sears, Karen E

2017-03-23

From bat wings to whale flippers, limb diversification has been crucial to the evolutionary success of mammals. We performed the first transcriptome-wide study of limb development in multiple species to explore the hypothesis that mammalian limb diversification has proceeded through the differential expression of conserved shared genes, rather than by major changes to limb patterning. Specifically, we investigated the manner in which the expression of shared genes has evolved within and among mammalian species. We assembled and compared transcriptomes of bat, mouse, opossum, and pig fore- and hind limbs at the ridge, bud, and paddle stages of development. Results suggest that gene expression patterns exhibit larger variation among species during later than earlier stages of limb development, while within species results are more mixed. Consistent with the former, results also suggest that genes expressed at later developmental stages tend to have a younger evolutionary age than genes expressed at earlier stages. A suite of key limb-patterning genes was identified as being differentially expressed among the homologous limbs of all species. However, only a small subset of shared genes is differentially expressed in the fore- and hind limbs of all examined species. Similarly, a small subset of shared genes is differentially expressed within the fore- and hind limb of a single species and among the forelimbs of different species. Taken together, results of this study do not support the existence of a phylotypic period of limb development ending at chondrogenesis, but do support the hypothesis that the hierarchical nature of development translates into increasing variation among species as development progresses.

In-hospital mortality for children with hypoplastic left heart syndrome after stage I surgical palliation: teaching versus nonteaching hospitals.

PubMed

Berry, Jay G; Cowley, Collin G; Hoff, Charles J; Srivastava, Rajendu

2006-04-01

Teaching hospitals are perceived to provide a higher quality of care for the treatment of rare disease and complex patients. A substantial proportion of stage I palliation for hypoplastic left heart syndrome (HLHS) may be performed in nonteaching hospitals. This study compares the in-hospital mortality of stage I palliation between teaching and nonteaching hospitals. The authors conducted a retrospective cohort study using the Kids' Inpatient Database 1997 and 2000. Patients with HLHS undergoing stage I palliation were identified using International Classification of Diseases, Ninth Revision, Clinical Modification diagnostic and procedural codes. Seven hundred fifty-four and 880 discharges of children with HLHS undergoing stage I palliation in 1997 and 2000, respectively, were identified. The in-hospital mortality for the study population was 28% in 1997 and 24% in 2000. Twenty percent of stage I palliation operations were performed in nonteaching hospitals in 1997. Two percent of operations were performed in nonteaching hospitals in 2000. In 1997 only, in-hospital mortality remained higher in nonteaching hospitals after controlling for stage I palliation hospital volume and condition-severity diagnoses. Low-volume hospitals performing stage I palliation were associated with increased in-hospital mortality in 1997 and 2000. Patients with HLHS undergoing stage I palliation in nonteaching hospitals experienced increased in-hospital mortality in 1997. A significant reduction in the number of stage I palliation procedures performed in nonteaching hospitals occurred between 1997 and 2000. This centralization of stage I palliation into teaching hospitals, along with advances in postoperative medical and surgical care for these children, was associated with a decrease in mortality. Patients in low-volume hospitals performing stage I palliation continued to experience increased mortality in 2000.

Engineered human skin substitutes undergo large-scale genomic reprogramming and normal skin-like maturation after transplantation to athymic mice.

PubMed

Klingenberg, Jennifer M; McFarland, Kevin L; Friedman, Aaron J; Boyce, Steven T; Aronow, Bruce J; Supp, Dorothy M

2010-02-01

Bioengineered skin substitutes can facilitate wound closure in severely burned patients, but deficiencies limit their outcomes compared with native skin autografts. To identify gene programs associated with their in vivo capabilities and limitations, we extended previous gene expression profile analyses to now compare engineered skin after in vivo grafting with both in vitro maturation and normal human skin. Cultured skin substitutes were grafted on full-thickness wounds in athymic mice, and biopsy samples for microarray analyses were collected at multiple in vitro and in vivo time points. Over 10,000 transcripts exhibited large-scale expression pattern differences during in vitro and in vivo maturation. Using hierarchical clustering, 11 different expression profile clusters were partitioned on the basis of differential sample type and temporal stage-specific activation or repression. Analyses show that the wound environment exerts a massive influence on gene expression in skin substitutes. For example, in vivo-healed skin substitutes gained the expression of many native skin-expressed genes, including those associated with epidermal barrier and multiple categories of cell-cell and cell-basement membrane adhesion. In contrast, immunological, trichogenic, and endothelial gene programs were largely lacking. These analyses suggest important areas for guiding further improvement of engineered skin for both increased homology with native skin and enhanced wound healing.

Eph/Ephrin signalling maintains eye field segregation from adjacent neural plate territories during forebrain morphogenesis

PubMed Central

Cavodeassi, Florencia; Ivanovitch, Kenzo; Wilson, Stephen W.

2013-01-01

During forebrain morphogenesis, there is extensive reorganisation of the cells destined to form the eyes, telencephalon and diencephalon. Little is known about the molecular mechanisms that regulate region-specific behaviours and that maintain the coherence of cell populations undergoing specific morphogenetic processes. In this study, we show that the activity of the Eph/Ephrin signalling pathway maintains segregation between the prospective eyes and adjacent regions of the anterior neural plate during the early stages of forebrain morphogenesis in zebrafish. Several Ephrins and Ephs are expressed in complementary domains in the prospective forebrain and combinatorial abrogation of their activity results in incomplete segregation of the eyes and telencephalon and in defective evagination of the optic vesicles. Conversely, expression of exogenous Ephs or Ephrins in regions of the prospective forebrain where they are not usually expressed changes the adhesion properties of the cells, resulting in segregation to the wrong domain without changing their regional fate. The failure of eye morphogenesis in rx3 mutants is accompanied by a loss of complementary expression of Ephs and Ephrins, suggesting that this pathway is activated downstream of the regional fate specification machinery to establish boundaries between domains undergoing different programmes of morphogenesis. PMID:24026122

Clear-cell differentiation and lymphatic invasion, but not the revised TNM classification, predict lymph node metastases in pT1 penile cancer: a clinicopathologic study of 76 patients from a low incidence area.

PubMed

Mannweiler, Sebastian; Sygulla, Stephan; Tsybrovskyy, Oleksiy; Razmara, Yas; Pummer, Karl; Regauer, Sigrid

2013-10-01

Prediction of lymph node (LN) metastases in penile invasive cancer relies on clinical features and histologic characteristics of the primary tumor. Correct prediction, however, is difficult, as only 50% patients undergoing lymphadenectomies will have LN metastases. In 2009, the tumor, nodes, metastases (TNM) classification for staging of early penile cancers was revised. We tested the predictive accuracy of the revised TNM classification in a low incidence area for penile carcinoma. The presence of LN metastases in 76 men with pT1 penile cancers was correlated with the 2009 TNM subclassification, which is based on a combined evaluation of tumor grade and lymphatic invasion, but also with individual parameters, such as histologic grade, lymphatic invasion, perineural invasion, invasion depth, growth pattern and human papilloma virus (HPV) status. 76pT1 penile cancers were reclassified into 31pT1a squamous cell carcinomas (SCC) and 45pT1b (41 SCC; 4 clear-cell carcinomas); 12/22 men (55%; 8 SCC, 4 clear-cell carcinomas) undergoing lymphadenectomy for enlarged inguinal lymph nodes had metastases, 54 patients without enlarged LN and lymphadenectomies had no LN metastases during follow-up of median 47 months. Statistically, clear cell differentiation of the primary carcinoma was highly associated with metastases (100% clear-cell carcinomas vs. 11% SCC) and poor survival (50% vs. 5.5%). Among conventional SCC, only lymphatic invasion showed a highly significant association with metastases with 100% specificity. The 2009 TNM classification, tumor grade alone, perineural invasion, growth pattern, invasion depth or HPV status could not predict LN status. Lymphadenectomy for enlarged LN resulted in 100% sensitivity and 42% predictive probability for identifying metastases and a 16% false positive rate. Statistically, survival correlated significantly with clear-cell differentiation and with lymphatic invasion in both clear-cell carcinomas and conventional SCC. Penile clear-cell carcinomas are more aggressive cancers than SCC. Our observation suggests a benefit of a prophylactic lymphadenectomy for patients with clear-cell carcinomas. Among conventional SCC, only lymphatic invasion predicted LN metastases. Neither tumor grade alone nor perineural invasion, growth pattern, depth of invasion, and subgrouping according to the revised TNM classification correlated with metastases. Clinical evaluation of the LN status was superior to histologic risk stratification. Copyright © 2013 Elsevier Inc. All rights reserved.

The temporal and spatial expression of Claudins in epidermal development and the accelerated program of epidermal differentiation in K14-CaSR transgenic mice.

PubMed

Troy, Tammy-Claire; Li, Yuhua; O'Malley, Lauren; Turksen, Kursad

2007-02-01

The importance of the epidermal permeability barrier (EPB) in protecting the mammalian species against harmful UV irradiation, microorganism invasion and water loss is well recognized, as is the role of calcium (Ca(2+)) in keratinocyte differentiation, cell-cell contact and the EPB. In a previous study, we reported that the overexpression of the Ca(2+)-sensing receptor (CaSR) in the undifferentiated basal cells of the epidermis induced a modified epidermal differentiation program including an accelerated EPB formation in transgenic mice, suggesting a role for CaSR signaling in the differentiation of embryonic epidermal cells during development. We now describe the expression profile of claudins (Cldns) and keratin markers in the accelerated EPB formation of K14-CaSR transgenic mice during development as compared to the wild type from E12.5 to newborn stages. Our data show that the transgenic epidermis undergoes an advanced epidermal differentiation program as compared to the wild type as evidenced morphologically as well as by the expression of K14, K1, loricrin, Cldn6, Cldn18 and Cldn11. In addition, we report for the first time the sequential expression of Cldns in epidermal development and describe that the localization of some Cldns change within the epidermis as it matures. Furthermore, we demonstrate that Cldn6 is expressed very early in epidermal morphogenesis, followed by Cldn18, Cldn11 and Cldn1.

Histogram analysis of apparent diffusion coefficient at 3.0 T in urinary bladder lesions: correlation with pathologic findings.

PubMed

Suo, Shi-Teng; Chen, Xiao-Xi; Fan, Yu; Wu, Lian-Ming; Yao, Qiu-Ying; Cao, Meng-Qiu; Liu, Qiang; Xu, Jian-Rong

2014-08-01

To investigate the potential value of histogram analysis of apparent diffusion coefficient (ADC) obtained at standard (700 s/mm(2)) and high (1500 s/mm(2)) b values on a 3.0-T scanner in the differentiation of bladder cancer from benign lesions and in assessing bladder tumors of different pathologic T stages and to evaluate the diagnostic performance of ADC-based histogram parameters. In all, 52 patients with bladder lesions, including benign lesions (n = 7) and malignant tumors (n = 45; T1 stage or less, 23; T2 stage, 7; T3 stage, 8; and T4 stage, 7), were retrospectively evaluated. Magnetic resonance examination at 3.0 T and diffusion-weighted imaging were performed. ADC maps were obtained at two b values (b = 700 and 1500 s/mm(2); ie, ADC-700 and ADC-1500). Parameters of histogram analysis included mean, kurtosis, skewness, and entropy. The correlations between these parameters and pathologic results were revealed. Receiver operating characteristic (ROC) curves were generated to determine the diagnostic value of histogram parameters. Significant differences were found in mean ADC-700, mean ADC-1500, skewness ADC-1500, and kurtosis ADC-1500 between bladder cancer and benign lesions (P = .002-.032). There were also significant differences in mean ADC-700, mean ADC-1500, and kurtosis ADC-1500 among bladder tumors of different pathologic T stages (P = .000-.046). No significant differences were observed in other parameters. Mean ADC-1500 and kurtosis ADC-1500 were significantly correlated with T stage, respectively (ρ = -0.614, P < .001; ρ = 0.374, P = .011). ROC analysis showed that the combination of mean ADC-1500 and kurtosis ADC-1500 has the maximal area under the ROC curve (AUC, 0.894; P < .001) in the differentiation of benign lesions and malignant tumors, with a sensitivity of 77.78% and specificity of 100%. AUCs for differentiating low- and high-stage tumors were 0.840 for mean ADC-1500 (P < .001) and 0.696 for kurtosis ADC-1500 (P = .015). Histogram analysis of ADC-1500 at 3.0 T can be useful in evaluation of bladder lesions. A combination of mean ADC-1500 and kurtosis ADC-1500 may be more beneficial in the differentiation of benign and malignant lesions. Mean ADC-1500 was the most promising parameter for differentiating low- from high-stage bladder cancer. Copyright © 2014 AUR. Published by Elsevier Inc. All rights reserved.

Tensile loading modulates bone marrow stromal cell differentiation and the development of engineered fibrocartilage constructs.

PubMed

Connelly, John T; Vanderploeg, Eric J; Mouw, Janna K; Wilson, Christopher G; Levenston, Marc E

2010-06-01

Mesenchymal progenitors such as bone marrow stromal cells (BMSCs) are an attractive cell source for fibrocartilage tissue engineering, but the types or combinations of signals required to promote fibrochondrocyte-specific differentiation remain unclear. The present study investigated the influences of cyclic tensile loading on the chondrogenesis of BMSCs and the development of engineered fibrocartilage. Cyclic tensile displacements (10%, 1 Hz) were applied to BMSC-seeded fibrin constructs for short (24 h) or extended (1-2 weeks) periods using a custom loading system. At early stages of chondrogenesis, 24 h of cyclic tension stimulated both protein and proteoglycan synthesis, but at later stages, tension increased protein synthesis only. One week of intermittent cyclic tension significantly increased the total sulfated glycosaminoglycan and collagen contents in the constructs, but these differences were lost after 2 weeks of loading. Constraining the gels during the extended culture periods prevented contraction of the fibrin matrix, induced collagen fiber alignment, and increased sulfated glycosaminoglycan release to the media. Cyclic tension specifically stimulated collagen I mRNA expression and protein synthesis, but had no effect on collagen II, aggrecan, or osteocalcin mRNA levels. Overall, these studies suggest that the combination of chondrogenic stimuli and tensile loading promotes fibrochondrocyte-like differentiation of BMSCs and has the potential to direct fibrocartilage development in vitro.

A PCR-Based Diagnostic System for Differentiating Two Weevil Species (Coleoptera: Curculionidae) of Economic Importance to the Chilean Citrus Industry.

PubMed

Aguirre, C; Olivares, N; Luppichini, P; Hinrichsen, P

2015-02-01

A PCR-based method was developed to identify Naupactus cervinus (Boheman) and Naupactus xanthographus (Germar), two curculionids affecting the citrus industry in Chile. The quarantine status of these two species depends on the country to which fruits are exported. This identification method was developed because it is not possible to discriminate between these two species at the egg stage. The method is based on the species-specific amplification of sequences of internal transcribed spacers, for which we cloned and sequenced these genome fragments from each species. We designed an identification system based on two duplex-PCR reactions. Each one contains the species-specific primer set and a second generic primer set that amplify a short 18S region common to coleopterans, to avoid false negatives. The marker system is able to differentiate each Naupactus species at any life stage, and with a diagnostic sensitivity to 0.045 ng of genomic DNA. This PCR kit was validated by samples collected from different citrus production areas throughout Chile and showed 100% accuracy in differentiating the two Naupactus species. © The Authors 2015. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Diplosporous development in Boehmeria tricuspis: Insights from de novo transcriptome assembly and comprehensive expression profiling

PubMed Central

Tang, Qing; Zang, Gonggu; Cheng, Chaohua; Luan, Mingbao; Dai, Zhigang; Xu, Ying; Yang, Zemao; Zhao, Lining; Su, Jianguang

2017-01-01

Boehmeria tricuspis includes sexually reproducing diploid and apomictic triploid individuals. Previously, we established that triploid B. tricuspis reproduces through obligate diplospory. To understand the molecular basis of apomictic development in B. tricuspis, we sequenced and compared transcriptomic profiles of the flowers of sexual and apomictic plants at four key developmental stages. A total of 283,341 unique transcripts were obtained from 1,463 million high-quality paired-end reads. In total, 18,899 unigenes were differentially expressed between the reproductive types at the four stages. By classifying the transcripts into gene ontology categories of differentially expressed genes, we showed that differential plant hormone signal transduction, cell cycle regulation, and transcription factor regulation are possibly involved in apomictic development and/or a polyploidization response in B. tricuspis. Furthermore, we suggest that specific gene families are possibly related to apomixis and might have important effects on diplosporous floral development. These results make a notable contribution to our understanding of the molecular basis of diplosporous development in B. tricuspis. PMID:28382950

Extracorporeal Membrane Oxygenation Outcomes After the Comprehensive Stage II Procedure in Patients With Single Ventricles.

PubMed

Gomez, Daniel; Duffy, Vicky; Hersey, Diane; Backes, Carl; Rycus, Peter; McConnell, Patrick; Voss, Jordan; Galantowicz, Mark; Cua, Clifford L

2017-01-01

Outcomes for extracorporeal membrane oxygenation (ECMO) have been described for patients with single ventricle physiology (SVP) undergoing cavopulmonary connection (Glenn procedure). An alternative surgical pathway for patients with SVP consists of an initial hybrid procedure followed by a comprehensive Stage II procedure. No data exist describing the outcomes of patients requiring ECMO after the comprehensive Stage II procedure. The goal of this study is to describe the outcomes for patients who required ECMO after the comprehensive Stage II procedure. Data from the Extracorporeal Life Support Organization (ELSO) registry from 2001 to 2015 for children undergoing the comprehensive Stage II procedure older than 3 months of age were retrospectively analyzed. Demographics and ECMO characteristics were recorded. A total of six children required ECMO support after the comprehensive Stage II procedure (2 males, 4 females). Four patients had the diagnosis of hypoplastic left heart syndrome and two patients had the diagnosis of an unbalanced atrioventricular septal defect. Bypass time was 242.8 ± 110.9 min and cross-clamp time was 91.2 ± 46.2 min for the surgical procedure. Weight was 5.8 ± 1.3 kg and age was 150.2 + 37.9 days at time of ECMO. ECMO duration was 276.0 ± 218.1 h. Complications during the ECMO run included hemorrhage in four patients (67%), renal dysfunction in two patients (33%), and neurologic injury in two patients (33%). Four patients (67%) were discharged alive after ECMO decannulation. Despite being a much more extensive surgical procedure, the morbidity and mortality after ECMO in patients undergoing the comprehensive Stage II procedure are similar to those in patients undergoing the Glenn procedure. If needed, ECMO support is reasonable for patients after the comprehensive Stage II procedure. © 2016 International Center for Artificial Organs and Transplantation and Wiley Periodicals, Inc.

Sexually dimorphic gene expressions in eels: useful markers for early sex assessment in a conservation context

PubMed Central

Geffroy, Benjamin; Guilbaud, Florian; Amilhat, Elsa; Beaulaton, Laurent; Vignon, Matthias; Huchet, Emmanuel; Rives, Jacques; Bobe, Julien; Fostier, Alexis; Guiguen, Yann; Bardonnet, Agnès

2016-01-01

Environmental sex determination (ESD) has been detected in a range of vertebrate reptile and fish species. Eels are characterized by an ESD that occurs relatively late, since sex cannot be histologically determined before individuals reach 28 cm. Because several eel species are at risk of extinction, assessing sex at the earliest stage is a crucial management issue. Based on preliminary results of RNA sequencing, we targeted genes susceptible to be differentially expressed between ovaries and testis at different stages of development. Using qPCR, we detected testis-specific expressions of dmrt1, amh, gsdf and pre-miR202 and ovary-specific expressions were obtained for zar1, zp3 and foxn5. We showed that gene expressions in the gonad of intersexual eels were quite similar to those of males, supporting the idea that intersexual eels represent a transitional stage towards testicular differentiation. To assess whether these genes would be effective early molecular markers, we sampled juvenile eels in two locations with highly skewed sex ratios. The combined expression of six of these genes allowed the discrimination of groups according to their potential future sex and thus this appears to be a useful tool to estimate sex ratios of undifferentiated juvenile eels. PMID:27658729

Sexually dimorphic gene expressions in eels: useful markers for early sex assessment in a conservation context

NASA Astrophysics Data System (ADS)

Geffroy, Benjamin; Guilbaud, Florian; Amilhat, Elsa; Beaulaton, Laurent; Vignon, Matthias; Huchet, Emmanuel; Rives, Jacques; Bobe, Julien; Fostier, Alexis; Guiguen, Yann; Bardonnet, Agnès

2016-09-01

Environmental sex determination (ESD) has been detected in a range of vertebrate reptile and fish species. Eels are characterized by an ESD that occurs relatively late, since sex cannot be histologically determined before individuals reach 28 cm. Because several eel species are at risk of extinction, assessing sex at the earliest stage is a crucial management issue. Based on preliminary results of RNA sequencing, we targeted genes susceptible to be differentially expressed between ovaries and testis at different stages of development. Using qPCR, we detected testis-specific expressions of dmrt1, amh, gsdf and pre-miR202 and ovary-specific expressions were obtained for zar1, zp3 and foxn5. We showed that gene expressions in the gonad of intersexual eels were quite similar to those of males, supporting the idea that intersexual eels represent a transitional stage towards testicular differentiation. To assess whether these genes would be effective early molecular markers, we sampled juvenile eels in two locations with highly skewed sex ratios. The combined expression of six of these genes allowed the discrimination of groups according to their potential future sex and thus this appears to be a useful tool to estimate sex ratios of undifferentiated juvenile eels.

Surfing the wave, cycle, life history, and genes/proteins expressed by testicular germ cells. Part 4: intercellular bridges, mitochondria, nuclear envelope, apoptosis, ubiquitination, membrane/voltage-gated channels, methylation/acetylation, and transcription factors.

PubMed

Hermo, Louis; Pelletier, R-Marc; Cyr, Daniel G; Smith, Charles E

2010-04-01

As germ cells divide and differentiate from spermatogonia to spermatozoa, they share a number of structural and functional features that are common to all generations of germ cells and these features are discussed herein. Germ cells are linked to one another by large intercellular bridges which serve to move molecules and even large organelles from the cytoplasm of one cell to another. Mitochondria take on different shapes and features and topographical arrangements to accommodate their specific needs during spermatogenesis. The nuclear envelope and pore complex also undergo extensive modifications concomitant with the development of germ cell generations. Apoptosis is an event that is normally triggered by germ cells and involves many proteins. It occurs to limit the germ cell pool and acts as a quality control mechanism. The ubiquitin pathway comprises enzymes that ubiquitinate as well as deubiquitinate target proteins and this pathway is present and functional in germ cells. Germ cells express many proteins involved in water balance and pH control as well as voltage-gated ion channel movement. In the nucleus, proteins undergo epigenetic modifications which include methylation, acetylation, and phosphorylation, with each of these modifications signaling changes in chromatin structure. Germ cells contain specialized transcription complexes that coordinate the differentiation program of spermatogenesis, and there are many male germ cell-specific differences in the components of this machinery. All of the above features of germ cells will be discussed along with the specific proteins/genes and abnormalities to fertility related to each topic. Copyright 2009 Wiley-Liss, Inc.

Modulation of CD4(+) T cell-dependent specific cytotoxic CD8(+) T cells differentiation and proliferation by the timing of increase in the pathogen load.

PubMed

Tzelepis, Fanny; Persechini, Pedro M; Rodrigues, Mauricio M

2007-04-25

Following infection with viruses, bacteria or protozoan parasites, naïve antigen-specific CD8(+) T cells undergo a process of differentiation and proliferation to generate effector cells. Recent evidences suggest that the timing of generation of specific effector CD8(+) T cells varies widely according to different pathogens. We hypothesized that the timing of increase in the pathogen load could be a critical parameter governing this process. Using increasing doses of the protozoan parasite Trypanosoma cruzi to infect C57BL/6 mice, we observed a significant acceleration in the timing of parasitemia without an increase in mouse susceptibility. In contrast, in CD8 deficient mice, we observed an inverse relationship between the parasite inoculum and the timing of death. These results suggest that in normal mice CD8(+) T cells became protective earlier, following the accelerated development of parasitemia. The evaluation of specific cytotoxic responses in vivo to three distinct epitopes revealed that increasing the parasite inoculum hastened the expansion of specific CD8(+) cytotoxic T cells following infection. The differentiation and expansion of T. cruzi-specific CD8(+) cytotoxic T cells is in fact dependent on parasite multiplication, as radiation-attenuated parasites were unable to activate these cells. We also observed that, in contrast to most pathogens, the activation process of T. cruzi-specific CD8(+) cytotoxic T cells was dependent on MHC class II restricted CD4(+) T cells. Our results are compatible with our initial hypothesis that the timing of increase in the pathogen load can be a critical parameter governing the kinetics of CD4(+) T cell-dependent expansion of pathogen-specific CD8(+) cytotoxic T cells.

[Metabolic therapy with iodine 131 in patients with chronic renal failure. Clinical case].

PubMed

Vázquez-Rodríguez Barbero, Inmaculada; Espadas-Maeso, María José; Muñoz-Morales, Ana; Flores-Gómez, Pilar; Serrano-Carretero, María Belén; Castedo-Sal, Juan José; Sánchez Rey-Castro, Elena; Zamorano-Córdoba, Antonio

2015-01-01

62 year-old male with CKD stage 5 in dialysis program since 2012 who underwent surgery for papillary thyroid carcinoma with lymph node metastasis. Subsequently, the patient was admitted to the Unit of Metabolic Therapy at his reference hospital, where he is administered 80 millicuries of iodine-131 as a treatment associated with the surgery, unable to have his conventional dialysis, nursing interventions required in various stages of implementation of the continuous extrarenal clearance techniques (TCDE) were performed. The following care values were addressed targeted to patients undergoing treatment with I(131): specific equipment, personal protective measures and major diagnoses and nursing interventions. TCDE, despite being attributed to very critical patients, they can be extrapolated to other patients in a given time. TCDE allowed these patients to undergo dialysis in a safe environment for staff, ensuring proper disposal of contaminated liquids. TCDE were an effective treatment for the removal of I131, a single ssesion being necessary to normalize the levels of radioactivity. Coordination between services allewed ensure proper and effective treatment for the patient. Copyright © 2015 Elsevier España, S.L.U. All rights reserved.

Optical spectroscopy for differentiation of liver tissue under distinct stages of fibrosis: an ex vivo study

NASA Astrophysics Data System (ADS)

Fabila, D. A.; Hernández, L. F.; de la Rosa, J.; Stolik, S.; Arroyo-Camarena, U. D.; López-Vancell, M. D.; Escobedo, G.

2013-11-01

Liver fibrosis is the decisive step towards the development of cirrhosis; its early detection affects crucially the diagnosis of liver disease, its prognosis and therapeutic decision making. Nowadays, several techniques are employed to this task. However, they have the limitation in estimating different stages of the pathology. In this paper we present a preliminary study to evaluate if optical spectroscopy can be employed as an auxiliary tool of diagnosis of biopsies of human liver tissue to differentiate the fibrosis stages. Ex vivo fluorescence and diffuse reflectance spectra were acquired from biopsies using a portable fiber-optic system. Empirical discrimination algorithms based on fluorescence intensity ratio at 500 nm and 680 nm as well as diffuse reflectance intensity at 650 nm were developed. Sensitivity and specificity of around 80% and 85% were respectively achieved. The obtained results show that combined use of fluorescence and diffuse reflectance spectroscopy could represent a novel and useful tool in the early evaluation of liver fibrosis.

Adipose tissue-derived mesenchymal stem cells in long-term dialysis patients display downregulation of PCAF expression and poor angiogenesis activation.

PubMed

Yamanaka, Shuichiro; Yokote, Shinya; Yamada, Akifumi; Katsuoka, Yuichi; Izuhara, Luna; Shimada, Yohta; Omura, Nobuo; Okano, Hirotaka James; Ohki, Takao; Yokoo, Takashi

2014-01-01

We previously demonstrated that mesenchymal stem cells (MSCs) differentiate into functional kidney cells capable of urine and erythropoietin production, indicating that they may be used for kidney regeneration. However, the viability of MSCs from dialysis patients may be affected under uremic conditions. In this study, we isolated MSCs from the adipose tissues of end-stage kidney disease (ESKD) patients undergoing long-term dialysis (KD-MSCs; mean: 72.3 months) and from healthy controls (HC-MSCs) to compare their viability. KD-MSCs and HC-MSCs were assessed for their proliferation potential, senescence, and differentiation capacities into adipocytes, osteoblasts, and chondrocytes. Gene expression of stem cell-specific transcription factors was analyzed by PCR array and confirmed by western blot analysis at the protein level. No significant differences of proliferation potential, senescence, or differentiation capacity were observed between KD-MSCs and HC-MSCs. However, gene and protein expression of p300/CBP-associated factor (PCAF) was significantly suppressed in KD-MSCs. Because PCAF is a histone acetyltransferase that mediates regulation of hypoxia-inducible factor-1α (HIF-1α), we examined the hypoxic response in MSCs. HC-MSCs but not KD-MSCs showed upregulation of PCAF protein expression under hypoxia. Similarly, HIF-1α and vascular endothelial growth factor (VEGF) expression did not increase under hypoxia in KD-MSCs but did so in HC-MSCs. Additionally, a directed in vivo angiogenesis assay revealed a decrease in angiogenesis activation of KD-MSCs. In conclusion, long-term uremia leads to persistent and systematic downregulation of PCAF gene and protein expression and poor angiogenesis activation of MSCs from patients with ESKD. Furthermore, PCAF, HIF-1α, and VEGF expression were not upregulated by hypoxic stimulation of KD-MSCs. These results suggest that the hypoxic response may be blunted in MSCs from ESKD patients.

Adipose Tissue-Derived Mesenchymal Stem Cells in Long-Term Dialysis Patients Display Downregulation of PCAF Expression and Poor Angiogenesis Activation

PubMed Central

Yamanaka, Shuichiro; Yokote, Shinya; Yamada, Akifumi; Katsuoka, Yuichi; Izuhara, Luna; Shimada, Yohta; Omura, Nobuo; Okano, Hirotaka James; Ohki, Takao; Yokoo, Takashi

2014-01-01

We previously demonstrated that mesenchymal stem cells (MSCs) differentiate into functional kidney cells capable of urine and erythropoietin production, indicating that they may be used for kidney regeneration. However, the viability of MSCs from dialysis patients may be affected under uremic conditions. In this study, we isolated MSCs from the adipose tissues of end-stage kidney disease (ESKD) patients undergoing long-term dialysis (KD-MSCs; mean: 72.3 months) and from healthy controls (HC-MSCs) to compare their viability. KD-MSCs and HC-MSCs were assessed for their proliferation potential, senescence, and differentiation capacities into adipocytes, osteoblasts, and chondrocytes. Gene expression of stem cell-specific transcription factors was analyzed by PCR array and confirmed by western blot analysis at the protein level. No significant differences of proliferation potential, senescence, or differentiation capacity were observed between KD-MSCs and HC-MSCs. However, gene and protein expression of p300/CBP-associated factor (PCAF) was significantly suppressed in KD-MSCs. Because PCAF is a histone acetyltransferase that mediates regulation of hypoxia-inducible factor-1α (HIF-1α), we examined the hypoxic response in MSCs. HC-MSCs but not KD-MSCs showed upregulation of PCAF protein expression under hypoxia. Similarly, HIF-1α and vascular endothelial growth factor (VEGF) expression did not increase under hypoxia in KD-MSCs but did so in HC-MSCs. Additionally, a directed in vivo angiogenesis assay revealed a decrease in angiogenesis activation of KD-MSCs. In conclusion, long-term uremia leads to persistent and systematic downregulation of PCAF gene and protein expression and poor angiogenesis activation of MSCs from patients with ESKD. Furthermore, PCAF, HIF-1α, and VEGF expression were not upregulated by hypoxic stimulation of KD-MSCs. These results suggest that the hypoxic response may be blunted in MSCs from ESKD patients. PMID:25025381

Caloric Restriction in Treating Patients With Stage 0-I Breast Cancer Undergoing Surgery and Radiation Therapy

ClinicalTrials.gov

2017-09-25

Ductal Breast Carcinoma in Situ; Invasive Ductal Breast Carcinoma; Invasive Lobular Breast Carcinoma; Lobular Breast Carcinoma in Situ; Recurrent Breast Cancer; Stage IA Breast Cancer; Stage IB Breast Cancer

Germinal center hypoxia potentiates immunoglobulin class switch recombination

PubMed Central

Abbott, Robert K.; Thayer, Molly; Labuda, Jasmine; Silva, Murillo; Philbrook, Phaethon; Cain, Derek W.; Kojima, Hidefumi; Hatfield, Stephen; Sethumadhavan, Shalini; Ohta, Akio; Reinherz, Ellis L.; Kelsoe, Garnett; Sitkovsky, Michail

2016-01-01

Germinal centers (GCs) are anatomic sites where B cells undergo secondary diversification to produce high affinity, class switched antibodies. We hypothesized that proliferating B cells in GCs create a hypoxic microenvironment that governs their further differentiation. Using molecular markers, we found GCs to be predominantly hypoxic. Compared to normoxia (21% O2), hypoxic culture conditions (1% O2) in vitro accelerated class switching and plasma cell formation and enhanced expression of GL-7 on B and CD4+ T cells. Reversal of GC hypoxia in vivo by breathing 60% O2 during immunization resulted in reduced frequencies of GC B cells, T follicular helper (TFH) cells and plasmacytes, as well as lower expression of ICOS on TFH. Importantly, this reversal of GC hypoxia decreased antigen-specific serum IgG1 and reduced the frequency of IgG1+ B cells within the antigen specific GC. Taken together, these observations reveal a critical role for hypoxia in GC B cell differentiation. PMID:27798169

Analysis of the SWI/SNF chromatin-remodeling complex during early heart development and BAF250a repression cardiac gene transcription during P19 cell differentiation

PubMed Central

Singh, Ajeet Pratap; Archer, Trevor K.

2014-01-01

The regulatory networks of differentiation programs and the molecular mechanisms of lineage-specific gene regulation in mammalian embryos remain only partially defined. We document differential expression and temporal switching of BRG1-associated factor (BAF) subunits, core pluripotency factors and cardiac-specific genes during post-implantation development and subsequent early organogenesis. Using affinity purification of BRG1 ATPase coupled to mass spectrometry, we characterized the cardiac-enriched remodeling complexes present in E8.5 mouse embryos. The relative abundance and combinatorial assembly of the BAF subunits provides functional specificity to Switch/Sucrose NonFermentable (SWI/SNF) complexes resulting in a unique gene expression profile in the developing heart. Remarkably, the specific depletion of the BAF250a subunit demonstrated differential effects on cardiac-specific gene expression and resulted in arrhythmic contracting cardiomyocytes in vitro. Indeed, the BAF250a physically interacts and functionally cooperates with Nucleosome Remodeling and Histone Deacetylase (NURD) complex subunits to repressively regulate chromatin structure of the cardiac genes by switching open and poised chromatin marks associated with active and repressed gene expression. Finally, BAF250a expression modulates BRG1 occupancy at the loci of cardiac genes regulatory regions in P19 cell differentiation. These findings reveal specialized and novel cardiac-enriched SWI/SNF chromatin-remodeling complexes, which are required for heart formation and critical for cardiac gene expression regulation at the early stages of heart development. PMID:24335282

Hydroxyapatite and Calcified Elastin Induce Osteoblast-like Differentiation in Rat Aortic Smooth Muscle Cells

PubMed Central

Lei, Yang; Sinha, Aditi; Nosoudi, Nasim; Grover, Ankit; Vyavahare, Naren

2014-01-01

Vascular calcification can be categorized into two different types. Intimal calcification related to atherosclerosis and elastin-specific medial arterial calcification (MAC). Osteoblast-like differentiation of vascular smooth muscle cells (VSMCs) has been shown in both types; however, how this relates to initiation of vascular calcification is unclear. We hypothesize that the initial deposition of hydroxyapatite-like mineral in MAC occurs on degraded elastin first and that causes osteogenic transformation of VSMCs. To test this, rat aortic smooth muscle cells (RASMCs) were cultured on hydroxyapatite crystals and calcified aortic elastin. Using RT-PCR and specific protein assays, we demonstrate that RASMCs lose their smooth muscle lineage markers like alpha smooth muscle actin (SMA) and myosin heavy chain (MHC) and undergo chondrogenic/osteogenic transformation. This is indicated by an increase in the expression of typical chondrogenic proteins such as aggrecan, collagen type II alpha 1(Col2a1) and bone proteins such as runt-related transcription factor 2 (RUNX2), alkaline phosphatase (ALP) and osteocalcin (OCN). Furthermore, when calcified conditions are removed, cells return to their original phenotype. Our data supports the hypothesis that elastin degradation and calcification precedes VSMCs' osteoblast-like differentiation. PMID:24447384

Intraoperative Raman Spectroscopy of Soft Tissue Sarcomas

PubMed Central

Nguyen, John Q.; Gowani, Zain S.; O’Connor, Maggie; Pence, Isaac J.; Nguyen, The-Quyen; Holt, Ginger E.; Schwartz, Herbert S.; Halpern, Jennifer L.; Mahadevan-Jansen, Anita

2017-01-01

Background and Objective Soft tissue sarcomas (STS) are a rare and heterogeneous group of malignant tumors that are often treated through surgical resection. Current intraoperative margin assessment methods are limited and highlight the need for an improved approach with respect to time and specificity. Here we investigate the potential of near-infrared Raman spectroscopy for the intraoperative differentiation of STS from surrounding normal tissue. Materials and Methods In vivo Raman measurements at 785 nm excitation were intraoperatively acquired from subjects undergoing STS resection using a probe based spectroscopy system. A multivariate classification algorithm was developed in order to automatically identify spectral features that can be used to differentiate STS from the surrounding normal muscle and fat. The classification algorithm was subsequently tested using leave-one-subject-out cross-validation. Results With the exclusion of well-differentiated liposarcomas, the algorithm was able to classify STS from the surrounding normal muscle and fat with a sensitivity and specificity of 89.5% and 96.4%, respectively. Conclusion These results suggest that single point near-infrared Raman spectroscopy could be utilized as a rapid and non-destructive surgical guidance tool for identifying abnormal tissue margins in need of further excision. PMID:27454580

Intraoperative Raman spectroscopy of soft tissue sarcomas.

PubMed

Nguyen, John Q; Gowani, Zain S; O'Connor, Maggie; Pence, Isaac J; Nguyen, The-Quyen; Holt, Ginger E; Schwartz, Herbert S; Halpern, Jennifer L; Mahadevan-Jansen, Anita

2016-10-01

Soft tissue sarcomas (STS) are a rare and heterogeneous group of malignant tumors that are often treated through surgical resection. Current intraoperative margin assessment methods are limited and highlight the need for an improved approach with respect to time and specificity. Here we investigate the potential of near-infrared Raman spectroscopy for the intraoperative differentiation of STS from surrounding normal tissue. In vivo Raman measurements at 785 nm excitation were intraoperatively acquired from subjects undergoing STS resection using a probe based spectroscopy system. A multivariate classification algorithm was developed in order to automatically identify spectral features that can be used to differentiate STS from the surrounding normal muscle and fat. The classification algorithm was subsequently tested using leave-one-subject-out cross-validation. With the exclusion of well-differentiated liposarcomas, the algorithm was able to classify STS from the surrounding normal muscle and fat with a sensitivity and specificity of 89.5% and 96.4%, respectively. These results suggest that single point near-infrared Raman spectroscopy could be utilized as a rapid and non-destructive surgical guidance tool for identifying abnormal tissue margins in need of further excision. Lasers Surg. Med. 48:774-781, 2016. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Inhibition of Apoptosis Overcomes Stage-Related Compatibility Barriers to Chimera Formation in Mouse Embryos.

PubMed

Masaki, Hideki; Kato-Itoh, Megumi; Takahashi, Yusuke; Umino, Ayumi; Sato, Hideyuki; Ito, Keiichi; Yanagida, Ayaka; Nishimura, Toshinobu; Yamaguchi, Tomoyuki; Hirabayashi, Masumi; Era, Takumi; Loh, Kyle M; Wu, Sean M; Weissman, Irving L; Nakauchi, Hiromitsu

2016-11-03

Cell types more advanced in development than embryonic stem cells, such as EpiSCs, fail to contribute to chimeras when injected into pre-implantation-stage blastocysts, apparently because the injected cells undergo apoptosis. Here we show that transient promotion of cell survival through expression of the anti-apoptotic gene BCL2 enables EpiSCs and Sox17 + endoderm progenitors to integrate into blastocysts and contribute to chimeric embryos. Upon injection into blastocyst, BCL2-expressing EpiSCs contributed to all bodily tissues in chimeric animals while Sox17 + endoderm progenitors specifically contributed in a region-specific fashion to endodermal tissues. In addition, BCL2 expression enabled rat EpiSCs to contribute to mouse embryonic chimeras, thereby forming interspecies chimeras that could survive to adulthood. Our system therefore provides a method to overcome cellular compatibility issues that typically restrict chimera formation. Application of this type of approach could broaden the use of embryonic chimeras, including region-specific chimeras, for basic developmental biology research and regenerative medicine. Copyright © 2016 Elsevier Inc. All rights reserved.

The structure of bradyzoite-specific enolase from Toxoplasma gondii reveals insights into its dual cytoplasmic and nuclear functions

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

Ruan, Jiapeng; Mouveaux, Thomas; Light, Samuel H.

2015-03-01

The second crystal structure of a parasite protein preferentially enriched in the brain cyst of T. gondii has been solved at 2.75 Å resolution. Bradyzoite enolase 1 is reported to have differential functions as a glycolytic enzyme and a transcriptional regulator in bradyzoites. In addition to catalyzing a central step in glycolysis, enolase assumes a remarkably diverse set of secondary functions in different organisms, including transcription regulation as documented for the oncogene c-Myc promoter-binding protein 1. The apicomplexan parasite Toxoplasma gondii differentially expresses two nuclear-localized, plant-like enolases: enolase 1 (TgENO1) in the latent bradyzoite cyst stage and enolase 2 (TgENO2)more » in the rapidly replicative tachyzoite stage. A 2.75 Å resolution crystal structure of bradyzoite enolase 1, the second structure to be reported of a bradyzoite-specific protein in Toxoplasma, captures an open conformational state and reveals that distinctive plant-like insertions are located on surface loops. The enolase 1 structure reveals that a unique residue, Glu164, in catalytic loop 2 may account for the lower activity of this cyst-stage isozyme. Recombinant TgENO1 specifically binds to a TTTTCT DNA motif present in the cyst matrix antigen 1 (TgMAG1) gene promoter as demonstrated by gel retardation. Furthermore, direct physical interactions of both nuclear TgENO1 and TgENO2 with the TgMAG1 gene promoter are demonstrated in vivo using chromatin immunoprecipitation (ChIP) assays. Structural and biochemical studies reveal that T. gondii enolase functions are multifaceted, including the coordination of gene regulation in parasitic stage development. Enolase 1 provides a potential lead in the design of drugs against Toxoplasma brain cysts.« less

IGFBP-2 Vaccine and Combination Chemotherapy in Treating Patients With Stage III-IV Ovarian, Fallopian Tube, or Primary Peritoneal Cancer Undergoing Surgery

ClinicalTrials.gov

2018-05-01

Stage III Fallopian Tube Cancer; Stage III Ovarian Cancer; Stage III Primary Peritoneal Cancer; Stage IIIA Fallopian Tube Cancer; Stage IIIA Ovarian Cancer; Stage IIIA Primary Peritoneal Cancer; Stage IIIB Fallopian Tube Cancer; Stage IIIB Ovarian Cancer; Stage IIIB Primary Peritoneal Cancer; Stage IIIC Fallopian Tube Cancer; Stage IIIC Ovarian Cancer; Stage IIIC Primary Peritoneal Cancer; Stage IV Fallopian Tube Cancer; Stage IV Ovarian Cancer; Stage IV Primary Peritoneal Cancer

Typical cerebral metabolic patterns in neurodegenerative brain diseases.

PubMed

Teune, Laura K; Bartels, Anna L; de Jong, Bauke M; Willemsen, Antoon T M; Eshuis, Silvia A; de Vries, Jeroen J; van Oostrom, Joost C H; Leenders, Klaus L

2010-10-30

The differential diagnosis of neurodegenerative brain diseases on clinical grounds is difficult, especially at an early disease stage. Several studies have found specific regional differences of brain metabolism applying [(18)F]-fluoro-deoxyglucose positron emission tomography (FDG-PET), suggesting that this method can assist in early differential diagnosis of neurodegenerative brain diseases.We have studied patients who had an FDG-PET scan on clinical grounds at an early disease stage and included those with a retrospectively confirmed diagnosis according to strictly defined clinical research criteria. Ninety-six patients could be included of which 20 patients with Parkinson's disease (PD), 21 multiple system atrophy (MSA), 17 progressive supranuclear palsy (PSP), 10 corticobasal degeneration (CBD), 6 dementia with Lewy bodies (DLB), 15 Alzheimer's disease (AD), and 7 frontotemporal dementia (FTD). FDG PET images of each patient group were analyzed and compared to18 healthy controls using Statistical Parametric Mapping (SPM5).Disease-specific patterns of relatively decreased metabolic activity were found in PD (contralateral parietooccipital and frontal regions), MSA (bilateral putamen and cerebellar hemispheres), PSP (prefrontal cortex and caudate nucleus, thalamus, and mesencephalon), CBD (contralateral cortical regions), DLB (occipital and parietotemporal regions), AD (parietotemporal regions), and FTD (frontotemporal regions).The integrated method addressing a spectrum of various neurodegenerative brain diseases provided means to discriminate patient groups also at early disease stages. Clinical follow-up enabled appropriate patient inclusion. This implies that an early diagnosis in individual patients can be made by comparing each subject's metabolic findings with a complete database of specific disease related patterns.

Development of ovary structures in the last larval and adult stages of psyllids (Insecta, Hemiptera, Sternorrhyncha: Psylloidea).

PubMed

Kot, Marta; Büning, Jürgen; Jankowska, Władysława; Drohojowska, Jowita; Szklarzewicz, Teresa

2016-07-01

The development and organization of the ovaries of ten species from four Psylloidea families (Psyllidae, Triozidae, Aphalaridae and Liviidae) have been investigated. The ovaries of the last larval stage (i.e. fifth instar) of all examined species are filled with numerous clusters of cystocytes which undergo synchronous incomplete mitotic division. Cystocytes of the given cluster are arranged into a rosette with polyfusome in the centre. These clusters are associated with single somatic cells. At the end of the fifth instar, the clusters begin to separate from each other, forming spherical ovarioles which are surrounded by a single layer of somatic cells. In the ovarioles of very young females all cystocytes enter the prophase of meiosis and differentiate shortly thereafter into oocytes and trophocytes (nurse cells). Meanwhile, somatic cells differentiate into cells of the inner epithelial sheath surrounding the trophocytes and into the prefollicular cells that encompass the oocytes. During this final differentiation, the trophocytes lose their cell membranes and become syncytial. Oocytes remain cellular and most of them (termed arrested oocytes) do not grow. In the ovarioles of older females, one oocyte encompassed by its follicle cells starts growing, still connected to the syncytial tropharium by a nutritive cord. After the short phase of previtellogenesis alone, the oocyte enters its vitellogenic the growth phase in the vitellarium. At that time, the second oocyte may enter the vitellarium and start its previtellogenic growth. In the light of the obtained results, the phylogeny of psyllids, as well as phylogenetic relationships between taxa of Hemiptera: Sternorrhyncha are discussed. Copyright © 2016 Elsevier Ltd. All rights reserved.

Midgut morphological changes and autophagy during metamorphosis in sand flies.

PubMed

Malta, Juliana; Heerman, Matthew; Weng, Ju Lin; Fernandes, Kenner M; Martins, Gustavo Ferreira; Ramalho-Ortigão, Marcelo

2017-06-01

During metamorphosis, holometabolous insects undergo significant remodeling of their midgut and become able to cope with changes in dietary requirements between larval and adult stages. At this stage, insects must be able to manage and recycle available food resources in order to develop fully into adults, especially when no nutrients are acquired from the environment. Autophagy has been previously suggested to play a crucial role during metamorphosis of the mosquito. Here, we investigate the overall morphological changes of the midgut of the sand fly during metamorphosis and assess the expression profiles of the autophagy-related genes ATG1, ATG6, and ATG8, which are associated with various steps of the autophagic process. Morphological changes in the midgut start during the fourth larval instar, with epithelial degeneration followed by remodeling via the differentiation of regenerative cells in pre-pupal and pupal stages. The changes in the midgut epithelium are paired with the up-regulation of ATG1, ATG6 and ATG8 during the larva-adult transition. Vein, a putative epidermal growth factor involved in regulating epithelial midgut regeneration, is also up-regulated. Autophagy has further been confirmed in sand flies via the presence of autophagosomes residing within the cytoplasmic compartment of the pupal stages. An understanding of the underlying mechanisms of this process should aid the future management of this neglected tropical vector.

Osteograft, plastic material for regenerative medicine

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

Zaidman, A. M., E-mail: AZaydman@niito.ru; Korel, A. V., E-mail: AKorel@niito.ru; Shchelkunova, E. I., E-mail: EShelkunova@niito.ru

Creating tissue-engineering constructs based on the mechanism of cartilage-bone evolution is promising for traumatology and orthopedics. Such a graft was obtained from a chondrograft by transdifferentiation. The hondrograft placed in osteogenic medium is undergoing osteogenic differentiation for 14–30 days. Tissue specificity of the osteograft was studied by morphology, immunohistochemistry, electron microscopy, and the expression of the corresponding genes was estimated. The expression of osteonectin, fibronectin, collagen of type I, izolektin and CD 44 is determined. Alkaline phosphatase and matrix vesicles are determined in osteoblasts. Calcificates are observed in the matrix. Chondrogenic proteins expression is absent. These findings evidence the tissuemore » specificity of the developed osteograft.« less

EF5 in Finding Oxygen in Tumor Cells of Patients Who Are Undergoing Surgery or Biopsy for Cervical, Endometrial, or Ovarian Epithelial Cancer

ClinicalTrials.gov

2013-01-15

Primary Peritoneal Cavity Cancer; Stage I Endometrial Carcinoma; Stage I Ovarian Epithelial Cancer; Stage IA Cervical Cancer; Stage IB Cervical Cancer; Stage II Endometrial Carcinoma; Stage II Ovarian Epithelial Cancer; Stage IIA Cervical Cancer; Stage IIB Cervical Cancer; Stage III Cervical Cancer; Stage III Endometrial Carcinoma; Stage III Ovarian Epithelial Cancer; Stage IV Endometrial Carcinoma; Stage IV Ovarian Epithelial Cancer; Stage IVA Cervical Cancer; Stage IVB Cervical Cancer

Minimally Invasive Surgical Staging in Early-stage Ovarian Carcinoma: A Systematic Review and Meta-analysis.

PubMed

Bogani, Giorgio; Borghi, Chiara; Leone Roberti Maggiore, Umberto; Ditto, Antonino; Signorelli, Mauro; Martinelli, Fabio; Chiappa, Valentina; Lopez, Carlos; Sabatucci, Ilaria; Scaffa, Cono; Indini, Alice; Ferrero, Simone; Lorusso, Domenica; Raspagliesi, Francesco

Few studies investigated the efficacy and safety of minimally invasive surgery for the treatment of early-stage epithelial ovarian cancer (eEOC). In this context, we aimed to review the current evidence comparing laparoscopy and the laparotomic approach for staging procedures in eEOC. This systematic review was registered in the International Prospective Register of Systematic Reviews. Overall, 3065 patients were included: 1450 undergoing laparoscopy and 1615 undergoing laparotomic staging. Patients undergoing laparoscopy experienced a longer (but not statistically significant) operative time (weighted mean difference [WMD] = 28.3 minutes; 95% confidence interval [CI], -2.59 to 59.2), a lower estimated blood loss (WMD = -156.5 mL; 95% CI, -216.4 to -96.5), a shorter length of hospital stay (WMD = -3.7 days; 95% CI, -5.2 to -2.1), and a lower postoperative complication rate (odds ratio [OR] = 0.48; 95% CI, 0.29-0.81) than patients undergoing laparotomy. The upstaging (OR = 0.81; 95% CI, 0.55-1.20) and cyst rupture (OR = 1.32; 95% CI, 0.52-3.38) rates were similar between groups. Laparoscopic staging is associated with a shorter time to chemotherapy than laparotomic procedures (WMD = -5.16 days; 95% CI, -8.68 to -1.64). Survival outcomes were not influenced by the route of surgery. Pooled data suggested that the minimally invasive surgical approach is equivalent to laparotomy for the treatment of eEOC and may be superior in terms of perioperative outcomes. However, because of the low level of evidence of the included studies, further randomized trials are warranted. Copyright © 2017 AAGL. Published by Elsevier Inc. All rights reserved.

Dental care protocol based on visual supports for children with autism spectrum disorders

PubMed Central

Mastroberardino, Stefano; Campus, Guglielmo; Olivari, Benedetta; Faggioli, Raffaella; Lenti, Carlo; Strohmenger, Laura

2015-01-01

Background Subjects with Autism Spectrum Disorders (ASDs) have often difficulties to accept dental treatments. The aim of this study is to propose a dental care protocol based on visual supports to facilitate children with ASDs to undergo to oral examination and treatments. Material and Methods 83 children (age range 6-12 years) with a signed consent form were enrolled; intellectual level, verbal fluency and cooperation grade were evaluated. Children were introduced into a four stages path in order to undergo: an oral examination (stage 1), a professional oral hygiene session (stage 2), sealants (stage 3), and, if necessary, a restorative treatment (stage 4). Each stage came after a visual training, performed by a psychologist (stage 1) and by parents at home (stages 2, 3 and 4). Association between acceptance rates at each stage and gender, intellectual level, verbal fluency and cooperation grade was tested with chi-square test if appropriate. Results Seventy-seven (92.8%) subjects overcame both stage 1 and 2. Six (7.2%) refused stage 3 and among the 44 subjects who need restorative treatments, only three refused it. The acceptance rate at each stage was statistically significant associated to the verbal fluency (p=0.02; p=0.04; p=0.01, respectively for stage 1, 3 and 4). In stage 2 all subjects accepted to move to the next stage. The verbal/intellectual/cooperation dummy variable was statistically associated to the acceptance rate (p<0.01). Conclusions The use of visual supports has shown to be able to facilitate children with ASDs to undergo dental treatments even in non-verbal children with a low intellectual level, underlining that behavioural approach should be used as the first strategy to treat patients with ASDs in dental setting. Key words:Autism spectrum disorders, behaviour management, paediatric dentistry, visual learning methods. PMID:26241453

Thyroid remnant ablation success and disease outcome in stage III or IV differentiated thyroid carcinoma: recombinant human thyrotropin versus thyroid hormone withdrawal.

PubMed

Vallejo Casas, Juan A; Mena Bares, Luisa M; Gálvez Moreno, Maria A; Moreno Ortega, Estefanía; Marlowe, Robert J; Maza Muret, Francisco R; Albalá González, María D

2016-06-01

Most publications to date compare outcomes after post-surgical thyroid remnant ablation stimulated by recombinant human thyrotropin (rhTSH) versus thyroid hormone withholding/withdrawal (THW) in low-recurrence risk differentiated thyroid carcinoma (DTC) patients. We sought to perform this comparison in high-risk patients. We retrospectively analyzed ~9-year single-center experience in 70 consecutive adults with initial UICC (Union for International Cancer Control) stage III/IV, M0 DTC undergoing rhTSH-aided (N.=54) or THW-aided (N.=16) high-activity ablation. Endpoints included ablation success and DTC outcome. Assessed ≥1 year post-ablation, ablation success comprised a) no visible scintigraphic thyroid bed uptake or pathological extra-thyroidal uptake; b) undetectable stimulated serum thyroglobulin (Tg) without interfering autoantibodies; c) both criteria. DTC outcome, determined at the latest visit, comprised either 1) "no evidence of disease" (NED): undetectable Tg, negative Tg autoantibodies, negative most recent whole-body scan, no suspicious findings clinically, on neck ultrasonography, or on other imaging; 2) persistent disease: failure to attain NED; or 3) recurrence: loss of NED. After the first ablative activity, ablation success by scintigraphic plus biochemical criteria was 64.8% in rhTSH patients, 56.3% in THW patients (P=NS). After 3.5-year versus 6.2-year median follow-up (P<0.05), DTC outcomes were NED, 85.2%, persistent disease, 13.0%, recurrence, 1.9%, in the rhTSH group and NED, 87.5%, persistent or recurrent disease, 6.3% each, in the THW group (P=NS). In patients with initial stage III/IV, M0 DTC, rhTSH-aided and THW-assisted ablation were associated with comparable remnant eradication or DTC cure rates.

A large scale Plasmodium vivax- Saimiri boliviensis trophozoite-schizont transition proteome

PubMed Central

Lapp, Stacey A.; Barnwell, John W.; Galinski, Mary R.

2017-01-01

Plasmodium vivax is a complex protozoan parasite with over 6,500 genes and stage-specific differential expression. Much of the unique biology of this pathogen remains unknown, including how it modifies and restructures the host reticulocyte. Using a recently published P. vivax reference genome, we report the proteome from two biological replicates of infected Saimiri boliviensis host reticulocytes undergoing transition from the late trophozoite to early schizont stages. Using five database search engines, we identified a total of 2000 P. vivax and 3487 S. boliviensis proteins, making this the most comprehensive P. vivax proteome to date. PlasmoDB GO-term enrichment analysis of proteins identified at least twice by a search engine highlighted core metabolic processes and molecular functions such as glycolysis, translation and protein folding, cell components such as ribosomes, proteasomes and the Golgi apparatus, and a number of vesicle and trafficking related clusters. Database for Annotation, Visualization and Integrated Discovery (DAVID) v6.8 enriched functional annotation clusters of S. boliviensis proteins highlighted vesicle and trafficking-related clusters, elements of the cytoskeleton, oxidative processes and response to oxidative stress, macromolecular complexes such as the proteasome and ribosome, metabolism, translation, and cell death. Host and parasite proteins potentially involved in cell adhesion were also identified. Over 25% of the P. vivax proteins have no functional annotation; this group includes 45 VIR members of the large PIR family. A number of host and pathogen proteins contained highly oxidized or nitrated residues, extending prior trophozoite-enriched stage observations from S. boliviensis infections, and supporting the possibility of oxidative stress in relation to the disease. This proteome significantly expands the size and complexity of the known P. vivax and Saimiri host iRBC proteomes, and provides in-depth data that will be valuable for ongoing research on this parasite’s biology and pathogenesis. PMID:28829774

Current strategies with 1-stage prosthetic breast reconstruction

PubMed Central

2015-01-01

Background 1-stage prosthetic breast reconstruction is gaining traction as a preferred method of breast reconstruction in select patients who undergo mastectomy for cancer or prevention. Methods Critical elements to the procedure including patient selection, technique, surgical judgment, and postoperative care were reviewed. Results Outcomes series reveal that in properly selected patients, direct-to-implant (DTI) reconstruction has similar low rates of complications and high rates of patient satisfaction compared to traditional 2-stage reconstruction. Conclusions 1-stage prosthetic breast reconstruction may be the procedure of choice in select patients undergoing mastectomy. Advantages include the potential for the entire reconstructive process to be complete in one surgery, the quick return to normal activities, and lack of donor site morbidity. PMID:26005643

Ultrafast hydrogen exchange reveals specific structural events during the initial stages of folding of cytochrome c.

PubMed

Fazelinia, Hossein; Xu, Ming; Cheng, Hong; Roder, Heinrich

2014-01-15

Many proteins undergo a sharp decrease in chain dimensions during early stages of folding, prior to the rate-limiting step in folding. However, it remains unclear whether compact states are the result of specific folding events or a general hydrophobic collapse of the poly peptide chain driven by the change in solvent conditions. To address this fundamental question, we extended the temporal resolution of NMR-detected H/D exchange labeling experiments into the microsecond regime by adopting a microfluidics approach. By observing the competition between H/D exchange and folding as a function of labeling pH, coupled with direct measurement of exchange rates in the unfolded state, we were able to monitor hydrogen-bond formation for over 50 individual backbone NH groups within the initial 140 microseconds of folding of horse cytochrome c. Clusters of solvent-shielded amide protons were observed in two α-helical segments in the C-terminal half of the protein, while the N-terminal helix remained largely unstructured, suggesting that proximity in the primary structure is a major factor in promoting helix formation and association at early stages of folding, while the entropically more costly long-range contacts between the N- and C-terminal helices are established only during later stages. Our findings clearly indicate that the initial chain condensation in cytochrome c is driven by specific interactions among a subset of α-helical segments rather than a general hydrophobic collapse.

Specific serine-proline phosphorylation and glycogen synthase kinase 3β-directed subcellular targeting of stathmin 3/Sclip in neurons.

PubMed

Devaux, Sara; Poulain, Fabienne E; Devignot, Véronique; Lachkar, Sylvie; Irinopoulou, Theano; Sobel, André

2012-06-22

During nervous system development, neuronal growth, migration, and functional morphogenesis rely on the appropriate control of the subcellular cytoskeleton including microtubule dynamics. Stathmin family proteins play major roles during the various stages of neuronal differentiation, including axonal growth and branching, or dendritic development. We have shown previously that stathmins 2 (SCG10) and 3 (SCLIP) fulfill distinct, independent and complementary regulatory roles in axonal morphogenesis. Although the two proteins have been proposed to display the four conserved phosphorylation sites originally identified in stathmin 1, we show here that they possess distinct phosphorylation sites within their specific proline-rich domains (PRDs) that are differentially regulated by phosphorylation by proline-directed kinases involved in the control of neuronal differentiation. ERK2 or CDK5 phosphorylate the two proteins but with different site specificities. We also show for the first time that, unlike stathmin 2, stathmin 3 is a substrate for glycogen synthase kinase (GSK) 3β both in vitro and in vivo. Interestingly, stathmin 3 phosphorylated at its GSK-3β target site displays a specific subcellular localization at neuritic tips and within the actin-rich peripheral zone of the growth cone of differentiating hippocampal neurons in culture. Finally, pharmacological inhibition of GSK-3β induces a redistribution of stathmin 3, but not stathmin 2, from the periphery toward the Golgi region of neurons. Stathmin proteins can thus be either regulated locally or locally targeted by specific phosphorylation, each phosphoprotein of the stathmin family fulfilling distinct and specific roles in the control of neuronal differentiation.

Rotating microgravity-bioreactor cultivation enhances the hepatic differentiation of mouse embryonic stem cells on biodegradable polymer scaffolds.

PubMed

Wang, Yingjie; Zhang, Yunping; Zhang, Shichang; Peng, Guangyong; Liu, Tao; Li, Yangxin; Xiang, Dedong; Wassler, Michael J; Shelat, Harnath S; Geng, Yongjian

2012-11-01

Embryonic stem (ES) cells are pluripotent cells that are capable of differentiating all the somatic cell lineages, including those in the liver tissue. We describe the generation of functional hepatic-like cells from mouse ES (mES) cells using a biodegradable polymer scaffold and a rotating bioreactor that allows simulated microgravity. Cells derived from ES cells cultured in the three-dimensional (3D) culture system with exogenous growth factors and hormones can differentiate into hepatic-like cells with morphologic characteristics of typical mature hepatocytes. Reverse-transcription polymerase chain-reaction testing, Western blot testing, immunostaining, and flow cytometric analysis show that these cells express hepatic-specific genes and proteins during differentiation. Differentiated cells on scaffolds further exhibit morphologic traits and biomarkers characteristic of liver cells, including albumin production, cytochrome P450 activity, and low-density lipoprotein uptake. When these stem cell-bearing scaffolds are transplanted into severe combined immunodeficient mice, the 3D constructs remained viable, undergoing further differentiation and maturation of hepatic-like cells in vivo. In conclusion, the growth and differentiation of ES cells in a biodegradable polymer scaffold and a rotating microgravity bioreactor can yield functional and organizational hepatocytes useful for research involving bioartificial liver and engineered liver tissue.

Uhrf1 controls the self-renewal versus differentiation of hematopoietic stem cells by epigenetically regulating the cell-division modes

PubMed Central

Zhao, Jingyao; Chen, Xufeng; Song, Guangrong; Zhang, Jiali; Liu, Haifeng; Liu, Xiaolong

2017-01-01

Hematopoietic stem cells (HSCs) are able to both self-renew and differentiate. However, how individual HSC makes the decision between self-renewal and differentiation remains largely unknown. Here we report that ablation of the key epigenetic regulator Uhrf1 in the hematopoietic system depletes the HSC pool, leading to hematopoietic failure and lethality. Uhrf1-deficient HSCs display normal survival and proliferation, yet undergo erythroid-biased differentiation at the expense of self-renewal capacity. Notably, Uhrf1 is required for the establishment of DNA methylation patterns of erythroid-specific genes during HSC division. The expression of these genes is enhanced in the absence of Uhrf1, which disrupts the HSC-division modes by promoting the symmetric differentiation and suppressing the symmetric self-renewal. Moreover, overexpression of one of the up-regulated genes, Gata1, in HSCs is sufficient to phenocopy Uhrf1-deficient HSCs, which show impaired HSC symmetric self-renewal and increased differentiation commitment. Taken together, our findings suggest that Uhrf1 controls the self-renewal versus differentiation of HSC through epigenetically regulating the cell-division modes, thus providing unique insights into the relationship among Uhrf1-mediated DNA methylation, cell-division mode, and HSC fate decision. PMID:27956603

Regulatory RNA Key Player in p53-Mediated Apoptosis in Embryonic Stem Cells | Center for Cancer Research

Cancer.gov

Embryonic stem cells (ESCs) must maintain the integrity of their genomes or risk passing potentially deleterious mutations on to numerous tissues. Thus, ESCs have a unique genome surveillance system and easily undergo apoptosis or differentiation when DNA damage is detected. The protein p53 is known to promote differentiation in mouse ESCs (mESCs), but its role in DNA damage-induced apoptosis (DIA) is unclear. p53 may have a pro-apoptotic function since it can regulate apoptotic genes in embryonal cells. Given that ESCs have a distinct transcriptional program, Jing Huang, Ph.D., of CCR’s Laboratory of Cancer Biology and Genetics, and his colleagues wondered whether p53 might regulate DIA in ESCs by utilizing the ESC-specific expression program.

Caspase inhibitors affect the kinetics and dimensions of tracheary elements in xylogenic Zinnia (Zinnia elegans) cell cultures.

PubMed

Twumasi, Peter; Iakimova, Elena T; Qian, Tian; van Ieperen, Wim; Schel, Jan H N; Emons, Anne Mie C; van Kooten, Olaf; Woltering, Ernst J

2010-08-06

The xylem vascular system is composed of fused dead, hollow cells called tracheary elements (TEs) that originate through trans-differentiation of root and shoot cambium cells. TEs undergo autolysis as they differentiate and mature. The final stage of the formation of TEs in plants is the death of the involved cells, a process showing some similarities to programmed cell death (PCD) in animal systems. Plant proteases with functional similarity to proteases involved in mammalian apoptotic cell death (caspases) are suggested as an integral part of the core mechanism of most PCD responses in plants, but participation of plant caspase-like proteases in TE PCD has not yet been documented. Confocal microscopic images revealed the consecutive stages of TE formation in Zinnia cells during trans-differentiation. Application of the caspase inhibitors Z-Asp-CH2-DCB, Ac-YVAD-CMK and Ac-DEVD-CHO affected the kinetics of formation and the dimensions of the TEs resulting in a significant delay of TE formation, production of larger TEs and in elimination of the 'two-wave' pattern of TE production. DNA breakdown and appearance of TUNEL-positive nuclei was observed in xylogenic cultures and this was suppressed in the presence of caspase inhibitors. To the best of our knowledge this is the first report showing that caspase inhibitors can modulate the process of trans-differentiation in Zinnia xylogenic cell cultures. As caspase inhibitors are closely associated with cell death inhibition in a variety of plant systems, this suggests that the altered TE formation results from suppression of PCD. The findings presented here are a first step towards the use of appropriate PCD signalling modulators or related molecular genetic strategies to improve the hydraulic properties of xylem vessels in favour of the quality and shelf life of plants or plant parts.

Donor-Dependent and Other Nondefined Factors Have Greater Influence on the Hepatic Phenotype Than the Starting Cell Type in Induced Pluripotent Stem Cell Derived Hepatocyte-Like Cells.

PubMed

Heslop, James A; Kia, Richard; Pridgeon, Christopher S; Sison-Young, Rowena L; Liloglou, Triantafillos; Elmasry, Mohamed; Fenwick, Stephen W; Mills, John S; Kitteringham, Neil R; Goldring, Chris E; Park, Bong K

2017-05-01

Drug-induced liver injury is the greatest cause of post-marketing drug withdrawal; therefore, substantial resources are directed toward triaging potentially dangerous new compounds at all stages of drug development. One of the major factors preventing effective screening of new compounds is the lack of a predictive in vitro model of hepatotoxicity. Primary human hepatocytes offer a metabolically relevant model for which the molecular initiating events of hepatotoxicity can be examined; however, these cells vary greatly between donors and dedifferentiate rapidly in culture. Induced pluripotent stem cell (iPSC)-derived hepatocyte-like cells (HLCs) offer a reproducible, physiologically relevant and genotypically normal model cell; however, current differentiation protocols produce HLCs with a relatively immature phenotype. During the reprogramming of somatic cells, the epigenome undergoes dramatic changes; however, this "resetting" is a gradual process, resulting in an altered differentiation propensity, skewed toward the lineage of origin, particularly in early passage cultures. We, therefore, performed a comparison of human hepatocyte- and dermal fibroblast-derived iPSCs, assessing the impact of epigenetic memory at all stages of HLC differentiation. These results provide the first isogenic assessment of the starting cell type in human iPSC-derived HLCs. Despite a trend toward improvement in hepatic phenotype in albumin secretion and gene expression, few significant differences in hepatic differentiation capacity were found between hepatocyte and fibroblast-derived iPSCs. We conclude that the donor and inter-clonal differences have a greater influence on the hepatocyte phenotypic maturity than the starting cell type. Therefore, it is not necessary to use human hepatocytes for generating iPSC-derived HLCs. Stem Cells Translational Medicine 2017;6:1321-1331. © 2017 The Authors Stem Cells Translational Medicine published by Wiley Periodicals, Inc. on behalf of AlphaMed Press.

Zebrafish no isthmus reveals a role for pax2.1 in tubule differentiation and patterning events in the pronephric primordia.

PubMed

Majumdar, A; Lun, K; Brand, M; Drummond, I A

2000-05-01

Pax genes are important developmental regulators and function at multiple stages of vertebrate kidney organogenesis. In this report, we have used the zebrafish pax2.1 mutant no isthmus to investigate the role for pax2.1 in development of the pronephros. We demonstrate a requirement for pax2.1 in multiple aspects of pronephric development including tubule and duct epithelial differentiation and cloaca morphogenesis. Morphological analysis demonstrates that noi(- )larvae specifically lack pronephric tubules while glomerular cell differentiation is unaffected. In addition, pax2.1 expression in the lateral cells of the pronephric primordium is required to restrict the domains of Wilms' tumor suppressor (wt1) and vascular endothelial growth factor (VEGF) gene expression to medial podocyte progenitors. Ectopic podocyte-specific marker expression in pronephric duct cells correlates with loss of expression of the pronephric tubule and duct-specific markers mAb 3G8 and a Na(+)/K(+) ATPase (&agr;)1 subunit. The results suggest that the failure in pronephric tubule differentiation in noi arises from a patterning defect during differentiation of the pronephric primordium and that mutually inhibitory regulatory interactions play an important role in defining the boundary between glomerular and tubule progenitors in the forming nephron.

Early In Vitro Differentiation of Mouse Definitive Endoderm Is Not Correlated with Progressive Maturation of Nuclear DNA Methylation Patterns

PubMed Central

Tajbakhsh, Jian; Gertych, Arkadiusz; Fagg, W. Samuel; Hatada, Seigo; Fair, Jeffrey H.

2011-01-01

The genome organization in pluripotent cells undergoing the first steps of differentiation is highly relevant to the reprogramming process in differentiation. Considering this fact, chromatin texture patterns that identify cells at the very early stage of lineage commitment could serve as valuable tools in the selection of optimal cell phenotypes for regenerative medicine applications. Here we report on the first-time use of high-resolution three-dimensional fluorescence imaging and comprehensive topological cell-by-cell analyses with a novel image-cytometrical approach towards the identification of in situ global nuclear DNA methylation patterns in early endodermal differentiation of mouse ES cells (up to day 6), and the correlations of these patterns with a set of putative markers for pluripotency and endodermal commitment, and the epithelial and mesenchymal character of cells. Utilizing this in vitro cell system as a model for assessing the relationship between differentiation and nuclear DNA methylation patterns, we found that differentiating cell populations display an increasing number of cells with a gain in DNA methylation load: first within their euchromatin, then extending into heterochromatic areas of the nucleus, which also results in significant changes of methylcytosine/global DNA codistribution patterns. We were also able to co-visualize and quantify the concomitant stochastic marker expression on a per-cell basis, for which we did not measure any correlation to methylcytosine loads or distribution patterns. We observe that the progression of global DNA methylation is not correlated with the standard transcription factors associated with endodermal development. Further studies are needed to determine whether the progression of global methylation could represent a useful signature of cellular differentiation. This concept of tracking epigenetic progression may prove useful in the selection of cell phenotypes for future regenerative medicine applications. PMID:21779341

Gender and Bladder Cancer: A Collaborative Review of Etiology, Biology, and Outcomes.

PubMed

Dobruch, Jakub; Daneshmand, Siamak; Fisch, Margit; Lotan, Yair; Noon, Aidan P; Resnick, Matthew J; Shariat, Shahrokh F; Zlotta, Alexandre R; Boorjian, Stephen A

2016-02-01

The incidence of bladder cancer is three to four times greater in men than in women. However, women are diagnosed with more advanced disease at presentation and have less favorable outcomes after treatment. To review the literature on potential biologic mechanisms underlying differential gender risk for bladder cancer, and evidence regarding gender disparities in bladder cancer presentation, management, and outcomes. A literature search of English-language publications that included an analysis of the association of gender with bladder cancer was performed using Pubmed. Ninety-seven articles were selected for analysis with the consensus of all authors. It has been shown that the gender difference in bladder cancer incidence is independent of differences in exposure risk, including smoking status. Potential molecular mechanisms include disparate metabolism of carcinogens by hepatic enzymes between men and women, resulting in differential exposure of the urothelium to carcinogens. In addition, the activity of the sex steroid hormone pathway may play a role in bladder cancer development, with demonstration that both androgens and estrogens have biologic effects in bladder cancer in vitro and in vivo. Importantly, gender differences exist in the timeliness and completeness of hematuria evaluation, with women experiencing a significantly greater delay in urologic referral and undergoing guideline-concordant imaging less frequently. Correspondingly, women have more advanced tumors at the time of bladder cancer diagnosis. Interestingly, higher cancer-specific mortality has been noted among women even after adjusting for tumor stage and treatment modality. Numerous potential biologic and epidemiologic factors probably underlie the gender differences observed for bladder cancer incidence, stage at diagnosis, and outcomes. Continued evaluation to define clinical applications for manipulation of the sex steroid pathway and to improve the standardization of hematuria evaluation in women may improve future patient outcomes and reduce these disparities. We describe the scientific basis and clinical evidence to explain the greater incidence of bladder cancer in men and the adverse presentation and outcomes for this disease in women. We identify goals for improving patient survival and reducing gender disparities in bladder cancer. Copyright © 2015 European Association of Urology. Published by Elsevier B.V. All rights reserved.

Oncologic outcomes in patients with nonurothelial bladder cancer.

PubMed

Patel, Sanjay G; Weiner, Adam Benjamin; Keegan, Kirk; Morgan, Todd

2018-01-01

We aimed to evaluate the relative prognostic impact of the most common variant histologies on disease-specific survival (DSS) in patients undergoing radical cystectomy. The Surveillance, Epidemiology, and End Result database was used to identify patients who underwent radical cystectomy for bladder cancer from 1990 to 2007. Patients with urothelial cell carcinoma (UCC), squamous cell carcinoma (SCC), adenocarcinoma (AC), sarcoma, small cell carcinoma, signet ring carcinoma, and spindle cell carcinoma were included in the study. Multivariable analysis was performed using Cox proportional hazards model to assess independent predictors of disease-specific survival (DSS). Mortality rates were estimated using Kaplan-Meier analyses. A total of 14,130 patients met inclusion criteria with the following histologies: UCC (90.1%), SCC (4.6%), AC, (2.3%), sarcoma (0.8%), small cell carcinoma (0.8%), signet ring carcinoma (0.5%), and spindle cell carcinoma (0.9%). Three-year DSS was most favorable in patients with UCC (63.7%; 95% confidence interval [62.9%-64.8%]) and AC (65.3% [59.3%-70.6%]), whereas 3-year DSS was the least favorable for small cell carcinoma (41.6% [31.3%-51.6%]) and sarcoma (45.4% [35.1%-55.1%]). In the multivariable analysis, independent predictors of DSS were age, marital status, grade, T-stage, N-stage, and variant histology. With respect to UCC, there was an increased risk of disease-specific death associated with all variants except AC. Sarcoma and spindle cell carcinoma were associated with the highest risk of death. With the exception of AC, the most common variant bladder cancer histologies are all independently associated with worse DSS relative to UCC in patients undergoing radical cystectomy.

Sulforaphane inhibits mitotic clonal expansion during adipogenesis through cell cycle arrest.

PubMed

Choi, Kyeong-Mi; Lee, Youn-Sun; Sin, Dong-Mi; Lee, Seunghyun; Lee, Mi Kyeong; Lee, Yong-Moon; Hong, Jin-Tae; Yun, Yeo-Pyo; Yoo, Hwan-Soo

2012-07-01

Obesity is a risk factor for numerous metabolic disorders such as type 2 diabetes, hypertension, and coronary heart disease. Adipocyte differentiation is triggered by adipocyte hyperplasia, which leads to obesity. In this study, the inhibitory effect of sulforaphane, an isothiocyanate, on adipogenesis in 3T3-L1 cells was investigated. Sulforaphane decreased the accumulation of lipid droplets stained with Oil Red O and inhibited the elevation of triglycerides in the adipocytes (half-maximal inhibitory concentration = 7.3 µmol/l). The expression of peroxisome proliferator-activated receptor γ (PPARγ) and CCAAT/enhancer-binding protein α (C/EBPα), major transcription factors for adipocyte differentiation, was significantly reduced by sulforaphane. The major effects of sulforaphane on the inhibition of adipocyte differentiation occurred during the early stage of adipogenesis. Thus, the expression of C/EBPβ, an early-stage biomarker of adipogenesis, decreased in a concentration-dependent manner when the adipocytes were exposed to sulforaphane (0, 5, 10, and 20 µmol/l). The proliferation of adipocytes treated with 20 µmol/l sulforaphane for 24 and 48 h was also suppressed. These results indicate that sulforaphane may specifically affect mitotic clonal expansion to inhibit adipocyte differentiation. Sulforaphane arrested the cell cycle at the G(0)/G(1) phase, increased p27 expression, and decreased retinoblastoma (Rb) phosphorylation. Additionally, sulforaphane modestly decreased the phosphorylation of ERK1/2 and Akt. Our results indicate that the inhibition of early-stage adipocyte differentiation by sulforaphane may be associated with cell cycle arrest at the G(0)/G(1) phase through upregulation of p27 expression.

Influence of wellness education on first-line icotinib hydrochloride patients with stage IV non-small cell lung cancer and their family caregivers.

PubMed

Yanwei, Li; Minghui, Fang; Manman, Quan; Zhuchun, Yan; Dongying, Liu; Zhanyu, Pan

2018-04-11

This study aims to examine the effects of wellness education (WE) intervention on the behavioral change, psychological status, performance status on patients with stage IV non-small cell lung cancer (NSCLC) undergoing icotinib hydrochloride treatment and their relationships with family caregivers. We conducted an intervention study involving 126 individuals with confirmed activating epidermal growth factor receptor mutation-positive stage IV NSCLC who received icotinib hydrochloride as first-line therapy between January 2014 and January 2016; their caregivers were also included in the study. For a period of 12 weeks, participants were randomly assigned into WE and control groups. The patients and family members in the WE group were provided with WE information about treatment, diet, social needs, rehabilitation, physical/mental health education, communication strategies, and patient care advice at least 3 times per week during treatment. Qualitative feedback of the participants was recorded during the intervention. Food Composition Database, the Family Environment Scale, patients/caregivers quality-of-life (Functional Assessment of Cancer Therapy-Lung/Caregiver Quality of Life Index-Cancer Scale), and Hospital Anxiety and Depression Scale (HADS) were measured at baseline and for 12 weeks. Data were analyzed to compare the different outcomes. Of the 126 caregivers (64 WE and 62 control), 120 completed the study. We observed significant differences between the WE group and control group with respect to low daily calorie intake (31.0% vs 77.4%, p < 0.05), smoking cessationaaa and awareness of cancer (85.48% vs 100%, p < 0.05). The WE group showed high ratings on awareness of cancer risk and benefit, as well as confidence relating to the behaviors of healthful diet and self-motivation to conduct cancer test. Family caregivers had high ratings on 30-minute daily moderate physical activity (p > 0.05). After 12 weeks, WE intervention had improved scores on Functional Assessment of Cancer Therapy-Lung-EWB and Caregiver Quality of Life Index-Cancer Scale adaptation. In addition, the patients also showed improvements in HADS. WE interventions in patients with stage IV NSCLC undergoing icotinib hydrochloride treatment and their family resulted in strong intentions to engage in health-promoting behaviors related to physical activity, smoking cessationaaa, and nutrition at the treatment period. WE intervention is a viable way to improve quality of life and HADS. Findings from this study suggest that WE interventions in patients' family with stage IV NSCLC undergoing icotinib hydrochloride treatment are significant improvements in both HADS and quality of life. These data also indicate that lung cancer disparities are unlikely to be associated with differential willingness to receive care but that Chinese may perceive financial and insurance ebarriers to treatment. Copyright © 2018 Elsevier Ltd. All rights reserved.

Health technology assessment of medical devices: a survey of non-European union agencies.

PubMed

Ciani, Oriana; Wilcher, Britni; Blankart, Carl Rudolf; Hatz, Maximilian; Rupel, Valentina Prevolnik; Erker, Renata Slabe; Varabyova, Yauheniya; Taylor, Rod S

2015-01-01

The aim of this study was to review and compare current health technology assessment (HTA) activities for medical devices across non-European Union HTA agencies. HTA activities for medical devices were evaluated from three perspectives: organizational structure, processes, and methods. Agencies were primarily selected upon membership of existing HTA networks. The data collection was performed in two stages: stage 1-agency Web-site assessment using a standardized questionnaire, followed by review and validation of the collected data by a representative of the agency; and stage 2-semi-structured telephone interviews with key informants of a sub-sample of agencies. In total, thirty-six HTA agencies across twenty non-EU countries assessing medical devices were included. Twenty-seven of thirty-six (75 percent) agencies were judged at stage 1 to have adopted HTA-specific approaches for medical devices (MD-specific agencies) that were largely organizational or procedural. There appeared to be few differences in the organization, process and methods between MD-specific and non-MD-specific agencies. Although the majority (69 percent) of both categories of agency had specific methods guidance or policy for evidence submission, only one MD-specific agency had developed methodological guidelines specific to medical devices. In stage 2, many MD-specific agencies cited insufficient resources (budget, skilled employees), lack of coordination (between regulator and reimbursement bodies), and the inability to generalize findings from evidence synthesis to be key challenges in the HTA of medical devices. The lack of evidence for differentiation in scientific methods for HTA of devices raises the question of whether HTA needs to develop new methods for medical devices but rather adapt existing methodological approaches. In contrast, organizational and/or procedural adaptation of existing HTA agency frameworks to accommodate medical devices appear relatively commonplace.

Inferring rules of lineage commitment in haematopoiesis.

PubMed

Pina, Cristina; Fugazza, Cristina; Tipping, Alex J; Brown, John; Soneji, Shamit; Teles, Jose; Peterson, Carsten; Enver, Tariq

2012-02-19

How the molecular programs of differentiated cells develop as cells transit from multipotency through lineage commitment remains unexplored. This reflects the inability to access cells undergoing commitment or located in the immediate vicinity of commitment boundaries. It remains unclear whether commitment constitutes a gradual process, or else represents a discrete transition. Analyses of in vitro self-renewing multipotent systems have revealed cellular heterogeneity with individual cells transiently exhibiting distinct biases for lineage commitment. Such systems can be used to molecularly interrogate early stages of lineage affiliation and infer rules of lineage commitment. In haematopoiesis, population-based studies have indicated that lineage choice is governed by global transcriptional noise, with self-renewing multipotent cells reversibly activating transcriptome-wide lineage-affiliated programs. We examine this hypothesis through functional and molecular analysis of individual blood cells captured from self-renewal cultures, during cytokine-driven differentiation and from primary stem and progenitor bone marrow compartments. We show dissociation between self-renewal potential and transcriptome-wide activation of lineage programs, and instead suggest that multipotent cells experience independent activation of individual regulators resulting in a low probability of transition to the committed state.

Raman spectroscopy of blood serum for Alzheimer's disease diagnostics: specificity relative to other types of dementia

PubMed Central

Ryzhikova, Elena; Kazakov, Oleksandr; Halamkova, Lenka; Celmins, Dzintra; Malone, Paula; Molho, Eric; Zimmerman, Earl A.; Lednev, Igor K.

2015-01-01

The key moment for efficiently and accurately diagnosing dementia occurs during the early stages. This is particularly true for Alzheimer's disease (AD). In this proof-of-concept study, we applied near infrared (NIR) Raman microspectroscopy of blood serum together with advanced multivariate statistics for the selective identification of AD. We analyzed data from 20 AD patients, 18 patients with other neurodegenerative dementias (OD) and 10 healthy control (HC) subjects. NIR Raman microspectroscopy differentiated patients with more than 95% sensitivity and specificity. We demonstrated the high discriminative power of artificial neural network (ANN) classification models, thus revealing the high potential of this developed methodology for the differential diagnosis of AD. Raman spectroscopic, blood-based tests may aid clinical assessments for the effective and accurate differential diagnosis of AD, decrease the labor, time and cost of diagnosis, and be useful for screening patient populations for AD development and progression. PMID:25256347

Pomegranate-Extract Pill in Preventing Tumor Growth in Patients With Localized Prostate Cancer Undergoing Active Surveillance

ClinicalTrials.gov

2018-03-02

PSA Level Less Than or Equal to Fifteen; PSA Level Less Than Ten; Stage I Prostate Cancer AJCC v7; Stage II Prostate Cancer AJCC v7; Stage IIA Prostate Cancer AJCC v7; Stage IIB Prostate Cancer AJCC v7

The Paired-box protein PAX-3 regulates the choice between lateral and ventral epidermal cell fates in C. elegans

PubMed Central

Thompson, Kenneth W.; Joshi, Pradeep; Dymond, Jessica S.; Gorrepati, Lakshmi; Smith, Harold; Krause, Michael; Eisenmann, David M.

2016-01-01

The development of the single cell layer skin or hypodermis of Caenorhabditis elegans is an excellent model for understanding cell fate specification and differentiation. Early in C. elegans embryogenesis, six rows of hypodermal cells adopt dorsal, lateral or ventral fates that go on to display distinct behaviors during larval life. Several transcription factors are known that function in specifying these major hypodermal cell fates, but our knowledge of the specification of these cell types is sparse, particularly in the case of the ventral hypodermal cells, which become Vulval Precursor Cells and form the vulval opening in response to extracellular signals. Previously, the gene pvl-4 was identified in a screen for mutants with defects in vulval development. We found by whole genome sequencing that pvl-4 is the Paired-box gene pax-3, which encodes the sole PAX-3 transcription factor homolog in C. elegans. pax-3 mutants show embryonic and larval lethality, and body morphology abnormalities indicative of hypodermal cell defects. We report that pax-3 is expressed in ventral P cells and their descendants during embryogenesis and early larval stages, and that in pax-3 reduction-of-function animals the ventral P cells undergo a cell fate transformation and express several markers of the lateral seam cell fate. Furthermore, forced expression of pax-3 in the lateral hypodermal cells causes them to lose expression of seam cell markers. We propose that pax-3 functions in the ventral hypodermal cells to prevent these cells from adopting the lateral seam cell fate. pax-3 represents the first gene required for specification solely of the ventral hypodermal fate in C. elegans providing insights into cell type diversification. PMID:26953187

The Paired-box protein PAX-3 regulates the choice between lateral and ventral epidermal cell fates in C. elegans.

PubMed

Thompson, Kenneth W; Joshi, Pradeep; Dymond, Jessica S; Gorrepati, Lakshmi; Smith, Harold E; Krause, Michael W; Eisenmann, David M

2016-04-15

The development of the single cell layer skin or hypodermis of Caenorhabditis elegans is an excellent model for understanding cell fate specification and differentiation. Early in C. elegans embryogenesis, six rows of hypodermal cells adopt dorsal, lateral or ventral fates that go on to display distinct behaviors during larval life. Several transcription factors are known that function in specifying these major hypodermal cell fates, but our knowledge of the specification of these cell types is sparse, particularly in the case of the ventral hypodermal cells, which become Vulval Precursor Cells and form the vulval opening in response to extracellular signals. Previously, the gene pvl-4 was identified in a screen for mutants with defects in vulval development. We found by whole genome sequencing that pvl-4 is the Paired-box gene pax-3, which encodes the sole PAX-3 transcription factor homolog in C. elegans. pax-3 mutants show embryonic and larval lethality, and body morphology abnormalities indicative of hypodermal cell defects. We report that pax-3 is expressed in ventral P cells and their descendants during embryogenesis and early larval stages, and that in pax-3 reduction-of-function animals the ventral P cells undergo a cell fate transformation and express several markers of the lateral seam cell fate. Furthermore, forced expression of pax-3 in the lateral hypodermal cells causes them to lose expression of seam cell markers. We propose that pax-3 functions in the ventral hypodermal cells to prevent these cells from adopting the lateral seam cell fate. pax-3 represents the first gene required for specification solely of the ventral hypodermal fate in C. elegans providing insights into cell type diversification. Copyright © 2016 Elsevier Inc. All rights reserved.

Fucoxanthin exerts differing effects on 3T3-L1 cells according to differentiation stage and inhibits glucose uptake in mature adipocytes

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

Kang, Seong-Il; Ko, Hee-Chul; Shin, Hye-Sun

2011-06-17

Highlights: {yields} Fucoxanthin enhances 3T3-L1 adipocyte differentiation at an early stage. {yields} Fucoxanthin inhibits 3T3-L1 adipocyte differentiation at intermediate and late stages. {yields} Fucoxanthin attenuates glucose uptake by inhibiting the phosphorylation of IRS in mature 3T3-L1 adipocytes. {yields} Fucoxanthin exerts its anti-obesity effect by inhibiting the differentiation of adipocytes at both intermediate and late stages, as well as glucose uptake in mature adipocytes. -- Abstract: Progression of 3T3-L1 preadipocyte differentiation is divided into early (days 0-2, D0-D2), intermediate (days 2-4, D2-D4), and late stages (day 4 onwards, D4-). In this study, we investigated the effects of fucoxanthin, isolated from themore » edible brown seaweed Petalonia binghamiae, on adipogenesis during the three differentiation stages of 3T3-L1 preadipocytes. When fucoxanthin was applied during the early stage of differentiation (D0-D2), it promoted 3T3-L1 adipocyte differentiation, as evidenced by increased triglyceride accumulation. At the molecular level, fucoxanthin increased protein expression of peroxisome proliferator-activated receptor {gamma} (PPAR{gamma}), CCAAT/enhancer-binding protein {alpha} (C/EBP{alpha}), sterol regulatory element-binding protein 1c (SREBP1c), and aP2, and adiponectin mRNA expression, in a dose-dependent manner. However, it reduced the expression of PPAR{gamma}, C/EBP{alpha}, and SREBP1c during the intermediate (D2-D4) and late stages (D4-D7) of differentiation. It also inhibited the uptake of glucose in mature 3T3-L1 adipocytes by reducing the phosphorylation of insulin receptor substrate 1 (IRS-1). These results suggest that fucoxanthin exerts differing effects on 3T3-L1 cells of different differentiation stages and inhibits glucose uptake in mature adipocytes.« less

Muscle development and differentiation in the urodele Ambystoma mexicanum.

PubMed

Banfi, Serena; Monti, Laura; Acquati, Francesco; Tettamanti, Gianluca; de Eguileor, Magda; Grimaldi, Annalisa

2012-05-01

Muscle differentiation has been widely described in zebrafish and Xenopus, but nothing is known about this process in amphibian urodeles. Both anatomical features and locomotor activity in urodeles are known to show intermediate features between fish and anurans. Therefore, a better understanding of myogenesis in urodeles could be useful to clarify the evolutionary changes that led to the formation of skeletal muscle in the trunk of land vertebrates. We report here a detailed morphological and molecular investigation on several embryonic stages of Ambystoma mexicanum and show that the first differentiating muscle fibers are the slow ones, originating from a myoblast population initially localized close to the notochord that forms a superficial layer on the somitic surface afterwards. Subsequently, fast fibers differentiation ensues. We also identified and cloned A. mexicanum Myf5 as a muscle-specific transcriptional factor likely involved in urodele muscle differentiation. © 2012 The Authors Development, Growth & Differentiation © 2012 Japanese Society of Developmental Biologists.

ACTOplus Met XR in Treating Patients With Stage I-IV Oral Cavity or Oropharynx Cancer Undergoing Definitive Treatment | Division of Cancer Prevention

Cancer.gov

This randomized phase IIb trial studies how well ACTOplus met XR works in treating in patients with stage I-IV oral cavity or oropharynx cancer that are undergoing definitive treatment. Chemoprevention is the use of drugs to keep oral cavity or oropharynx cancer from forming or coming back. The use of ACTOplus met XR may slow disease progression in patients with oral cavity or

Incorporating robotic-assisted surgery for endometrial cancer staging: Analysis of morbidity and costs.

PubMed

Bogani, Giorgio; Multinu, Francesco; Dowdy, Sean C; Cliby, William A; Wilson, Timothy O; Gostout, Bobbie S; Weaver, Amy L; Borah, Bijan J; Killian, Jill M; Bijlani, Akash; Angioni, Stefano; Mariani, Andrea

2016-05-01

To evaluate how the introduction of robotic-assisted surgery affects treatment-related morbidity and cost of endometrial cancer (EC) staging. We retrospectively reviewed the records of consecutive patients with stage I-III EC undergoing surgical staging between 2007 and 2012 at our institution. Costs (from surgery to 30days after surgery) were set based on the Medicare cost-to-charge ratio for each year and inflated to 2014 values. Inverse probability weighting (IPW) was used to decrease the allocation bias when comparing outcomes between surgical groups. We focused our analysis on the 251 EC patients who had robotic-assisted surgery and the 384 who had open staging. During the study period, the use of robotic-assisted surgery increased and open staging decreased (P<0.001). Correcting group imbalances by using IPW methodology, we observed that patients undergoing robotic-assisted staging had a significantly lower postoperative complication rate, lower blood transfusion rate, longer median operating time, shorter median length of stay, and lower readmission rate than patients undergoing open staging (all P<0.001). Overall 30-day costs were similar between the 2 groups, with robotic-assisted surgery having significantly higher median operating room costs ($2820 difference; P<0.001) but lower median room and board costs ($2929 difference; P<0.001) than open surgery. Increasing experience with robotic-assisted staging was significantly associated with a decrease in median operating time (P=0.002) and length of stay (P=0.003). The implementation of robotic-assisted surgery for EC staging improves patient outcomes. It provides women the benefits of minimally invasive surgery without increasing costs and potentially improves patient turnover. Copyright © 2016 Elsevier Inc. All rights reserved.

Complications After Total Skin-Sparing Mastectomy and Expander-Implant Reconstruction: Effects of Radiation Therapy on the Stages of Reconstruction.

PubMed

Peled, Anne Warren; Sears, Marie; Wang, Frederick; Foster, Robert D; Alvarado, Michael; Wong, Jasmine; Ewing, Cheryl A; Sbitany, Hani; Esserman, Laura J; Fowble, Barbara

2018-01-01

Many patients undergoing total-skin sparing mastectomy (TSSM) and 2-staged expander-implant (TE-I) reconstruction require postmastectomy radiation therapy (PMRT). Additionally, many patients undergoing TSSM for recurrent cancer have a history of lumpectomy and radiation therapy (XRT). Few studies have looked at the impact of XRT on the stages of TE-I reconstruction. Patients undergoing TSSM and immediate TE-I reconstruction between 2006 and 2013 were identified from a prospectively maintained database. Rates of TE-I loss and severe infection requiring intravenous antibiotics were compared in patients with prior XRT (85 cases) and PMRT (133 cases). Complications were divided by stage of reconstruction: first stage (TSSM and TE placement) and second stage (TE-I exchange). Mean follow-up time was 2.5 years. Patients with prior XRT had more complications after the first stage of reconstruction than the second (TE-I loss: 15% vs 5%, P = 0.03; infection: 20% vs 8%, P = 0.04). Patients receiving PMRT had low complication rates after the first stage, when they had not yet received radiation (TE-I loss: 2%; infection: 5%). However, complication rates after TE-I exchange (TE-I loss, 18%; infection, 31%) were significantly higher, and nearly 4-fold higher than patients with prior XRT. Patients with prior XRT are at high risk for complications after the first stage of TE-I reconstruction after TSSM; however, the risk of complications at the second stage is comparable to patients without radiation exposure and significantly lower than patients receiving PMRT. Patients receiving radiation therapy should be given appropriate preoperative counseling regarding their risks.

The ubiquitin ligase LIN41/TRIM71 targets p53 to antagonize cell death and differentiation pathways during stem cell differentiation

PubMed Central

Nguyen, Duong Thi Thuy; Richter, Daniel; Michel, Geert; Mitschka, Sibylle; Kolanus, Waldemar; Cuevas, Elisa; Gregory Wulczyn, F

2017-01-01

Rapidity and specificity are characteristic features of proteolysis mediated by the ubiquitin-proteasome system. Therefore, the UPS is ideally suited for the remodeling of the embryonic stem cell proteome during the transition from pluripotent to differentiated states and its inverse, the generation of inducible pluripotent stem cells. The Trim-NHL family member LIN41 is among the first E3 ubiquitin ligases to be linked to stem cell pluripotency and reprogramming. Initially discovered in C. elegans as a downstream target of the let-7 miRNA, LIN41 is now recognized as a critical regulator of stem cell fates as well as the timing of neurogenesis. Despite being indispensable for embryonic development and neural tube closure in mice, the underlying mechanisms for LIN41 function in these processes are poorly understood. To better understand the specific contributions of the E3 ligase activity for the stem cell functions of LIN41, we characterized global changes in ubiquitin or ubiquitin-like modifications using Lin41-inducible mouse embryonic stem cells. The tumor suppressor protein p53 was among the five most strongly affected proteins in cells undergoing neural differentiation in response to LIN41 induction. We show that LIN41 interacts with p53, controls its abundance by ubiquitination and antagonizes p53-dependent pro-apoptotic and pro-differentiation responses. In vivo, the lack of LIN41 is associated with upregulation of Grhl3 and widespread caspase-3 activation, two downstream effectors of p53 with essential roles in neural tube closure. As Lin41-deficient mice display neural tube closure defects, we conclude that LIN41 is critical for the regulation of p53 functions in cell fate specification and survival during early brain development. PMID:28430184

Student Employment: Social Differentials and Field-Specific Developments in Higher Education

ERIC Educational Resources Information Center

Jacob, Marita; Gerth, Maria; Weiss, Felix

2018-01-01

In this article, we examine social origin differences in employment patterns across different stages of higher education and compare these differences between vocational and academic fields of study. Using data from a large-scale German student survey, we study the development of inequality, according to social origins, in student employment from…

Mutations in gasdermin 3 cause aberrant differentiation of the hair follicle and sebaceous gland.

PubMed

Lunny, Declan P; Weed, Erica; Nolan, Patrick M; Marquardt, Andreas; Augustin, Martin; Porter, Rebecca M

2005-03-01

Defolliculated (Dfl) is a spontaneous mouse mutant with a hair-loss phenotype that includes altered sebaceous gland differentiation, short hair shafts, aberrant catagen stage of the hair cycle, and eventual loss of the hair follicle. Recently a similar mutant, finnegan (Fgn), with an identical phenotype was discovered during a phenotypic screen for mutations induced by chemical mutagenesis. The gene underlying the phenotype of both finnegan and defolliculated has been mapped to chromosome 11 and here we show that both mice harbor mutations in gasdermin 3 (Gsdm3), a gene of unknown function. Gsdm3(Dfl) is a B2 insertion near the 3' splice site of exon 7 and Gsdm3(Fgn) is a point mutation T278P. To investigate the role of the gasdermin gene family an antiserum was raised to a peptide highly homologous to all three mouse gasdermins and human gasdermin. Immunohistochemical analysis revealed that gasdermins are expressed specifically in cells at advanced stages of differentiation in the upper epidermis, the differentiating inner root sheath and hair shaft and in the most mature sebocytes of the sebaceous gland and preputial, meibomium, ceruminous gland, and anal glands. This expression pattern suggests a role for gasdermins in differentiation of the epidermis and its appendages.

Silver nanoparticles induce developmental stage-specific embryonic phenotypes in zebrafish.

PubMed

Lee, Kerry J; Browning, Lauren M; Nallathamby, Prakash D; Osgood, Christopher J; Xu, Xiao-Hong Nancy

2013-12-07

Much is anticipated from the development and deployment of nanomaterials in biological organisms, but concerns remain regarding their biocompatibility and target specificity. Here we report our study of the transport, biocompatibility and toxicity of purified and stable silver nanoparticles (Ag NPs, 13.1 ± 2.5 nm in diameter) upon the specific developmental stages of zebrafish embryos using single NP plasmonic spectroscopy. We find that single Ag NPs passively diffuse into five different developmental stages of embryos (cleavage, early-gastrula, early-segmentation, late-segmentation, and hatching stages), showing stage-independent diffusion modes and diffusion coefficients. Notably, the Ag NPs induce distinctive stage and dose-dependent phenotypes and nanotoxicity, upon their acute exposure to the Ag NPs (0-0.7 nM) for only 2 h. The late-segmentation embryos are most sensitive to the NPs with the lowest critical concentration (CNP,c < 0.02 nM) and highest percentages of cardiac abnormalities, followed by early-segmentation embryos (CNP,c < 0.02 nM), suggesting that disruption of cell differentiation by the NPs causes the most toxic effects on embryonic development. The cleavage-stage embryos treated with the NPs develop into a wide variety of phenotypes (abnormal finfold, tail/spinal cord flexure, cardiac malformation/edema, yolk sac edema, and acephaly). These organ structures are not yet developed in cleavage-stage embryos, suggesting that the earliest determinative events to create these structures are ongoing, and disrupted by NPs, which leads to the downstream effects. In contrast, the hatching embryos are most resistant to the Ag NPs, and majority of embryos (94%) develop normally, and none of them develop abnormally. Interestingly, early-gastrula embryos are less sensitive to the NPs than cleavage and segmentation stage embryos, and do not develop abnormally. These important findings suggest that the Ag NPs are not simple poisons, and they can target specific pathways in development, and potentially enable target specific study and therapy for early embryonic development.

The role of early development in mammalian limb diversification: a descriptive comparison of early limb development between the Natal long-fingered bat (Miniopterus natalensis) and the mouse (Mus musculus).

PubMed

Hockman, Dorit; Mason, Mandy K; Jacobs, David S; Illing, Nicola

2009-04-01

Comparative embryology expands our understanding of unique limb structures, such as that found in bats. Bat forelimb digits 2 to 5 are differentially elongated and joined by webbing, while the hindlimb digits are of similar length in many species. We compare limb development between the mouse and the Natal long-fingered bat, Miniopterus natalensis, to pinpoint the stage at which their limbs begin to differ. The bat forelimb differs from the mouse at Carollia stage (CS) 14 with the appearance of the wing membrane primordia. This difference is enhanced at CS 15 with the posterior expansion of the hand plate. The bat hindlimb begins to differ from the mouse between CS 15 and 16 when the foot plate undergoes a proximal expansion resulting in digit primordia of very similar length. Our findings support recent gene expression studies, which reveal a role for early patterning in the development of the bat limb. Copyright 2009 Wiley-Liss, Inc.

Nintedanib Compared With Placebo in Treating Against Radiation-Induced Pneumonitis in Patients With Non-small Cell Lung Cancer That Cannot Be Removed by Surgery and Are Undergoing Chemoradiation Therapy

ClinicalTrials.gov

2017-07-08

Radiation-Induced Pneumonitis; Stage IIA Non-Small Cell Lung Carcinoma; Stage IIB Non-Small Cell Lung Carcinoma; Stage IIIA Non-Small Cell Lung Cancer; Stage IIIB Non-Small Cell Lung Cancer; Stage IV Non-Small Cell Lung Cancer

Comprehensive Patient Questionnaires in Predicting Complications in Older Patients With Gynecologic Cancer Undergoing Surgery

ClinicalTrials.gov

2018-02-14

Endometrial Serous Adenocarcinoma; Fallopian Tube Carcinoma; Ovarian Carcinoma; Primary Peritoneal Carcinoma; Stage IIIA Uterine Corpus Cancer AJCC v7; Stage IIIB Uterine Corpus Cancer AJCC v7; Stage IIIC Uterine Corpus Cancer AJCC v7; Stage IVA Uterine Corpus Cancer AJCC v7; Stage IVB Uterine Corpus Cancer AJCC v7

A Genome-Wide Screen Indicates Correlation between Differentiation and Expression of Metabolism Related Genes

PubMed Central

Shende, Akhilesh; Singh, Anupama; Meena, Anil; Ghosal, Ritika; Ranganathan, Madhav; Bandyopadhyay, Amitabha

2013-01-01

Differentiated tissues may be considered as materials with distinct properties. The differentiation program of a given tissue ensures that it acquires material properties commensurate with its function. It may be hypothesized that some of these properties are acquired through production of tissue-specific metabolites synthesized by metabolic enzymes. To establish correlation between metabolism and organogenesis we have carried out a genome-wide expression study of metabolism related genes by RNA in-situ hybridization. 23% of the metabolism related genes studied are expressed in a tissue-restricted but not tissue-exclusive manner. We have conducted the screen on whole mount chicken (Gallus gallus) embryos from four distinct developmental stages to correlate dynamic changes in expression patterns of metabolic enzymes with spatio-temporally unique developmental events. Our data strongly suggests that unique combinations of metabolism related genes, and not specific metabolic pathways, are upregulated during differentiation. Further, expression of metabolism related genes in well established signaling centers that regulate different aspects of morphogenesis indicates developmental roles of some of the metabolism related genes. The database of tissue-restricted expression patterns of metabolism related genes, generated in this study, should serve as a resource for systematic identification of these genes with tissue-specific functions during development. Finally, comprehensive understanding of differentiation is not possible unless the downstream genes of a differentiation cascade are identified. We propose, metabolic enzymes constitute a significant portion of these downstream target genes. Thus our study should help elucidate different aspects of tissue differentiation. PMID:23717462

A genome-wide screen indicates correlation between differentiation and expression of metabolism related genes.

PubMed

Roy, Priti; Kumar, Brijesh; Shende, Akhilesh; Singh, Anupama; Meena, Anil; Ghosal, Ritika; Ranganathan, Madhav; Bandyopadhyay, Amitabha

2013-01-01

Differentiated tissues may be considered as materials with distinct properties. The differentiation program of a given tissue ensures that it acquires material properties commensurate with its function. It may be hypothesized that some of these properties are acquired through production of tissue-specific metabolites synthesized by metabolic enzymes. To establish correlation between metabolism and organogenesis we have carried out a genome-wide expression study of metabolism related genes by RNA in-situ hybridization. 23% of the metabolism related genes studied are expressed in a tissue-restricted but not tissue-exclusive manner. We have conducted the screen on whole mount chicken (Gallus gallus) embryos from four distinct developmental stages to correlate dynamic changes in expression patterns of metabolic enzymes with spatio-temporally unique developmental events. Our data strongly suggests that unique combinations of metabolism related genes, and not specific metabolic pathways, are upregulated during differentiation. Further, expression of metabolism related genes in well established signaling centers that regulate different aspects of morphogenesis indicates developmental roles of some of the metabolism related genes. The database of tissue-restricted expression patterns of metabolism related genes, generated in this study, should serve as a resource for systematic identification of these genes with tissue-specific functions during development. Finally, comprehensive understanding of differentiation is not possible unless the downstream genes of a differentiation cascade are identified. We propose, metabolic enzymes constitute a significant portion of these downstream target genes. Thus our study should help elucidate different aspects of tissue differentiation.

STAT signaling in mammary gland differentiation, cell survival and tumorigenesis.

PubMed

Haricharan, S; Li, Y

2014-01-25

The mammary gland is a unique organ that undergoes extensive and profound changes during puberty, menstruation, pregnancy, lactation and involution. The changes that take place during puberty involve large-scale proliferation and invasion of the fat-pad. During pregnancy and lactation, the mammary cells are exposed to signaling pathways that inhibit apoptosis, induce proliferation and invoke terminal differentiation. Finally, during involution the mammary gland is exposed to milk stasis, programmed cell death and stromal reorganization to clear the differentiated milk-producing cells. Not surprisingly, the signaling pathways responsible for bringing about these changes in breast cells are often subverted during the process of tumorigenesis. The STAT family of proteins is involved in every stage of mammary gland development, and is also frequently implicated in breast tumorigenesis. While the roles of STAT3 and STAT5 during mammary gland development and tumorigenesis are well studied, others members, e.g. STAT1 and STAT6, have only recently been observed to play a role in mammary gland biology. Continued investigation into the STAT protein network in the mammary gland will likely yield new biomarkers and risk factors for breast cancer, and may also lead to novel prophylactic or therapeutic strategies against breast cancer. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Acute Kidney Injury in Patients Undergoing the Extracardiac Fontan Operation With and Without the Use of Cardiopulmonary Bypass.

PubMed

Algaze, Claudia A; Koth, Andrew M; Faberowski, Lisa W; Hanley, Frank L; Krawczeski, Catherine D; Axelrod, David M

2017-01-01

To describe the prevalence and risk factors for acute kidney injury in patients undergoing the extracardiac Fontan operation with and without cardiopulmonary bypass, and to determine whether acute kidney injury is associated with duration of mechanical ventilation, cardiovascular ICU and hospital postoperative length of stay, and early mortality. Single-center retrospective cohort study. Pediatric cardiovascular ICU, university-affiliated children's hospital. Patients with a preoperative creatinine before undergoing first-time extracardiac Fontan between January 1, 2004, and April 30, 2012. None. Acute kidney injury occurred in 55 of 138 patients (39.9%), including 41 (29.7%) with stage 1, six (4.4%) with stage 2, and eight (5.8%) with stage 3 acute kidney injury. Cardiopulmonary bypass was strongly associated with a higher risk of any acute kidney injury (adjusted odds ratio, 4.8 [95% CI, 1.4-16.0]; p = 0.01) but not stage 2/3 acute kidney injury. Lower renal perfusion pressure on the day of surgery (postoperative day, 0) was associated with a higher risk of stage 2/3 acute kidney injury (adjusted odds ratio, 1.2 [95% CI, 1.0-1.5]; p = 0.03). Higher vasoactive-inotropic score on postoperative day 0 was associated with a higher risk for stage 2/3 acute kidney injury (adjusted odds ratio, 1.9 [95% CI, 1.0-3.4]; p = 0.04). Stage 2/3 acute kidney injury was associated with longer cardiovascular ICU length of stay (mean, 7.3 greater d [95% CI, 3.4-11.3]; p < 0.001) and hospital postoperative length of stay (mean, 6.4 greater d [95% CI, 0.06-12.5]; p = 0.04). Postoperative acute kidney injury in patients undergoing the extracardiac Fontan operation is common and is associated with lower postoperative renal perfusion pressure and higher vasoactive-inotropic score. Cardiopulmonary bypass was strongly associated with any acute kidney injury, although not stage 2/3 acute kidney injury. Stage 2/3 acute kidney injury is a compelling risk factor for longer cardiovascular ICU and hospital postoperative length of stay. Increased attention to and management of renal perfusion pressure may reduce postoperative acute kidney injury and improve outcomes.

Reconstructing the regulatory circuit of cell fate determination in yeast mating response.

PubMed

Shao, Bin; Yuan, Haiyu; Zhang, Rongfei; Wang, Xuan; Zhang, Shuwen; Ouyang, Qi; Hao, Nan; Luo, Chunxiong

2017-07-01

Massive technological advances enabled high-throughput measurements of proteomic changes in biological processes. However, retrieving biological insights from large-scale protein dynamics data remains a challenging task. Here we used the mating differentiation in yeast Saccharomyces cerevisiae as a model and developed integrated experimental and computational approaches to analyze the proteomic dynamics during the process of cell fate determination. When exposed to a high dose of mating pheromone, the yeast cell undergoes growth arrest and forms a shmoo-like morphology; however, at intermediate doses, chemotropic elongated growth is initialized. To understand the gene regulatory networks that control this differentiation switch, we employed a high-throughput microfluidic imaging system that allows real-time and simultaneous measurements of cell growth and protein expression. Using kinetic modeling of protein dynamics, we classified the stimulus-dependent changes in protein abundance into two sources: global changes due to physiological alterations and gene-specific changes. A quantitative framework was proposed to decouple gene-specific regulatory modes from the growth-dependent global modulation of protein abundance. Based on the temporal patterns of gene-specific regulation, we established the network architectures underlying distinct cell fates using a reverse engineering method and uncovered the dose-dependent rewiring of gene regulatory network during mating differentiation. Furthermore, our results suggested a potential crosstalk between the pheromone response pathway and the target of rapamycin (TOR)-regulated ribosomal biogenesis pathway, which might underlie a cell differentiation switch in yeast mating response. In summary, our modeling approach addresses the distinct impacts of the global and gene-specific regulation on the control of protein dynamics and provides new insights into the mechanisms of cell fate determination. We anticipate that our integrated experimental and modeling strategies could be widely applicable to other biological systems.

PPARbeta agonists trigger neuronal differentiation in the human neuroblastoma cell line SH-SY5Y.

PubMed

Di Loreto, S; D'Angelo, B; D'Amico, M A; Benedetti, E; Cristiano, L; Cinque, B; Cifone, M G; Cerù, M P; Festuccia, C; Cimini, A

2007-06-01

Neuroblastomas are pediatric tumors originating from immature neuroblasts in the developing peripheral nervous system. Differentiation therapies could help lowering the high mortality due to rapid tumor progression to advanced stages. Oleic acid has been demonstrated to promote neuronal differentiation in neuronal cultures. Herein we report on the effects of oleic acid and of a specific synthetic PPARbeta agonist on cell growth, expression of differentiation markers and on parameters responsible for the malignancy such as adhesion, migration, invasiveness, BDNF, and TrkB expression of SH-SY5Y neuroblastoma cells. The results obtained demonstrate that many, but not all, oleic acid effects are mediated by PPARbeta and support a role for PPARbeta in neuronal differentiation strongly pointing towards PPAR ligands as new therapeutic strategies against progression and recurrences of neuroblastoma.

Transcriptome profiles of embryos before and after cleavage in Eriocheir sinensis: identification of developmental genes at the earliest stages

NASA Astrophysics Data System (ADS)

Hui, Min; Cui, Zhaoxia; Liu, Yuan; Song, Chengwen

2017-07-01

In crab, embryogenesis is a complicated developmental program marked by a series of critical events. RNA-Sequencing technology offers developmental biologists a way to identify many more developmental genes than ever before. Here, we present a comprehensive analysis of the transcriptomes of Eriocheir sinensis oosperms (Os) and embryos at the 2-4 cell stage (Cs), which are separated by a cleavage event. A total of 18 923 unigenes were identified, and 403 genes matched with gene ontology (GO) terms related to developmental processes. In total, 432 differentially expressed genes (DEGs) were detected between the two stages. Nine DEGs were specifically expressed at only one stage. These DEGs may be relevant to stage-specific molecular events during development. A number of DEGs related to `hedgehog signaling pathway', `Wnt signaling pathway' `germplasm', `nervous system', `sensory perception' and `segment polarity' were identified as being up-regulated at the Cs stage. The results suggest that these embryonic developmental events begin before the early cleavage event in crabs, and that many of the genes expressed in the two transcriptomes might be maternal genes. Our study provides ample information for further research on the molecular mechanisms underlying crab development.

Impact of Increasing Age on Cause-Specific Mortality and Morbidity in Patients With Stage I Non-Small-Cell Lung Cancer: A Competing Risks Analysis.

PubMed

Eguchi, Takashi; Bains, Sarina; Lee, Ming-Ching; Tan, Kay See; Hristov, Boris; Buitrago, Daniel H; Bains, Manjit S; Downey, Robert J; Huang, James; Isbell, James M; Park, Bernard J; Rusch, Valerie W; Jones, David R; Adusumilli, Prasad S

2017-01-20

Purpose To perform competing risks analysis and determine short- and long-term cancer- and noncancer-specific mortality and morbidity in patients who had undergone resection for stage I non-small-cell lung cancer (NSCLC). Patients and Methods Of 5,371 consecutive patients who had undergone curative-intent resection of primary lung cancer at our institution (2000 to 2011), 2,186 with pathologic stage I NSCLC were included in the analysis. All preoperative clinical variables known to affect outcomes were included in the analysis, specifically, Charlson comorbidity index, predicted postoperative (ppo) diffusing capacity of the lung for carbon monoxide, and ppo forced expiratory volume in 1 second. Cause-specific mortality analysis was performed with competing risks analysis. Results Of 2,186 patients, 1,532 (70.1%) were ≥ 65 years of age, including 638 (29.2%) ≥ 75 years of age. In patients < 65, 65 to 74, and ≥ 75 years of age, 5-year lung cancer-specific cumulative incidence of death (CID) was 7.5%, 10.7%, and 13.2%, respectively (overall, 10.4%); noncancer-specific CID was 1.8%, 4.9%, and 9.0%, respectively (overall, 5.3%). In patients ≥ 65 years of age, for up to 2.5 years after resection, noncancer-specific CID was higher than lung cancer-specific CID; the higher noncancer-specific, early-phase mortality was enhanced in patients ≥ 75 years of age than in those 65 to 74 years of age. Multivariable analysis showed that low ppo diffusing capacity of lung for carbon monoxide was an independent predictor of severe morbidity ( P < .001), 1-year mortality ( P < .001), and noncancer-specific mortality ( P < .001), whereas low ppo forced expiratory volume in 1 second was an independent predictor of lung cancer-specific mortality ( P = .002). Conclusion In patients who undergo curative-intent resection of stage I NSCLC, noncancer-specific mortality is a significant competing event, with an increasing impact as patient age increases.

Oncofertility in patients with stage I epithelial ovarian cancer: fertility-sparing surgery in young women of reproductive age.

PubMed

Jiang, Xuan; Yang, Jiaxin; Yu, Mei; Xie, Weimin; Cao, Dongyan; Wu, Ming; Pan, Lingya; Huang, Huifang; You, Yan; Shen, Keng

2017-08-15

Fertility-sparing surgery is indicated for patients with stage I epithelial ovarian cancers. We sought to evaluate the clinical outcomes and oncofertility in a cohort of patients of reproductive age with stage I epithelial ovarian cancer (EOC). Overall, 108 patients of reproductive age (≤ 40 years) diagnosed with stage I EOC who were treated at Peking Union Medical College Hospital between 1999 and 2013 were included in the study. The Kaplan-Meier model and Cox regression analyses were used for the survival analysis. The type of surgery included fertility-sparing surgery (FSS) (48.1%) and radical surgery (RS) (51.9%). After a median follow-up of 83 months, we observed that grade 3 or clear-cell carcinoma was the only independent risk factor for disease-free survival and tumor-specific survival in the multivariate analysis. Patients with grade 3 or clear-cell carcinoma tended to be older than 30 years, have endometriosis, and undergo RS (p < 0.05). Fertility-sparing surgery did not affect disease-free survival or tumor-specific survival among patients of reproductive age with stage I EOC and among high-risk patients with stage IC2-3, grade 3, or clear-cell carcinoma. Thirty-four out of 52 (65.4%) FSS patients attempted to get pregnant. Twenty-eight (82.4%) achieved a successful pregnancy with a full-term delivery. Grade 3 or clear-cell carcinoma was the only independent risk factor for survival of patients of reproductive age with stage I EOC. FSS can be safely performed on patients of reproductive age with grade 1-2, stage I EOC. The safety of FSS for grade 3 and clear-cell carcinoma warrants further investigation.

Use of transcriptome sequencing to understand the pistillate flowering in hickory (Carya cathayensis Sarg.).

PubMed

Huang, You-Jun; Liu, Li-Li; Huang, Jian-Qin; Wang, Zheng-Jia; Chen, Fang-Fang; Zhang, Qi-Xiang; Zheng, Bing-Song; Chen, Ming

2013-10-10

Different from herbaceous plants, the woody plants undergo a long-period vegetative stage to achieve floral transition. They then turn into seasonal plants, flowering annually. In this study, a preliminary model of gene regulations for seasonal pistillate flowering in hickory (Carya cathayensis) was proposed. The genome-wide dynamic transcriptome was characterized via the joint-approach of RNA sequencing and microarray analysis. Differential transcript abundance analysis uncovered the dynamic transcript abundance patterns of flowering correlated genes and their major functions based on Gene Ontology (GO) analysis. To explore pistillate flowering mechanism in hickory, a comprehensive flowering gene regulatory network based on Arabidopsis thaliana was constructed by additional literature mining. A total of 114 putative flowering or floral genes including 31 with differential transcript abundance were identified in hickory. The locations, functions and dynamic transcript abundances were analyzed in the gene regulatory networks. A genome-wide co-expression network for the putative flowering or floral genes shows three flowering regulatory modules corresponding to response to light abiotic stimulus, cold stress, and reproductive development process, respectively. Totally 27 potential flowering or floral genes were recruited which are meaningful to understand the hickory specific seasonal flowering mechanism better. Flowering event of pistillate flower bud in hickory is triggered by several pathways synchronously including the photoperiod, autonomous, vernalization, gibberellin, and sucrose pathway. Totally 27 potential flowering or floral genes were recruited from the genome-wide co-expression network function module analysis. Moreover, the analysis provides a potential FLC-like gene based vernalization pathway and an 'AC' model for pistillate flower development in hickory. This work provides an available framework for pistillate flower development in hickory, which is significant for insight into regulation of flowering and floral development of woody plants.

Use of transcriptome sequencing to understand the pistillate flowering in hickory (Carya cathayensis Sarg.)

PubMed Central

2013-01-01

Background Different from herbaceous plants, the woody plants undergo a long-period vegetative stage to achieve floral transition. They then turn into seasonal plants, flowering annually. In this study, a preliminary model of gene regulations for seasonal pistillate flowering in hickory (Carya cathayensis) was proposed. The genome-wide dynamic transcriptome was characterized via the joint-approach of RNA sequencing and microarray analysis. Results Differential transcript abundance analysis uncovered the dynamic transcript abundance patterns of flowering correlated genes and their major functions based on Gene Ontology (GO) analysis. To explore pistillate flowering mechanism in hickory, a comprehensive flowering gene regulatory network based on Arabidopsis thaliana was constructed by additional literature mining. A total of 114 putative flowering or floral genes including 31 with differential transcript abundance were identified in hickory. The locations, functions and dynamic transcript abundances were analyzed in the gene regulatory networks. A genome-wide co-expression network for the putative flowering or floral genes shows three flowering regulatory modules corresponding to response to light abiotic stimulus, cold stress, and reproductive development process, respectively. Totally 27 potential flowering or floral genes were recruited which are meaningful to understand the hickory specific seasonal flowering mechanism better. Conclusions Flowering event of pistillate flower bud in hickory is triggered by several pathways synchronously including the photoperiod, autonomous, vernalization, gibberellin, and sucrose pathway. Totally 27 potential flowering or floral genes were recruited from the genome-wide co-expression network function module analysis. Moreover, the analysis provides a potential FLC-like gene based vernalization pathway and an 'AC’ model for pistillate flower development in hickory. This work provides an available framework for pistillate flower development in hickory, which is significant for insight into regulation of flowering and floral development of woody plants. PMID:24106755

LRF-mediated Dll4 repression in erythroblasts is necessary for hematopoietic stem cell maintenance

PubMed Central

Lee, Sung-Uk; Maeda, Manami; Ishikawa, Yuichi; Li, Sierra Min; Wilson, Anne; Jubb, Adrian M.; Sakurai, Nagisa; Weng, Lihong; Fiorini, Emma; Radtke, Freddy; Yan, Minhong; MacDonald, H. Robson; Chen, Ching-Cheng

2013-01-01

Hematopoietic stem cells (HSCs) are the most primitive cells in the hematopoietic system and are under tight regulation for self-renewal and differentiation. Notch signals are essential for the emergence of definitive hematopoiesis in mouse embryos and are critical regulators of lymphoid lineage fate determination. However, it remains unclear how Notch regulates the balance between HSC self-renewal and differentiation in the adult bone marrow (BM). Here we report a novel mechanism that prevents HSCs from undergoing premature lymphoid differentiation in BM. Using a series of in vivo mouse models and functional HSC assays, we show that leukemia/lymphoma related factor (LRF) is necessary for HSC maintenance by functioning as an erythroid-specific repressor of Delta-like 4 (Dll4) expression. Lrf deletion in erythroblasts promoted up-regulation of Dll4 in erythroblasts, sensitizing HSCs to T-cell instructive signals in the BM. Our study reveals novel cross-talk between HSCs and erythroblasts, and sheds a new light on the regulatory mechanisms regulating the balance between HSC self-renewal and differentiation. PMID:23134786

STIM1 and TRPV4 regulate fluid flow-induced calcium oscillation at early and late stages of osteoclast differentiation.

PubMed

Li, Ping; Bian, Xueyan; Liu, Chenglin; Wang, Shurong; Guo, Mengmeng; Tao, Yingjie; Huo, Bo

2018-05-01

Bone resorption is mainly mediated by osteoclasts (OCs), whose formation and function are regulated by intracellular Ca 2+ oscillation. Our previous studies demonstrated that fluid shear stress (FSS) lead to Ca 2+ oscillation through mechanosensitive cation-selective channels. However, the specific channels responsible for this FSS-induced Ca 2+ oscillation remain unknown. In the present study, we examined the expression of several Ca 2+ channels in OCs, including STIM1, ORAI1, TRPV1, TRPV4, TRPV5, and TRPV6, by western blotting and reverse transcription-polymerase chain reaction. The results showed that STIM1 was highly expressed in early stage OCs, while TRPV4 was highly expressed in late stage OCs. We observed intracellular Ca 2+ responses in OCs that were mechanically stimulated by FSS. When we blocked STIM1-dependent store-operated Ca 2+ entry or inhibited TRPV4 using siRNA or drug inhibition, FSS-induced Ca 2+ oscillations were almost undetectable in early and late stage OCs, respectively. These results indicate that STIM1 and TRPV4 act as mechanical transduction channels for OCs during the early and late differentiation stages, respectively, suggesting that these calcium channel could serve as markers of osteoclastogenesis or bone resorption. Copyright © 2017 Elsevier Ltd. All rights reserved.

Lineage specific expression of Polycomb Group Proteins in human embryonic stem cells in vitro.

PubMed

Pethe, Prasad; Pursani, Varsha; Bhartiya, Deepa

2015-05-01

Human embryonic (hES) stem cells are an excellent model to study lineage specification and differentiation into various cell types. Differentiation necessitates repression of specific genes not required for a particular lineage. Polycomb Group (PcG) proteins are key histone modifiers, whose primary function is gene repression. PcG proteins form complexes called Polycomb Repressive Complexes (PRCs), which catalyze histone modifications such as H2AK119ub1, H3K27me3, and H3K9me3. PcG proteins play a crucial role during differentiation of stem cells. The expression of PcG transcripts during differentiation of hES cells into endoderm, mesoderm, and ectoderm lineage is yet to be shown. In-house derived hES cell line KIND1 was differentiated into endoderm, mesoderm, and ectoderm lineages; followed by characterization using RT-PCR for HNF4A, CDX2, MEF2C, TBX5, SOX1, and MAP2. qRT-PCR and western blotting was performed to compare expression of PcG transcripts and proteins across all the three lineages. We observed that cells differentiated into endoderm showed upregulation of RING1B, BMI1, EZH2, and EED transcripts. Mesoderm differentiation was characterized by significant downregulation of all PcG transcripts during later stages. BMI1 and RING1B were upregulated while EZH2, SUZ12, and EED remained low during ectoderm differentiation. Western blotting also showed distinct expression of BMI1 and EZH2 during differentiation into three germ layers. Our study shows that hES cells differentiating into endoderm, mesoderm, and ectoderm lineages show distinct PcG expression profile at transcript and protein level. © 2015 International Federation for Cell Biology.

Prevalence of chronic kidney disease among patients undergoing transradial percutaneous coronary interventions.

PubMed

Hossain, Mohammad A; Quinlan, Amy; Heck-Kanellidis, Jennifer; Calderon, Dawn; Patel, Tejas; Gandhi, Bhavika; Patel, Shrinil; Hetavi, Mahida; Costanzo, Eric J; Cosentino, James; Patel, Chirag; Dewan, Asa; Kuo, Yen-Hong; Salman, Loay; Vachharajani, Tushar J

2018-07-01

While transradial approach to conduct percutaneous coronary interventions offers multiple advantages, the procedure can cause radial artery damage and occlusion. Because radial artery is the preferred site for the creation of an arteriovenous fistula to provide dialysis, patients with chronic kidney disease are particularly dependent on radial artery for their long-term survival. In this retrospective study, we investigated the prevalence of chronic kidney disease in patients undergoing coronary interventions via radial artery. Stage of chronic kidney disease was based on estimated glomerular filtration rate and National Kidney Foundation - Kidney Disease Outcomes Quality Initiative guidelines. A total of 497 patients undergoing transradial percutaneous coronary interventions were included. Over 70.4% (350/497) of the patients had chronic kidney disease. Stage II chronic kidney disease was observed in 243 (69%) patients (estimated glomerular filtration rate = 76.0 ± 8.4 mL/min). Stage III was observed in 93 (27%) patients (estimated glomerular filtration rate = 49 ± 7.5 mL/min). Stage IV chronic kidney disease was observed in 5 (1%) patients (estimated glomerular filtration rate = 25.6 ± 4.3 mL/min) and Stage V chronic kidney disease was observed in 9 (3%) patients (estimated glomerular filtration rate = 9.3 ± 3.5 mL/min). Overall, 107 of 350 patients (30%) had advanced chronic kidney disease, that is, stage III-V chronic kidney disease. Importantly, 14 of the 107 (13%) patients had either stage IV or V chronic kidney disease. This study finds that nearly one-third of the patients undergoing transradial percutaneous coronary interventions have advanced chronic kidney disease. Because many of these patients may require dialysis, the use of radial artery to conduct percutaneous coronary interventions must be carefully considered in chronic kidney disease population.

Expression of CDX-2 and Ki-67 in different grades of colorectal adenocarcinomas.

PubMed

Sen, Anway; Mitra, Sumit; Das, Ram Narayan; Dasgupta, Shatavisha; Saha, Koushik; Chatterjee, Uttara; Mukherjee, Krishnendu; Datta, Chhanda; Chattopadhyay, Bitan K

2015-01-01

CDX2 is a caudal homeobox gene essential for intestinal differentiation and is specifically expressed in colorectal adenocarcinomas. Its role in colorectal carcinogenesis is not fully elucidated. To study the expression pattern of CDX2 and Ki-67 in different grades of colorectal adenocarcinomas and to observe the relationship of their staining patterns in various tumor stages and to look for correlation if any, between Ki-67 labeling index (Ki-67 LI) and CDX2 expression. A total of 74 cases were enrolled. Detailed clinical profile, peroperative findings, histological grading and staging were noted. Immunohistochemistry for CDX2 and Ki-67 was done, and Ki-67 LI was calculated. CDX2 staining was graded semi-quantitatively, and statistical analysis was done. Age of presentation ranged from 20 to 75 years, and the male:female ratio was 1.83:1. There were 8, 47 and 13 cases of well, moderate and poorly differentiated adenocarcinomas, respectively. The mean Ki-67 LI of well, moderate and poorly differentiated adenocarcinomas were 14.25, 31.34 and 43.08 respectively, and their difference was statistically significant, correlation was also noted with stage. CDX2 expression appeared to be stronger in poorly differentiated cases, but there was no significant difference in its expression in the different grades and stages. There was no correlation between Ki-67 LI and CDX2 immunostaining pattern. The lymph node metastasis showed CDX2 positivity in all the cases. Expression of CDX2 does not significantly change with the grade of colorectal adenocarcinomas. However, it is an important diagnostic marker in metastatic colonic lesions. The Ki-67 LI, on the other hand, showed a strong correlation with histopathological grades.

Distribution of fluorescently labeled tubulin injected into sand dollar eggs from fertilization through cleavage.

PubMed

Hamaguchi, Y; Toriyama, M; Sakai, H; Hiramoto, Y

1985-04-01

Porcine brain tubulin labeled with fluorescein isothiocyanate (FITC) was able to polymerize by itself and co-polymerize with tubulin purified from starfish sperm flagella. When we injected the FITC-labeled tubulin into unfertilized eggs of the sand dollar, Clypeaster japonicus, and the eggs were then fertilized, the labeled tubulin was incorporated into the sperm aster. When injected into fertilized eggs at streak stage, the tubulin was quickly incorporated into each central region of growing asters. It was clearly visualized that the labeled tubulin, upon reaching metaphase, accumulated in the mitotic apparatus and later disappeared over the cytoplasm during interphase. The accumulation of the fluorescence in the mitotic apparatus was observed repeatedly at successive cleavage. After lysis of the fertilized eggs with a microtubule-stabilizing solution, fluorescent fibrous structures around the nucleus and those of the sperm aster and the mitotic apparatus were preserved and coincided with the fibrous structures observed by polarization and differential interference microscopy. We found the FITC-labeled tubulin to be incorporated into the entire mitotic apparatus within 20-30 s when injected into the eggs at metaphase or anaphase. This rapid incorporation of the labeled tubulin into the mitotic apparatus suggests that the equilibrium between mitotic microtubules and tubulin is attained very rapidly in the living eggs. Axonemal tubulin purified from starfish sperm flagella and labeled with FITC was also incorporated into microtubular structures in the same fashion as the FITC-labeled brain tubulin. These results suggest that even FITC-labeled heterogeneous tubulins undergo spatial and stage-specific regulation of assembly-disassembly in the same manner as does sand dollar egg tubulin.

Distribution of fluorescently labeled tubulin injected into sand dollar eggs from fertilization through cleavage

PubMed Central

1985-01-01

Porcine brain tubulin labeled with fluorescein isothiocyanate (FITC) was able to polymerize by itself and co-polymerize with tubulin purified from starfish sperm flagella. When we injected the FITC- labeled tubulin into unfertilized eggs of the sand dollar, Clypeaster japonicus, and the eggs were then fertilized, the labeled tubulin was incorporated into the sperm aster. When injected into fertilized eggs at streak stage, the tubulin was quickly incorporated into each central region of growing asters. It was clearly visualized that the labeled tubulin, upon reaching metaphase, accumulated in the mitotic apparatus and later disappeared over the cytoplasm during interphase. The accumulation of the fluorescence in the mitotic apparatus was observed repeatedly at successive cleavage. After lysis of the fertilized eggs with a microtubule-stabilizing solution, fluorescent fibrous structures around the nucleus and those of the sperm aster and the mitotic apparatus were preserved and coincided with the fibrous structures observed by polarization and differential interference microscopy. We found the FITC-labeled tubulin to be incorporated into the entire mitotic apparatus within 20-30 s when injected into the eggs at metaphase or anaphase. This rapid incorporation of the labeled tubulin into the mitotic apparatus suggests that the equilibrium between mitotic microtubules and tubulin is attained very rapidly in the living eggs. Axonemal tubulin purified from starfish sperm flagella and labeled with FITC was also incorporated into microtubular structures in the same fashion as the FITC-labeled brain tubulin. These results suggest that even FITC-labeled heterogeneous tubulins undergo spatial and stage-specific regulation of assembly-disassembly in the same manner as does sand dollar egg tubulin. PMID:3920225

Activation of the mouse Oct4 promoter in medaka embryonic stem cells and its use for ablation of spontaneous differentiation.

PubMed

Hong, Yunhan; Winkler, Christoph; Liu, Tongming; Chai, Guixuan; Schartl, Manfred

2004-07-01

The determination and maintenance of the cell fate is ultimately due to differential gene activity. In the mouse, expression of the transcription factor Oct4 is high in totipotent inner cell mass, germ cells and undifferentiated embryonic stem (ES) cells, but dramatically reduced or extinct upon differentiation. Here, we show that medaka blastula embryos and cells of the ES cell line MES1 are able to activate the Oct4 promoter. Ectopic expression of a fusion gene for beta-galactosidase and neomycin resistance from the Oct4 promoter conferred resistance to G418. G418 selection led to a homogeneous population of undifferentiated ES cells which were able to undergo induced or directed differentiation into various cell types including neuron-like cells and melanocytes. Furthermore, GFP-labeled GOF18geo-MES1 cells after differentiation ablation were able to contribute to a wide variety of organ systems derived from all the three germ layers. Most importantly, we show that drug ablation of differentiation on the basis of Oct4 promoter is a useful tool to improve ES cell cultivation and chimera formation: MES1 cells after differentiation ablation appeared to be better donors than the parental MES1 line, as the permissive number of input donor cells increases from 100 to 200, resulting in an enhanced degree of chimerism. Taken together, some transcription factors and cis-acting regulatory sequences controlling totipotency-specific gene expression appear to be conserved between mammals and fish, and medaka ES cells offer an in vitro system for characterizing the expression of totipotency-specific genes such as putative Oct4 homologs from fish.

Graphene supports in vitro proliferation and osteogenic differentiation of goat adult mesenchymal stem cells: potential for bone tissue engineering.

PubMed

Elkhenany, Hoda; Amelse, Lisa; Lafont, Andersen; Bourdo, Shawn; Caldwell, Marc; Neilsen, Nancy; Dervishi, Enkeleda; Derek, Oshin; Biris, Alexandru S; Anderson, David; Dhar, Madhu

2015-04-01

Current treatments for bone loss injuries involve autologous and allogenic bone grafts, metal alloys and ceramics. Although these therapies have proved useful, they suffer from inherent challenges, and hence, an adequate bone replacement therapy has not yet been found. We hypothesize that graphene may be a useful nanoscaffold for mesenchymal stem cells and will promote proliferation and differentiation into bone progenitor cells. In this study, we evaluate graphene, a biocompatible inert nanomaterial, for its effect on in vitro growth and differentiation of goat adult mesenchymal stem cells. Cell proliferation and differentiation are compared between polystyrene-coated tissue culture plates and graphene-coated plates. Graphitic materials are cytocompatible and support cell adhesion and proliferation. Importantly, cells seeded on to oxidized graphene films undergo osteogenic differentiation in fetal bovine serum-containing medium without the addition of any glucocorticoid or specific growth factors. These findings support graphene's potential to act as an osteoinducer and a vehicle to deliver mesenchymal stem cells, and suggest that the combination of graphene and goat mesenchymal stem cells provides a promising construct for bone tissue engineering. Copyright © 2014 John Wiley & Sons, Ltd.

Induced-Pluripotent-Stem-Cell-Derived Primitive Macrophages Provide a Platform for Modeling Tissue-Resident Macrophage Differentiation and Function.

PubMed

Takata, Kazuyuki; Kozaki, Tatsuya; Lee, Christopher Zhe Wei; Thion, Morgane Sonia; Otsuka, Masayuki; Lim, Shawn; Utami, Kagistia Hana; Fidan, Kerem; Park, Dong Shin; Malleret, Benoit; Chakarov, Svetoslav; See, Peter; Low, Donovan; Low, Gillian; Garcia-Miralles, Marta; Zeng, Ruizhu; Zhang, Jinqiu; Goh, Chi Ching; Gul, Ahmet; Hubert, Sandra; Lee, Bernett; Chen, Jinmiao; Low, Ivy; Shadan, Nurhidaya Binte; Lum, Josephine; Wei, Tay Seok; Mok, Esther; Kawanishi, Shohei; Kitamura, Yoshihisa; Larbi, Anis; Poidinger, Michael; Renia, Laurent; Ng, Lai Guan; Wolf, Yochai; Jung, Steffen; Önder, Tamer; Newell, Evan; Huber, Tara; Ashihara, Eishi; Garel, Sonia; Pouladi, Mahmoud A; Ginhoux, Florent

2017-07-18

Tissue macrophages arise during embryogenesis from yolk-sac (YS) progenitors that give rise to primitive YS macrophages. Until recently, it has been impossible to isolate or derive sufficient numbers of YS-derived macrophages for further study, but data now suggest that induced pluripotent stem cells (iPSCs) can be driven to undergo a process reminiscent of YS-hematopoiesis in vitro. We asked whether iPSC-derived primitive macrophages (iMacs) can terminally differentiate into specialized macrophages with the help of growth factors and organ-specific cues. Co-culturing human or murine iMacs with iPSC-derived neurons promoted differentiation into microglia-like cells in vitro. Furthermore, murine iMacs differentiated in vivo into microglia after injection into the brain and into functional alveolar macrophages after engraftment in the lung. Finally, iPSCs from a patient with familial Mediterranean fever differentiated into iMacs with pro-inflammatory characteristics, mimicking the disease phenotype. Altogether, iMacs constitute a source of tissue-resident macrophage precursors that can be used for biological, pathophysiological, and therapeutic studies. Copyright © 2017 Elsevier Inc. All rights reserved.

A Noninvasive In Vitro Monitoring System Reporting Skeletal Muscle Differentiation.

PubMed

Öztürk-Kaloglu, Deniz; Hercher, David; Heher, Philipp; Posa-Markaryan, Katja; Sperger, Simon; Zimmermann, Alice; Wolbank, Susanne; Redl, Heinz; Hacobian, Ara

2017-01-01

Monitoring of cell differentiation is a crucial aspect of cell-based therapeutic strategies depending on tissue maturation. In this study, we have developed a noninvasive reporter system to trace murine skeletal muscle differentiation. Either a secreted bioluminescent reporter (Metridia luciferase) or a fluorescent reporter (green fluorescent protein [GFP]) was placed under the control of the truncated muscle creatine kinase (MCK) basal promoter enhanced by variable numbers of upstream MCK E-boxes. The engineered pE3MCK vector, coding a triple tandem of E-Boxes and the truncated MCK promoter, showed twentyfold higher levels of luciferase activation compared with a Cytomegalovirus (CMV) promoter. This newly developed reporter system allowed noninvasive monitoring of myogenic differentiation in a straining bioreactor. Additionally, binding sequences of endogenous microRNAs (miRNAs; seed sequences) that are known to be downregulated in myogenesis were ligated as complementary seed sequences into the reporter vector to reduce nonspecific signal background. The insertion of seed sequences improved the signal-to-noise ratio up to 25% compared with pE3MCK. Due to the highly specific, fast, and convenient expression analysis for cells undergoing myogenic differentiation, this reporter system provides a powerful tool for application in skeletal muscle tissue engineering.

Analysis of integrated multiple 'omics' datasets reveals the mechanisms of initiation and determination in the formation of tuberous roots in Rehmannia glutinosa.

PubMed

Li, Mingjie; Yang, Yanhui; Li, Xinyu; Gu, Li; Wang, Fengji; Feng, Fajie; Tian, Yunhe; Wang, Fengqing; Wang, Xiaoran; Lin, Wenxiong; Chen, Xinjian; Zhang, Zhongyi

2015-09-01

All tuberous roots in Rehmannia glutinosa originate from the expansion of fibrous roots (FRs), but not all FRs can successfully transform into tuberous roots. This study identified differentially expressed genes and proteins associated with the expansion of FRs, by comparing the tuberous root at expansion stages (initiated tuberous root, ITRs) and FRs at the seedling stage (initiated FRs, IFRs). The role of miRNAs in the expansion of FRs was also explored using the sRNA transcriptome and degradome to identify miRNAs and their target genes that were differentially expressed between ITRs and FRs at the mature stage (unexpanded FRs, UFRs, which are unable to expand into ITRs). A total of 6032 genes and 450 proteins were differentially expressed between ITRs and IFRs. Integrated analyses of these data revealed several genes and proteins involved in light signalling, hormone response, and signal transduction that might participate in the induction of tuberous root formation. Several genes related to cell division and cell wall metabolism were involved in initiating the expansion of IFRs. Of 135 miRNAs differentially expressed between ITRs and UFRs, there were 27 miRNAs whose targets were specifically identified in the degradome. Analysis of target genes showed that several miRNAs specifically expressed in UFRs were involved in the degradation of key genes required for the formation of tuberous roots. As far as could be ascertained, this is the first time that the miRNAs that control the transition of FRs to tuberous roots in R. glutinosa have been identified. This comprehensive analysis of 'omics' data sheds new light on the mechanisms involved in the regulation of tuberous roots formation. © The Author 2015. Published by Oxford University Press on behalf of the Society for Experimental Biology. All rights reserved. For permissions, please email: journals.permissions@oup.com.

Identification of multiple dmrt1s in catfish: localization, dimorphic expression pattern, changes during testicular cycle and after methyltestosterone treatment.

PubMed

Raghuveer, K; Senthilkumaran, B

2009-05-01

The double sex and mab-3 related (DM) transcription factor 1 (dmrt1) plays an important role in testicular differentiation. Here, we report cloning of multiple dmrt1s, a full-length and two alternative spliced forms from adult catfish (Clarias gariepinus) testis, which encode predicted proteins of 287 (dmrt1a), 253 (dmrt1b) and 233 (dmrt1c) amino acid residues respectively. Interestingly, dmrt1c lacks the majority of the DM domain. Multiple dmrt1s (dmrt1a and dmrt1c) were obtained from Clarias batrachus as well. Tissue distribution (transcript and protein) of catfish dmrt1 revealed exclusive expression in testis. Semi-quantitative RT-PCR revealed the presence of multiple dmrt1s with high levels of dmrt1a in adult testis but not in ovary. Real-time RT-PCR analysis during testicular cycle showed higher levels of dmrt1 transcripts in preparatory and pre-spawning when compared with spawning and post-spawning phases. Immunocytochemical and immunofluorescence localization revealed the presence of catfish Dmrt1 protein in spermatogonia and spermatocytes, which indicates plausible role in spermatogenesis. Histological analysis indicated initiation of gonadal sex differentiation in catfish around 40-50 days after hatching. The potential role for dmrt1 in testicular differentiation is evident from its stage-dependent elevated expression in developing testis. Furthermore, dimorphic expressions of dmrt1s were evident at different stages of gonadal development or recrudescence in catfish. Treatment of methyl testosterone (MT) during early stages of gonadal sex differentiation resulted in adult males. Interestingly, we also obtained MT-treated fishes having ova-testis gonads. Analysis of dmrt1, sox9a, foxl2 and cyp19a1 expression patterns in MT-treated gonads revealed tissue-specific pattern. These results together suggest that multiple dmrt1s are testis-specific markers in catfish.

The chemistry of caries: remineralization and demineralization events with direct clinical relevance.

PubMed

González-Cabezas, Carlos

2010-07-01

Dental caries is a site-specific disease that undergoes many cycles of demineralization and remineralization during lesion development. Because of its developmental characteristics dynamics, the caries lesion can be arrested and even repaired at its early stages without operative intervention by increasing the net mineral gain during the demineralization and remineralization cycles. This result can be accomplished by reducing the effect of etiological factors such as cariogenic biofilms and diet, and increasing the efficacy of remineralizing agents such as saliva and fluoride. Copyright 2010 Elsevier Inc. All rights reserved.

Comprehensive Plasma Metabolomic Analyses of Atherosclerotic Progression Reveal Alterations in Glycerophospholipid and Sphingolipid Metabolism in Apolipoprotein E-deficient Mice

PubMed Central

Dang, Vi T.; Huang, Aric; Zhong, Lexy H.; Shi, Yuanyuan; Werstuck, Geoff H.

2016-01-01

Atherosclerosis is the major underlying cause of most cardiovascular diseases. Despite recent advances, the molecular mechanisms underlying the pathophysiology of atherogenesis are not clear. In this study, comprehensive plasma metabolomics were used to investigate early-stage atherosclerotic development and progression in chow-fed apolipoprotein E-deficient mice at 5, 10 and 15 weeks of age. Comprehensive plasma metabolomic profiles, based on 4365 detected metabolite features, differentiate atherosclerosis-prone from atherosclerosis-resistant models. Metabolites in the sphingomyelin pathway were significantly altered prior to detectable lesion formation and at all subsequent time-points. The cytidine diphosphate-diacylglycerol pathway was up-regulated during stage I of atherosclerosis, while metabolites in the phosphatidylethanolamine and glycosphingolipid pathways were augmented in mice with stage II lesions. These pathways, involving glycerophospholipid and sphingolipid metabolism, were also significantly affected during the course of atherosclerotic progression. Our findings suggest that distinct plasma metabolomic profiles can differentiate the different stages of atherosclerotic progression. This study reveals that alteration of specific, previously unreported pathways of glycerophospholipid and sphingolipid metabolism are associated with atherosclerosis. The clear difference in the level of several metabolites supports the use of plasma lipid profiling as a diagnostic tool of atherogenesis. PMID:27721472

Similarities in neural activations of face and Chinese character discrimination.

PubMed

Liu, Jiangang; Tian, Jie; Li, Jun; Gong, Qiyong; Lee, Kang

2009-02-18

This study compared Chinese participants' visual discrimination of Chinese faces with that of Chinese characters, which are highly similar to faces on a variety of dimensions. Both Chinese faces and characters activated the bilateral middle fusiform with high levels of correlations. These findings suggest that although the expertise systems for faces and written symbols are known to be anatomically differentiated at the later stages of processing to serve face processing or written-symbol-specific processing purposes, they may share similar neural structures in the ventral occipitotemporal cortex at the stages of visual processing.

Pulmonary Rehabilitation in Improving Lung Function in Patients With Locally Advanced Non-Small Cell Lung Cancer Undergoing Chemoradiation

ClinicalTrials.gov

2017-04-12

Cachexia; Fatigue; Pulmonary Complications; Radiation Toxicity; Recurrent Non-small Cell Lung Cancer; Stage IIIA Non-small Cell Lung Cancer; Stage IIIB Non-small Cell Lung Cancer; Stage IV Non-small Cell Lung Cancer

Canine Platelet Lysate Is Inferior to Fetal Bovine Serum for the Isolation and Propagation of Canine Adipose Tissue- and Bone Marrow-Derived Mesenchymal Stromal Cells

PubMed Central

Russell, Keith A.; Gibson, Thomas W. G.; Chong, Andrew; Co, Carmon; Koch, Thomas G.

2015-01-01

Background Mesenchymal stromal cells (MSC) are increasingly investigated for their clinical utility in dogs. Fetal bovine serum (FBS) is a common culture supplement used for canine MSC expansion. However, FBS content is variable, its clinical use carries risk of an immune response, and its cost is increasing due to global demand. Platelet lysate (PL) has proven to be a suitable alternative to FBS for expansion of human MSC. Hypothesis and Objectives We hypothesized that canine adipose tissue (AT) and bone marrow (BM) MSC could be isolated and expanded equally in PL and FBS at conventionally-used concentrations with differentiation of these MSC unaffected by choice of supplement. Our objectives were to evaluate the use of canine PL in comparison with FBS at four stages: 1) isolation, 2) proliferation, 3) spontaneous differentiation, and 4) directed differentiation. Results 1) Medium with 10% PL was unable to isolate MSC. 2) MSC, initially isolated in FBS-supplemented media, followed a dose-dependent response with no significant difference between PL and FBS cultures at up to 20% (AT) or 30% (BM) enrichment. Beyond these respective peaks, proliferation fell in PL cultures only, while a continued dose-dependent proliferation response was noted in FBS cultures. 3) Further investigation indicated PL expansion culture was inducing spontaneous adipogenesis in concentrations as low as 10% and as early as 4 days in culture. 4) MSC isolated in FBS, but expanded in either FBS or PL, maintained ability to undergo directed adipogenesis and osteogenesis, but not chondrogenesis. Conclusions/Significance Canine PL did not support establishment of MSC colonies from AT and BM, nor expansion of MSC, which appear to undergo spontaneous adipogenesis in response to PL exposure. In vivo studies are warranted to determine if concurrent use of MSC with any platelet-derived products such as platelet-rich plasma are associated with synergistic, neutral or antagonistic effects. PMID:26353112