Donor age and C1orf132/MIR29B2C determine age-related methylation signature of blood after allogeneic hematopoietic stem cell transplantation.

PubMed

Spólnicka, Magdalena; Piekarska, Renata Zbieć; Jaskuła, Emilia; Basak, Grzegorz W; Jacewicz, Renata; Pięta, Agnieszka; Makowska, Żanetta; Jedrzejczyk, Maciej; Wierzbowska, Agnieszka; Pluta, Agnieszka; Robak, Tadeusz; Berent, Jarosław; Branicki, Wojciech; Jędrzejczak, Wiesław; Lange, Andrzej; Płoski, Rafał

2016-01-01

Our recent study demonstrated that DNA methylation status in a set of CpGs located in ELOVL2, C1orf132, TRIM59, KLF14, and FHL2 can accurately predict calendar age in blood. In the present work, we used these markers to evaluate the effect of allogeneic hematopoietic stem cell transplantation (HSCT) on the age-related methylation signature of human blood. DNA methylation in 32 CpGs was investigated in 16 donor-recipient pairs using pyrosequencing. DNA was isolated from the whole blood collected from recipients 27-360 days (mean 126) after HSCT and from the donors shortly before the HSCT. It was found that in the recipients, the predicted age did not correlate with their calendar age but was correlated with the calendar age (r = 0.94, p = 4 × 10(-8)) and predicted age (r = 0.97, p = 5 × 10(-10)) of a respective donor. Despite this strong correlation, the predicted age of a recipient was consistently lower than the predicted age of a donor by 3.7 years (p = 7.8 × 10(-4)). This shift was caused by hypermethylation of the C1orf132 CpGs, for C1orf132 CpG_1. Intriguingly, the recipient-donor methylation difference correlated with calendar age of the donor (r = 0.76, p = 6 × 10(-4)). This finding could not trivially be explained by shifts of the major cellular factions of blood. We confirm the single previous report that after HSCT, the age of the donor is the major determinant of age-specific methylation signature in recipient's blood. A novel finding is the unique methylation dynamics of C1orf132 which encodes MIR29B2C implicated in the self-renewing of hematopoietic stem cells. This observation suggests that C1orf132 could influence graft function after HSCT.

Nuclease footprint analyses of the interactions between RNase P ribozyme and a model mRNA substrate.

PubMed Central

Trang, P; Hsu, A W; Liu, F

1999-01-01

RNase P ribozyme cleaves an RNA helix substrate which resembles the acceptor stem and T-stem structures of its natural tRNA substrate. By linking the ribozyme covalently to a sequence (guide sequence) complementary to a target RNA, the catalytic RNA can be converted into a sequence-specific ribozyme, M1GS RNA. We have previously shown that M1GS RNA can efficiently cleave the mRNA sequence encoding thymidine kinase (TK) of herpes simplex virus 1. In this study, a footprint procedure using different nucleases was carried out to map the regions of a M1GS ribozyme that potentially interact with the TK mRNA substrate. The ribozyme regions that are protected from nuclease degradation in the presence of the TK mRNA substrate include those that interact with the acceptor stem and T-stem, the 3' terminal CCA sequence and the cleavage site of a tRNA substrate. However, some of the protected regions (e.g. P13 and P14) are unique and not among those protected in the presence of a tRNA substrate. Identification of the regions that interact with a mRNA substrate will allow us to study how M1GS RNA recognizes a mRNA substrate and facilitate the development of mRNA-cleaving ribozymes for gene-targeting applications. PMID:10556315

Improving the efficiency of CHO cell line generation using glutamine synthetase gene knockout cells.

PubMed

Fan, Lianchun; Kadura, Ibrahim; Krebs, Lara E; Hatfield, Christopher C; Shaw, Margaret M; Frye, Christopher C

2012-04-01

Although Chinese hamster ovary (CHO) cells, with their unique characteristics, have become a major workhorse for the manufacture of therapeutic recombinant proteins, one of the major challenges in CHO cell line generation (CLG) is how to efficiently identify those rare, high-producing clones among a large population of low- and non-productive clones. It is not unusual that several hundred individual clones need to be screened for the identification of a commercial clonal cell line with acceptable productivity and growth profile making the cell line appropriate for commercial application. This inefficiency makes the process of CLG both time consuming and laborious. Currently, there are two main CHO expression systems, dihydrofolate reductase (DHFR)-based methotrexate (MTX) selection and glutamine synthetase (GS)-based methionine sulfoximine (MSX) selection, that have been in wide industrial use. Since selection of recombinant cell lines in the GS-CHO system is based on the balance between the expression of the GS gene introduced by the expression plasmid and the addition of the GS inhibitor, L-MSX, the expression of GS from the endogenous GS gene in parental CHOK1SV cells will likely interfere with the selection process. To study endogenous GS expression's potential impact on selection efficiency, GS-knockout CHOK1SV cell lines were generated using the zinc finger nuclease (ZFN) technology designed to specifically target the endogenous CHO GS gene. The high efficiency (∼2%) of bi-allelic modification on the CHO GS gene supports the unique advantages of the ZFN technology, especially in CHO cells. GS enzyme function disruption was confirmed by the observation of glutamine-dependent growth of all GS-knockout cell lines. Full evaluation of the GS-knockout cell lines in a standard industrial cell culture process was performed. Bulk culture productivity improved two- to three-fold through the use of GS-knockout cells as parent cells. The selection stringency was significantly increased, as indicated by the large reduction of non-producing and low-producing cells after 25 µM L-MSX selection, and resulted in a six-fold efficiency improvement in identifying similar numbers of high-productive cell lines for a given recombinant monoclonal antibody. The potential impact of GS-knockout cells on recombinant protein quality is also discussed. Copyright © 2011 Wiley Periodicals, Inc.

A protein/antibiotic releasing poly(lactic-co-glycolic acid)/lecithin scaffold for bone repair applications.

PubMed

Shi, Xuetao; Wang, Yingjun; Ren, Li; Huang, Wei; Wang, Dong-An

2009-05-21

Novel poly(lactic-co-glycolic acid) (PLGA)-hybridizing-lecithin scaffolds loaded with drug or protein were prepared with water/oil/water techniques and sintering microspheres technique. In such fabricated composite scaffolds (abbreviated "PLGA/Lec-SMS"), the introduction of lecithin component has been proven capable of largely enhancing Gentamicin (GS) and protein (Bovine Serum Albumin) encapsulation efficiency. The in vitro GS and BSA releasing profiles of PLGA/Lec-SMS system were plotted basing over 60 days' and 18 days' data collection, respectively. It indicates a sustained releasing tendency despite a burst at the very beginning. The antibacterial properties of GS-laden scaffolds were determined in vitro, and the antibacterial activity of scaffolds was enhanced by incorporating lecithin into PLGA bulks. Additionally, mesenchymal stem cells (MSCs) were seeded onto PLGA-SMS and PLGA/Lec-SMS in vitro. The outcome confirmed PLGA/Lec(5%)-SMS functions to improve MSC proliferation and also to enhance general ALP production and calcium secretion which is the vital markers for osteogenesis. In conclusion, this newly designed antibiotic releasing PLGA/Lec-SMS is promising for bone-repairing therapeutics.

Glucose dependence of glycogen synthase activity regulation by GSK3 and MEK/ERK inhibitors and angiotensin-(1-7) action on these pathways in cultured human myotubes.

PubMed

Montori-Grau, Marta; Tarrats, Núria; Osorio-Conles, Oscar; Orozco, Anna; Serrano-Marco, Lucía; Vázquez-Carrera, Manuel; Gómez-Foix, Anna M

2013-05-01

Glycogen synthase (GS) is activated by glucose/glycogen depletion in skeletal muscle cells, but the contributing signaling pathways, including the chief GS regulator GSK3, have not been fully defined. The MEK/ERK pathway is known to regulate GSK3 and respond to glucose. The aim of this study was to elucidate the GSK3 and MEK/ERK pathway contribution to GS activation by glucose deprivation in cultured human myotubes. Moreover, we tested the glucose-dependence of GSK3 and MEK/ERK effects on GS and angiotensin (1-7) actions on these pathways. We show that glucose deprivation activated GS, but did not change phospho-GS (Ser640/1), GSK3β activity or activity-activating phosphorylation of ERK1/2. We then treated glucose-replete and -depleted cells with SB415286, U0126, LY294 and rapamycin to inhibit GSK3, MEK1/2, PI3K and mTOR, respectively. SB415286 activated GS and decreased the relative phospho-GS (Ser640/1) level, more in glucose-depleted than -replete cells. U0126 activated GS and reduced the phospho-GS (Ser640/1) content significantly in glucose-depleted cells, while GSK3β activity tended to increase. LY294 inactivated GS in glucose-depleted cells only, without affecting relative phospho-GS (Ser640/1) level. Rapamycin had no effect on GS activation. Angiotensin-(1-7) raised phospho-ERK1/2 but not phospho-GSK3β (Ser9) content, while it inactivated GS and increased GS phosphorylation on Ser640/1, in glucose-replete cells. In glucose-depleted cells, angiotensin-(1-7) effects on ERK1/2 and GS were reverted, while relative phospho-GSK3β (Ser9) content decreased. In conclusion, activation of GS by glucose deprivation is not due to GS Ser640/1 dephosphorylation, GSK3β or ERK1/2 regulation in cultured myotubes. However, glucose depletion enhances GS activation/Ser640/1 dephosphorylation due to both GSK3 and MEK/ERK inhibition. Angiotensin-(1-7) inactivates GS in glucose-replete cells in association with ERK1/2 activation, not with GSK3 regulation, and glucose deprivation reverts both hormone effects. Thus, the ERK1/2 pathway negatively regulates GS activity in myotubes, without involving GSK3 regulation, and as a function of the presence of glucose. Copyright © 2013 Elsevier Inc. All rights reserved.

Natural biosorbents (garlic stem and horse chesnut shell) for removal of chromium(VI) from aqueous solutions.

PubMed

Parlayıcı, Şerife; Pehlivan, Erol

2015-12-01

The biosorption of Cr(VI) by the garlic stem (GS)-Allium sativum L. and horse chesnut shell (HCS)-Aesculus hippocastanum plant residues in a batch type reactor was studied in detail for the purpose of wastewater treatment. The influence of initial Cr(VI) concentration, time, and pH was investigated to optimize Cr(VI) removal from aqueous solutions and equilibrium isotherms and kinetic data. This influence was evaluated. The adsorption capacity of the GS and the HCS for Cr(VI) was determined with the Langmuir and Freundlich isotherm models, and the data was fitted to the Langmuir. The adsorption capacity of the GS and the HCS was found to be 103.09 and 142.85 mg/g of adsorbent from a solution containing 3000 ppm of Cr(VI), respectively. The GS's capacity was considerably lower than that of the HCS in its natural form. Gibbs free energy was spontaneous for all interactions, and the adsorption process exhibited exothermic enthalpy values. The HCS was shown to be a promising biosorbent for Cr(VI) removal from aqueous solutions.

Design and synthesis of 2-oxindole based multi-targeted inhibitors of PDK1/Akt signaling pathway for the treatment of glioblastoma multiforme.

PubMed

Sestito, Simona; Nesi, Giulia; Daniele, Simona; Martelli, Alma; Digiacomo, Maria; Borghini, Alice; Pietra, Daniele; Calderone, Vincenzo; Lapucci, Annalina; Falasca, Marco; Parrella, Paola; Notarangelo, Angelantonio; Breschi, Maria C; Macchia, Marco; Martini, Claudia; Rapposelli, Simona

2015-11-13

Aggressive behavior and diffuse infiltrative growth are the main features of Glioblastoma multiforme (GBM), together with the high degree of resistance and recurrence. Evidence indicate that GBM-derived stem cells (GSCs), endowed with unlimited proliferative potential, play a critical role in tumor development and maintenance. Among the many signaling pathways involved in maintaining GSC stemness, tumorigenic potential, and anti-apoptotic properties, the PDK1/Akt pathway is a challenging target to develop new potential agents able to affect GBM resistance to chemotherapy. In an effort to find new PDK1/Akt inhibitors, we rationally designed and synthesized a small family of 2-oxindole derivatives. Among them, compound 3 inhibited PDK1 kinase and downstream effectors such as CHK1, GS3Kα and GS3Kβ, which contribute to GCS survival. Compound 3 appeared to be a good tool for studying the role of the PDK1/Akt pathway in GCS self-renewal and tumorigenicity, and might represent the starting point for the development of more potent and focused multi-target therapies for GBM. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

Glutamine synthetase gene knockout-human embryonic kidney 293E cells for stable production of monoclonal antibodies.

PubMed

Yu, Da Young; Lee, Sang Yoon; Lee, Gyun Min

2018-05-01

Previously, it was inferred that a high glutamine synthetase (GS) activity in human embryonic kidney (HEK) 293E cells results in elevated resistance to methionine sulfoximine (MSX) and consequently hampers GS-mediated gene amplification and selection by MSX. To overcome this MSX resistance in HEK293E cells, a GS-knockout HEK293E cell line was generated using the CRISPR/Cas9 system to target the endogenous human GS gene. The GS-knockout in the HEK293E cell line (RK8) was confirmed by Western blot analysis of GS and by observation of glutamine-dependent growth. Unlike the wild type HEK293E cells, the RK8 cells were successfully used as host cells to generate a recombinant HEK293E cell line (rHEK293E) producing a monoclonal antibody (mAb). When the RK8 cells were transfected with the GS expression vector containing the mAb gene, rHEK293E cells producing the mAb could be selected in the absence as well as in the presence of MSX. The gene copies and mRNA expression levels of the mAb in rHEK293E cells were also quantified using qRT-PCR. Taken together, the GS-knockout HEK293E cell line can be used as host cells to generate stable rHEK293E cells producing a mAb through GS-mediated gene selection in the absence as well as in the presence of MSX. © 2018 Wiley Periodicals, Inc.

Limitations to the development of recombinant human embryonic kidney 293E cells using glutamine synthetase-mediated gene amplification: Methionine sulfoximine resistance.

PubMed

Yu, Da Young; Noh, Soo Min; Lee, Gyun Min

2016-08-10

To investigate the feasibility of glutamine synthetase (GS)-mediated gene amplification in HEK293 cells for the high-level stable production of therapeutic proteins, HEK293E cells were transfected by the GS expression vector containing antibody genes and were selected at various methionine sulfoximine (MSX) concentrations in 96-well plates. For a comparison, CHOK1 cells were transfected by the same GS expression vector and selected at various MSX concentrations. Unlike CHOK1 cells, HEK293E cells producing high levels of antibodies were not selected at all. For HEK293E cells, the number of wells with the cell pool did not decrease with an increase in the concentration of MSX up to 500μM MSX. A q-RT-PCR analysis confirmed that the antibody genes in the HEK293E cells, unlike the CHOK1 cells, were not amplified after increasing the MSX concentration. It was found that the GS activity in HEK293E cells was much higher than that in CHOK1 cells (P<0.05). In a glutamine-free medium, the GS activity of HEK293E cells was approximately 4.8 times higher than that in CHOK1 cells. Accordingly, it is inferred that high GS activity of HEK293E cells results in elevated resistance to MSX and therefore hampers GS-mediated gene amplification by MSX. Thus, in order to apply the GS-mediated gene amplification system to HEK293 cells, the endogenous GS expression level in HEK293 cells needs to be minimized by knock-out or down-regulation methods. Copyright © 2016 Elsevier B.V. All rights reserved.

Epigallocatechin Gallate-Modified Gelatin Sponges Treated by Vacuum Heating as a Novel Scaffold for Bone Tissue Engineering.

PubMed

Honda, Yoshitomo; Takeda, Yoshihiro; Li, Peiqi; Huang, Anqi; Sasayama, Satoshi; Hara, Eiki; Uemura, Naoya; Ueda, Mamoru; Hashimoto, Masanori; Arita, Kenji; Matsumoto, Naoyuki; Hashimoto, Yoshiya; Baba, Shunsuke; Tanaka, Tomonari

2018-04-11

Chemical modification of gelatin using epigallocatechin gallate (EGCG) promotes bone formation in vivo. However, further improvements are required to increase the mechanical strength and bone-forming ability of fabricated EGCG-modified gelatin sponges (EGCG-GS) for practical applications in regenerative therapy. In the present study, we investigated whether vacuum heating-induced dehydrothermal cross-linking of EGCG-GS enhances bone formation in critical-sized rat calvarial defects. The bone-forming ability of vacuum-heated EGCG-GS (vhEGCG-GS) and other sponges was evaluated by micro-computed tomography and histological staining. The degradation of sponges was assessed using protein assays, and cell morphology and proliferation were verified by scanning electron microscopy and immunostaining using osteoblastic UMR106 cells in vitro. Four weeks after the implantation of sponges, greater bone formation was detected for vhEGCG-GS than for EGCG-GS or vacuum-heated gelatin sponges (dehydrothermal cross-linked sponges without EGCG). In vitro experiments revealed that the relatively low degradability of vhEGCG-GS supports cell attachment, proliferation, and cell-cell communication on the matrix. These findings suggest that vacuum heating enhanced the bone forming ability of EGCG-GS, possibly via the dehydrothermal cross-linking of EGCG-GS, which provides a scaffold for cells, and by maintaining the pharmacological effect of EGCG.

Electron Transport Controls Glutamine Synthetase Activity in the Facultative Heterotrophic Cyanobacterium Synechocystis sp. PCC 6803.

PubMed Central

Reyes, J. C.; Crespo, J. L.; Garcia-Dominguez, M.; Florencio, F. J.

1995-01-01

Glutamine synthetase (GS) from Synechocystis sp. PCC 6803 was inactivated in vivo by transferring cells from light to darkness or by incubation with the photosynthetic inhibitor 3-(3,4-dichlorophenyl)-1,1-dimethylurea but not with 2,5-dibromo-3-methyl-6-isopropyl-p-benzoquinone. Addition of glucose prevented both dark and 3-(3,4-dichlorophenyl)-1,1-dimethylurea GS inactivation. In a Synechocystis psbE-psbF mutant (T1297) lacking photosystem II, glucose was required to maintain active GS, even in the light. However, in nitrogen-starved T1297 cells the removal of glucose did not affect GS activity. The fact that dark-inactivated GS was reactivated in vitro by the same treatments that reactivate the ammonium-inactivated GS points out that both nitrogen metabolism and redox state of the cells lead to the same molecular regulatory mechanism in the control of GS activity. Using GS antibodies we detected that dark-inactivated GS displayed a different electrophoretic migration with respect to the active form in nondenaturing polyacrylamide gel electrophoresis but not in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The possible pathway to modulate GS activity by the electron transport flow in Synechocystis cells is discussed. PMID:12228640

Guggulsterone decreases proliferation and metastatic behavior of pancreatic cancer cells by modulating JAK/STAT and Src/FAK signaling.

PubMed

Macha, Muzafar A; Rachagani, Satyanarayana; Gupta, Suprit; Pai, Priya; Ponnusamy, Moorthy P; Batra, Surinder K; Jain, Maneesh

2013-12-01

Inadequate efficacy, high toxicity and drug resistance associated with existing chemotherapeutic agents mandate a need for novel therapeutic strategies for highly aggressive Pancreatic Cancer (PC). Guggulsterone (GS) exhibits potent anti-proliferative effects against various cancer cells and has emerged as an attractive candidate for use in complementary or preventive cancer therapies. However, the knowledge regarding the therapeutic potential of GS in PC is still limited and needs to be explored. We studied the effect of GS on PC cell growth, motility and invasion and elucidated the molecular mechanisms associated with its anti-tumor effects. Treatment of Capan1 and CD18/HPAF PC cells with GS resulted in dose- and time-dependent growth inhibition and decreased colony formation. Further, GS treatment induced apoptosis and cell cycle arrest as assessed by Annexin-V assay and FACS analysis. Increased apoptosis following GS treatment was accompanied with Bad dephosphorylation and its translocation to the mitochondria, increased Caspase-3 activation, decreased Cyclin D1, Bcl-2 and xIAP expression. Additionally, GS treatment decreased motility and invasion of PC cells by disrupting cytoskeletal organization, inhibiting activation of FAK and Src signaling and decreased MMP9 expression. More importantly, GS treatment decreased mucin MUC4 expression in Capan1 and CD18/HPAF cells through transcriptional regulation by inhibiting Jak/STAT pathway. In conclusion, our results support the utility of GS as a potential therapeutic agent for lethal PC. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

Guggulsterone decreases proliferation and metastatic behavior of pancreatic cancer cells by modulating JAK/STAT and Src/FAK signaling

PubMed Central

Macha, Muzafar A.; Rachagani, Satyanarayana; Gupta, Suprit; Pai, Priya; Ponnusamy, Moorthy P.; Batra, Surinder K.; Jain, Maneesh

2013-01-01

Inadequate efficacy, high toxicity and drug resistance associated with existing chemotherapeutic agents mandate a need for novel therapeutic strategies for highly aggressive pancreatic cancer (PC). Guggulsterone (GS) exhibits potent anti-proliferative effects against various cancer cells and has emerged as an attractive candidate for use in complementary or preventive cancer therapies. However, the knowledge regarding the therapeutic potential of GS in PC is still limited and needs to be explored. We studied the effect of GS on PC cell growth, motility and invasion and elucidated the molecular mechanisms associated with its anti-tumor effects. Treatment of Capan1 and CD18/HPAF PC cells with GS resulted in dose- and time-dependent growth inhibition and decreased colony formation. Further, GS treatment induced apoptosis and cell cycle arrest as assessed by Annexin-V assay and FACS analysis. Increased apoptosis following GS treatment was accompanied with Bad dephosphorylation and its translocation to the mitochondria, increased Caspase-3 activation, decreased Cyclin D1, Bcl-2 and xIAP expression. Additionally, GS treatment decreased motility and invasion of PC cells by disrupting cytoskeletal organization, inhibiting activation of FAK and Src signaling and decreased MMP9 expression. More importantly, GS treatment decreased mucin MUC4 expression in Capan1 and CD18/HPAF cells through transcriptional regulation by inhibiting Jak/STAT pathway. In conclusion, our results support the utility of GS as a potential therapeutic agent for lethal PC. PMID:23920124

Genomic selection for quantitative adult plant stem rust resistance in wheat

USDA-ARS?s Scientific Manuscript database

Quantitative adult plant resistance (APR) to stem rust (Puccinia graminis f. sp. tritici) is an important breeding target in wheat (Triticum aestivum L.) and a potential target for genomic selection (GS). To evaluate the relative importance of known APR loci in applying genomic selection, we charact...

Toll-Like Receptor 7 Agonist GS-9620 Induces HIV Expression and HIV-Specific Immunity in Cells from HIV-Infected Individuals on Suppressive Antiretroviral Therapy.

PubMed

Tsai, Angela; Irrinki, Alivelu; Kaur, Jasmine; Cihlar, Tomas; Kukolj, George; Sloan, Derek D; Murry, Jeffrey P

2017-04-15

Antiretroviral therapy can suppress HIV replication to undetectable levels but does not eliminate latent HIV, thus necessitating lifelong therapy. Recent efforts to target this persistent reservoir have focused on inducing the expression of latent HIV so that infected cells may be recognized and eliminated by the immune system. Toll-like receptor (TLR) activation stimulates antiviral immunity and has been shown to induce HIV from latently infected cells. Activation of TLR7 leads to the production of several stimulatory cytokines, including type I interferons (IFNs). In this study, we show that the selective TLR7 agonist GS-9620 induced HIV in peripheral blood mononuclear cells (PBMCs) from HIV-infected individuals on suppressive antiretroviral therapy. GS-9620 increased extracellular HIV RNA 1.5- to 2-fold through a mechanism that required type I IFN signaling. GS-9620 also activated HIV-specific T cells and enhanced antibody-mediated clearance of HIV-infected cells. Activation by GS-9620 in combination with HIV peptide stimulation increased CD8 T cell degranulation, production of intracellular cytokines, and cytolytic activity. T cell activation was again dependent on type I IFNs produced by plasmacytoid dendritic cells. GS-9620 induced phagocytic cell maturation and improved effector-mediated killing of HIV-infected CD4 T cells by the HIV envelope-specific broadly neutralizing antibody PGT121. Collectively, these data show that GS-9620 can activate HIV production and improve the effector functions that target latently infected cells. GS-9620 may effectively complement orthogonal therapies designed to stimulate antiviral immunity, such as therapeutic vaccines or broadly neutralizing antibodies. Clinical studies are under way to determine if GS-9620 can target HIV reservoirs. IMPORTANCE Though antiretroviral therapies effectively suppress viral replication, they do not eliminate integrated proviral DNA. This stable intermediate of viral infection is persistently maintained in reservoirs of latently infected cells. Consequently, lifelong therapy is required to maintain viral suppression. Ultimately, new therapies that specifically target and eliminate the latent HIV reservoir are needed. Toll-like receptor agonists are potent enhancers of innate antiviral immunity that can also improve the adaptive immune response. Here, we show that a highly selective TLR7 agonist, GS-9620, activated HIV from peripheral blood mononuclear cells isolated from HIV-infected individuals with suppressed infection. GS-9620 also improved immune effector functions that specifically targeted HIV-infected cells. Previously published studies on the compound in other chronic viral infections show that it can effectively induce immune activation at safe and tolerable clinical doses. Together, the results of these studies suggest that GS-9620 may be useful for treating HIV-infected individuals on suppressive antiretroviral therapy. Copyright © 2017 Tsai et al.

Toll-Like Receptor 7 Agonist GS-9620 Induces HIV Expression and HIV-Specific Immunity in Cells from HIV-Infected Individuals on Suppressive Antiretroviral Therapy

PubMed Central

Tsai, Angela; Irrinki, Alivelu; Kaur, Jasmine; Cihlar, Tomas; Kukolj, George

2017-01-01

ABSTRACT Antiretroviral therapy can suppress HIV replication to undetectable levels but does not eliminate latent HIV, thus necessitating lifelong therapy. Recent efforts to target this persistent reservoir have focused on inducing the expression of latent HIV so that infected cells may be recognized and eliminated by the immune system. Toll-like receptor (TLR) activation stimulates antiviral immunity and has been shown to induce HIV from latently infected cells. Activation of TLR7 leads to the production of several stimulatory cytokines, including type I interferons (IFNs). In this study, we show that the selective TLR7 agonist GS-9620 induced HIV in peripheral blood mononuclear cells (PBMCs) from HIV-infected individuals on suppressive antiretroviral therapy. GS-9620 increased extracellular HIV RNA 1.5- to 2-fold through a mechanism that required type I IFN signaling. GS-9620 also activated HIV-specific T cells and enhanced antibody-mediated clearance of HIV-infected cells. Activation by GS-9620 in combination with HIV peptide stimulation increased CD8 T cell degranulation, production of intracellular cytokines, and cytolytic activity. T cell activation was again dependent on type I IFNs produced by plasmacytoid dendritic cells. GS-9620 induced phagocytic cell maturation and improved effector-mediated killing of HIV-infected CD4 T cells by the HIV envelope-specific broadly neutralizing antibody PGT121. Collectively, these data show that GS-9620 can activate HIV production and improve the effector functions that target latently infected cells. GS-9620 may effectively complement orthogonal therapies designed to stimulate antiviral immunity, such as therapeutic vaccines or broadly neutralizing antibodies. Clinical studies are under way to determine if GS-9620 can target HIV reservoirs. IMPORTANCE Though antiretroviral therapies effectively suppress viral replication, they do not eliminate integrated proviral DNA. This stable intermediate of viral infection is persistently maintained in reservoirs of latently infected cells. Consequently, lifelong therapy is required to maintain viral suppression. Ultimately, new therapies that specifically target and eliminate the latent HIV reservoir are needed. Toll-like receptor agonists are potent enhancers of innate antiviral immunity that can also improve the adaptive immune response. Here, we show that a highly selective TLR7 agonist, GS-9620, activated HIV from peripheral blood mononuclear cells isolated from HIV-infected individuals with suppressed infection. GS-9620 also improved immune effector functions that specifically targeted HIV-infected cells. Previously published studies on the compound in other chronic viral infections show that it can effectively induce immune activation at safe and tolerable clinical doses. Together, the results of these studies suggest that GS-9620 may be useful for treating HIV-infected individuals on suppressive antiretroviral therapy. PMID:28179531

Inhibition of nitrogen-fixing activity of the cyanobiont affects the localization of glutamine synthetase in hair cells of Azolla.

PubMed

Uheda, Eiji; Maejima, Kazuhiro

2009-10-15

In the Azolla-Anabaena association, the host plant Azolla efficiently incorporates and assimilates ammonium ions that are released from the nitrogen-fixing cyanobiont, probably via glutamine synthetase (GS; EC 6.3.1.2) in hair cells, which are specialized cells protruding into the leaf cavity. In order to clarify the regulatory mechanism underlying ammonium assimilation in the Azolla-Anabaena association, Azolla plants were grown under an argon environment (Ar), in which the nitrogen-fixing activity of the cyanobiont was inhibited specifically and completely. The localization of GS in hair cells was determined by immunoelectron microscopy and quantitative analysis of immunogold labeling. Azolla plants grew healthily under Ar when nitrogen sources, such as NO(3)(-) and NH(4)(+), were provided in the growth medium. Both the number of cyanobacterial cells per leaf and the heterocyst frequency of the plants under Ar were similar to those of plants in a nitrogen environment (N(2)). In hair cells of plants grown under Ar, regardless of the type of nitrogen source provided, only weak labeling of GS was observed in the cytoplasm and in chloroplasts. In contrast, in hair cells of plants grown under N(2), abundant labeling of GS was observed in both sites. These findings indicate that specific inhibition of the nitrogen-fixing activity of the cyanobiont affects the localization of GS isoenzymes. Ammonium fixed and released by the cyanobiont could stimulate GS synthesis in hair cells. Simultaneously, the abundant GS, probably GS1, in these cells, could assimilate ammonium rapidly.

Glial architecture of the ghost shark (Callorhinchus milii, Holocephali, Chondrichthyes) as revealed by different immunohistochemical markers.

PubMed

Ari, Csilla; Kálmán, Mihály

2008-09-15

This article presents the first study on the glial architecture of a representative species of Holocephali, Callorhinchus milii (ghost shark). Holocephali are a small subclass of Chondrichthyes, with only a few extant genera, and those are considered to have a brain organization more similar to squalomorph sharks than to galeomorph sharks, skates, and rays. Three different astroglial markers--glial fibrillary acidic protein, S-100 protein, and glutamine synthetase (GS)--were investigated by immunohistochemical methods, applying both diaminobenzidine (DAB) and fluorescent techniques. They revealed similar glial structures, although most of them were detected by immunohistochemical reaction against GS and visualized by DAB. The predominant elements were radial ependymoglia spanning the area between the ventricular and meningeal surfaces, as in squalomorph sharks. Other similar features were the light appearance of myelinated neural tracts devoid of immunoreactivity, and the glial architecture of the reticular formation of the brain stem, cerebellum, and tectum, the latter with recognizable layers. The immunoreactivity of the vascular walls was similar; however, it is believed that different cell types form the blood-brain barrier in chimeras and in elasmobranchs. Some glial structures, however, resembled those of skates, rays, and galeomorph sharks. In C. milii astrocyte-like elements were observed in the telencephalon, using GS and S-100, although typical astrocyte-rich regions were not found. In some areas, especially the telencephalon, not only endfeet but also cell bodies were observed to be attached to the meningeal surface, with processes extending into the brain substance.

Methionine sulfoximine supplementation enhances productivity in GS-CHOK1SV cell lines through glutathione biosynthesis.

PubMed

Feary, Marc; Racher, Andrew J; Young, Robert J; Smales, C Mark

2017-01-01

In Lonza Biologics' GS Gene Expression System™, recombinant protein-producing GS-CHOK1SV cell lines are generated by transfection with an expression vector encoding both GS and the protein product genes followed by selection in MSX and glutamine-free medium. MSX is required to inhibit endogenous CHOK1SV GS, and in effect create a glutamine auxotrophy in the host that can be complemented by the expression vector encoded GS in selected cell lines. However, MSX is not a specific inhibitor of GS as it also inhibits the activity of GCL (a key enzyme in the glutathione biosynthesis pathway) to a similar extent. Glutathione species (GSH and GSSG) have been shown to provide both oxidizing and reducing equivalents to ER-resident oxidoreductases, raising the possibility that selection for transfectants with increased GCL expression could result in the isolation of GS-CHOKISV cell lines with improved capacity for recombinant protein production. In this study we have begun to address the relationship between MSX supplementation, the amount of intracellular GCL subunit and mAb production from a panel of GS-CHOK1SV cell lines. We then evaluated the influence of reduced GCL activity on batch culture of an industrially relevant mAb-producing GS-CHOK1SV cell line. To the best of our knowledge, this paper describes for the first time the change in expression of GCL subunits and recombinant mAb production in these cell lines with the degree of MSX supplementation in routine subculture. Our data also shows that partial inhibition of GCL activity in medium containing 75 µM MSX increases mAb productivity, and its more specific inhibitor BSO used at a concentration of 80 µM in medium increases the specific rate of mAb production eight-fold and the concentration in harvest medium by two-fold. These findings support a link between the inhibition of glutathione biosynthesis and recombinant protein production in industrially relevant systems and provide a process-driven method for increasing mAb productivity from GS-CHOK1SV cell lines. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 33:17-25, 2017. © 2016 American Institute of Chemical Engineers.

Astrocytic glutamine synthetase is expressed in the neuronal somatic layers and down-regulated proportionally to neuronal loss in the human epileptic hippocampus.

PubMed

Papageorgiou, Ismini E; Valous, Nektarios A; Lahrmann, Bernd; Janova, Hana; Klaft, Zin-Juan; Koch, Arend; Schneider, Ulf C; Vajkoczy, Peter; Heppner, Frank L; Grabe, Niels; Halama, Niels; Heinemann, Uwe; Kann, Oliver

2018-05-01

Human mesial temporal lobe epilepsy (MTLE) features subregion-specific hippocampal neurodegeneration and reactive astrogliosis, including up-regulation of the glial fibrillary acidic protein (GFAP) and down-regulation of glutamine synthetase (GS). However, the regional astrocytic expression pattern of GFAP and GS upon MTLE-associated neurodegeneration still remains elusive. We assessed GFAP and GS expression in strict correlation with the local neuronal number in cortical and hippocampal surgical specimens from 16 MTLE patients using immunohistochemistry, stereology and high-resolution image analysis for digital pathology and whole-slide imaging. In the cortex, GS-positive (GS+) astrocytes are dominant in all neuronal layers, with a neuron to GS+ cell ratio of 2:1. GFAP-positive (GFAP+) cells are widely spaced, with a GS+ to GFAP+ cell ratio of 3:1-5:1. White matter astrocytes, on the contrary, express mainly GFAP and, to a lesser extent, GS. In the hippocampus, the neuron to GS+ cell ratio is approximately 1:1. Hippocampal degeneration is associated with a reduction of GS+ astrocytes, which is proportional to the degree of neuronal loss and primarily present in the hilus. Up-regulation of GFAP as a classical hallmark of reactive astrogliosis does not follow the GS-pattern and is prominent in the CA1. Reactive alterations were proportional to the neuronal loss in the neuronal somatic layers (stratum pyramidale and hilus), while observed to a lesser extent in the axonal/dendritic layers (stratum radiatum, molecular layer). We conclude that astrocytic GS is expressed in the neuronal somatic layers and, upon neurodegeneration, is down-regulated proportionally to the degree of neuronal loss. © 2018 Wiley Periodicals, Inc.

Removal of either N-glycan site from the envelope receptor binding domain of Moloney and Friend but not AKV mouse ecotropic gammaretroviruses alters receptor usage

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

Knoper, Ryan C.; Ferrarone, John; Yan Yuhe

2009-09-01

Three N-linked glycosylation sites were removed from the envelope glycoproteins of Friend, Moloney, and AKV mouse ecotropic gammaretroviruses: gs1 and gs2, in the receptor binding domain; and gs8, in a region implicated in post-binding cell fusion. Mutants were tested for their ability to infect rodent cells expressing 4 CAT-1 receptor variants. Three mutants (Mo-gs1, Mo-gs2, and Fr-gs1) infect NIH 3T3 and rat XC cells, but are severely restricted in Mus dunni cells and Lec8, a Chinese hamster cell line susceptible to ecotropic virus. This restriction is reproduced in ferret cells expressing M. dunni dCAT-1, but not in cells expressing NIHmore » 3T3 mCAT-1. Virus binding assays, pseudotype assays, and the use of glycosylation inhibitors further suggest that restriction is primarily due to receptor polymorphism and, in M. dunni cells, to glycosylation of cellular proteins. Virus envelope glycan size or type does not affect infectivity. Thus, host range variation due to N-glycan deletion is receptor variant-specific, cell-specific, virus type-specific, and glycan site-specific.« less

Piezo type mechanosensitive ion channel component 1 functions as a regulator of the cell fate determination of mesenchymal stem cells.

PubMed

Sugimoto, Asuna; Miyazaki, Aya; Kawarabayashi, Keita; Shono, Masayuki; Akazawa, Yuki; Hasegawa, Tomokazu; Ueda-Yamaguchi, Kimiko; Kitamura, Takamasa; Yoshizaki, Keigo; Fukumoto, Satoshi; Iwamoto, Tsutomu

2017-12-18

The extracellular environment regulates the dynamic behaviors of cells. However, the effects of hydrostatic pressure (HP) on cell fate determination of mesenchymal stem cells (MSCs) are not clearly understood. Here, we established a cell culture chamber to control HP. Using this system, we found that the promotion of osteogenic differentiation by HP is depend on bone morphogenetic protein 2 (BMP2) expression regulated by Piezo type mechanosensitive ion channel component 1 (PIEZO1) in MSCs. The PIEZO1 was expressed and induced after HP loading in primary MSCs and MSC lines, UE7T-13 and SDP11. HP and Yoda1, an activator of PIEZO1, promoted BMP2 expression and osteoblast differentiation, whereas inhibits adipocyte differentiation. Conversely, PIEZO1 inhibition reduced osteoblast differentiation and BMP2 expression. Furthermore, Blocking of BMP2 function by noggin inhibits HP induced osteogenic maker genes expression. In addition, in an in vivo model of medaka with HP loading, HP promoted caudal fin ray development whereas inhibition of piezo1 using GsMTx4 suppressed its development. Thus, our results suggested that PIEZO1 is responsible for HP and could functions as a factor for cell fate determination of MSCs by regulating BMP2 expression.

Ovarian cancer therapeutic potential of glutamine depletion based on GS expression.

PubMed

Furusawa, Akiko; Miyamoto, Morikazu; Takano, Masashi; Tsuda, Hitoshi; Song, Yong Sang; Aoki, Daisuke; Miyasaka, Naoyuki; Inazawa, Johji; Inoue, Jun

2018-05-28

Amino acids (AAs) are biologically important nutrient compounds necessary for the survival of any cell. Of the 20 AAs, cancer cells depend on the uptake of several extracellular AAs for survival. However, which extracellular AA is indispensable for the survival of cancer cells and the molecular mechanism involved have not been fully defined. In this study, we found that the reduction of cell survival caused by glutamine (Gln) depletion is inversely correlated with the expression level of glutamine synthetase (GS) in ovarian cancer (OVC) cells. GS expression was downregulated in 45 of 316 OVC cases (14.2%). The depletion of extracellular Gln by treatment with l-asparaginase, in addition to inhibiting Gln uptake via the knockdown of a Gln transporter, led to the inhibition of cell growth in OVC cells with low expression of GS (GSlow-OVC cells). Furthermore, the re-expression of GS in GSlow-OVC cells induced the inhibition of tumor growth in vitro and in vivo. Thus, these findings provide novel insight into the development of an OVC therapy based on the requirement of Gln.

Differential expression of multiple glutamine synthetase genes in air-breathing magur catfish, Clarias magur and their induction under hyper-ammonia stress.

PubMed

Banerjee, Bodhisattwa; Koner, Debaprasad; Bhuyan, Gitalee; Saha, Nirmalendu

2018-06-01

The present study demonstrates the unique presence of three different gs genes (cmgs01, cmgs02, and cmgs03) in air-breathing ureogenic magur catfish (Clarias magur), which is otherwise reported to be encoded by a single gene in higher vertebrates. Of these three genes, two (cmgs01and cmgs03) were identified as 'liver' form, predominantly expressed in liver cells, and the third one as 'brain' form (cmgs02), expressed chiefly in brain cells. Molecular characterization studies have revealed conservation of homologous active site residues in all the three gs genes. In silico analysis, accompanied by GS enzyme assay and Western blot analysis of different GS isoforms in different subcellular fractions indicated the mitochondrial localization of cmGS01 and cmGS03 in liver and kidney cells and cytosolic localization of cmGS02 in brain cells. Further, exposure of magur catfish to high external ammonia (HEA; 25 mM NH 4 Cl) led to a significant induction of multiple gs genes as evidenced by higher expression of different gs mRNAs at variable levels in different tissues. The cmgs01 and cmgs03 mRNA levels elevated significantly in liver, kidney, muscle, and gills, whereas the cmgs02 mRNA level increased considerably in the brain after 14 days of exposure to HEA. These increases in mRNA levels were associated with a significant rise in cmGS01 and cmGS03 proteins in liver, kidney, muscle, and gills, and the cmGS02 protein in the brain after 14 days of exposure to HEA. Therefore, it can be concluded that the unique differential expression of three gs genes and their induction under high ammonia level probably helps in detoxification of ammonia to glutamine and further to urea via the ornithine-urea cycle in ureogenic as well as non-ureogenic tissues of these magur catfish. Copyright © 2017. Published by Elsevier B.V.

14-3-3 zeta is a molecular target in guggulsterone induced apoptosis in head and neck cancer cells.

PubMed

Macha, Muzafar A; Matta, Ajay; Chauhan, Ss; Siu, Kw Michael; Ralhan, Ranju

2010-11-30

The five-year survival rates for head and neck squamous cell carcinoma (HNSCC) patients are less than 50%, and the prognosis has not improved, despite advancements in standard multi-modality therapies. Hence major emphasis is being laid on identification of novel molecular targets and development of multi-targeted therapies. 14-3-3 zeta, a multifunctional phospho-serine/phospho-threonine binding protein, is emerging as an effector of pro-survival signaling by binding to several proteins involved in apoptosis (Bad, FKHRL1 and ASK1) and may serve as an appropriate target for head and neck cancer therapy. Herein, we determined effect of guggulsterone (GS), a farnesoid X receptor antagonist, on 14-3-3 zeta associated molecular pathways for abrogation of apoptosis in head and neck cancer cells. Head and neck cancer cells were treated with guggulsterone (GS). Effect of GS-treatment was evaluated using cell viability (MTT) assay and apoptosis was verified by annexin V, DNA fragmentation and M30 CytoDeath antibody assay. Mechanism of GS-induced apoptosis was determined by western blotting and co-IP assays using specific antibodies. Using in vitro models of head and neck cancer, we showed 14-3-3 zeta as a key player regulating apoptosis in GS treated SCC4 cells. Treatment with GS releases BAD from the inhibitory action of 14-3-3 zeta in proliferating HNSCC cells by activating protein phosphatase 2A (PP2A). These events initiate the intrinsic mitochondrial pathway of apoptosis, as revealed by increased levels of cytochrome c in cytoplasmic extracts of GS-treated SCC4 cells. In addition, GS treatment significantly reduced the expression of anti-apoptotic proteins, Bcl-2, xIAP, Mcl1, survivin, cyclin D1 and c-myc, thus committing cells to apoptosis. These events were followed by activation of caspase 9, caspase 8 and caspase 3 leading to cleavage of its downstream target, poly-ADP-ribose phosphate (PARP). GS targets 14-3-3 zeta associated cellular pathways for reducing proliferation and inducing apoptosis in head and neck cancer cells, warranting its investigation for use in treatment of head and neck cancer.

Anti-HBV response to toll-like receptor 7 agonist GS-9620 is associated with intrahepatic aggregates of T cells and B cells.

PubMed

Li, Li; Barry, Vivian; Daffis, Stephane; Niu, Congrong; Huntzicker, Erik; French, Dorothy M; Mikaelian, Igor; Lanford, Robert E; Delaney, William E; Fletcher, Simon P

2018-05-01

GS-9620, an oral agonist of toll-like receptor 7, is in clinical development for the treatment of chronic hepatitis B (CHB). GS-9620 was previously shown to induce prolonged suppression of serum viral DNA and antigens in the chimpanzee and woodchuck models of CHB. Herein, we investigated the immunomodulatory mechanisms underlying these antiviral effects. Archived liver biopsies and paired peripheral blood mononuclear cell samples from a previous chimpanzee study were analyzed by RNA sequencing, quantitative reverse transcription PCR, immunohistochemistry (IHC) and in situ hybridization (ISH). GS-9620 treatment of CHB chimpanzees induced an intrahepatic transcriptional profile significantly enriched with genes associated with hepatitis B virus (HBV) clearance in acutely infected chimpanzees. Type I and II interferon, CD8 + T cell and B cell transcriptional signatures were associated with treatment response, together with evidence of hepatocyte death and liver regeneration. IHC and ISH confirmed an increase in intrahepatic CD8 + T cell and B cell numbers during treatment, and revealed that GS-9620 transiently induced aggregates predominantly comprised of CD8 + T cells and B cells in portal regions. There were no follicular dendritic cells or IgG-positive cells in these lymphoid aggregates and very few CD11b + myeloid cells. There was no change in intrahepatic natural killer cell number during GS-9620 treatment. The antiviral response to GS-9620 treatment in CHB chimpanzees was associated with an intrahepatic interferon response and formation of lymphoid aggregates in the liver. Our data indicate these intrahepatic structures are not fully differentiated follicles containing germinal center reactions. However, the temporal correlation between development of these T and B cell aggregates and the antiviral response to treatment suggests they play a role in promoting an effective immune response against HBV. New therapies to treat chronic hepatitis B (CHB) are urgently needed. In this study we performed a retrospective analysis of liver and blood samples from a chimpanzee model of CHB to help understand how GS-9620, a drug in clinical trials, suppressed hepatitis B virus (HBV). We found that the antiviral response to GS-9620 was associated with accumulation of immune cells in the liver that can either kill cells infected with HBV or can produce antibodies that may prevent HBV from infecting new liver cells. These findings have important implications for how GS-9620 may be used in patients and may also help guide the development of new therapies to treat chronic HBV infection. Copyright © 2017 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

Manipulation of the apoplastic pH of intact plants mimics stomatal and growth responses to water availability and microclimatic variation.

PubMed

Wilkinson, Sally; Davies, William J

2008-01-01

The apoplastic pH of intact Forsythiaxintermedia (cv. Lynwood) and tomato (Solanum lycopersicum) plants has been manipulated using buffered foliar sprays, and thereby stomatal conductance (g(s)), leaf growth rate, and plant water loss have been controlled. The more alkaline the pH of the foliar spray, the lower the g(s) and/or leaf growth rate subsequently measured. The most alkaline pH that was applied corresponds to that measured in sap extracted from shoots of tomato and Forsythia plants experiencing, respectively, soil drying or a relatively high photon flux density (PFD), vapour pressure deficit (VPD), and temperature in the leaf microclimate. The negative correlation between PFD/VPD/temperature and g(s) determined in well-watered Forsythia plants exposed to a naturally varying summer microclimate was eliminated by spraying the plants with relatively alkaline but not acidic buffers, providing evidence for a novel pH-based signalling mechanism linking the aerial microclimate with stomatal aperture. Increasing the pH of the foliar spray only reduced g(s) in plants of the abscisic acid (ABA)-deficient flacca mutant of tomato when ABA was simultaneously sprayed onto leaves or injected into stems. In well-watered Forsythia plants exposed to a naturally varying summer microclimate (variable PFD, VPD, and temperature), xylem pH and leaf ABA concentration fluctuated but were positively correlated. Manipulation of foliar apoplastic pH also affected the response of g(s) and leaf growth to ABA injected into stems of intact Forsythia plants. The techniques used here to control physiology and water use in intact growing plants could easily be applied in a horticultural context.

Guggulsterone (GS) inhibits smokeless tobacco and nicotine-induced NF-κB and STAT3 pathways in head and neck cancer cells.

PubMed

Macha, Muzafar A; Matta, Ajay; Chauhan, S S; Siu, K W Michael; Ralhan, Ranju

2011-03-01

Understanding the molecular pathways perturbed in smokeless tobacco- (ST) associated head and neck squamous cell carcinoma (HNSCC) is critical for identifying novel complementary agents for effective disease management. Activation of nuclear factor-kappaB (NF-κB) and cyclooxygenase-2 (COX-2) was reported in ST-associated HNSCC by us [Sawhney,M. et al. (2007) Expression of NF-kappaB parallels COX-2 expression in oral precancer and cancer: association with smokeless tobacco. Int. J. Cancer, 120, 2545-2556]. In search of novel agents for treatment of HNSCC, we investigated the potential of guggulsterone (GS), (4,17(20)-pregnadiene-3,16-dione), a biosafe nutraceutical, in inhibiting ST- and nicotine-induced activation of NF-κB and signal transducer and activator of transcription (STAT) 3 pathways in HNSCC cells. GS inhibited the activation of NF-κB and STAT3 proteins in head and neck cancer cells. This inhibition of NF-κB by GS resulted from decreased phosphorylation and degradation of nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor, alpha the inhibitory subunit of NF-κB. Importantly, treatment of HNSCC cells with GS abrogated both ST- and nicotine-induced nuclear activation of NF-κB and pSTAT3 proteins and their downstream targets COX-2 and vascular endothelial growth factor. Furthermore, GS treatment decreased the levels of ST- and nicotine-induced secreted interleukin-6 in culture media of HNSCC cells. In conclusion, our findings demonstrated that GS treatment abrogates the effects of ST and nicotine on activation of NF-κB and STAT3 pathways in HNSCC cells that contribute to inflammatory and angiogenic responses as well as its progression and metastasis. These findings provide a biologic rationale for further clinical investigation of GS as an effective complementary agent for inhibiting ST-induced head and neck cancer.

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

Varma, Shailly; Shrivastav, Anuraag; Health Research Division, Saskatchewan Cancer Agency, Saskatoon, SK S7N 4H4

Protein kinase B (Akt/PKB) is a Ser/Thr kinase that is involved in the regulation of cell proliferation/survival through mammalian target of rapamycin (mTOR) and the regulation of glycogen metabolism through glycogen synthase kinase 3{beta} (GSK-3{beta}) and glycogen synthase (GS). Rapamycin is an inhibitor of mTOR. The objective of this study was to investigate the effects of rapamycin pretreatment on the insulin mediated phosphorylation of Akt/PKB phosphorylation and GS activity in parental HepG2 and HepG2 cells with overexpression of constitutively active Akt1/PKB-{alpha} (HepG2-CA-Akt/PKB). Rapamycin pretreatment resulted in a decrease (20-30%) in the insulin mediated phosphorylation of Akt1 (Ser 473) in parentalmore » HepG2 cells but showed an upregulation of phosphorylation in HepG2-CA-Akt/PKB cells. Rictor levels were decreased (20-50%) in parental HepG2 cells but were not significantly altered in the HepG2-CA-Akt/PKB cells. Furthermore, rictor knockdown decreased the phosphorylation of Akt (Ser 473) by 40-60% upon rapamycin pretreatment. GS activity followed similar trends as that of phosphorylated Akt and so with rictor levels in these cells pretreated with rapamycin; parental HepG2 cells showed a decrease in GS activity, whereas as HepG2-CA-Akt/PKB cells showed an increase in GS activity. The changes in the levels of phosphorylated Akt/PKB (Ser 473) correlated with GS and protein phoshatase-1 activity.« less

Lignin metabolism involves Botrytis cinerea BcGs1- induced defense response in tomato.

PubMed

Yang, Chenyu; Liang, Yingbo; Qiu, Dewen; Zeng, Hongmei; Yuan, Jingjing; Yang, Xiufen

2018-06-04

BcGs1, a cell wall-degrading enzyme (CWDE), was originally derived from Botrytis cinerea. Our previous study revealed that BcGs1 could trigger defense responses and protect plants against various pathogens. We researched the defense response mechanism underlying this BcGs1 elicitation in tomato. We revealed that the two domains were required for BcGs1's full necrosis activity. According to analysis and quantitative real-time PCR of the up-regulated proteins and genes filtered by iTRAQ-based quantitative proteome approach, oxidative metabolism and phenylpropanoid metabolism were speculated to be involved in BcGs1-triggered defense response in tomato. Furthermore, experimental evidence showed that BcGs1 triggered reactive oxygen species (ROS) burst and increased the level of phenylalanine-ammonia lyase (PAL) and peroxidase (POD) enzyme activity, as well as lignin accumulation. Moreover, histochemical analysis revealed that infiltration of BcGs1 in tomato leaves exhibited cell wall thickening compared with untreated plants. The results suggested that BcGs1 activated the basal defense response included lignin metabolism contributed to BcGs1-induced resistance to Botrytis. cinerea infection in tomato.

Secondary NAD+ deficiency in the inherited defect of glutamine synthetase.

PubMed

Hu, Liyan; Ibrahim, Khalid; Stucki, Martin; Frapolli, Michele; Shahbeck, Noora; Chaudhry, Farrukh A; Görg, Boris; Häussinger, Dieter; Penberthy, W Todd; Ben-Omran, Tawfeg; Häberle, Johannes

2015-11-01

Glutamine synthetase (GS) deficiency is an ultra-rare inborn error of amino acid metabolism that has been described in only three patients so far. The disease is characterized by neonatal onset of severe encephalopathy, low levels of glutamine in blood and cerebrospinal fluid, chronic moderate hyperammonemia, and an overall poor prognosis in the absence of an effective treatment. Recently, enteral glutamine supplementation was shown to be a safe and effective therapy for this disease but there are no data available on the long-term effects of this intervention. The amino acid glutamine, severely lacking in this disorder, is central to many metabolic pathways in the human organism and is involved in the synthesis of nicotinamide adenine dinucleotide (NAD(+)) starting from tryptophan or niacin as nicotinate, but not nicotinamide. Using fibroblasts, leukocytes, and immortalized peripheral blood stem cells (PBSC) from a patient carrying a GLUL gene point mutation associated with impaired GS activity, we tested whether glutamine deficiency in this patient results in NAD(+) depletion and whether it can be rescued by supplementation with glutamine, nicotinamide or nicotinate. The present study shows that congenital GS deficiency is associated with NAD(+) depletion in fibroblasts, leukocytes and PBSC, which may contribute to the severe clinical phenotype of the disease. Furthermore, it shows that NAD(+) depletion can be rescued by nicotinamide supplementation in fibroblasts and leukocytes, which may open up potential therapeutic options for the treatment of this disorder.

Apical localization of glutamate in GLAST-1, glutamine synthetase positive ciliary body nonpigmented epithelial cells

PubMed Central

Langford, Marlyn P; Gosslee, Jeffrey M; Liang, Chanping; Chen, Dequan; Redens, Thomas B.; Welbourne, Tomas C

2007-01-01

The distribution of glutamate (Glu), the Glu transporter GLAST-1, and glutamine synthetase (GS) in human and monkey anterior uveal tissue, as well as serum (S) to aqueous humor (AH) Glu and glutamine (Gln) gradients were investigated. Cross-linked Glu (xGlu), GLAST-1, and GS were detected using the immunofluorescent antibody technique. S/AH Glu, Gln, and alanine (Ala) concentrations were quantified by high performance liquid chromatography. xGlu immunoreactivity was detected in melanocytes, posterior pigmented epithelial/dilator muscle cells, vascular endothelial cells, and lymphocytes of the iris, as well as the pigmented (PE) and nonpigmented epithelial (NPE) cells and muscle cells of ciliary body. xGlu immunoreactivity was highly concentrated at the apices of GLAST-1, GS positive ciliary body NPE cells, and in GLAST-1 positive iris melanocytes and iris dilator muscle cells. AH Glu concentrations were lower (p < 0.001), while Gln was higher in monkey (p = 0.01) and human cataractous (p = 0.15) AH than serum. The results indicate that Glu is concentrated within GLAST-1, GS positive NPE cells and are consistent with the suggestion that Glu and Gln concentrations in AH may be due in part to GLAST-1 and GS activity in iris and ciliary body epithelial cells. PMID:19668465

Effect of emodin on mobility signal transduction system of gallbladder smooth muscle in Guinea pig with cholelithiasis.

PubMed

Fang, Bang-Jiang; Shen, Jun-Yi; Zhang, Hua; Zhou, Shuang; Lyu, Chuan-Zhu; Xie, Yi-Qiang

2016-10-01

To study the effect of emodin on protein and gene expressions of the massagers in mobility signal transduction system of cholecyst smooth muscle cells in guinea pig with cholesterol calculus. The guinea pigs were randomly divided into 4 groups, such as control group, gall-stone (GS) group, emodin group and ursodeoxycholic acid (UA) group. Cholesterol calculus models were induced in guinea pigs of GS, emodin and UA groups by lithogenic diet, while emodin or UA were given to the corresponding group for 7 weeks. The histomorphological and ultrastructure change of gallbladder were detected by microscope and electron microscope, the content of plasma cholecystokinin (CCK) and [Ca 2+ ] i were analyzed successively by radioimmunoassay and flow cytometry. The protein and mRNA of Gsα, Giα and Cap in cholecyst cells were determined by western blotting and real time polymerase chain reaction (RT-PCR). Emodin or UA can relieve pathogenic changes in epithelial cells and muscle cells in gallbladder of guinea pig with cholesterol calculus by microscope and transmission electron microscope. In the cholecyst cells of GS group, CCK levels in plasma and [Ca 2+ ] i decreased, the protein and mRNA of GS were down-regulated, the protein and mRNA of Gi and Cap were up-regulated. Emodin significantly decreased the formative rate of gallstone, improved the pathogenic change in epithelial cells and muscle cells, increased CCK levels in plasma and [Ca 2+ ] i in cholecyst cells, enhanced the protein and mRNA of Gs in cholecyst cells, reduced the protein and mRNA of Gi and Cap in cholecyst cells in guinea pig with cholesterol calculus. The dysfunction of gallbladder contraction gives rise to the disorders of mobility signal transduction system in cholecyst smooth muscle cells, including low content of plasma CCK and [Ca 2+ ] i in cholecyst cells, abnormal protein and mRNA of Gs, Gi and Cap. Emodin can enhance the contractibility of gallbladder and alleviate cholestasis by regulating plasma CCK levels, [Ca 2+ ] i in cholecyst cells and the protein and mRNA of Gs, Gi and Cap. Copyright © 2016 Hainan Medical University. Production and hosting by Elsevier B.V. All rights reserved.

The Stable Level of Glutamine synthetase 2 Plays an Important Role in Rice Growth and in Carbon-Nitrogen Metabolic Balance

PubMed Central

Bao, Aili; Zhao, Zhuqing; Ding, Guangda; Shi, Lei; Xu, Fangsen; Cai, Hongmei

2015-01-01

Glutamine synthetase 2 (GS2) is a key enzyme involved in the ammonium metabolism in plant leaves. In our previous study, we obtained GS2-cosuppressed plants, which displayed a normal growth phenotype at the seedling stage, while at the tillering stage they showed a chlorosis phenotype. In this study, to investigate the chlorosis mechanism, we systematically analyzed the plant growth, carbon-nitrogen metabolism and gene expressions between the GS2-cosuppressed rice and wild-type plants. The results revealed that the GS2-cosuppressed plants exhibited a poor plant growth phenotype and a poor nitrogen transport ability, which led to nitrogen accumulation and a decline in the carbon/nitrogen ratio in the stems. Interestingly, there was a higher concentration of soluble proteins and a lower concentration of carbohydrates in the GS2-cosuppressed plants at the seedling stage, while a contrasting result was displayed at the tillering stage. The analysis of the metabolic profile showed a significant increase of sugars and organic acids. Additionally, gene expression patterns were different in root and leaf of GS2-cosuppressed plants between the seedling and tillering stage. These results indicated the important role of a stable level of GS2 transcription during normal rice development and the importance of the carbon-nitrogen metabolic balance in rice growth. PMID:26053400

The Stable Level of Glutamine synthetase 2 Plays an Important Role in Rice Growth and in Carbon-Nitrogen Metabolic Balance.

PubMed

Bao, Aili; Zhao, Zhuqing; Ding, Guangda; Shi, Lei; Xu, Fangsen; Cai, Hongmei

2015-06-04

Glutamine synthetase 2 (GS2) is a key enzyme involved in the ammonium metabolism in plant leaves. In our previous study, we obtained GS2-cosuppressed plants, which displayed a normal growth phenotype at the seedling stage, while at the tillering stage they showed a chlorosis phenotype. In this study, to investigate the chlorosis mechanism, we systematically analyzed the plant growth, carbon-nitrogen metabolism and gene expressions between the GS2-cosuppressed rice and wild-type plants. The results revealed that the GS2-cosuppressed plants exhibited a poor plant growth phenotype and a poor nitrogen transport ability, which led to nitrogen accumulation and a decline in the carbon/nitrogen ratio in the stems. Interestingly, there was a higher concentration of soluble proteins and a lower concentration of carbohydrates in the GS2-cosuppressed plants at the seedling stage, while a contrasting result was displayed at the tillering stage. The analysis of the metabolic profile showed a significant increase of sugars and organic acids. Additionally, gene expression patterns were different in root and leaf of GS2-cosuppressed plants between the seedling and tillering stage. These results indicated the important role of a stable level of GS2 transcription during normal rice development and the importance of the carbon-nitrogen metabolic balance in rice growth.

Comprehensive characterization of glutamine synthetase-mediated selection for the establishment of recombinant CHO cells producing monoclonal antibodies.

PubMed

Noh, Soo Min; Shin, Seunghyeon; Lee, Gyun Min

2018-03-29

To characterize a glutamine synthetase (GS)-based selection system, monoclonal antibody (mAb) producing recombinant CHO cell clones were generated by a single round of selection at various methionine sulfoximine (MSX) concentrations (0, 25, and 50 μM) using two different host cell lines (CHO-K1 and GS-knockout CHO). Regardless of the host cell lines used, the clones selected at 50 μM MSX had the lowest average specific growth rate and the highest average specific production rates of toxic metabolic wastes, lactate and ammonia. Unlike CHO-K1, high producing clones could be generated in the absence of MSX using GS-knockout CHO with an improved selection stringency. Regardless of the host cell lines used, the clones selected at various MSX concentrations showed no significant difference in the GS, heavy chain, and light chain gene copies (P > 0.05). Furthermore, there was no correlation between the specific mAb productivity and these three gene copies (R 2  ≤ 0.012). Taken together, GS-mediated gene amplification does not occur in a single round of selection at a MSX concentration up to 50 μM. The use of the GS-knockout CHO host cell line facilitates the rapid generation of high producing clones with reduced production of lactate and ammonia in the absence of MSX.

[Nevoid basal-cell carcinoma syndrome (Gorlin Syndrome): report of two cases and review of the literature].

PubMed

Castro-Mujica, María Del Carmen; Barletta-Carrillo, Claudia; Poterico, Julio A; Acosta, Marisa; Valer, Jesús; Cruz, Miguel De La

2017-01-01

Gorlin syndrome (GS) is a genetic disorder with an autosomal dominant inheritance pattern, with complete penetrance and variable expressivity. GS is caused by germline mutations in the genes PTCH1 or SUFU, which are components of the Sonic hedgehog molecular pathway. GS is characterized by the presence of multiple nevoid basal cell carcinomas, odontogenic cysts, calcification of the brain sickle, and lesions in the palms and soles. This study is the first to report cases in Peru of patients with GS who underwent genetic evaluation and counseling. We present two GS cases that meet the clinical criteria for the syndrome and review the literature.

Stem girdling evidences a trade-off between cambial activity and sprouting and dramatically reduces plant transpiration due to feedback inhibition of photosynthesis and hormone signaling

PubMed Central

López, Rosana; Brossa, Ricard; Gil, Luis; Pita, Pilar

2015-01-01

The photosynthesis source–sink relationship in young Pinus canariensis seedlings was modified by stem girdling to investigate sprouting and cambial activity, feedback inhibition of photosynthesis, and stem and root hydraulic capacity. Removal of bark tissue showed a trade-off between sprouting and diameter growth. Above the girdle, growth was accelerated but the number of sprouts was almost negligible, whereas below the girdle the response was reversed. Girdling resulted in a sharp decrease in whole plant transpiration and root hydraulic conductance. The reduction of leaf area after girdling was strengthened by the high levels of abscisic acid found in buds which pointed to stronger bud dormancy, preventing a new needle flush. Accumulation of sugars in leaves led to a coordinated reduction in net photosynthesis (AN) and stomatal conductance (gS) in the short term, but later (gS below 0.07 mol m-2 s-1) AN decreased faster. The decrease in maximal efficiency of photosystem II (FV/FM) and the operating quantum efficiency of photosystem II (ΦPSII) in girdled plants could suggest photoprotection of leaves, as shown by the vigorous recovery of AN and ΦPSII after reconnection of the phloem. Stem girdling did not affect xylem embolism but increased stem hydraulic conductance above the girdle. This study shows that stem girdling affects not only the carbon balance, but also the water status of the plant. PMID:25972884

Stem girdling evidences a trade-off between cambial activity and sprouting and dramatically reduces plant transpiration due to feedback inhibition of photosynthesis and hormone signaling.

PubMed

López, Rosana; Brossa, Ricard; Gil, Luis; Pita, Pilar

2015-01-01

The photosynthesis source-sink relationship in young Pinus canariensis seedlings was modified by stem girdling to investigate sprouting and cambial activity, feedback inhibition of photosynthesis, and stem and root hydraulic capacity. Removal of bark tissue showed a trade-off between sprouting and diameter growth. Above the girdle, growth was accelerated but the number of sprouts was almost negligible, whereas below the girdle the response was reversed. Girdling resulted in a sharp decrease in whole plant transpiration and root hydraulic conductance. The reduction of leaf area after girdling was strengthened by the high levels of abscisic acid found in buds which pointed to stronger bud dormancy, preventing a new needle flush. Accumulation of sugars in leaves led to a coordinated reduction in net photosynthesis (AN) and stomatal conductance (gS) in the short term, but later (gS below 0.07 mol m(-2) s(-1)) AN decreased faster. The decrease in maximal efficiency of photosystem II (FV/FM) and the operating quantum efficiency of photosystem II (ΦPSII) in girdled plants could suggest photoprotection of leaves, as shown by the vigorous recovery of AN and ΦPSII after reconnection of the phloem. Stem girdling did not affect xylem embolism but increased stem hydraulic conductance above the girdle. This study shows that stem girdling affects not only the carbon balance, but also the water status of the plant.

TaGS5-3A, a grain size gene selected during wheat improvement for larger kernel and yield.

PubMed

Ma, Lin; Li, Tian; Hao, Chenyang; Wang, Yuquan; Chen, Xinhong; Zhang, Xueyong

2016-05-01

Grain size is a dominant component of grain weight in cereals. Earlier studies have shown that OsGS5 plays a major role in regulating both grain size and weight in rice via promotion of cell division. In this study, we isolated TaGS5 homoeologues in wheat and mapped them on chromosomes 3A, 3B and 3D. Temporal and spatial expression analysis showed that TaGS5 homoeologues were preferentially expressed in young spikes and developing grains. Two alleles of TaGS5-3A, TaGS5-3A-T and TaGS5-3A-G were identified in wheat accessions, and a functional marker was developed to discriminate them. Association analysis revealed that TaGS5-3A-T was significantly correlated with larger grain size and higher thousand kernel weight. Biochemical assays showed that TaGS5-3A-T possesses a higher enzymatic activity than TaGS5-3A-G. Transgenic rice lines overexpressing TaGS5-3A-T also exhibited larger grain size and higher thousand kernel weight than TaGS5-3A-G lines, and the transcript levels of cell cycle-related genes in TaGS5-3A-T lines were higher than those in TaGS5-3A-G lines. Furthermore, systematic evolution analysis in diploid, tetraploid and hexaploid wheat showed that TaGS5-3A underwent strong artificial selection during wheat polyploidization events and the frequency changes of two alleles demonstrated that TaGS5-3A-T was favoured in global modern wheat cultivars. These results suggest that TaGS5-3A is a positive regulator of grain size and its favoured allele TaGS5-3A-T exhibits a larger potential application in wheat high-yield breeding. © 2015 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

Recovery of diurnal depression of leaf hydraulic conductance in a subtropical woody bamboo species: embolism refilling by nocturnal root pressure.

PubMed

Yang, Shi-Jian; Zhang, Yong-Jiang; Sun, Mei; Goldstein, Guillermo; Cao, Kun-Fang

2012-04-01

Despite considerable investigations of diurnal water use characteristics in different plant functional groups, the research on daily water use strategies of woody bamboo grasses remains lacking. We studied the daily water use and gas exchange of Sinarundinaria nitida (Mitford) Nakai, an abundant subtropical bamboo species in Southwest China. We found that the stem relative water content (RWC) and stem hydraulic conductivity (K(s)) of this bamboo species did not decrease significantly during the day, whereas the leaf RWC and leaf hydraulic conductance (K(leaf)) showed a distinct decrease at midday, compared with the predawn values. Diurnal loss of K(leaf) was coupled with a midday decline in stomatal conductance (g(s)) and CO(2) assimilation. The positive root pressures in the different habitats were of sufficient magnitude to refill the embolisms in leaves. We concluded that (i) the studied bamboo species does not use stem water storage for daily transpiration; (ii) diurnal down-regulation in K(leaf) and gs has the function to slow down potential water loss in stems and protect the stem hydraulic pathway from cavitation; (iii) since K(leaf) did not recover during late afternoon, refilling of embolism in bamboo leaves probably fully depends on nocturnal root pressure. The embolism refilling mechanism by root pressure could be helpful for the growth and persistence of this woody monocot species.

Poly-L-glutamate/glutamine synthesis in the cell wall of Mycobacterium bovis is regulated in response to nitrogen availability

PubMed Central

2013-01-01

Background The cell wall of pathogenic mycobacteria is known to possess poly-L-glutamine (PLG) layer. PLG synthesis has been directly linked to glutamine synthetase (GS) enzyme. glnA1 gene encodes for GS enzyme in mycobacteria. PLG layer is absent in cell wall of avirulent Mycobacterium smegmatis, although M. smegmatis strain expressing GS enzyme of pathogenic mycobacteria can synthesize PLG layer in the cell wall. The role of GS enzyme has been extensively studied in Mycobacterium tuberculosis, however, little is known about GS enzyme in other mycobacterial species. Mycobacterium bovis, as an intracellular pathogen encounters nitrogen stress inside macrophages, thus it has developed nitrogen assimilatory pathways to survive in adverse conditions. We have investigated the expression and activity of M. bovis GS in response to nitrogen availability and effect on synthesis of PLG layer in the cell wall. M. smegmatis was used as a model to study the behaviour of glnA1 locus of M. bovis. Results We observed that GS expression and activity decreased significantly in high nitrogen grown conditions. In high nitrogen conditions, the amount of PLG in cell wall was drastically reduced (below detectable limits) as compared to low nitrogen condition in M. bovis and in M. smegmatis strain complemented with M. bovis glnA1. Additionally, biofilm formation by M. smegmatis strain complemented with M. bovis glnA1 was increased than the wild type M. smegmatis strain. Conclusions The physiological regulation of GS in M. bovis was found to be similar to that reported in other mycobacteria but this data revealed that PLG synthesis in the cell wall of pathogenic mycobacteria occurs only in nitrogen limiting conditions and on the contrary high nitrogen conditions inhibit PLG synthesis. This indicates that PLG synthesis may be a form of nitrogen assimilatory pathway during ammonium starvation in virulent mycobacteria. Also, we have found that M. smegmatis complemented with M. bovis glnA1 was more efficient in biofilm formation than the wild type strain. This indicates that PLG layer favors biofilm formation. This study demonstrate that the nitrogen availability not only regulates GS expression and activity in M. bovis but also affects cell surface properties by modulating synthesis of PLG. PMID:24112767

Antimicrobial Properties and Cytotoxicity of Sulfated (1,3)-β-D-Glucan from the Mycelium of the Mushroom Ganoderma lucidum.

PubMed

Wan-Mohtar, Wan Abd Al Qadr Imad; Young, Louise; Abbott, Gráinne M; Clements, Carol; Harvey, Linda M; McNeil, Brian

2016-06-28

Ganoderma lucidum BCCM 31549 has a long established role for its therapeutic activities. In this context, much interest has focused on the possible functions of the (1,3)-β-D-glucan (G) produced by these cultures in a stirred-tank bioreactor and extracted from their underutilized mycelium. In the existing study, we report on the systematic production of G, and its sulfated derivative (GS). The aim of this study was to investigate G and its GS from G. lucidum in terms of their antibacterial properties and cytotoxicity spectrum against human prostate cells (PN2TA) and human caucasian histiocytic lymphoma cells (U937). (1)H NMR for both G and GS compounds showed β-glycosidic linkages and structural similarities when compared with two standards (laminarin and fucoidan). The existence of characteristic absorptions at 1,170 and 867 cm(-1) in the FTIR (Fourier Transform Infrared Spectroscopy) for GS demonstrated the successful sulfation of G. Only GS exhibited antimicrobial activity against a varied range of test bacteria of relevance to foodstuffs and human health. Moreover, both G and GS did not show any cytotoxic effects on PN2TA cells, thus helping demonstrate the safety of these polymers. Moreover, GS showed 40% antiproliferation against cancerous U937 cells at the low concentration (60 μg/ ml) applied in this study compared with G (10%). Together, this demonstrates that sulfation clearly improved the solubility and therapeutic activities of G. The water-soluble GS demonstrates the potential multifunctional effects of these materials in foodstuffs.

Display of fungal hydrophobin on the Pichia pastoris cell surface and its influence on Candida antarctica lipase B

PubMed Central

Wang, Pan; He, Jie; Sun, Yufei; Reynolds, Matthew; Zhang, Li; Han, Shuangyan; Liang, Shuli; Sui, Haixin; Lin, Ying

2016-01-01

To modify the Pichia pastoris cell surface, two classes of hydrophobins, SC3 from Schizophyllum commune and HFBI from Trichoderma reesei, were separately displayed on the cell wall. There was an observable increase in the hydrophobicity of recombinant strains. Candida antarctica lipase B (CALB) was then co-displayed on the modified cells, generating strains GS115/SC3-61/CALB-51 and GS115/HFBI-61/CALB-51. Interestingly, the hydrolytic and synthetic activities of strain GS115/HFBI-61/CALB-51 increased by 37% and 109%, respectively, but decreased by 26% and 43%, respectively, in strain GS115/SC3-61/CALB-51 compared with the hydrophobin-minus recombinant strain GS115/CALB-GCW51. The amount of glycerol by-product from the transesterification reaction adsorbed on the cell surface was significantly decreased following hydrophobin modification, removing the glycerol barrier and allowing substrates to access the active sites of lipases. Electron micrographs indicated that the cell wall structures of both recombinant strains appeared altered, including changes to the inner glucan layer and outer mannan layer. These results suggest that the display of hydrophobins can change the surface structure and hydrophobic properties of P. pastoris, and affect the catalytic activities of CALB displayed on the surface of P. pastoris cells. PMID:26969039

Glutamine synthetase expression as a valuable marker of epilepsy and longer survival in newly diagnosed glioblastoma multiforme

PubMed Central

Rosati, Anna; Poliani, Pietro Luigi; Todeschini, Alice; Cominelli, Manuela; Medicina, Daniela; Cenzato, Marco; Simoncini, Edda Lucia; Magrini, Stefano Maria; Buglione, Michela; Grisanti, Salvatore; Padovani, Alessandro

2013-01-01

Abstract Background Glutamine synthetase (GS) is an astrocytic enzyme catalyzing the conversion of glutamate and ammonia to glutamine. Its up-regulation has been related to higher tumor proliferation and poor prognosis in extra-cerebral tumors. We have previously reported a GS deficiency in patients with glioblastoma multiforme (GBM) who also developed epilepsy, which is a favorable prognostic factor in glioma. Here, we investigated the prognostic value of GS expression in patients with GBM with or without epilepsy and its correlation with survival. Methods We conducted a clinical and histopathological study on 83 (52 males) consecutive patients with newly diagnosed GBM. Immunohistochemical expression of GS was scored semi-quantitatively on the basis of cell number, staining intensity, and distribution of immunoreactive cells. Several clinical and neuropathological variables were analyzed in relation to survival and GS expression. Results Median age at diagnosis was 62 years. At the last evaluation, with a median follow-up of 11.5 months (range, 1.5–58 months), 5 patients (6%) were still alive and 78 (94%) were dead. GS expression patterns in neoplastic cells were inversely correlated to the presence of epilepsy (P < .0001 for intensity and P < .009 for homogeneity of GS distribution, respectively). Univariate analysis showed that RPA score, epilepsy, O6-methylguanine-DNA methyltransferase (MGM)T status, application of Stupp protocol, and GS intensity pattern had a significant impact on survival. Absent/low intensity of GS expression was significantly associated with a longer survival in both uni- (19 vs 8 months; P < .0005) and multivariate (P = .003) analyses. Conclusions Absent/low-intensity GS expression pattern represents a valuable biomarker of both epilepsy and overall survival in GBM. PMID:23410662

Mapping of Variable DNA Methylation Across Multiple Cell Types Defines a Dynamic Regulatory Landscape of the Human Genome.

PubMed

Gu, Junchen; Stevens, Michael; Xing, Xiaoyun; Li, Daofeng; Zhang, Bo; Payton, Jacqueline E; Oltz, Eugene M; Jarvis, James N; Jiang, Kaiyu; Cicero, Theodore; Costello, Joseph F; Wang, Ting

2016-04-07

DNA methylation is an important epigenetic modification involved in many biological processes and diseases. Many studies have mapped DNA methylation changes associated with embryogenesis, cell differentiation, and cancer at a genome-wide scale. Our understanding of genome-wide DNA methylation changes in a developmental or disease-related context has been steadily growing. However, the investigation of which CpGs are variably methylated in different normal cell or tissue types is still limited. Here, we present an in-depth analysis of 54 single-CpG-resolution DNA methylomes of normal human cell types by integrating high-throughput sequencing-based methylation data. We found that the ratio of methylated to unmethylated CpGs is relatively constant regardless of cell type. However, which CpGs made up the unmethylated complement was cell-type specific. We categorized the 26,000,000 human autosomal CpGs based on their methylation levels across multiple cell types to identify variably methylated CpGs and found that 22.6% exhibited variable DNA methylation. These variably methylated CpGs formed 660,000 variably methylated regions (VMRs), encompassing 11% of the genome. By integrating a multitude of genomic data, we found that VMRs enrich for histone modifications indicative of enhancers, suggesting their role as regulatory elements marking cell type specificity. VMRs enriched for transcription factor binding sites in a tissue-dependent manner. Importantly, they enriched for GWAS variants, suggesting that VMRs could potentially be implicated in disease and complex traits. Taken together, our results highlight the link between CpG methylation variation, genetic variation, and disease risk for many human cell types. Copyright © 2016 Gu et al.

Water-soluble fractions from defatted sesame seeds protect human neuroblast cells against peroxyl radicals and hydrogen peroxide-induced oxidative stress.

PubMed

Ben Othman, Sana; Katsuno, Nakako; Kitayama, Akemi; Fujimura, Makoto; Kitaguchi, Kohji; Yabe, Tomio

2016-09-01

Oxidative stress is involved in the development of aging-related diseases, such as neurodegenerative diseases. Dietary antioxidants that can protect neuronal cells from oxidative damage play an important role in preventing such diseases. Previously, we reported that water-soluble fractions purified from defatted sesame seed flour exhibit good antioxidant activity in vitro. In the present study, we investigated the protective effects of white and gold sesame seed water-soluble fractions (WS-wsf and GS-wsf, respectively) against 2,2'-azobis(2-amidinopropane) dihydrochloride (AAPH) and hydrogen peroxide (H2O2) induced oxidative stress in human neuroblast SH-SY5Y cells. Pretreatment with WS-wsf and GS-wsf did not protect cells against AAPH-induced cytotoxicity, while simultaneous co-treatment with AAPH significantly improved cell viability and inhibited membrane lipid peroxidation. These results suggest that WS-wsf and GS-wsf protect cells from AAPH-induced extracellular oxidative damage via direct scavenging of peroxyl radicals. When oxidative stress was induced by H2O2, pretreatment WS-wsf and GS-wsf significantly enhanced cell viability. These results suggest that in addition to radical scavenging, WS-wsf and GS-wsf enhance cellular resistance to intracellular oxidative stress by activation of the Nrf-2/ARE pathway as confirmed by the increased Nrf2 protein level in the nucleus and increased heme oxygenase 1 (HO-1) mRNA expression. The roles of ferulic and vanillic acids as bioactive antioxidants in these fractions were also confirmed. In conclusion, our results indicated that WS-wsf and GS-wsf, which showed antioxidant activity in vitro, are also efficient antioxidants in a cell system protecting SH-SY5Y cells against both extracellular and intracellular oxidative stress.

Glutamine Synthetase Isoenzymes in the Green Soil Alga Stichococcus bacillaris Naeg.

PubMed

Ahmad, I; Hellebust, J A

1987-02-01

Two forms of glutamine synthetase (GS(1) and GS(2)) have been separated from cells of Stichococcus bacillaris by fast protein liquid chromatography. The activities of the two isoenzymes were influenced by the composition of the media employed; thiol reagents were essential for stabilizing GS(2) but they suppressed GS(1) activity. The activity of each isoenzyme was, therefore, determined following separate purification procedures. Growth conditions influenced both isoenzymes; GS(2) showed maximum activity under photoautotrophic conditions, whereas GS(1) showed maximum activity under heterotrophic conditions.

Glutamine Synthetase Isoenzymes in the Green Soil Alga Stichococcus bacillaris Naeg. 1

PubMed Central

Ahmad, Iftikhar; Hellebust, Johan A.

1987-01-01

Two forms of glutamine synthetase (GS1 and GS2) have been separated from cells of Stichococcus bacillaris by fast protein liquid chromatography. The activities of the two isoenzymes were influenced by the composition of the media employed; thiol reagents were essential for stabilizing GS2 but they suppressed GS1 activity. The activity of each isoenzyme was, therefore, determined following separate purification procedures. Growth conditions influenced both isoenzymes; GS2 showed maximum activity under photoautotrophic conditions, whereas GS1 showed maximum activity under heterotrophic conditions. PMID:16665232

Glutamine Acts as a Neuroprotectant against DNA Damage, Beta-Amyloid and H2O2-Induced Stress

PubMed Central

Chen, Jianmin; Herrup, Karl

2012-01-01

Glutamine is the most abundant free amino acid in the human blood stream and is ‘conditionally essential’ to cells. Its intracellular levels are regulated both by the uptake of extracellular glutamine via specific transport systems and by its intracellular synthesis by glutamine synthetase (GS). Adding to the regulatory complexity, when extracellular glutamine is reduced GS protein levels rise. Unfortunately, this excess GS can be maladaptive. GS overexpression is neurotoxic especially if the cells are in a low-glutamine medium. Similarly, in low glutamine, the levels of multiple stress response proteins are reduced rendering cells hypersensitive to H2O2, zinc salts and DNA damage. These altered responses may have particular relevance to neurodegenerative diseases of aging. GS activity and glutamine levels are lower in the Alzheimer's disease (AD) brain, and a fraction of AD hippocampal neurons have dramatically increased GS levels compared with control subjects. We validated the importance of these observations by showing that raising glutamine levels in the medium protects cultured neuronal cells against the amyloid peptide, Aβ. Further, a 10-day course of dietary glutamine supplementation reduced inflammation-induced neuronal cell cycle activation, tau phosphorylation and ATM-activation in two different mouse models of familial AD while raising the levels of two synaptic proteins, VAMP2 and synaptophysin. Together, our observations suggest that healthy neuronal cells require both intracellular and extracellular glutamine, and that the neuroprotective effects of glutamine supplementation may prove beneficial in the treatment of AD. PMID:22413000

Inhibition of Gαs/cAMP Signaling Decreases TCR-Stimulated IL-2 transcription in CD4(+) T Helper Cells.

PubMed

Hynes, Thomas R; Yost, Evan A; Yost, Stacy M; Hartle, Cassandra M; Ott, Braden J; Berlot, Catherine H

2015-07-06

The role of cAMP in regulating T cell activation and function has been controversial. cAMP is generally known as an immunosuppressant, but it is also required for generating optimal immune responses. As the effect of cAMP is likely to depend on its cellular context, the current study investigated whether the mechanism of activation of Gαs and adenylyl cyclase influences their effect on T cell receptor (TCR)-stimulated interleukin-2 (IL-2) mRNA levels. The effect of blocking Gs-coupled receptor (GsPCR)-mediated Gs activation on TCR-stimulated IL-2 mRNA levels in CD4(+) T cells was compared with that of knocking down Gαs expression or inhibiting adenylyl cyclase activity. The effect of knocking down Gαs expression on TCR-stimulated cAMP accumulation was compared with that of blocking GsPCR signaling. ZM-241385, an antagonist to the Gs-coupled A2A adenosine receptor (A2AR), enhanced TCR-stimulated IL-2 mRNA levels in primary human CD4(+) T helper cells and in Jurkat T cells. A dominant negative Gαs construct, GαsDN3, also enhanced TCR-stimulated IL-2 mRNA levels. Similar to GsPCR antagonists, GαsDN3 blocked GsPCR-dependent activation of both Gαs and Gβγ. In contrast, Gαs siRNA and 2',5'-dideoxyadenosine (ddA), an adenylyl cyclase inhibitor, decreased TCR-stimulated IL-2 mRNA levels. Gαs siRNA, but not GαsDN3, decreased TCR-stimulated cAMP synthesis. Potentiation of IL-2 mRNA levels by ZM-241385 required at least two days of TCR stimulation, and addition of ddA after three days of TCR stimulation enhanced IL-2 mRNA levels. GsPCRs play an inhibitory role in the regulation of TCR-stimulated IL-2 mRNA levels whereas Gαs and cAMP can play a stimulatory one. Additionally, TCR-dependent activation of Gαs does not appear to involve GsPCRs. These results suggest that the context of Gαs/cAMP activation and the stage of T cell activation and differentiation determine the effect on TCR-stimulated IL-2 mRNA levels.

Docosahexaenoic acid alters Gsα localization in lipid raft and potentiates adenylate cyclase.

PubMed

Zhu, Zhuoran; Tan, Zhoubin; Li, Yan; Luo, Hongyan; Hu, Xinwu; Tang, Ming; Hescheler, Jürgen; Mu, Yangling; Zhang, Lanqiu

2015-01-01

Supplementation with docosahexaenoic acid (DHA), an ω-3 polyunsaturated fatty acid (PUFA), recently has become popular for the amelioration of depression; however the molecular mechanism of DHA action remains unclear. The aim of this study was to investigate the mechanism underlying the antidepressant effect of DHA by evaluating Gsα localization in lipid raft and the activity of adenylate cyclase in an in vitro glioma cell model. Lipid raft fractions from C6 glioma cells treated chronically with DHA were isolated by sucrose gradient ultracentrifugation. The content of Gsα in lipid raft was analyzed by immunoblotting and colocalization of Gsα with lipid raft was subjected to confocal microscopic analysis. The intracellular cyclic adenosine monophosphate (cAMP) level was determined by cAMP immunoassay kit. DHA decreased the amount of Gsα in lipid raft, whereas whole cell lysate Gsα was not changed. Confocal microscopic analysis demonstrated that colocalization of Gsα with lipid raft was decreased, whereas DHA increased intracellular cAMP accumulation in a dose-dependent manner. Interestingly, we found that DHA increased the lipid raft level, instead of disrupting it. The results of this study suggest that DHA may exert its antidepressant effect by translocating Gsα from lipid raft and potentiating the activity of adenylate cyclase. Importantly, the reduced Gsα in lipid raft by DHA is independent of disruption of lipid raft. Overall, the study provides partial preclinical evidence supporting a safe and effective therapy using DHA for depression. Copyright © 2015 Elsevier Inc. All rights reserved.

Targeting of glutamine transporter ASCT2 and glutamine synthetase suppresses gastric cancer cell growth.

PubMed

Ye, Jianxin; Huang, Qiang; Xu, Jie; Huang, Jinsheng; Wang, Jinzhou; Zhong, Wenjing; Chen, Wannan; Lin, Xinjian; Lin, Xu

2018-05-01

Glutamine (Gln) is essential for the proliferation of most cancer cells, making it an appealing target for cancer therapy. However, the role of Gln in gastric cancer (GC) metabolism is unknown and Gln-targeted therapy against GC remains scarce. The aim of this study was to investigate the relevance of Gln in GC growth and targeting. Expression of Gln transporter ASCT2 and glutamine synthetase (GS) in the parental and molecularly engineered GC cells or in human GC specimens was determined by RT-PCR and western blot analysis or immunohistochemistry. Cell proliferation and survival was assessed by CCK-8 assay and colony formation assay. Intracellular Gln content was measured by a HPLC system. Effects of ASCT2 and/or GS inhibitor on tumor growth were investigated in xenograft models. A significant heterogeneity of GC cells was observed with respect to their response to the treatment of ASCT2 inhibitor benzylserine (BenSer). Gln deprivation did not affect the BenSer-resistant cell growth due to endogenous GS expression, whose inhibition remarkably reduced cell proliferation. The differential in vitro sensitivity correlated with overall intracellular Gln content. Combined therapy with both ASCT2 and GS inhibitors produced a greater therapeutic efficacy than the treatment of either inhibitor alone. Furthermore, 77% human GC tissues were found to express moderate and high levels of ASCT2, 12% of which also co-expressed relatively high levels of GS. Gln mediates GC growth and the therapeutic efficacy of Gln-targeted treatment relies on distinct ASCT2 and GS expression pattern in specific gastric cancer groups.

Early physiological flood tolerance is followed by slow post-flooding root recovery in the dryland riparian tree Eucalyptus camaldulensis subsp. refulgens.

PubMed

Argus, R E; Colmer, T D; Grierson, P F

2015-06-01

We investigated physiological and morphological responses to flooding and recovery in Eucalyptus camaldulensis subsp. refulgens, a riparian tree species from a dryland region prone to intense episodic floods. Seedlings in soil flooded for 88 d produced extensive adventitious roots, displayed stem hypertrophy (stem diameter increased by 93%) and increased root porosity owing to aerenchyma formation. Net photosynthesis (Pn) and stomatal conductance (gs) were maintained for at least 2 weeks of soil flooding, contrasting with previous studies of other subspecies of E. camaldulensis. Gradual declines followed in both gs (30% less than controls) and Pn (19% less). Total leaf soluble sugars did not differ between flooded and control plants. Root mass did not recover 32 d after flooding ceased, but gs was not lower than controls, suggesting the root system was able to functionally compensate. However, the limited root growth during recovery after flooding was surprising given the importance of extensive root systems in dryland environments. We conclude that early flood tolerance could be an adaptation to capitalize on scarce water resources in a water-limited environment. Overall, our findings highlight the need to assess flooding responses in relation to a species' fitness for particular flood regimes or ecological niches. © 2014 John Wiley & Sons Ltd.

Antihyperglycemic drug Gymnema sylvestre also shows anticancer potentials in human melanoma A375 cells via reactive oxygen species generation and mitochondria-dependent caspase pathway.

PubMed

Chakraborty, Debrup; Ghosh, Samrat; Bishayee, Kausik; Mukherjee, Avinaba; Sikdar, Sourav; Khuda-Bukhsh, Anisur Rahman

2013-09-01

Ethanolic extract of Gymnema sylvestre (GS) leaves is used as a potent antidiabetic drug in various systems of alternative medicine, including homeopathy. The present study was aimed at examining if GS also had anticancer potentials, and if it had, to elucidate its possible mechanism of action. We initially tested possible anticancer potential of GS on A375 cells (human skin melanoma) through MTT assay and determined cytotoxicity levels in A375 and normal liver cells; we then thoroughly studied its apoptotic effects on A375 cells through protocols such as Hoechst 33258, H2DCFDA, and rhodamine 123 staining and conducted ELISA for cytochrome c, caspase 3, and PARP activity levels; we determined the mRNA level expression of cytochrome c, caspase 3, Bcl2, Bax, PARP, ICAD, and EGFR signaling genes through semiquantitative reverse transcriptase polymerase chain reaction and conducted Western blot analysis of caspase 3 and PARP. We also analyzed cell cycle events, determined reactive oxygen species accumulation, measured annexin V-FITC/PI and rhodamine 123 intensity by flow cytometry. Compared with both normal liver cells and drug-untreated A375, the mortality of GS-treated A375 cells increased in a dose-dependent manner. Additionally, GS induced nuclear DNA fragmentation and showed an increased level of mRNA expression of apoptotic signal related genes cytochrome c, caspase 3, PARP, Bax, and reduced expression level of ICAD, EGFR, and the anti-apoptotic gene Bcl2. Overall results indicate GS to have significant anticancer effect on A375 cells apart from its reported antidiabetic effect, indicating possibility of its palliative use in patients with symptoms of both the diseases.

Constitutive stimulatory G protein activity in limb mesenchyme impairs bone growth.

PubMed

Karaca, Anara; Malladi, Vijayram Reddy; Zhu, Yan; Tafaj, Olta; Paltrinieri, Elena; Wu, Joy Y; He, Qing; Bastepe, Murat

2018-05-01

GNAS mutations leading to constitutively active stimulatory G protein alpha-subunit (Gsα) cause different tumors, fibrous dysplasia of bone, and McCune-Albright syndrome, which are typically not associated with short stature. Enhanced signaling of the parathyroid hormone/parathyroid hormone-related peptide receptor, which couples to multiple G proteins including Gsα, leads to short bones with delayed endochondral ossification. It has remained unknown whether constitutive Gsα activity also impairs bone growth. Here we generated mice expressing a constitutively active Gsα mutant (Gsα-R201H) conditionally upon Cre recombinase (cGsα R201H mice). Gsα-R201H was expressed in cultured bone marrow stromal cells from cGsα R201H mice upon adenoviral-Cre transduction. When crossed with mice in which Cre is expressed in a tamoxifen-regulatable fashion (CAGGCre-ER™), tamoxifen injection resulted in mosaic expression of the transgene in double mutant offspring. We then crossed the cGsα R201H mice with Prx1-Cre mice, in which Cre is expressed in early limb-bud mesenchyme. The double mutant offspring displayed short limbs at birth, with narrow hypertrophic chondrocyte zones in growth plates and delayed formation of secondary ossification center. Consistent with enhanced Gsα signaling, bone marrow stromal cells from these mice demonstrated increased levels of c-fos mRNA. Our findings indicate that constitutive Gsα activity during limb development disrupts endochondral ossification and bone growth. Given that Gsα haploinsufficiency also leads to short bones, as in patients with Albright's hereditary osteodystrophy, these results suggest that a tight control of Gsα activity is essential for normal growth plate physiology. Copyright © 2018 Elsevier Inc. All rights reserved.

PI3Kδ inhibitor idelalisib in combination with BTK inhibitor ONO/GS-4059 in diffuse large B cell lymphoma with acquired resistance to PI3Kδ and BTK inhibitors.

PubMed

Yahiaoui, Anella; Meadows, Sarah A; Sorensen, Rick A; Cui, Zhi-Hua; Keegan, Kathleen S; Brockett, Robert; Chen, Guang; Quéva, Christophe; Li, Li; Tannheimer, Stacey L

2017-01-01

Activated B-cell-like diffuse large B-cell lymphoma relies on B-cell receptor signaling to drive proliferation and survival. Downstream of the B-cell receptor, the key signaling kinases Bruton's tyrosine kinase and phosphoinositide 3-kinase δ offer opportunities for therapeutic intervention by agents such as ibrutinib, ONO/GS-4059, and idelalisib. Combination therapy with such targeted agents could provide enhanced efficacy due to complimentary mechanisms of action. In this study, we describe both the additive interaction of and resistance mechanisms to idelalisib and ONO/GS-4059 in a model of activated B-cell-like diffuse large B-cell lymphoma. Significant tumor regression was observed with a combination of PI3Kδ and Bruton's tyrosine kinase inhibitors in the mouse TMD8 xenograft. Acquired resistance to idelalisib in the TMD8 cell line occurred by loss of phosphatase and tensin homolog and phosphoinositide 3-kinase pathway upregulation, but not by mutation of PIK3CD. Sensitivity to idelalisib could be restored by combining idelalisib and ONO/GS-4059. Further evaluation of targeted inhibitors revealed that the combination of idelalisib and the phosphoinositide-dependent kinase-1 inhibitor GSK2334470 or the AKT inhibitor MK-2206 could partially overcome resistance. Characterization of acquired Bruton's tyrosine kinase inhibitor resistance revealed a novel tumor necrosis factor alpha induced protein 3 mutation (TNFAIP3 Q143*), which led to a loss of A20 protein, and increased p-IκBα. The combination of idelalisib and ONO/GS-4059 partially restored sensitivity in this resistant line. Additionally, a mutation in Bruton's tyrosine kinase at C481F was identified as a mechanism of resistance. The combination activity observed with idelalisib and ONO/GS-4059, taken together with the ability to overcome resistance, could lead to a new therapeutic option in activated B-cell-like diffuse large B-cell lymphoma. A clinical trial is currently underway to evaluate the combination of idelalisib and ONO/GS-4059 (NCT02457598).

Repurposing Treprostinil for Enhancing Hematopoietic Progenitor Cell Transplantation

PubMed Central

Kazemi, Zahra; Bergmayr, Christian; Prchal-Murphy, Michaela; Javaheri, Tahereh; Themanns, Madeleine; Pham, Ha T. T.; Strohmaier, Wolfgang; Sexl, Veronika; Zebedin-Brandl, Eva

2016-01-01

Activation of Gs-coupled receptors enhances engraftment of hematopoietic stem and progenitor cells (HSPCs). We tested the hypothesis that treprostinil, a prostacyclin analog approved for the treatment of pulmonary hypertension, can be repurposed to improve hematopoietic stem cell transplantation. Murine and human HSPCs were isolated from bone marrow and umbilical cord blood, respectively. Prostanoid receptor agonists and the combination thereof with forskolin were tested for their capacity to stimulate [3H]cAMP accumulation in HSPCs. Three independent approaches were employed to verify the ability of agonist-activated HSPCs to reconstitute the bone marrow in lethally irradiated recipient mice. The underlying mechanism was explored in cellular migration assays and by blocking C-X-C motif chemokine receptor 4 (CXCR4). Among several prostanoid agonists tested in combination with forskolin, treprostinil was most efficacious in raising intracellular cAMP levels in murine and human HPSCs. Injection of murine and human HSPCs, which had been pretreated with treprostinil and forskolin, enhanced survival of lethally irradiated recipient mice. Survival was further improved if recipient mice were subcutaneously administered treprostinil (0.15 mg kg−1 8 h−1) for 10 days. This regimen also reduced the number of HSPCs required to rescue lethally irradiated mice. Enhanced survival of recipient mice was causally related to treprostinil-enhanced CXCR4-dependent migration of HSPCs. Treprostinil stimulates the engraftment of human and murine hematopoietic stem cells without impairing their capacity for self-renewal. The investigated dose range corresponds to the dose approved for human use. Hence, these findings may be readily translated into a clinical application. PMID:26989084

The Positive Regulatory Roles of the TIFY10 Proteins in Plant Responses to Alkaline Stress

PubMed Central

Zhu, Dan; Li, Rongtian; Liu, Xin; Sun, Mingzhe; Wu, Jing; Zhang, Ning; Zhu, Yanming

2014-01-01

The TIFY family is a novel plant-specific protein family, and is characterized by a conserved TIFY motif (TIFF/YXG). Our previous studies indicated the potential roles of TIFY10/11 proteins in plant responses to alkaline stress. In the current study, we focused on the regulatory roles and possible physiological and molecular basis of the TIFY10 proteins in plant responses to alkaline stress. We demonstrated the positive function of TIFY10s in alkaline responses by using the AtTIFY10a and AtTIFY10b knockout Arabidopsis, as evidenced by the relatively lower germination rates of attify10a and attify10b mutant seeds under alkaline stress. We also revealed that ectopic expression of GsTIFY10a in Medicago sativa promoted plant growth, and increased the NADP-ME activity, citric acid content and free proline content but decreased the MDA content of transgenic plants under alkaline stress. Furthermore, expression levels of the stress responsive genes including NADP-ME, CS, H+-ppase and P5CS were also up-regulated in GsTIFY10a transgenic plants under alkaline stress. Interestingly, GsTIFY10a overexpression increased the jasmonate content of the transgenic alfalfa. In addition, we showed that neither GsTIFY10a nor GsTIFY10e exhibited transcriptional activity in yeast cells. However, through Y2H and BiFc assays, we demonstrated that GsTIFY10a, not GsTIFY10e, could form homodimers in yeast cells and in living plant cells. As expected, we also demonstrated that GsTIFY10a and GsTIFY10e could heterodimerize with each other in both yeast and plant cells. Taken together, our results provided direct evidence supporting the positive regulatory roles of the TIFY10 proteins in plant responses to alkaline stress. PMID:25375909

A Rare Allele of GS2 Enhances Grain Size and Grain Yield in Rice.

PubMed

Hu, Jiang; Wang, Yuexing; Fang, Yunxia; Zeng, Longjun; Xu, Jie; Yu, Haiping; Shi, Zhenyuan; Pan, Jiangjie; Zhang, Dong; Kang, Shujing; Zhu, Li; Dong, Guojun; Guo, Longbiao; Zeng, Dali; Zhang, Guangheng; Xie, Lihong; Xiong, Guosheng; Li, Jiayang; Qian, Qian

2015-10-05

Grain size determines grain weight and affects grain quality. Several major quantitative trait loci (QTLs) regulating grain size have been cloned; however, our understanding of the underlying mechanism that regulates the size of rice grains remains fragmentary. Here, we report the cloning and characterization of a dominant QTL, grain size on chromosome 2 (GS2), which encodes Growth-Regulating Factor 4 (OsGRF4), a transcriptional regulator. GS2 localizes to the nucleus and may act as a transcription activator. A rare mutation of GS2 affecting the binding site of a microRNA, OsmiR396c, causes elevated expression of GS2/OsGRF4. The increase in GS2 expression leads to larger cells and increased numbers of cells, which thus enhances grain weight and yield. The introduction of this rare allele of GS2/OsGRF4 into rice cultivars could significantly enhance grain weight and increase grain yield, with possible applications in breeding high-yield rice varieties. Copyright © 2015 The Author. Published by Elsevier Inc. All rights reserved.

Effects of kinetics of light-induced stomatal responses on photosynthesis and water-use efficiency.

PubMed

McAusland, Lorna; Vialet-Chabrand, Silvère; Davey, Philip; Baker, Neil R; Brendel, Oliver; Lawson, Tracy

2016-09-01

Both photosynthesis (A) and stomatal conductance (gs ) respond to changing irradiance, yet stomatal responses are an order of magnitude slower than photosynthesis, resulting in noncoordination between A and gs in dynamic light environments. Infrared gas exchange analysis was used to examine the temporal responses and coordination of A and gs to a step increase and decrease in light in a range of different species, and the impact on intrinsic water use efficiency was evaluated. The temporal responses revealed a large range of strategies to save water or maximize photosynthesis in the different species used in this study but also displayed an uncoupling of A and gs in most of the species. The shape of the guard cells influenced the rapidity of response and the overall gs values achieved, with different impacts on A and Wi . The rapidity of gs in dumbbell-shaped guard cells could be attributed to size, whilst in elliptical-shaped guard cells features other than anatomy were more important for kinetics. Our findings suggest significant variation in the rapidity of stomatal responses amongst species, providing a novel target for improving photosynthesis and water use. © 2016 The Authors. New Phytologist © 2016 New Phytologist Trust.

Anti-tumor efficacy study of the Bruton's tyrosine kinase (BTK) inhibitor, ONO/GS-4059, in combination with the glycoengineered type II anti-CD20 monoclonal antibody obinutuzumab (GA101) demonstrates superior in vivo efficacy compared to ONO/GS-4059 in combination with rituximab.

PubMed

Yasuhiro, Tomoko; Sawada, Wako; Klein, Christian; Kozaki, Ryohei; Hotta, Shingo; Yoshizawa, Toshio

2017-03-01

The activated B-cell diffuse large B-cell-like lymphoma (ABC-DLBCL) correlates with poor prognosis. The B-cell receptor signaling pathway is known to be dysregulated in NHL/CLL and given BTK is a downstream mediator of BCR signaling, BTK constitutes an interesting and obvious therapeutic target. Given the high potency and selectivity of the BTK inhibitor, ONO/GS-4059, it was hypothesized that, the anti-tumor activity of ONO/GS-4059 could be further enhanced by combining it with the anti-CD20 Abs, rituximab (RTX) or obinutuzumab (GA101). ONO/GS-4059 combined with GA101 or RTX was significantly better than the respective monotherapy with tumor growth inhibition (TGI) of 90% for the GA101 combination and 86% for the RTX combination. In contrast, ibrutinib (PCI-32765) combined with RTX did not result in improved efficacy compared with respective monotherapy. Taken together these data indicate that the combination of ONO/GS-4059 with rituximab and particularly obinutuzumab may be an effective treatment for ABC-DLBCL.

A new gallium complex inhibits tumor cell invasion and matrix metalloproteinase MMP-14 expression and activity.

PubMed

Mohsen, Ahmed; Collery, Philippe; Garnotel, Roselyne; Brassart, Bertrand; Etique, Nicolas; Mohamed Sabry, Gilane; Elsherif Hassan, Rasha; Jeannesson, Pierre; Desmaële, Didier; Morjani, Hamid

2017-08-16

In this study, we investigated the effect of [N-(5-chloro-2-hydroxyphenyl)-l-aspartato] chlorogallate (GS2), a new water soluble gallium complex, on cell invasion and on the expression and activity of matrix metalloproteinases (MMPs) in human metastatic HT-1080 fibrosarcoma and MDA-MB 231 breast carcinoma cells. The effect on cell invasion was studied using a modified Boyden chamber coated with a type-I collagen. We analyzed the effect of GS2 on MMP-2, MMP-9, and MMP-14 via zymography and enzymatic assay using high affinity fluorogenic substrates. The expression of MMP mRNA was analyzed via qRT-PCR. GS2 induced a decrease in cell invasion. A dose-dependent inhibition effect was observed on the activities of MMP-2, MMP-9, and MMP-14 with the IC 50 values of 168, 82, and 20 μM, respectively. A decrease in the expression of MMP-14 mRNA was observed in both cell lines, whereas the expression of MMP-2 and MMP-9 mRNA was decreased only in the MDA-MB231 cells. Data obtained for the expression of MMP-14 mRNA were confirmed via Western blotting. In fact, MMP-14 expression was decreased in the presence of GS2. Overall, these data show that GS2 is a promising compound for anti-invasive and anticancer therapy.

Confirmation and mechanism of glyphosate resistance in tall windmill grass (Chloris elata) from Brazil.

PubMed

Brunharo, Caio Acg; Patterson, Eric L; Carrijo, Daniela R; de Melo, Marcel Sc; Nicolai, Marcelo; Gaines, Todd A; Nissen, Scott J; Christoffoleti, Pedro J

2016-09-01

Overreliance on glyphosate as a single tool for weed management in agricultural systems in Brazil has selected glyphosate-resistant populations of tall windmill grass (Chloris elata Desv.). Two C. elata populations, one glyphosate resistant (GR) and one glyphosate susceptible (GS), were studied in detail for a dose-response experiment and for resistance mechanism. The dose causing 50% reduction in dry weight was 620 g a.e. ha(-1) for GR and 114 g ha(-1) for GS, resulting in an R/S ratio of 5.4. GS had significantly higher maximum (14) C-glyphosate absorption into the treated leaf (51.3%) than GR (39.5%), a difference of 11.8% in maximum absorption. GR also retained more (14) C-glyphosate in the treated leaf (74%) than GS (51%), and GR translocated less glyphosate (27%) to other plant parts (stems, roots and root exudation) than GS (36%). There were no mutations at the Pro106 codon in the gene encoding 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS). There was no difference in EPSPS genomic copy number or EPSPS transcription between GS and GR populations. Based on these data, reduced glyphosate absorption and increased glyphosate retention in the treated leaf contribute to glyphosate resistance in this C. elata population from Brazil. © 2015 Society of Chemical Industry. © 2015 Society of Chemical Industry.

Amino- and carboxy-terminal deletion mutants of Gs alpha are localized to the particulate fraction of transfected COS cells

PubMed Central

1992-01-01

To elucidate the structural basis for membrane attachment of the alpha subunit of the stimulatory G protein (Gs alpha), mutant Gs alpha cDNAs with deletions of amino acid residues in the amino and/or carboxy termini were transiently expressed in COS-7 cells. The particulate and soluble fractions prepared from these cells were analyzed by immunoblot using peptide specific antibodies to monitor distribution of the expressed proteins. Transfection of mutant forms of Gs alpha with either 26 amino terminal residues deleted (delta 3-28) or with 59 amino terminal residues deleted (delta 1-59) resulted in immunoreactive proteins which localized primarily to the particulate fraction. Similarly, mutants with 10 (delta 385-394), 32 (delta 353-384), or 42 (delta 353-394) amino acid residues deleted from the carboxy terminus also localized to the particulate fraction, as did a mutant form of Gs alpha lacking amino acid residues at both the amino and carboxy termini (delta 3-28)/(delta 353-384). Mutant and wild type forms of Gs alpha demonstrated a similar degree of tightness in their binding to membranes as demonstrated by treatment with 2.5 M NaCl or 6 M urea, but some mutant forms were relatively resistant compared with wild type Gs alpha to solubilization by 15 mM NaOH or 1% sodium cholate. We conclude that: (a) deletion of significant portions of the amino and/or carboxyl terminus of Gs alpha is still compatible with protein expression; (b) deletion of these regions is insufficient to cause cytosolic localization of the expressed protein. The basis of Gs alpha membrane targeting remains to be elucidated. PMID:1400589

A high-risk 70-gene signature is not associated with the detection of tumor cell dissemination to the bone marrow.

PubMed

Walter, Vincent P; Taran, Florin-Andrei; Wallwiener, Markus; Walter, Christina; Grischke, Eva-Maria; Wallwiener, Diethelm; Brucker, Sara Y; Hartkopf, Andreas D

2018-06-01

The 70-gene signature (70-GS) is a prognostic tool, grouping patients in risk groups to assess their need for adjuvant chemotherapy. Tumor cell dissemination to the bone marrow is a marker of minimal residual disease and associated with impaired survival. In this study, we aimed to evaluate whether 70-GS is associated with the presence of disseminated tumor cells (DTCs) in the bone marrow of patients with early breast cancer. In patients with hormone receptor-positive HER2-negative early breast cancer, the 70-GS was obtained and the presence of DTCs was immunohistochemically evaluated using cytokeratin staining with the A45-B/B3 antibody. 149 patients were included into the analysis. 40 (27%) had a high-risk 70-GS and 35 (23%) had detectable DTCs in their bone marrow. 9 (22%) of the 40 patients with high-risk 70-GS and 26 (24%) of the 109 patients with a low-risk 70-GS were positive for DTCs (p = 0.863). As both 70-GS and DTC detection are known prognostic factors but do not seem to correlate, a follow-up on a larger cohort is warranted to evaluate if a combination of the two is able to better stratify the relapse risk in early breast cancer patients.

Three Cytosolic Glutamine Synthetase Isoforms Located in Different Order Veins Work Together for N Remobilization and Seed Filling in Arabidopsis.

PubMed

Moison, Michael; Marmagne, Anne; Dinant, Sylvie; Soulay, Fabienne; Azzopardi, Marianne; Lothier, Jérémy; Citerne, Sylvie; Morin, Halima; Legay, Nicolas; Chardon, Fabien; Avice, Jean-Christophe; Reisdorf-Cren, Michèle; Masclaux-Daubresse, Céline

2018-06-05

Glutamine synthetase (GS) is central for ammonium assimilation and consists of cytosolic (GS1) and chloroplastic (GS2) isoenzymes. During plant ageing, GS2 protein decreases due to chloroplast degradation, and GS1 activity increases to support glutamine biosynthesis and N remobilization from senescing leaves. The role of the different Arabidopsis GS1 isoforms in nitrogen remobilization was examined using 15N tracing experiments. Only the triple gln1;1-gln1;2-gln1;3 mutation affecting the three GLN1;1, GLN1;2 and GLN1;3 genes reduced significantly N remobilization, seed yield, seed weight, harvest index and vegetative biomass. The triple gln1;1-gln1;2-gln1;3 mutant accumulated large amount of ammonium that cannot be assimilated by GS1. Alternative ammonium assimilation through asparagine biosynthesis was increased and related to higher ASN2 asparagine synthetase transcript levels. The GS2 transcript, protein and activity levels were also increased to compensate for the lack of GS1-related glutamine biosynthesis. Localization of the different GLN1 genes showed that they are all expressed in the phloem companion cells but in different order veins. In conclusion, our study shows that glutamine biosynthesis for N-remobilization occurs in all the order veins (major and minor) in leaves, is mainly catalysed by the three major GS1 isoforms (GLN1;1, GLN1;2 and GLN1;3) and is alternatively supported by AS2 in the veins and GS2 in the mesophyll cells.

Glutamine synthetase, heat shock protein-70, and glypican-3 in intrahepatic cholangiocarcinoma and tumors metastatic to liver.

PubMed

Lagana, Stephen M; Moreira, Roger K; Remotti, Helen E; Bao, Fei

2013-05-01

Glutamine synthetase (GS), heat shock protein-70 (HSP-70), and glypican-3 (GPC-3) are markers best characterized in hepatocellular lesions, where they are useful in distinguishing hepatocellular carcinoma from dysplastic nodules. Their staining patterns in intrahepatic cholangiocarcinoma (IH-ChCa) and metastatic tumors in liver are not well described. Tissue microarrays containing 41 IH-ChCa and 24 metastatic tumors in liver were stained with commercially available antibodies to GS, HSP-70, and GPC-3. Five percent staining of tumor cells was considered positive for HSP-70 and GPC-3. For GS, 50% was the cut-off. GS reactivity was present in 31 of 41 IH-ChCa (76%), with the median amount of staining being 65% of tumor cells. HSP-70 reactivity was present in 36 of 41 IH-ChCa (88%) with the median amount of staining being 75% of tumor cells. GPC-3 reactivity was absent from all IH-ChCa. Twenty-seven of 41 IH-ChCa cases were positive for both GS and HSP-70 (66%). GS reactivity was present in 17 of 24 tumors metastatic to liver (71%), with the median amount of staining being 50% of tumor cells. HSP-70 reactivity was present in 21 of 24 tumors metastatic to liver (88%) with the median amount of staining being 80% of tumor cells. GPC-3 reactivity was present in 2 of 24 tumors metastatic to liver (8%) with one showing 5% staining and the other showing 50% staining of tumor cells. Fifteen of 24 cases were positive for both GS and HSP-70 (63%), and 2 cases were positive for all 3 markers (8%). Of the panel of immunostains currently commonly used to distinguish hepatocellular carcinoma from dysplastic hepatocytic nodules, only GPC-3 did not react frequently with metastatic tumors and IH-ChCa, although there was staining in 2 metastatic tumors. GS and HSP-70 are typically positive in IH-ChCa and metastatic tumors. Nothing should be inferred about the histogenesis of a tumor based on positive staining with either of these 2 markers, which currently have no role in tumor of unknown origin panels.

Ses proteins as possible targets for vaccine development against Staphylococcus epidermidis infections.

PubMed

Hofmans, Dorien; Khodaparast, Laleh; Khodaparast, Ladan; Vanstreels, Els; Shahrooei, Mohammad; Van Eldere, Johan; Van Mellaert, Lieve

2018-05-09

The opportunistic pathogen Staphylococcus epidermidis is progressively involved in device-related infections. Since these infections involve biofilm formation, antibiotics are not effective. Conversely, a vaccine can be advantageous to prevent these infections. In view of vaccine development, predicted surface proteins were evaluated on their potential as a vaccine target. Immunoglobulins directed against S. epidermidis surface proteins SesB, M, O, Q and R, were used to firstly affirm their surface location. Further, inhibitory effects of these IgGs on biofilm formation were determined in vitro on polystyrene and polyurethane surfaces and in vivo using a subcutaneous catheter mouse model. We also examined the opsonophagocytic capacity of these IgGs. Surface localization of the five Ses proteins was demonstrated both for planktonic and sessile cells, though to a variable extent. Ses-specific IgGs added to planktonic cells had a variable inhibitory effect on cell adhesion to polystyrene, while only anti-SesO IgGs decreased cell attachment to polyurethane catheters. Although phagocytic killing was only obtained after opsonisation with SesB-specific IgGs, a significant reduction of in vivo formed biofilms was observed after administration of SesB-, SesM- and SesO-specific IgGs. Regardless of their characterization or function, S. epidermidis surface proteins can be adequate targets for vaccine development aiming the prevention of device-related infections caused by invasive S. epidermidis strains. Copyright © 2018 Elsevier Ltd. All rights reserved.

Synthesis and optimization of cholesterol-based diquaternary ammonium Gemini Surfactant (Chol-GS) as a new gene delivery vector.

PubMed

Kim, Bieong-Kil; Doh, Kyung-Oh; Bae, Yun-Ui; Seu, Young-Bae

2011-01-01

Amongst a number of potential nonviral vectors, cationic liposomes have been actively researched, with both gemini surfactants and bola amphiphiles reported as being in possession of good structures in terms of cell viability and in vitro transfection. In this study, a cholesterol-based diquaternary ammonium gemini surfactant (Chol-GS) was synthesized and assessed as a novel nonviral gene vector. Chol-GS was synthesized from cholesterol by way of four reaction steps. The optimal efficiency was found to be at a weight ratio of 1:4 of lipid:DOPE (1,2-dioleoyl-L-alpha- glycero-3-phosphatidylethanolamine), and at a ratio of between 10:1~15:1 of liposome:DNA. The transfection efficiency was compared with commercial liposomes and with Lipofectamine, 1,2-dimyristyloxypropyl-3-dimethylhydroxyethylammonium bromide (DMRIE-C), and N-[1-(2,3-dioleoyloxy)propyl]- N,N,N-trimethylammonium chloride (DOTAP). The results indicate that the efficiency of Chol-GS is greater than that of all the tested commercial liposomes in COS7 and Huh7 cells, and higher than DOTAP and Lipofectamine in A549 cells. Confirmation of these findings was observed through the use of green fluorescent protein expression. Chol-GS exhibited a moderate level of cytotoxicity, at optimum concentrations for efficient transfection, indicating cell viability. Hence, the newly synthesized Chol-GS liposome has the potential of being an excellent nonviral vector for gene delivery.

De Novo Glutamine Synthesis

PubMed Central

He, Qiao; Shi, Xinchong; Zhang, Linqi; Yi, Chang; Zhang, Xuezhen

2016-01-01

Purpose: The aim of this study was to investigate the role of de novo glutamine (Gln) synthesis in the proliferation of C6 glioma cells and its detection with 13N-ammonia. Methods: Chronic Gln-deprived C6 glioma (0.06C6) cells were established. The proliferation rates of C6 and 0.06C6 cells were measured under the conditions of Gln deprivation along with or without the addition of ammonia or glutamine synthetase (GS) inhibitor. 13N-ammonia uptake was assessed in C6 cells by gamma counting and in rats with C6 and 0.06C6 xenografts by micro–positron emission tomography (PET) scanning. The expression of GS in C6 cells and xenografts was assessed by Western blotting and immunohistochemistry, respectively. Results: The Gln-deprived C6 cells showed decreased proliferation ability but had a significant increase in GS expression. Furthermore, we found that low concentration of ammonia was sufficient to maintain the proliferation of Gln-deprived C6 cells, and 13N-ammonia uptake in C6 cells showed Gln-dependent decrease, whereas inhibition of GS markedly reduced the proliferation of C6 cells as well as the uptake of 13N-ammoina. Additionally, microPET/computed tomography exhibited that subcutaneous 0.06C6 xenografts had higher 13N-ammonia uptake and GS expression in contrast to C6 xenografts. Conclusion: De novo Gln synthesis through ammonia–glutamate reaction plays an important role in the proliferation of C6 cells. 13N-ammonia can be a potential metabolic PET tracer for Gln-dependent tumors. PMID:27118759

Glutathione-S-conjugate transport in plants

DOEpatents

Rea, Philip A.; Lu, Yu-Ping; Li, Ze-Sheng

2000-01-01

The invention includes an isolated DNA encoding a plant GS-X pump polypeptide and an isolated preparation of a plant GS-X pump polypeptide. Also included is an isolated preparation of a nucleic acid which is antisense in orientation to a portion or all of a plant GS-X pump gene. The invention also includes a cells, vectors and transgenic plants having an isolated DNA encoding a plant GS-X pump and methods of use thereof. In addition, the invention relates to plant GS-X pump promoter sequences and the uses thereof.

Inhibition of intracerebral glioblastoma growth by targeting the insulin-like growth factor 1 receptor involves different context-dependent mechanisms

PubMed Central

Zamykal, Martin; Martens, Tobias; Matschke, Jakob; Günther, Hauke S.; Kathagen, Annegret; Schulte, Alexander; Peters, Regina; Westphal, Manfred; Lamszus, Katrin

2015-01-01

Background Signaling by insulin-like growth factor 1 receptor (IGF-1R) can contribute to the formation and progression of many diverse tumor types, including glioblastoma. We investigated the effect of the IGF-1R blocking antibody IMC-A12 on glioblastoma growth in different in vivo models. Methods U87 cells were chosen to establish rapidly growing, angiogenesis-dependent tumors in the brains of nude mice, and the GS-12 cell line was used to generate highly invasive tumors. IMC-A12 was administered using convection-enhanced local delivery. Tumor parameters were quantified histologically, and the functional relevance of IGF-1R activation was analyzed in vitro. Results IMC-A12 treatment inhibited the growth of U87 and GS-12 tumors by 75% and 50%, respectively. In GS-12 tumors, the invasive tumor extension and proliferation rate were significantly reduced by IMC-A12 treatment, while apoptosis was increased. In IMC-A12–treated U87 tumors, intratumoral vascularization was markedly decreased, and tumor cell proliferation was moderately reduced. Flow cytometry showed that <2% of U87 cells but >85% of GS-12 cells expressed IGF-1R. Activation of IGF-1R by IGF-1 and IGF-2 in GS-12 cells was blocked by IMC-A12. Both ligands stimulated GS-12 cell proliferation, and IGF-2 also stimulated migration. IMC-A12 inhibited these stimulatory effects and increased apoptosis. In U87 cells, stimulation with either ligand had no functional effect. Conclusions IGF-1R blockade can inhibit glioblastoma growth by different mechanisms, including direct effects on the tumor cells as well as indirect anti-angiogenic effects. Hence, blocking IGF-1R may be useful to target both the highly proliferative, angiogenesis-dependent glioblastoma core component as well as the infiltrative periphery. PMID:25543125

Electrical signaling, stomatal conductance, ABA and Ethylene content in avocado trees in response to root hypoxia

PubMed Central

Gurovich, Luis; Schaffer, Bruce; García, Nicolás; Iturriaga, Rodrigo

2009-01-01

Avocado (Persea americana Mill.) trees are among the most sensitive of fruit tree species to root hypoxia as a result of flooded or poorly drained soil. Similar to drought stress, an early physiological response to root hypoxia in avocado is a reduction of stomatal conductance. It has been previously determined in avocado trees that an extracellular electrical signal between the base of stem and leaves is produced and related to reductions in stomatal conductance in response to drought stress. The current study was designed to determine if changes in the extracellular electrical potential between the base of the stem and leaves in avocado trees could also be detected in response to short-term (min) or long-term (days) root hypoxia, and if these signals could be related to stomatal conductance (gs), root and leaf ABA and ACC concentrations, ethylene emission from leaves and leaf abscission. In contrast to previous observations for drought-stressed trees, short-term or long-term root hypoxia did not stimulate an electrical potential difference between the base of the stem and leaves. Short-term hypoxia did not result in a significant decrease in gs compared with plants in the control treatment, and no differences in ABA concentration were found between plants subjected to hypoxia and control plants. Long-term hypoxia in the root zone resulted in a significant decrease in gs, increased leaf ethylene and increased leaf abscission. The results indicate that for avocado trees exposed to root hypoxia, electrical signals do not appear to be the primary root-to-shoot communication mechanism involved in signaling for stomatal closure as a result of hypoxia in the root zone. PMID:19649181

Reduction of ammonia and lactate through the coupling of glutamine synthetase selection and downregulation of lactate dehydrogenase-A in CHO cells.

PubMed

Noh, Soo Min; Park, Jin Hyoung; Lim, Myung Sin; Kim, Jong Won; Lee, Gyun Min

2017-02-01

Chinese hamster ovary (CHO) cell cultivation for production of therapeutic proteins is accompanied by production of metabolic wastes, mostly ammonia and lactate. To reduce ammonia production, the glutamine synthetase (GS) system was used to develop therapeutic monoclonal antibody (mAb)-producing CHO cells (SM-0.025). Additionally, the lactate dehydrogenase-A (LDH-A) was downregulated with shRNA to reduce lactate production in SM-0.025. The resulting mAb-producing cell lines (#2, #46, and #52) produced less ammonia than the host cell line during the exponential phase due to GS protein overexpression. LDH-A downregulation in SM-0.025 not only reduced lactate production but also further reduced ammonia production. Among the three LDH-A-downregulated clones, clone #2 had the highest mAb production along with significantly reduced specific lactate and ammonia production rates compared to those in SM-0.025. Waste reduction increased the galactosylation level of N-glycosylation, which improved mAb quality. LDH-A downregulation was also successfully applied to the host cell lines (CHO K1 and GS knockout CHO-K1). However, LDH-A downregulated host cells could not survive the pool-selection process wherein glutamine was excluded and methionine sulfoximine was added to the media. Taken together, LDH-A downregulation in the mAb-producing cell line generated with the GS system successfully reduced both ammonia and lactate levels, improving mAb galactosylation. However, LDH-A downregulation could not be applied to host cell lines because it hampered the selection process of the GS system.

A phase 1 clinical trial of the selective BTK inhibitor ONO/GS-4059 in relapsed and refractory mature B-cell malignancies.

PubMed

Walter, Harriet S; Rule, Simon A; Dyer, Martin J S; Karlin, Lionel; Jones, Ceri; Cazin, Bruno; Quittet, Philippe; Shah, Nimish; Hutchinson, Claire V; Honda, Hideyuki; Duffy, Kevin; Birkett, Joseph; Jamieson, Virginia; Courtenay-Luck, Nigel; Yoshizawa, Toshio; Sharpe, John; Ohno, Tomoya; Abe, Shinichiro; Nishimura, Akihisa; Cartron, Guillaume; Morschhauser, Franck; Fegan, Christopher; Salles, Gilles

2016-01-28

We report the results of a multicenter phase 1 dose-escalation study of the selective Bruton tyrosine kinase (BTK) inhibitor ONO/GS-4059 in 90 patients with relapsed/refractory B-cell malignancies. There were 9 dose-escalation cohorts ranging from 20 mg to 600 mg once daily with twice-daily regimens of 240 mg and 300 mg. Twenty-four of 25 evaluable chronic lymphocytic leukemia (CLL) patients (96%) responded to ONO/GS-4059, with a median treatment duration of 80 weeks; 21 CLL patients remain on treatment. Lymph node responses were rapid and associated with a concurrent lymphocytosis. Eleven of 12 evaluable patients with mantle cell lymphoma (92%) responded (median treatment duration, 40 weeks). Eleven of 31 non-germinal center B-cell diffuse large B-cell lymphoma patients (35%) responded but median treatment duration was 12 weeks due to development of progressive disease. ONO/GS-4059 was very well tolerated with 75% of adverse events (AEs) being Common Toxicity Criteria for Adverse Events version 4.0 grade 1 or grade 2. Grade 3/4 AEs were mainly hematologic and recovered spontaneously during therapy. One CLL patient experienced a grade 3 treatment-related bleeding event (spontaneous muscle hematoma) but no clinically significant diarrhea, cardiac dysrhythmias, or arthralgia were observed. No maximal tolerated dose (MTD) was reached in the CLL cohort. In the non-Hodgkin lymphoma cohort, 4 patients developed a dose-limiting toxicity, yielding an MTD of 480 mg once daily. ONO/GS-4059 has significant activity in relapsed/refractory B-cell malignancies without major drug-related toxicity. The selectivity of ONO/GS-4059 should confer advantages in combination therapies. This trial was registered at www.clinicaltrials.gov as #NCT01659255. © 2016 by The American Society of Hematology.

GS143, an I{kappa}B ubiquitination inhibitor, inhibits allergic airway inflammation in mice

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

Hirose, Koichi; Wakashin, Hidefumi; Oki, Mie

2008-09-26

Asthma is characterized by airway inflammation with intense eosinophil infiltration and mucus hyper-production, in which antigen-specific Th2 cells play critical roles. Nuclear factor-{kappa}B (NF-{kappa}B) pathway has been demonstrated to be essential for the production of Th2 cytokines and chemokines in the airways in murine asthma models. In the present study, we examined the effect of GS143, a novel small-molecule inhibitor of I{kappa}B ubiquitination, on antigen-induced airway inflammation and Th2 cytokine production in mice. Intranasal administration of GS143 prior to antigen challenge suppressed antigen-induced NF-{kappa}B activation in the lung of sensitized mice. Intranasal administration of GS143 also inhibited antigen-induced eosinophil andmore » lymphocyte recruitment into the airways as well as the expression of Th2 cytokines and eotaxin in the airways. Moreover, GS143 inhibited antigen-induced differentiation of Th2 cells but not of Th1 cells in vitro. Taken together, these results suggest that I{kappa}B ubiquitination inhibitor may have therapeutic potential against asthma.« less

Radioimmunoassay of the marjor group specific protein of endogenous baboon type C viruses: relation to the RD-114/CCC group and detection of antigen in normal baboon tissues

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

Sherr, C.J.; Todaro, G.J.

1974-09-01

The major group specific (gs) protein of the endogenous baboon type C virus M7 was purified to homogeneity by gel filtration and isoelectric focusing. The protein has a molecular weight of approximately 33,000, as determined by electrophoresis in polyacrylamide gels containing sodium dodecyl sulfate, and an isoelectric point (pl) of 7.1, different from the pls of similarly purified gs proteins from six other mammalian type C viruses. Detergent disrupted M7 virus, whether grown in canine thymus or human rhabdomyosarcoma cells, fully displaced radiolabeled M7 gs protein from antigen-antibody complexes in a competitive radioimmunoassay. No antigenic differences were detected among themore » gs proteins of five independent isolates of baboon type C viruses grown in various cultured cell lines. The gs proteins of six independently isolated feline viruses of the RD-114/CCC group were antigenically related to, but could be distinguished from, the gs proteins of baboon type C viruses. No significant cross-reactions were observed in the radioimmunoassay for M7 gs protein using several other type C viruses, including two previously isolated from a woolly monkey and a gibbon ape. Group specific antigen was found in normal baboon testicular and splenic tissues using the M7 radioimmunoassay; no gs antigen could be detected in these same tissues using a radioimmunoassay for the gs protein of the woolly monkey type C virus. No gs antigen was found in baboon liver or in the tissues of several other primates. (auth)« less

Guggulsterone Targets Smokeless Tobacco Induced PI3K/Akt Pathway in Head and Neck Cancer Cells

PubMed Central

Macha, Muzafar A.; Matta, Ajay; Chauhan, Shyam Singh; Siu, K. W. Michael; Ralhan, Ranju

2011-01-01

Background Epidemiological association of head and neck cancer with smokeless tobacco (ST) emphasizes the need to unravel the molecular mechanisms implicated in cancer development, and identify pharmacologically safe agents for early intervention and prevention of disease recurrence. Guggulsterone (GS), a biosafe nutraceutical, inhibits the PI3K/Akt pathway that plays a critical role in HNSCC development. However, the potential of GS to suppress ST and nicotine (major component of ST) induced HNSCC remains unexplored. We hypothesized GS can abrogate the effects of ST and nicotine on apoptosis in HNSCC cells, in part by activation of PI3K/Akt pathway and its downstream targets, Bax and Bad. Methods and Results Our results showed ST and nicotine treatment resulted in activation of PI3K, PDK1, Akt, and its downstream proteins - Raf, GSK3β and pS6 while GS induced a time dependent decrease in activation of PI3K/Akt pathway. ST and nicotine treatment also resulted in induction of Bad and Bax phosphorylation, increased the association of Bad with 14-3-3ζresulting in its sequestration in the cytoplasm of head and neck cancer cells, thus blocking its pro-apoptotic function. Notably, GS pre-treatment inhibited ST/nicotine induced activation of PI3K/Akt pathway, and inhibited the Akt mediated phosphorylation of Bax and Bad. Conclusions In conclusion, GS treatment not only inhibited proliferation, but also induced apoptosis by abrogating the effects of ST / nicotine on PI3K/Akt pathway in head and neck cancer cells. These findings provide a rationale for designing future studies to evaluate the chemopreventive potential of GS in ST / nicotine associated head and neck cancer. PMID:21383988

Guggulsterone targets smokeless tobacco induced PI3K/Akt pathway in head and neck cancer cells.

PubMed

Macha, Muzafar A; Matta, Ajay; Chauhan, Shyam Singh; Siu, K W Michael; Ralhan, Ranju

2011-02-24

Epidemiological association of head and neck cancer with smokeless tobacco (ST) emphasizes the need to unravel the molecular mechanisms implicated in cancer development, and identify pharmacologically safe agents for early intervention and prevention of disease recurrence. Guggulsterone (GS), a biosafe nutraceutical, inhibits the PI3K/Akt pathway that plays a critical role in HNSCC development. However, the potential of GS to suppress ST and nicotine (major component of ST) induced HNSCC remains unexplored. We hypothesized GS can abrogate the effects of ST and nicotine on apoptosis in HNSCC cells, in part by activation of PI3K/Akt pathway and its downstream targets, Bax and Bad. Our results showed ST and nicotine treatment resulted in activation of PI3K, PDK1, Akt, and its downstream proteins--Raf, GSK3β and pS6 while GS induced a time dependent decrease in activation of PI3K/Akt pathway. ST and nicotine treatment also resulted in induction of Bad and Bax phosphorylation, increased the association of Bad with 14-3-3ζresulting in its sequestration in the cytoplasm of head and neck cancer cells, thus blocking its pro-apoptotic function. Notably, GS pre-treatment inhibited ST/nicotine induced activation of PI3K/Akt pathway, and inhibited the Akt mediated phosphorylation of Bax and Bad. In conclusion, GS treatment not only inhibited proliferation, but also induced apoptosis by abrogating the effects of ST/nicotine on PI3K/Akt pathway in head and neck cancer cells. These findings provide a rationale for designing future studies to evaluate the chemopreventive potential of GS in ST/nicotine associated head and neck cancer.

Screening for Glycosylphosphatidylinositol-Modified Cell Wall Proteins in Pichia pastoris and Their Recombinant Expression on the Cell Surface

PubMed Central

Zhang, Li; Liang, Shuli; Zhou, Xinying; Jin, Zi; Jiang, Fengchun; Han, Shuangyan; Zheng, Suiping

2013-01-01

Glycosylphosphatidylinositol (GPI)-anchored glycoproteins have various intrinsic functions in yeasts and different uses in vitro. In the present study, the genome of Pichia pastoris GS115 was screened for potential GPI-modified cell wall proteins. Fifty putative GPI-anchored proteins were selected on the basis of (i) the presence of a C-terminal GPI attachment signal sequence, (ii) the presence of an N-terminal signal sequence for secretion, and (iii) the absence of transmembrane domains in mature protein. The predicted GPI-anchored proteins were fused to an alpha-factor secretion signal as a substitute for their own N-terminal signal peptides and tagged with the chimeric reporters FLAG tag and mature Candida antarctica lipase B (CALB). The expression of fusion proteins on the cell surface of P. pastoris GS115 was determined by whole-cell flow cytometry and immunoblotting analysis of the cell wall extracts obtained by β-1,3-glucanase digestion. CALB displayed on the cell surface of P. pastoris GS115 with the predicted GPI-anchored proteins was examined on the basis of potential hydrolysis of p-nitrophenyl butyrate. Finally, 13 proteins were confirmed to be GPI-modified cell wall proteins in P. pastoris GS115, which can be used to display heterologous proteins on the yeast cell surface. PMID:23835174

Long-acting peptidomimergic control of gigantism caused by pituitary acidophilic stem cell adenoma.

PubMed

Maheshwari, H G; Prezant, T R; Herman-Bonert, V; Shahinian, H; Kovacs, K; Melmed, S

2000-09-01

Gigantism is caused by GH hypersecretion occurring before epiphyseal long bone closure and usually is associated with pituitary adenoma. A 15-yr-old female patient presented with accelerated growth due to a large pituitary tumor that was surgically resected to relieve pressure effects. Second surgery to remove residual tumor tissue was followed by administration of octreotide LAR, a long-acting depot somatostatin analog, together with long-acting cabergoline. Height was over the 95th percentile, with evidence of a recent growth spurt. Serum GH levels were more than 60 ng/mL (normal, <10 ng/mL) with no suppression to 75 g oral glucose, and serum PRL (>8,000 ng/mL; normal, <23 ng/mL) and insulin-like growth factor I levels (845 ng/mL; age-matched normal, 242-660 ng/mL) were elevated. Histology, immunostaining, and electron microscopy demonstrated a pituitary acidophil stem cell adenoma. Tumor tissue expressed both somatostatin receptor type 2 and dopamine receptor type 2. The Gs alpha subunit, GHRH receptor, and MEN1 genes were intact, and tumor tissue abundantly expressed pituitary tumor transforming gene (PTTG). Serum GH and PRL levels were controlled after two surgeries, and with continued cabergoline and octreotide LAR GH, PRL, and insulin-like growth factor I levels were normalized. In conclusion, administration of long-acting somatostatin analog every 4 weeks in combination with a long-acting dopamine agonist biweekly controlled biochemical parameters and accelerated growth in a patient with gigantism caused by a rare pituitary acidophil stem cell adenoma.

The color and size of chili peppers (Capsicum annuum) influence Hep-G2 cell growth.

PubMed

Popovich, David G; Sia, Sharon Y; Zhang, Wei; Lim, Mon L

2014-11-01

Four types of chili (Capsicum annuum) extracts, categorized according to color; green and red, and size; small and large were studied in Hep-G2 cells. Red small (RS) chili had an LC50 value of 0.378 ± 0.029 compared to green big (GB) 1.034 ± 0.061 and green small (GS) 1.070 ± 0.21 mg/mL. Red big (RB) was not cytotoxic. Capsaicin content was highest in RS and produced a greater percentage sub-G1 cells (6.47 ± 1.8%) after 24 h compared to GS (2.96 ± 1.3%) and control (1.29 ± 0.8%) cells. G2/M phase was reduced by GS compared to RS and control cells. RS at the LC50 concentration contained 1.6 times the amount of pure capsaicin LC50 to achieve the same effect of capsaicin alone. GS and GB capsaicin content at the LC50 value was lower (0.2 and 0.66, respectively) compared to the amount of capsaicin to achieve a similar reduction in cell growth.

The Glycine soja NAC transcription factor GsNAC019 mediates the regulation of plant alkaline tolerance and ABA sensitivity.

PubMed

Cao, Lei; Yu, Yang; Ding, Xiaodong; Zhu, Dan; Yang, Fan; Liu, Beidong; Sun, Xiaoli; Duan, Xiangbo; Yin, Kuide; Zhu, Yanming

2017-10-01

Overexpression of Gshdz4 or GsNAC019 enhanced alkaline tolerance in transgenic Arabidopsis. We proved that Gshdz4 up-regulated both GsNAC019 and GsRD29B but GsNAC019 may repress the GsRD29B expression under alkaline stress. Wild soybean (Glycine soja) has a high tolerance to environmental challenges. It is a model species for dissecting the molecular mechanisms of salt-alkaline stresses. Although many NAC transcription factors play important roles in response to multiple abiotic stresses, such as salt, osmotic and cold, their mode of action in alkaline stress resistance is largely unknown. In our study, we identified a G. soja NAC gene, GsNAC019, which is a homolog of the Arabidopsis AtNAC019 gene. GsNAC019 was highly up-regulated by 50 mM NaHCO 3 treatment in the roots of wild soybean. Further investigation showed that a well-characterized transcription factor, Gshdz4 protein, bound the cis-acting element sequences (CAATA/TA), which are located in the promoter of the AtNAC019/GsNAC019 genes. Overexpression of Gshdz4 positively regulated AtNAC019 expression in transgenic Arabidopsis, implying that AtNAC019/GsNAC019 may be the target genes of Gshdz4. GsNAC019 was demonstrated to be a nuclear-localized protein in onion epidermal cells and possessed transactivation activity in yeast cells. Moreover, overexpression of GsNAC019 in Arabidopsis resulted in enhanced tolerance to alkaline stress at the seedling and mature stages, but reduced ABA sensitivity. The closest Arabidopsis homolog mutant plants of Gshdz4, GsNAC019 and GsRD29B containing athb40, atnac019 and atrd29b were sensitive to alkaline stress. Overexpression or the closest Arabidopsis homolog mutant plants of the GsNAC019 gene in Arabidopsis positively or negatively regulated the expression of stress-related genes, such as AHA2, RD29A/B and KIN1. Moreover, this mutation could phenotypically promoted or compromised plant growth under alkaline stress, implying that GsNAC019 may contribute to alkaline stress tolerance via the ABA signal transduction pathway and regulate expression of the downstream stress-related genes.

Leaf δ18O of remaining trees is affected by thinning intensity in a semiarid pine forest.

PubMed

Moreno-Gutiérrez, Cristina; Barberá, Gonzalo G; Nicolás, Emilio; DE Luis, Martín; Castillo, Víctor M; Martínez-Fernández, Faustino; Querejeta, José I

2011-06-01

Silvicultural thinning usually improves the water status of remaining trees in water-limited forests. We evaluated the usefulness of a dual stable isotope approach (δ¹³C, δ¹⁸O) for comparing the physiological performance of remaining trees between forest stands subjected to two different thinning intensities (moderate versus heavy) in a 60-year-old Pinus halepensis Mill. plantation in semiarid southeastern Spain. We measured bulk leaf δ¹³C and δ¹⁸O, foliar elemental concentrations, stem water content, stem water δ¹⁸O (δ¹⁸O(stem water)), tree ring widths and leaf gas exchange rates to assess the influence of forest stand density on tree performance. Remaining trees in low-density stands (heavily thinned) showed lower leaf δ¹⁸O, and higher stomatal conductance (g(s)), photosynthetic rate and radial growth than those in moderate-density stands (moderately thinned). By contrast, leaf δ¹³C, intrinsic water-use efficiency, foliar elemental concentrations and δ¹⁸O(stem water) were unaffected by stand density. Lower foliar δ¹⁸O in heavily thinned stands reflected higher g(s) of remaining trees due to decreased inter-tree competition for water, whereas higher photosynthetic rate was largely attributable to reduced stomatal limitation to CO₂ uptake. The dual isotope approach provided insight into the early (12 months) effects of stand density manipulation on the physiological performance of remaining trees. © 2011 Blackwell Publishing Ltd.

GS-2, a pyrazolo[1,5-a]indole derivative with inhibitory activity of topoisomerases, exerts its potent cytotoxic activity by ROS generation.

PubMed

Ji, Yuan Yuan; Zhu, Yong Ming; Wang, Jian Wen

2013-11-01

Pyrazolo[1,5-a]indole derivatives, a new type of topoisomerase (topo) inhibitor, demonstrate a broad spectrum of antitumor activities. However, the mechanism underlying the induced cytotoxicity remains unclear. In this study, we investigated whether GS-2, one of the derivatives, altered the levels of ROS in breast cancer MDA-231 cells and whether these ROS contributed to the observed antitumoral activity. Our data revealed that GS-2 caused a time- and dose-dependent elevation of intracellular ROS level in MDA-231 cells. GS-2 subsequently elicited notable inhibition on the expression of topos, DNA damage, activation of caspase-3, -9. The loss of mitochondrial membrane potential (MMP) was observed during the induction. The addition of N-acetyl cysteine (NAC, a well-known antioxidant) could effectively attenuate the GS-2-induced ROS enhancement and subsequent apoptosis. NAC attenuated the induced inhibition on expression of topos, indicating that topos might be the target of GS-2-induced ROS. The finding of the induced ROS provides new evidence for the molecular mechanisms of antitumor activity of pyrazolo[1,5-a]indole derivatives. Copyright © 2013 Elsevier B.V. All rights reserved.

FMISO accumulation in tumor is dependent on glutathione conjugation capacity in addition to hypoxic state.

PubMed

Masaki, Yukiko; Shimizu, Yoichi; Yoshioka, Takeshi; Nishijima, Ken-Ichi; Zhao, Songji; Higashino, Kenichi; Numata, Yoshito; Tamaki, Nagara; Kuge, Yuji

2017-10-01

18 F-fluoromisonidazole (FMISO), a well-known PET imaging probe for diagnosis of hypoxia, is believed to accumulate in hypoxic cells via covalent binding with macromolecules after reduction of the nitro group. Previously, we showed the majority of 18 F-FMISO was incorporated into low-molecular-weight metabolites in hypoxic tumors, and the glutathione conjugate of reduced FMISO (amino-FMISO-GS) distributed in the tumor hypoxic regions as revealed by imaging mass spectrometry (IMS). The present study was conducted to clarify whether FMISO is metabolized to amino-FMISO-GS within tumor cells and how amino-FMISO-GS contributes to FMISO accumulation in hypoxic cells. We also evaluated the relationship between FMISO accumulation and the glutathione conjugation-related factors in the cells. Tumor cells (FaDu, LOVO, and T24) were treated with 18 F-FMISO and incubated under normoxic or hypoxic conditions for 4 h. The FMISO metabolites were analyzed with LC-ESI-MS. Several glutathione conjugation-related factors of tumor cells were evaluated in vitro. FaDu tumor-bearing mice were intravenously injected with 18 F-FMISO and the tumors were excised at 4 h post-injection. Autoradiography, IMS and histologic studies were performed. Amino-FMISO-GS was the main contributor to FMISO incorporated in hypoxic FaDu cells in vitro and in vivo. Total FMISO uptake levels and amino-FMISO-GS levels were highest in FaDu, followed by LOVO, and then T24 (total uptake: 0.851 ± 0.009 (FaDu), 0.617 ± 0.021 (LOVO) and 0.167 ± 0.006 (T24) % dose/mg protein; amino-FMISO-GS: 0.502 ± 0.035 (FaDu), 0.158 ± 0.013 (LOVO), and 0.007 ± 0.001 (T24) % dose/mg protein). The glutathione level of FaDu was significantly higher than those of LOVO and T24. The enzyme activity of glutathione-S-transferase catalyzing the glutathione conjugation reaction in FaDu was similar levels to that in LOVO, and was higher than that in T24. Quantitative RT-PCR analysis revealed that the expression levels of efflux transporters of the glutathione conjugate (multidrug resistance-associated protein 1) were lowest in FaDu, followed by LOVO, and then T24. FMISO accumulates in hypoxic cells through reductive metabolism followed by glutathione conjugation. We illustrated the possibility that increased production and decreased excretion of amino-FMISO-GS contribute to FMISO accumulation in tumor cells under hypoxic conditions.

Supplementation with N-3 Long-Chain Polyunsaturated Fatty Acids or Olive Oil in Men and Women with Renal Disease Induces Differential Changes in the DNA Methylation of FADS2 and ELOVL5 in Peripheral Blood Mononuclear Cells

PubMed Central

Hoile, Samuel P.; Clarke-Harris, Rebecca; Huang, Rae-Chi; Calder, Philip C.; Mori, Trevor A.; Beilin, Lawrence J.; Lillycrop, Karen A.; Burdge, Graham C.

2014-01-01

Background Studies in animal models and in cultured cells have shown that fatty acids can induce alterations in the DNA methylation of specific genes. There have been no studies of the effects of fatty acid supplementation on the epigenetic regulation of genes in adult humans. Methods and Results We investigated the effect of supplementing renal patients with 4 g daily of either n-3 long-chain polyunsaturated fatty acids (n-3 LCPUFA) or olive oil (OO) for 8 weeks on the methylation status of individual CpG loci in the 5′ regulatory region of genes involved in PUFA biosynthesis in peripheral blood mononuclear cells from men and women (aged 53 to 63 years). OO and n-3 LCPUFA each altered (>10% difference in methylation) 2/22 fatty acid desaturase (FADS)-2 CpGs, while n-3 LCPUFA, but not OO, altered (>10%) 1/12 ELOVL5 CpGs in men. OO altered (>6%) 8/22 FADS2 CpGs and (>3%) 3/12 elongase (ELOVL)-5 CpGs, while n-3 LCPUFA altered (>5%) 3/22 FADS2 CpGs and 2/12 (>3%) ELOVL5 CpGs in women. FADS1 or ELOVL2 methylation was unchanged. The n-3 PUFA supplementation findings were replicated in blood DNA from healthy adults (aged 23 to 30 years). The methylation status of the altered CpGs in FADS2 and ELOVL5 was associated negatively with the level of their transcripts. Conclusions These findings show that modest fatty acid supplementation can induce altered methylation of specific CpG loci in adult humans, contingent on the nature of the supplement and on sex. This has implications for understanding the effect of fatty acids on PUFA metabolism and cell function. PMID:25329159

Epigenetic variability in cells of normal cytology is associated with the risk of future morphological transformation.

PubMed

Teschendorff, Andrew E; Jones, Allison; Fiegl, Heidi; Sargent, Alexandra; Zhuang, Joanna J; Kitchener, Henry C; Widschwendter, Martin

2012-03-27

Recently, it has been proposed that epigenetic variation may contribute to the risk of complex genetic diseases like cancer. We aimed to demonstrate that epigenetic changes in normal cells, collected years in advance of the first signs of morphological transformation, can predict the risk of such transformation. We analyzed DNA methylation (DNAm) profiles of over 27,000 CpGs in cytologically normal cells of the uterine cervix from 152 women in a prospective nested case-control study. We used statistics based on differential variability to identify CpGs associated with the risk of transformation and a novel statistical algorithm called EVORA (Epigenetic Variable Outliers for Risk prediction Analysis) to make predictions. We observed many CpGs that were differentially variable between women who developed a non-invasive cervical neoplasia within 3 years of sample collection and those that remained disease-free. These CpGs exhibited heterogeneous outlier methylation profiles and overlapped strongly with CpGs undergoing age-associated DNA methylation changes in normal tissue. Using EVORA, we demonstrate that the risk of cervical neoplasia can be predicted in blind test sets (AUC = 0.66 (0.58 to 0.75)), and that assessment of DNAm variability allows more reliable identification of risk-associated CpGs than statistics based on differences in mean methylation levels. In independent data, EVORA showed high sensitivity and specificity to detect pre-invasive neoplasia and cervical cancer (AUC = 0.93 (0.86 to 1) and AUC = 1, respectively). We demonstrate that the risk of neoplastic transformation can be predicted from DNA methylation profiles in the morphologically normal cell of origin of an epithelial cancer. Having profiled only 0.1% of CpGs in the human genome, studies of wider coverage are likely to yield improved predictive and diagnostic models with the accuracy needed for clinical application. The ARTISTIC trial is registered with the International Standard Randomised Controlled Trial Number ISRCTN25417821.

Epigenetic variability in cells of normal cytology is associated with the risk of future morphological transformation

PubMed Central

2012-01-01

Background Recently, it has been proposed that epigenetic variation may contribute to the risk of complex genetic diseases like cancer. We aimed to demonstrate that epigenetic changes in normal cells, collected years in advance of the first signs of morphological transformation, can predict the risk of such transformation. Methods We analyzed DNA methylation (DNAm) profiles of over 27,000 CpGs in cytologically normal cells of the uterine cervix from 152 women in a prospective nested case-control study. We used statistics based on differential variability to identify CpGs associated with the risk of transformation and a novel statistical algorithm called EVORA (Epigenetic Variable Outliers for Risk prediction Analysis) to make predictions. Results We observed many CpGs that were differentially variable between women who developed a non-invasive cervical neoplasia within 3 years of sample collection and those that remained disease-free. These CpGs exhibited heterogeneous outlier methylation profiles and overlapped strongly with CpGs undergoing age-associated DNA methylation changes in normal tissue. Using EVORA, we demonstrate that the risk of cervical neoplasia can be predicted in blind test sets (AUC = 0.66 (0.58 to 0.75)), and that assessment of DNAm variability allows more reliable identification of risk-associated CpGs than statistics based on differences in mean methylation levels. In independent data, EVORA showed high sensitivity and specificity to detect pre-invasive neoplasia and cervical cancer (AUC = 0.93 (0.86 to 1) and AUC = 1, respectively). Conclusions We demonstrate that the risk of neoplastic transformation can be predicted from DNA methylation profiles in the morphologically normal cell of origin of an epithelial cancer. Having profiled only 0.1% of CpGs in the human genome, studies of wider coverage are likely to yield improved predictive and diagnostic models with the accuracy needed for clinical application. Trial registration The ARTISTIC trial is registered with the International Standard Randomised Controlled Trial Number ISRCTN25417821. PMID:22453031

Next Generation Lithium-Ion Cell For Satellite Applications

NASA Astrophysics Data System (ADS)

Inoue, Takefumi; Segawa, Masazumi; Yoshida, Hiroaki; Takeda, Koichi

2011-10-01

GS Yuasa Technology has standard line up cells for satellite applications since 1999. The design of these cells is not changed and their production will continue. GS Yuasa is now developing higher performance Next Generation Lithium-ion cells. These cells have an improved positive material, negative material, electrolyte, and separator and have demonstrated excellent capacity retention with very low DC resistance growth during life testing. The new cell has approximately 40% more EOL energy (Wh/kg), and slightly lighter weight while using the same size components as our existing products.

Changes in leaf organisation, photosynthetic performance and wood formation during ex vitro acclimatisation of black mulberry (Morus nigra L.).

PubMed

Misalová, A; Durkovic, J; Mamonová, M; Priwitzer, T; Lengyelová, A; Hladká, D; Lux, A

2009-09-01

Changes in anatomical organisation of the leaf, photosynthetic performance and wood formation were examined to evaluate the temporal and spatial patterns of acclimatisation of micropropagated slow-growing black mulberry (Morus nigra L.) plantlets to the ex vitro environment. Leaf structure differentiation, the rates of net photosynthesis (P(n)), transpiration (E) and stomatal conductance (g(s)), and secondary xylem growth were determined in the course of a 56-day acclimatisation. Differentiation of palisade parenchyma was observed 7 days after transfer. At this stage, the rates of P(n), E and g(s) reached maximum values, after which the rates of all three gas exchange parameters gradually decreased. The highest proportion of woody area occupied by vessels was also observed 7 days after transfer. An important feature of developing woody tissue is the difference in patterns of vessel distribution from the characteristic differentiation patterns of earlywood and latewood vessels in mature wood of ring-porous trees. Vessels with lumen areas over 3000 microm(2) were only differentiated in acclimatised plantlets, whereas vessels in stems sampled on days 0 and 7 had very small lumen areas of up to 560 microm(2). Full acclimatisation, observed 56 days after transfer to the ex vitro environment, was associated with the rapid growth of new in vivo formed leaves, very low rates of E and g(s), and much increased secondary xylem tissue within the stem area.

Cost-effective three dimensional Ag/polymer dyes/graphene-carbon spheres hybrids for high performance nonenzymatic sensor and its application in living cell H2O2 detection.

PubMed

Lu, Baoping; Yuan, Xuna; Ren, Yuehong; Shi, Qinghua; Wang, Song; Dong, Jinlong; Nan, Ze-Dong

2018-05-03

We describe a facile method to synthesize a new type of catalyst by electrodepositing Ag nanocrystals (AgNCs) on the different polymer dyes, Poly (methylene blue) (PMB) or Poly (4-(2-Pyridylazo)-Resorcinol) (PAR) modified graphene‑carbon spheres (GS) hybrids. The self-assembled GS take dual advantages of carbon spheres and graphene. Carbon spheres acts as nano-spacers prevent the aggregation of graphene and guarantee the fast electron transfer of GS. Secondly, polymerized dyes used here are beneficial for AgNCs growing as a linker. The effects of dyes on the growth habits, morphologies and catalytic properties for AgNCs were investigated. A novel electrochemical nonenzymatic sensor for hydrogen peroxide (H 2 O 2 ) detection is fabricated based on the Ag/Polymer dyes/GS ternary composites modified glass carbon electrode (GCE) for the first time. It was found that the proposed electrodes, especially for Ag/PMB/GS/GCE, displayed a peculiar electrocatalytic activity towards H 2 O 2 reduction synergistically as compared to Ag/PAR/GS/GCE or Ag/GS/GCE alone. Ag/PMB/GS/GCE showed a linear response over the H 2 O 2 concentration range of 0.5 to 1112 μM. The detection limit and sensitivity is 0.15 μM and 400 μA mM -1  cm -2 , respectively. These outstanding results enable the practical application of Ag/PMB/GS/GCE for the H 2 O 2 tracking released from MCF-7 (human breast cancer cells) with satisfactory results. Copyright © 2018 Elsevier B.V. All rights reserved.

Changes in DNA Methylation from Age 18 to Pregnancy in Type 1, 2, and 17 T Helper and Regulatory T-Cells Pathway Genes

PubMed Central

Iqbal, Sabrina; Lockett, Gabrielle A.; Arshad, S. Hasan; Zhang, Hongmei; Kaushal, Akhilesh; Tetali, Sabarinath R.; Mukherjee, Nandini

2018-01-01

To succeed, pregnancies need to initiate immune biases towards T helper 2 (Th2) responses, yet little is known about what establishes this bias. Using the Illumina 450 K platform, we explored changes in DNA methylation (DNAm) of Th1, Th2, Th17, and regulatory T cell pathway genes before and during pregnancy. Female participants were recruited at birth (1989), and followed through age 18 years and their pregnancy (2011–2015). Peripheral blood DNAm was measured in 245 girls at 18 years; from among these girls, the DNAm of 54 women was repeatedly measured in the first (weeks 8–21, n = 39) and second (weeks 22–38, n = 35) halves of pregnancy, respectively. M-values (logit-transformed β-values of DNAm) were analyzed: First, with repeated measurement models, cytosine–phosphate–guanine sites (CpGs) of pathway genes in pregnancy and at age 18 (nonpregnant) were compared for changes (p ≤ 0.05). Second, we tested how many of the 348 pathway-related CpGs changed compared to 10 randomly selected subsets of all other CpGs and compared to 10 randomly selected subsets of other CD4+-related CpGs (348 in each subset). Contrasted to the nonpregnant state, 27.7% of Th1-related CpGs changed in the first and 36.1% in the second half of pregnancy. Among the Th2 pathway CpGs, proportions of changes were 35.1% (first) and 33.8% (second half). The methylation changes suggest involvement of both Th1 and Th2 pathway CpGs in the immune bias during pregnancy. Changes in regulatory T cell and Th17 pathways need further exploration. PMID:29415463

Assessing the osteoblast transcriptome in a model of enhanced bone formation due to constitutive Gs-G protein signaling in osteoblasts

PubMed Central

Wattanachanya, Lalita; Wang, Liping; Millard, Susan M.; Lu, Wei-Dar; O’Carroll, Dylan; Hsiao, Edward C.; Conklin, Bruce R.; Nissenson, Robert A.

2015-01-01

G protein–coupled receptor (GPCR) signaling in osteoblasts (OBs) is an important regulator of bone formation. We previously described a mouse model expressing Rs1, an engineered constitutively active Gs-coupled GPCR, under the control of the 2.3-kb Col I promoter. These mice showed a dramatic age-dependent increase in trabecular bone of femurs. Here, we further evaluated the effects of enhanced Gs signaling in OBs on intramembranous bone formation by examining calvariae of 1- and 9-week-old Col1(2.3)/Rs1 mice and characterized the in vivo gene expression specifically occurring in osteoblasts with activated Gs G protein–coupled receptor signaling, at the cellular level rather than in a whole bone. Rs1 calvariae displayed a dramatic increase in bone volume with partial loss of cortical structure. By immunohistochemistry, Osterix was detected in cells throughout the inter-trabecular space while Osteocalcin was expressed predominantly in cells along bone surfaces, suggesting the role of paracrine mediators secreted from OBs driven by 2.3-kb Col I promoter could influence early OB commitment, differentiation, and/or proliferation. Gene expression analysis of calvarial OBs revealed that genes affected by Rs1 signaling include those encoding proteins important for cell differentiation, cytokines and growth factors, angiogenesis, coagulation, and energy metabolism. The set of Gs-GPCRs and other GPCRs that may contribute to the observed skeletal phenotype and candidate paracrine mediators of the effect of Gs signaling in OBs were also determined. Our results identify novel detailed in vivo cellular changes of the anabolic response of the skeleton to Gs signaling in mature OBs. PMID:25704759

A novel glycogen synthase kinase-3 inhibitor optimized for acute myeloid leukemia differentiation activity

PubMed Central

Stetson, Lindsay; Ignatz-Hoover, James; Moreton, Stephen; Chakrabarti, Amit; Xia, Zhiqiang; Karan, Goutam; de Lima, Marcos; Agrawal, Mukesh K; Wald, David N

2016-01-01

Standard therapies used for the treatment of Acute Myeloid Leukemia (AML) are cytotoxic agents that target rapidly proliferating cells. Unfortunately, this therapeutic approach has limited efficacy and significant toxicity and the majority of AML patients still die of their disease. In contrast to the poor prognosis of most AML patients, most individuals with a rare subtype of AML, Acute Promyelocytic Leukemia (APL), can be cured by differentiation therapy using regimens containing all-trans retinoic acid. GSK3 has previously been identified as a therapeutic target in AML where its inhibition can lead to the differentiation and growth arrest of leukemic cells. Unfortunately, existing GSK3 inhibitors lead to suboptimal differentiation activity making them less useful as clinical AML differentiation agents. Here we describe the discovery of a novel GSK3 inhibitor, GS87. GS87 was discovered in efforts to optimize GSK3 inhibition for AML differentiation activity. Despite GS87's dramatic ability to induce AML differentiation, kinase profiling reveals its high specificity in targeting GSK3 as compared to other kinases. GS87 demonstrates high efficacy in a mouse AML model system and unlike current AML therapeutics, exhibits little effect on normal bone marrow cells. GS87 induces potent differentiation by more effectively activating GSK3-dependent signaling components including MAPK signaling as compared to other GSK3 inhibitors. GS87 is a novel GSK3 inhibitor with therapeutic potential as a differentiation agent for non-promyelocytic AML. PMID:27196775

Loss of Gi G-Protein-Coupled Receptor Signaling in Osteoblasts Accelerates Bone Fracture Healing.

PubMed

Wang, Liping; Hsiao, Edward C; Lieu, Shirley; Scott, Mark; O'Carroll, Dylan; Urrutia, Ashley; Conklin, Bruce R; Colnot, Celine; Nissenson, Robert A

2015-10-01

G-protein-coupled receptors (GPCRs) are key regulators of skeletal homeostasis and are likely important in fracture healing. Because GPCRs can activate multiple signaling pathways simultaneously, we used targeted disruption of G(i) -GPCR or activation of G(s) -GPCR pathways to test how each pathway functions in the skeleton. We previously demonstrated that blockade of G(i) signaling by pertussis toxin (PTX) transgene expression in maturing osteoblastic cells enhanced cortical and trabecular bone formation and prevented age-related bone loss in female mice. In addition, activation of G(s) signaling by expressing the G(s) -coupled engineered receptor Rs1 in maturing osteoblastic cells induced massive trabecular bone formation but cortical bone loss. Here, we test our hypothesis that the G(i) and G(s) pathways also have distinct functions in fracture repair. We applied closed, nonstabilized tibial fractures to mice in which endogenous G(i) signaling was inhibited by PTX, or to mice with activated G(s) signaling mediated by Rs1. Blockade of endogenous G(i) resulted in a smaller callus but increased bone formation in both young and old mice. PTX treatment decreased expression of Dkk1 and increased Lef1 mRNAs during fracture healing, suggesting a role for endogenous G(i) signaling in maintaining Dkk1 expression and suppressing Wnt signaling. In contrast, adult mice with activated Gs signaling showed a slight increase in the initial callus size with increased callus bone formation. These results show that G(i) blockade and G(s) activation of the same osteoblastic lineage cell can induce different biological responses during fracture healing. Our findings also show that manipulating the GPCR/cAMP signaling pathway by selective timing of G(s) and G(i) -GPCR activation may be important for optimizing fracture repair. © 2015 American Society for Bone and Mineral Research.

Therapeutic efficacy of the small molecule GS-5734 against Ebola virus in rhesus monkeys.

PubMed

Warren, Travis K; Jordan, Robert; Lo, Michael K; Ray, Adrian S; Mackman, Richard L; Soloveva, Veronica; Siegel, Dustin; Perron, Michel; Bannister, Roy; Hui, Hon C; Larson, Nate; Strickley, Robert; Wells, Jay; Stuthman, Kelly S; Van Tongeren, Sean A; Garza, Nicole L; Donnelly, Ginger; Shurtleff, Amy C; Retterer, Cary J; Gharaibeh, Dima; Zamani, Rouzbeh; Kenny, Tara; Eaton, Brett P; Grimes, Elizabeth; Welch, Lisa S; Gomba, Laura; Wilhelmsen, Catherine L; Nichols, Donald K; Nuss, Jonathan E; Nagle, Elyse R; Kugelman, Jeffrey R; Palacios, Gustavo; Doerffler, Edward; Neville, Sean; Carra, Ernest; Clarke, Michael O; Zhang, Lijun; Lew, Willard; Ross, Bruce; Wang, Queenie; Chun, Kwon; Wolfe, Lydia; Babusis, Darius; Park, Yeojin; Stray, Kirsten M; Trancheva, Iva; Feng, Joy Y; Barauskas, Ona; Xu, Yili; Wong, Pamela; Braun, Molly R; Flint, Mike; McMullan, Laura K; Chen, Shan-Shan; Fearns, Rachel; Swaminathan, Swami; Mayers, Douglas L; Spiropoulou, Christina F; Lee, William A; Nichol, Stuart T; Cihlar, Tomas; Bavari, Sina

2016-03-17

The most recent Ebola virus outbreak in West Africa, which was unprecedented in the number of cases and fatalities, geographic distribution, and number of nations affected, highlights the need for safe, effective, and readily available antiviral agents for treatment and prevention of acute Ebola virus (EBOV) disease (EVD) or sequelae. No antiviral therapeutics have yet received regulatory approval or demonstrated clinical efficacy. Here we report the discovery of a novel small molecule GS-5734, a monophosphoramidate prodrug of an adenosine analogue, with antiviral activity against EBOV. GS-5734 exhibits antiviral activity against multiple variants of EBOV and other filoviruses in cell-based assays. The pharmacologically active nucleoside triphosphate (NTP) is efficiently formed in multiple human cell types incubated with GS-5734 in vitro, and the NTP acts as an alternative substrate and RNA-chain terminator in primer-extension assays using a surrogate respiratory syncytial virus RNA polymerase. Intravenous administration of GS-5734 to nonhuman primates resulted in persistent NTP levels in peripheral blood mononuclear cells (half-life, 14 h) and distribution to sanctuary sites for viral replication including testes, eyes, and brain. In a rhesus monkey model of EVD, once-daily intravenous administration of 10 mg kg(-1) GS-5734 for 12 days resulted in profound suppression of EBOV replication and protected 100% of EBOV-infected animals against lethal disease, ameliorating clinical disease signs and pathophysiological markers, even when treatments were initiated three days after virus exposure when systemic viral RNA was detected in two out of six treated animals. These results show the first substantive post-exposure protection by a small-molecule antiviral compound against EBOV in nonhuman primates. The broad-spectrum antiviral activity of GS-5734 in vitro against other pathogenic RNA viruses, including filoviruses, arenaviruses, and coronaviruses, suggests the potential for wider medical use. GS-5734 is amenable to large-scale manufacturing, and clinical studies investigating the drug safety and pharmacokinetics are ongoing.

Glutamine Synthetase Is a Genetic Determinant of Cell Type–Specific Glutamine Independence in Breast Epithelia

PubMed Central

Kung, Hsiu-Ni; Marks, Jeffrey R.; Chi, Jen-Tsan

2011-01-01

Although significant variations in the metabolic profiles exist among different cells, little is understood in terms of genetic regulations of such cell type–specific metabolic phenotypes and nutrient requirements. While many cancer cells depend on exogenous glutamine for survival to justify the therapeutic targeting of glutamine metabolism, the mechanisms of glutamine dependence and likely response and resistance of such glutamine-targeting strategies among cancers are largely unknown. In this study, we have found a systematic variation in the glutamine dependence among breast tumor subtypes associated with mammary differentiation: basal- but not luminal-type breast cells are more glutamine-dependent and may be susceptible to glutamine-targeting therapeutics. Glutamine independence of luminal-type cells is associated mechanistically with lineage-specific expression of glutamine synthetase (GS). Luminal cells can also rescue basal cells in co-culture without glutamine, indicating a potential for glutamine symbiosis within breast ducts. The luminal-specific expression of GS is directly induced by GATA3 and represses glutaminase expression. Such distinct glutamine dependency and metabolic symbiosis is coupled with the acquisition of the GS and glutamine independence during the mammary differentiation program. Understanding the genetic circuitry governing distinct metabolic patterns is relevant to many symbiotic relationships among different cells and organisms. In addition, the ability of GS to predict patterns of glutamine metabolism and dependency among tumors is also crucial in the rational design and application of glutamine and other metabolic pathway targeted therapies. PMID:21852960

Corruption of the intra-gene DNA methylation architecture is a hallmark of cancer.

PubMed

Bartlett, Thomas E; Zaikin, Alexey; Olhede, Sofia C; West, James; Teschendorff, Andrew E; Widschwendter, Martin

2013-01-01

Epigenetic processes--including DNA methylation--are increasingly seen as having a fundamental role in chronic diseases like cancer. It is well known that methylation levels at particular genes or loci differ between normal and diseased tissue. Here we investigate whether the intra-gene methylation architecture is corrupted in cancer and whether the variability of levels of methylation of individual CpGs within a defined gene is able to discriminate cancerous from normal tissue, and is associated with heterogeneous tumour phenotype, as defined by gene expression. We analysed 270985 CpGs annotated to 18272 genes, in 3284 cancerous and 681 normal samples, corresponding to 14 different cancer types. In doing so, we found novel differences in intra-gene methylation pattern across phenotypes, particularly in those genes which are crucial for stem cell biology; our measures of intra-gene methylation architecture are a better determinant of phenotype than measures based on mean methylation level alone (K-S test [Formula: see text] in all 14 diseases tested). These per-gene methylation measures also represent a considerable reduction in complexity, compared to conventional per-CpG beta-values. Our findings strongly support the view that intra-gene methylation architecture has great clinical potential for the development of DNA-based cancer biomarkers.

Cytotoxicity of β-D-glucose/sucrose-coated silver nanoparticles depends on cell type, nanoparticles concentration and time of incubation

NASA Astrophysics Data System (ADS)

Vergallo, Cristian; Panzarini, Elisa; Carata, Elisabetta; Ahmadi, Meysam; Mariano, Stefania; Tenuzzo, Bernardetta Anna; Dini, Luciana

2016-06-01

The use of silver NanoParticles (AgNPs) in several consumer commercialized products, like food contact materials, medical devices and cosmetics has increased significantly, owing to their antibacterial and antifungal properties. Even though the NPs are widely diffused, due to the great variety in size, coating or shape, controversial data on their possible detrimental health effects still exist. Herein, by performing an easy and fast green method synthesis, we used β-D-glucose/sucrose to stabilize AgNPs and avoid the release of cytotoxic soluble silver ions Ag+ in the culture medium. The cytotoxic effects of these β-D-Glucose/Sucrose-Coated AgNPs (AgNPs-GS) was assessed on two cell culture models, which are human liver HepG2 and human Peripheral Blood Lymphocytes (PBLs) cells. AgNPs-GS, as determined by Transmission Electron Microscopy (TEM) analyses, had an average diameter of 30±5 nm, a spherical shape and were well-dispersed in the freshly-prepared solution. In addition, they were found spectrophotometrically stable throughout the experiment. Cytotoxicity, determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction assay, was evaluated by using two AgNPs-GS amounts, indicated as highest (10×103 of NPs/cell) and lowest (2×103 NPs/cell) concentration for 6, 12 and 24 h. The highest concentration of AgNPs-GS was significantly cytotoxic for both HepG2 and PBLs cells at all times, when compared with the negative control; conversely, the lowest amount of AgNPs-GS was toxic only for HepG2 cells. A significant increase of Reactive Oxygen Species (ROS) levels, determined by Nitro Blue Tetrazolium (NBT) reduction assay, was observed only in PBLs after treatment with NPs, by reaching maximum levels after the incubation with the lowest amount of NPs for 24 h. Significant morphological changes, depending on NPs/cell amount, characteristic of cell toxicity, like shape, cytoplasm, and nucleus alterations, were observed in lymphocytes and HepG2 cells exposed to AgNPs. The results indicate that HepG2 cells are more responsive to AgNPs treatment than PBLs. It is generally believed that cellular oxidative stress induces toxicity of NPs; however, in this study we did not detect any AgNPs-induced oxidative stress in HepG2, thus suggesting an alternative mechanism of toxicity in this cell line. In the whole, our findings suggest that AgNPs-GS-induced toxicity strictly depends on cell type, NPs amount and time of treatment.

Effects of High Glucose Levels and Glycated Serum on GIP Responsiveness in the Pancreatic Beta Cell Line HIT-T15.

PubMed

Puddu, Alessandra; Sanguineti, Roberta; Montecucco, Fabrizio; Viviani, Giorgio Luciano

2015-01-01

Glucose-dependent insulinotropic peptide (GIP) is an incretin hormone produced in the gastrointestinal tract that stimulates glucose dependent insulin secretion. Impaired incretin response has been documented in diabetic patients and was mainly related to the inability of the pancreatic beta cells to secrete insulin in response to GIP. Advanced Glycation End Products (AGEs) have been shown to play an important role in pancreatic beta cell dysfunction. The aim of this study is to investigate whether the exposure to AGEs can induce GIP resistance in the pancreatic beta cell line HIT-T15. Cells were cultured for 5 days in low (CTR) or high glucose (HG) concentration in the presence of AGEs (GS) to evaluate the expression of GIP receptor (GIPR), the intracellular signaling activated by GIP, and secretion of insulin in response to GIP. The results showed that incubation with GS alone altered intracellular GIP signaling and decreased insulin secretion as compared to CTR. GS in combination with HG reduced the expression of GIPR and PI3K and abrogated GIP-induced AKT phosphorylation and GIP-stimulated insulin secretion. In conclusion, we showed that treatment with GS is associated with the loss of the insulinotropic effect of GIP in hyperglycemic conditions.

Characterisation of the insulinotropic activity of an aqueous extract of Gymnema sylvestre in mouse beta-cells and human islets of Langerhans.

PubMed

Liu, Bo; Asare-Anane, Henry; Al-Romaiyan, Altaf; Huang, Guocai; Amiel, Stephanie A; Jones, Peter M; Persaud, Shanta J

2009-01-01

Leaves of the Gymnema sylvestre (GS) plant have been used to treat diabetes mellitus for millennia, but the previously documented insulin secretagogue effects of GS extracts in vitro may be non-physiological through damage to the beta-cells. We have now examined the effects of a novel GS extract (termed OSA) on insulin secretion from the MIN6 beta-cell line and isolated human islets of Langerhans. Insulin secretion from MIN6 cells was stimulated by OSA in a concentration-dependent manner, with low concentrations (0.06-0.25 mg/ml) having no deleterious effects on MIN6 cell viability, while higher concentrations (> or = 0.5 mg/ml) caused increased Trypan blue uptake. OSA increased beta-cell Ca2+ levels, an effect that was mediated by Ca2+ influx through voltage-operated calcium channels. OSA also reversibly stimulated insulin secretion from isolated human islets and its insulin secretagogue effects in MIN6 cells and human islets were partially dependent on the presence of extracellular Ca2+. These data indicate that low concentrations of the GS isolate OSA stimulate insulin secretion in vitro, at least in part as a consequence of Ca2+ influx, without compromising beta-cell viability. Identification of the component of the OSA extract that stimulates regulated insulin exocytosis, and further investigation of its mode(s) of action, may provide promising lead targets for Type 2 diabetes therapy. 2009 S. Karger AG, Basel.

Immunocytochemical localization of glutamic acid decarboxylase (GAD) and glutamine synthetase (GS) in the area postrema of the cat. Light and electron microscopy

NASA Technical Reports Server (NTRS)

D'Amelio, Fernando E.; Mehler, William R.; Gibbs, Michael A.; Eng, Lawrence F.; Wu, Jang-Yen

1987-01-01

Morphological evidence is presented of the existence of the putative neurotransmitter gamma-aminobutyric acid (GABA) in axon terminals and of glutamine synthetase (GS) in ependymoglial cells and astroglial components of the area postrema (AP) of the cat. Purified antiserum directed against the GABA biosynthetic enzyme glutamic acid decarboxylase (GAD) and GS antiserum were used. The results showed that punctate structures of variable size corresponding to axon terminals exhibited GAD-immunoreactivity and were distributed in varying densities. The greatest accumulation occurred in the caudal and middle segment of the AP and particularly in the area subpostrema, where the aggregation of terminals was extremely dense. The presence of both GAD-immunoreactive profiles and GS-immunostained ependymoglial cells and astrocytes in the AP provide further evidence of the functional correlation between the two enzymes.

Prospective assessment of a gene signature potentially predictive of clinical benefit in metastatic melanoma patients following MAGE-A3 immunotherapeutic (PREDICT).

PubMed

Saiag, P; Gutzmer, R; Ascierto, P A; Maio, M; Grob, J-J; Murawa, P; Dreno, B; Ross, M; Weber, J; Hauschild, A; Rutkowski, P; Testori, A; Levchenko, E; Enk, A; Misery, L; Vanden Abeele, C; Vojtek, I; Peeters, O; Brichard, V G; Therasse, P

2016-10-01

Genomic profiling of tumor tissue may aid in identifying predictive or prognostic gene signatures (GS) in some cancers. Retrospective gene expression profiling of melanoma and non-small-cell lung cancer led to the characterization of a GS associated with clinical benefit, including improved overall survival (OS), following immunization with the MAGE-A3 immunotherapeutic. The goal of the present study was to prospectively evaluate the predictive value of the previously characterized GS. An open-label prospective phase II trial ('PREDICT') in patients with MAGE-A3-positive unresectable stage IIIB-C/IV-M1a melanoma. Of 123 subjects who received the MAGE-A3 immunotherapeutic, 71 (58.7%) displayed the predictive GS (GS+). The 1-year OS rate was 83.1%/83.3% in the GS+/GS- populations. The rate of progression-free survival at 12 months was 5.8%/4.1% in GS+/GS- patients. The median time-to-treatment failure was 2.7/2.4 months (GS+/GS-). There was one complete response (GS-) and two partial responses (GS+). The MAGE-A3 immunotherapeutic was similarly immunogenic in both populations and had a clinically acceptable safety profile. Treatment of patients with MAGE-A3-positive unresectable stage IIIB-C/IV-M1a melanoma with the MAGE-A3 immunotherapeutic demonstrated an overall 1-year OS rate of 83.5%. GS- and GS+ patients had similar 1-year OS rates, indicating that in this study, GS was not predictive of outcome. Unexpectedly, the objective response rate was lower in this study than in other studies carried out in the same setting with the MAGE-A3 immunotherapeutic. Investigation of a GS to predict clinical benefit to adjuvant MAGE-A3 immunotherapeutic treatment is ongoing in another melanoma study.This study is registered at www.clinicatrials.gov NCT00942162. © The Author 2016. Published by Oxford University Press on behalf of the European Society for Medical Oncology.

Molecular cloning and characterization of glycogen synthase in Eriocheir sinensis.

PubMed

Li, Ran; Zhu, Li-Na; Ren, Li-Qi; Weng, Jie-Yang; Sun, Jin-Sheng

2017-12-01

Glycogen plays an important role in glucose and energy homeostasis at cellular and organismal levels. In glycogen synthesis, glycogen synthase (GS) is a rate-limiting enzyme catalysing the addition of α-1,4-linked glucose units from (UDP) 3 -glucose to a nascent glycogen chain using glycogenin (GN) as a primer. While studies on mammalian liver GS (GYS2) are numerous, enzymes from crustaceans, which also use glycogen and glucose as their main energy source, have received less attention. In the present study, we amplified full-length GS cDNA from Eriocheir sinensis. Tissue expression profiling revealed the highest expression of GS in the hepatopancreas. During moulting, GS expression and activity declined, and glycogen levels in the hepatopancreas were reduced. Recombinant GS was expressed in Escherichia coli Rosetta (DE3), and induction at 37°C or 16°C yielded EsGS in insoluble inclusion bodies (EsGS-I) or in soluble form (EsGS-S), respectively. Enzyme activity was measured in a cell-free system containing glucose-6-phosphate (G6P), and both forms possessed glycosyltransferase activity, but refolded EsGS-I was more active. Enzyme activity of both GS and EsGS-I in the hepatopancreas was optimum at 25°C, which is coincident with the optimum growth temperature of Chinese mitten crab, and higher (37°C) or lower (16°C) temperatures resulted in lower enzyme activity. Taken together, the results suggest that GS may be important for maintaining normal physiological functions such as growth and reproduction. Copyright © 2017 Elsevier Inc. All rights reserved.

Binding of /sup 18/F by cell membranes and cell walls of Streptococcus mutans

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

Yotis, W.W.; Zeb, M.; McNulty, J.

1983-07-01

The binding of /sup 18/F to isolated cell membranes and cell walls of Streptococcus mutans GS-5 or other bacteria was assayed. The attachment of /sup 18/F to these cell envelopes proceeded slowly and reached equilibrium within 60 min. /sup 18/F binding was stimulated by Ca/sup 2 +/ (1 mM). The binding of /sup 18/F to cellular components was dependent upon the pH, as well as the amount of /sup 18/F and dose of the binder employed. The binding of /sup 18/F by cell walls prepared from fluoride-sensitive and fluoride-resistant cells of S. salivarius and S. mutans did not differ significantly.more » The pretreatment of cell walls or cell membranes for 60 min at 30 degrees C with 1 mg of RNase, DNase, or trypsin per ml did not influence the binding of /sup 18/F by the walls and membranes of S. mutans GS-5. However, prior exposure of cell membranes to sodium dodecyl sulfate caused a significant reduction in the number of /sup 18/F atoms bound by the membranes. In saturated assay systems, cell membranes of S. mutans GS-5 bound 10(15) to 10(16) atoms of /sup 18/F per mg (dry weight), whereas cell walls from S. mutans GS-5, FA-1, and HS-6 or Actinomyces viscosus T14V and T14AV bound 10(12) to 10(13) atoms of /sup 18/F per mg (dry weight). /sup 18/F in this quantity (10(12) to 10(13) atoms) cannot be detected with the fluoride electrode. The data provide, for the first time, a demonstration of /sup 18/F binding by cell membranes and walls of oral flora.« less

Investigations of the toxic effects of glycans-based silver nanoparticles on different types of human cells

NASA Astrophysics Data System (ADS)

Panzarini, E.; Mariano, S.; Dini, L.

2017-08-01

The effects of glycans-capped AgNPs (30±5 nm average diameter, spherical shape) on biocompatibility and uptake was studied in relation to the glycan capping (glucose AgNPs-G, glucose/sucrose AgNPs-GS, glucose/fructose AgNPs-GF), and to the cell types (HeLa cells, lymphocytes, and HepG2 cells). Glycan capping and type of cells drive morphological changes, viability loss and type and extent of cell death induction; in addition cells response is largely influenced by the AgNPs amount. The MTT photometric method to determine cell metabolism and the analysis of the membrane integrity by Annexin V-Propidium Iodide labelling were used to quantify cell viability and cell death with different concentrations of NPs. It turns out that i) AgNPs-GF are the most toxic, whereas ii) AgNPs-GS are the less toxic NPs, probably due to the stability of glucose/sucrose capping up to 5 days in culture medium; iii) HepG2 cells are the most sensitive to the presence of NPs. A deeper investigation is necessary to explain the interesting PBLs proliferation increase observed in the presence of AgNPs-GS.

Human depression: a new approach in quantitative psychiatry

PubMed Central

2010-01-01

The biomolecular approach to major depression disorder is explained by the different steps that involve cell membrane viscosity, Gsα protein and tubulin. For the first time it is hypothesised that a biomolecular pathway exists, moving from cell membrane viscosity through Gsα protein and Tubulin, which can condition the conscious state and is measurable by electroencephalogram study of the brain's γ wave synchrony. PMID:20525273

Glycans coated silver nanoparticles induces autophagy and necrosis in HeLa cells

NASA Astrophysics Data System (ADS)

Panzarini, Elisa; Mariano, Stefania; Dini, Luciana

2015-06-01

This study reports the induction of autophagy by two concentrations (2×103 or 2×104 NPs/cell) of 30 nm sized β-D-Glucose- and β-D-Glucose/Sucrose-coated silver NanoParticles (AgNPs-G and AgNPs-GS respectively) in HeLa cells treated for 6, 12, 24 and 48 hrs. Cell viability was assessed by Neutral Red (NR) test and morphological evaluation. In addition ROS generation (NBT test) and induction of apoptosis/necrosis (Annexin V/Propidium Iodide-Annexin V/PI staining) and autophagy (Monodansylcadaverine-MDC staining) were evaluated. Cytotoxicity, ROS generation and morphology changes depend on NPs type and amount, and incubation time. As a general result, AgNPs-G are more toxic than AgNPs-GS. Moreover, the lowest AgNPs-GS concentration is ineffective on cell viability and ROS generation. Only 10% and 25% of viable HeLa cells were found at the end of incubation time in the presence of higher amount of AgNPs - G and AgNPs-GS respectively and in parallel ROS generation is induced. To elucidate the type of cell death, Annexin V/PI and MDC staining was performed. Interestingly, irrespective of coating type and NPs amount the percentage of apoptotic cells (Annexin V+/PI-) is similar to viable HeLa cells. At contrary, we observed a NPs amount dependent autophagy and necrosis induction. In fact, the lower amount of NPs induces autophagy (MDC+/PI- cells) whereas the higher one induces necrosis (Annexin V+/PI+ cells). Our findings suggest that AgNPs-induced cytotoxicity depends on AgNPs amount and type and provide preliminary evidence of induction of autophagy in HeLa cells cultured in the presence of AgNPs.

In vitro Fab display: a cell-free system for IgG discovery

PubMed Central

Stafford, Ryan L.; Matsumoto, Marissa L.; Yin, Gang; Cai, Qi; Fung, Juan Jose; Stephenson, Heather; Gill, Avinash; You, Monica; Lin, Shwu-Hwa; Wang, Willie D.; Masikat, Mary Rose; Li, Xiaofan; Penta, Kalyani; Steiner, Alex R.; Baliga, Ramesh; Murray, Christopher J.; Thanos, Christopher D.; Hallam, Trevor J.; Sato, Aaron K.

2014-01-01

Selection technologies such as ribosome display enable the rapid discovery of novel antibody fragments entirely in vitro. It has been assumed that the open nature of the cell-free reactions used in these technologies limits selections to single-chain protein fragments. We present a simple approach for the selection of multi-chain proteins, such as antibody Fab fragments, using ribosome display. Specifically, we show that a two-chain trastuzumab (Herceptin) Fab domain can be displayed in a format which tethers either the heavy or light chain to the ribosome while retaining functional antigen binding. Then, we constructed synthetic Fab HC and LC libraries and performed test selections against carcinoembryonic antigen (CEA) and vascular endothelial growth factor (VEGF). The Fab selection output was reformatted into full-length immunoglobulin Gs (IgGs) and directly expressed at high levels in an optimized cell-free system for immediate screening, purification and characterization. Several novel IgGs were identified using this cell-free platform that bind to purified CEA, CEA positive cells and VEGF. PMID:24586053

Nonantibiotic macrolides restore airway macrophage phagocytic function with potential anti-inflammatory effects in chronic lung diseases

PubMed Central

Tran, Hai B.; Hamon, Rhys; Roscioli, Eugene; Hodge, Greg; Jersmann, Hubertus; Ween, Miranda; Reynolds, Paul N.; Yeung, Arthur; Treiberg, Jennifer; Wilbert, Sibylle

2017-01-01

We reported defective efferocytosis associated with cigarette smoking and/or airway inflammation in chronic lung diseases, including chronic obstructive pulmonary disease, severe asthma, and childhood bronchiectasis. We also showed defects in phagocytosis of nontypeable Haemophilus influenzae (NTHi), a common colonizer of the lower airway in these diseases. These defects could be substantially overcome with low-dose azithromycin; however, chronic use may induce bacterial resistance. The aim of the present study was therefore to investigate two novel macrolides—2′-desoxy-9-(S)-erythromycylamine (GS-459755) and azithromycin-based 2′-desoxy molecule (GS-560660)—with significantly diminished antibiotic activity against Staphylococcus aureus, Streptococcus pneumonia, Moraxella catarrhalis, and H. influenzae. We tested their effects on efferocytosis, phagocytosis of NTHi, cell viability, receptors involved in recognition of apoptotic cells and/or NTHi (flow cytometry), secreted and cleaved intracellular IL-1β (cytometric bead array, immunofluorescence/confocal microscopy), and nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing 3 (NLRP3) using primary alveolar macrophages and THP-1 macrophages ± 10% cigarette smoke extract. Dose-response experiments showed optimal prophagocytic effects of GS-459755 and GS-560660 at concentrations of 0.5–1 µg/ml compared with our findings with azithromycin. Both macrolides significantly improved phagocytosis of apoptotic cells and NTHi (e.g., increases in efferocytosis and phagocytosis of NTHi: GS-459755, 23 and 22.5%, P = 0.043; GS-560660, 23.5 and 22%, P = 0.043, respectively). Macrophage viability remained >85% following 24 h exposure to either macrolide at concentrations up to 20 µg/ml. Secreted and intracellular-cleaved IL-1β was decreased with both macrolides with no significant changes in recognition molecules c-mer proto-oncogene tyrosine kinase; scavenger receptor class A, member 1; Toll-like receptor 2/4; or CD36. Particulate cytoplasmic immunofluorescence of NLRP3 inflammasome was also reduced significantly. We conclude that GS-459755 and GS-560660 may be useful for reducing airway inflammation in chronic lung diseases without inducing bacterial resistance. PMID:28258107

Nonantibiotic macrolides restore airway macrophage phagocytic function with potential anti-inflammatory effects in chronic lung diseases.

PubMed

Hodge, Sandra; Tran, Hai B; Hamon, Rhys; Roscioli, Eugene; Hodge, Greg; Jersmann, Hubertus; Ween, Miranda; Reynolds, Paul N; Yeung, Arthur; Treiberg, Jennifer; Wilbert, Sibylle

2017-05-01

We reported defective efferocytosis associated with cigarette smoking and/or airway inflammation in chronic lung diseases, including chronic obstructive pulmonary disease, severe asthma, and childhood bronchiectasis. We also showed defects in phagocytosis of nontypeable Haemophilus influenzae (NTHi), a common colonizer of the lower airway in these diseases. These defects could be substantially overcome with low-dose azithromycin; however, chronic use may induce bacterial resistance. The aim of the present study was therefore to investigate two novel macrolides-2'-desoxy-9-(S)-erythromycylamine (GS-459755) and azithromycin-based 2'-desoxy molecule (GS-560660)-with significantly diminished antibiotic activity against Staphylococcus aureus , Streptococcus pneumonia , Moraxella catarrhalis , and H. influenzae We tested their effects on efferocytosis, phagocytosis of NTHi, cell viability, receptors involved in recognition of apoptotic cells and/or NTHi (flow cytometry), secreted and cleaved intracellular IL-1β (cytometric bead array, immunofluorescence/confocal microscopy), and nucleotide-binding oligomerization domain-like receptor family pyrin domain-containing 3 (NLRP3) using primary alveolar macrophages and THP-1 macrophages ± 10% cigarette smoke extract. Dose-response experiments showed optimal prophagocytic effects of GS-459755 and GS-560660 at concentrations of 0.5-1 µg/ml compared with our findings with azithromycin. Both macrolides significantly improved phagocytosis of apoptotic cells and NTHi (e.g., increases in efferocytosis and phagocytosis of NTHi: GS-459755, 23 and 22.5%, P = 0.043; GS-560660, 23.5 and 22%, P = 0.043, respectively). Macrophage viability remained >85% following 24 h exposure to either macrolide at concentrations up to 20 µg/ml. Secreted and intracellular-cleaved IL-1β was decreased with both macrolides with no significant changes in recognition molecules c-mer proto-oncogene tyrosine kinase; scavenger receptor class A, member 1; Toll-like receptor 2/4; or CD36. Particulate cytoplasmic immunofluorescence of NLRP3 inflammasome was also reduced significantly. We conclude that GS-459755 and GS-560660 may be useful for reducing airway inflammation in chronic lung diseases without inducing bacterial resistance. Copyright © 2017 the American Physiological Society.

Chondroitin and glucosamine sulfate in combination decrease the pro-resorptive properties of human osteoarthritis subchondral bone osteoblasts: a basic science study

PubMed Central

Tat, Steeve Kwan; Pelletier, Jean-Pierre; Vergés, Josep; Lajeunesse, Daniel; Montell, Eulàlia; Fahmi, Hassan; Lavigne, Martin; Martel-Pelletier, Johanne

2007-01-01

Early in the pathological process of osteoarthritis (OA), subchondral bone remodelling, which is related to altered osteoblast metabolism, takes place. In the present study, we explored in human OA subchondral bone whether chondroitin sulfate (CS), glucosamine sulfate (GS), or both together affect the major bone biomarkers, osteoprotegerin (OPG), receptor activator of nuclear factor-kappa B ligand (RANKL), and the pro-resorptive activity of OA osteoblasts. The effect of CS (200 μg/mL), GS (50 and 200 μg/mL), or both together on human OA subchondral bone osteoblasts, in the presence or absence of 1,25(OH)2D3 (vitamin D3) (50 nM), was determined on the bone biomarkers alkaline phosphatase and osteocalcin, on the expression (mRNA) and production (enzyme-linked immunosorbent assay) of bone remodelling factors OPG and RANKL, and on the pro-resorptive activity of these cells. For the latter experiments, human OA osteoblasts were incubated with differentiated peripheral blood mononuclear cells on a sub-micron synthetic calcium phosphate thin film. Data showed that CS and GS affected neither basal nor vitamin D3-induced alkaline phosphatase or osteocalcin release. Interestingly, OPG expression and production under basal conditions or vitamin D3 treatment were upregulated by CS and by both CS and GS incubated together. Under basal conditions, RANKL expression was significantly reduced by CS and by both drugs incubated together. Under vitamin D3, these drugs also showed a decrease in RANKL level, which, however, did not reach statistical significance. Importantly, under basal conditions, CS and both compounds combined significantly upregulated the expression ratio of OPG/RANKL. Vitamin D3 decreased this ratio, and GS further decreased it. Both drugs reduced the resorption activity, and statistical significance was reached for GS and when CS and GS were incubated together. Our data indicate that CS and GS do not overly affect cell integrity or bone biomarkers. Yet CS and both compounds together increase the expression ratio of OPG/RANKL, suggesting a positive effect on OA subchondral bone structural changes. This was confirmed by the decreased resorptive activity for the combination of CS and GS. These data are of major significance and may help to explain how these two drugs exert a positive effect on OA pathophysiology. PMID:17996099

Nuclear Expression of GS28 Protein: A Novel Biomarker that Predicts Prognosis in Colorectal Cancers

PubMed Central

Lee, Sung Hak; Yoo, Hyung Jae; Rim, Do Eun; Cui, Yinji; Lee, Ahwon; Jung, Eun Sun; Oh, Seung Taek; Kim, Jun Gi; Kwon, Oh-Joo; Kim, Su Young; Jeong, Seong-Whan

2017-01-01

Aims: GS28 (Golgi SNARE protein, 28 kDa), a member of the soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNARE) protein family, plays a critical role in mammalian endoplasmic reticulum (ER)-Golgi or intra-Golgi vesicle transport. To date, few researches on the GS28 protein in human cancer tissues have been reported. In this study, we assessed the prognostic value of GS28 in patients with colorectal cancer (CRC). Methods and results: We screened for GS28 expression using immunohistochemistry in 230 surgical CRC specimens. The CRCs were right-sided and left-sided in 28.3% (65/230) and 71.3% (164/230) of patients, respectively. GS28 staining results were available in 214 cases. Among these, there were 26 nuclear predominant cases and 188 non-nuclear predominant cases. Stromal GS28 expression was noted in 152 cases of CRC. GS28 nuclear predominant immunoreactivity was significantly associated with advanced tumour stage (p = 0.045) and marginally associated with perineural invasion (p = 0.064). Decreased GS28 expression in the stromal cells was significantly associated with lymph node metastasis (N stage; p = 0.036). GS28 expression was not associated with epidermal growth factor receptor (EGFR) immunohistochemical positivity or KRAS mutation status. Investigation of the prognostic value of GS28 with Kaplan-Meier analysis revealed a correlation with overall survival (p = 0.004). Cases with GS28 nuclear predominant expression had significantly poorer overall survival than those with a non-nuclear predominant pattern. Conclusions: Taken together, these results indicate that GS28 nuclear predominant expression could serve as a prognostic marker for CRC and may help in identifying aggressive forms of CRC. PMID:28638266

Outgrowth of Rice Tillers Requires Availability of Glutamine in the Basal Portions of Shoots.

PubMed

Ohashi, Miwa; Ishiyama, Keiki; Kojima, Soichi; Konishi, Noriyuki; Sasaki, Kazuhiro; Miyao, Mitsue; Hayakawa, Toshihiko; Yamaya, Tomoyuki

2018-05-09

Our previous studies concluded that metabolic disorder in the basal portions of rice shoots caused by a lack of cytosolic glutamine synthetase1;2 (GS1;2) resulted in a severe reduction in the outgrowth of tillers. Rice mutants lacking GS1;2 (gs1;2 mutants) showed a remarkable reduction in the contents of both glutamine and asparagine in the basal portions of shoots. In the current study, we attempted to reveal the mechanisms for this decrease in asparagine content using rice mutants lacking either GS1;2 or asparagine synthetase 1 (AS1). The contributions of the availability of glutamine and asparagine to the outgrowth of rice tillers were investigated. Rice has two AS genes, and the enzymes catalyse asparagine synthesis from glutamine. In the basal portions of rice shoots, expression of OsAS1, the major species in this tissue, was reduced in gs1;2 mutants, whereas OsAS2 expression was relatively constant. OsAS1 was expressed in phloem companion cells of the nodal vascular anastomoses connected to the axillary bud vasculatures in the basal portions of wild-type shoots, whereas cell-specific expression was markedly reduced in gs1;2 mutants. OsAS1 was up-regulated significantly by NH 4 + supply in the wild type but not in gs1;2 mutants. When GS reactions were inhibited by methionine sulfoximine, OsAS1 was up-regulated by glutamine but not by NH 4 + . The rice mutants lacking AS1 (as1 mutants) showed a decrease in asparagine content in the basal portions of shoots. However, glutamine content and tiller number were less affected by the lack of AS1. These results indicate that in phloem companion cells of the nodal vascular anastomoses, asparagine synthesis is largely dependent on glutamine or its related metabolite-responsive AS1. Thus, the decrease in glutamine content caused by a lack of GS1;2 is suggested to result in low expression of OsAS1, decreasing asparagine content. However, the availability of asparagine generated from AS1 reactions is apparently less effective for the outgrowth of tillers. With respect to the tiller number and the contents of glutamine and asparagine in gs1;2 and as1 mutants, the availability of glutamine rather than asparagine in basal portions of rice shoots may be required for the outgrowth of rice tillers.

DNA methylation analysis reveals distinct methylation signatures in pediatric germ cell tumors.

PubMed

Amatruda, James F; Ross, Julie A; Christensen, Brock; Fustino, Nicholas J; Chen, Kenneth S; Hooten, Anthony J; Nelson, Heather; Kuriger, Jacquelyn K; Rakheja, Dinesh; Frazier, A Lindsay; Poynter, Jenny N

2013-06-27

Aberrant DNA methylation is a prominent feature of many cancers, and may be especially relevant in germ cell tumors (GCTs) due to the extensive epigenetic reprogramming that occurs in the germ line during normal development. We used the Illumina GoldenGate Cancer Methylation Panel to compare DNA methylation in the three main histologic subtypes of pediatric GCTs (germinoma, teratoma and yolk sac tumor (YST); N = 51) and used recursively partitioned mixture models (RPMM) to test associations between methylation pattern and tumor and demographic characteristics. We identified genes and pathways that were differentially methylated using generalized linear models and Ingenuity Pathway Analysis. We also measured global DNA methylation at LINE1 elements and evaluated methylation at selected imprinted loci using pyrosequencing. Methylation patterns differed by tumor histology, with 18/19 YSTs forming a distinct methylation class. Four pathways showed significant enrichment for YSTs, including a human embryonic stem cell pluripotency pathway. We identified 190 CpG loci with significant methylation differences in mature and immature teratomas (q < 0.05), including a number of CpGs in stem cell and pluripotency-related pathways. Both YST and germinoma showed significantly lower methylation at LINE1 elements compared with normal adjacent tissue while there was no difference between teratoma (mature and immature) and normal tissue. DNA methylation at imprinted loci differed significantly by tumor histology and location. Understanding methylation patterns may identify the developmental stage at which the GCT arose and the at-risk period when environmental exposures could be most harmful. Further, identification of relevant genetic pathways could lead to the development of new targets for therapy.

A High Throughput Screening Assay System for the Identification of Small Molecule Inhibitors of gsp

PubMed Central

Bhattacharyya, Nisan; Hu, Xin; Chen, Catherine Z.; Mathews Griner, Lesley A.; Zheng, Wei; Inglese, James; Austin, Christopher P.; Marugan, Juan J.; Southall, Noel; Neumann, Susanne; Northup, John K.; Ferrer, Marc; Collins, Michael T.

2014-01-01

Mis-sense mutations in the α-subunit of the G-protein, Gsα, cause fibrous dysplasia of bone/McCune-Albright syndrome. The biochemical outcome of these mutations is constitutively active Gsα and increased levels of cAMP. The aim of this study was to develop an assay system that would allow the identification of small molecule inhibitors specific for the mutant Gsα protein, the so-called gsp oncogene. Commercially available Chinese hamster ovary cells were stably transfected with either wild-type (WT) or mutant Gsα proteins (R201C and R201H). Stable cell lines with equivalent transfected Gsα protein expression that had relatively lower (WT) or higher (R201C and R201H) cAMP levels were generated. These cell lines were used to develop a fluorescence resonance energy transfer (FRET)–based cAMP assay in 1536-well microplate format for high throughput screening of small molecule libraries. A small molecule library of 343,768 compounds was screened to identify modulators of gsp activity. A total of 1,356 compounds with inhibitory activity were initially identified and reconfirmed when tested in concentration dose responses. Six hundred eighty-six molecules were selected for further analysis after removing cytotoxic compounds and those that were active in forskolin-induced WT cells. These molecules were grouped by potency, efficacy, and structural similarities to yield 22 clusters with more than 5 of structurally similar members and 144 singleton molecules. Seven chemotypes of the major clusters were identified for further testing and analyses. PMID:24667240

Biological study of the effect of water soluble [N-(2-hydroxybenzyl)-L-aspartato] gallium complexes on breast carcinoma and fibrosarcoma cells.

PubMed

Mohsen, Ahmed; Saby, Charles; Collery, Philippe; Sabry, Gilane Mohamed; Hassan, Rasha Elsherif; Badawi, Abdelfattah; Jeannesson, Pierre; Desmaële, Didier; Morjani, Hamid

2016-10-01

Two water soluble gallium complexes described as [Ga(III)LCl], where L is the deprotonated form of N-2-hydroxybenzyl aspartic acid derivatives, were synthesized and characterized by (1)H NMR, (13)C NMR, FT-IR, mass spectrometry, and elemental analysis. The 2-(5-chloro-2-hydroxybenzylamino)succinic acid derivative (GS2) has been found to be a promising anticancer drug candidate. This compound was found to be more cytotoxic against human breast carcinoma MDA-MB231 and fibrosarcoma HT-1080 cell lines than the unsubstituted derivative and GaCl3. GS2 was able to induce apoptosis through downregulation of AKT phosphorylation, G2M arrest in cell cycle, and caspase 3/7 pathway. This gallium complex was found to induce an increase in mitochondrial ROS level in HT-1080 cells but not in MDA-MB231 cells. This suggests that the mechanism of action of GS2 would not be mediated by the drug-induced oxidative stress but probably by directly and indirectly inhibiting the AKT cell-signaling pathway.

Cell size, genome size and the dominance of Angiosperms

NASA Astrophysics Data System (ADS)

Simonin, K. A.; Roddy, A. B.

2016-12-01

Angiosperms are capable of maintaining the highest rates of photosynthetic gas exchange of all land plants. High rates of photosynthesis depends mechanistically both on efficiently transporting water to the sites of evaporation in the leaf and on regulating the loss of that water to the atmosphere as CO2 diffuses into the leaf. Angiosperm leaves are unique in their ability to sustain high fluxes of liquid and vapor phase water transport due to high vein densities and numerous, small stomata. Despite the ubiquity of studies characterizing the anatomical and physiological adaptations that enable angiosperms to maintain high rates of photosynthesis, the underlying mechanism explaining why they have been able to develop such high leaf vein densities, and such small and abundant stomata, is still incomplete. Here we ask whether the scaling of genome size and cell size places a fundamental constraint on the photosynthetic metabolism of land plants, and whether genome downsizing among the angiosperms directly contributed to their greater potential and realized primary productivity relative to the other major groups of terrestrial plants. Using previously published data we show that a single relationship can predict guard cell size from genome size across the major groups of terrestrial land plants (e.g. angiosperms, conifers, cycads and ferns). Similarly, a strong positive correlation exists between genome size and both stomatal density and vein density that together ultimately constrains maximum potential (gs, max) and operational stomatal conductance (gs, op). Further the difference in the slopes describing the covariation between genome size and both gs, max and gs, op suggests that genome downsizing brings gs, op closer to gs, max. Taken together the data presented here suggests that the smaller genomes of angiosperms allow their final cell sizes to vary more widely and respond more directly to environmental conditions and in doing so bring operational photosynthetic metabolism closer to maximum potential photosynthesis.EndFragment

Adaptive Calcified Matrix Response of Dental Pulp to Bacterial Invasion Is Associated with Establishment of a Network of Glial Fibrillary Acidic Protein+/Glutamine Synthetase+ Cells

PubMed Central

Farahani, Ramin M.; Nguyen, Ky-Anh; Simonian, Mary; Hunter, Neil

2010-01-01

We report evidence for anatomical and functional changes of dental pulp in response to bacterial invasion through dentin that parallel responses to noxious stimuli reported in neural crest-derived sensory tissues. Sections of resin-embedded carious adult molar teeth were prepared for immunohistochemistry, in situ hybridization, ultrastructural analysis, and microdissection to extract mRNA for quantitative analyses. In odontoblasts adjacent to the leading edge of bacterial invasion in carious teeth, expression levels of the gene encoding dentin sialo-protein were 16-fold greater than in odontoblasts of healthy teeth, reducing progressively with distance from this site of the carious lesion. In contrast, gene expression for dentin matrix protein-1 by odontoblasts was completely suppressed in carious teeth relative to healthy teeth. These changes in gene expression were related to a gradient of deposited reactionary dentin that displayed a highly modified structure. In carious teeth, interodontoblastic dentin sialo-protein− cells expressing glutamine synthetase (GS) showed up-regulation of glial fibrillary acidic protein (GFAP). These cells extended processes that associated with odontoblasts. Furthermore, connexin 43 established a linkage between adjacent GFAP+/GS+ cells in carious teeth only. These findings indicate an adaptive pulpal response to encroaching caries that includes the deposition of modified, calcified, dentin matrix associated with networks of GFAP+/GS+ interodontoblastic cells. A regulatory role for the networks of GFAP+/GS+ cells is proposed, mediated by the secretion of glutamate to modulate odontoblastic response. PMID:20802180

High performance sulfur graphite full cell for next generation sulfur Li-ion battery

NASA Astrophysics Data System (ADS)

Wu, Yunwen; Momma, Toshiyuki; Yokoshima, Tokihiko; Nara, Hiroki; Osaka, Tetsuya

2018-06-01

Sulfur (S) Li-ion battery which use the metallic Li free anode is deemed as a promising solution to conquer the hazards originating from Li metal. However, stable cycling performance and low production price of the S Li-ion battery still remain challenging. Here, we propose a S-LixC full cell system by paring a S cathode and a pre-lithiated graphite anode which is cheap and commercially available. It shows stable cycling performance with a capacity around 1300 mAh (g-S)-1 at 0.2 C-rate and 1000 mAh (g-S)-1 at 0.5 C-rate. In addition, 0.1% per cycle capacity fading rate with a capacity retention of 880 mAh (g-S)-1 after 400 cycles at 0.2 C-rate has been achieved. The pre-formed solid electrolyte interphase (SEI) layer on the pre-lithaited graphite anode largely contributes to the high capacity performance. Notably, a 10-times-enlarged scale of S-LixC laminate type full cell has been assembled with high capacity performance (around 1000 mAh (g-S)-1) even after high rate cycling.

The role of gluten in a pound cake system: A model approach based on gluten-starch blends.

PubMed

Wilderjans, Edith; Pareyt, Bram; Goesaert, Hans; Brijs, Kristof; Delcour, Jan A

2008-10-15

In order to evaluate the role of gluten in cake-making, gluten-starch (GS) blends with different ratios of gluten to starch were tested in a research pound cake formula. The viscosities of batters made from commercial GS blends in the otherwise standardised formula increased with their gluten content. High viscosities during heating provide the batters with the capacity to retain expanding air nuclei, and thereby led to desired product volumes. In line with the above, increasing gluten levels in the cake recipes led to a more extended oven spring period. Cakes with a starch content exceeding 92.5% in the GS blend suffered from substantial collapse during cooling. They had a coarse crumb with a solid gummy layer at the bottom. Image analysis showed statistical differences in numbers of cells per cm(2), cell to total area ratio and mean cell area (p<0.05). Both density and mean cell area were related to gluten level. Moreover, mean cell area and cell to total area ratio were the highest for cakes with the lowest density and highest gluten levels. Relative sodium dodecyl sulfate (SDS, 2.0%) buffer (pH 6.8) extractabilities of protein from cakes baked with the different GS blends decreased with gluten content and were strongly correlated with the intensity of collapse. Taken together, the results teach that protein gives the cakes resistance to collapse, resulting in desirable volumes and an optimal grain structure with uniform cell distribution. Copyright © 2008 Elsevier Ltd. All rights reserved.

Is the WBE model appropriate for semi-arid shrubs subjected to clear cutting?

PubMed

Issoufou, Hassane Bil-Assanou; Rambal, Serge; Le Dantec, Valérie; Oï, Monique; Laurent, Jean-Paul; Saadou, Mahamane; Seghieri, Josiane

2015-02-01

It is crucial to understand the adaptive mechanisms of woody plants facing periodic drought to assess their vulnerability to the increasing climate variability predicted in the Sahel. Guiera senegalensis J.F.Gmel is a semi-evergreen Combretaceae commonly found in Sahelian rangelands, fallows and crop fields because of its value as an agroforestry species. We compared canopy leafing, and allometric measurements of leaf area, stem area and stem length and their relationships with leaf water potential, stomatal conductance (gs) and soil-to-leaf hydraulic conductance (KS-L), in mature and current-year resprouts of G. senegalensis in Sahelian Niger. In mature shrubs, seasonal drought reduced the ratio of leaf area to cross-sectional stem area (AL : AS), mainly due to leaf shedding. The canopy of the current-year resprouts remained permanently leafed as the shrubs produced leaves and stems continuously, and their AL : AS ratio increased throughout the dry season. Their KS-L increased, whereas gs decreased. West, Brown and Enquist's (WBE) model can thus describe allometric trends in the seasonal life cycle of undisturbed mature shrubs, but not that of resprouts. Annual clear cutting drives allometric scaling relationships away from theoretical WBE predictions in the current-year resprouts, with scaling exponents 2.5 times greater than those of mature shrubs. High KS-L (twice that of mature shrubs) supports this intensive regeneration process. The adaptive strategy described here is probably common to many woody species that have to cope with both severe seasonal drought and regular disturbance over the long term. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Mineral nitrogen sources differently affect root glutamine synthetase isoforms and amino acid balance among organs in maize.

PubMed

Prinsi, Bhakti; Espen, Luca

2015-04-03

Glutamine synthetase (GS) catalyzes the first step of nitrogen assimilation in plant cell. The main GS are classified as cytosolic GS1 and plastidial GS2, of which the functionality is variable according to the nitrogen sources, organs and developmental stages. In maize (Zea mays L.) one gene for GS2 and five genes for GS1 subunits are known, but their roles in root metabolism are not yet well defined. In this work, proteomic and biochemical approaches have been used to study root GS enzymes and nitrogen assimilation in maize plants re-supplied with nitrate, ammonium or both. The plant metabolic status highlighted the relevance of root system in maize nitrogen assimilation during both nitrate and ammonium nutrition. The analysis of root proteomes allowed a study to be made of the accumulation and phosphorylation of six GS proteins. Three forms of GS2 were identified, among which only the phosphorylated one showed an accumulation trend consistent with plastidial GS activity. Nitrogen availabilities enabled increments in root total GS synthetase activity, associated with different GS1 isoforms according to the nitrogen sources. Nitrate nutrition induced the specific accumulation of GS1-5 while ammonium led to up-accumulation of both GS1-1 and GS1-5, highlighting co-participation. Moreover, the changes in thermal sensitivity of root GS transferase activity suggested differential rearrangements of the native enzyme. The amino acid accumulation and composition in roots, xylem sap and leaves deeply changed in response to mineral sources. Glutamine showed the prevalent changes in all nitrogen nutritions. Besides, the ammonium nutrition was associated with an accumulation of asparagine and reducing sugars and a drop in glutamic acid level, significantly alleviated by the co-provision with nitrate. This work provides new information about the multifaceted regulation of the GS enzyme in maize roots, indicating the involvement of specific isoenzymes/isoforms, post-translational events and biochemical factors. For the first time, the proteomic approach allowed to discriminate the individual contribution of the GS1 isoforms, highlighting the participation of GS1-5 in nitrate metabolism. Moreover, the results give new insights about the influence of amino acid metabolism in plant C/N balance.

Annual Progress Report FY-91. Volume 1 and 2.

DTIC Science & Technology

1992-03-12

Pulmonary Med Tech 11 0644 GS Berger, TA Allergy Microbiologist 12 0403 GS Billups, L Flow Cytom Chemist 12 1320 GS Vacant Pulmonary Kyle Metabolic Unit...Reactions 11 (11/89) 3349 Salata, Kalman PhD. Mitogen-Inducible T Suppressor Cell 12 Assay by Flow Cytometry (12/89) 3350 Salata, Kalman PhD. Flow ...3/90) 3354 Salata, Kalman PhD. Two Way Mixed Lymphocyte Culture: 17 Analysis by Two Color Flow Cytometry (4/90) 3355 Salata, Kalman PhD. Effect of

Proliferative human cell sources applied as biocomponent in bioartificial livers: a review.

PubMed

Nibourg, Geert A A; Chamuleau, Robert A F M; van Gulik, Thomas M; Hoekstra, Ruurdtje

2012-07-01

Bioartificial livers (BALs) are urgently needed to bridge severe liver failure patients to liver transplantation or liver regeneration. When based on primary hepatocytes, their efficacy has been shown in animal experiments and their safety was confirmed in clinical trials. However, a proliferative human cell source with therapeutic functionality is needed to secure availability and move BAL application forward. This review compares the performance of BALs based on proliferative human biocomponents and primary hepatocytes. This review evaluates relevant studies identified by searching the MEDLINE database until July 2011 and some of our own unpublished data. All the discussed hepatocyte-like biocomponents show deficiencies in their hepatic functionality compared with primary hepatocytes, particularly functions occurring late in liver development. Nonetheless, the HepaRG, HepG2-GS-CYP3A4, and mesenchymal stem cells show efficacy in a statistically well-powered animal model of acute liver failure, when applied in a BAL device. Various methods to gain higher functionality of BALs, including genetic modification, the usage of combinatory cell sources, and improvement of culture methods, have scarcely been applied, but may further pave the path for BAL application. Clinical implementation of a BAL based on a human proliferative biocomponent is still several years away.

RoMo: An efficient strategy for functional mosaic analysis via stochastic Cre recombination and gene targeting in the ROSA26 locus.

PubMed

Movahedi, Kiavash; Wiegmann, Robert; De Vlaminck, Karen; Van Ginderachter, Jo A; Nikolaev, Viacheslav O

2018-07-01

Functional mosaic analysis allows for the direct comparison of mutant cells with differentially marked control cells in the same organism. While this offers a powerful approach for elucidating the role of specific genes or signalling pathways in cell populations of interest, genetic strategies for generating functional mosaicism remain challenging. We describe a novel and streamlined approach for functional mosaic analysis, which combines stochastic Cre/lox recombination with gene targeting in the ROSA26 locus. With the RoMo strategy a cell population of interest is randomly split into a cyan fluorescent and red fluorescent subset, of which the latter overexpresses a chosen transgene. To integrate this approach into high-throughput gene targeting initiatives, we developed a procedure that utilizes Gateway cloning for the generation of new targeting vectors. RoMo can be used for gain-of-function experiments or for altering signaling pathways in a mosaic fashion. To demonstrate this, we developed RoMo-dnGs mice, in which Cre-recombined red fluorescent cells co-express a dominant-negative Gs protein. RoMo-dnGs mice allowed us to inhibit G protein-coupled receptor activation in a fraction of cells, which could then be directly compared to differentially marked control cells in the same animal. We demonstrate how RoMo-dnGs mice can be used to obtain mosaicism in the brain and in peripheral organs for various cell types. RoMo offers an efficient new approach for functional mosaic analysis that extends the current toolbox and may reveal important new insights into in vivo gene function. © 2018 Wiley Periodicals, Inc.

Murine Denys-Drash syndrome: evidence of podocyte de-differentiation and systemic mediation of glomerulosclerosis.

PubMed

Patek, Charles E; Fleming, Stewart; Miles, Colin G; Bellamy, Christopher O; Ladomery, Michael; Spraggon, Lee; Mullins, John; Hastie, Nicholas D; Hooper, Martin L

2003-09-15

Denys-Drash syndrome (DDS) is caused by dominant mutations of the Wilms' tumour suppressor gene, WT1, and characterized by a nephropathy involving diffuse mesangial sclerosis, male pseudohermaphroditism and/or Wilms' tumourigenesis. Previously, we reported that heterozygosity for the Wt1tmT396 mutation induces DDS in heterozygous and chimeric (Wt1tmT396/+<-->+/+) mice. In the present study, the fate of Wt1 mutant cells in chimeric kidneys was assessed by in situ marker analysis, and immunocytochemistry was used to re-examine the claim that glomerulosclerosis (GS) is caused by loss of WT1 and persistent Pax-2 expression by podocytes. Wt1 mutant cells colonized glomeruli efficiently, including podocytes, but some sclerotic glomeruli contained no detectable Wt1 mutant cells. The development of GS was preceded by widespread loss of ZO-1 signal in podocytes (even in kidneys where <5% of glomeruli contained Wt1 mutant podocytes), increased intra-renal renin expression, and de novo podocyte TGF-beta1 expression, but not podocyte Pax-2 expression or loss of WT1, synaptopodin, alpha-actinin-4 or nephrin expression. However, podocytes in partially sclerotic glomeruli that still expressed WT1 at high levels showed reduced vimentin expression, cell cycle re-entry, and re-expressed desmin, cytokeratin and Pax-2. The results suggest that: (i) GS is not due to loss of WT1 expression by podocytes; (ii) podocyte Pax-2 expression reflects re-expression rather than persistent expression, and is the consequence of GS; (iii) GS is mediated systemically and the mechanism involves activation of the renin-angiotensin system; and (iv) podocytes undergo typical maturational changes but subsequently de-differentiate and revert to an immature phenotype during disease progression.

Panax ginseng exerts antiproliferative effects on rat hepatocarcinogenesis.

PubMed

Kim, Hyemee; Lee, Hae-Jeung; Kim, Dae Joong; Kim, Tae Myoung; Moon, Hyun-Seuk; Choi, Haymie

2013-09-01

It has been proposed that ginseng has chemopreventive effects against several types of cancer in animals and humans. However, the mechanisms underlying the chemopreventive activities of fresh ginseng against hepatocarcinogenesis have not yet been elucidated. Therefore, we hypothesized that these ginseng species may prevent hepatocarcinogenesis but that the chemopreventive mechanisms may differ by species. To determine the chemopreventive and therapeutic potential of 3 different types of fresh ginseng on hepatocarcinogenesis, Sprague-Dawley rats were injected with diethylnitrosamine and fed diets containing 2% Panax japonicus CA Meyer (JN), P. quinquefolius L (QQ), or P. ginseng CA Meyer (GS) for 10 weeks. Glutathione S-transferase P form (GST-P)-positive foci, a stable marker for rat hepatocarcinogenesis, were shown in all carcinogen-injected rats; but only the GS diet significantly reduced the area and number (62% and 68%, respectively; P < .05) of GST-P-positive foci compared with the diethylnitrosamine control group. In addition, the number of proliferating cell nuclear antigen-positive hepatocytes in the GST-P-positive area was significantly decreased in the GS group but not in the JN or QQ groups. Using cDNA microarray analyses to investigate the underlying molecular mechanisms, we observed that the p53 signaling pathway was altered by the GS diet and that the expression of Cyclin D1, Cyclin G1, Cdc2a, and Igf-1, which are involved in the p53 signaling pathway, was downregulated by the GS diet. Our data demonstrate, for the first time, that GS, but not JN or QQ, induces cell cycle arrest in hepatocarcinogenesis. This study suggests that fresh GS has potential chemopreventive effects and may prove to be a therapeutic agent against hepatocarcinogenesis. © 2013.

CpG methylation patterns and decitabine treatment response in acute myeloid leukemia cells and normal hematopoietic precursors

PubMed Central

Negrotto, Soledad; Ng, Kwok Peng; Jankowska, Ania M.; Bodo, Juraj; Gopalan, Banu; Guinta, Kathryn; Mulloy, James C.; Hsi, Eric; Maciejewski, Jaroslaw; Saunthararajah, Yogen

2011-01-01

The DNA hypomethylating drug decitabine maintains normal hematopoietic stem cell (HSC) self-renewal but induces terminal differentiation in acute myeloid leukemia (AML) cells. The basis for these contrasting cell-fates, and for selective CpG hypomethylation by decitabine, is poorly understood. Promoter CpGs, with methylation measured by microarray, were classified by the direction of methylation change with normal myeloid maturation. In AML cells, the methylation pattern at maturation-responsive CpG suggested at least partial maturation. Consistent with partial maturation, in gene expression analyses, AML cells expressed high levels of the key lineage-specifying factor CEBPA, but relatively low levels of the key late-differentiation driver CEBPE. In methylation analysis by mass-spectrometry, CEBPE promoter CpG that are usually hypomethylated during granulocyte maturation were significantly hypermethylated in AML cells. Decitabine treatment induced cellular differentiation of AML cells, and the largest methylation decreases were at CpG that are hypomethylated with myeloid maturation, including CEBPE promoter CpG. In contrast, decitabine-treated normal HSC retained immature morphology, and methylation significantly decreased at CpG that are less methylated in immature cells. High expression of lineage-specifying factor and aberrant epigenetic repression of some key late-differentiation genes distinguishes AML cells from normal HSC and could explain the contrasting differentiation and methylation responses to decitabine. PMID:21836612

An integrated -omics analysis of the epigenetic landscape of gene expression in human blood cells.

PubMed

Kennedy, Elizabeth M; Goehring, George N; Nichols, Michael H; Robins, Chloe; Mehta, Divya; Klengel, Torsten; Eskin, Eleazar; Smith, Alicia K; Conneely, Karen N

2018-06-19

Gene expression can be influenced by DNA methylation 1) distally, at regulatory elements such as enhancers, as well as 2) proximally, at promoters. Our current understanding of the influence of distal DNA methylation changes on gene expression patterns is incomplete. Here, we characterize genome-wide methylation and expression patterns for ~ 13 k genes to explore how DNA methylation interacts with gene expression, throughout the genome. We used a linear mixed model framework to assess the correlation of DNA methylation at ~ 400 k CpGs with gene expression changes at ~ 13 k transcripts in two independent datasets from human blood cells. Among CpGs at which methylation significantly associates with transcription (eCpGs), > 50% are distal (> 50 kb) or trans (different chromosome) to the correlated gene. Many eCpG-transcript pairs are consistent between studies and ~ 90% of neighboring eCpGs associate with the same gene, within studies. We find that enhancers (P < 5e-18) and microRNA genes (P = 9e-3) are overrepresented among trans eCpGs, and insulators and long intergenic non-coding RNAs are enriched among cis and distal eCpGs. Intragenic-eCpG-transcript correlations are negative in 60-70% of occurrences and are enriched for annotated gene promoters and enhancers (P < 0.002), highlighting the importance of intragenic regulation. Gene Ontology analysis indicates that trans eCpGs are enriched for transcription factor genes and chromatin modifiers, suggesting that some trans eCpGs represent the influence of gene networks and higher-order transcriptional control. This work sheds new light on the interplay between epigenetic changes and gene expression, and provides useful data for mining biologically-relevant results from epigenome-wide association studies.

Control of Breast Tumor Cell Growth by Dietary Indoles

DTIC Science & Technology

1997-09-01

N-nitrosodimethylamine metabolites to mouse liver macromolecules. Chemico-Biol. Interactions 48, 81-90. 5. Bailey, G.S., Hendricks , J.D., Shelton...Food Chem. Toxicol. 21, 31-36. 7. Dashwood, R.H., Arbogast, D.N., Fong, A.T., Hendricks , J.D. and Bailey, G.S. (1988) Mechanisms of... penicillin , 50 units/ml streptomycin, and 2 mM L-glutamine. MDA- MB-231 cells were grown in DMEM supplemented with 10% FBS, 50 units/ml penicillin , 50

Immunological relationships between glucosyltransferases from Streptococcus mutans serotypes.

PubMed

Kuramitsu, H; Ingersoll, L

1976-09-01

Partially purified glycosyltransferase enzymes for Streptococcus mutans GS-5 (serotype c) have been utilized to prepare antibodies directed against the soluble glucan-synthesizing activity, GTF-B, and the insoluble-soluble glucan synthetic activity, GTF-A. Anti-GTF-A inhibited insoluble glucan formation catalyzed by the extracellular enzymes from strains GS-5 and FA-1 (serotype b) to a much greater extent than that of strains HS-6 (serotype a) or OMZ-176 (serotype d). This antibody fraction also inhibited both the cell-associated glucosyltransferase activities as well as the sucrose-mediated adherence of cells to glass surfaces by strains GS-5 and FA-1 but not that of strains HS-6 and OMZ-176. Anti-GTF-B inhibited soluble glucan formation catalyzed by the extracellular enzymes of strains GS-5 but not that of strain HS-6, FA-1, or OMZ-176. However, this antibody fraction did not strongly inhibit either the cell-associated glycosyltransferase activity or cellular adherence of any of the four strains. These results with body antibody fractions were also correlated with the ability of the antibodies to agglutinate the cells and form precipitin bands after immunodiffusion with the extracellular enzymes. Antibody prepared against the homogeneous soluble glucan-synthesizing enzyme demonstrated similar effects to the anti-GTF-B fraction. These results are discussed in terms of the antigenic relationships existing between the glucosyltransferases from different serotypes of S. mutans.

Molecular characterization of gluten hydrolysing Bacillus sp. and their efficacy and biotherapeutic potential as probiotics using Caco-2 cell line.

PubMed

Rashmi, B S; Gayathri, D

2017-09-01

To isolate and characterize indigenous gluten hydrolysing bacteria from wheat sourdough and curd samples and further evaluation of their probiotic potentiality. Indigenous gluten hydrolysing isolates GS 33, GS 143, GS 181 and GS 188 were identified as Bacillus sp. by molecular-typing methods and studied extensively for their functional and probiotic attributes. All the tested isolates could survive at pH 2 and toxicity of 0·3% bile and also exhibited cell surface hydrophobicity and autoaggregation phenotype. The isolates were adhered strongly to Caco-2 cells and coaggregated with Escherichia coli MTCC 433 and Listeria monocytogenes MTCC 1143 preventing pathogen invasion into Caco-2 cells in vitro. In addition, the minimum inhibitory concentration of selected antibiotics for all the investigated gluten hydrolysing isolates was within the breakpoint values as recommended by the European Food Safety Authority. The indigenous high intensity gluten hydrolysing bacteria exhibited high resistance to gastric and pancreatic stress and possessed antibacterial, aggregation, adhesion and pathogen exclusion properties, and as a potential probiotics, either alone or in consortium would be useful in the development of gluten-free wheat foods. Exploring new indigenous gluten hydrolysing bacteria from wheat sourdough and curd samples would be beneficial in developing gluten-free wheat foods using potential indigenous probiotics. © 2017 The Society for Applied Microbiology.

Medulloblastoma in a toddler with Gorlin syndrome.

PubMed

Al-Rahawan, Mohamad G; Trevino, Sorleen; Jacob, Roy; Murray, Jeffrey C; Al-Rahawan, Mohamad M

2018-04-01

Gorlin syndrome (GS) is a rare hereditary multisystem disorder caused by mutations in PTCH1, PTCH2 , or SUFU . It is characterized by multiple anomalies and an increased risk of developing various tumors. Basal cell carcinoma is most common, and medulloblastoma (MB) is especially frequent in patients with SUFU mutations. MB treatment often includes radiation therapy in patients older than 3 years; however, such treatment is very toxic to patients with GS. Most reported cases of MB in patients with GS present after GS is diagnosed. We report a male toddler with multicentric posterior fossa tumor and calcifications along the falx cerebri, suggesting MB and GS. Pathology revealed nodular MB. His testing confirmed a germline SUFU mutation. His tumor resolved with three induction cycles of chemotherapy, but he died of respiratory failure due to infection at 20 months of age. Overlooking calcifications along the falx cerebri in children with MB can induce significant morbidity.

Passively Administered Pooled Human Immunoglobulins Exert IL-10 Dependent Anti-Inflammatory Effects that Protect against Fatal HSV Encephalitis

PubMed Central

Ramakrishna, Chandran; Newo, Alain N. S.; Shen, Yueh-Wei; Cantin, Edouard

2011-01-01

HSV-1 is the leading cause of sporadic encephalitis in humans. HSV infection of susceptible 129S6 mice results in fatal encephalitis (HSE) caused by massive inflammatory brainstem lesions comprising monocytes and neutrophils. During infection with pathogenic microorganisms or autoimmune disease, IgGs induce proinflammatory responses and recruit innate effector cells. In contrast, high dose intravenous immunoglobulins (IVIG) are an effective treatment for various autoimmune and inflammatory diseases because of potent anti-inflammatory effects stemming in part from sialylated IgGs (sIgG) present at 1–3% in IVIG. We investigated the ability of IVIG to prevent fatal HSE when given 24 h post infection. We discovered a novel anti-inflammatory pathway mediated by low-dose IVIG that protected 129S6 mice from fatal HSE by modulating CNS inflammation independently of HSV specific antibodies or sIgG. IVIG suppressed CNS infiltration by pathogenic CD11b+ Ly6Chigh monocytes and inhibited their spontaneous degranulation in vitro. FcγRIIb expression was required for IVIG mediated suppression of CNS infiltration by CD45+ Ly6Clow monocytes but not for inhibiting development of Ly6Chigh monocytes. IVIG increased accumulation of T cells in the CNS, and the non-sIgG fraction induced a dramatic expansion of FoxP3+ CD4+ T regulatory cells (Tregs) and FoxP3− ICOS+ CD4+ T cells in peripheral lymphoid organs. Tregs purified from HSV infected IVIG treated, but not control, mice protected adoptively transferred mice from fatal HSE. IL-10, produced by the ICOS+ CD4+ T cells that accumulated in the CNS of IVIG treated, but not control mice, was essential for induction of protective anti-inflammatory responses. Our results significantly enhance understanding of IVIG's anti-inflammatory and immunomodulatory capabilities by revealing a novel sIgG independent anti-inflammatory pathway responsible for induction of regulatory T cells that secrete the immunosuppressive cytokine IL-10 and further reveal the therapeutic potential of IVIG for treating viral induced inflammatory diseases. PMID:21655109

The Molecular Basis of TnrA Control by Glutamine Synthetase in Bacillus subtilis*

PubMed Central

Hauf, Ksenia; Kayumov, Airat; Gloge, Felix; Forchhammer, Karl

2016-01-01

TnrA is a master regulator of nitrogen assimilation in Bacillus subtilis. This study focuses on the mechanism of how glutamine synthetase (GS) inhibits TnrA function in response to key metabolites ATP, AMP, glutamine, and glutamate. We suggest a model of two mutually exclusive GS conformations governing the interaction with TnrA. In the ATP-bound state (A-state), GS is catalytically active but unable to interact with TnrA. This conformation was stabilized by phosphorylated l-methionine sulfoximine (MSX), fixing the enzyme in the transition state. When occupied by glutamine (or its analogue MSX), GS resides in a conformation that has high affinity for TnrA (Q-state). The A- and Q-state are mutually exclusive, and in agreement, ATP and glutamine bind to GS in a competitive manner. At elevated concentrations of glutamine, ATP is no longer able to bind GS and to bring it into the A-state. AMP efficiently competes with ATP and prevents formation of the A-state, thereby favoring GS-TnrA interaction. Surface plasmon resonance analysis shows that TnrA bound to a positively regulated promoter fragment binds GS in the Q-state, whereas it rapidly dissociates from a negatively regulated promoter fragment. These data imply that GS controls TnrA activity at positively controlled promoters by shielding the transcription factor in the DNA-bound state. According to size exclusion and multiangle light scattering analysis, the dodecameric GS can bind three TnrA dimers. The highly interdependent ligand binding properties of GS reveal this enzyme as a sophisticated sensor of the nitrogen and energy state of the cell to control the activity of DNA-bound TnrA. PMID:26635369

Fertilization intensifies drought stress: water use and stomatal conductance of Pinus taeda in a midrotation fertilization and throughfall reduction experiment

Treesearch

Eric J. Ward; Jean-Christophe Domec; Marshall A. Laviner; Thomas R. Fox; Ge Sun; Steve McNulty; John King; Asko Noormets

2015-01-01

While mid-rotation fertilization increases productivity in many southern pine forests, it remains unclear what impact such management may have on stand water use. We examined the impact of nutrient and water availability on stem volume, leaf area, transpiration per unit ground area (EC) and canopy conductance per unit leaf area (GS...

GsCBRLK, a calcium/calmodulin-binding receptor-like kinase, is a positive regulator of plant tolerance to salt and ABA stress.

PubMed

Yang, Liang; Ji, Wei; Zhu, Yanming; Gao, Peng; Li, Yong; Cai, Hua; Bai, Xi; Guo, Dianjing

2010-05-01

Calcium/calmodulin-dependent kinases play vital roles in protein phosphorylation in eukaryotes, yet little is known about the phosphorylation process of calcium/calmodulin-dependent protein kinase and its role in stress signal transduction in plants. A novel plant-specific calcium-dependent calmodulin-binding receptor-like kinase (GsCBRLK) has been isolated from Glycine soja. A subcellular localization study using GFP fusion protein indicated that GsCBRLK is localized in the plasma membrane. Binding assays demonstrated that calmodulin binds to GsCBRLK with an affinity of 25.9 nM in a calcium-dependent manner and the binding motif lies between amino acids 147 to169 within subdomain II of the kinase domain. GsCBRLK undergoes autophosphorylation and Myelin Basis Protein phosphorylation in the presence of calcium. It was also found that calcium/calmodulin positively regulates GsCBRLK kinase activity through direct interaction between the calmodulin-binding domain and calmodulin. So, it is likely that GsCBRLK responds to an environmental stimulus in two ways: by increasing the protein expression level and by regulating its kinase activity through the calcium/calmodulin complex. Furthermore, cold, salinity, drought, and ABA stress induce GsCBRLK gene transcripts. Over-expression of GsCBRLK in transgenic Arabidopsis resulted in enhanced plant tolerance to high salinity and ABA and increased the expression pattern of a number of stress gene markers in response to ABA and high salt. These results identify GsCBRLK as a molecular link between the stress- and ABA-induced calcium/calmodulin signal and gene expression in plant cells.

The mechanisms of detoxification of As(III), dimethylarsinic acid (DMA) and As(V) in the microalga Chlorella vulgaris.

PubMed

Pantoja Munoz, L; Purchase, D; Jones, H; Raab, A; Urgast, D; Feldmann, J; Garelick, H

2016-06-01

The response of Chlorella vulgaris when challenged by As(III), As(V) and dimethylarsinic acid (DMA) was assessed through experiments on adsorption, efflux and speciation of arsenic (reduction, oxidation, methylation and chelation with glutathione/phytochelatin [GSH/PC]). Our study indicates that at high concentrations of phosphate (1.62mM of HPO4(2-)), upon exposure to As(V), cells are able to shift towards methylation of As(V) rather than PC formation. Treatment with As(V) caused a moderate decrease in intracellular pH and a strong increase in the concentration of free thiols (GSH). Passive surface adsorption was found to be negligible for living cells exposed to DMA and As(V). However, adsorption of As(III) was observed to be an active process in C. vulgaris, because it did not show saturation at any of the exposure periods. Chelation of As(III) with GS/PC and to a lesser extent hGS/hPC is a major detoxification mechanism employed by C. vulgaris cells when exposed to As(III). The increase of bound As-GS/PC complexes was found to be strongly related to an increase in concentration of As(III) in media. C. vulgaris cells did not produce any As-GS/PC complex when exposed to As(V). This may indicate that a reduction step is needed for As(V) complexation with GSH/PC. C. vulgaris cells formed DMAS(V)-GS upon exposure to DMA independent of the exposure period. As(III) triggers the formation of arsenic complexes with PC and homophytochelatins (hPC) and their compartmentalisation to vacuoles. A conceptual model was devised to explain the mechanisms involving ABCC1/2 transport. The potential of C. vulgaris to bio-remediate arsenic from water appeared to be highly selective and effective without the potential hazard of reducing As(V) to As(III), which is more toxic to humans. Copyright © 2016 Elsevier B.V. All rights reserved.

Heterocyclic cationic gemini surfactants: a comparative overview of their synthesis, self-assembling, physicochemical, and biological properties.

PubMed

Sharma, Vishnu Dutt; Ilies, Marc A

2014-01-01

Gemini surfactants (GS) are presently receiving substantial attention due to their special self-assembling properties and unique interfacial activity. This comprehensive review is focused on positively charged heterocyclic GS, presenting their major synthetic access routes and examining the impact of structural elements on physicochemical and aggregation properties of this class of amphiphiles. Interaction of geminis surfactants with cells and their biological properties as novel transfection agents are emphasized through a detailed structure-activity relationship analysis. Throughout the review we have also presented the properties of selected ammonium GS, simple surfactants and lipid congeners, in order to emphasize the advantages conferred by using heterocyclic polar heads in GS design. © 2012 Wiley Periodicals, Inc.

Stochastic nanoroughness modulates neuron-astrocyte interactions and function via mechanosensing cation channels.

PubMed

Blumenthal, Nils R; Hermanson, Ola; Heimrich, Bernd; Shastri, V Prasad

2014-11-11

Extracellular soluble signals are known to play a critical role in maintaining neuronal function and homeostasis in the CNS. However, the CNS is also composed of extracellular matrix macromolecules and glia support cells, and the contribution of the physical attributes of these components in maintenance and regulation of neuronal function is not well understood. Because these components possess well-defined topography, we theorize a role for topography in neuronal development and we demonstrate that survival and function of hippocampal neurons and differentiation of telencephalic neural stem cells is modulated by nanoroughness. At roughnesses corresponding to that of healthy astrocytes, hippocampal neurons dissociated and survived independent from astrocytes and showed superior functional traits (increased polarity and calcium flux). Furthermore, telencephalic neural stem cells differentiated into neurons even under exogenous signals that favor astrocytic differentiation. The decoupling of neurons from astrocytes seemed to be triggered by changes to astrocyte apical-surface topography in response to nanoroughness. Blocking signaling through mechanosensing cation channels using GsMTx4 negated the ability of neurons to sense the nanoroughness and promoted decoupling of neurons from astrocytes, thus providing direct evidence for the role of nanotopography in neuron-astrocyte interactions. We extrapolate the role of topography to neurodegenerative conditions and show that regions of amyloid plaque buildup in brain tissue of Alzheimer's patients are accompanied by detrimental changes in tissue roughness. These findings suggest a role for astrocyte and ECM-induced topographical changes in neuronal pathologies and provide new insights for developing therapeutic targets and engineering of neural biomaterials.

Half molecular exchange of IgGs in the blood of healthy humans: chimeric lambda-kappa-immunoglobulins containing HL fragments of antibodies of different subclasses (IgG1-IgG4).

PubMed

Sedykh, Sergey E; Lekchnov, Evgenii A; Prince, Viktor V; Buneva, Valentina N; Nevinsky, Georgy A

2016-10-20

In the classic paradigm, immunoglobulins represent products of clonal B cell populations, each producing antibodies recognizing a single antigen (monospecific). There is a common belief that IgGs in mammalian biological fluids are monospecific molecules having stable structures and two identical antigen-binding sites. But the issue concerning the possibility of exchange by HL-fragments between the antibody molecules in human blood is still unexplored. Different physico-chemical and immunological methods for analysis of half-molecule exchange between human blood IgGs were used. Using eighteen blood samples of healthy humans we have shown unexpected results for the first time: blood antibodies undergo extensive post-transcriptional half-molecule exchange and IgG pools on average consist of 62.4 ± 6.5% IgGs containing kappa light chains (kappa-kappa-IgGs), 29.8.6 ± 5.4% lambda light chains (lambda-lambda-IgGs), and 8.8 ± 2.7% (range 2.6-16.8%) IgGs containing both kappa- and lambda-light chains. Kappa-kappa-IgGs and lambda-lambda-IgGs contained on average (%): IgG1 (36.0 and 32.3), IgG2 (50.9 and 51.4), IgG3 (9.7 and 9.9), and IgG4 (6.5 and 5.7), while chimeric kappa-lambda-IgGs consisted of (%): 25.5 ± 4.2 IgG1, 50.8 ± 3.9 IgG2, 9.1 ± 2.1 IgG3, and 14.5 ± 2.2 IgG4. Our unexpected data are indicative of the possibility of half-molecule exchange between blood IgGs of various subclasses, raised against different antigens. The existence of blood chimeric bifunctional IgGs with different binding sites destroys the classic paradigm. Due to the phenomenon of polyspecificity and cross-reactivity of bifunctional IgGs containing HL-fragments of different types to different antigens, such IgGs may be important in human blood for widening their different biological functions.

Epigenetic Biomarker to Support Classification into Pluripotent and Non-Pluripotent Cells

NASA Astrophysics Data System (ADS)

Lenz, Michael; Goetzke, Roman; Schenk, Arne; Schubert, Claudia; Veeck, Jürgen; Hemeda, Hatim; Koschmieder, Steffen; Zenke, Martin; Schuppert, Andreas; Wagner, Wolfgang

2015-03-01

Quality control of human induced pluripotent stem cells (iPSCs) can be performed by several methods. These methods are usually relatively labor-intensive, difficult to standardize, or they do not facilitate reliable quantification. Here, we describe a biomarker to distinguish between pluripotent and non-pluripotent cells based on DNA methylation (DNAm) levels at only three specific CpG sites. Two of these CpG sites were selected by their discriminatory power in 258 DNAm profiles - they were either methylated in pluripotent or non-pluripotent cells. The difference between these two β-values provides an Epi-Pluri-Score that was validated on independent DNAm-datasets (264 pluripotent and 1,951 non-pluripotent samples) with 99.9% specificity and 98.9% sensitivity. This score was complemented by a third CpG within the gene POU5F1 (OCT4), which better demarcates early differentiation events. We established pyrosequencing assays for the three relevant CpG sites and thereby correctly classified DNA of 12 pluripotent cell lines and 31 non-pluripotent cell lines. Furthermore, DNAm changes at these three CpGs were tracked in the course of differentiation of iPSCs towards mesenchymal stromal cells. The Epi-Pluri-Score does not give information on lineage-specific differentiation potential, but it provides a simple, reliable, and robust biomarker to support high-throughput classification into either pluripotent or non-pluripotent cells.

Food antigen-induced immune responses in Crohn's disease patients and experimental colitis mice.

PubMed

Kawaguchi, Takaaki; Mori, Maiko; Saito, Keiko; Suga, Yasuyo; Hashimoto, Masaki; Sako, Minako; Yoshimura, Naoki; Uo, Michihide; Danjo, Keiko; Ikenoue, Yuka; Oomura, Kaori; Shinozaki, Junko; Mitsui, Akira; Kajiura, Takayuki; Suzuki, Manabu; Takazoe, Masakazu

2015-04-01

In Crohn's disease (CD), the involvement of food antigens in immune responses remains unclear. The objective of this study was to detect immune responses against food antigens in CD patients and examine the mechanism in a mouse model of colitis. We enrolled 98 CD patients, 50 ulcerative colitis patients, and 52 healthy controls (HCs) to compare the levels of serum immunoglobulin (Ig)Gs against 88 foods. The presence of serum IgGs against foods was also examined in interleukin (IL)-10 knockout (KO) mice in which CD4(+) T cell activation by antigenic food protein was assessed. Mice transferred with IL-10 KO cells received diets with or without food antigens, and the development of colitis was evaluated. The prevalence of IgGs against various foods, especially vegetables, grains, and nuts, was significantly higher in CD patients than in HCs. Similarly, the prevalence of IgGs against food proteins was higher in IL-10 KO mice than in BALB/c mice. Beta-conglycinin, identified as an antigenic food proteins in IL-10 KO mice, induced CD4(+) T cell production of interferon-γ and IL-17 through dendritic cell antigen presentation. Elimination of the food antigens ameliorated the development of colitis in mice without altering the composition of their intestinal microbiota. In CD colitis mice, intestinal inflammation via CD4(+) T cell hyperactivation was induced by food antigens associated with high serum IgG levels and was ameliorated by the elimination of food antigens. This disrupted immunological tolerance to food antigen, which might act as an exacerbating factor, remains to be elucidated in CD patients.

Preparation and characterization of gellan gum/glucosamine/clioquinol film as oral cancer treatment patch.

PubMed

Tsai, Wanchi; Tsai, Huifang; Wong, Yinuan; Hong, Juiyen; Chang, Shwujen; Lee, Mingwei

2018-01-01

To administer cancer drugs with improved convenience to patients and to enhance the bioavailability of cancer drugs for oral cancer therapy, this study prepared gellan gum/glucosamine/clioquinol (GG/GS/CQ) film as the oral cancer treatment patch. GG/GS/CQ film fabricated through the EDC-mediated coupling reactions (GG/GS/CQ/EDC film). The film of the physicochemical properties and drug release kinetics were studied. The effectiveness of GG/GS/CQ/EDC film as oral cancer treatment patch were evaluated with the animal model. The results confirmed that CQ can be incorporated via EDC-mediated covalent conjugation to gellan gum/glucosamine. Mechanical testing revealed that the maximum tensile strength and elongation percentage at break were 1.91kgf/mm 2 and 5.01% for GG/GS/CQ/EDC film. After a drug release experiment lasting 45days, 86.8% of CQ was released from GG/GS/CQ/EDC film. The Huguchi model fit the GG/GS/CQ/EDC drug release data with high correlation coefficients (R 2 =0.9994, respectively). The effect of the CQ dose on oral cancer cells (OC-2) was tested, and the IC 50 of CQ alone and CQ with 10μM CuCl 2 were 9.59 and 2.22μM, respectively. The animal testing indicated that GG/GS/CQ/EDC film was decreased epidermal growth factor receptor (EGFR) expression and suppress tumor progression. These findings provide insights into a possible use for GG/GS/CQ/EDC film for oral ca in clinical practice. The GG/GS/CQ/EDC film is suitable as the dressing for use in the treatment of early-stage cancer or as wound care after surgery in late-stage of oral cancer treatment. Copyright © 2017 Elsevier B.V. All rights reserved.

Separation and identification of selenotrisulfides in epithelial cell homogenates by LC-ICP-MS and LC-ESI-MS after incubation with selenite.

PubMed

Gabel-Jensen, Charlotte; Gammelgaard, Bente; Bendahl, Lars; Stürup, Stefan; Jøns, Ole

2006-02-01

To elucidate how selenite is metabolised in the intestine after oral intake, it was incubated with homogenized epithelial cells from pigs. When the metabolites were analysed by LC-ICP-MS, two major selenium metabolites were separated in the supernatant from the homogenate. These metabolites were formed instantly but disappeared within 15 min. No other selenium-containing compounds appeared during this time. Hence, the secondary reaction products were either volatilised or precipitated. To verify the identity of the compounds, a larger amount of selenite was incubated with epithelial cells. The presence of Cys-Se-SG and GS-Se-SG was verified by LC-ESI-MS. Selenotrisulfides were synthesized by reaction of L-cysteine and L-glutathione with sodium selenite. The reaction mixture contained three main products: selenodicysteine (Cys-Se-Cys), selenocysteine glutathione (Cys-Se-SG), and selenodiglutathione (GS-Se-SG). The two transient selenium compounds in the epithelial cell incubation mixture co-eluted with the synthesized Cys-Se-SG and GS-Se-SG, respectively. The identities of these compounds were verified by LC-ESI-MS. Hence, these selenium metabolites have now been identified by ESI-MS after isolation from epithelial cells.

Measurement of leaf hydraulic conductance and stomatal conductance and their responses to irradiance and dehydration using the Evaporative Flux Method (EFM).

PubMed

Sack, Lawren; Scoffoni, Christine

2012-12-31

Water is a key resource, and the plant water transport system sets limits on maximum growth and drought tolerance. When plants open their stomata to achieve a high stomatal conductance (gs) to capture CO2 for photosynthesis, water is lost by transpiration(1,2). Water evaporating from the airspaces is replaced from cell walls, in turn drawing water from the xylem of leaf veins, in turn drawing from xylem in the stems and roots. As water is pulled through the system, it experiences hydraulic resistance, creating tension throughout the system and a low leaf water potential (Ψ(leaf)). The leaf itself is a critical bottleneck in the whole plant system, accounting for on average 30% of the plant hydraulic resistance(3). Leaf hydraulic conductance (K(leaf) = 1/ leaf hydraulic resistance) is the ratio of the water flow rate to the water potential gradient across the leaf, and summarizes the behavior of a complex system: water moves through the petiole and through several orders of veins, exits into the bundle sheath and passes through or around mesophyll cells before evaporating into the airspace and being transpired from the stomata. K(leaf) is of strong interest as an important physiological trait to compare species, quantifying the effectiveness of the leaf structure and physiology for water transport, and a key variable to investigate for its relationship to variation in structure (e.g., in leaf venation architecture) and its impacts on photosynthetic gas exchange. Further, K(leaf) responds strongly to the internal and external leaf environment(3). K(leaf) can increase dramatically with irradiance apparently due to changes in the expression and activation of aquaporins, the proteins involved in water transport through membranes(4), and K(leaf) declines strongly during drought, due to cavitation and/or collapse of xylem conduits, and/or loss of permeability in the extra-xylem tissues due to mesophyll and bundle sheath cell shrinkage or aquaporin deactivation(5-10). Because K(leaf) can constrain gs and photosynthetic rate across species in well watered conditions and during drought, and thus limit whole-plant performance they may possibly determine species distributions especially as droughts increase in frequency and severity(11-14). We present a simple method for simultaneous determination of K(leaf) and gs on excised leaves. A transpiring leaf is connected by its petiole to tubing running to a water source on a balance. The loss of water from the balance is recorded to calculate the flow rate through the leaf. When steady state transpiration (E, mmol • m(-2) • s(-1)) is reached, gs is determined by dividing by vapor pressure deficit, and K(leaf) by dividing by the water potential driving force determined using a pressure chamber (K(leaf)= E /- Δψ(leaf), MPa)(15). This method can be used to assess K(leaf) responses to different irradiances and the vulnerability of K(leaf) to dehydration(14,16,17).

Fourier transform infrared spectroscopic study of the interactions of a strongly antimicrobial but weakly hemolytic analogue of gramicidin S with lipid micelles and lipid bilayer membranes.

PubMed

Lewis, Ruthven N A H; Kiricsi, Monika; Prenner, Elmar J; Hodges, Robert S; McElhaney, Ronald N

2003-01-21

Cyclo[VKLdKVdYPLKVKLdYP] (GS14dK(4)), a synthetic tetradecameric ring-size analogue of the naturally occurring antimicrobial peptide gramicidin S (GS), retains the strong antimicrobial activity of GS but is 15-20 times less hemolytic. To characterize its interaction with lipid membranes and to understand the molecular basis of its capacity to lyse bacterial cells, in preference to erythrocytes, we have investigated the interactions of GS14dK(4) with detergent micelles and with lipid bilayer model membranes by Fourier transform infrared spectroscopy and compared our results with those of a similar study of GS [Lewis, R. N. A. H., et al. (1999) Biochemistry 38, 15193-15203]. In both aqueous and organic solvent solutions, GS14dK(4) adopts a beta-sheet conformation that is somewhat distorted and more sensitive to the polarity of its environment than GS. Like GS, GS14dK(4) is completely or partially excluded from gel-state lipid bilayers but interacts strongly with liquid-crystalline lipid bilayers and detergent micelle, and interacts more strongly with more fluid liquid-crystalline lipid systems. However, its interactions are more strongly influenced by membrane lipid order and fluidity, and unlike GS, it is essentially excluded from cholesterol-containing phospholipid bilayers. Also, GS14dK(4) is excluded from cationic lipid bilayers, but partitions more strongly and/or penetrates more deeply into anionic lipid bilayers than into those composed of either zwitterionic or nonionic lipids. Anionic lipids also facilitate GS14dK(4) interactions with multicomponent lipid bilayers which are predominantly zwitterionic or nonionic. Although GS14dK(4) generally penetrates and/or partitions into zwitterionic or uncharged lipid bilayers less strongly than does GS, its greater size and altered distribution of positive charges make it intrinsically more perturbing with regard to membrane organization once associated with lipid bilayers. This fact, combined with its relatively strong interactions with anionic phospholipids, may explain why GS14dK(4) retains relatively high antimicrobial activity. However, its low hemolytic activity is probably largely attributable to its low propensity to penetrate and/or partition into cholesterol-containing zwitterionic lipid membranes.

Inhibition of Enterococcus faecium adherence to collagen by antibodies against high-affinity binding subdomains of Acm.

PubMed

Nallapareddy, Sreedhar R; Sillanpää, Jouko; Ganesh, Vannakambadi K; Höök, Magnus; Murray, Barbara E

2007-06-01

Strains of Enterococcus faecium express a cell wall-anchored protein, Acm, which mediates adherence to collagen. Here, we (i) identify the minimal and high-affinity binding subsegments of Acm and (ii) show that anti-Acm immunoglobulin Gs (IgGs) purified against these subsegments reduced E. faecium TX2535 strain collagen adherence up to 73 and 50%, respectively, significantly more than the total IgGs against the full-length Acm A domain (28%) (P < 0.0001). Blocking Acm adherence with functional subsegment-specific antibodies raises the possibility of their use as therapeutic or prophylactic agents.

Detection of Cyclooxygenase-2-Derived Oxygenation Products of the Endogenous Cannabinoid 2-Arachidonoylglycerol in Mouse Brain.

PubMed

Morgan, Amanda J; Kingsley, Philip J; Mitchener, Michelle M; Altemus, Megan; Patrick, Toni A; Gaulden, Andrew D; Marnett, Lawrence J; Patel, Sachin

2018-05-09

Cyclooxygenase-2 (COX-2) catalyzes the formation of prostaglandins, which are involved in immune regulation, vascular function, and synaptic signaling. COX-2 also inactivates the endogenous cannabinoid (eCB) 2-arachidonoylglycerol (2-AG) via oxygenation of its arachidonic acid backbone to form a variety of prostaglandin glyceryl esters (PG-Gs). Although this oxygenation reaction is readily observed in vitro and in intact cells, detection of COX-2-derived 2-AG oxygenation products has not been previously reported in neuronal tissue. Here we show that 2-AG is metabolized in the brain of transgenic COX-2-overexpressing mice and mice treated with lipopolysaccharide to form multiple species of PG-Gs that are detectable only when monoacylglycerol lipase is concomitantly blocked. Formation of these PG-Gs is prevented by acute pharmacological inhibition of COX-2. These data provide evidence that neuronal COX-2 is capable of oxygenating 2-AG to form a variety PG-Gs in vivo and support further investigation of the physiological functions of PG-Gs.

A crop loss-related forecasting model for sclerotinia stem rot in winter oilseed rape.

PubMed

Koch, S; Dunker, S; Kleinhenz, B; Röhrig, M; Tiedemann, A von

2007-09-01

Sclerotinia stem rot (SSR) is an increasing threat to winter oilseed rape (OSR) in Germany and other European countries due to the growing area of OSR cultivation. A forecasting model was developed to provide decision support for the fungicide spray against SSR at flowering. Four weather variables-air temperature, relative humidity, rainfall, and sunshine duration-were used to calculate the microclimate in the plant canopy. From data reinvestigated in a climate chamber study, 7 to 11 degrees C and 80 to 86% relative humidity (RH) were established as minimum conditions for stem infection with ascospores and expressed as an index to discriminate infection hours (Inh). Disease incidence (DI) significantly correlated with Inh occurring post-growth stage (GS) 58 (late bud stage) (r(2) = 0.42, P /= Inh(i). Historical field data (1994 to 2004) were used to assess the impact of agronomic factors on SSR incidence. A 2-year crop rotation enhanced disease risk and, therefore, lowered the infection threshold in the model by a factor of 0.8, whereas in 4-year rotations, the threshold was elevated by a factor 1.3. Number of plants per square meter, nitrogen fertilization, and soil management did not have significant effects on DI. In an evaluation of SkleroPro with 76 historical (1994 to 2004) and 32 actual field experiments conducted in 2005, the percentage of economically correct decisions was 70 and 81%, respectively. Compared with the common practice of routine sprays, this corresponded to savings in fungicides of 39 and 81% and to increases in net return for the grower of 23 and 45 euro/ha, respectively. This study demonstrates that, particularly in areas with abundant inoculum, the level of SSR in OSR can be predicted from conditions of stem infection during late bud or flowering with sufficient accuracy, and does not require simulation of apothecial development and ascospore dispersal. SkleroPro is the first crop-loss-related forecasting model for a Sclerotinia disease, with the potential of being widely used in agricultural practice, accessible through the Internet. Its concept, components, and implementation may be useful in developing forecasting systems for Sclerotinia diseases in other crops or climates.

Nonsteroidal anti-inflammatory drugs (NSAID) sparing effects of glucosamine hydrochloride through N-glycosylation inhibition; strategy to rescue stomach from NSAID damage.

PubMed

Park, S H; Hong, H; Han, Y M; Kangwan, N; Kim, S J; Kim, E H; Hahm, K B

2013-04-01

Gastrointestinal or cardiovascular complications limit nonsteroidal anti-inflammatory drugs (NSAID) prescription. Glucosamine hydrochloride (GS-HCl) alternatively chosen, but debates still exist in its clinical efficiency. COX-2 instability through inhibiting COX-2 N-glycosylation of GS-HCl raised the possibility of NSAID sparing effect. Study was done to determine whether combination treatment of glucosamine and NSAID contributes to gastric safety through NSAID sparing effect. IEC-6 cells were stimulated with TNF-α and compared the expressions of inflammatory mediators after indomethacin alone or combination of indomethacin and GS-HCl by Western blotting and RT-PCR. C57BL/6 mice injected with type II collagen to induce arthritis were treated with indomethacin alone or combination of reduced dose of indomethacin and GS-HCl after 3 weeks. TNF-α increased the expression of COX-2, iNOS and inflammatory cytokines, but GS-HCl significantly attenuated TNF-α-induced COX-2 expression. Decreased COX-2 after GS-HCl was caused by N-glycosylation inhibition as much as tunicamycin. Combination of reduced dose of indomethacin and GS-HCl significantly reduced the expressions of ICAM-1, VCAM-1, IL-8, IL-1β, MMP-2, MMP-7, MMP-9, and MMP-11 mRNA as well as NF-κB activation better than high dose indomethacin alone. These NSAID sparing effect of GS-HCl was further proven in collagen-induced arthritis model. Combination of GS-HCl and 2.5 mg/kg indomethacin showed significant protection from gastric damages as well as efficacious anti-arthritic effect. Taken together, COX-2 N-glycosylation inhibition by GS-HCl led to indomethacin sparing effects, based on which combination of GS-HCl and reduced dose of NSAID can provide the strategy to secure stomach from NSAID-induced gastric damage as well as excellent anti-arthritic effects.

A truncated form of CD9-partner 1 (CD9P-1), GS-168AT2, potently inhibits in vivo tumour-induced angiogenesis and tumour growth

PubMed Central

Colin, S; Guilmain, W; Creoff, E; Schneider, C; Steverlynck, C; Bongaerts, M; Legrand, E; Vannier, J P; Muraine, M; Vasse, M; Al-Mahmood, S

2011-01-01

Background: Tetraspanins are transmembrane proteins known to contribute to angiogenesis. CD9 partner-1 (CD9P-1/EWI-F), a glycosylated type 1 transmembrane immunoglobulin, is a member of the tetraspanin web, but its role in angiogenesis remains to be elucidated. Methods: We measured the expression of CD9P-1 under angiogenic and angiostatic conditions, and the influence of its knockdown onto capillary structures formation by human endothelial cells (hECs). A truncated form of CDP-1, GS-168AT2, was produced and challenged vs hEC proliferation, migration and capillaries' formation. Its association with CD9P-1, CD9, CD81 and CD151 and the expressions of these later at hEC surface were analysed. Finally, its effects onto in vivo tumour-induced angiogenesis and tumour growth were investigated. Results: Vascular endothelial growth factor (VEGF)-induced capillary tube-like formation was inhibited by tumour necrosis factor α and was associated with a rise in CD9P-1 mRNA expression (P<0.05); accordingly, knockdown of CD9P-1 inhibited VEGF-dependent in vitro angiogenesis. GS-168AT2 dose-dependently inhibited in vitro angiogenesis, hEC migration and proliferation (P<0.05). Co-precipitation experiments suggest that GS-168AT2 corresponds to the sequence by which CD9P-1 physiologically associates with CD81. GS-168AT2 induced the depletion of CD151, CD9 and CD9P-1 from hEC surface, correlating with GS-168AT2 degradation. Finally, in vivo injections of GS-168AT2 inhibited tumour-associated angiogenesis by 53.4±9.5% (P=0.03), and reduced tumour growth of Calu 6 tumour xenografts by 73.9±16.4% (P=0.007) without bodyweight loss. Conclusion: The truncated form of CD9P-1, GS-168AT2, potently inhibits angiogenesis and cell migration by at least the downregulation of CD151 and CD9, which provides the first evidences for the central role of CD9P-1 in tumour-associated angiogenesis and tumour growth. PMID:21863033

Functioning and nonfunctioning thyroid adenomas involve different molecular pathogenetic mechanisms.

PubMed

Tonacchera, M; Vitti, P; Agretti, P; Ceccarini, G; Perri, A; Cavaliere, R; Mazzi, B; Naccarato, A G; Viacava, P; Miccoli, P; Pinchera, A; Chiovato, L

1999-11-01

The molecular biology of follicular cell growth in thyroid nodules is still poorly understood. Because gain-of-function (activating) mutations of the thyroid-stimulating hormone receptor (TShR) and/or Gs alpha genes may confer TSh-independent growth advantage to neoplastic thyroid cells, we searched for somatic mutations of these genes in a series of hyperfunctioning and nonfunctioning follicular thyroid adenomas specifically selected for their homogeneous gross anatomy (single nodule in an otherwise normal thyroid gland). TShR gene mutations were identified by direct sequencing of exons 9 and 10 of the TShR gene in genomic DNA obtained from surgical specimens. Codons 201 and 227 of the Gs alpha gene were also analyzed. At histology, all hyperfunctioning nodules and 13 of 15 nonfunctioning nodules were diagnosed as follicular adenomas. Two nonfunctioning thyroid nodules, although showing a prevalent microfollicular pattern of growth, had histological features indicating malignant transformation (a minimally invasive follicular carcinoma and a focal papillary carcinoma). Activating mutations of the TShR gene were found in 12 of 15 hyperfunctioning follicular thyroid adenomas. In one hyperfunctioning adenoma, which was negative for TShR mutations, a mutation in codon 227 of the Gs alpha gene was identified. At variance with hyperfunctioning thyroid adenomas, no mutation of the TShR or Gs alpha genes was detected in nonfunctioning thyroid nodules. In conclusion, our findings clearly define a different molecular pathogenetic mechanism in hyperfunctioning and nonfunctioning follicular thyroid adenomas. Activation of the cAMP cascade, which leads to proliferation but maintains differentiation of follicular thyroid cells, typically occurs in hyperfunctioning thyroid adenomas. Oncogenes other than the TShR and Gs alpha genes are probably involved in nonfunctioning follicular adenomas.

A Restricted Repertoire of De Novo Mutations in ITPR1 Cause Gillespie Syndrome with Evidence for Dominant-Negative Effect

PubMed Central

McEntagart, Meriel; Williamson, Kathleen A.; Rainger, Jacqueline K.; Wheeler, Ann; Seawright, Anne; De Baere, Elfride; Verdin, Hannah; Bergendahl, L. Therese; Quigley, Alan; Rainger, Joe; Dixit, Abhijit; Sarkar, Ajoy; López Laso, Eduardo; Sanchez-Carpintero, Rocio; Barrio, Jesus; Bitoun, Pierre; Prescott, Trine; Riise, Ruth; McKee, Shane; Cook, Jackie; McKie, Lisa; Ceulemans, Berten; Meire, Françoise; Temple, I. Karen; Prieur, Fabienne; Williams, Jonathan; Clouston, Penny; Németh, Andrea H.; Banka, Siddharth; Bengani, Hemant; Handley, Mark; Freyer, Elisabeth; Ross, Allyson; van Heyningen, Veronica; Marsh, Joseph A.; Elmslie, Frances; FitzPatrick, David R.

2016-01-01

Gillespie syndrome (GS) is characterized by bilateral iris hypoplasia, congenital hypotonia, non-progressive ataxia, and progressive cerebellar atrophy. Trio-based exome sequencing identified de novo mutations in ITPR1 in three unrelated individuals with GS recruited to the Deciphering Developmental Disorders study. Whole-exome or targeted sequence analysis identified plausible disease-causing ITPR1 mutations in 10/10 additional GS-affected individuals. These ultra-rare protein-altering variants affected only three residues in ITPR1: Glu2094 missense (one de novo, one co-segregating), Gly2539 missense (five de novo, one inheritance uncertain), and Lys2596 in-frame deletion (four de novo). No clinical or radiological differences were evident between individuals with different mutations. ITPR1 encodes an inositol 1,4,5-triphosphate-responsive calcium channel. The homo-tetrameric structure has been solved by cryoelectron microscopy. Using estimations of the degree of structural change induced by known recessive- and dominant-negative mutations in other disease-associated multimeric channels, we developed a generalizable computational approach to indicate the likely mutational mechanism. This analysis supports a dominant-negative mechanism for GS variants in ITPR1. In GS-derived lymphoblastoid cell lines (LCLs), the proportion of ITPR1-positive cells using immunofluorescence was significantly higher in mutant than control LCLs, consistent with an abnormality of nuclear calcium signaling feedback control. Super-resolution imaging supports the existence of an ITPR1-lined nucleoplasmic reticulum. Mice with Itpr1 heterozygous null mutations showed no major iris defects. Purkinje cells of the cerebellum appear to be the most sensitive to impaired ITPR1 function in humans. Iris hypoplasia is likely to result from either complete loss of ITPR1 activity or structure-specific disruption of multimeric interactions. PMID:27108798

Lamin A/C might be involved in the EMT signalling pathway.

PubMed

Zuo, Lingkun; Zhao, Huanying; Yang, Ronghui; Wang, Liyong; Ma, Hui; Xu, Xiaoxue; Zhou, Ping; Kong, Lu

2018-07-15

We have previously reported a heterogeneous expression pattern of the nuclear membrane protein lamin A/C in low- and high-Gleason score (GS) prostate cancer (PC) tissues, and we have now found that this change is not associated with LMNA mutations. This expression pattern appears to be similar to the process of epithelial to mesenchymal transition (EMT) or to that of mesenchymal to epithelial transition (MET). The role of lamin A/C in EMT or MET in PC remains unclear. Therefore, we first investigated the expression levels of and the associations between lamin A/C and several common EMT markers, such as E-cadherin, N-cadherin, β-catenin, snail, slug and vimentin in PC tissues with different GS values and in different cell lines with varying invasion abilities. Our results suggest that lamin A/C might constitute a type of epithelial marker that better signifies EMT and MET in PC tissue, since a decrease in lamin A/C expression in GS 4 + 5 cases is likely associated with the EMT process, while the re-expression of lamin A/C in GS 5 + 4 cases is likely linked with MET. The detailed GS better exhibited the changes in lamin A/C and the EMT markers examined. Lamin A/C overexpression or knockdown had an impact on EMT biomarkers in a cell model by direct regulation of β-catenin. Hence, we suggest that lamin A/C might serve as a reliable epithelial biomarker for the distinction of PC cell differentiation and might also be a fundamental factor in the occurrence of EMT or MET in PC. Copyright © 2018. Published by Elsevier B.V.

Gorlin-Goltz syndrome: a rare case report.

PubMed

Mohan, Ravi Prakash Sasankoti; Verma, Sankalp; Agarwal, Neha; Singh, Udita

2013-06-27

Gorlin-Goltz syndrome (GS), also known as nevoid basal cell carcinoma syndrome, is an infrequent multisystem disease inherited in a dominant autosomal way, which shows a high level of penetrance and variable expressiveness. It is characterised by keratocystic odontogenic tumours (KCOT) in the jaw, multiple basal cell nevi carcinomas and skeletal abnormalities. This syndrome may be diagnosed early by a dentist by routine radiographical examinations in the first decade of life, since the KCOTs are usually one of the first manifestations of the syndrome. This article describes an 11-year-old boy with GS.

Gorlin-Goltz syndrome: a rare case report

PubMed Central

Mohan, Ravi Prakash Sasankoti; Verma, Sankalp; Agarwal, Neha; Singh, Udita

2013-01-01

Gorlin-Goltz syndrome (GS), also known as nevoid basal cell carcinoma syndrome, is an infrequent multisystem disease inherited in a dominant autosomal way, which shows a high level of penetrance and variable expressiveness. It is characterised by keratocystic odontogenic tumours (KCOT) in the jaw, multiple basal cell nevi carcinomas and skeletal abnormalities. This syndrome may be diagnosed early by a dentist by routine radiographical examinations in the first decade of life, since the KCOTs are usually one of the first manifestations of the syndrome. This article describes an 11-year-old boy with GS. PMID:23814215

Multi-level storage and ultra-high speed of superlattice-like Ge50Te50/Ge8Sb92 thin film for phase-change memory application.

PubMed

Wu, Weihua; Chen, Shiyu; Zhai, Jiwei; Liu, Xinyi; Lai, Tianshu; Song, Sannian; Song, Zhitang

2017-10-06

Superlattice-like Ge 50 Te 50 /Ge 8 Sb 92 (SLL GT/GS) thin film was systematically investigated for multi-level storage and ultra-fast switching phase-change memory application. In situ resistance measurement indicates that SLL GT/GS thin film exhibits two distinct resistance steps with elevated temperature. The thermal stability of the amorphous state and intermediate state were evaluated with the Kissinger and Arrhenius plots. The phase-structure evolution revealed that the amorphous SLL GT/GS thin film crystallized into rhombohedral Sb phase first, then the rhombohedral GeTe phase. The microstructure, layered structure, and interface stability of SLL GT/GS thin film was confirmed by using transmission electron microscopy. The transition speed of crystallization and amorphization was measured by the picosecond laser pump-probe system. The volume variation during the crystallization was obtained from x-ray reflectivity. Phase-change memory (PCM) cells based on SLL GT/GS thin film were fabricated to verify the multi-level switching under an electrical pulse as short as 30 ns. These results illustrate that the SLL GT/GS thin film has great potentiality in high-density and high-speed PCM applications.

Secreted Respiratory Syncytial Virus G Glycoprotein Induces Interleukin-5 (IL-5), IL-13, and Eosinophilia by an IL-4-Independent Mechanism

PubMed Central

Johnson, Teresa R.; Graham, Barney S.

1999-01-01

The attachment glycoprotein G of respiratory syncytial virus (RSV) is produced as both membrane-anchored and secreted forms by infected cells. Immunization with secreted RSV G (Gs) or formalin-inactivated alumprecipitated RSV (FI-RSV) predisposes mice to immune responses involving a Th2 cell phenotype which results in more severe illness and pathology, decreased viral clearance, and increased pulmonary eosinophilia upon subsequent RSV challenge. These responses are associated with increased interleukin-4 (IL-4) production in FI-RSV-primed mice, and the responses are IL-4 dependent. RNase protection assays demonstrated that similar levels of IL-4 mRNA were induced after RSV challenge in mice primed with vaccinia virus expressing Gs (vvGs) or a construct expressing only membrane-anchored G (vvGr). However, upon RSV challenge, vvGs-primed mice produced significantly greater levels of IL-5 and IL-13 mRNA and protein than vvGr-primed mice. Administration of neutralizing anti-IL-4 antibody 11.B11 during vaccinia virus priming did not alter the levels of vvGs-induced IL-5, IL-13, pulmonary eosinophilia, illness, or RSV titers upon RSV challenge, although immunoglobulin G (IgG) isotype profiles revealed that more IgG2a was produced. vvGs-priming of IL-4-deficient mice demonstrated that G-induced airway eosinophilia was not dependent on IL-4. In contrast, airway eosinophilia induced by FI-RSV priming was significantly reduced in IL-4-deficient mice. Thus we conclude that, in contrast to FI-RSV, the secreted form of RSV G can directly induce IL-5 and IL-13, producing pulmonary eosinophilia and enhanced illness in RSV-challenged mice by an IL-4-independent mechanism. PMID:10482601

Characterization of two minor saponins from Cordia piauhiensis by 1H and 13C NMR spectroscopy.

PubMed

Santos, Renata P; Silveira, Edilberto R; Lemos, Telma Leda G; Viana, Francisco Arnaldo; Braz-Filho, Raimundo; Pessoa, Otília Deusdênia L

2005-06-01

A careful NMR analysis with full assignment of the 1H and 13C spectral data for two minor saponins isolated from stems of Cordia piauhiensis is reported. These saponins were isolated by high-performance liquid chromatography and characterized as 3beta-O-[alpha-L-rhamnopyranosyl-(1 --> 2)-beta-D-glucopyranosyl]pomolic acid 28-O-[beta-D-glucopyranosyl-(1 --> 6)-beta-D-glucopyranosyl] ester (1) and 3beta-O-[alpha-L-rhamnopyranosyl-(1 --> 2)-beta-D-glucopyranosyl]oleanolic acid 28-O-[beta-D-xylopyranosyl-(1 --> 2)-beta-D-glucopyranosyl-(1 --> 6)-beta-D-glucopyranosyl] ester (2). Their structures were established using a combination of 1D and 2D (1H, 1H-COSY, TOCSY, NOESY, gs-HMQC and gs-HMBC) NMR techniques, electrospray ionization mass spectrometry and chemical evidence. Copyright 2005 John Wiley & Sons, Ltd.

Differential immune response of mallard duck peripheral blood mononuclear cells to two highly pathogenic avian influenza H5N1 viruses with distinct pathogenicity in mallard ducks.

PubMed

Cui, Zhu; Hu, Jiao; He, Liang; Li, Qunhui; Gu, Min; Wang, Xiaoquan; Hu, Shunlin; Liu, Huimou; Liu, Wenbo; Liu, Xiaowen; Liu, Xiufan

2014-02-01

CK10 and GS10 are two H5N1 highly pathogenic influenza viruses of similar genetic background but differ in their pathogenicity in mallard ducks. CK10 is highly pathogenic whereas GS10 is low pathogenic. In this study, strong inflammatory response in terms of the expression level of several cytokines was observed in mallard duck peripheral blood mononuclear cells (PBMC) infected with CK10 while mild response was triggered in those by GS10 infection. Two remarkable and intense peaks of immune response were induced by CK10 infection within 24 hours (at 8 and 24 hours post infection, respectively) without reducing the virus replication. Our observations indicated that sustained and intense innate immune responses may be central to the high pathogenicity caused by CK10 in ducks.

Decreased fat accumulation in 3T3-L1 pre-adipocytes treated with extracts of heat processed soy flour and breads

USDA-ARS?s Scientific Manuscript database

The antiadipogenic effects of a gluten-free soy bread developed from soy flour pretreated by germination (GS), steaming (SS), or roasting (RS) were evaluated in an in vitro adipocyte cell model. Previously we had shown that soy bread prepared by GS, SS or RS flours had acceptable sensory attributes ...

BcGs1, a glycoprotein from Botrytis cinerea, elicits defence response and improves disease resistance in host plants.

PubMed

Zhang, Yi; Zhang, Yunhua; Qiu, Dewen; Zeng, Hongmei; Guo, Lihua; Yang, Xiufen

2015-02-20

In this study, a necrosis-inducing protein was purified from the culture filtrate of the necrotrophic fungus Botrytis cinerea BC-98 strain. Secreted proteins were collected and fractionated by liquid chromatography. The fraction with the highest necrosis-inducing activity was further purified. A glycoprotein named BcGs1 was identified by 2D electrophoresis and mass spectrometry. The BcGs1 protein consisted of 672 amino acids with a theoretical molecular weight of 70.487 kDa. Functional domain analysis indicated that BcGs1 was a glucan 1,4-alpha-glucosidase, a cell wall-degrading enzyme, with a Glyco_hydro_15 domain and a CBM20_glucoamylase domain. The BcGs1 protein caused necrotic lesions that mimicked a typical hypersensitive response and H2O2 production in tomato and tobacco leaves. BcGs1-treated plants exhibited resistance to B. cinerea, Pseudomonas syringae pv. tomato DC3000 and tobacco mosaic virus in systemic leaves. In addition, BcGs1 triggered elevation of the transcript levels of the defence-related genes PR-1a, TPK1b and Prosystemin. This is the first report of a Botrytis glucan 1,4-alpha-glucosidase triggering host plant immunity as an elicitor. These results lay a foundation for further study of the comprehensive interaction between plants and necrotrophic fungi. Copyright © 2015 Elsevier Inc. All rights reserved.

Isolated dorsal root ganglion neurones inhibit receptor-dependent adenylyl cyclase activity in associated glial cells

PubMed Central

Ng, KY; Yeung, BHS; Wong, YH; Wise, H

2013-01-01

Background and Purpose Hyper-nociceptive PGE2 EP4 receptors and prostacyclin (IP) receptors are present in adult rat dorsal root ganglion (DRG) neurones and glial cells in culture. The present study has investigated the cell-specific expression of two other Gs-protein coupled hyper-nociceptive receptor systems: β-adrenoceptors and calcitonin gene-related peptide (CGRP) receptors in isolated DRG cells and has examined the influence of neurone–glial cell interactions in regulating adenylyl cyclase (AC) activity. Experimental Approach Agonist-stimulated AC activity was determined in mixed DRG cell cultures from adult rats and compared with activity in DRG neurone-enriched cell cultures and pure DRG glial cell cultures. Key Results Pharmacological analysis showed the presence of Gs-coupled β2-adrenoceptors and CGRP receptors, but not β1-adrenoceptors, in all three DRG cell preparations. Agonist-stimulated AC activity was weakest in DRG neurone-enriched cell cultures. DRG neurones inhibited IP receptor-stimulated glial cell AC activity by a process dependent on both cell–cell contact and neurone-derived soluble factors, but this is unlikely to involve purine or glutamine receptor activation. Conclusions and Implications Gs-coupled hyper-nociceptive receptors are readily expressed on DRG glial cells in isolated cell cultures and the activity of CGRP, EP4 and IP receptors, but not β2-adrenoceptors, in glial cells is inhibited by DRG neurones. Studies using isolated DRG cells should be aware that hyper-nociceptive ligands may stimulate receptors on glial cells in addition to neurones, and that variable numbers of neurones and glial cells will influence absolute measures of AC activity and affect downstream functional responses. PMID:22924655

Broad-spectrum antiviral GS-5734 inhibits both epidemic and zoonotic coronaviruses

PubMed Central

Sheahan, Timothy P.; Sims, Amy C.; Graham, Rachel L.; Menachery, Vineet D.; Gralinski, Lisa E.; Case, James B.; Leist, Sarah R.; Pyrc, Krzysztof; Feng, Joy Y.; Trantcheva, Iva; Bannister, Roy; Park, Yeojin; Babusis, Darius; Clarke, Michael O.; Mackman, Richard L.; Spahn, Jamie E.; Palmiotti, Christopher A.; Siegel, Dustin; Ray, Adrian S.; Cihlar, Tomas; Jordan, Robert; Denison, Mark R.; Baric, Ralph S.

2017-01-01

Emerging viral infections are difficult to control as heterogeneous members periodically cycle in and out of humans and zoonotic hosts, complicating the development of specific antiviral therapies and vaccines. Coronaviruses (CoVs) have a proclivity to spread rapidly into new host species causing severe disease. SARS-CoV and MERS-CoV successively emerged causing severe epidemic respiratory disease in immunologically naïve human populations throughout the globe. Broad-spectrum therapies capable of inhibiting CoV infections would address an immediate unmet medical need and could be invaluable in the treatment of emerging and endemic CoV infections. Here we show that a nucleotide prodrug GS-5734, currently in clinical development for treatment of Ebola virus disease, can inhibit SARS-CoV and MERS-CoV replication in multiple in vitro systems including primary human airway epithelial cell cultures with submicromolar IC50 values. GS-5734 was also effective against bat-CoVs, prepandemic bat-CoVs and circulating contemporary human CoV in primary human lung cells, thus demonstrating broad-spectrum anti-CoV activity. In a mouse model of SARS-CoV pathogenesis, prophylactic and early therapeutic administration of GS-5734 significantly reduced lung viral load and improved clinical signs of disease as well as respiratory functions. These data provide substantive evidence that GS-5734 may prove effective against endemic MERS-CoV in the Middle East, circulating human CoV, and possibly most importantly, emerging CoV of the future. PMID:28659436

Stomatal characteristics and infection biology of Pyrenopeziza betulicola in Betula pendula trees grown under elevated CO2 and O3.

PubMed

Riikonen, Johanna; Syrjälä, Leena; Tulva, Ingmar; Mänd, Pille; Oksanen, Elina; Poteri, Marja; Vapaavuori, Elina

2008-11-01

Two silver birch clones were exposed to ambient and elevated concentrations of CO(2) and O(3), and their combination for 3 years, using open-top chambers. We evaluated the effects of elevated CO(2) and O(3) on stomatal conductance (g(s)), density (SD) and index (SI), length of the guard cells, and epidermal cell size and number, with respect to crown position and leaf type. The relationship between the infection biology of the fungus (Pyrenopeziza betulicola) causing leaf spot disease and stomatal characteristics was also studied. Leaf type was an important determinant of O(3) response in silver birch, while crown position and clone played only a minor role. Elevated CO(2) reduced the g(s), but had otherwise no significant effect on the parameters studied. No significant interactions between elevated CO(2) and O(3) were found. The infection biology of P. betulicola was not correlated with SD or g(s), but it did occasionally correlate positively with the length of the guard cells.

Mechanistic Characterization of GS-9190 (Tegobuvir), a Novel Nonnucleoside Inhibitor of Hepatitis C Virus NS5B Polymerase▿

PubMed Central

Shih, I-hung; Vliegen, Inge; Peng, Betty; Yang, Huiling; Hebner, Christy; Paeshuyse, Jan; Pürstinger, Gerhard; Fenaux, Martijn; Tian, Yang; Mabery, Eric; Qi, Xiaoping; Bahador, Gina; Paulson, Matthew; Lehman, Laura S.; Bondy, Steven; Tse, Winston; Reiser, Hans; Lee, William A.; Schmitz, Uli; Neyts, Johan; Zhong, Weidong

2011-01-01

GS-9190 (Tegobuvir) is a novel imidazopyridine inhibitor of hepatitis C virus (HCV) RNA replication in vitro and has demonstrated potent antiviral activity in patients chronically infected with genotype 1 (GT1) HCV. GS-9190 exhibits reduced activity against GT2a (JFH1) subgenomic replicons and GT2a (J6/JFH1) infectious virus, suggesting that the compound's mechanism of action involves a genotype-specific viral component. To further investigate the GS-9190 mechanism of action, we utilized the susceptibility differences between GT1b and GT2a by constructing a series of replicon chimeras where combinations of 1b and 2a nonstructural proteins were encoded within the same replicon. The antiviral activities of GS-9190 against the chimeric replicons were reduced to levels comparable to that of the wild-type GT2a replicon in chimeras expressing GT2a NS5B. GT1b replicons in which the β-hairpin region (amino acids 435 to 455) was replaced by the corresponding sequence of GT2a were markedly less susceptible to GS-9190, indicating the importance of the thumb subdomain of the polymerase in this effect. Resistance selection in GT1b replicon cells identified several mutations in NS5B (C316Y, Y448H, Y452H, and C445F) that contributed to the drug resistance phenotype. Reintroduction of these mutations into wild-type replicons conferred resistance to GS-9190, with the number of NS5B mutations correlating with the degree of resistance. Analysis of GS-9190 cross-resistance against previously reported NS5B drug-selected mutations showed that the resistance pattern of GS-9190 is different from other nonnucleoside inhibitors. Collectively, these data demonstrate that GS-9190 represents a novel class of nonnucleoside polymerase inhibitors that interact with NS5B likely through involvement of the β-hairpin in the thumb subdomain. PMID:21746939

L-Theanine Content and Related Gene Expression: Novel Insights into Theanine Biosynthesis and Hydrolysis among Different Tea Plant (Camellia sinensis L.) Tissues and Cultivars

PubMed Central

Liu, Zhi-Wei; Wu, Zhi-Jun; Li, Hui; Wang, Yong-Xin; Zhuang, Jing

2017-01-01

L-Theanine content has tissues and cultivars specificity in tea plant (Camellia sinensis L.), the correlations of theanine metabolic related genes expression profiles with theanine contents were explored in this study. L-theanine contents in the bud and 1st leaf, 2nd leaf, 3rd leaf, old leaf, stem, and lateral root were determined by HPLC from three C. sinensis cultivars, namely ‘Huangjinya’, ‘Anjibaicha’, and ‘Yingshuang’, respectively. The theanine contents in leaves and root of ‘Huangjinya’ were the highest, followed by ‘Anjibaicha’, and ‘Yingshuang’. The theanine contents in the leaves reduced as the leaf mature gradually, and in stem were the least. Seventeen genes encoding enzymes involved in theanine metabolism were identified from GenBank and our tea transcriptome database, including CsTS1, CsTS2, CsGS1, CsGS2, CsGOGAT-Fe, CsGOGAT-NAD(P)H, CsGDH1, CsGDH2, CsALT, CsSAMDC, CsADC, CsCuAO, CsPAO, CsNiR, CsNR, CsGGT1, and CsGGT3. The transcript profiles of those seventeen genes in the different tissues of three tea plant cultivars were analyzed comparatively. Among the different cultivars, the transcript levels of most selected genes in ‘Huangjinya’ were significantly higher than that in the ‘Anjibaicha’ and ‘Yingshuang’. Among the different tissues, the transcript levels of CsTS2, CsGS1, and CsGDH2 almost showed positive correlation with the theanine contents, while the other genes showed negative correlation with the theanine contents in most cases. The theanine contents showed correlations with related genes expression levels among cultivars and tissues of tea plant, and were determined by the integrated effect of the metabolic related genes. PMID:28439281

Direct Loading and Tunable Release of Antibiotics from Polyelectrolyte Multilayers To Reduce Bacterial Adhesion and Biofilm Formation.

PubMed

Wang, Bailiang; Jin, Tingwei; Xu, Qingwen; Liu, Huihua; Ye, Zi; Chen, Hao

2016-05-18

Bacteria adhesion on the surface of biomaterials and following biofilm formation are important problems in biomedical applications. The charged antibiotics with small molar mass can hardly deposit alternately with polymers into multilayered films to load the drug. Herein, the (poly(acrylic acid)-gentamicin/poly(ethylenimine))n ((PAA-GS/PEI)n) multilayer film was designed and constructed via a layer-by-layer self-assembly method. Low molar mass GS cations were first combined with polyanion PAA and self-assembled with PEI to form multilayer films showing exponential growth behavior. The GS dosage could be adjusted by changing the layer number of films. Furthermore, the thermal cross-linking method was used to control the release rate of GS in PBS buffer. Owing to the diffusion of GS, a zone of inhibition of about 7.0 mm showed the efficient disinfection activity of the multilayer film. It could also be seen from the biofilm inhibition assay that the multilayer film effectively inhibited bacterial adhesion and biofilm formation. As the drug loading dosage was 160 μg/cm(2), the multilayer films showed very low cytotoxicity against human lens epithelial cells. The present work provides an easy way to load GS into multilayer films which can be applied to surface modification of implants and biomedical devices.

Facile fabrication of 3D porous MnO@GS/CNT architecture as advanced anode materials for high-performance lithium-ion battery.

PubMed

Wang, Junyong; Deng, Qinglin; Li, Mengjiao; Wu, Cong; Jiang, Kai; Hu, Zhigao; Chu, Junhao

2018-08-03

To overcome inferior rate capability and cycle stability of MnO-based anode materials for lithium-ion batteries (LIBs), we reported a novel 3D porous MnO@GS/CNT composite, consisting of MnO nanoparticles homogeneously distributed on the conductive interconnected framework based on 2D graphene sheets (GS) and 1D carbon nanotubes (CNTs). The distinctive architecture offers highly interpenetrated network along with efficient porous channels for fast electron transfer and ionic diffusion as well as abundant stress buffer space to accommodate the volume expansion of the MnO nanoparticles. The MnO@GS/CNT anode exhibits an ultrahigh capacity of 1115 mAh g -1 at 0.2 A g -1 after 150 cycles and outstanding rate capacity of 306 mAh g -1 at 10.0 A g -1 . Moreover, a stable capacity of 405 mAh g -1 after 3200 cycles can still be achieved, even at a large current density of 5.0 A g -1 . When coupled with LiMn 2 O 4 (LMO) cathode, the LMO [Formula: see text] MnO@GS/CNT full cell characterizes an excellent cycling stability and rate capability, indicating the promising application of MnO@GS/CNT anode in the next-generation LIBs.

A light-dependent ammonia-assimilating mechanism in the ctenidia of a giant clam

NASA Astrophysics Data System (ADS)

Hiong, Kum C.; Choo, Celine Y. L.; Boo, Mel V.; Ching, Biyun; Wong, Wai P.; Chew, Shit F.; Ip, Yuen K.

2017-03-01

Giant clams harbor symbiotic zooxanthellae and manifest light-enhanced calcification. In light, they also increase the absorption and assimilation of exogenous ammonia, but the roles of the host and symbionts are unclear and the reason for light dependency remains enigmatic. Here, we report the complete coding cDNA sequence of a glutamine synthetase (GS), from the ctenidia (gill) of Tridacna squamosa. The deduced GS amino acid sequence was closely related to other animal GSs, indicating a host origin. The GS/GS transcript level and protein abundance increased significantly in the ctenidia after 12 h of light exposure. These results denote the ctenidia as the site of light-enhanced ammonia absorption with the absorbed ammonia being assimilated to glutamine by the host GS. Immunofluorescence microscopy confirmed that GS was expressed predominantly in the cytoplasm of the epithelial cells separating the tertiary water channels from the hemolymph inside the ctenidia. Hence, unlike symbiotic cnidarians, the host clam probably supplies exogenous nitrogen as glutamine from the ctenidia, through the hemolymph and tubular fluid, to the nitrogen-deficient zooxanthellae which reside extracellularly in the extensible outer mantle. This is the first report on light-dependent gene and protein expression of a host enzyme involved in nitrogen metabolism in an alga-invertebrate association.

Synthetic Human Monoclonal Antibodies toward Staphylococcal Enterotoxin B (SEB) Protective against Toxic Shock Syndrome*

PubMed Central

Karauzum, Hatice; Chen, Gang; Abaandou, Laura; Mahmoudieh, Mahta; Boroun, Atefeh R.; Shulenin, Sergey; Devi, V. Sathya; Stavale, Eric; Warfield, Kelly L.; Zeitlin, Larry; Roy, Chad J.; Sidhu, Sachdev S.; Aman, M. Javad

2012-01-01

Staphylococcal enterotoxin B (SEB) is a potent toxin that can cause toxic shock syndrome and act as a lethal and incapacitating agent when used as a bioweapon. There are currently no vaccines or immunotherapeutics available against this toxin. Using phage display technology, human antigen-binding fragments (Fabs) were selected against SEB, and proteins were produced in Escherichia coli cells and characterized for their binding affinity and their toxin neutralizing activity in vitro and in vivo. Highly protective Fabs were converted into full-length IgGs and produced in mammalian cells. Additionally, the production of anti-SEB antibodies was explored in the Nicotiana benthamiana plant expression system. Affinity maturation was performed to produce optimized lead anti-SEB antibody candidates with subnanomolar affinities. IgGs produced in N. benthamiana showed characteristics comparable with those of counterparts produced in mammalian cells. IgGs were tested for their therapeutic efficacy in the mouse toxic shock model using different challenge doses of SEB and a treatment with 200 μg of IgGs 1 h after SEB challenge. The lead candidates displayed full protection from lethal challenge over a wide range of SEB challenge doses. Furthermore, mice that were treated with anti-SEB IgG had significantly lower IFNγ and IL-2 levels in serum compared with mock-treated mice. In summary, these anti-SEB monoclonal antibodies represent excellent therapeutic candidates for further preclinical and clinical development. PMID:22645125

An optimized library for reference-based deconvolution of whole-blood biospecimens assayed using the Illumina HumanMethylationEPIC BeadArray.

PubMed

Salas, Lucas A; Koestler, Devin C; Butler, Rondi A; Hansen, Helen M; Wiencke, John K; Kelsey, Karl T; Christensen, Brock C

2018-05-29

Genome-wide methylation arrays are powerful tools for assessing cell composition of complex mixtures. We compare three approaches to select reference libraries for deconvoluting neutrophil, monocyte, B-lymphocyte, natural killer, and CD4+ and CD8+ T-cell fractions based on blood-derived DNA methylation signatures assayed using the Illumina HumanMethylationEPIC array. The IDOL algorithm identifies a library of 450 CpGs, resulting in an average R 2  = 99.2 across cell types when applied to EPIC methylation data collected on artificial mixtures constructed from the above cell types. Of the 450 CpGs, 69% are unique to EPIC. This library has the potential to reduce unintended technical differences across array platforms.

FLiGS Score: A New Method of Outcome Assessment for Lip Carcinoma–Treated Patients

PubMed Central

Grassi, Rita; Toia, Francesca; Di Rosa, Luigi; Cordova, Adriana

2015-01-01

Background: Lip cancer and its treatment have considerable functional and cosmetic effects with resultant nutritional and physical detriments. As we continue to investigate new treatment regimens, we are simultaneously required to assess postoperative outcomes to design interventions that lessen the adverse impact of this disease process. We wish to introduce Functional Lip Glasgow Scale (FLiGS) score as a new method of outcome assessment to measure the effect of lip cancer and its treatment on patients’ daily functioning. Methods: Fifty patients affected by lip squamous cell carcinoma were recruited between 2009 and 2013. Patients were asked to fill the FLiGS questionnaire before surgery, 1 month, 6 months, and 1 year after surgery. The subscores were used to calculate a total FLiGS score of global oral disability. Statistical analysis was performed to test validity and reliability. Results: FLiGS scores improved significantly from preoperative to 12 months postoperative values (P = 0.000). Statistical evidence of validity was provided through rs (Spearman correlation coefficient) that resulted >0.30 for all surveys and for which P < 0.001. FLiGS score reliability was shown through examination of internal consistency and test-retest reliability. Conclusions: FLiGS score is a simple way of assessing functional impairment related to lip cancer before and after surgery; it is sensitive, valid, reliable, and clinically relevant: it provides useful information to orient the physician in the postoperative management and in the rehabilitation program. PMID:26034652

Gender-specific effects of endogenous testosterone: female alpha-estrogen receptor-deficient C57Bl/6J mice develop glomerulosclerosis.

PubMed

Elliot, S J; Berho, M; Korach, K; Doublier, S; Lupia, E; Striker, G E; Karl, M

2007-08-01

Young female mice on a C57Bl/6J (B6) background are considered glomerulosclerosis (GS)-resistant but aging B6 mice develop mild GS. Estrogen deficiency accelerates while estrogen replacement retards GS in young sclerosis-prone oligosyndactyly mutant mice on an ROP background. To explore the effects of sex hormones on glomerular structure and function in the context of gender and genetic background, we studied mice in which the estrogen-receptor (ER) genes alpha- or -beta were deleted (alpha- or betaER knockout (KO)) and crossed into the B6 background. We also studied ovariectomized (Ovx) B6 mice given testosterone. Male and female betaERKO and male alphaERKO mice had no glomerular dysfunction at 9 months of age; however, alphaERKO female mice displayed albuminuria and GS. Ovx prevented glomerular dysfunction in alphaERKO female mice by eliminating endogenous testosterone production while exogenous testosterone induced GS in Ovx B6 mice. Androgen receptor (AR) expression and function was found in microdissected glomeruli and cultured mesangial cells. Testosterone compared to placebo increased both AR expression and TGF-beta1 mRNA levels in glomeruli isolated from female B6 mice. Estrogen deficiency had no deleterious effects on the glomeruli in B6 mice. Our study shows that genetic traits strongly influence the GS-promoting effects of estrogen deficiency while testosterone induces GS in a gender-specific manner.

Abnormal Glycogen Storage by Retinal Neurons in Diabetes.

PubMed

Gardiner, Tom A; Canning, Paul; Tipping, Nuala; Archer, Desmond B; Stitt, Alan W

2015-12-01

It is widely held that neurons of the central nervous system do not store glycogen and that accumulation of the polysaccharide may cause neurodegeneration. Since primary neural injury occurs in diabetic retinopathy, we examined neuronal glycogen status in the retina of streptozotocin-induced diabetic and control rats. Glycogen was localized in eyes of streptozotocin-induced diabetic and control rats using light microscopic histochemistry and electron microscopy, and correlated with immunohistochemical staining for glycogen phosphorylase and phosphorylated glycogen synthase (pGS). Electron microscopy of 2-month-old diabetic rats (n = 6) showed massive accumulations of glycogen in the perinuclear cytoplasm of many amacrine neurons. In 4-month-old diabetic rats (n = 11), quantification of glycogen-engorged amacrine cells showed a mean of 26 cells/mm of central retina (SD ± 5), compared to 0.5 (SD ± 0.2) in controls (n = 8). Immunohistochemical staining for glycogen phosphorylase revealed strong expression in amacrine and ganglion cells of control retina, and increased staining in cell processes of the inner plexiform layer in diabetic retina. In control retina, the inactive pGS was consistently sequestered within the cell nuclei of all retinal neurons and the retinal pigment epithelium (RPE), but in diabetics nuclear pGS was reduced or lost in all classes of retinal cell except the ganglion cells and cone photoreceptors. The present study identifies a large population of retinal neurons that normally utilize glycogen metabolism but show pathologic storage of the polysaccharide during uncontrolled diabetes.

Cancer-associated fibroblasts regulate keratinocyte cell-cell adhesion via TGF-β-dependent pathways in genotype-specific oral cancer.

PubMed

Cirillo, N; Hassona, Y; Celentano, A; Lim, K P; Manchella, S; Parkinson, E K; Prime, S S

2017-01-01

The interrelationship between malignant epithelium and the underlying stroma is of fundamental importance in tumour development and progression. In the present study, we used cancer-associated fibroblasts (CAFs) derived from genetically unstable oral squamous cell carcinomas (GU-OSCC), tumours that are characterized by the loss of genes such as TP53 and p16 INK4A and with extensive loss of heterozygosity, together with CAFs from their more genetically stable (GS) counterparts that have wild-type TP53 and p16 INK4A and minimal loss of heterozygosity (GS-OSCC). Using a systems biology approach to interpret the genome-wide transcriptional profile of the CAFs, we show that transforming growth factor-β (TGF-β) family members not only had biological relevance in silico but also distinguished GU-OSCC-derived CAFs from GS-OSCC CAFs and fibroblasts from normal oral mucosa. In view of the close association between TGF-β family members, we examined the expression of TGF-β1 and TGF-β2 in the different fibroblast subtypes and showed increased levels of active TGF-β1 and TGF-β2 in CAFs from GU-OSCC. CAFs from GU-OSCC, but not GS-OSCC or normal fibroblasts, induced epithelial-mesenchymal transition and down-regulated a broad spectrum of cell adhesion molecules resulting in epithelial dis-cohesion and invasion of target keratinocytes in vitro in a TGF-β-dependent manner. The results demonstrate that the TGF-β family of cytokines secreted by CAFs derived from genotype-specific oral cancer (GU-OSCC) promote, at least in part, the malignant phenotype by weakening intercellular epithelial adhesion. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

The role of glutamine synthetase and glutamate dehydrogenase in nitrogen assimilation and possibilities for improvement in the nitrogen utilization of crops.

PubMed

Miflin, Ben J; Habash, Dimah Z

2002-04-01

This short review outlines the central role of glutamine synthetase (GS) in plant nitrogen metabolism and discusses some possibilities for crop improvement. GS functions as the major assimilatory enzyme for ammonia produced from N fixation, and nitrate or ammonia nutrition. It also reassimilates ammonia released as a result of photorespiration and the breakdown of proteins and nitrogen transport compounds. GS is distributed in different subcellular locations (chloroplast and cytoplasm) and in different tissues and organs. This distribution probably changes as a function of the development of the tissue, for example, GS1 appears to play a key role in leaf senescence. The enzyme is the product of multiple genes with complex promoters that ensure the expression of the genes in an organ- and tissue-specific manner and in response to a number of environmental variables affecting the nutritional status of the cell. GS activity is also regulated post-translationally in a manner that involves 14-3-3 proteins and phosphorylation. GS and plant nitrogen metabolism is best viewed as a complex matrix continually changing during the development cycle of plants. Along with GS, a number of other enzymes play key roles in maintaining the balance of carbon and nitrogen. It is proposed that one of these is glutamate dehydrogenase (GDH). There is considerable evidence for a GDH shunt to return the carbon in amino acids back into reactions of carbon metabolism and the tri-carboxylic acid cycle. Results with transgenic plants containing transferred GS genes suggest that there may be ways in which it is possible to improve the efficiency with which crop plants use nitrogen. Marker-assisted breeding may also bring about such improvements.

Purification and growth of melanocortin 1 receptor (Mc1r)-defective primary murine melanocytes is dependent on stem cell factor from keratinocyte-conditioned media

PubMed Central

Scott, Timothy L.; Wakamatsu, Kazumasa; Ito, Shosuke; D’Orazio, John A.

2015-01-01

Summary The melanocortin 1 receptor (MC1R) is a transmembrane Gs-coupled surface protein found on melanocytes that binds melanocyte stimulating hormone (MSH) and mediates activation of adenylyl cyclase and generation of the second messenger cAMP. MC1R regulates growth and differentiation of melanocytes and protects against carcinogenesis. Persons with loss-of-function polymorphisms of MC1R tend to be UV-sensitive (fair-skinned and with a poor tanning response) and are at high risk for melanoma. Mechanistic studies of the role of MC1R in melanocytic UV responses, however, have been hindered in part because Mc1r-defective primary murine melanocytes have been difficult to culture in vitro. Until now, effective growth of murine melanocytes has depended on cAMP stimulation with adenylyl cyclase activating or phosphodiesterase inhibiting agents. However, rescuing cAMP in the setting of defective MC1R signaling would be expected to confound experiments directly testing MC1R function on melanocytic UV responses. Here we report a novel method of culturing primary murine melanocytes in the absence of pharmacologic cAMP stimulation by incorporating conditioned supernatants containing stem cell factor (SCF) derived from primary keratinocytes. Importantly, this method seems to permit similar pigment expression by cultured melanocytes as that found in the skin of their parental murine strains. This novel approach will allow mechanistic investigation into MC1R’s role in protection against UV-mediated carcinogenesis and determination of the role of melanin pigment subtypes on UV-mediated melanocyte responses. PMID:19633898

Pituitary hyperplasia and gigantism in mice caused by a cholera toxin transgene.

PubMed

Burton, F H; Hasel, K W; Bloom, F E; Sutcliffe, J G

1991-03-07

Cyclic AMP is thought to act as an intracellular second messenger, mediating the physiological response of many cell types to extracellular signals. In the pituitary, growth hormone (GH)-producing cells (somatotrophs) proliferate and produce GH in response to hypothalamic GH-releasing factor, which binds a receptor that stimulates Gs protein activation of adenylyl cyclase. We have now determined whether somatotroph proliferation and GH production are stimulated by cAMP alone, or require concurrent, non-Gs-mediated induction of other regulatory molecules by designing a transgene to induce chronic supraphysiological concentrations of cAMP in somatotrophs. The rat GH promoter was used to express an intracellular form of cholera toxin, a non-cytotoxic and irreversible activator of Gs. Introduction of this transgene into mice caused gigantism, elevated serum GH levels, somatotroph proliferation and pituitary hyperplasia. These results support the direct triggering of these events by cAMP, and illustrate the utility of cholera toxin transgenes as a tool for physiological engineering.

CpG Oligodeoxynucleotides Facilitate Delivery of Whole Inactivated H9N2 Influenza Virus via Transepithelial Dendrites of Dendritic Cells in Nasal Mucosa

PubMed Central

Qin, Tao; Yin, Yinyan; Yu, Qinghua; Huang, Lulu; Wang, Xiaoqing; Lin, Jian

2015-01-01

ABSTRACT The spread of the low-pathogenicity avian H9N2 influenza virus has seriously increased the risk of a new influenza pandemic. Although whole inactivated virus (WIV) vaccine via intranasal pathway is the effective method of blocking virus transmission, the mucosal barrier seems to be a major factor hampering its development. CpG oligodeoxynucleotides, a known adjuvant, can target downstream dendritic cells (DCs) and effectively enhance the mucosal and systemic immune responses. However, the ability of CpGs to assist H9N2 WIV in transepithelial transport remains unknown. Here, in vitro and in vivo, we showed that CpGs provided assistance for H9N2 WIV in recruiting DCs to the nasal epithelial cells (ECs) and forming transepithelial dendrites (TEDs) to capture luminal viruses. CD103+ DCs participated in this process. Chemokine CCL20 from nasal ECs played a key role in driving DC recruitment and TED formation. Virus-loaded DCs quickly migrated into the draining cervical lymph nodes (CLNs) for antigen presentation. In addition, the competence of CpGs was independent of direct epithelial transport via the transcellular or paracellular pathway. Taken together, our data demonstrated that CpGs enhanced the transport of H9N2 WIV via TEDs of nasal DCs, which might be a novel mechanism for optimal adaptive immune responses. IMPORTANCE This paper demonstrates by both an in vivo and an in vitro coculture model that CpG oligodeoxynucleotides, known as an adjuvant generally targeting downstream immune responses, also are crucial for the transport of H9N2 WIV across nasal epithelial cells (ECs) via the uptake of transepithelial dendrites (TEDs). Our results prove for the first time to our knowledge that the immune-potentiating mechanism of CpGs is based on strengthening the transepithelial uptake of H9N2 WIV in nasal mucosa. These findings provide a fresh perspective for further improvement of intranasal influenza vaccines, which are urgently needed in the face of the potential threat of H9N2 influenza. PMID:25810544

Differentiation to adipocytes in accompanied by an increase in the amounts of Gi- and Go-proteins in 3T3-L1 cells

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

Watkins, D.C.; Northup, J.K.; Malbon, C.C.

Treatment of cultures of 3T3-L1 cells with methylisobutyl-xanthine and dexamethasone has been shown to result in accumulation of lipid and conversion to the morphology of adipocytes in more than 90% of the cells. The status of the stimulatory (Gs), inhibitory (Gi) and Go-proteins during the course of 3T3-L1 differentiation was examined. The amount of alpha subunit of Gs (..cap alpha..Gs), assayed by radiolabeling in the presence of cholera toxin and (/sup 32/P)NAD/sup +/, increased upon differentiation as previously described by others. The amounts of ..cap alpha..Gi and ..cap alpha..Go assayed by radiolabeling in the presence of pertussis toxin and (/supmore » 32/P)NAD/sup +/ increased 3-fold upon differentiation. Immunoblots of cell membranes subjected to gel electrophoresis in sodium dodecyl sulfate were probed with two rabbit antisera raised against bovine brain ..cap alpha..Go and with one raised against the..beta..-subunit of the bovine rod-outer-segment G-protein, referred to as transducin. The immunoblotting data confirm the increase upon differentiation of ..cap alpha..Go and also demonstrate an increase in the amount of the ..beta..-subunit. Thus differentiation of 3T3-L1 cells is accompanied by dramatic changes in the complexion of G-proteins in the membranes.« less

Oral administration of Lentinus edodes β-glucans ameliorates DSS-induced ulcerative colitis in mice via MAPK-Elk-1 and MAPK-PPARγ pathways.

PubMed

Shi, Limin; Lin, Qinlu; Yang, Tao; Nie, Ying; Li, Xinhua; Liu, Bo; Shen, Junjun; Liang, Ying; Tang, Yiping; Luo, Feijun

2016-11-09

To evaluate the anti-inflammatory effect of β-glucans from Lentinus edodes, and its molecular mechanism, the dextran sulfate sodium salt (DSS) induced colitis model of mice and the LPS-stimulated RAW264.7 cell inflammation model were used in this study. 40 ICR male mice were randomly divided into 4 groups: Control, DSS (DSS treated only), DSS + low-βGs (500 mg kg -1 d -1 ) and DSS + high-βGs (1000 mg kg -1 d -1 ). The body weight of the mice with Lentinus edodes β-glucan supplementation increased significantly compared to the DSS group and the disease activity index (DAI) was improved in both βG-treated groups. Compared with the DSS group, histopathological analysis showed that the infiltration of inflammatory cells of both βG-treated groups decreased significantly in colonic tissues. Furthermore, oral administration of β-glucans decreases the concentration of malondialdehyde (MDA) and myeloperoxidase (MPO) and inhibits the expression of iNOS and several inflammatory factors: TNF-α, IL-1β and IL-6 as well as nitric oxide (NO) of the colonic tissues. The mitogen-activated protein kinase (MAPK) pathway is closely related to the expression of pro-inflammatory factors. In the DSS-induced colitis model and the LPS-stimulated RAW264.7 cell model, βGs inhibited the expression of pro-inflammatory factors and blocked the phosphorylation of JNK/ERK1/2 and p38; βGs also suppress the phosphorylation of Elk-1 at Ser84 and the phosphorylation of PPARγ at Ser112. Altogether, these results suggest that Lentinus edodes βGs could inhibit the DSS-induced ulcerative colitis and decrease inflammatory factor expressions. The molecular mechanism may be involved in suppressing MAPK signaling and inactivation of Elk-1 and activation of PPARγ.

Overexpression of glutamine synthetase is associated with beta-catenin-mutations in mouse liver tumors during promotion of hepatocarcinogenesis by phenobarbital.

PubMed

Loeppen, Sandra; Schneider, Daniela; Gaunitz, Frank; Gebhardt, Rolf; Kurek, Raffael; Buchmann, Albrecht; Schwarz, Michael

2002-10-15

Phenobarbital (PB) is an antiepileptic drug that promotes hepatocarcinogenesis in rodents when administered subsequent to an initiating carcinogen like N-nitrosodiethylamine (DEN). In the mouse, the promotional effect of PB on liver tumor development results from a selective stimulation of clonal outgrowth of hepatocytes harboring activating mutations in the beta-catenin gene. Because glutamine synthetase (GS) has recently been shown to be a putative transcriptional target of beta-catenin, expression of GS during PB-mediated promotion of mouse hepatocarcinogenesis was investigated. Preneoplastic and neoplastic liver lesions were induced in 6-week-old male mice by a single injection of 90 micro g/g body weight of DEN, and groups of mice were subsequently kept on PB-containing (0.05%) or control diet for 39 weeks. In PB-treated mice, 46 of 51 lesions ( approximately 90%) were GS-positive in contrast to only 16 of 46 ( approximately 35%) in mice not treated with PB. Approximately 33% of liver was occupied by neoplastic tissue in PB-treated mice, of which >80% was GS positive. By contrast, only approximately 3.5% of liver consisted of neoplastic tissue in mice treated with DEN only, and approximately 25% of this was GS positive. We have previously shown that beta-catenin mutations are present in approximately 80% of liver tumors from PB-treated mice but are absent in liver tumors from mice treated with DEN only. By analyzing a panel of larger liver tumors, we now observed that tumors harboring beta-catenin mutations were GS positive, whereas tumors without beta-catenin mutations were GS negative. Similarly, tumors from an additional mouse carcinogenicity experiment where PB inhibited rather than promoted hepatocarcinogenesis were mostly GS negative. These data suggest that promotion of hepatocarcinogenesis by PB confers beta-catenin-mutated tumor cells with a selective advantage by up-regulation of GS expression.

Effects of Acutely Elevated Hydrostatic Pressure in a Rat Ex Vivo Retinal Preparation

PubMed Central

Yoshitomi, Takeshi; Zorumski, Charles F.; Izumi, Yukitoshi

2010-01-01

Purpose. A new experimental glaucoma model was developed by using an ex vivo rat retinal preparation to examine the effects of elevated hydrostatic pressure on retinal morphology and glutamine synthetase (GS) activity. Methods. Ex vivo rat retinas were exposed to elevated hydrostatic pressure for 24 hours in the presence of glutamate or glutamate receptor antagonists and examined histologically. GS activity was assessed by colorimetric assay. Results. Pressure elevation induced axonal swelling in the nerve fiber layer. Axonal swelling was prevented by a combination of non-N-methyl-d-aspartate (non-NMDA) receptor antagonist and an NMDA receptor antagonist, indicating that the damage results from activation of both types of glutamate receptor. When glial function was preserved, the typical changes induced by glutamate consisted of reversible Müller cell swelling resulting from excessive glial glutamate uptake. The irreversible Müller cell swelling in hyperbaric conditions may indicate that pressure disrupts glutamate metabolism. Indeed, elevated pressure inhibited GS activity. In addition, glutamate exposure after termination of pressure exposure exhibited apparent Müller cell swelling. Conclusions. These results suggest that the neural degeneration observed during pressure elevation is caused by impaired glial glutamate metabolism after uptake. PMID:20688725

Quality-Assurance Plan for Water-Quality Activities of the U.S. Geological Survey in Miami, Florida

DTIC Science & Technology

2003-01-01

Jacqueline Lima GS-12 GS-10 GS-09 GS-09 GS-08 GS-07 GS-06 GS-06 GS-04 GS-04Eleanor Seymore Surface-Water Data Collections & Jon Woolverton Jorge Agis...Thomas J. Smith, III Gordon Anderson Kevin Whelan Don De Angelis Allison Snow GS-13 GS-07 GS-05 GS-12 GS-13 GS-07 GS-12 GS-13 GS-09 GS-07 GS-14 GS-07...fan, active, inactive, inventory only playa , month day year stream channel, digital rec- order, North American Datum of 1927 North American Datum of

Clinical Investigation Program. Annual Progress Report. Volume 1

DTIC Science & Technology

1994-01-20

Suport Labs Resch Chemist 13 0644 GS Salata, KF Allergy Microbiologist 12 0403 CS Billups, L Flow Cytom Microbiologist 12 0403 GS Dobek, AS Inf Disease 5...continued to increase laboratory research support to principal investigators throughout the medical center. The DCI Flow Cytometry Laboratory provided...Kalman PhD. Mitogen-Inducible T Suppressor Cell 12 Assay by Flow Cytometry (12/89) * Reference is to page number(s) in Volume II. 30 PROTOCOL NUMBER

Gastric submucosa is inferior to the liver as transplant site for autologous islet transplantation in pancreatectomized diabetic Beagles.

PubMed

Yin, Zhu-Zeng; Wang, Shu-Sen; Li, Qiang; Huang, Ying; Chen, Li; Chen, Gang; Liu, Rong; Wang, Xi-Mo

2016-08-01

Intraportal transplantation of islets is no longer considered to be an ideal procedure and finding the extrahepatic alternative site is becoming a subject of high priority. Herein, in this study, we would introduce our initial outcomes of using gastric submucosa (GS) and liver as sites of islet autotransplantation in pancreatectomized diabetic Beagles. Total pancreatectomy was performed in Beagles and then their own islets extracted from the excised pancreas were transplanted into GS (GS group, n=8) or intrahepatic via portal vein (PV group, n=5). Forty-eight hours post transplantation, graft containing tissue harvested from the recipients revealed the presence of insulin-positive cells. All recipients in GS group achieved euglycemia within 1 day, but returned to a diabetic state at 6 to 8 days post-transplantation (mean survival time, 7.16±0.69 days). However, all of the animals kept normoglycemic until 85 to 155 days post-transplantation in PV group (mean survival time, 120±28.58 days; P<0.01 vs. GS group). The results of intravenous glucose tolerance test (IVGTT) confirmed that the marked improvement in glycometabolism was obtained in intrahepatic islet autotransplantation. Thus, our findings indicate that the liver is still superior to the GS as the site of islet transplantation, at least in our islet autotransplant model in pancreatectomized diabetic Beagles.

Albumin/gentamicin microspheres produced by supercritical assisted atomization: optimization of size, drug loading and release.

PubMed

Della Porta, G; Adami, R; Del Gaudio, P; Prota, L; Aquino, R; Reverchon, E

2010-11-01

In this work, the supercritical assisted atomization (SAA) is proposed, for the first time, not only as a micronization technology but also as a thermal coagulation process for the production of bovine serum albumin (BSA) microspheres charged with Gentamicin sulfate (GS). Particularly, different water solutions of BSA/GS were processed by SAA to produce protein microspheres with different size and antibiotic content. SAA precipitation temperature was selected in the range 100-130 °C to generate protein coagulation and to recover micronized BSA in form of hydrophobic aggregates; GS loading was varied between 10% and 50% (w/w) with an encapsulation efficiency which often reached 100%. In all cases, spherical and noncoalescing particles were successfully produced with a mean particle size of 2 µm and with a standard deviation of about ±1 µm. The microspheres also showed a good stability and constant water content after 60 days of storage. The release profiles of the entrapped drug were monitored using Franz cells to evaluate the possible application of the produced microspheres in wound dressing formulations. Particularly, the microspheres with a BSA/GS ratio of 4:1 after the first burst effect (of 40% of GS loaded) were able to release the GS continuously over 10 days. © 2010 Wiley-Liss, Inc. and the American Pharmacists Association

A Novel Regulatory Function of Sweet Taste-Sensing Receptor in Adipogenic Differentiation of 3T3-L1 Cells

PubMed Central

Masubuchi, Yosuke; Nakagawa, Yuko; Ma, Jinhui; Sasaki, Tsutomu; Kitamura, Tadahiro; Yamamoto, Yoritsuna; Kurose, Hitoshi; Kojima, Itaru; Shibata, Hiroshi

2013-01-01

Background Sweet taste receptor is expressed not only in taste buds but also in nongustatory organs such as enteroendocrine cells and pancreatic beta-cells, and may play more extensive physiological roles in energy metabolism. Here we examined the expression and function of the sweet taste receptor in 3T3-L1 cells. Methodology/Principal Findings In undifferentiated preadipocytes, both T1R2 and T1R3 were expressed very weakly, whereas the expression of T1R3 but not T1R2 was markedly up-regulated upon induction of differentiation (by 83.0 and 3.8-fold, respectively at Day 6). The α subunits of Gs (Gαs) and G14 (Gα14) but not gustducin were expressed throughout the differentiation process. The addition of sucralose or saccharin during the first 48 hours of differentiation considerably reduced the expression of peroxisome proliferator activated receptor γ (PPARγ and CCAAT/enhancer-binding protein α (C/EBPα at Day 2, the expression of aP2 at Day 4 and triglyceride accumulation at Day 6. These anti-adipogenic effects were attenuated by short hairpin RNA-mediated gene-silencing of T1R3. In addition, overexpression of the dominant-negative mutant of Gαs but not YM-254890, an inhibitor of Gα14, impeded the effects of sweeteners, suggesting a possible coupling of Gs with the putative sweet taste-sensing receptor. In agreement, sucralose and saccharin increased the cyclic AMP concentration in differentiating 3T3-L1 cells and also in HEK293 cells heterologously expressing T1R3. Furthermore, the anti-adipogenic effects of sweeteners were mimicked by Gs activation with cholera toxin but not by adenylate cyclase activation with forskolin, whereas small interfering RNA-mediated knockdown of Gαs had the opposite effects. Conclusions 3T3-L1 cells express a functional sweet taste-sensing receptor presumably as a T1R3 homomer, which mediates the anti-adipogenic signal by a Gs-dependent but cAMP-independent mechanism. PMID:23336004

p97/VCP promotes degradation of CRBN substrate glutamine synthetase and neosubstrates

PubMed Central

Nguyen, Thang Van; Li, Jing; Lu, Chin-Chun (Jean); Mamrosh, Jennifer L.; Lu, Gang; Cathers, Brian E.; Deshaies, Raymond J.

2017-01-01

Glutamine synthetase (GS) plays an essential role in metabolism by catalyzing the synthesis of glutamine from glutamate and ammonia. Our recent study showed that CRBN, a direct protein target for the teratogenic and antitumor activities of immunomodulatory drugs such as thalidomide, lenalidomide, and pomalidomide, recognizes an acetyl degron of GS, resulting in ubiquitylation and degradation of GS in response to glutamine. Here, we report that valosin-containing protein (VCP)/p97 promotes the degradation of ubiquitylated GS, resulting in its accumulation in cells with compromised p97 function. Notably, p97 is also required for the degradation of all four known CRBN neo-substrates [Ikaros family zinc finger proteins 1 (IKZF1) and 3 (IKZF3), casein kinase 1α (CK1α), and the translation termination factor GSPT1] whose ubiquitylation is induced by immunomodulatory drugs. Together, these data point to an unexpectedly intimate relationship between the E3 ubiquitin ligase CRL4CRBN and p97 pathways. PMID:28320958

Types of Stem Cells

MedlinePlus

... Cell Glossary Search Toggle Nav Types of Stem Cells Stem cells are the foundation from which all ... About Stem Cells > Types of Stem Cells Stem cells Stem cells are the foundation for every organ ...

The Influence of Thyroid-Stimulating Hormone and Thyroid-Stimulating Hormone Receptor Antibodies on Osteoclastogenesis

PubMed Central

Morshed, Syed; Latif, Rauf; Zaidi, Mone; Davies, Terry F.

2011-01-01

Background We have shown that thyroid-stimulating hormone (TSH) has a direct inhibitory effect on osteoclastic bone resorption and that TSH receptor (TSHR) null mice display osteoporosis. To determine the stage of osteoclast development at which TSH may exert its effect, we examined the influence of TSH and agonist TSHR antibodies (TSHR-Ab) on osteoclast differentiation from murine embryonic stem (ES) cells to gain insight into bone remodeling in hyperthyroid Graves' disease. Methods Osteoclast differentiation was initiated in murine ES cell cultures through exposure to macrophage colony stimulation factor, receptor activator of nuclear factor кB ligand, vitamin D, and dexamethasone. Results Tartrate resistant acid phosphatase (TRAP)-positive osteoclasts formed in ∼12 days. This coincided with the expected downregulation of known markers of self renewal and pluripotency (including Oct4, Sox2, and REX1). Both TSH and TSHR-Abs inhibited osteoclastogenesis as evidenced by decreased development of TRAP-positive cells (∼40%–50% reduction, p = 0.0047), and by decreased expression, in a concentration-dependent manner, of osteoclast differentiation markers (including the calcitonin receptor, TRAP, cathepsin K, matrix metallo-proteinase-9, and carbonic anhydrase II). Similar data were obtained using serum immunoglobulin-Gs (IgGs) from patients with hyperthyroid Graves' disease and known TSHR-Abs. TSHR stimulators inhibited tumor necrosis factor-alpha mRNA and protein expression, but increased the expression of osteoprotegerin (OPG), an antiosteoclastogenic human soluble receptor activator of nuclear factor кB ligand receptor. Neutralizing antibody to OPG reversed the inhibitory effect of TSH on osteoclast differentiation evidencing that the TSH effect was at least in part mediated by increased OPG. Conclusion These data establish ES-derived osteoclastogenesis as an effective model system to study the regulation of osteoclast differentiation in early development. The results support the observations that TSH has a bone protective action by negatively regulating osteoclastogenesis. Further, our results implicate TSHR-Abs in offering skeletal protection in hyperthyroid Graves' disease, even in the face of high thyroid hormone and low TSH levels. PMID:21745106

Stomatal conductance, mesophyll conductance, and transpiration efficiency in relation to leaf anatomy in rice and wheat genotypes under drought.

PubMed

Ouyang, Wenjing; Struik, Paul C; Yin, Xinyou; Yang, Jianchang

2017-11-02

Increasing leaf transpiration efficiency (TE) may provide leads for growing rice like dryland cereals such as wheat (Triticum aestivum). To explore avenues for improving TE in rice, variations in stomatal conductance (gs) and mesophyll conductance (gm) and their anatomical determinants were evaluated in two cultivars from each of lowland, aerobic, and upland groups of Oryza sativa, one cultivar of O. glaberrima, and two cultivars of T. aestivum, under three water regimes. The TE of upland rice, O. glaberrima, and wheat was more responsive to the gm/gs ratio than that of lowland and aerobic rice. Overall, the explanatory power of the particular anatomical trait varied among species. Low stomatal density mostly explained the low gs in drought-tolerant rice, whereas rice genotypes with smaller stomata generally responded more strongly to drought. Compared with rice, wheat had a higher gm, which was associated with thicker mesophyll tissue, mesophyll and chloroplasts more exposed to intercellular spaces, and thinner cell walls. Upland rice, O. glaberrima, and wheat cultivars minimized the decrease in gm under drought by maintaining high ratios of chloroplasts to exposed mesophyll cell walls. Rice TE could be improved by increasing the gm/gs ratio via modifying anatomical traits. © The Author 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology.

miRNA-regulated cancer stem cells: understanding the property and the role of miRNA in carcinogenesis.

PubMed

Chakraborty, Chiranjib; Chin, Kok-Yong; Das, Srijit

2016-10-01

Over the last few years, microRNAs (miRNA)-controlled cancer stem cells have drawn enormous attention. Cancer stem cells are a small population of tumor cells that possess the stem cell property of self-renewal. Recent data shows that miRNA regulates this small population of stem cells. In the present review, we explained different characteristics of cancer stem cells as well as miRNA regulation of self-renewal and differentiation in cancer stem cells. We also described the migration and tumor formation. Finally, we described the different miRNAs that regulate various types of cancer stem cells, such as prostate cancer stem cells, head and neck cancer stem cells, breast cancer stem cells, colorectal cancer stem cells, lung cancer stem cells, gastric cancer stem cells, pancreatic cancer stem cells, etc. Extensive research is needed in order to employ miRNA-based therapeutics to control cancer stem cell population in various cancers in the future.

What is a stem cell?

PubMed

Slack, Jonathan M W

2018-05-15

The historical roots of the stem cell concept are traced with respect to its usage in embryology and in hematology. The modern consensus definition of stem cells, comprising both pluripotent stem cells in culture and tissue-specific stem cells in vivo, is explained and explored. Methods for identifying stem cells are discussed with respect to cell surface markers, telomerase, label retention and transplantability, and properties of the stem cell niche are explored. The CreER method for identifying stem cells in vivo is explained, as is evidence in favor of a stochastic rather than an obligate asymmetric form of cell division. In conclusion, it is found that stem cells do not possess any unique and specific molecular markers; and stem cell behavior depends on the environment of the cell as well as the stem cell's intrinsic qualities. Furthermore, the stochastic mode of division implies that stem cell behavior is a property of a cell population not of an individual cell. In this sense, stem cells do not exist in isolation but only as a part of multicellular system. This article is categorized under: Adult Stem Cells, Tissue Renewal, and Regeneration > Tissue Stem Cells and Niches Adult Stem Cells, Tissue Renewal, and Regeneration > Methods and Principles Adult Stem Cells, Tissue Renewal, and Regeneration > Environmental Control of Stem Cells. © 2018 Wiley Periodicals, Inc.

Increasing Minority Participation and Matriculation in the Geosciences at El Paso Community College

NASA Astrophysics Data System (ADS)

Villalobos, J. I.

2011-12-01

Community colleges currently serve 44% of all undergraduate students and 45% of all of all first time freshmen in the US. Hispanics now constitute 15% of the general population and 19% of the college population in the US. This increase has led to more institutions emerging as HSI (Hispanic Serving Institution) by the federal government. These facts illustrate the potential community colleges hold to encourage STEM (Science Technology Engineering and Math) majors to minorities as well as non-minorities. But the reality is the number of STEM degrees awarded at community colleges has not followed the same trends in enrollment. El Paso Community College (EPCC) currently enrolls 27,000 students with 85% of the student body being Hispanic. More than 130 programs of study are offered including an Associate of Science degree in Geological Sciences. Over the past three years we have implemented several initiatives in our effort to increase the number of Geological Science (GS) majors at EPCC. These efforts are aimed to decrease attrition rates of science majors by; streamlining the GS degree plan along with the process of course registration, introduce field-based research projects to students to allow hands on research, develop a work relationships with students and university faculty, increase the number of geology courses offered at EPCC including a field-based capstone course (GEOL 2407- Geological Field Methods), and strengthening the educational-bridge between the geological science departments of EPCC and University of Texas at El Paso.

Confocal and dermoscopic features of basal cell carcinoma in Gorlin-Goltz syndrome: A case report.

PubMed

Casari, Alice; Argenziano, Giuseppe; Moscarella, Elvira; Lallas, Aimilios; Longo, Caterina

2017-05-01

Gorlin-Goltz (GS) syndrome is an autosomal dominant disease linked to a mutation in the PTCH gene. Major criteria include the onset of multiple basal cell carcinoma (BCC), keratocystic odontogenic tumours in the jaws and bifid ribs. Dermoscopy and reflectance confocal microscopy represent imaging tools that are able to increase the diagnostic accuracy of skin cancer in a totally noninvasive manner, without performing punch biopsies. Here we present a case of a young woman in whom the combined approach of dermoscopy and RCM led to the identification of multiple small inconspicuous lesions as BCC and thus to the diagnosis of GS syndrome. © 2016 The Australasian College of Dermatologists.

Complexity and Selectivity of γ-Secretase Cleavage on Multiple Substrates: Consequences in Alzheimer's Disease and Cancer.

PubMed

Medoro, Alessandro; Bartollino, Silvia; Mignogna, Donatella; Passarella, Daniela; Porcile, Carola; Pagano, Aldo; Florio, Tullio; Nizzari, Mario; Guerra, Germano; Di Marco, Roberto; Intrieri, Mariano; Raimo, Gennaro; Russo, Claudio

2018-01-01

The processing of the amyloid-β protein precursor (AβPP) by β- and γ-secretases is a pivotal event in the genesis of Alzheimer's disease (AD). Besides familial mutations on the AβPP gene, or upon its overexpression, familial forms of AD are often caused by mutations or deletions in presenilin 1 (PSEN1) and 2 (PSEN2) genes: the catalytic components of the proteolytic enzyme γ-secretase (GS). The "amyloid hypothesis", modified over time, states that the aberrant processing of AβPP by GS induces the formation of specific neurotoxic soluble amyloid-β (Aβ) peptides which, in turn, cause neurodegeneration. This theory, however, has recently evidenced significant limitations and, in particular, the following issues are debated: 1) the concept and significance of presenilin's "gain of function" versus "loss of function"; and 2) the presence of several and various GS substrates, which interact with AβPP and may influence Aβ formation. The latter consideration is suggestive: despite the increasing number of GS substrates so far identified, their reciprocal interaction with AβPP itself, even in the AD field, is significantly unexplored. On the other hand, GS is also an important pharmacological target in the cancer field; inhibitors or GS activity are investigated in clinical trials for treating different tumors. Furthermore, the function of AβPP and PSENs in brain development and in neuronal migration is well known. In this review, we focused on a specific subset of GS substrates that directly interact with AβPP and are involved in its proteolysis and signaling, by evaluating their role in neurodegeneration and in cell motility or proliferation, as a possible connection between AD and cancer.

Improved fatigue resistance in Gsα-deficient and aging mouse skeletal muscles due to adaptive increases in slow fibers

PubMed Central

Feng, Han-Zhong; Chen, Min; Weinstein, Lee S.

2011-01-01

Genetically modified mice with deficiency of the G protein α-subunit (Gsα) in skeletal muscle showed metabolic abnormality with reduced glucose tolerance, low muscle mass, and low contractile force, along with a fast-to-slow-fiber-type switch (Chen M, Feng HZ, Gupta D, Kelleher J, Dickerson KE, Wang J, Hunt D, Jou W, Gavrilova O, Jin JP, Weinstein LS. Am J Physiol Cell Physiol 296: C930–C940, 2009). Here we investigated a hypothesis that the switching to more slow fibers is an adaptive response with specific benefit. The results showed that, corresponding to the switch of myosin isoforms, the thin-filament regulatory proteins troponin T and troponin I both switched to their slow isoforms in the atrophic soleus muscle of 3-mo-old Gsα-deficient mice. This fiber-type switch involving coordinated changes of both thick- and thin-myofilament proteins progressed in the Gsα-deficient soleus muscles of 18- to 24-mo-old mice, as reflected by the expression of solely slow isoforms of myosin and troponin. Compared with age-matched controls, Gsα-deficient soleus muscles with higher proportion of slow fibers exhibited slower contractile and relaxation kinetics and lower developed force, but significantly increased resistance to fatigue, followed by a better recovery. Gsα-deficient soleus muscles of neonatal and 3-wk-old mice did not show the increase in slow fibers. Therefore, the fast-to-slow-fiber-type switch in Gsα deficiency at older ages was likely an adaptive response. The benefit of higher fatigue resistance in adaption to metabolic deficiency and aging provides a mechanism to sustain skeletal muscle function in diabetic patients and elderly individuals. PMID:21680879

Involvement of indole-3-acetic acid produced by Azospirillum brasilense in accumulating intracellular ammonium in Chlorella vulgaris.

PubMed

Meza, Beatriz; de-Bashan, Luz E; Bashan, Yoav

2015-01-01

Accumulation of intracellular ammonium and activities of the enzymes glutamine synthetase (GS) and glutamate dehydrogenase (GDH) were measured when the microalgae Chlorella vulgaris was immobilized in alginate with either of two wild type strains of Azospirillum brasilense or their corresponding indole-3-acetic acid (IAA)-attenuated mutants. After 48 h of immobilization, both wild types induced higher levels of intracellular ammonium in the microalgae than their respective mutants; the more IAA produced, the higher the intracellular ammonium accumulated. Accumulation of intracellular ammonium in the cells of C. vulgaris followed application of four levels of exogenous IAA reported for A. brasilense and its IAA-attenuated mutants, which had a similar pattern for the first 24 h. This effect was transient and disappeared after 48 h of incubation. Immobilization of C. vulgaris with any bacteria strain induced higher GS activity. The bacterial strains also had GS activity, comparable to the activity detected in C. vulgaris, but weaker than when immobilized with the bacteria. When net activity was calculated, the wild type always induced higher GS activity than IAA-attenuated mutants. GDH activity in most microalgae/bacteria interactions resembled GS activity. When complementing IAA-attenuated mutants with exogenous IAA, GS activity in co-immobilized cultures matched those of the wild type A. brasilense immobilized with the microalga. Similarity occurred when the net GS activity was measured, and was higher with greater quantities of exogenous IAA. It is proposed that IAA produced by A. brasilense is involved in ammonium uptake and later assimilation by C. vulgaris. Copyright © 2014 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.

Generation and analysis of the thiazide-sensitive Na+ -Cl- cotransporter (Ncc/Slc12a3) Ser707X knockin mouse as a model of Gitelman syndrome.

PubMed

Yang, Sung-Sen; Lo, Yi-Fen; Yu, I-Shing; Lin, Shu-Wha; Chang, Tai-Hsiang; Hsu, Yu-Juei; Chao, Tai-Kuang; Sytwu, Huey-Kang; Uchida, Shinichi; Sasaki, Sei; Lin, Shih-Hua

2010-12-01

Gitelman syndrome (GS) is characterized by salt-losing hypotension, hypomagnesemia, hypokalemic metabolic alkalosis, and hypocalciuria. To better model human GS caused by a specific mutation in the thiazide-sensitive Na(+) -Cl(-) cotransporter (NCC) gene SLC12A3, we generated a nonsense Ncc Ser707X knockin mouse corresponding to human p.Ser710X (c.2135C>A), a recurrent mutation with severe phenotypes in Chinese GS patients. Compared with wild-type or heterozygous littermates, homozygous (Hom) knockin mice fully recapitulated the phenotype of human GS. The markedly reduced Ncc mRNA and virtually absent Ncc protein expression in kidneys of Hom mice was primarily due to nonsense-mediated mRNA decay (NMD) surveillance mechanisms. Expression of epithelial Na(+) channel (Enac), Ca(2+) channels (Trpv5 and Trpv6), and K(+) channels (Romk1 and maxi-K) were significantly increased. Late distal convoluted tubules (DCT) volume was increased and DCT cell ultrastructure appeared intact. High K(+) intake could not correct hypokalemia but caused a further increase in maxi-K but not Romk1 expression. Renal tissue from a patient with GS also showed the enhanced TRPV5 and ROMK1 expression in distal tubules. We suggest that the upregulation of TRPV5/6 and of ROMK1 and Maxi-K may contribute to hypocalciuria and hypokalemia in Ncc Ser707X knockin mice and human GS, respectively. © 2010 Wiley-Liss, Inc.

A high cell density transient transfection system for therapeutic protein expression based on a CHO GS-knockout cell line: process development and product quality assessment.

PubMed

Rajendra, Yashas; Hougland, Maria D; Alam, Riazul; Morehead, Teresa A; Barnard, Gavin C

2015-05-01

Transient gene expression (TGE) is a rapid method for the production of recombinant proteins in mammalian cells. While the volumetric productivity of TGE has improved significantly over the past decade, most methods involve extensive cell line engineering and plasmid vector optimization in addition to long fed batch cultures lasting up to 21 days. Our colleagues have recently reported the development of a CHO K1SV GS-KO host cell line. By creating a bi-allelic glutamine synthetase knock out of the original CHOK1SV host cell line, they were able to improve the efficiency of generating high producing stable CHO lines for drug product manufacturing. We developed a TGE method using the same CHO K1SV GS-KO host cell line without any further cell line engineering. We also refrained from performing plasmid vector engineering. Our objective was to setup a TGE process to mimic protein quality attributes obtained from stable CHO cell line. Polyethyleneimine (PEI)-mediated transfections were performed at high cell density (4 × 10(6) cells/mL) followed by immediate growth arrest at 32 °C for 7 days. Optimizing DNA and PEI concentrations proved to be important. Interestingly, found the direct transfection method (where DNA and PEI were added sequentially) to be superior to the more common indirect method (where DNA and PEI are first pre-complexed). Moreover, the addition of a single feed solution and a polar solvent (N,N dimethylacetamide) significantly increased product titers. The scalability of process from 2 mL to 2 L was demonstrated using multiple proteins and multiple expression volumes. Using this simple, short, 7-day TGE process, we were able to successfully produce 54 unique proteins in a fraction of the time that would have been required to produce the respective stable CHO cell lines. The list of 54 unique proteins includes mAbs, bispecific antibodies, and Fc-fusion proteins. Antibody titers of up to 350 mg/L were achieved with the simple 7-day process. Titers were increased to 1 g/L by extending the culture to 16 days. We also present two case studies comparing product quality of material generated by transient HEK293, transient CHO K1SV GS-KO, and stable CHO K1SV KO pool. Protein from transient CHO was more representative of stable CHO protein compared to protein produced from HEK293. © 2014 Wiley Periodicals, Inc.

Impact of concurrent overexpression of cytosolic glutamine synthetase (GS1) and sucrose phosphate synthase (SPS) on growth and development in transgenic tobacco.

PubMed

Seger, Mark; Gebril, Sayed; Tabilona, Jules; Peel, Amanda; Sengupta-Gopalan, Champa

2015-01-01

The outcome of simultaneously increasing SPS and GS activities in transgenic tobacco, suggests that sucrose is the major determinant of growth and development, and is not affected by changes in N assimilation. Carbon (C) and nitrogen (N) are the major components required for plant growth and the metabolic pathways for C and N assimilation are very closely interlinked. Maintaining an appropriate balance or ratio of sugar to nitrogen metabolites in the cell, is important for the regulation of plant growth and development. To understand how C and N metabolism interact, we manipulated the expression of key genes in C and N metabolism individually and concurrently and checked for the repercussions. Transgenic tobacco plants with a cytosolic soybean glutamine synthetase (GS1) gene and a sucrose phosphate synthase (SPS) gene from maize, both driven by the CaMV 35S promoter were produced. Co-transformants, with both the transgenes were produced by sexual crosses. While GS is the key enzyme in N assimilation, involved in the synthesis of glutamine, SPS plays a key role in C metabolism by catalyzing the synthesis of sucrose. Moreover, to check if nitrate has any role in this interaction, the plants were grown under both low and high nitrogen. The SPS enzyme activity in the SPS and SPS/GS1 co-transformants were the same under both nitrogen regimens. However, the GS activity was lower in the co-transformants compared to the GS1 transformants, specifically under low nitrogen conditions. The GS1/SPS transformants showed a phenotype similar to the SPS transformants, suggesting that sucrose is the major determinant of growth and development in tobacco, and its effect is only marginally affected by increased N assimilation. Sucrose may be functioning in a metabolic capacity or as a signaling molecule.

Dirhamnolipids secreted from Pseudomonas aeruginosa modify anjpegungal susceptibility of Aspergillus fumigatus by inhibiting β1,3 glucan synthase activity

PubMed Central

Briard, Benoit; Rasoldier, Vero; Bomme, Perrine; ElAouad, Noureddine; Guerreiro, Catherine; Chassagne, Pierre; Muszkieta, Laetitia; Latgé, Jean-Paul; Mulard, Laurence; Beauvais, Anne

2017-01-01

Pseudomonas aeruginosa and Aspergillus fumigatus are the two microorganisms responsible for most of the chronic infections in cystic fibrosis patients. P. aeruginosa is known to produce quorum-sensing controlled rhamnolipids during chronic infections. Here we show that the dirhamnolipids secreted from P. aeruginosa (i) induce A. fumigatus to produce an extracellular matrix, rich in galactosaminogalactan, 1,8-dihydroxynaphthalene (DHN)- and pyo-melanin, surrounding their hyphae, which facilitates P. aeruginosa binding and (ii) inhibit A. fumigatus growth by blocking β1,3 glucan synthase (GS) activity, thus altering the cell wall architecture. A. fumigatus in the presence of diRhls resulted in a growth phenotype similar to that upon its treatment with anjpegungal echinocandins, showing multibranched hyphae and thicker cell wall rich in chitin. The diRhl structure containing two rhamnose moieties attached to fatty acyl chain is essential for the interaction with β1,3 GS; however, the site of action of diRhls on GS is different from that of echinocandins, and showed synergistic anjpegungal effect with azoles. PMID:28338676

Differential marker expression by cultures rich in mesenchymal stem cells

PubMed Central

2013-01-01

Background Mesenchymal stem cells have properties that make them amenable to therapeutic use. However, the acceptance of mesenchymal stem cells in clinical practice requires standardized techniques for their specific isolation. To date, there are no conclusive marker (s) for the exclusive isolation of mesenchymal stem cells. Our aim was to identify markers differentially expressed between mesenchymal stem cell and non-stem cell mesenchymal cell cultures. We compared and contrasted the phenotype of tissue cultures in which mesenchymal stem cells are rich and rare. By initially assessing mesenchymal stem cell differentiation, we established that bone marrow and breast adipose cultures are rich in mesenchymal stem cells while, in our hands, foreskin fibroblast and olfactory tissue cultures contain rare mesenchymal stem cells. In particular, olfactory tissue cells represent non-stem cell mesenchymal cells. Subsequently, the phenotype of the tissue cultures were thoroughly assessed using immuno-fluorescence, flow-cytometry, proteomics, antibody arrays and qPCR. Results Our analysis revealed that all tissue cultures, regardless of differentiation potential, demonstrated remarkably similar phenotypes. Importantly, it was also observed that common mesenchymal stem cell markers, and fibroblast-associated markers, do not discriminate between mesenchymal stem cell and non-stem cell mesenchymal cell cultures. Examination and comparison of the phenotypes of mesenchymal stem cell and non-stem cell mesenchymal cell cultures revealed three differentially expressed markers – CD24, CD108 and CD40. Conclusion We indicate the importance of establishing differential marker expression between mesenchymal stem cells and non-stem cell mesenchymal cells in order to determine stem cell specific markers. PMID:24304471

Granulocytic sarcoma in non-leukaemic child involving maxillary sinus with long term follow up: A rare case report.

PubMed

Sharma, Aman; Singh, Harkanwal Preet; Gupta, Anish Ashok; Garg, Parveen; Moon, Ninad Joshirao; Chavan, Rahul

2014-01-01

Granulocytic sarcoma (GS) is a rare extramedullary malignant tumor composed of immature myeloid cells. It is strongly associated with acute myeloid leukaemia, chronic myeloproliferative diseases. Occurrence of GS in the oral cavity is extremely uncommon. Present case reported an unusual occurrence of GS without leukemia involving maxillary sinus of a child. The patient underwent chemotherapy followed by radiotherapy with complete remission. A long-term follow-up of the patient was carried without any evidence of recurrence with special focus on diagnostic difficulties. The present case highlights the perplexity in diagnosing such lesions with emphasis on the need of careful interpretation of all clinical, radiographic, histopathological and immunohistochemical details as it is one of the most frequently misdiagnosed disorder.

Human placenta: relative content of antibodies of different classes and subclasses (IgG1-IgG4) containing lambda- and kappa-light chains and chimeric lambda-kappa-immunoglobulins.

PubMed

Lekchnov, Evgenii A; Sedykh, Sergey E; Dmitrenok, Pavel S; Buneva, Valentina N; Nevinsky, Georgy A

2015-06-01

The specific organ placenta is much more than a filter: it is an organ that protects, feeds and regulates the growth of the embryo. Affinity chromatography, ELISA, SDS-PAGE and matrix-assisted laser desorption ionization mass spectrometry were used. Using 10 intact human placentas deprived of blood, a quantitative analysis of average relative content [% of total immunoglobulins (Igs)] was carried out for the first time: (92.7), IgA (2.4), IgM (2.5), kappa-antibodies (51.4), lambda-antibodies (48.6), IgG1 (47.0), IgG2 (39.5), IgG3 (8.8) and IgG4 (4.3). It was shown for the first time that placenta contains sIgA (2.5%). In the classic paradigm, Igs represent products of clonal B-cell populations, each producing antibodies recognizing a single antigen. There is a common belief that IgGs in mammalian biological fluids are monovalent molecules having stable structures and two identical antigen-binding sites. However, similarly to human milk Igs, placenta antibodies undergo extensive half-molecule exchange and the IgG pool consists of 43.5 ± 15.0% kappa-kappa-IgGs and 41.6 ± 17.0% lambda-lambda-IgGs, while 15.0 ± 4.0% of the IgGs contained both kappa- and lambda-light chains. Kappa-kappa-IgGs and lambda-lambda-IgGs contained, respectively (%): IgG1 (47.7 and 34.4), IgG2 (36.3 and 44.5), IgG3 (7.4 and 11.8) and IgG4 (7.5 and 9.1), while chimeric kappa-lambda-IgGs consisted of (%): 43.5 IgG1, 41.0 IgG2, 5.6 IgG3 and 7.9 IgG4. Our data are indicative of the possibility of half-molecule exchange between placenta IgGs of various subclasses, raised against different antigens, which explains a very well-known polyspecificity and cross-reactivity of different human IgGs. © The Japanese Society for Immunology. 2015. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Improved stress tolerance and productivity in transgenic rice plants constitutively expressing the Oryza sativa glutathione synthetase OsGS under paddy field conditions.

PubMed

Park, Seong-Im; Kim, Young-Saeng; Kim, Jin-Ju; Mok, Ji-Eun; Kim, Yul-Ho; Park, Hyang-Mi; Kim, Il-Sup; Yoon, Ho-Sung

2017-08-01

Reactive oxygen species, which increase under various environmental stresses, have deleterious effects on plants. An important antioxidant, glutathione, is used to detoxify reactive oxygen species in plant cells and is mainly produced by two enzymes: gamma-glutamylcysteine synthetase (γ-ECS) and glutathione synthetase (GS). To evaluate the functional roles of the glutathione synthetase gene (OsGS) in rice, we generated four independent transgenic rice plants (TG1-TG4) that overexpressed OsGS under the control of the constitutively expressed OsCc1 promoter. When grown under natural paddy field conditions, the TG rice plants exhibited greater growth development, higher chlorophyll content, and higher GSH/GSSH ratios than control wild-type (WT) rice plants. Subsequently, the TG rice plants enhanced redox homeostasis by preventing hydroperoxide-mediated membrane damage, which improved their adaptation to environmental stresses. As a result, TG rice plants improved rice grain yield and total biomass following increases in panicle number and number of spikelets per panicle, despite differences in climate during the cultivation periods of 2014 and 2015. Overall, our results indicate that OsGS overexpression improved redox homeostasis by enhancing the glutathione pool, which resulted in greater tolerance to environmental stresses in the paddy fields. Copyright © 2017. Published by Elsevier GmbH.

Gorlin syndrome with an ovarian leiomyoma associated with a PTCH1 second hit.

PubMed

Akizawa, Yoshika; Miyashita, Toshiyuki; Sasaki, Ryo; Nagata, Reiko; Aoki, Ryoko; Ishitani, Ken; Nagashima, Yoji; Matsui, Hideo; Saito, Kayoko

2016-04-01

We describe a Gorlin syndrome (GS) case with two different second hit mutations of PTCH1, one in a keratocystic odontogenic tumor (KCOT) and the other in an ovarian leiomyoma. GS is a rare genetic condition manifesting as multiple basal cell nevi associated with other features such as medulloblastomas, skeletal abnormalities, and ovarian fibromas. A 21-year-old Japanese woman with a history of two KCOTs was diagnosed with GS according to clinical criteria. A PTCH1 mutation, c.1427del T, was detected in peripheral blood. A novel PTCH1 mutation, c.264_265insAATA, had been found in the maxillary KCOT as a second hit mutation. More recently, the ovarian tumor was detected during a gynecological examination. Laparoscopic adnexectomy was performed, and the pathological diagnosis of the ovarian tumor was leiomyoma. Interestingly, another novel mutation, loss of heterozygosity spanning from 9q22.32 to 9q31.2, including PTCH1 and 89 other genes, was detected in this ovarian tumor, providing evidence of a second hit mutation. This is the first report describing a GS-associated ovarian tumor carrying a second hit in the PTCH1 region. We anticipate that accumulation of more cases will clarify the importance of second hit mutations in ovarian tumor formation in GS. © 2016 Wiley Periodicals, Inc.

Amnionless function is required for cubilin brush-border expression and intrinsic factor-cobalamin (vitamin B12) absorption in vivo

PubMed Central

He, Qianchuan; Madsen, Mette; Kilkenney, Adam; Gregory, Brittany; Christensen, Erik I.; Vorum, Henrik; Højrup, Peter; Schäffer, Alejandro A.; Kirkness, Ewen F.; Tanner, Stephan M.; de la Chapelle, Albert; Giger, Urs; Moestrup, Søren K.; Fyfe, John C.

2005-01-01

Amnionless (AMN) and cubilin gene products appear to be essential functional subunits of an endocytic receptor called cubam. Mutation of either gene causes autosomal recessive Imerslund-Gräsbeck syndrome (I-GS, OMIM no. 261100) in humans, a disorder characterized by selective intestinal malabsorption of cobalamin (vitamin B12) and urinary loss of several specific low-molecular-weight proteins. Vital insight into the molecular pathology of I-GS has been obtained from studies of dogs with a similar syndrome. In this work, we show that I-GS segregates in a large canine kindred due to an in-frame deletion of 33 nucleotides in exon 10 of AMN. In a second, unrelated I-GS kindred, affected dogs exhibit a homozygous substitution in the AMN translation initiation codon. Studies in vivo demonstrated that both mutations abrogate AMN expression and block cubilin processing and targeting to the apical membrane. The essential features of AMN dysfunction observed in vivo are recapitulated in a heterologous cell-transfection system, thus validating the system for analysis of AMN-cubilin interactions. Characterization of canine AMN mutations that cause I-GS establishes the canine model as an ortholog of the human disorder well suited to studies of AMN function and coevolution with cubilin. (Blood. 2005;106:1447-1453) PMID:15845892

Learn About Stem Cells

MedlinePlus

... Handbook Stem Cell Glossary Search Toggle Nav Stem Cell Basics Stem cells are the foundation from which ... Home > Learn About Stem Cells > Stem Cell Basics Cells in the human body The human body comprises ...

Erythroid differentiation of human induced pluripotent stem cells is independent of donor cell type of origin.

PubMed

Dorn, Isabel; Klich, Katharina; Arauzo-Bravo, Marcos J; Radstaak, Martina; Santourlidis, Simeon; Ghanjati, Foued; Radke, Teja F; Psathaki, Olympia E; Hargus, Gunnar; Kramer, Jan; Einhaus, Martin; Kim, Jeong Beom; Kögler, Gesine; Wernet, Peter; Schöler, Hans R; Schlenke, Peter; Zaehres, Holm

2015-01-01

Epigenetic memory in induced pluripotent stem cells, which is related to the somatic cell type of origin of the stem cells, might lead to variations in the differentiation capacities of the pluripotent stem cells. In this context, induced pluripotent stem cells from human CD34(+) hematopoietic stem cells might be more suitable for hematopoietic differentiation than the commonly used fibroblast-derived induced pluripotent stem cells. To investigate the influence of an epigenetic memory on the ex vivo expansion of induced pluripotent stem cells into erythroid cells, we compared induced pluripotent stem cells from human neural stem cells and human cord blood-derived CD34(+) hematopoietic stem cells and evaluated their potential for differentiation into hematopoietic progenitor and mature red blood cells. Although genome-wide DNA methylation profiling at all promoter regions demonstrates that the epigenetic memory of induced pluripotent stem cells is influenced by the somatic cell type of origin of the stem cells, we found a similar hematopoietic induction potential and erythroid differentiation pattern of induced pluripotent stem cells of different somatic cell origin. All human induced pluripotent stem cell lines showed terminal maturation into normoblasts and enucleated reticulocytes, producing predominantly fetal hemoglobin. Differences were only observed in the growth rate of erythroid cells, which was slightly higher in the induced pluripotent stem cells derived from CD34(+) hematopoietic stem cells. More detailed methylation analysis of the hematopoietic and erythroid promoters identified similar CpG methylation levels in the induced pluripotent stem cell lines derived from CD34(+) cells and those derived from neural stem cells, which confirms their comparable erythroid differentiation potential. Copyright© Ferrata Storti Foundation.

Effects of photodynamic therapy on dermal fibroblasts from xeroderma pigmentosum and Gorlin-Goltz syndrome patients.

PubMed

Zamarrón, Alicia; García, Marta; Río, Marcela Del; Larcher, Fernando; Juarranz, Ángeles

2017-09-29

PDT is widely applied for the treatment of non-melanoma skin cancer pre-malignant and malignant lesions (actinic keratosis, basal cell carcinoma and in situ squamous cell carcinoma). In photodynamic therapy (PDT) the interaction of a photosensitizer (PS), light and oxygen leads to the formation of reactive oxygen species (ROS) and thus the selective tumor cells eradication. Xeroderma pigmentosum (XP) and Gorlin-Goltz Syndrome (GS) patients are at high risk of developing skin cancer in sun-exposed areas. Therefore, the use of PDT as a preventive treatment may constitute a very promising therapeutic modality for these syndromes. Given the demonstrated role of cancer associated fibroblasts (CAFs) in tumor progression and the putative CAFs features of some cancer-prone genodermatoses fibroblasts, in this study, we have further characterized the phenotype of XP and GS dermal fibroblasts and evaluated their response to methyl-δ-aminolevulinic acid (MAL)-PDT compared to that of dermal fibroblasts obtained from healthy donors. We show here that XP/GS fibroblasts display clear features of CAFs and present a significantly higher response to PDT, even after being stimulated with UV light, underscoring the value of this therapeutic approach for these rare skin conditions and likely to other forms of skin cancer were CAFs play a major role.

Effects of photodynamic therapy on dermal fibroblasts from xeroderma pigmentosum and Gorlin-Goltz syndrome patients

PubMed Central

Zamarrón, Alicia; García, Marta; Río, Marcela Del; Larcher, Fernando; Juarranz, Ángeles

2017-01-01

PDT is widely applied for the treatment of non-melanoma skin cancer pre-malignant and malignant lesions (actinic keratosis, basal cell carcinoma and in situ squamous cell carcinoma). In photodynamic therapy (PDT) the interaction of a photosensitizer (PS), light and oxygen leads to the formation of reactive oxygen species (ROS) and thus the selective tumor cells eradication. Xeroderma pigmentosum (XP) and Gorlin-Goltz Syndrome (GS) patients are at high risk of developing skin cancer in sun-exposed areas. Therefore, the use of PDT as a preventive treatment may constitute a very promising therapeutic modality for these syndromes. Given the demonstrated role of cancer associated fibroblasts (CAFs) in tumor progression and the putative CAFs features of some cancer-prone genodermatoses fibroblasts, in this study, we have further characterized the phenotype of XP and GS dermal fibroblasts and evaluated their response to methyl-δ-aminolevulinic acid (MAL)-PDT compared to that of dermal fibroblasts obtained from healthy donors. We show here that XP/GS fibroblasts display clear features of CAFs and present a significantly higher response to PDT, even after being stimulated with UV light, underscoring the value of this therapeutic approach for these rare skin conditions and likely to other forms of skin cancer were CAFs play a major role. PMID:29100394

[Progress in stem cells and regenerative medicine].

PubMed

Wang, Libin; Zhu, He; Hao, Jie; Zhou, Qi

2015-06-01

Stem cells have the ability to differentiate into all types of cells in the body and therefore have great application potential in regenerative medicine, in vitro disease modelling and drug screening. In recent years, stem cell technology has made great progress, and induced pluripotent stem cell technology revolutionizes the whole stem cell field. At the same time, stem cell research in our country has also achieved great progress and becomes an indispensable power in the worldwide stem cell research field. This review mainly focuses on the research progress in stem cells and regenerative medicine in our country since the advent of induced pluripotent stem cell technology, including induced pluripotent stem cells, transdifferentiation, haploid stem cells, and new gene editing tools.

Application of Graphene Based Nanotechnology in Stem Cells Research.

PubMed

Hu, Shanshan; Zeng, Yongxiang; Yang, Shuying; Qin, Han; Cai, He; Wang, Jian

2015-09-01

The past several years have witnessed significant advances in stem cell therapy, tissue engineering and regenerative medicine. Graphene, with its unique properties such as high electrical conductivity, elasticity and good molecule absorption, have potential for creating the next generation of biomaterials. This review summarizes the interrelationship between graphene and stem cells. The analysis of graphene when applied on mesenchymal stem cells, neural stem cells, induced pluripotent stem cells, embryonic stem cells, periodontal ligament stem cells, human adipose-derived stem cells and cancer stem cells, and how graphene influences cell behavior and differentiation are discussed in details.

A revisionist history of adult marrow stem cell biology or 'they forgot about the discard'.

PubMed

Quesenberry, P; Goldberg, L

2017-08-01

The adult marrow hematopoietic stem cell biology has largely been based on studies of highly purified stem cells. This is unfortunate because during the stem cell purification the great bulk of stem cells are discarded. These cells are actively proliferating. The final purified stem cell is dormant and not representative of the whole stem cell compartment. Thus, a large number of studies on the cellular characteristics, regulators and molecular details of stem cells have been carried on out of non-represented cells. Niche studies have largely pursued using these purified stem cells and these are largely un-interpretable. Other considerations include the distinction between baseline and transplant stem cells and the modulation of stem cell phenotype by extracellular vesicles, to cite a non-inclusive list. Work needs to proceed on characterizing the true stem cell population.

Perspectives on stem cell therapy for cardiac regeneration. Advances and challenges.

PubMed

Choi, Sung Hyun; Jung, Seok Yun; Kwon, Sang-Mo; Baek, Sang Hong

2012-01-01

Ischemic heart disease (IHD) accelerates cardiomyocyte loss, but the developing stem cell research could be useful for regenerating a variety of tissue cells, including cardiomyocytes. Diverse sources of stem cells for IHD have been reported, including embryonic stem cells, induced pluripotent stem cells, skeletal myoblasts, bone marrow-derived stem cells, mesenchymal stem cells, and cardiac stem cells. However, stem cells have unique advantages and disadvantages for cardiac tissue regeneration, which are important considerations in determining the specific cells for improving cell survival and long-term engraftment after transplantation. Additionally, the dosage and administration method of stem cells need to be standardized to increase stability and efficacy for clinical applications. Accordingly, this review presents a summary of the stem cell therapies that have been studied for cardiac regeneration thus far, and discusses the direction of future cardiac regeneration research for stem cells.

Influence of arbuscular mycorrhizal fungi and treated wastewater on water relations and leaf structure alterations of Viburnum tinus L. plants during both saline and recovery periods.

PubMed

Gómez-Bellot, María José; Nortes, Pedro Antonio; Ortuño, María Fernanda; Romero, Cristina; Fernández-García, Nieves; Sánchez-Blanco, María Jesús

2015-09-01

Nowadays, irrigation with low quality water is becoming an alternative to satisfy the needs of crops. However, some plant species have to deal with high salinity of reclaimed water, by adapting their physiological behaviour during both saline and recovery periods and developing morphological changes in their leaves. The application of arbuscular mycorrhizal fungi (AMF) could also be a suitable option to mitigate the negative effects of this kind of water, although the effectiveness of plant-AMF association is influenced by many factors. In this work, during forty weeks, the combined effect of Glomus iranicum var. tenuihypharum and two types of water: control, C, EC<0.9 dS m(-1) and reclaimed water, RW (with EC: 4 dS m(-1) during a first saline period and EC: 6 dS m(-1) during a second saline period) was evaluated for laurustinus plants (Viburnum tinus L.) transplanted in soil. This was followed by a recovery period of eight weeks, when all the plants were irrigated in the control irrigation conditions. Seasonal and daily changes in stem water potential (Ψstem), stomatal conductance (gs), photosynthesis (Pn) and leaf internal CO2 concentration (Ci) of laurustinus plants were evaluated. Leaf structure alterations, nutrient imbalance, height and leaf hydraulic conductivity (Kleaf) were also determined. Due to the high difficulty of absorbing water from the soil, RW plants showed a high volumetric water content (θv) in soil. The stem water potential and the stomatal conductance (gs) values were reduced in RW plants throughout the second saline period. These decreases were also found during the day. Leaf Ca(2+)/Na(+) and K(+)/Na(+) ratios diminished in RW plants respect to the C plants due to the Na(+) accumulation, although height and chlorophyll content values did not show statistical differences. Leaves from RW plants showed a significantly thicker mesophyll than Control leaves as a consequence of high EC. The area of palisade parenchyma (PP) increased while the area of spongy parenchyma (SP) decreased in RW leaves with respect to the C leaves. These structural changes could be considered as a strategy to maximize photosynthesis potential in saline conditions. Mycorrhizal inoculation improved the water status of both C and RW plants by increasing their Ψstem and gs values. As regards leaf structure, AMF showed an opposite effect to salinity for PP and SP. At the end of the recovery period, hardly any statistical differences of physiological parameters were found between treatments, although a tendency to improve them was observed in inoculated plants. In any case, the leaf structural changes and the great reduction in Kleaf observed at Ψleaf below -1.5 MPa would constitute an important mechanism for laurustinus plants to reduce the water loses produced by salinity. Copyright © 2015 Elsevier GmbH. All rights reserved.

Stem Cells

MedlinePlus

Stem cells are cells with the potential to develop into many different types of cells in the body. They serve as a repair ... body. There are two main types of stem cells: embryonic stem cells and adult stem cells. Stem ...

The Role of Integrin α6 (CD49f) in Stem Cells: More than a Conserved Biomarker.

PubMed

Krebsbach, Paul H; Villa-Diaz, Luis G

2017-08-01

Stem cells have the capacity for self-renewal and differentiation into specialized cells that form and repopulated all tissues and organs, from conception to adult life. Depending on their capacity for differentiation, stem cells are classified as totipotent (ie, zygote), pluripotent (ie, embryonic stem cells), multipotent (ie, neuronal stem cells, hematopoietic stem cells, epithelial stem cells, etc.), and unipotent (ie, spermatogonial stem cells). Adult or tissue-specific stem cells reside in specific niches located in, or nearby, their organ or tissue of origin. There, they have microenvironmental support to remain quiescent, to proliferate as undifferentiated cells (self-renewal), and to differentiate into progenitors or terminally differentiated cells that migrate from the niche to perform specialized functions. The presence of proteins at the cell surface is often used to identify, classify, and isolate stem cells. Among the diverse groups of cell surface proteins used for these purposes, integrin α6, also known as CD49f, may be the only biomarker commonly found in more than 30 different populations of stem cells, including some cancer stem cells. This broad expression among stem cell populations indicates that integrin α6 may play an important and conserved role in stem cell biology, which is reaffirmed by recent demonstrations of its role maintaining self-renewal of pluripotent stem cells and breast and glioblastoma cancer stem cells. Therefore, this review intends to highlight and synthesize new findings on the importance of integrin α6 in stem cell biology.

Scaling Sap Flow Results Over Wide Areas Using High-Resolution Aerial Multispectral Digital Imaging, Leaf Area Index (LAI) and MODIS Satellite Imagery in Saltcedar Stands on the Lower Colorado River

NASA Astrophysics Data System (ADS)

Murray, R.; Neale, C.; Nagler, P. L.; Glenn, E. P.

2008-12-01

Heat-balance sap flow sensors provide direct estimates of water movement through plant stems and can be used to accurately measure leaf-level transpiration (EL) and stomatal conductance (GS) over time scales ranging from 20-minutes to a month or longer in natural stands of plants. However, their use is limited to relatively small branches on shrubs or trees, as the gauged stem section needs to be uniformly heated by the heating coil to produce valid measurements. This presents a scaling problem in applying the results to whole plants, stands of plants, and larger landscape areas. We used high-resolution aerial multispectral digital imaging with green, red and NIR bands as a bridge between ground measurements of EL and GS, and MODIS satellite imagery of a flood plain on the Lower Colorado River dominated by saltcedar (Tamarix ramosissima). Saltcedar is considered to be a high-water-use plant, and saltcedar removal programs have been proposed to salvage water. Hence, knowledge of actual saltcedar ET rates is needed on western U.S. rivers. Scaling EL and GS to large landscape units requires knowledge of leaf area index (LAI) over large areas. We used a LAI model developed for riparian habitats on Bosque del Apache, New Mexico, to estimate LAI at our study site on the Colorado River. We compared the model estimates to ground measurements of LAI, determined with a Li-Cor LAI-2000 Plant Canopy Analyzer calibrated by leaf harvesting to determine Specific Leaf Area (SLA) (m2 leaf area per g dry weight leaves) of the different species on the floodplain. LAI could be adequately predicted from NDVI from aerial multispectral imagery and could be cross-calibrated with MODIS NDVI and EVI. Hence, we were able to project point measurements of sap flow and LAI over multiple years and over large areas of floodplain using aerial multispectral imagery as a bridge between ground and satellite data. The methods are applicable to riparian corridors throughout the western U.S.

Urinary exosomes in the diagnosis of Gitelman and Bartter syndromes.

PubMed

Corbetta, Samuele; Raimondo, Francesca; Tedeschi, Silvana; Syrèn, Marie-Louise; Rebora, Paola; Savoia, Andrea; Baldi, Lorenza; Bettinelli, Alberto; Pitto, Marina

2015-04-01

Gitelman syndrome (GS) and Bartter syndrome (BS) are hereditary salt-losing tubulopathies (SLTs) resulting from defects of renal proteins involved in electrolyte reabsorption, as for sodium-chloride cotransporter (NCC) and furosemide-sensitive sodium-potassium-chloride cotransporter (NKCC2) cotransporters, affected in GS and BS Type 1 patients, respectively. Currently, definitive diagnosis is obtained through expensive and time-consuming genetic testing. Urinary exosomes (UE), nanovesicles released by every epithelial cell facing the urinary space, represent an ideal source of markers for renal dysfunction and injury, because UE molecular composition stands for the cell of origin. On these assumptions, the aim of this work is to evaluate the relevance of UE for the diagnosis of SLTs. UE were purified from second morning urines collected from 32 patients with genetically proven SLTs (GS, BS1, BS2 and BS3 patients), 4 with unclassified SLTs and 22 control subjects (age and sex matched). The levels of NCC and NKCC2 were evaluated in UE by SDS-PAGE/western blotting with specific antibodies. Due to their location on the luminal side of tubular cells, NCC and NKCC2 are well represented in UE proteome. The NCC signal is significantly decreased/absent in UE of Gitelman patients compared with control subjects (Mann-Whitney t-test, P < 0.001) and, similarly, the NKCC2 in those of Bartter type 1 (P < 0.001). The difference in the levels of the two proteins allows recognition of Gitelman and Bartter type 1 patients from controls and, combined with clinical data, from other Bartter patients. Moreover, the receiver operating characteristic curve analysis using UE NCC densitometric values showed a good discriminating power of the test comparing GS patients versus controls and BS patients (area under the curve value = 0.92; sensitivity 84.2% and specificity 88.6%). UE phenotyping may be useful in the diagnosis of GS and BS, thus providing an alternative/complementary, urine-based diagnostic tool for SLT patient recognition and a diagnostic guidance in complex cases. © The Author 2014. Published by Oxford University Press on behalf of ERA-EDTA. All rights reserved.

Rapid actions of calcitriol and its side chain analogues CB1093 and GS1500 on intracellular calcium levels in skeletal muscle cells: a comparative study

PubMed Central

Vazquez, Guillermo; Sellés, Juana; de Boland, Ana Russo; Boland, Ricardo

1999-01-01

The ability of synthetic analogues of the secosteroid hormone 1α,25-dihydroxy-vitamin-D3 [calcitriol, CT; 1,25(OH)2D3] to exert non-genomic (rapid) effects on target cells has been scarcely studied. To evaluate the pharmacological potential of the CT side-chain analogues CB1093 and GS1500, we compared their fast effects on intracellular calcium concentration ([Ca2+]i) in chick skeletal muscle cells with those elicited by the natural hormone.Both analogues, similarly to CT, specifically induced rapid (30–60 s) and sustained rises in [Ca2+]i levels. CB1093 and GS1500 were more potent than the natural hormone at concentrations as low as 10−13 M (4.5 fold stimulation) and 10−12 M (2.5 fold), respectively, whereas higher concentrations (10−9–10−8 M) of CT were more effective than the analogues in elevating [Ca2+]i. Cyclic AMP was markedly increased by both analogues pointing for a role of this messenger in the fast actions of the synthetic compounds.In Ca2+ free medium CT and analogues elicited a transient elevation in [Ca2+]i. The PLC inhibitors U73122 (2 μM) and neomycin (0.5 mM), as well as depletion of intracellular stores with thapsigargin (1 μM), completely prevented CB1093/GS1500-dependent changes in [Ca2+]i suggesting that, similarly to CT, these analogues mobilized Ca2+ from an IP3/thapsigargin-sensitive store.The voltage-dependent calcium channel (VDCC) blocker nifedipine (2 μM) reduced by 50–60% the influx phase of the [Ca2+]i response to CB1093 and GS1500, indicating that VDCC contributed partially to Ca2+ entry. The Ca2+ readdition protocol suggested that analogue-dependent activation of a SOC entry pathway accounted, to the same extent as for CT, for the remaining non-VDCC mediated Ca2+ influx. PMID:10372825

Drosophila's contribution to stem cell research.

PubMed

Singh, Gyanesh

2015-01-01

The discovery of Drosophila stem cells with striking similarities to mammalian stem cells has brought new hope for stem cell research. Recent developments in Drosophila stem cell research is bringing wider opportunities for contemporary stem cell biologists. In this regard, Drosophila germ cells are becoming a popular model of stem cell research. In several cases, genes that controlled Drosophila stem cells were later discovered to have functional homologs in mammalian stem cells. Like mammals, Drosophila germline stem cells (GSCs) are controlled by both intrinsic as well as external signals. Inside the Drosophila testes, germline and somatic stem cells form a cluster of cells (the hub). Hub cells depend on JAK-STAT signaling, and, in absence of this signal, they do not self-renew. In Drosophila, significant changes occur within the stem cell niche that contributes to a decline in stem cell number over time. In case of aging Drosophila, somatic niche cells show reduced DE-cadherin and unpaired (Upd) proteins. Unpaired proteins are known to directly decrease stem cell number within the niches, and, overexpression of upd within niche cells restored GSCs in older males also . Stem cells in the midgut of Drosophila are also very promising. Reduced Notch signaling was found to increase the number of midgut progenitor cells. On the other hand, activation of the Notch pathway decreased proliferation of these cells. Further research in this area should lead to the discovery of additional factors that regulate stem and progenitor cells in Drosophila.

Drosophila's contribution to stem cell research

PubMed Central

Singh, Gyanesh

2016-01-01

The discovery of Drosophila stem cells with striking similarities to mammalian stem cells has brought new hope for stem cell research. Recent developments in Drosophila stem cell research is bringing wider opportunities for contemporary stem cell biologists. In this regard, Drosophila germ cells are becoming a popular model of stem cell research. In several cases, genes that controlled Drosophila stem cells were later discovered to have functional homologs in mammalian stem cells. Like mammals, Drosophila germline stem cells (GSCs) are controlled by both intrinsic as well as external signals. Inside the Drosophila testes, germline and somatic stem cells form a cluster of cells (the hub). Hub cells depend on JAK-STAT signaling, and, in absence of this signal, they do not self-renew. In Drosophila, significant changes occur within the stem cell niche that contributes to a decline in stem cell number over time. In case of aging Drosophila, somatic niche cells show reduced DE-cadherin and unpaired (Upd) proteins. Unpaired proteins are known to directly decrease stem cell number within the niches, and, overexpression of upd within niche cells restored GSCs in older males also . Stem cells in the midgut of Drosophila are also very promising. Reduced Notch signaling was found to increase the number of midgut progenitor cells. On the other hand, activation of the Notch pathway decreased proliferation of these cells. Further research in this area should lead to the discovery of additional factors that regulate stem and progenitor cells in Drosophila. PMID:26180635

Current overview on dental stem cells applications in regenerative dentistry.

PubMed

Bansal, Ramta; Jain, Aditya

2015-01-01

Teeth are the most natural, noninvasive source of stem cells. Dental stem cells, which are easy, convenient, and affordable to collect, hold promise for a range of very potential therapeutic applications. We have reviewed the ever-growing literature on dental stem cells archived in Medline using the following key words: Regenerative dentistry, dental stem cells, dental stem cells banking, and stem cells from human exfoliated deciduous teeth. Relevant articles covering topics related to dental stem cells were shortlisted and the facts are compiled. The objective of this review article is to discuss the history of stem cells, different stem cells relevant for dentistry, their isolation approaches, collection, and preservation of dental stem cells along with the current status of dental and medical applications.

Study of lignification by noninvasive techniques in growing maize internodes. An investigation by Fourier transform infrared cross-polarization-magic angle spinning 13C-nuclear magnetic resonance spectroscopy and immunocytochemical transmission electron microscopy.

PubMed Central

Joseleau, J P; Ruel, K

1997-01-01

Noninvasive techniques were used for the study in situ of lignification in the maturing cell walls of the maize (Zea mays L.) stem. Within the longitudinal axis of a developing internode all of the stages of lignification can be found. The synthesis of the three types of lignins, p-hydroxyphenylpropane (H), guaiacyl (G), and syringyl (S), was investigated in situ by cross-polarization-magic angle spinning 13C-solid-state nuclear magnetic resonance, Fourier transform infrared spectroscopy, and immunocytochemical electron microscopy. The first lignin appearing in the parenchyma is of the G-type preceeding the incorporation of S nuclei in the later stages. However, in vascular bundles, typical absorption bands of S nuclei are visible in the Fourier transform infrared spectra at the earliest stage of lignification. Immunocytochemical determination of the three types of lignin in transmission electron microscopy was possible thanks to the use of antisera prepared against synthetic H, G, and the mixed GS dehydrogenative polymers (K. Ruel, O. Faix, J.P. Joseleau [1994] J Trace Microprobe Tech 12: 247-265). The specificity of the immunological probes demonstrated that there are differences in the relative temporal synthesis of the H, G, and GS lignins in the different tissues undergoing lignification. Considering the intermonomeric linkages predominating in the antigens used for the preparation of the immunological probes, the relative intensities of the labeling obtained provided, for the first time to our knowledge, information about the macromolecular nature of lignins (condensed versus noncondensed) in relation to their ultrastructural localization and development stage. PMID:9232887

The longest telomeres: a general signature of adult stem cell compartments

PubMed Central

Flores, Ignacio; Canela, Andres; Vera, Elsa; Tejera, Agueda; Cotsarelis, George; Blasco, María A.

2008-01-01

Identification of adult stem cells and their location (niches) is of great relevance for regenerative medicine. However, stem cell niches are still poorly defined in most adult tissues. Here, we show that the longest telomeres are a general feature of adult stem cell compartments. Using confocal telomere quantitative fluorescence in situ hybridization (telomapping), we find gradients of telomere length within tissues, with the longest telomeres mapping to the known stem cell compartments. In mouse hair follicles, we show that cells with the longest telomeres map to the known stem cell compartments, colocalize with stem cell markers, and behave as stem cells upon treatment with mitogenic stimuli. Using K15-EGFP reporter mice, which mark hair follicle stem cells, we show that GFP-positive cells have the longest telomeres. The stem cell compartments in small intestine, testis, cornea, and brain of the mouse are also enriched in cells with the longest telomeres. This constitutes the description of a novel general property of adult stem cell compartments. Finally, we make the novel finding that telomeres shorten with age in different mouse stem cell compartments, which parallels a decline in stem cell functionality, suggesting that telomere loss may contribute to stem cell dysfunction with age. PMID:18283121

Energy-based culture medium design for biomanufacturing optimization: A case study in monoclonal antibody production by GS-NS0 cells.

PubMed

Quiroga-Campano, Ana L; Panoskaltsis, Nicki; Mantalaris, Athanasios

2018-03-02

Demand for high-value biologics, a rapidly growing pipeline, and pressure from competition, time-to-market and regulators, necessitate novel biomanufacturing approaches, including Quality by Design (QbD) principles and Process Analytical Technologies (PAT), to facilitate accelerated, efficient and effective process development platforms that ensure consistent product quality and reduced lot-to-lot variability. Herein, QbD and PAT principles were incorporated within an innovative in vitro-in silico integrated framework for upstream process development (UPD). The central component of the UPD framework is a mathematical model that predicts dynamic nutrient uptake and average intracellular ATP content, based on biochemical reaction networks, to quantify and characterize energy metabolism and its adaptive response, metabolic shifts, to maintain ATP homeostasis. The accuracy and flexibility of the model depends on critical cell type/product/clone-specific parameters, which are experimentally estimated. The integrated in vitro-in silico platform and the model's predictive capacity reduced burden, time and expense of experimentation resulting in optimal medium design compared to commercially available culture media (80% amino acid reduction) and a fed-batch feeding strategy that increased productivity by 129%. The framework represents a flexible and efficient tool that transforms, improves and accelerates conventional process development in biomanufacturing with wide applications, including stem cell-based therapies. Copyright © 2018. Published by Elsevier Inc.

Context clues: the importance of stem cell-material interactions

PubMed Central

Murphy, William L.

2014-01-01

Understanding the processes by which stem cells give rise to de novo tissues is an active focus of stem cell biology and bioengineering disciplines. Instructive morphogenic cues surrounding the stem cell during morphogenesis create what is referred to as the stem cell microenvironment. An emerging paradigm in stem cell bioengineering involves “biologically driven assembly,” in which stem cells are encouraged to largely define their own morphogenesis processes. However, even in the case of biologically driven assembly, stem cells do not act alone. The properties of the surrounding microenvironment can be critical regulators of cell fate. Stem cell-material interactions are among the most well-characterized microenvironmental effectors of stem cell fate, and they establish a signaling “context” that can define the mode of influence for morphogenic cues. Here we describe illustrative examples of cell-material interactions that occur during in vitro stem cell studies, with an emphasis on how cell-material interactions create instructive contexts for stem cell differentiation and morphogenesis. PMID:24369691

Cancer stem cells and differentiation therapy.

PubMed

Jin, Xiong; Jin, Xun; Kim, Hyunggee

2017-10-01

Cancer stem cells can generate tumors from only a small number of cells, whereas differentiated cancer cells cannot. The prominent feature of cancer stem cells is its ability to self-renew and differentiate into multiple types of cancer cells. Cancer stem cells have several distinct tumorigenic abilities, including stem cell signal transduction, tumorigenicity, metastasis, and resistance to anticancer drugs, which are regulated by genetic or epigenetic changes. Like normal adult stem cells involved in various developmental processes and tissue homeostasis, cancer stem cells maintain their self-renewal capacity by activating multiple stem cell signaling pathways and inhibiting differentiation signaling pathways during cancer initiation and progression. Recently, many studies have focused on targeting cancer stem cells to eradicate malignancies by regulating stem cell signaling pathways, and products of some of these strategies are in preclinical and clinical trials. In this review, we describe the crucial features of cancer stem cells related to tumor relapse and drug resistance, as well as the new therapeutic strategy to target cancer stem cells named "differentiation therapy."

Clinical trials for stem cell transplantation: when are they needed?

PubMed

Van Pham, Phuc

2016-04-27

In recent years, both stem cell research and the clinical application of these promising cells have increased rapidly. About 1000 clinical trials using stem cells have to date been performed globally. More importantly, more than 10 stem cell-based products have been approved in some countries. With the rapid growth of stem cell applications, some countries have used clinical trials as a tool to diminish the rate of clinical stem cell applications. However, the point at which stem cell clinical trials are essential remains unclear. This commentary discusses when stem cell clinical trials are essential for stem cell transplantation therapies.

Stem cells - biological update and cell therapy progress

PubMed Central

GIRLOVANU, MIHAI; SUSMAN, SERGIU; SORITAU, OLGA; RUS-CIUCA, DAN; MELINCOVICI, CARMEN; CONSTANTIN, ANNE-MARIE; MIHU, CARMEN MIHAELA

2015-01-01

In recent years, the advances in stem cell research have suggested that the human body may have a higher plasticity than it was originally expected. Until now, four categories of stem cells were isolated and cultured in vivo: embryonic stem cells, fetal stem cells, adult stem cells and induced pluripotent stem cells (hiPSCs). Although multiple studies were published, several issues concerning the stem cells are still debated, such as: the molecular mechanisms of differentiation, the methods to prevent teratoma formation or the ethical and religious issues regarding especially the embryonic stem cell research. The direct differentiation of stem cells into specialized cells: cardiac myocytes, neural cells, pancreatic islets cells, may represent an option in treating incurable diseases such as: neurodegenerative diseases, type I diabetes, hematologic or cardiac diseases. Nevertheless, stem cell-based therapies, based on stem cell transplantation, remain mainly at the experimental stages and their major limitation is the development of teratoma and cancer after transplantation. The induced pluripotent stem cells (hiPSCs) represent a prime candidate for future cell therapy research because of their significant self-renewal and differentiation potential and the lack of ethical issues. This article presents an overview of the biological advances in the study of stem cells and the current progress made in the field of regenerative medicine. PMID:26609255

Nerve Impulses in Plants

ERIC Educational Resources Information Center

Blatt, F. J.

1974-01-01

Summarizes research done on the resting and action potential of nerve impulses, electrical excitation of nerve cells, electrical properties of Nitella, and temperature effects on action potential. (GS)

Establishment of mouse expanded potential stem cells

PubMed Central

Gao, Xuefei; Antunes, Liliana; Yu, Yong; Zhu, Zhexin; Wang, Juexuan; Kolodziejczyk, Aleksandra A.; Campos, Lia S.; Wang, Cui; Yang, Fengtang; Zhong, Zhen; Fu, Beiyuan; Eckersley-Maslin, Melanie A.; Woods, Michael; Tanaka, Yosuke; Chen, Xi; Wilkinson, Adam C.; Bussell, James; White, Jacqui; Ramirez-Solis, Ramiro; Reik, Wolf; Göttgens, Berthold; Teichmann, Sarah A.; Tam, Patrick P. L.; Nakauchi, Hiromitsu; Zou, Xiangang; Lu, Liming; Liu, Pentao

2018-01-01

Mouse embryonic stem cells derived from the epiblast1 contribute to the somatic lineages and the germline but are excluded from the extra-embryonic tissues that are derived from the trophectoderm and the primitive endoderm2 upon reintroduction to the blastocyst. Here we report that cultures of expanded potential stem cells can be established from individual eight-cell blastomeres, and by direct conversion of mouse embryonic stem cells and induced pluripotent stem cells. Remarkably, a single expanded potential stem cell can contribute both to the embryo proper and to the trophectoderm lineages in a chimaera assay. Bona fide trophoblast stem cell lines and extra-embryonic endoderm stem cells can be directly derived from expanded potential stem cells in vitro. Molecular analyses of the epigenome and single-cell transcriptome reveal enrichment for blastomere-specific signature and a dynamic DNA methylome in expanded potential stem cells. The generation of mouse expanded potential stem cells highlights the feasibility of establishing expanded potential stem cells for other mammalian species. PMID:29019987

Construction of High Drug Loading and Enzymatic Degradable Multilayer Films for Self-Defense Drug Release and Long-Term Biofilm Inhibition.

PubMed

Wang, Bailiang; Liu, Huihua; Sun, Lin; Jin, Yingying; Ding, Xiaoxu; Li, Lingli; Ji, Jian; Chen, Hao

2018-01-08

Bacterial infections and biofilm formation on the surface of implants are important issues that greatly affect biomedical applications and even cause device failure. Construction of high drug loading systems on the surface and control of drug release on-demand is an efficient way to lower the development of resistant bacteria and biofilm formation. In the present study, (montmorillonite/hyaluronic acid-gentamicin) 10 ((MMT/HA-GS) 10 ) organic/inorganic hybrid multilayer films were alternately self-assembled on substrates. The loading dosage of GS was as high as 0.85 mg/cm 2 , which could be due the high specific surface area of MMT. The obtained multilayer film with high roughness gradually degraded in hyaluronidase (HAS) solutions or a bacterial infection microenvironment, which caused the responsive release of GS. The release of GS showed dual enzyme and bacterial infection responsiveness, which also indicated good drug retention and on-demand self-defense release properties of the multilayer films. Moreover, the GS release responsiveness to E. coli showed higher sensitivity than that to S. aureus. There was only ∼5 wt % GS release from the film in PBS after 48 h of immersion, and the amount quickly increased to 30 wt % in 10 5 CFU/mL of E. coli. Importantly, the high drug dosage, smart drug release, and film peeling from the surface contributed to the efficient antibacterial properties and long-term biofilm inhibition functions. Both in vitro and in vivo antibacterial tests indicated efficient sterilization function and good mammalian cell and tissue compatibility.

Nonantibiotic macrolides prevent human neutrophil elastase-induced mucus stasis and airway surface liquid volume depletion.

PubMed

Tarran, Robert; Sabater, Juan R; Clarke, Tainya C; Tan, Chong D; Davies, Catrin M; Liu, Jia; Yeung, Arthur; Garland, Alaina L; Stutts, M Jackson; Abraham, William M; Phillips, Gary; Baker, William R; Wright, Clifford D; Wilbert, Sibylle

2013-06-01

Mucus clearance is an important component of the lung's innate defense system. A failure of this system brought on by mucus dehydration is common to both cystic fibrosis (CF) and chronic obstructive pulmonary disease (COPD). Mucus clearance rates are regulated by the volume of airway surface liquid (ASL) and by ciliary beat frequency (CBF). Chronic treatment with macrolide antibiotics is known to be beneficial to both CF and COPD patients. However, chronic macrolide usage may induce bacterial resistance. We have developed a novel macrolide, 2'-desoxy-9-(S)-erythromycylamine (GS-459755), that has significantly diminished antibiotic activity against Staphylococcus aureus, Streptococcus pneumonia, Moraxella catarrhalis, and Haemophilus influenzae. Since neutrophilia frequently occurs in chronic lung disease and human neutrophil elastase (HNE) induces mucus stasis by activating the epithelial sodium channel (ENaC), we tested the ability of GS-459755 to protect against HNE-induced mucus stasis. GS-459755 had no effect on HNE activity. However, GS-459755 pretreatment protected against HNE-induced ASL volume depletion in human bronchial epithelial cells (HBECs). The effect of GS-459755 on ASL volume was dose dependent (IC₅₀ ~3.9 μM) and comparable to the antibacterial macrolide azithromycin (IC₅₀ ~2.4 μM). Macrolides had no significant effect on CBF or on transepithelial water permeability. However, the amiloride-sensitive transepithelial voltage, a marker of ENaC activity, was diminished by macrolide pretreatment. We conclude that GS-459755 may limit HNE-induced activation of ENaC and may be useful for the treatment of mucus dehydration in CF and COPD without inducing bacterial resistance.

Adult Stem Cell Therapy for Stroke: Challenges and Progress

PubMed Central

Bang, Oh Young; Kim, Eun Hee; Cha, Jae Min; Moon, Gyeong Joon

2016-01-01

Stroke is one of the leading causes of death and physical disability among adults. It has been 15 years since clinical trials of stem cell therapy in patients with stroke have been conducted using adult stem cells like mesenchymal stem cells and bone marrow mononuclear cells. Results of randomized controlled trials showed that adult stem cell therapy was safe but its efficacy was modest, underscoring the need for new stem cell therapy strategies. The primary limitations of current stem cell therapies include (a) the limited source of engraftable stem cells, (b) the presence of optimal time window for stem cell therapies, (c) inherited limitation of stem cells in terms of growth, trophic support, and differentiation potential, and (d) possible transplanted cell-mediated adverse effects, such as tumor formation. Here, we discuss recent advances that overcome these hurdles in adult stem cell therapy for stroke. PMID:27733032

Two sides of the same coin? Unraveling subtle differences between human embryonic and induced pluripotent stem cells by Raman spectroscopy.

PubMed

Parrotta, Elvira; De Angelis, Maria Teresa; Scalise, Stefania; Candeloro, Patrizio; Santamaria, Gianluca; Paonessa, Mariagrazia; Coluccio, Maria Laura; Perozziello, Gerardo; De Vitis, Stefania; Sgura, Antonella; Coluzzi, Elisa; Mollace, Vincenzo; Di Fabrizio, Enzo Mario; Cuda, Giovanni

2017-11-28

Human pluripotent stem cells, including embryonic stem cells and induced pluripotent stem cells, hold enormous promise for many biomedical applications, such as regenerative medicine, drug testing, and disease modeling. Although induced pluripotent stem cells resemble embryonic stem cells both morphologically and functionally, the extent to which these cell lines are truly equivalent, from a molecular point of view, remains controversial. Principal component analysis and K-means cluster analysis of collected Raman spectroscopy data were used for a comparative study of the biochemical fingerprint of human induced pluripotent stem cells and human embryonic stem cells. The Raman spectra analysis results were further validated by conventional biological assays. Raman spectra analysis revealed that the major difference between human embryonic stem cells and induced pluripotent stem cells is due to the nucleic acid content, as shown by the strong positive peaks at 785, 1098, 1334, 1371, 1484, and 1575 cm -1 , which is enriched in human induced pluripotent stem cells. Here, we report a nonbiological approach to discriminate human induced pluripotent stem cells from their native embryonic stem cell counterparts.

Assessment of Heparanase-Mediated Angiogenesis Using Microvascular Endothelial Cells: Identification of λ-Carrageenan Derivative as a Potent Anti Angiogenic Agent.

PubMed

Poupard, Nicolas; Badarou, Pamela; Fasani, Fabienne; Groult, Hugo; Bridiau, Nicolas; Sannier, Frédéric; Bordenave-Juchereau, Stéphanie; Kieda, Claudine; Piot, Jean-Marie; Grillon, Catherine; Fruitier-Arnaudin, Ingrid; Maugard, Thierry

2017-05-09

Heparanase is overexpressed by tumor cells and degrades the extracellular matrix proteoglycans through cleavage of heparan sulfates (HS), allowing pro-angiogenic factor release and thus playing a key role in tumor angiogenesis and metastasis. Here we propose new HS analogs as potent heparanase inhibitors: Heparin as a positive control, Dextran Sulfate, λ-Carrageenan, and modified forms of them obtained by depolymerization associated to glycol splitting (RD-GS). After heparanase activity assessment, 11 kDa RD-GS-λ-Carrageenan emerged as the most effective heparanase inhibitor with an IC 50 of 7.32 ng/mL compared to 10.7 ng/mL for the 16 kDa unfractionated heparin. The fractionated polysaccharides were then tested in a heparanase-rich medium-based in vitro model, mimicking tumor microenvironment, to determine their effect on microvascular endothelial cells (HSkMEC) angiogenesis. As a preliminary study, we identified that under hypoxic and nutrient poor conditions, MCF-7 cancer cells released much more mature heparanase in their supernatant than in normal conditions. Then a Matrigel TM assay using HSkMEC cultured under hypoxic conditions in the presence (or not) of this heparanase-rich supernatant was realized. Adding heparanase-rich media strongly enhanced angiogenic network formation with a production of twice more pseudo-vessels than with the control. When sulfated polysaccharides were tested in this angiogenesis assay, RD-GS-λ-Carrageenan was identified as a promising anti-angiogenic agent.

A family business: stem cell progeny join the niche to regulate homeostasis.

PubMed

Hsu, Ya-Chieh; Fuchs, Elaine

2012-01-23

Stem cell niches, the discrete microenvironments in which the stem cells reside, play a dominant part in regulating stem cell activity and behaviours. Recent studies suggest that committed stem cell progeny become indispensable components of the niche in a wide range of stem cell systems. These unexpected niche inhabitants provide versatile feedback signals to their stem cell parents. Together with other heterologous cell types that constitute the niche, they contribute to the dynamics of the microenvironment. As progeny are often located in close proximity to stem cell niches, similar feedback regulations may be the underlying principles shared by different stem cell systems.

A family business: stem cell progeny join the niche to regulate homeostasis

PubMed Central

Hsu, Ya-Chieh; Fuchs, Elaine

2012-01-01

Stem cell niches, the discrete microenvironments in which the stem cells reside, play a dominant part in regulating stem cell activity and behaviours. Recent studies suggest that committed stem cell progeny become indispensable components of the niche in a wide range of stem cell systems. These unexpected niche inhabitants provide versatile feedback signals to their stem cell parents. Together with other heterologous cell types that constitute the niche, they contribute to the dynamics of the microenvironment. As progeny are often located in close proximity to stem cell niches, similar feedback regulations may be the underlying principles shared by different stem cell systems. PMID:22266760

Stem Cell Therapy for Erectile Dysfunction.

PubMed

Matz, Ethan L; Terlecki, Ryan; Zhang, Yuanyuan; Jackson, John; Atala, Anthony

2018-04-06

The prevalence of erectile dysfunction (ED) is substantial and continues to rise. Current therapeutics for ED consist of oral medications, intracavernosal injections, vacuum erection devices, and penile implants. While such options may manage the disease state, none of these modalities, however, restore function. Stem cell therapy has been evaluated for erectile restoration in animal models. These cells have been derived from multiple tissues, have varied potential, and may function via local engraftment or paracrine signaling. Bone marrow-derived stem cells (BMSC) and adipose-derived stem cells (ASC) have both been used in these models with noteworthy effects. Herein, we will review the pathophysiology of ED, animal models, current and novel stem-cell based therapeutics, clinical trials and areas for future research. The relevant literature and contemporary data using keywords, "stem cells and erectile dysfunction" was reviewed. Examination of evidence supporting the association between erectile dysfunction and adipose derived stem cells, bone marrow derived stem cells, placental stem cells, urine stem cells and stem cell therapy respectively. Placental-derived stem cells and urine-derived stem cells possess many similar properties as BMSC and ASC, but the methods of acquisition are favorable. Human clinical trials have already demonstrated successful use of stem cells for improvement of erectile function. The future of stem cell research is constantly being evaluated, although, the evidence suggests a place for stem cells in erectile dysfunction therapeutics. Matz EL, Terlecki R, Zhang Y, et al. Stem Cell Therapy for Erectile Dysfunction. Sex Med Rev 2018;XX:XXX-XXX. Copyright © 2018 International Society for Sexual Medicine. Published by Elsevier Inc. All rights reserved.

A new prospect in cancer therapy: targeting cancer stem cells to eradicate cancer.

PubMed

Chen, Li-Sha; Wang, An-Xin; Dong, Bing; Pu, Ke-Feng; Yuan, Li-Hua; Zhu, Yi-Min

2012-12-01

According to the cancer stem cell theory, cancers can be initiated by cancer stem cells. This makes cancer stem cells prime targets for therapeutic intervention. Eradicating cancer stem cells by efficient targeting agents may have the potential to cure cancer. In this review, we summarize recent breakthroughs that have improved our understanding of cancer stem cells, and we discuss the therapeutic strategy of targeting cancer stem cells, a promising future direction for cancer stem cell research.

Coordination of Leaf Photosynthesis, Transpiration, and Structural Traits in Rice and Wild Relatives (Genus Oryza)1[W][OA

PubMed Central

Giuliani, Rita; Koteyeva, Nuria; Voznesenskaya, Elena; Evans, Marc A.; Cousins, Asaph B.; Edwards, Gerald E.

2013-01-01

The genus Oryza, which includes rice (Oryza sativa and Oryza glaberrima) and wild relatives, is a useful genus to study leaf properties in order to identify structural features that control CO2 access to chloroplasts, photosynthesis, water use efficiency, and drought tolerance. Traits, 26 structural and 17 functional, associated with photosynthesis and transpiration were quantified on 24 accessions (representatives of 17 species and eight genomes). Hypotheses of associations within, and between, structure, photosynthesis, and transpiration were tested. Two main clusters of positively interrelated leaf traits were identified: in the first cluster were structural features, leaf thickness (Thickleaf), mesophyll (M) cell surface area exposed to intercellular air space per unit of leaf surface area (Smes), and M cell size; a second group included functional traits, net photosynthetic rate, transpiration rate, M conductance to CO2 diffusion (gm), stomatal conductance to gas diffusion (gs), and the gm/gs ratio. While net photosynthetic rate was positively correlated with gm, neither was significantly linked with any individual structural traits. The results suggest that changes in gm depend on covariations of multiple leaf (Smes) and M cell (including cell wall thickness) structural traits. There was an inverse relationship between Thickleaf and transpiration rate and a significant positive association between Thickleaf and leaf transpiration efficiency. Interestingly, high gm together with high gm/gs and a low Smes/gm ratio (M resistance to CO2 diffusion per unit of cell surface area exposed to intercellular air space) appear to be ideal for supporting leaf photosynthesis while preserving water; in addition, thick M cell walls may be beneficial for plant drought tolerance. PMID:23669746

Adult bone marrow-derived stem cells for organ regeneration and repair.

PubMed

Tögel, Florian; Westenfelder, Christof

2007-12-01

Stem cells have been recognized as a potential tool for the development of innovative therapeutic strategies. There are in general two types of stem cells, embryonic and adult stem cells. While embryonic stem cell therapy has been riddled with problems of allogeneic rejection and ethical concerns, adult stem cells have long been used in the treatment of hematological malignancies. With the recognition of additional, potentially therapeutic characteristics, bone marrow-derived stem cells have become a tool in regenerative medicine. The bone marrow is an ideal source of stem cells because it is easily accessible and harbors two types of stem cells. Hematopoietic stem cells give rise to all blood cell types and have been shown to exhibit plasticity, while multipotent marrow stromal cells are the source of osteocytes, chondrocytes, and fat cells and have been shown to support and generate a large number of different cell types. This review describes the general characteristics of these stem cell populations and their current and potential future applications in regenerative medicine. 2007 Wiley-Liss, Inc

Stem cells.

PubMed

Behr, Björn; Ko, Sae Hee; Wong, Victor W; Gurtner, Geoffrey C; Longaker, Michael T

2010-10-01

Stem cells are self-renewing cells capable of differentiating into multiple cell lines and are classified according to their origin and their ability to differentiate. Enormous potential exists in use of stem cells for regenerative medicine. To produce effective stem cell-based treatments for a range of diseases, an improved understanding of stem cell biology and better control over stem cell fate are necessary. In addition, the barriers to clinical translation, such as potential oncologic properties of stem cells, need to be addressed. With renewed government support and continued refinement of current stem cell methodologies, the future of stem cell research is exciting and promises to provide novel reconstructive options for patients and surgeons limited by traditional paradigms.

Some Ethical Concerns About Human Induced Pluripotent Stem Cells.

PubMed

Zheng, Yue Liang

2016-10-01

Human induced pluripotent stem cells can be obtained from somatic cells, and their derivation does not require destruction of embryos, thus avoiding ethical problems arising from the destruction of human embryos. This type of stem cell may provide an important tool for stem cell therapy, but it also results in some ethical concerns. It is likely that abnormal reprogramming occurs in the induction of human induced pluripotent stem cells, and that the stem cells generate tumors in the process of stem cell therapy. Human induced pluripotent stem cells should not be used to clone human beings, to produce human germ cells, nor to make human embryos. Informed consent should be obtained from patients in stem cell therapy.

Laser biomodulation on stem cells

NASA Astrophysics Data System (ADS)

Liu, Timon C.; Duan, Rui; Li, Yan; Li, Xue-Feng; Tan, Li-Ling; Liu, Songhao

2001-08-01

Stem cells are views from the perspectives of their function, evolution, development, and cause. Counterintuitively, most stem cells may arise late in development, to act principally in tissue renewal, thus ensuring an organisms long-term survival. Surprisingly, recent reports suggest that tissue-specific adult stem cells have the potential to contribute to replenishment of multiple adult tissues. Stem cells are currently in the news for two reasons: the successful cultivation of human embryonic stem cell lines and reports that adult stem cells can differentiate into developmentally unrelated cell types, such as nerve cells into blood cells. The spotlight on stem cells has revealed gaps in our knowledge that must be filled if we are to take advantage of their full potential for treating devastating degenerative diseases such as Parkinsons's disease and muscular dystrophy. We need to know more about the intrinsic controls that keep stem cells as stem cells or direct them along particular differentiation pathways. Such intrinsic regulators are, in turn, sensitive to the influences of the microenvironment, or niche, where stem cells normally reside. Both intrinsic and extrinsic signals regular stem cell fate and some of these signals have now been identified. Vacek et al and Wang et al have studied the effect of low intensity laser on the haemopoietic stem cells in vitro. There experiments show there is indeed the effect of low intensity laser on the haemopoietic stem cells in vitro, and the present effect is the promotion of haemopoietic stem cells proliferation. In other words, low intensity laser irradiation can act as an extrinsic signal regulating stem cell fate. In this paper, we study how low intensity laser can be used to regulate stem cell fate from the viewpoint of collective phototransduction.

Fourteen new generic and ten new specific synonymies in Pholcidae (Araneae), and transfer of Mystes Bristowe to Filistatidae.

PubMed

Huber, Bernhard A; Colmenares, Pío A; Ramirez, Martin J

2014-08-08

Between 1998 and 2011, the Venezuelan arachnologist Manuel Ángel González-Sponga (GS) published a series of taxonomic papers devoted to the Pholcidae of Venezuela. Of his 22 new genera, 20 were monotypic when described, suggesting a high percentage of synonyms. We studied his descriptions and as far as accessible his type specimens and propose the following new generic synonymies: Autana GS, 2011 = Mesabolivar GS, 1998; Ayomania GS, 2005 and Venezuela Koçak & Kemal, 2008 (new replacement names for Falconia GS, 2003) = Mecolaesthus Simon, 1893; Carbonaria GS, 2009 = Mecolaesthus Simon, 1893; Caruaya GS, 2011 = Mesabolivar GS, 1998; Coroia GS, 2005 = Artema Walckenaer, 1837; Maimire GS, 2009 = Mecolaesthus Simon, 1893; Moraia GS, 2011 = Mecolaesthus Simon, 1893; Nasuta GS, 2009 = Mecolaesthus Simon, 1893; Portena GS, 2011 = Metagonia Simon, 1893; Rioparaguanus GS, 2005 = Mesabolivar GS, 1998; Tonoro GS, 2009 = Litoporus Simon, 1893; Sanluisi GS, 2003 = Mecolaesthus Simon, 1893. Three of the type species are also specific synonyms: Autana autanensis GS, 2011 = Mesabolivar aurantiacus (Mello-Leitão, 1930); Coroia magna GS, 2005 = Artema atlanta Walckenaer, 1837; Tonoro multispinae GS, 2009 = Litoporus uncatus (Simon, 1893). Six species that González-Sponga described under Blechroscelis (a genus previously synonymized with Priscula Simon, 1893) are all synonyms of Mesabolivar eberhardi Huber, 2000 (B. acuoso GS, 2011; B. araguanus GS, 2011; B. blechroscelis GS, 2011; B. copeyensis GS, 2011; B. cordillerano GS, 2011; B. andinensis GS, 2011). In addition, and unrelated to González-Sponga's work, we synonymize the Central Asian monotypic genus Ceratopholcus Spassky, 1934 with Crossopriza Simon, 1893; we synonymize the Chinese species Pholcus acerosus Peng & Zhang, 2011 with Pholcus fragillimus Strand, 1907 and remove the Malaysian monotypic genus Mystes Bristowe, 1938, previously thought to be the only East Asian representative of the subfamily Ninetinae, to the family Filistatidae. 

Potential antitumor therapeutic strategies of human amniotic membrane and amniotic fluid-derived stem cells.

PubMed

Kang, N-H; Hwang, K-A; Kim, S U; Kim, Y-B; Hyun, S-H; Jeung, E-B; Choi, K-C

2012-08-01

As stem cells are capable of self-renewal and can generate differentiated progenies for organ development, they are considered as potential source for regenerative medicine and tissue replacement after injury or disease. Along with this capacity, stem cells have the therapeutic potential for treating human diseases including cancers. According to the origins, stem cells are broadly classified into two types: embryonic stem cells (ESCs) and adult stem cells. In terms of differentiation potential, ESCs are pluripotent and adult stem cells are multipotent. Amnion, which is a membranous sac that contains the fetus and amniotic fluid and functions in protecting the developing embryo during gestation, is another stem cell source. Amnion-derived stem cells are classified as human amniotic membrane-derived epithelial stem cells, human amniotic membrane-derived mesenchymal stem cells and human amniotic fluid-derived stem cells. They are in an intermediate stage between pluripotent ESCs and lineage-restricted adult stem cells, non-tumorigenic, and contribute to low immunogenicity and anti-inflammation. Furthermore, they are easily available and do not cause any controversial issues in their recovery and applications. Not only are amnion-derived stem cells applicable in regenerative medicine, they have anticancer capacity. In non-engineered stem cells transplantation strategies, amnion-derived stem cells effectively target the tumor and suppressed the tumor growth by expressing cytotoxic cytokines. Additionally, they also have a potential as novel delivery vehicles transferring therapeutic genes to the cancer formation sites in gene-directed enzyme/prodrug combination therapy. Owing to their own advantageous properties, amnion-derived stem cells are emerging as a new candidate in anticancer therapy.

In vitro differentiation of primordial germ cells and oocyte-like cells from stem cells.

PubMed

Costa, José J N; Souza, Glaucinete B; Soares, Maria A A; Ribeiro, Regislane P; van den Hurk, Robert; Silva, José R V

2018-02-01

Infertility is the result of failure due to an organic disorder of the reproductive organs, especially their gametes. Recently, much progress has been made on generating germ cells, including oocytes, from various types of stem cells. This review focuses on advances in female germ cell differentiation from different kinds of stem cells, with emphasis on embryonic stem cells, adult stem cells, and induced pluripotent stem cells. The advantages and disadvantages of the derivation of female germ cells from several types of stem cells are also highlighted, as well as the ability of stem cells to generate mature and functional female gametes. This review shows that stem cell therapies have opened new frontiers in medicine, especially in the reproductive area, with the possibility of regenerating fertility.

Syntheses and Immunological Evaluation of Self-Adjuvanting Clustered N-Acetyl and N-Propionyl Sialyl-Tn Combined with A T-helper Cell Epitope as Antitumor Vaccine Candidates.

PubMed

Chang, Tsung-Che; Manabe, Yoshiyuki; Fujimoto, Yukari; Ohshima, Shino; Kametani, Yoshie; Kabayama, Kazuya; Nimura, Yuka; Lin, Chun-Cheng; Fukase, Koichi

2018-05-16

Sialyl-Tn (STn) is a tumor-associated carbohydrate antigen (TACA) rarely observed on healthy tissues. We synthesized two fully synthetic N-acetyl and N-propionyl STn trimer (triSTn) vaccines possessing a T-helper epitope and a TLR2 agonist, since the clustered STn antigens are highly expressed on many cancer cells. Immunization of both vaccines in mice induced the anti-triSTn IgG antibodies, which recognized triSTn-expressing cell lines PANC-1 and HepG2. The N-propionyl triSTn vaccine induced the triSTn-specific IgGs, while IgGs induced by the N-acetyl triSTn vaccine were less specific. These results illustrated that N-propionyl triSTn is a valuable unnatural TACA for anticancer vaccines. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Reduced hematopoietic stem cell frequency predicts outcome in acute myeloid leukemia.

PubMed

Wang, Wenwen; Stiehl, Thomas; Raffel, Simon; Hoang, Van T; Hoffmann, Isabel; Poisa-Beiro, Laura; Saeed, Borhan R; Blume, Rachel; Manta, Linda; Eckstein, Volker; Bochtler, Tilmann; Wuchter, Patrick; Essers, Marieke; Jauch, Anna; Trumpp, Andreas; Marciniak-Czochra, Anna; Ho, Anthony D; Lutz, Christoph

2017-09-01

In patients with acute myeloid leukemia and low percentages of aldehyde-dehydrogenase-positive cells, non-leukemic hematopoietic stem cells can be separated from leukemic cells. By relating hematopoietic stem cell frequencies to outcome we detected poor overall- and disease-free survival of patients with low hematopoietic stem cell frequencies. Serial analysis of matched diagnostic and follow-up samples further demonstrated that hematopoietic stem cells increased after chemotherapy in patients who achieved durable remissions. However, in patients who eventually relapsed, hematopoietic stem cell numbers decreased dramatically at the time of molecular relapse demonstrating that hematopoietic stem cell levels represent an indirect marker of minimal residual disease, which heralds leukemic relapse. Upon transplantation in immune-deficient mice cases with low percentages of hematopoietic stem cells of our cohort gave rise to leukemic or no engraftment, whereas cases with normal hematopoietic stem cell levels mostly resulted in multi-lineage engraftment. Based on our experimental data, we propose that leukemic stem cells have increased niche affinity in cases with low percentages of hematopoietic stem cells. To validate this hypothesis, we developed new mathematical models describing the dynamics of healthy and leukemic cells under different regulatory scenarios. These models suggest that the mechanism leading to decreases in hematopoietic stem cell frequencies before leukemic relapse must be based on expansion of leukemic stem cells with high niche affinity and the ability to dislodge hematopoietic stem cells. Thus, our data suggest that decreasing numbers of hematopoietic stem cells indicate leukemic stem cell persistence and the emergence of leukemic relapse. Copyright© 2017 Ferrata Storti Foundation.

Evaluation of the secretion and release of vascular endothelial growth factor from two-dimensional culture and three-dimensional cell spheroids formed with stem cells and osteoprecursor cells.

PubMed

Lee, Hyunjin; Lee, Sung-Il; Ko, Youngkyung; Park, Jun-Beom

2018-05-18

Co-culture has been applied in cell therapy, including stem cells, and has been reported to give enhanced functionality. In this study, stem-cell spheroids were formed in concave micromolds at different ratios of stem cells to osteoprecursor cells, and the amount of secretion of vascular endothelial growth factor (VEGF) was evaluated. Gingiva-derived stem cells and osteoprecursor cells in the amount of 6 × 105 were seeded on a 24-well culture plate or concave micromolds. The ratios of stem cells to osteoprecursor cells included: 0:4 (group 1), 1:3 (group 2), 2:2 (group 3), 3:1 (group 4), and 4:0 (group 5). The morphology of cells in a 2-dimensional culture (groups 1-5) showed a fibroblast-like appearance. The secretion of VEGF increased with the increase in stem cells, and a statistically significant increase was noted in groups 3, 4 and 5 when compared with the media-only group (p < 0.05). Osteoprecursor cells formed spheroids in concave microwells, and no noticeable change in the morphology was noted with the increase in stem cells. Spheroids containing stem cells were positive for the stem-cell markers SSEA-4. The secretion of VEGF from cell spheroids increased with the increase in stem cells. This study showed that cell spheroids formed with stem cells and osteoprecursor cells with different ratios, using microwells, had paracrine effects on the stem cells. The secretion of VEGF increased with the increase in stem cells. This stem-cell spheroid may be applied for tissue-engineering purposes.

Growth and physiological responses of isohydric and anisohydric poplars to drought

DOE PAGES

Attia, Ziv; Domec, Jean-Christophe; Oren, Ram; ...

2015-05-07

Understanding how different plants prioritize carbon gain and drought vulnerability under a variable water supply is important for predicting which trees will maximize woody biomass production under different environmental conditions. Here, Populus balsamifera (BS, isohydric genotype), P. simonii (SI, previously uncharacterized stomatal behaviour), and their cross, P. balsamifera x simonii (BSxSI, anisohydric genotype) were studied to assess the physiological basis for biomass accumulation and water-use efficiency across a range of water availabilities. Under ample water, whole plant stomatal conductance (g s), transpiration (E), and growth rates were higher in anisohydric genotypes (SI and BSxSI) than in isohydric poplars (BS). Undermore » drought, all genotypes regulated the leaf to stem water potential gradient via changes in gs, synchronizing leaf hydraulic conductance (K leaf) and E: isohydric plants reduced K leaf, g s, and E, whereas anisohydric genotypes maintained high K leaf and E, which reduced both leaf and stem water potentials. Nevertheless, SI poplars reduced their plant hydraulic conductance (K plant) during water stress and, unlike, BSxSI plants, recovered rapidly from drought. Low gs of the isohydric BS under drought reduced CO 2 assimilation rates and biomass potential under moderate water stress. While anisohydric genotypes had the fastest growth under ample water and higher photosynthetic rates under increasing water stress, isohydric poplars had higher water-use efficiency. Overall, the results indicate three strategies for how closely related biomass species deal with water stress: survival-isohydric (BS), sensitive-anisohydric (BSxSI), and resilience-anisohydric (SI). Lastly, we discuss implications for woody biomass growth, water-use efficiency, and survival under variable environmental conditions.« less

The Role of Stem Cells in Aesthetic Surgery: Fact or Fiction?

PubMed Central

McArdle, Adrian; Senarath-Yapa, Kshemendra; Walmsley, Graham G.; Hu, Michael; Atashroo, David A.; Tevlin, Ruth; Zielins, Elizabeth; Gurtner, Geoffrey C.; Wan, Derrick C.; Longaker, Michael T.

2014-01-01

Stem cells are attractive candidates for the development of novel therapies, targeting indications that involve functional restoration of defective tissue. Although most stem cell therapies are new and highly experimental, there are clinics around the world that exploit vulnerable patients with the hope of offering supposed stem cell therapies, many of which operate without credible scientific merit, oversight, or other patient protection. We review the potential, as well as drawbacks, for incorporation of stem cells in cosmetic procedures. A review of FDA-approved indications and ongoing clinical trials with adipose stem cells is provided. Furthermore, a “snapshot” analysis of websites using the search terms “stem cell therapy” or “stem cell treatment” or “stem cell facelift” was performed. Despite the protective net cast by regulatory agencies such as the FDA and professional societies such as the American Society of Plastic Surgeons, we are witnessing worrying advertisements for procedures such as stem cell facelifts, stem cell breast augmentations, and even stem cell vaginal rejuvenation. The marketing and promotion of stem cell procedures in aesthetic surgery is not adequately supported by clinical evidence in the majority of cases. Stem cells offer tremendous potential, but the marketplace is saturated with unsubstantiated and sometimes fraudulent claims that may place patients at risk. With plastic surgeons at the forefront of stem cell-based regenerative medicine, it is critically important that we provide an example of a rigorous approach to research, data collection, and advertising of stem cell therapies. PMID:24732654

Delayed diagnosis of Gorlin syndrome: Learning from mistakes!

PubMed

Ramanathan, Subramaniyan; Kumar, Devendra; Al Heidous, Mahmoud; Palaniappan, Yegu

2015-01-01

Gorlin syndrome (GS) is a rare inherited multisystem disorder with predisposition to basal cell carcinomas and various other neoplasms. Characteristic features include falx calcification, multiple odontogenic keratocysts (OKCs), early onset medulloblastoma, craniofacial and skeletal malformations, cardiac and ovarian fibroma. We present a case of GS in a 9-year-old girl with recurrent dental infections which was overlooked for 8 years. Diagnosis was finally suggested by the incidental detection of multiple OKCs and ovarian fibromas on follow-up magnetic resonance imaging performed for surveillance of previous operated brain tumor.

A WUSCHEL-Independent Stem Cell Specification Pathway Is Repressed by PHB, PHV and CNA in Arabidopsis.

PubMed

Lee, Chunghee; Clark, Steven E

2015-01-01

The homeostatic maintenance of stem cells that carry out continuous organogenesis at the shoot meristem is crucial for plant development. Key known factors act to signal between the stem cells and an underlying group of cells thought to act as the stem cell niche. In Arabidopsis thaliana the homeodomain transcription factor WUSCHEL (WUS) is essential for stem cell initiation and maintenance at shoot and flower meristems. Recent data suggest that the WUS protein may move from the niche cells directly into the stem cells to maintain stem cell identity. Here we provide evidence for a second, previously unknown, pathway for stem cell specification at shoot and flower meristems that bypasses the requirement for WUS. We demonstrate that this novel stem cell specification pathway is normally repressed by the activity of the HD-zip III transcription factors PHABULOSA (PHB), PHAVOLUTA (PHV) and CORONA (CNA). When de-repressed, this second stem cell pathway leads to an accumulation of stem cells and an enlargement of the stem cell niche. When de-repressed in a wus mutant background, this second stem cell pathway leads to functional meristems with largely normal cell layering and meristem morphology, activation of WUS cis regulatory elements, and extensive, but not indeterminate, organogenesis. Thus, WUS is largely dispensable for stem cell specification and meristem function, suggesting a set of key stem cell specification factors, competitively regulated by WUS and PHB/PHV/CNA, remain unidentified.

A WUSCHEL-Independent Stem Cell Specification Pathway Is Repressed by PHB, PHV and CNA in Arabidopsis

PubMed Central

Lee, Chunghee; Clark, Steven E.

2015-01-01

The homeostatic maintenance of stem cells that carry out continuous organogenesis at the shoot meristem is crucial for plant development. Key known factors act to signal between the stem cells and an underlying group of cells thought to act as the stem cell niche. In Arabidopsis thaliana the homeodomain transcription factor WUSCHEL (WUS) is essential for stem cell initiation and maintenance at shoot and flower meristems. Recent data suggest that the WUS protein may move from the niche cells directly into the stem cells to maintain stem cell identity. Here we provide evidence for a second, previously unknown, pathway for stem cell specification at shoot and flower meristems that bypasses the requirement for WUS. We demonstrate that this novel stem cell specification pathway is normally repressed by the activity of the HD-zip III transcription factors PHABULOSA (PHB), PHAVOLUTA (PHV) and CORONA (CNA). When de-repressed, this second stem cell pathway leads to an accumulation of stem cells and an enlargement of the stem cell niche. When de-repressed in a wus mutant background, this second stem cell pathway leads to functional meristems with largely normal cell layering and meristem morphology, activation of WUS cis regulatory elements, and extensive, but not indeterminate, organogenesis. Thus, WUS is largely dispensable for stem cell specification and meristem function, suggesting a set of key stem cell specification factors, competitively regulated by WUS and PHB/PHV/CNA, remain unidentified. PMID:26011610

Generation, characterization and potential therapeutic applications of mature and functional hepatocytes from stem cells.

PubMed

Zhang, Zhenzhen; Liu, Jianfang; Liu, Yang; Li, Zheng; Gao, Wei-Qiang; He, Zuping

2013-02-01

Liver cancer is the sixth most common tumor in the world and the majority of patients with this disease usually die within 1 year. The effective treatment for end-stage liver disease (also known as liver failure), including liver cancer or cirrhosis, is liver transplantation. However, there is a severe shortage of liver donors worldwide, which is the major handicap for the treatment of patients with liver failure. Scarcity of liver donors underscores the urgent need of using stem cell therapy to the end-stage liver disease. Notably, hepatocytes have recently been generated from hepatic and extra-hepatic stem cells. We have obtained mature and functional hepatocytes from rat hepatic stem cells. Here, we review the advancements on hepatic differentiation from various stem cells, including hepatic stem cells, embryonic stem cells, the induced pluripotent stem cells, hematopoietic stem cells, mesenchymal stem cells, and probably spermatogonial stem cells. The advantages, disadvantages, and concerns on differentiation of these stem cells into hepatic cells are highlighted. We further address the methodologies, phenotypes, and functional characterization on the differentiation of numerous stem cells into hepatic cells. Differentiation of stem cells into mature and functional hepatocytes, especially from an extra-hepatic stem cell source, would circumvent the scarcity of liver donors and human hepatocytes, and most importantly it would offer an ideal and promising source of hepatocytes for cell therapy and tissue engineering in treating liver disease. Copyright © 2012 Wiley Periodicals, Inc.

Gorlin syndrome and desmoplastic medulloblastoma: Report of 3 cases with unfavorable clinical course and novel mutations.

PubMed

Gururangan, Sridharan; Robinson, Giles; Ellison, David W; Wu, Gang; He, Xuelian; Lu, Q Richard; McLendon, Roger; Grant, Gerald; Driscoll, Timothy; Neuberg, Ronnie

2015-10-01

We present three cases of genetically confirmed Gorlin syndrome with desmoplastic medulloblastoma (DMB) in whom tumor recurred despite standard therapy. One patient was found to have a novel germline missense PTCH1 mutation. Molecular analysis of recurrent tumor using fluorescent in situ hybridization (FISH) revealed PTEN and/ or PTCH1 loss in 2 patients. Whole exome sequencing (WES) of tumor in one patient revealed loss of heterozygosity of PTCH1 and a mutation of GNAS gene in its non-coding 3' -untranslated region (UTR) with corresponding decreased protein expression. While one patient died despite high-dose chemotherapy (HDC) plus stem cell rescue (ASCR) and palliative radiotherapy, two patients are currently alive for 18+ and 120+ months respectively following retrieval therapy that did not include irradiation. Infants with DMB and GS should be treated aggressively with chemotherapy at diagnosis to prevent relapse but radiotherapy should be avoided. The use of molecular prognostic markers for DMB should be routinely used to identify the subset of tumors that might have an aggressive course. © 2015 Wiley Periodicals, Inc.

Stem cells in dentistry--part I: stem cell sources.

PubMed

Egusa, Hiroshi; Sonoyama, Wataru; Nishimura, Masahiro; Atsuta, Ikiru; Akiyama, Kentaro

2012-07-01

Stem cells can self-renew and produce different cell types, thus providing new strategies to regenerate missing tissues and treat diseases. In the field of dentistry, adult mesenchymal stem/stromal cells (MSCs) have been identified in several oral and maxillofacial tissues, which suggests that the oral tissues are a rich source of stem cells, and oral stem and mucosal cells are expected to provide an ideal source for genetically reprogrammed cells such as induced pluripotent stem (iPS) cells. Furthermore, oral tissues are expected to be not only a source but also a therapeutic target for stem cells, as stem cell and tissue engineering therapies in dentistry continue to attract increasing clinical interest. Part I of this review outlines various types of intra- and extra-oral tissue-derived stem cells with regard to clinical availability and applications in dentistry. Additionally, appropriate sources of stem cells for regenerative dentistry are discussed with regard to differentiation capacity, accessibility and possible immunomodulatory properties. Copyright © 2012 Japan Prosthodontic Society. Published by Elsevier Ltd. All rights reserved.

Plant stem cell niches.

PubMed

Stahl, Yvonne; Simon, Rüdiger

2005-01-01

Stem cells are required to support the indeterminate growth style of plants. Meristems are a plants stem cell niches that foster stem cell survival and the production of descendants destined for differentiation. In shoot meristems, stem cell fate is decided at the populational level. The size of the stem cell domain at the meristem tip depends on signals that are exchanged with cells of the organizing centre underneath. In root meristems, individual stem cells are controlled by direct interaction with cells of the quiescent centre that lie in the immediate neighbourhood. Analysis of the interactions and signaling processes in the stem cell niches has delivered some insights into the molecules that are involved and revealed that the two major niches for plant stem cells are more similar than anticipated.

Stem cells in the Drosophila digestive system.

PubMed

Zeng, Xiankun; Chauhan, Chhavi; Hou, Steven X

2013-01-01

Adult stem cells maintain tissue homeostasis by continuously replenishing damaged, aged and dead cells in any organism. Five types of region and organ-specific multipotent adult stem cells have been identified in the Drosophila digestive system: intestinal stem cells (ISCs) in the posterior midgut; hindgut intestinal stem cells (HISCs) at the midgut/hindgut junction; renal and nephric stem cells (RNSCs) in the Malpighian Tubules; type I gastric stem cells (GaSCs) at foregut/midgut junction; and type II gastric stem cells (GSSCs) at the middle of the midgut. Despite the fact that each type of stem cell is unique to a particular organ, they share common molecular markers and some regulatory signaling pathways. Due to the simpler tissue structure, ease of performing genetic analysis, and availability of abundant mutants, Drosophila serves as an elegant and powerful model system to study complex stem cell biology. The recent discoveries, particularly in the Drosophila ISC system, have greatly advanced our understanding of stem cell self-renewal, differentiation, and the role of stem cells play in tissue homeostasis/regeneration and adaptive tissue growth.

Induced cancer stem cells generated by radiochemotherapy and their therapeutic implications.

PubMed

Chen, Xiewan; Liao, Rongxia; Li, Dezhi; Sun, Jianguo

2017-03-07

Local and distant recurrence of malignant tumors following radio- and/or chemotherapy correlates with poor prognosis of patients. Among the reasons for cancer recurrence, preexisting cancer stem cells (CSCs) are considered the most likely cause due to their properties of self-renewal, pluripotency, plasticity and tumorigenicity. It has been demonstrated that preexisting cancer stem cells derive from normal stem cells and differentiated somatic cells that undergo transformation and dedifferentiation respectively under certain conditions. However, recent studies have revealed that cancer stem cells can also be induced from non-stem cancer cells by radiochemotherapy, constituting the subpopulation of induced cancer stem cells (iCSCs). These findings suggest that radiochemotherapy has the side effect of directly transforming non-stem cancer cells into induced cancer stem cells, possibly contributing to tumor recurrence and metastasis. Therefore, drugs targeting cancer stem cells or preventing dedifferentiation of non-stem cancer cells can be combined with radiochemotherapy to improve its antitumor efficacy. The current review is to investigate the mechanisms by which induced cancer stem cells are generated by radiochemotherapy and hence provide new strategies for cancer treatment.

Stem cells in gastroenterology and hepatology

PubMed Central

Quante, Michael; Wang, Timothy C.

2010-01-01

Cellular and tissue regeneration in the gastrointestinal tract and liver depends on stem cells with properties of longevity, self-renewal and multipotency. Progress in stem cell research and the identification of potential esophageal, gastric, intestinal, colonic, hepatic and pancreatic stem cells provides hope for the use of stem cells in regenerative medicine and treatments for disease. Embryonic stem cells and induced pluripotent stem cells have the potential to give rise to any cell type in the human body, but their therapeutic application remains challenging. The use of adult or tissue-restricted stem cells is emerging as another possible approach for the treatment of gastrointestinal diseases. The same self-renewal properties that allow stem cells to remain immortal and generate any tissue can occasionally make their proliferation difficult to control and make them susceptible to malignant transformation. This Review provides an overview of the different types of stem cell, focusing on tissue-restricted adult stem cells in the fields of gastroenterology and hepatology and summarizing the potential benefits and risks of using stems cells to treat gastroenterological and liver disorders. PMID:19884893

Resveratrol Prevents Ammonia Toxicity in Astroglial Cells

PubMed Central

Guerra, Maria Cristina; Leite, Marina Concli; Souza, Diogo Onofre; Gonçalves, Carlos-Alberto; Gottfried, Carmem

2012-01-01

Ammonia is implicated as a neurotoxin in brain metabolic disorders associated with hyperammonemia. Acute ammonia toxicity can be mediated by an excitotoxic mechanism, oxidative stress and nitric oxide (NO) production. Astrocytes interact with neurons, providing metabolic support and protecting against oxidative stress and excitotoxicity. Astrocytes also convert excess ammonia and glutamate into glutamine via glutamine synthetase (GS). Resveratrol, a polyphenol found in grapes and red wines, exhibits antioxidant and anti-inflammatory properties and modulates glial functions, such as glutamate metabolism. We investigated the effect of resveratrol on the production of reactive oxygen species (ROS), GS activity, S100B secretion, TNF-α, IL-1β and IL-6 levels in astroglial cells exposed to ammonia. Ammonia induced oxidative stress, decreased GS activity and increased cytokines release, probably by a mechanism dependent on protein kinase A (PKA) and extracellular signal-regulated kinase (ERK) pathways. Resveratrol prevented ammonia toxicity by modulating oxidative stress, glial and inflammatory responses. The ERK and nuclear factor-κB (NF-κB) are involved in the protective effect of resveratrol on cytokines proinflammatory release. In contrast, other antioxidants (e.g., ascorbic acid and trolox) were not effective against hyperammonemia. Thus, resveratrol could be used to protect against ammonia-induced neurotoxicity. PMID:23284918

Immunolocalization of a Unique Form of Maize Kernel Glutamine Synthetase Using a Monoclonal Antibody.

PubMed Central

Muhitch, M. J.; Felker, F. C.; Taliercio, E. W.; Chourey, P. S.

1995-01-01

The pedicel (basal maternal tissue) of maize (Zea mays L.) kernels contains a physically and kinetically unique form of glutamine synthetase (GSp1) that is involved in the conversion of transport forms of nitrogen into glutamine for uptake by the developing endosperm (M.J. Muhitch [1989] Plant Physiol 91: 868-875). A monoclonal antibody has been raised against this kernel-specific GS that does not cross-react either with a second GS isozyme found in the pedicel or with the GS isozymes from the embryo, roots, or leaves. When used as a probe for tissue printing, the antibody labeled the pedicel tissue uniformly and also labeled some of the pericarp surrounding the lower endosperm. Silver-enhanced immunogold staining of whole-kernel paraffin sections revealed the presence of GSp1 in both the vascular tissue that terminates in the pedicel and the pedicel parenchyma cells, which are located between the vascular tissue and the basal endosperm transfer cells. Light staining of the subaleurone was also noted. The tissue-specific localization of GSp1 within the pedicel is consistent with its role in the metabolism of nitrogenous transport compounds as they are unloaded from the phloem. PMID:12228400

Construction of a multiplex mutation hot spot PCR panel: the first step towards colorectal cancer genotyping on the GS Junior platform.

PubMed

Péterfia, Bálint; Kalmár, Alexandra; Patai, Árpád V; Csabai, István; Bodor, András; Micsik, Tamás; Wichmann, Barnabás; Egedi, Krisztina; Hollósi, Péter; Kovalszky, Ilona; Tulassay, Zsolt; Molnár, Béla

2017-01-01

Background: To support cancer therapy, development of low cost library preparation techniques for targeted next generation sequencing (NGS) is needed. In this study we designed and tested a PCR-based library preparation panel with limited target area for sequencing the top 12 somatic mutation hot spots in colorectal cancer on the GS Junior instrument. Materials and Methods: A multiplex PCR panel was designed to amplify regions of mutation hot spots in 12 selected genes ( APC, BRAF, CTNNB1, EGFR, FBXW7, KRAS, NRAS, MSH6, PIK3CA, SMAD2, SMAD4, TP53 ). Amplicons were sequenced on a GS Junior instrument using ligated and barcoded adaptors. Eight samples were sequenced in a single run. Colonic DNA samples (8 normal mucosa; 33 adenomas; 17 adenocarcinomas) as well as HT-29 and Caco-2 cell lines with known mutation profiles were analyzed. Variants found by the panel on APC, BRAF, KRAS and NRAS genes were validated by conventional sequencing. Results: In total, 34 kinds of mutations were detected including two novel mutations ( FBXW7 c.1740:C>G and SMAD4 c.413C>G) that have not been recorded in mutation databases, and one potential germline mutation ( APC ). The most frequently mutated genes were APC, TP53 and KRAS with 30%, 15% and 21% frequencies in adenomas and 29%, 53% and 29% frequencies in carcinomas, respectively. In cell lines, all the expected mutations were detected except for one located in a homopolymer region. According to re-sequencing results sensitivity and specificity was 100% and 92% respectively. Conclusions: Our NGS-based screening panel denotes a promising step towards low cost colorectal cancer genotyping on the GS Junior instrument. Despite the relatively low coverage, we discovered two novel mutations and obtained mutation frequencies comparable to literature data. Additionally, as an advantage, this panel requires less template DNA than sequence capture colon cancer panels currently available for the GS Junior instrument.

Lower Oncogenic Potential of Human Mesenchymal Stem Cells Derived from Cord Blood Compared to Induced Pluripotent Stem Cells

PubMed Central

Foroutan, T.; Najmi, M.; Kazemi, N.; Hasanlou, M.; Pedram, A.

2015-01-01

Background: In regenerative medicine, use of each of the mesenchymal stem cells derived from bone marrow, cord blood, and adipose tissue, has several cons and pros. Mesenchymal stem cells derived from cord blood have been considered the best source for precursor transplantation. Direct reprogramming of a somatic cell into induced pluripotent stem cells by over-expression of 6 transcription factors Oct4, Sox2, Klf4, lin28, Nanog, and c-Myc has great potential for regenerative medicine, eliminating the ethical issues of embryonic stem cells and the rejection problems of using non-autologous cells. Objective: To compare reprogramming and pluripotent markers OCT4, Sox-2, c-Myc, Klf4, Nanog, and lin28 in mesenchymal stem cells derived from cord blood and induced pluripotent stem cells. Methods: We analyzed the expression level of OCT4, Sox-2, c-Myc, Klf4, Nanog and lin28 genes in human mesenchymal stem cells derived from cord blood and induced pluripotent stem cells by cell culture and RT-PCR. Results: The expression level of pluripotent genes OCT4 and Sox-2, Nanog and lin28 in mesenchymal stem cells derived from cord blood were significantly higher than those in induced pluripotent stem cells. In contrast to OCT-4A and Sox-2, Nanog and lin28, the expression level of oncogenic factors c-Myc and Klf4 were significantly higher in induced pluripotent stem cells than in mesenchymal stem cells derived from cord blood. Conclusion: It could be concluded that mesenchymal stem cells derived from human cord blood have lower oncogenic potential compared to induced pluripotent stem cells. PMID:26306155

Eckol suppresses maintenance of stemness and malignancies in glioma stem-like cells

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

Hyun, Kyung-Hwan; Yoon, Chang-Hwan; Kim, Rae-Kwon

A subpopulation of cancer cells with stem cell properties is responsible for tumor maintenance and progression, and may contribute to resistance to anticancer treatments. Thus, compounds that target cancer stem-like cells could be usefully applied to destroy cancer. In this study, we investigated the effect of Eckol, a phlorotannin compound, on stemness and malignancies in glioma stem-like cells. To determine whether Eckol targets glioma stem-like cells, we examined whether Eckol treatment could change the expression levels of glioma stem-like cell markers and self-renewal-related proteins as well as the sphere forming ability, and the sensitivity to anticancer treatments. Alterations in themore » malignant properties of sphere-derived cells by Eckol were also investigated by soft-agar colony forming assay, by xenograft assay in nude mice, and by cell invasion assay. Treatment of sphere-forming glioma cells with Eckol effectively decreased the sphere formation as well as the CD133{sup +} cell population. Eckol treatment suppressed expression of the glioma stem-like cell markers and the self-renewal-related proteins without cell death. Moreover, treatment of glioma stem-like cells with Eckol significantly attenuated anchorage-independent growth on soft agar and tumor formation in xenograft mice. Importantly, Eckol treatment effectively reduced the resistance of glioma stem-like cells to ionizing radiation and temozolomide. Treatment of glioma stem-like cells with Eckol markedly blocked both phosphoinositide 3-kinase-Akt and Ras-Raf-1-Erk signaling pathways. These results indicate that the natural phlorotannin Eckol suppresses stemness and malignancies in glioma stem-like cells, and thereby makes glioma stem-like cells more sensitive to anticancer treatments, providing novel therapeutic strategies targeting specifically cancer stem-like cells.« less

Flagellin preconditioning enhances the efficacy of mesenchymal stem cells in an irradiation-induced proctitis model.

PubMed

Linard, Christine; Strup-Perrot, Carine; Lacave-Lapalun, Jean-Victor; Benderitter, Marc

2016-09-01

The success of mesenchymal stem cell transplantation for proctitis depends not only on cell donors but also on host microenvironmental factors, which play a major role in conditioning mesenchymal stem cell immunosuppressive action and repair. This study sought to determine if flagellin, a TLR5 ligand, can enhance the mesenchymal stem cell treatment efficacy in radiation-induced proctitis. With the use of a colorectal model of 27 Gy irradiation in rats, we investigated and compared the effects on immune capacity and remodeling at 28 d after irradiation of the following: 1) systemic mesenchymal stem cell (5 × 10(6)) administration at d 7 after irradiation, 2) administration of flagellin at d 3 and systemic mesenchymal stem cell administration at d 7, and 3) in vitro preconditioning of mesenchymal stem cells with flagellin, 24 h before their administration on d 7. The mucosal CD8(+) T cell population was normalized after treatment with flagellin-preconditioned mesenchymal stem cells or flagellin plus mesenchymal stem cells, whereas mesenchymal stem cells alone did not alter the radiation-induced elevation of CD8(+) T cell frequency. Mesenchymal stem cell treatment returned the irradiation-elevated frequency of CD25(+) cells in the mucosa-to-control levels, whereas both flagellin-preconditioned mesenchymal stem cell and flagellin-plus-mesenchymal stem cell treatment each significantly increased not only CD25(+) cell frequency but also forkhead box p3 and IL-2Rα expression. Specifically, IL-10 was overexpressed after flagellin-preconditioned mesenchymal stem cell treatment. Analysis of collagen expression showed that the collagen type 1/collagen type 3 ratio, an indicator of wound-healing maturation, was low in the irradiated and mesenchymal stem cell-treated groups and returned to the normal level only after the flagellin-preconditioned mesenchymal stem cell treatment. This was associated with a reduction in myofibroblast accumulation. In a proctitis model, flagellin-preconditioned mesenchymal stem cells improved colonic immune capacity and enhanced tissue remodeling. © Society for Leukocyte Biology.

Epidermal stem cells: location, potential and contribution to cancer.

PubMed

Ambler, C A; Määttä, A

2009-01-01

Epidermal stem cells have been classically characterized as slow-cycling, long-lived cells that reside in discrete niches in the skin. Gene expression studies of niche-resident cells have revealed a number of stem cell markers and regulators, including the Wnt/beta-catenin, Notch, p63, c-Myc and Hedgehog pathways. A new study challenges the traditional developmental paradigm of slow-cycling stem cells and rapid-cycling transit amplifying cells in some epidermal regions, and there is mounting evidence to suggest that multi-lineage epidermal progenitors can be isolated from highly proliferative, non-niche regions. Whether there is a unique microenvironment surrounding these progenitors remains to be determined. Interestingly, cancer stem cells derived from epidermal tumours exist independent of the classic skin stem cell niche, yet also have stem cell properties, including multi-lineage differentiation. This review summarizes recent studies identifying the location and regulators of mouse and human epidermal stem cells and highlights the strategies used to identify cancer stem cells, including expression of normal epidermal stem cell markers, expression of cancer stem cell markers identified in other epidermal tumours and characterization of side-population tumour cells.

Inhibitor of apoptosis signal-regulating kinase 1 protects against acetaminophen-induced liver injury

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

Xie, Yuchao; Ramachandran, Anup; Breckenridge, David G.

Metabolic activation and oxidant stress are key events in the pathophysiology of acetaminophen (APAP) hepatotoxicity. The initial mitochondrial oxidative stress triggered by protein adduct formation is amplified by c-jun-N-terminal kinase (JNK), resulting in mitochondrial dysfunction and ultimately cell necrosis. Apoptosis signal-regulating kinase 1 (ASK1) is considered the link between oxidant stress and JNK activation. The objective of the current study was to assess the efficacy and mechanism of action of the small-molecule ASK1 inhibitor GS-459679 in a murine model of APAP hepatotoxicity. APAP (300 mg/kg) caused extensive glutathione depletion, JNK activation and translocation to the mitochondria, oxidant stress and livermore » injury as indicated by plasma ALT activities and area of necrosis over a 24 h observation period. Pretreatment with 30 mg/kg of GS-459679 almost completely prevented JNK activation, oxidant stress and injury without affecting the metabolic activation of APAP. To evaluate the therapeutic potential of GS-459679, mice were treated with APAP and then with the inhibitor. Given 1.5 h after APAP, GS-459679 was still protective, which was paralleled by reduced JNK activation and p-JNK translocation to mitochondria. However, GS-459679 treatment was not more effective than N-acetylcysteine, and the combination of GS-459679 and N-acetylcysteine exhibited similar efficacy as N-acetylcysteine monotherapy, suggesting that GS-459769 and N-acetylcysteine affect the same pathway. Importantly, inhibition of ASK1 did not impair liver regeneration as indicated by PCNA staining. In conclusion, the ASK1 inhibitor GS-459679 protected against APAP toxicity by attenuating JNK activation and oxidant stress in mice and may have therapeutic potential for APAP overdose patients. - Highlights: • Two ASK1 inhibitors protected against acetaminophen-induced liver injury. • The ASK1 inhibitors protect when used as pre- or post-treatment. • Protection by ASK1 inhibitor is not due to inhibition of APAP metabolism. • The ASK1 inhibitor prevents JNK activation and translocation to mitochondria. • Treatment with ASK1 inhibitors does not impair liver regeneration after APAP.« less

MicroRNAs: key regulators of stem cells.

PubMed

Gangaraju, Vamsi K; Lin, Haifan

2009-02-01

The hallmark of a stem cell is its ability to self-renew and to produce numerous differentiated cells. This unique property is controlled by dynamic interplays between extrinsic signalling, epigenetic, transcriptional and post-transcriptional regulations. Recent research indicates that microRNAs (miRNAs) have an important role in regulating stem cell self-renewal and differentiation by repressing the translation of selected mRNAs in stem cells and differentiating daughter cells. Such a role has been shown in embryonic stem cells, germline stem cells and various somatic tissue stem cells. These findings reveal a new dimension of gene regulation in controlling stem cell fate and behaviour.

[Progress in epidermal stem cells].

PubMed

Wang, Li-Juan; Wang, You-Liang; Yang, Xiao

2010-03-01

Mammalian skin epidermis contains different epidermal stem cell pools which contribute to the homeostasis and repair of skin epithelium. Epidermal stem cells possess two essential features common to all stem cells: self-renewal and differentiation. Disturbing the balance between self-renewal and differentiation of epidermal stem cell often causes tumors or other skin diseases. Epidermal stem cell niches provide a special microenvironment that maintains a balance of stem cell quiescence and activity. This review primarily concentrates on the following points of the epidermal stem cells: the existing evidences, the self-renewal and differentiation, the division pattern, the signal pathways regulating self-renewal and differentiation, and the microenvironment (niche) and macroenvironment maintaining the homeostasis of stem cells.

[Research progress of intervertebral disc endogenous stem cells for intervertebral disc regeneration].

PubMed

Liang, Hang; Deng, Xiangyu; Shao, Zengwu

2017-10-01

To summarize the research progress of intervertebral disc endogenous stem cells for intervertebral disc regeneration and deduce the therapeutic potential of endogenous repair for intervertebral disc degeneration. The original articles about intervertebral disc endogenous stem cells for intervertebral disc regeneration were extensively reviewed; the reparative potential in vivo and the extraction and identification in vitro of intervertebral disc endogenous stem cells were analyzed; the prospect of endogenous stem cells for intervertebral disc regeneration was predicted. Stem cell niche present in the intervertebral discs, from which stem cells migrate to injured tissues and contribute to tissues regeneration under certain specific microenvironment. Moreover, the migration of stem cells is regulated by chemokines system. Tissue specific progenitor cells have been identified and successfully extracted and isolated. The findings provide the basis for biological therapy of intervertebral disc endogenous stem cells. Intervertebral disc endogenous stem cells play a crucial role in intervertebral disc regeneration. Therapeutic strategy of intervertebral disc endogenous stem cells is proven to be a promising biological approach for intervertebral disc regeneration.

Amnion-derived stem cells: in quest of clinical applications

PubMed Central

2011-01-01

In the promising field of regenerative medicine, human perinatal stem cells are of great interest as potential stem cells with clinical applications. Perinatal stem cells could be isolated from normally discarded human placentae, which are an ideal cell source in terms of availability, the fewer number of ethical concerns, less DNA damage, and so on. Numerous studies have demonstrated that some of the placenta-derived cells possess stem cell characteristics like pluripotent differentiation ability, particularly in amniotic epithelial (AE) cells. Term human amniotic epithelium contains a relatively large number of stem cell marker-positive cells as an adult stem cell source. In this review, we introduce a model theory of why so many AE cells possess stem cell characteristics. We also describe previous work concerning the therapeutic applications and discuss the pluripotency of the AE cells and potential pitfalls for amnion-derived stem cell research. PMID:21596003

The role of stem cells in aesthetic surgery: fact or fiction?

PubMed

McArdle, Adrian; Senarath-Yapa, Kshemendra; Walmsley, Graham G; Hu, Michael; Atashroo, David A; Tevlin, Ruth; Zielins, Elizabeth; Gurtner, Geoffrey C; Wan, Derrick C; Longaker, Michael T

2014-08-01

Stem cells are attractive candidates for the development of novel therapies, targeting indications that involve functional restoration of defective tissue. Although most stem cell therapies are new and highly experimental, there are clinics around the world that exploit vulnerable patients with the hope of offering supposed stem cell therapies, many of which operate without credible scientific merit, oversight, or other patient protection. The authors review the potential and the drawbacks of incorporation of stem cells in cosmetic procedures. A review of U.S. Food and Drug Administration-approved indications and ongoing clinical trials with adipose stem cells is provided. Furthermore, a "snapshot" analysis of Web sites using the search terms "stem cell therapy" or "stem cell treatment" or "stem cell facelift" was performed. Despite the protective net cast by regulatory agencies such as the U.S. Food and Drug Administration and professional societies such as the American Society of Plastic Surgeons, the authors are witnessing worrying advertisements for procedures such as stem cell face lifts, stem cell breast augmentations, and even stem cell vaginal rejuvenation. The marketing and promotion of stem cell procedures in aesthetic surgery is not adequately supported by clinical evidence in the majority of cases. Stem cells offer tremendous potential, but the marketplace is saturated with unsubstantiated and sometimes fraudulent claims that may place patients at risk. With plastic surgeons at the forefront of stem cell-based regenerative medicine, it is critically important that they provide an example of a rigorous approach to research, data collection, and advertising of stem cell therapies.

Polymer microarray technology for stem cell engineering

PubMed Central

Coyle, Robert; Jia, Jia; Mei, Ying

2015-01-01

Stem cells hold remarkable promise for applications in tissue engineering and disease modeling. During the past decade, significant progress has been made in developing soluble factors (e.g., small molecules and growth factors) to direct stem cells into a desired phenotype. However, the current lack of suitable synthetic materials to regulate stem cell activity has limited the realization of the enormous potential of stem cells. This can be attributed to a large number of materials properties (e.g., chemical structures and physical properties of materials) that can affect stem cell fate. This makes it challenging to design biomaterials to direct stem cell behavior. To address this, polymer microarray technology has been developed to rapidly identify materials for a variety of stem cell applications. In this article, we summarize recent developments in polymer array technology and their applications in stem cell engineering. Statement of significance Stem cells hold remarkable promise for applications in tissue engineering and disease modeling. In the last decade, significant progress has been made in developing chemically defined media to direct stem cells into a desired phenotype. However, the current lack of the suitable synthetic materials to regulate stem cell activities has been limiting the realization of the potential of stem cells. This can be attributed to the number of variables in material properties (e.g., chemical structures and physical properties) that can affect stem cells. Polymer microarray technology has shown to be a powerful tool to rapidly identify materials for a variety of stem cell applications. Here we summarize recent developments in polymer array technology and their applications in stem cell engineering. PMID:26497624

Stem cells in kidney regeneration.

PubMed

Yokote, Shinya; Yokoo, Takashi

2012-01-01

Currently many efforts are being made to apply regenerative medicine to kidney diseases using several types of stem/progenitor cells, such as mesenchymal stem cells, renal stem/progenitor cells, embryonic stem cells and induced pluripotent stem cells. Stem cells have the ability to repair injured organs and ameliorate damaged function. The strategy for kidney tissue repair is the recruitment of stem cells and soluble reparative factors to the kidney to elicit tissue repair and the induction of dedifferentiation of resident renal cells. On the other hand, where renal structure is totally disrupted, absolute kidney organ regeneration is needed to rebuild a whole functional kidney. In this review, we describe current advances in stem cell research for kidney tissue repair and de novo organ regeneration.

Stem Cell Sciences plc.

PubMed

Daniels, Sebnem

2006-09-01

Stem Cell Sciences' core objective is to develop safe and effective stem cell-based therapies for currently incurable diseases. In order to achieve this goal, Stem Cell Sciences recognizes the need for multiple technologies and a globally integrated stem cell initiative. The key challenges for the successful application of stem cells in the clinic is the need for a reproducible supply of pure, fully characterized stem cells that have been grown in suitable conditions for use in the clinic.

Formation of different abzymes in autoimmune-prone MRL-lpr/lpr mice is associated with changes in colony formation of haematopoietic progenitors

PubMed Central

Andryushkova, Alexandra A; Kuznetsova, Irina A; Bineva, Valentina N; Toporkova, Ludmila B; Sakhno, Ludmila V; Tikhonova, Marina A; Chernykh, Elena R; Orlovskaya, Irina A; Nevinsky, Georgy A

2007-01-01

Abstract It was shown that IgGs from the sera of 2–7-month-old control non-autoimmune (CBA x C57BL)F1 and BALB/c mice and 2–3-month-old autoimmune prone MRL-lpr/lpr mice (conditionally healthy mice) are catalytically inactive. During spontaneous development of deep systemic lupus erythematosus (SLE)-like pathology a specific reorganization of immune system of these mice leads to conditions associated with a production of IgGs hydrolyzing DNA, ATP and polysaccharides with low catalytic activities (conditionally pre-diseased mice).A significant increase in DNase, ATPase and amylase IgG relative activities associated with a transition from pre-diseased to deep diseased mice is correlated with additional changes in differentiation and proliferation of mice bone marrow haematopoietic stem cells (HSCs) and lymphocyte proliferation in different organs.The highest increase in all abzyme activities was found in mice immunized with DNA, which in comparison with pre-diseased and diseased mice are characterized by a different profile of HSC differentiation and by a suppression of cell apoptosis. Abzyme activities in the serum of pregnant females were comparable with those for pre-diseased mice, but the profile of HSC differentiation and cell apoptosis levels in pregnant and pre-diseased mice were quite different. Right after the beginning of lactation (4 days after delivery) and in a late time of lactation (14 days after delivery) there was an observed increase in cell apoptosis and two different stages of significant change in the HSC differentiation profiles; the first stage was accompanied with a significant increase and the second with a remarkable decrease in abzyme activities. Overall, all mouse groups investigated are characterized by a specific relationship between abzyme activities, HSC differentiation profiles, levels of lymphocyte proliferation, and cell apoptosis in different organs. From our point of view, the appearance of ATPase, DNase activities may be considered the earliest statistically significant marker of mouse spontaneous SLE and a further significant increase in their activities correlates with the appearance of SLE visible markers and with an increase in concentrations of anti-DNA Abs and urine protein. However, development of autoimmune (AI)-reactions and the increase in the sera anti-DNA antibodies (Abs) and in the abzyme activities in pregnant and lactating mice do not associate with SLE visible markers and proteinuria. The possible differences in immune system reorganizations during pre-disease, disease, pregnancy and lactation leading to production of different auto-antibodies and abzymes are discussed. PMID:17635644

Fusion with stem cell makes the hepatocellular carcinoma cells similar to liver tumor-initiating cells.

PubMed

Wang, Ran; Chen, Shuxun; Li, Changxian; Ng, Kevin Tak Pan; Kong, Chi-wing; Cheng, Jinping; Cheng, Shuk Han; Li, Ronald A; Lo, Chung Mau; Man, Kwan; Sun, Dong

2016-02-04

Cell fusion is a fast and highly efficient technique for cells to acquire new properties. The fusion of somatic cells with stem cells can reprogram somatic cells to a pluripotent state. Our research on the fusion of stem cells and cancer cells demonstrates that the fused cells can exhibit stemness and cancer cell-like characteristics. Thus, tumor-initiating cell-like cells are generated. We employed laser-induced single-cell fusion technique to fuse the hepatocellular carcinoma cells and human embryonic stem cells (hESC). Real-time RT-PCR, flow cytometry and in vivo tumorigenicity assay were adopted to identify the gene expression difference. We successfully produced a fused cell line that coalesces the gene expression information of hepatocellular carcinoma cells and stem cells. Experimental results showed that the fused cells expressed cancer and stemness markers as well as exhibited increased resistance to drug treatment and enhanced tumorigenesis. Fusion with stem cells transforms liver cancer cells into tumor initiating-like cells. Results indicate that fusion between cancer cell and stem cell may generate tumor initiating-like cells.

Stem cell-derived vascular endothelial cells and their potential application in regenerative medicine

USDA-ARS?s Scientific Manuscript database

Although a 'vascular stem cell' population has not been identified or generated, vascular endothelial and mural cells (smooth muscle cells and pericytes) can be derived from currently known pluripotent stem cell sources, including human embryonic stem cells and induced pluripotent stem cells. We rev...

Hematopoietic cell differentiation from embryonic and induced pluripotent stem cells

PubMed Central

2013-01-01

Pluripotent stem cells, both embryonic stem cells and induced pluripotent stem cells, are undifferentiated cells that can self-renew and potentially differentiate into all hematopoietic lineages, such as hematopoietic stem cells (HSCs), hematopoietic progenitor cells and mature hematopoietic cells in the presence of a suitable culture system. Establishment of pluripotent stem cells provides a comprehensive model to study early hematopoietic development and has emerged as a powerful research tool to explore regenerative medicine. Nowadays, HSC transplantation and hematopoietic cell transfusion have successfully cured some patients, especially in malignant hematological diseases. Owing to a shortage of donors and a limited number of the cells, hematopoietic cell induction from pluripotent stem cells has been regarded as an alternative source of HSCs and mature hematopoietic cells for intended therapeutic purposes. Pluripotent stem cells are therefore extensively utilized to facilitate better understanding in hematopoietic development by recapitulating embryonic development in vivo, in which efficient strategies can be easily designed and deployed for the generation of hematopoietic lineages in vitro. We hereby review the current progress of hematopoietic cell induction from embryonic stem/induced pluripotent stem cells. PMID:23796405

Stem Cell Basics

MedlinePlus

... Tips Info Center Research Topics Federal Policy Glossary Stem Cell Information General Information Clinical Trials Funding Information Current ... Basics » Stem Cell Basics I. Back to top Stem Cell Basics I. Introduction: What are stem cells, and ...

TOPICAL REVIEW: Stem cells engineering for cell-based therapy

NASA Astrophysics Data System (ADS)

Taupin, Philippe

2007-09-01

Stem cells carry the promise to cure a broad range of diseases and injuries, from diabetes, heart and muscular diseases, to neurological diseases, disorders and injuries. Significant progresses have been made in stem cell research over the past decade; the derivation of embryonic stem cells (ESCs) from human tissues, the development of cloning technology by somatic cell nuclear transfer (SCNT) and the confirmation that neurogenesis occurs in the adult mammalian brain and that neural stem cells (NSCs) reside in the adult central nervous system (CNS), including that of humans. Despite these advances, there may be decades before stem cell research will translate into therapy. Stem cell research is also subject to ethical and political debates, controversies and legislation, which slow its progress. Cell engineering has proven successful in bringing genetic research to therapy. In this review, I will review, in two examples, how investigators are applying cell engineering to stem cell biology to circumvent stem cells' ethical and political constraints and bolster stem cell research and therapy.

From Banking to International Governance: Fostering Innovation in Stem Cell Research

PubMed Central

Isasi, Rosario; Knoppers, Bartha M.

2011-01-01

Stem cell banks are increasingly recognized as an essential resource of biological materials for both basic and translational stem cell research. By providing transnational access to quality controlled and ethically sourced stem cell lines, stem cell banks seek to foster international collaboration and innovation. However, given that national stem cell banks operate under different policy, regulatory and commercial frameworks, the transnational sharing of stem cell materials and data can be complicating. This paper will provide an overview of the most pressing challenges regarding the governance of stem cell banks, and the difficulties in designing regulatory and commercial frameworks that foster stem cell research. Moreover, the paper will shed light on the numerous international initiatives that have arisen to help harmonize and standardize stem cell banking and research processes to overcome such challenges. PMID:21904557

Stem Cells Transplantation in the Treatment of Patients with Liver Failure.

PubMed

Tao, Ya-Chao; Wang, Meng-Lan; Chen, En-Qiang; Tang, Hong

2018-02-23

Liver failure is a life-threatening liver disease encompassing severe acute deterioration of liver function. Emergency liver transplantation is the only curative treatment for liver failure, but is restricted by the severe shortage of organ donors. Stem cell, including embroyonic stem cells, induced pluripotent stem cells, mesenchymal stem cells, hematopoietic stem cells and hepatic progenitor cells, have capacity to proliferate and differentiate and could be used in a variety of liver diseases including hereditary liver diseases, cirrhosis and liver failure. We summarized the basic experimental and clinical advances of stem cell transplantation in liver failure treatment, and also discussed the advantages and disadvantage of different stem cells subtype in this field, aiming to provide a perspective on the stem cell-based therapy for liver failure. Stem cells, especially mesenchymal stem cells (mainly low immunogenicity and paracrine characteristics) and induced pluripotent stem cells (generation of desired cell type from somatic cell), are feasible candidates for cell therapy in the treatment of liver failure, but there are some drawbacks remaining to be resolved, such as low engraftment, cryotpreservation methods and tumorigenesis. Stem cell transplantation is a promising but challenging strategy and paves a new way for curing liver failure. But more efforts need to be made to overcome problems before this new strategy could be safely and effectively applied to humans. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Recent Progress in Stem Cell Modification for Cardiac Regeneration

PubMed Central

Voronina, Natalia; Steinhoff, Gustav

2018-01-01

During the past decades, stem cell-based therapy has acquired a promising role in regenerative medicine. The application of novel cell therapeutics for the treatment of cardiovascular diseases could potentially achieve the ambitious aim of effective cardiac regeneration. Despite the highly positive results from preclinical studies, data from phase I/II clinical trials are inconsistent and the improvement of cardiac remodeling and heart performance was found to be quite limited. The major issues which cardiac stem cell therapy is facing include inefficient cell delivery to the site of injury, accompanied by low cell retention and weak effectiveness of remaining stem cells in tissue regeneration. According to preclinical and clinical studies, various stem cells (adult stem cells, embryonic stem cells, and induced pluripotent stem cells) represent the most promising cell types so far. Beside the selection of the appropriate cell type, researchers have developed several strategies to produce “second-generation” stem cell products with improved regenerative capacity. Genetic and nongenetic modifications, chemical and physical preconditioning, and the application of biomaterials were found to significantly enhance the regenerative capacity of transplanted stem cells. In this review, we will give an overview of the recent developments in stem cell engineering with the goal to facilitate stem cell delivery and to promote their cardiac regenerative activity. PMID:29535769

Eat, breathe, ROS: controlling stem cell fate through metabolism.

PubMed

Kubli, Dieter A; Sussman, Mark A

2017-05-01

Research reveals cardiac regeneration exists at levels previously deemed unattainable. Clinical trials using stem cells demonstrate promising cardiomyogenic and regenerative potential but insufficient contractile recovery. Incomplete understanding of the biology of administered cells likely contributes to inconsistent patient outcomes. Metabolism is a core component of many well-characterized stem cell types, and metabolic changes fundamentally alter stem cell fate from self-renewal to lineage commitment, and vice versa. However, the metabolism of stem cells currently studied for cardiac regeneration remains incompletely understood. Areas covered: Key metabolic features of stem cells are reviewed and unique stem cell metabolic characteristics are discussed. Metabolic changes altering stem cell fate are considered from quiescence and self-renewal to lineage commitment. Key metabolic concepts are applied toward examining cardiac regeneration through stem cell-based approaches, and clinical implications of current cell therapies are evaluated to identify potential areas of improvement. Expert commentary: The metabolism and biology of stem cells used for cardiac therapy remain poorly characterized. A growing appreciation for the fundamental relationship between stem cell functionality and metabolic phenotype is developing. Future studies unraveling links between cardiac stem cell metabolism and regenerative potential may considerably improve treatment strategies and therapeutic outcomes.

Eat, breathe, ROS: controlling stem cell fate through metabolism

PubMed Central

Kubli, Dieter A.; Sussman, Mark A.

2017-01-01

Introduction Research reveals cardiac regeneration exists at levels previously deemed unattainable. Clinical trials using stem cells demonstrate promising cardiomyogenic and regenerative potential but insufficient contractile recovery. Incomplete understanding of the biology of administered cells likely contributes to inconsistent patient outcomes. Metabolism is a core component of many well-characterized stem cell types, and metabolic changes fundamentally alter stem cell fate from self-renewal to lineage commitment, and vice versa. However, the metabolism of stem cells currently studied for cardiac regeneration remains incompletely understood. Areas covered Key metabolic features of stem cells are reviewed and unique stem cell metabolic characteristics are discussed. Metabolic changes altering stem cell fate are considered from quiescence and self-renewal to lineage commitment. Key metabolic concepts are applied toward examining cardiac regeneration through stem cell-based approaches, and clinical implications of current cell therapies are evaluated to identify potential areas of improvement. Expert commentary The metabolism and biology of stem cells used for cardiac therapy remain poorly characterized. A growing appreciation for the fundamental relationship between stem cell functionality and metabolic phenotype is developing. Future studies unraveling links between cardiac stem cell metabolism and regenerative potential may considerably improve treatment strategies and therapeutic outcomes. PMID:28406333

Therapeutic strategies involving uterine stem cells in reproductive medicine.

PubMed

Simoni, Michael; Taylor, Hugh S

2018-06-01

The current review provides an update on recent advances in stem cell biology relevant to female reproduction. Stem cells are undifferentiated cells that often serve as a reservoir of cells to regenerate tissue in settings or injury or cell loss. The endometrium has progenitor stem cells that can replace all of the endometrium during each menstrual cycle. In addition, multipotent endometrial cells replace these progenitor cells when depleted. Recruitment of stem cells from outside of the uterus occurs in setting of increased demand such as ischemia or injury. Bone marrow-derived multipotent stem cells are recruited to the uterus by estrogen or injury-induced expression of the chemokine CXCL12. In the setting of overwhelming injury, especially in the setting of low estrogen levels, there may be insufficient stem cell recruitment to adequately repair the uterus resulting in conditions such as Asherman syndrome or other endometrial defects. In contrast, excessive recruitment of stem cells underlies endometriosis. Enhanced understanding of stem-cell mobilization, recruitment, and engraftment has created the possibility of improved therapy for endometrial defects and endometriosis through enhanced manipulation of stem-cell trafficking. Further, the normal endometrium is a rich source of multipotent stem cells that can be used for numerous applications in regenerative medicine beyond reproduction. A better understanding of reproductive stem-cell biology may allow improved treatment of endometrial disease such as Asherman syndrome and other endometrial receptivity defects. Inhibiting stem-cell mobilization may also be helpful in endometriosis therapy. Finally, endometrial derived multipotent stem cells may play a crucial role in cell therapy for regenerative medicine.

Gene screening of Wharton's jelly derived stem cells.

PubMed

Mechiche Alami, S; Velard, F; Draux, F; Siu Paredes, F; Josse, J; Lemaire, F; Gangloff, S C; Graesslin, O; Laurent-Maquin, D; Kerdjoudj, H

2014-01-01

Stem cells are the most powerful candidate for the treatment of various diseases. Suitable stem cell source should be harvested with minimal invasive procedure, found in great quantity, and transplanted with no risk of immune response and tumor formation. Fetal derived stem cells have been introduced as an excellent alternative to adult and embryonic stem cells use, but unfortunately, their degree of "stemness" and molecular characterization is still unclear. Several studies have been performed deciphering whether fetal stem cells meet the needs of regenerative medicine. We believe that a transcriptomic screening of Wharton's jelly stem cells will bring insights on cell population features.

Stem Cell Banking for Regenerative and Personalized Medicine

PubMed Central

Harris, David T.

2014-01-01

Regenerative medicine, tissue engineering and gene therapy offer the opportunity to treat and cure many of today’s intractable afflictions. These approaches to personalized medicine often utilize stem cells to accomplish these goals. However, stem cells can be negatively affected by donor variables such as age and health status at the time of collection, compromising their efficacy. Stem cell banking offers the opportunity to cryogenically preserve stem cells at their most potent state for later use in these applications. Practical stem cell sources include bone marrow, umbilical cord blood and tissue, and adipose tissue. Each of these sources contains stem cells that can be obtained from most individuals, without too much difficulty and in an economical fashion. This review will discuss the advantages and disadvantages of each stem cell source, factors to be considered when contemplating banking each stem cell source, the methodology required to bank each stem cell source, and finally, current and future clinical uses of each stem cell source. PMID:28548060

Tanovea® for the treatment of lymphoma in dogs.

PubMed

De Clercq, Erik

2018-05-17

Tanovea® (first named GS-9219, then VDC-1101, generic name: rabacfosadine) is a pro-prodrug or "double" prodrug of PMEG [9-(2-phosphonylmethoxyethyl)guanine], which has been conditionally approved by the US FDA (Food and Drug Administration) for the treatment of lymphoma in dogs. Tanovea has been demonstrated to be effective against non-Hodgkin's lymphoma (NHL) in dogs, as well as canine cutaneous T-cell lymphoma, spontaneous canine multiple myeloma, naïve canine multicentric lymphoma and relapsed canine B-cell lymphoma. As a double prodrug of PMEG, GS-9219 is first converted intracellularly by hydrolysis to cPr-PMEDAP, then deaminated to PMEG, which is then phosphorylated twice to its active metabolite PMEGpp, acting at the level of the cellular DNA polymerases. Copyright © 2018 Elsevier Inc. All rights reserved.

Broadband Gerchberg-Saxton algorithm for freeform diffractive spectral filter design.

PubMed

Vorndran, Shelby; Russo, Juan M; Wu, Yuechen; Pelaez, Silvana Ayala; Kostuk, Raymond K

2015-11-30

A multi-wavelength expansion of the Gerchberg-Saxton (GS) algorithm is developed to design and optimize a surface relief Diffractive Optical Element (DOE). The DOE simultaneously diffracts distinct wavelength bands into separate target regions. A description of the algorithm is provided, and parameters that affect filter performance are examined. Performance is based on the spectral power collected within specified regions on a receiver plane. The modified GS algorithm is used to design spectrum splitting optics for CdSe and Si photovoltaic (PV) cells. The DOE has average optical efficiency of 87.5% over the spectral bands of interest (400-710 nm and 710-1100 nm). Simulated PV conversion efficiency is 37.7%, which is 29.3% higher than the efficiency of the better performing PV cell without spectrum splitting optics.

Nine Things to Know About Stem Cell Treatments

MedlinePlus

... Toggle Nav Nine Things To Know About Stem Cell Treatments Home > Stem Cells and Medicine > Nine Things ... About Stem Cell Treatments Many clinics offering stem cell treatments make claims that are not supported by ...

Cancer (stem) cell differentiation: An inherent or acquired property?

PubMed

Mohr, Marieke; Zänker, Kurt S; Dittmar, Thomas

2015-12-01

There is a growing list of data indicating that cancer (stem) cells could functionally adapt foreign tissue features, such as endothelial-like cells or neuroendocrine cells, express lineage markers or could differentiate into various lineages in response to appropriate differentiation criteria. The finding that cancer (stem) cells may possess some kind of differentiation capacity poses the question whether this might be an inherent or acquired property. Cancer stem cells share stem cell characteristics and may thus possess an inherent differentiation capacity enabling the cells to respond to various differentiation stimuli. Considering the plasticity of cancer (stem) cells, even non-tumorigenic (and putatively non-differentiable) tumor cells could give rise to tumorigenic tumor stem cells, exhibiting stem cell characteristics including an inherent differentiation capacity. On the contrary, cancer (stem) cells may have acquired differentiation capacity as a consequence of a previous cell fusion event with cell types exhibiting differentiation potential and being fusogenic, such as macrophages or stem cells. Of pivotal interest in a tumor context are macrophages, which chiefly foster the chronically inflamed tumor microenvironment. Because chronically inflamed tissue is a well-known trigger for cell fusion and both macrophages and stem cells are highly fusogenic we conclude that cell fusion events between these cell types and cancer (stem) cells should frequently occur, thereby giving rise to hybrid cells exhibiting not only novel properties, like an enhanced metastatogenic phenotype, but also parental characteristics, such as differentiation capacity. Conceivably, the combination of both properties might be advantageous for metastasizing cancer (stem) cells to adapt better and faster to a foreign organ tissue environment. Copyright © 2015 Elsevier Ltd. All rights reserved.

Sox10+ adult stem cells contribute to biomaterial encapsulation and microvascularization

PubMed Central

Wang, Dong; Wang, Aijun; Wu, Fan; Qiu, Xuefeng; Li, Ye; Chu, Julia; Huang, Wen-Chin; Xu, Kang; Gong, Xiaohua; Li, Song

2017-01-01

Implanted biomaterials and biomedical devices generally induce foreign body reaction and end up with encapsulation by a dense avascular fibrous layer enriched in extracellular matrix. Fibroblasts/myofibroblasts are thought to be the major cell type involved in encapsulation, but it is unclear whether and how stem cells contribute to this process. Here we show, for the first time, that Sox10+ adult stem cells contribute to both encapsulation and microvessel formation. Sox10+ adult stem cells were found sparsely in the stroma of subcutaneous loose connective tissues. Upon subcutaneous biomaterial implantation, Sox10+ stem cells were activated and recruited to the biomaterial scaffold, and differentiated into fibroblasts and then myofibroblasts. This differentiation process from Sox10+ stem cells to myofibroblasts could be recapitulated in vitro. On the other hand, Sox10+ stem cells could differentiate into perivascular cells to stabilize newly formed microvessels. Sox10+ stem cells and endothelial cells in three-dimensional co-culture self-assembled into microvessels, and platelet-derived growth factor had chemotactic effect on Sox10+ stem cells. Transplanted Sox10+ stem cells differentiated into smooth muscle cells to stabilize functional microvessels. These findings demonstrate the critical role of adult stem cells in tissue remodeling and unravel the complexity of stem cell fate determination. PMID:28071739

Tumor suppressors Sav/Scrib and oncogene Ras regulate stem cell transformation in adult Drosophila Malpighian Tubules

PubMed Central

Zeng, Xiankun; Singh, Shree Ram; Hou, David; Hou, Steven X.

2012-01-01

An increasing body of evidence suggests that tumors might originate from a few transformed cells that share many properties with normal stem cells. However, it remains unclear how normal stem cells are transformed into cancer stem cells. Here, we demonstrated that mutations causing the loss of tumor suppressor Sav or Scrib or activation of the oncogene Ras transform normal stem cells into cancer stem cells through a multistep process in the adult Drosophila Malpighian Tubules (MTs). In wild-type MTs, each stem cell generates one self-renewing and one differentiating daughter cell. However, in flies with loss-of-function sav or scrib or gain-of-function Ras mutations, both daughter cells grew and behaved like stem cells, leading to the formation of tumors in MTs. Ras functioned downstream of Sav and Scrib in regulating the stem cell transformation. The Ras-transformed stem cells exhibited many of the hallmarks of cancer, such as increased proliferation, reduced cell death, and failure to differentiate. We further demonstrated that several signal transduction pathways (including MEK/MAPK, RhoA, PKA, and TOR) mediate Rasṕ function in the stem cell transformation. Therefore, we have identified a molecular mechanism that regulates stem cell transformation, and this finding may lead to strategies for preventing tumor formation in certain organs. PMID:20432470

The king is dead, long live the king: entering a new era of stem cell research and clinical development.

PubMed

Ichim, Thomas; Riordan, Neil H; Stroncek, David F

2011-12-20

In mid November the biopharma industry was shocked by the announcement from Geron that they were ending work on embryonic stem cell research and therapy. For more than 10 years the public image of all stem cell research has been equated with embryonic stem cells. Unfortunately, a fundamentally important medical and financial fact was being ignored: embryonic stem cell therapy is extremely immature. In parallel to efforts in embryonic stem cell research and development, scientists and physicians in the field of adult stem cells realized that the natural role of adult stem cells in the body is to promote healing and to act like endogenous "repair cells" and, as a result, numerous companies have entered the field of adult stem cell therapy with the goal of expanding numbers of adult stem cells for administration to patients with various conditions. In contrast to embryonic stem cells, which are extremely expensive and potentially dangerous, adult cell cells are inexpensive and have an excellent safety record when used in humans. Many studies are now showing that adult stem cells are practical, patient-applicable, therapeutics that are very close to being available for incorporation into the practice of medicine. These events signal the entrance of the field of stem cells into a new era: an era where hype and misinformation no longer triumph over economic and medical realities.

Evidence for functional pre-coupled complexes of receptor heteromers and adenylyl cyclase.

PubMed

Navarro, Gemma; Cordomí, Arnau; Casadó-Anguera, Verónica; Moreno, Estefanía; Cai, Ning-Sheng; Cortés, Antoni; Canela, Enric I; Dessauer, Carmen W; Casadó, Vicent; Pardo, Leonardo; Lluís, Carme; Ferré, Sergi

2018-03-28

G protein-coupled receptors (GPCRs), G proteins and adenylyl cyclase (AC) comprise one of the most studied transmembrane cell signaling pathways. However, it is unknown whether the ligand-dependent interactions between these signaling molecules are based on random collisions or the rearrangement of pre-coupled elements in a macromolecular complex. Furthermore, it remains controversial whether a GPCR homodimer coupled to a single heterotrimeric G protein constitutes a common functional unit. Using a peptide-based approach, we here report evidence for the existence of functional pre-coupled complexes of heteromers of adenosine A 2A receptor and dopamine D 2 receptor homodimers coupled to their cognate Gs and Gi proteins and to subtype 5 AC. We also demonstrate that this macromolecular complex provides the necessary frame for the canonical Gs-Gi interactions at the AC level, sustaining the ability of a Gi-coupled GPCR to counteract AC activation mediated by a Gs-coupled GPCR.

Control of stem cell fate by engineering their micro and nanoenvironment

PubMed Central

Griffin, Michelle F; Butler, Peter E; Seifalian, Alexander M; Kalaskar, Deepak M

2015-01-01

Stem cells are capable of long-term self-renewal and differentiation into specialised cell types, making them an ideal candidate for a cell source for regenerative medicine. The control of stem cell fate has become a major area of interest in the field of regenerative medicine and therapeutic intervention. Conventional methods of chemically inducing stem cells into specific lineages is being challenged by the advances in biomaterial technology, with evidence highlighting that material properties are capable of driving stem cell fate. Materials are being designed to mimic the clues stem cells receive in their in vivo stem cell niche including topographical and chemical instructions. Nanotopographical clues that mimic the extracellular matrix (ECM) in vivo have shown to regulate stem cell differentiation. The delivery of ECM components on biomaterials in the form of short peptides sequences has also proved successful in directing stem cell lineage. Growth factors responsible for controlling stem cell fate in vivo have also been delivered via biomaterials to provide clues to determine stem cell differentiation. An alternative approach to guide stem cells fate is to provide genetic clues including delivering DNA plasmids and small interfering RNAs via scaffolds. This review, aims to provide an overview of the topographical, chemical and molecular clues that biomaterials can provide to guide stem cell fate. The promising features and challenges of such approaches will be highlighted, to provide directions for future advancements in this exciting area of stem cell translation for regenerative medicine. PMID:25621104

The Neurovascular Properties of Dental Stem Cells and Their Importance in Dental Tissue Engineering

PubMed Central

Ratajczak, Jessica; Bronckaers, Annelies; Dillen, Yörg; Gervois, Pascal; Vangansewinkel, Tim; Driesen, Ronald B.; Wolfs, Esther; Lambrichts, Ivo

2016-01-01

Within the field of tissue engineering, natural tissues are reconstructed by combining growth factors, stem cells, and different biomaterials to serve as a scaffold for novel tissue growth. As adequate vascularization and innervation are essential components for the viability of regenerated tissues, there is a high need for easily accessible stem cells that are capable of supporting these functions. Within the human tooth and its surrounding tissues, different stem cell populations can be distinguished, such as dental pulp stem cells, stem cells from human deciduous teeth, stem cells from the apical papilla, dental follicle stem cells, and periodontal ligament stem cells. Given their straightforward and relatively easy isolation from extracted third molars, dental stem cells (DSCs) have become an attractive source of mesenchymal-like stem cells. Over the past decade, there have been numerous studies supporting the angiogenic, neuroprotective, and neurotrophic effects of the DSC secretome. Together with their ability to differentiate into endothelial cells and neural cell types, this makes DSCs suitable candidates for dental tissue engineering and nerve injury repair. PMID:27688777

Multipotent Stem Cell and Reproduction.

PubMed

Khanlarkhani, Neda; Baazm, Maryam; Mohammadzadeh, Farzaneh; Najafi, Atefeh; Mehdinejadiani, Shayesteh; Sobhani, Aligholi

Stem cells are self-renewing and undifferentiated cell types that can be differentiate into functional cells. Stem cells can be classified into two main types based on their source of origin: Embryonic and Adult stem cells. Stem cells also classified based on the range of differentiation potentials into Totipotent, Pluripotent, Multipotent, and Unipotent. Multipotent stem cells have the ability to differentiate into all cell types within one particular lineage. There are plentiful advantages and usages for multipotent stem cells. Multipotent Stem cells act as a significant key in procedure of development, tissue repair, and protection. The accessibility and adaptability of these amazing cells create them a great therapeutic choice for different part of medical approaches, and it becomes interesting topic in the scientific researches to found obvious method for the most advantageous use of MSC-based therapies. Recent studies in the field of stem cell biology have provided new perspectives and opportunities for the treatment of infertility disorders.

Strategies to improve homing of mesenchymal stem cells for greater efficacy in stem cell therapy.

PubMed

Naderi-Meshkin, Hojjat; Bahrami, Ahmad Reza; Bidkhori, Hamid Reza; Mirahmadi, Mahdi; Ahmadiankia, Naghmeh

2015-01-01

Stem/progenitor cell-based therapeutic approach in clinical practice has been an elusive dream in medical sciences, and improvement of stem cell homing is one of major challenges in cell therapy programs. Stem/progenitor cells have a homing response to injured tissues/organs, mediated by interactions of chemokine receptors expressed on the cells and chemokines secreted by the injured tissue. For improvement of directed homing of the cells, many techniques have been developed either to engineer stem/progenitor cells with higher amount of chemokine receptors (stem cell-based strategies) or to modulate the target tissues to release higher level of the corresponding chemokines (target tissue-based strategies). This review discusses both of these strategies involved in the improvement of stem cell homing focusing on mesenchymal stem cells as most frequent studied model in cellular therapies. © 2014 International Federation for Cell Biology.

College Students' Conceptions of Stem Cells, Stem Cell Research, and Cloning

ERIC Educational Resources Information Center

Concannon, James P.; Siegel, Marcelle A.; Halverson, Kristy; Freyermuth, Sharyn

2010-01-01

In this study, we examined 96 undergraduate non-science majors' conceptions of stem cells, stem cell research, and cloning. This study was performed at a large, Midwest, research extensive university. Participants in the study were asked to answer 23 questions relating to stem cells, stem cell research, and cloning in an on-line assessment before…

Stem cell biobanks.

PubMed

Bardelli, Silvana

2010-04-01

Stem cells contribute to innate healing and harbor a promising role for regenerative medicine. Stem cell banking through long-term storage of different stem cell platforms represents a fundamental source to preserve original features of stem cells for patient-specific clinical applications. Stem cell research and clinical translation constitute fundamental and indivisible modules catalyzed through biobanking activity, generating a return of investment.

Federal Employees: Appointees Converted to Career Positions, July through September 1988

DTIC Science & Technology

1989-01-13

Media) GS-1035-13/2 GS- 1082 -12/5 Program Analyst yes Program Analyst Temporary GS-345-12/1 GS-345-12/1 Legislative Affairs yes Congressional Liaison...Officer GS-301-13/1 GM-345-14 GS-14/1 equivalent pay MERIT SYSTEMS PROTECTION BOARD Writer/Editor yes Writer/Editor Temporary GS- 1082 -12/1 GS- 1082 -12/1

Muscle Stem Cells: A Model System for Adult Stem Cell Biology.

PubMed

Cornelison, Ddw; Perdiguero, Eusebio

2017-01-01

Skeletal muscle stem cells, originally termed satellite cells for their position adjacent to differentiated muscle fibers, are absolutely required for the process of skeletal muscle repair and regeneration. In the last decade, satellite cells have become one of the most studied adult stem cell systems and have emerged as a standard model not only in the field of stem cell-driven tissue regeneration but also in stem cell dysfunction and aging. Here, we provide background in the field and discuss recent advances in our understanding of muscle stem cell function and dysfunction, particularly in the case of aging, and the potential involvement of muscle stem cells in genetic diseases such as the muscular dystrophies.

Redox regulation of plant stem cell fate.

PubMed

Zeng, Jian; Dong, Zhicheng; Wu, Haijun; Tian, Zhaoxia; Zhao, Zhong

2017-10-02

Despite the importance of stem cells in plant and animal development, the common mechanisms of stem cell maintenance in both systems have remained elusive. Recently, the importance of hydrogen peroxide (H 2 O 2 ) signaling in priming stem cell differentiation has been extensively studied in animals. Here, we show that different forms of reactive oxygen species (ROS) have antagonistic roles in plant stem cell regulation, which were established by distinct spatiotemporal patterns of ROS-metabolizing enzymes. The superoxide anion (O2·-) is markedly enriched in stem cells to activate WUSCHEL and maintain stemness, whereas H 2 O 2 is more abundant in the differentiating peripheral zone to promote stem cell differentiation. Moreover, H 2 O 2 negatively regulates O2·- biosynthesis in stem cells, and increasing H 2 O 2 levels or scavenging O2·- leads to the termination of stem cells. Our results provide a mechanistic framework for ROS-mediated control of plant stem cell fate and demonstrate that the balance between O2·- and H 2 O 2 is key to stem cell maintenance and differentiation. © 2017 The Authors.

Ocular Stem Cell Research from Basic Science to Clinical Application: A Report from Zhongshan Ophthalmic Center Ocular Stem Cell Symposium

PubMed Central

Ouyang, Hong; Goldberg, Jeffrey L.; Chen, Shuyi; Li, Wei; Xu, Guo-Tong; Li, Wei; Zhang, Kang; Nussenblatt, Robert B.; Liu, Yizhi; Xie, Ting; Chan, Chi-Chao; Zack, Donald J.

2016-01-01

Stem cells hold promise for treating a wide variety of diseases, including degenerative disorders of the eye. The eye is an ideal organ for stem cell therapy because of its relative immunological privilege, surgical accessibility, and its being a self-contained system. The eye also has many potential target diseases amenable to stem cell-based treatment, such as corneal limbal stem cell deficiency, glaucoma, age-related macular degeneration (AMD), and retinitis pigmentosa (RP). Among them, AMD and glaucoma are the two most common diseases, affecting over 200 million people worldwide. Recent results on the clinical trial of retinal pigment epithelial (RPE) cells from human embryonic stem cells (hESCs) and induced pluripotent stem cells (iPSCs) in treating dry AMD and Stargardt’s disease in the US, Japan, England, and China have generated great excitement and hope. This marks the beginning of the ocular stem cell therapy era. The recent Zhongshan Ophthalmic Center Ocular Stem Cell Symposium discussed the potential applications of various stem cell types in stem cell-based therapies, drug discoveries and tissue engineering for treating ocular diseases. PMID:27102165

StemTextSearch: Stem cell gene database with evidence from abstracts.

PubMed

Chen, Chou-Cheng; Ho, Chung-Liang

2017-05-01

Previous studies have used many methods to find biomarkers in stem cells, including text mining, experimental data and image storage. However, no text-mining methods have yet been developed which can identify whether a gene plays a positive or negative role in stem cells. StemTextSearch identifies the role of a gene in stem cells by using a text-mining method to find combinations of gene regulation, stem-cell regulation and cell processes in the same sentences of biomedical abstracts. The dataset includes 5797 genes, with 1534 genes having positive roles in stem cells, 1335 genes having negative roles, 1654 genes with both positive and negative roles, and 1274 with an uncertain role. The precision of gene role in StemTextSearch is 0.66, and the recall is 0.78. StemTextSearch is a web-based engine with queries that specify (i) gene, (ii) category of stem cell, (iii) gene role, (iv) gene regulation, (v) cell process, (vi) stem-cell regulation, and (vii) species. StemTextSearch is available through http://bio.yungyun.com.tw/StemTextSearch.aspx. Copyright © 2017. Published by Elsevier Inc.

Functional intercalated nanocomposites with chitosan-glutathione-glycylsarcosine and layered double hydroxides for topical ocular drug delivery.

PubMed

Xu, Tingting; Xu, Xiaoyue; Gu, Yan; Fang, Lei; Cao, Feng

2018-01-01

To enhance ocular bioavailability, the traditional strategies have focused on prolonging precorneal retention and improving corneal permeability by nano-carriers with positive charge, thiolated polymer, absorption enhancer and so on. Glycylsarcosine (GS) as an active target ligand of the peptide tranpsporter-1 (PepT-1), could specific interact with the PepT-1 on the cornea and guide the nanoparticles to the treating site. The objective of the study was to explore the active targeting intercalated nanocomposites based on chitosan-glutathione-glycylsarcosine (CG-GS) and layered double hydroxides (LDH) as novel carriers for the treatment of mid-posterior diseases. CG-GS-LDH intercalated nanocomposites were prepared by the coprecipitation hydrothermal method. In vivo precorneal retention study, ex vivo fluorescence images, in vivo experiment for distribution and irritation were studied in rabbits. The cytotoxicity and cellular uptake were studied in human corneal epithelial primary cells (HCEpiC). CG-GS-LDH nanocomposites were prepared successfully and characterized by FTIR and XRD. Experiments with rabbits showed longer precorneal retention and higher distribution of fluorescence probe/model drug. In vitro cytological study, CG-GS-LDH nanocomposites exhibited enhanced cellular uptake compared to pure drug solution. Furthermore, the investigation of cellular uptake mechanisms demonstrated that both the active transport by PepT-1 and clathrin-mediated endocytosis were involved in the internalization of CG-GS-LDH intercalated nanocomposites. An ocular irritation study and a cytotoxicity test indicated that these nanocomposites produced no significant irritant effects. The active targeting intercalated nanocomposites could have great potential for topical ocular drug delivery due to the capacity for prolonging the retention on the ocular surface, enhancing the drug permeability through the cornea, and efficiently delivering the drug to the targeted site.

The anti-caries activity and toxicity of an experimental propolis-containing varnish.

PubMed

DE Luca, Mariana Passos; Freires, Irlan Almeida; Gala-García, Alfonso; Santos, Vagner Rodrigues; Vale, Miriam Pimenta; Alencar, Severino Matias de; Rosalen, Pedro Luiz

2017-06-05

We investigated the anti-caries effects of an experimental propolis varnish in vivo, and further tested its toxicity against fibroblasts. Fifty-six SPF female Wistar rats were infected with Streptococcus mutans UA159 (SM) and allocated into four groups (n = 14/group): G1, propolis varnish (15%/PV); G2, chitosan varnish (CV/vehicle); G3, gold standard (GS/Duraphat®); and G4, untreated. The animals received a single varnish application on their molars and were submitted to a high cariogenic challenge (Diet-2000, 56% sucrose, and 5% sucrose-added water, ad libitum) for 4 weeks. Total cultivable microbiota and SM were counted, and smooth-surface and sulcal caries were scored. PV, CV and GS cytotoxic effects were tested against fibroblasts. The data were analyzed using ANOVA with the Tukey-Kramer test (p ≤ 0.05). Total microbiota and SM counts did not differ among the treatments (p = 0.78), or in relation to the untreated group (p = 0.52). PV reduced development of smooth-surface enamel caries compared with the untreated group (p = 0.0018), with no significant difference from GS (p = 0.92); however, the PV effects were no longer observed when the dentin was affected. Neither PV nor GS prevented enamel sulcal lesion onset, but GS significantly reduced the severity of dentinal sulcal lesions (p < 0.0001). No significant difference was observed in fibroblast viability between PV and GS (p < 0.0001). In conclusion, PV prevented smooth-surface enamel caries and showed low cell toxicity. Nevertheless, due to the high cariogenic challenge, its effects were not sustained throughout the experiment. Further studies are encouraged to establish a protocol to sustain the long-term anti-caries activity of PV in the oral cavity.

Functional intercalated nanocomposites with chitosan-glutathione-glycylsarcosine and layered double hydroxides for topical ocular drug delivery

PubMed Central

Gu, Yan

2018-01-01

Background To enhance ocular bioavailability, the traditional strategies have focused on prolonging precorneal retention and improving corneal permeability by nano-carriers with positive charge, thiolated polymer, absorption enhancer and so on. Glycylsarcosine (GS) as an active target ligand of the peptide tranpsporter-1 (PepT-1), could specific interact with the PepT-1 on the cornea and guide the nanoparticles to the treating site. Purpose The objective of the study was to explore the active targeting intercalated nanocomposites based on chitosan-glutathione-glycylsarcosine (CG-GS) and layered double hydroxides (LDH) as novel carriers for the treatment of mid-posterior diseases. Materials and methods CG-GS-LDH intercalated nanocomposites were prepared by the coprecipitation hydrothermal method. In vivo precorneal retention study, ex vivo fluorescence images, in vivo experiment for distribution and irritation were studied in rabbits. The cytotoxicity and cellular uptake were studied in human corneal epithelial primary cells (HCEpiC). Results CG-GS-LDH nanocomposites were prepared successfully and characterized by FTIR and XRD. Experiments with rabbits showed longer precorneal retention and higher distribution of fluorescence probe/model drug. In vitro cytological study, CG-GS-LDH nanocomposites exhibited enhanced cellular uptake compared to pure drug solution. Furthermore, the investigation of cellular uptake mechanisms demonstrated that both the active transport by PepT-1 and clathrin-mediated endocytosis were involved in the internalization of CG-GS-LDH intercalated nanocomposites. An ocular irritation study and a cytotoxicity test indicated that these nanocomposites produced no significant irritant effects. Conclusions The active targeting intercalated nanocomposites could have great potential for topical ocular drug delivery due to the capacity for prolonging the retention on the ocular surface, enhancing the drug permeability through the cornea, and efficiently delivering the drug to the targeted site. PMID:29491707

A 14-3-3 Family Protein from Wild Soybean (Glycine Soja) Regulates ABA Sensitivity in Arabidopsis

PubMed Central

Sun, Xiaoli; Sun, Mingzhe; Jia, Bowei; Chen, Chao; Qin, Zhiwei; Yang, Kejun; Shen, Yang; Meiping, Zhang; Mingyang, Cong; Zhu, Yanming

2015-01-01

It is widely accepted that the 14-3-3 family proteins are key regulators of multiple stress signal transduction cascades. By conducting genome-wide analysis, researchers have identified the soybean 14-3-3 family proteins; however, until now, there is still no direct genetic evidence showing the involvement of soybean 14-3-3s in ABA responses. Hence, in this study, based on the latest Glycine max genome on Phytozome v10.3, we initially analyzed the evolutionary relationship, genome organization, gene structure and duplication, and three-dimensional structure of soybean 14-3-3 family proteins systematically. Our results suggested that soybean 14-3-3 family was highly evolutionary conserved and possessed segmental duplication in evolution. Then, based on our previous functional characterization of a Glycine soja 14-3-3 protein GsGF14o in drought stress responses, we further investigated the expression characteristics of GsGF14o in detail, and demonstrated its positive roles in ABA sensitivity. Quantitative real-time PCR analyses in Glycine soja seedlings and GUS activity assays in PGsGF14O:GUS transgenic Arabidopsis showed that GsGF14o expression was moderately and rapidly induced by ABA treatment. As expected, GsGF14o overexpression in Arabidopsis augmented the ABA inhibition of seed germination and seedling growth, promoted the ABA induced stomata closure, and up-regulated the expression levels of ABA induced genes. Moreover, through yeast two hybrid analyses, we further demonstrated that GsGF14o physically interacted with the AREB/ABF transcription factors in yeast cells. Taken together, results presented in this study strongly suggested that GsGF14o played an important role in regulation of ABA sensitivity in Arabidopsis. PMID:26717241

Nonantibiotic macrolides prevent human neutrophil elastase-induced mucus stasis and airway surface liquid volume depletion

PubMed Central

Sabater, Juan R.; Clarke, Tainya C.; Tan, Chong D.; Davies, Catrin M.; Liu, Jia; Yeung, Arthur; Garland, Alaina L.; Stutts, M. Jackson; Abraham, William M.; Phillips, Gary; Baker, William R.; Wright, Clifford D.; Wilbert, Sibylle

2013-01-01

Mucus clearance is an important component of the lung's innate defense system. A failure of this system brought on by mucus dehydration is common to both cystic fibrosis (CF) and chronic obstructive pulmonary disease (COPD). Mucus clearance rates are regulated by the volume of airway surface liquid (ASL) and by ciliary beat frequency (CBF). Chronic treatment with macrolide antibiotics is known to be beneficial to both CF and COPD patients. However, chronic macrolide usage may induce bacterial resistance. We have developed a novel macrolide, 2′-desoxy-9-(S)-erythromycylamine (GS-459755), that has significantly diminished antibiotic activity against Staphylococcus aureus, Streptococcus pneumonia, Moraxella catarrhalis, and Haemophilus influenzae. Since neutrophilia frequently occurs in chronic lung disease and human neutrophil elastase (HNE) induces mucus stasis by activating the epithelial sodium channel (ENaC), we tested the ability of GS-459755 to protect against HNE-induced mucus stasis. GS-459755 had no effect on HNE activity. However, GS-459755 pretreatment protected against HNE-induced ASL volume depletion in human bronchial epithelial cells (HBECs). The effect of GS-459755 on ASL volume was dose dependent (IC50 ∼3.9 μM) and comparable to the antibacterial macrolide azithromycin (IC50 ∼2.4 μM). Macrolides had no significant effect on CBF or on transepithelial water permeability. However, the amiloride-sensitive transepithelial voltage, a marker of ENaC activity, was diminished by macrolide pretreatment. We conclude that GS-459755 may limit HNE-induced activation of ENaC and may be useful for the treatment of mucus dehydration in CF and COPD without inducing bacterial resistance. PMID:23542952

Application of Stem Cell Technology in Dental Regenerative Medicine.

PubMed

Feng, Ruoxue; Lengner, Chistopher

2013-07-01

In this review, we summarize the current literature regarding the isolation and characterization of dental tissue-derived stem cells and address the potential of these cell types for use in regenerative cell transplantation therapy. Looking forward, platforms for the delivery of stem cells via scaffolds and the use of growth factors and cytokines for enhancing dental stem cell self-renewal and differentiation are discussed. We aim to understand the developmental origins of dental tissues in an effort to elucidate the molecular pathways governing the genesis of somatic dental stem cells. The advantages and disadvantages of several dental stem cells are discussed, including the developmental stage and specific locations from which these cells can be purified. In particular, stem cells from human exfoliated deciduous teeth may act as a very practical and easily accessibly reservoir for autologous stem cells and hold the most value in stem cell therapy. Dental pulp stem cells and periodontal ligament stem cells should also be considered for their triple lineage differentiation ability and relative ease of isolation. Further, we address the potentials and limitations of induced pluripotent stem cells as a cell source in dental regenerative. From an economical and a practical standpoint, dental stem cell therapy would be most easily applied in the prevention of periodontal ligament detachment and bone atrophy, as well as in the regeneration of dentin-pulp complex. In contrast, cell-based tooth replacement due to decay or other oral pathology seems, at the current time, an untenable approach.

The UK Stem Cell Bank: a UK government-funded, international resource center for stem cell research.

PubMed

Stacey, Glyn; Hunt, Charles J

2006-01-01

The UK Stem Cell Bank is a UK Research Council-funded initiative that aims to provide ethically sourced and quality controlled stocks of cells for researchers and also establish seed stocks of cell lines for clinical trials. Whilst the Bank is prohibited from carrying out basic stem cell research (to avoid conflicts of interest) it is working to improve stem cell banking procedures including cryopreservation, characterization and quality control. The Bank also supports training activities and has provided the hub for the International Stem Cell Initiative, which includes 17 expert stem cell centers aiming to characterize a large number of human embryonic stem cell lines in a standardized way to improve our understanding of the characteristics of these cells.

Methods for Stem Cell Production and Therapy

NASA Technical Reports Server (NTRS)

Valluri, Jagan V. (Inventor); Claudio, Pier Paolo (Inventor)

2015-01-01

The present invention relates to methods for rapidly expanding a stem cell population with or without culture supplements in simulated microgravity conditions. The present invention relates to methods for rapidly increasing the life span of stem cell populations without culture supplements in simulated microgravity conditions. The present invention also relates to methods for increasing the sensitivity of cancer stem cells to chemotherapeutic agents by culturing the cancer stem cells under microgravity conditions and in the presence of omega-3 fatty acids. The methods of the present invention can also be used to proliferate cancer cells by culturing them in the presence of omega-3 fatty acids. The present invention also relates to methods for testing the sensitivity of cancer cells and cancer stem cells to chemotherapeutic agents by culturing the cancer cells and cancer stem cells under microgravity conditions. The methods of the present invention can also be used to produce tissue for use in transplantation by culturing stem cells or cancer stem cells under microgravity conditions. The methods of the present invention can also be used to produce cellular factors and growth factors by culturing stem cells or cancer stem cells under microgravity conditions. The methods of the present invention can also be used to produce cellular factors and growth factors to promote differentiation of cancer stem cells under microgravity conditions.

Dental pulp stem cells in regenerative dentistry.

PubMed

Casagrande, Luciano; Cordeiro, Mabel M; Nör, Silvia A; Nör, Jacques E

2011-01-01

Stem cells constitute the source of differentiated cells for the generation of tissues during development, and for regeneration of tissues that are diseased or injured postnatally. In recent years, stem cell research has grown exponentially owing to the recognition that stem cell-based therapies have the potential to improve the life of patients with conditions that span from Alzheimer's disease to cardiac ischemia to bone or tooth loss. Growing evidence demonstrates that stem cells are primarily found in niches and that certain tissues contain more stem cells than others. Among these tissues, the dental pulp is considered a rich source of mesenchymal stem cells that are suitable for tissue engineering applications. It is known that dental pulp stem cells have the potential to differentiate into several cell types, including odontoblasts, neural progenitors, osteoblasts, chondrocytes, and adipocytes. The dental pulp stem cells are highly proliferative. This characteristic facilitates ex vivo expansion and enhances the translational potential of these cells. Notably, the dental pulp is arguably the most accessible source of postnatal stem cells. Collectively, the multipotency, high proliferation rates, and accessibility make the dental pulp an attractive source of mesenchymal stem cells for tissue regeneration. This review discusses fundamental concepts of stem cell biology and tissue engineering within the context of regenerative dentistry.

Translating stem cell therapies: the role of companion animals in regenerative medicine

PubMed Central

Volk, Susan W.; Theoret, Christine

2013-01-01

Veterinarians and veterinary medicine have been integral to the development of stem cell therapies. The contributions of large animal experimental models to the development and refinement of modern hematopoietic stem cell transplantation were noted nearly five decades ago. More recent advances in adult stem cell/regenerative cell therapies continue to expand knowledge of the basic biology and clinical applications of stem cells. A relatively liberal legal and ethical regulation of stem cell research in veterinary medicine has facilitated the development and in some instances clinical translation of a variety of cell-based therapies involving hematopoietic (HSC) and mesenchymal stem cells (MSC) as well as other adult regenerative cells and recently embryonic stem cells (ESC) and induced pluripotent stem cells (iPSC). In fact, many of the pioneering developments in these fields of stem cell research have been achieved through collaborations of veterinary and human scientists. This review aims to provide an overview of the contribution of large animal veterinary models in advancing stem cell therapies for both human and clinical veterinary applications. Moreover, in the context of the “One Health Initiative”, the role veterinary patients may play in the future evolution of stem cell therapies for both human and animal patients will be explored. PMID:23627495

Wnt6 maintains anterior escort cells as an integral component of the germline stem cell niche

PubMed Central

2018-01-01

ABSTRACT Stem cells reside in a niche, a local environment whose cellular and molecular complexity is still being elucidated. In Drosophila ovaries, germline stem cells depend on cap cells for self-renewing signals and physical attachment. Germline stem cells also contact the anterior escort cells, and here we report that anterior escort cells are absolutely required for germline stem cell maintenance. When escort cells die from impaired Wnt signaling or hid expression, the loss of anterior escort cells causes loss of germline stem cells. Anterior escort cells function as an integral niche component by promoting DE-cadherin anchorage and by transiently expressing the Dpp ligand to promote full-strength BMP signaling in germline stem cells. Anterior escort cells are maintained by Wnt6 ligands produced by cap cells; without Wnt6 signaling, anterior escort cells die leaving vacancies in the niche, leading to loss of germline stem cells. Our data identify anterior escort cells as constituents of the germline stem cell niche, maintained by a cap cell-produced Wnt6 survival signal. PMID:29361569

Wnt6 maintains anterior escort cells as an integral component of the germline stem cell niche.

PubMed

Wang, Xiaoxi; Page-McCaw, Andrea

2018-02-07

Stem cells reside in a niche, a local environment whose cellular and molecular complexity is still being elucidated. In Drosophila ovaries, germline stem cells depend on cap cells for self-renewing signals and physical attachment. Germline stem cells also contact the anterior escort cells, and here we report that anterior escort cells are absolutely required for germline stem cell maintenance. When escort cells die from impaired Wnt signaling or hid expression, the loss of anterior escort cells causes loss of germline stem cells. Anterior escort cells function as an integral niche component by promoting DE-cadherin anchorage and by transiently expressing the Dpp ligand to promote full-strength BMP signaling in germline stem cells. Anterior escort cells are maintained by Wnt6 ligands produced by cap cells; without Wnt6 signaling, anterior escort cells die leaving vacancies in the niche, leading to loss of germline stem cells. Our data identify anterior escort cells as constituents of the germline stem cell niche, maintained by a cap cell-produced Wnt6 survival signal. © 2018. Published by The Company of Biologists Ltd.

21st Nantes Actualités Transplantation: "When Stem Cells Meet Immunology".

PubMed

Anegon, Ignacio; Nguyen, Tuan Huy

2017-01-01

"When Stem Cells Meet Immunology" has been the topic of the 21st annual "Nantes Actualités en Transplantation" meeting (June 9-10, 2016, Nantes, France). This meeting brought together pioneers and leading experts in the fields of stem cells, biomaterials and immunoregulation. Presentations covered multipotent (mesenchymal and hematopoietic) and pluripotent stem cells (embryonic and induced) for regenerative medicine of incurable diseases, immunotherapy and blood transfusions. An additional focus had been immune rejections and responses of allogeneic or autologous stem cells. Conversely, stem cells are also able to directly modulate the immune response through the production of immunoregulatory molecules. Moreover, stem cells may also provide an unlimited source of immune cells (DCs, NK cells, B cells, and T cells) that can operate as "super" immune cells, for example, through genetic engineering with chimeric antigen receptors.This meeting report puts presentations into an overall context highlighting new potential biomarkers for potency prediction of mesenchymal stem cell-derived and pluripotent stem cell-derived multicellular organoids. Finally, we propose future directions arising from the flourishing encounter of stem cell and immune biology.

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.

Glutamine metabolism and cycling in Neurospora crassa.

PubMed Central

Mora, J

1990-01-01

Evidence for the existence of a glutamine cycle in Neurospora crassa is reviewed. Through this cycle glutamine is converted into glutamate by glutamate synthase and catabolized by the glutamine transaminase-omega-amidase pathway, the products of which (2-oxoglutarate and ammonium) are the substrates for glutamate dehydrogenase-NADPH, which synthesizes glutamate. In the final step ammonium is assimilated into glutamine by the action of a glutamine synthetase (GS), which is formed by two distinct polypeptides, one catalytically very active (GS beta), and the other (GS alpha) less active but endowed with the capacity to modulate the activity of GS alpha. Glutamate synthase uses the amide nitrogen of glutamine to synthesize glutamate; glutamate dehydrogenase uses ammonium, and both are required to maintain the level of glutamate. The energy expended in the synthesis of glutamine drives the cycle. The glutamine cycle is not futile, because it is necessary to drive an effective carbon flow to support growth; in addition, it facilitates the allocation of nitrogen or carbon according to cellular demands. The glutamine cycle which dissipates energy links catabolism and anabolism and, in doing so, buffers variations in the nutrient supply and drives energy generation and carbon flow for optimal cell function. PMID:2145504

Effect of aging on stem cells

PubMed Central

Ahmed, Abu Shufian Ishtiaq; Sheng, Matilda HC; Wasnik, Samiksha; Baylink, David J; Lau, Kin-Hing William

2017-01-01

Pluripotent stem cells have the remarkable self-renewal ability and are capable of differentiating into multiple diverse cells. There is increasing evidence that the aging process can have adverse effects on stem cells. As stem cells age, their renewal ability deteriorates and their ability to differentiate into the various cell types is altered. Accordingly, it is suggested aging-induced deterioration of stem cell functions may play a key role in the pathophysiology of the various aging-associated disorders. Understanding the role of the aging process in deterioration of stem cell function is crucial, not only in understanding the pathophysiology of aging-associated disorders, but also in future development of novel effective stem cell-based therapies to treat aging-associated diseases. This review article first focuses on the basis of the various aging disease-related stem cell dysfunction. It then addresses the several concepts on the potential mechanism that causes aging-related stem cell dysfunction. It also briefly discusses the current potential therapies under development for aging-associated stem cell defects. PMID:28261550

Engineering Stem Cells for Biomedical Applications

PubMed Central

Yin, Perry T.; Han, Edward

2018-01-01

Stem cells are characterized by a number of useful properties, including their ability to migrate, differentiate, and secrete a variety of therapeutic molecules such as immunomodulatory factors. As such, numerous pre-clinical and clinical studies have utilized stem cell-based therapies and demonstrated their tremendous potential for the treatment of various human diseases and disorders. Recently, efforts have focused on engineering stem cells in order to further enhance their innate abilities as well as to confer them with new functionalities, which can then be used in various biomedical applications. These engineered stem cells can take on a number of forms. For instance, engineered stem cells encompass the genetic modification of stem cells as well as the use of stem cells for gene delivery, nanoparticle loading and delivery, and even small molecule drug delivery. The present Review gives an in-depth account of the current status of engineered stem cells, including potential cell sources, the most common methods used to engineer stem cells, and the utilization of engineered stem cells in various biomedical applications, with a particular focus on tissue regeneration, the treatment of immunodeficiency diseases, and cancer. PMID:25772134

Therapeutic potential of dental stem cells

PubMed Central

Chalisserry, Elna Paul; Nam, Seung Yun; Park, Sang Hyug; Anil, Sukumaran

2017-01-01

Stem cell biology has become an important field in regenerative medicine and tissue engineering therapy since the discovery and characterization of mesenchymal stem cells. Stem cell populations have also been isolated from human dental tissues, including dental pulp stem cells, stem cells from human exfoliated deciduous teeth, stem cells from apical papilla, dental follicle progenitor cells, and periodontal ligament stem cells. Dental stem cells are relatively easily obtainable and exhibit high plasticity and multipotential capabilities. The dental stem cells represent a gold standard for neural-crest-derived bone reconstruction in humans and can be used for the repair of body defects in low-risk autologous therapeutic strategies. The bioengineering technologies developed for tooth regeneration will make substantial contributions to understand the developmental process and will encourage future organ replacement by regenerative therapies in a wide variety of organs such as the liver, kidney, and heart. The concept of developing tooth banking and preservation of dental stem cells is promising. Further research in the area has the potential to herald a new dawn in effective treatment of notoriously difficult diseases which could prove highly beneficial to mankind in the long run. PMID:28616151

Microarray platform affords improved product analysis in mammalian cell growth studies

PubMed Central

Li, Lingyun; Migliore, Nicole; Schaefer, Eugene; Sharfstein, Susan T.; Dordick, Jonathan S.; Linhardt, Robert J.

2014-01-01

High throughput (HT) platforms serve as cost-efficient and rapid screening method for evaluating the effect of cell culture conditions and screening of chemicals. The aim of the current study was to develop a high-throughput cell-based microarray platform to assess the effect of culture conditions on Chinese hamster ovary (CHO) cells. Specifically, growth, transgene expression and metabolism of a GS/MSX CHO cell line, which produces a therapeutic monoclonal antibody, was examined using microarray system in conjunction with conventional shake flask platform in a non-proprietary medium. The microarray system consists of 60 nl spots of cells encapsulated in alginate and separated in groups via an 8-well chamber system attached to the chip. Results show the non-proprietary medium developed allows cell growth, production and normal glycosylation of recombinant antibody and metabolism of the recombinant CHO cells in both the microarray and shake flask platforms. In addition, 10.3 mM glutamate addition to the defined base media results in lactate metabolism shift in the recombinant GS/MSX CHO cells in the shake flask platform. Ultimately, the results demonstrate that the high-throughput microarray platform has the potential to be utilized for evaluating the impact of media additives on cellular processes, such as, cell growth, metabolism and productivity. PMID:24227746

Single-cell sequencing in stem cell biology.

PubMed

Wen, Lu; Tang, Fuchou

2016-04-15

Cell-to-cell variation and heterogeneity are fundamental and intrinsic characteristics of stem cell populations, but these differences are masked when bulk cells are used for omic analysis. Single-cell sequencing technologies serve as powerful tools to dissect cellular heterogeneity comprehensively and to identify distinct phenotypic cell types, even within a 'homogeneous' stem cell population. These technologies, including single-cell genome, epigenome, and transcriptome sequencing technologies, have been developing rapidly in recent years. The application of these methods to different types of stem cells, including pluripotent stem cells and tissue-specific stem cells, has led to exciting new findings in the stem cell field. In this review, we discuss the recent progress as well as future perspectives in the methodologies and applications of single-cell omic sequencing technologies.

The Implications of the Cancer Stem Cell Hypothesis for Neuro-Oncology and Neurology.

PubMed

Rich, Jeremy N

2008-05-01

The cancer stem cell hypothesis posits that cancers contain a subset of neoplastic cells that propagate and maintain tumors through sustained self-renewal and potent tumorigenecity. Recent excitement has been generated by a number of reports that have demonstrated the existence of cancer stem cells in several types of brain tumors. Brain cancer stem cells - also called tumor initiating cells or tumor propagating cells - share features with normal neural stem cells but do not necessarily originate from stem cells. Although most cancers have only a small fraction of cancer stem cells, these tumor cells have been shown in laboratory studies to contribute to therapeutic resistance, formation of new blood vessels to supply the tumor, and tumor spread. As malignant brain tumors rank among the deadliest of all neurologic diseases, the identification of new cellular targets may have profound implications in neuro-oncology. Novel drugs that target stem cell pathways active in brain tumors have been efficacious against cancer stem cells suggesting that anti-cancer stem cell therapies may advance brain tumor therapy. The cancer stem cell hypothesis may have several implications for other neurologic diseases as caution must be exercised in activating stem cell maintenance pathways in cellular therapies for neurodegenerative diseases. The ability for a small fraction of cells to determine the overall course of a disease may also inform new paradigms of disease that may translate into improved patient outcomes.

GS-Nitroxide (JP4-039)-Mediated Radioprotection of Human Fanconi Anemia Cell Lines

PubMed Central

Bernard, Mark E.; Kim, Hyun; Berhane, Hebist; Epperly, Michael W.; Franicola, Darcy; Zhang, Xichen; Houghton, Frank; Shields, Donna; Wang, Hong; Bakkenist, Christopher J.; Frantz, Marie-Celine; Forbeck, Erin M.; Goff, Julie P.; Wipf, Peter; Greenberger, Joel S.

2011-01-01

Fanconi anemia (FA) is an inherited disorder characterized by defective DNA repair and cellular sensitivity to DNA crosslinking agents. Clinically, FA is associated with high risk for marrow failure, leukemia and head and neck squamous cell carcinoma (HNSCC). Radiosensitivity in FA patients compromises the use of total-body irradiation for hematopoietic stem cell transplantation and radiation therapy for HNSCC. A radioprotector for the surrounding tissue would therefore be very valuable during radiotherapy for HNSCC. Clonogenic radiation survival curves were determined for pre- or postirradiation treatment with the parent nitroxide Tempol or JP4-039 in cells of four FA patient-derived cell lines and two transgene-corrected subclonal lines. FancG–/– (PD326) and FancD2–/– (PD20F) patient lines were more sensitive to the DNA crosslinking agent mitomycin C (MMC) than their transgene-restored subclonal cell lines (both P < 0.0001). FancD2–/– cells were more radiosensitive than the transgene restored subclonal cell line (ñ = 2.0 ± 0.7 and 4.7 ± 2.2, respectively, P = 0.03). In contrast, FancG–/– cells were radioresistant relative to the transgene-restored subclonal cell line (ñ = 9.4 ± 1.5 and 2.2 ± 05, respectively, P = 0.001). DNA strand breaks measured by the comet assay correlated with radiosensitivity. Cell lines from a Fanc-C and Fanc-A patients showed radiosensitivity similar to that of Fanc-D2–/– cells. A fluorophore-tagged JP4-039 (BODIPY-FL) analog targeted the mitochondria of the cell lines. Preirradiation or postirradiation treatment with JP4-039 at a lower concentration than Tempol significantly increased the radioresistance and stabilized the antioxidant stores of all cell lines. Tempol increased the toxicity of MMC in FancD2–/– cells. These data provide support for the potential clinical use of JP4-039 for normal tissue radioprotection during chemoradiotherapy in FA patients. PMID:21939290

GS-nitroxide (JP4-039)-mediated radioprotection of human Fanconi anemia cell lines.

PubMed

Bernard, Mark E; Kim, Hyun; Berhane, Hebist; Epperly, Michael W; Franicola, Darcy; Zhang, Xichen; Houghton, Frank; Shields, Donna; Wang, Hong; Bakkenist, Christopher J; Frantz, Marie-Celine; Forbeck, Erin M; Goff, Julie P; Wipf, Peter; Greenberger, Joel S

2011-11-01

Fanconi anemia (FA) is an inherited disorder characterized by defective DNA repair and cellular sensitivity to DNA crosslinking agents. Clinically, FA is associated with high risk for marrow failure, leukemia and head and neck squamous cell carcinoma (HNSCC). Radiosensitivity in FA patients compromises the use of total-body irradiation for hematopoietic stem cell transplantation and radiation therapy for HNSCC. A radioprotector for the surrounding tissue would therefore be very valuable during radiotherapy for HNSCC. Clonogenic radiation survival curves were determined for pre- or postirradiation treatment with the parent nitroxide Tempol or JP4-039 in cells of four FA patient-derived cell lines and two transgene-corrected subclonal lines. FancG(-/-) (PD326) and FancD2(-/-) (PD20F) patient lines were more sensitive to the DNA crosslinking agent mitomycin C (MMC) than their transgene-restored subclonal cell lines (both P < 0.0001). FancD2(-/-) cells were more radiosensitive than the transgene restored subclonal cell line (ñ = 2.0 ± 0.7 and 4.7 ± 2.2, respectively, P = 0.03). In contrast, FancG(-/-) cells were radioresistant relative to the transgene-restored subclonal cell line (ñ = 9.4 ± 1.5 and 2.2 ± 05, respectively, P = 0.001). DNA strand breaks measured by the comet assay correlated with radiosensitivity. Cell lines from a Fanc-C and Fanc-A patients showed radiosensitivity similar to that of Fanc-D2(-/-) cells. A fluorophore-tagged JP4-039 (BODIPY-FL) analog targeted the mitochondria of the cell lines. Preirradiation or postirradiation treatment with JP4-039 at a lower concentration than Tempol significantly increased the radioresistance and stabilized the antioxidant stores of all cell lines. Tempol increased the toxicity of MMC in FancD2(-/-) cells. These data provide support for the potential clinical use of JP4-039 for normal tissue radioprotection during chemoradiotherapy in FA patients.

Can bone marrow differentiate into renal cells?

PubMed

Imai, Enyu; Ito, Takahito

2002-10-01

A considerable plasticity of adult stem cells has been confirmed in a wide variety of tissues. In particular, the pluripotency of bone marrow-derived stem cells may influence the regeneration of injured tissues and may provide novel avenues in regenerative medicine. Bone marrow contains at least hematopoietic and mesenchymal stem cells, and both can differentiate into a wide range of differentiated cells. Side population (SP) cells, which are originally defined in bone marrow cells by high efflux of DNA-binding dye, seem to be a new class of multipotent stem cells. Irrespective of the approach used to obtain stem cells, the fates of marrow-derived cells following bone marrow transplantation can be traced by labeling donor cells with green fluorescence protein or by identifying donor Y chromosome in female recipients. So far, bone marrow-derived cells have been reported to differentiate into renal cells, including mesangial cells, endothelial cells, podocytes, and tubular cells in the kidney, although controversy exists. Further studies are required to address this issue. Cell therapy will be promising when we learn to control stem cells such as bone marrow-derived stem cells, embryonic stem cells, and resident stem cells in the kidney. Identification of factors that support stem cells or promote their differentiation should provide a relevant step towards cell therapy.

[The emerging technology of tissue engineering : Focus on stem cell niche].

PubMed

Schlötzer-Schrehardt, U; Freudenberg, U; Kruse, F E

2017-04-01

Limbal stem cells reside in a highly specialized complex microenvironment that is known as the stem cell niche, an anatomically protected region at the bottom of the Palisades of Vogt, where the stem cells are located and where their quiescence, proliferation and differentiation are maintained in balance. Besides the epithelial stem and progenitor cell clusters, the limbal niche comprises several types of supporting niche cells and a specific extracellular matrix mediating biochemical and biophysical signals. Stem cell-based tissue engineering aims to mimic the native stem cell niche and to present appropriate microenvironmental cues in a controlled and reproducible fashion in order to maintain stem cell function within the graft. Current therapeutic approaches for ex vivo expansion of limbal stem cells only take advantage of surrogate niches. However, new insights into the molecular composition of the limbal niche and innovative biosynthetic scaffolds have stimulated novel strategies for niche-driven stem cell cultivation. Promising experimental approaches include collagen-based organotypic coculture systems of limbal epithelial stem cells with their niche cells and biomimetic hydrogel platforms prefunctionalized with appropriate biomolecular and biophysical signals. Future translation of these novel regenerative strategies into clinical application is expected to improve long-term outcomes of limbal stem cell transplantation for ocular surface reconstruction.

Combination cell therapy with mesenchymal stem cells and neural stem cells for brain stroke in rats.

PubMed

Hosseini, Seyed Mojtaba; Farahmandnia, Mohammad; Razi, Zahra; Delavari, Somayeh; Shakibajahromi, Benafsheh; Sarvestani, Fatemeh Sabet; Kazemi, Sepehr; Semsar, Maryam

2015-05-01

Brain stroke is the second most important events that lead to disability and morbidity these days. Although, stroke is important, there is no treatment for curing this problem. Nowadays, cell therapy has opened a new window for treating central nervous system disease. In some previous studies the Mesenchymal stem cells and neural stem cells. In this study, we have designed an experiment to assess the combination cell therapy (Mesenchymal and Neural stem cells) effects on brain stroke. The Mesenchymal stem cells were isolated from adult rat bone marrow and the neural stem cells were isolated from ganglion eminence of rat embryo 14 days. The Mesenchymal stem cells were injected 1 day after middle cerebral artery occlusion (MCAO) and the neural stem cells transplanted 7 day after MCAO. After 28 days, the neurological outcomes and brain lesion volumes were evaluated. Also, the activity of Caspase 3 was assessed in different groups. The group which received combination cell therapy had better neurological examination and less brain lesion. Also the combination cell therapy group had the least Caspase 3 activity among the groups. The combination cell therapy is more effective than Mesenchymal stem cell therapy and neural stem cell therapy separately in treating the brain stroke in rats.

Application of Stem Cells in Oral Disease Therapy: Progresses and Perspectives

PubMed Central

Yang, Bo; Qiu, Yi; Zhou, Niu; Ouyang, Hong; Ding, Junjun; Cheng, Bin; Sun, Jianbo

2017-01-01

Stem cells are undifferentiated and pluripotent cells that can differentiate into specialized cells with a more specific function. Stem cell therapies become preferred methods for the treatment of multiple diseases. Oral and maxillofacial defect is one kind of the diseases that could be most possibly cured by stem cell therapies. Here we discussed oral diseases, oral adult stem cells, iPS cells, and the progresses/challenges/perspectives of application of stem cells for oral disease treatment. PMID:28421002

Diploid, but not haploid, human embryonic stem cells can be derived from microsurgically repaired tripronuclear human zygotes

PubMed Central

Fan, Yong; Li, Rong; Huang, Jin; Yu, Yang; Qiao, Jie

2013-01-01

Human embryonic stem cells have shown tremendous potential in regenerative medicine, and the recent progress in haploid embryonic stem cells provides new insights for future applications of embryonic stem cells. Disruption of normal fertilized embryos remains controversial; thus, the development of a new source for human embryonic stem cells is important for their usefulness. Here, we investigated the feasibility of haploid and diploid embryo reconstruction and embryonic stem cell derivation using microsurgically repaired tripronuclear human zygotes. Diploid and haploid zygotes were successfully reconstructed, but a large proportion of them still had a tripolar spindle assembly. The reconstructed embryos developed to the blastocyst stage, although the loss of chromosomes was observed in these zygotes. Finally, triploid and diploid human embryonic stem cells were derived from tripronuclear and reconstructed zygotes (from which only one pronucleus was removed), but haploid human embryonic stem cells were not successfully derived from the reconstructed zygotes when two pronuclei were removed. Both triploid and diploid human embryonic stem cells showed the general characteristics of human embryonic stem cells. These results indicate that the lower embryo quality resulting from abnormal spindle assembly contributed to the failure of the haploid embryonic stem cell derivation. However, the successful derivation of diploid embryonic stem cells demonstrated that microsurgical tripronuclear zygotes are an alternative source of human embryonic stem cells. In the future, improving spindle assembly will facilitate the application of triploid zygotes to the field of haploid embryonic stem cells. PMID:23255130

Simple Multi-level Microchannel Fabrication by Pseudo-Grayscale Backside Diffused Light Lithography.

PubMed

Lai, David; Labuz, Joseph M; Kim, Jiwon; Luker, Gary D; Shikanov, Ariella; Takayama, Shuichi

2013-11-14

Photolithography of multi-level channel features in microfluidics is laborious and/or costly. Grayscale photolithography is mostly used with positive photoresists and conventional front side exposure, but the grayscale masks needed are generally costly and positive photoresists are not commonly used in microfluidic rapid prototyping. Here we introduce a simple and inexpensive alternative that uses pseudo-grayscale (pGS) photomasks in combination with backside diffused light lithography (BDLL) and the commonly used negative photoresist, SU-8. BDLL can produce smooth multi-level channels of gradually changing heights without use of true grayscale masks because of the use of diffused light. Since the exposure is done through a glass slide, the photoresist is cross-linked from the substrate side up enabling well-defined and stable structures to be fabricated from even unspun photoresist layers. In addition to providing unique structures and capabilities, the method is compatible with the "garage microfluidics" concept of creating useful tools at low cost since pGS BDLL can be performed with the use of only hot plates and a UV transilluminator: equipment commonly found in biology labs. Expensive spin coaters or collimated UV aligners are not needed. To demonstrate the applicability of pGS BDLL, a variety of weir-type cell traps were constructed with a single UV exposure to separate cancer cells (MDA-MB-231, 10-15 μm in size) from red blood cells (RBCs, 2-8 μm in size) as well as follicle clusters (40-50 μm in size) from cancer cells (MDA-MB-231, 10-15 μm in size).

Nano scaffolds and stem cell therapy in liver tissue engineering

NASA Astrophysics Data System (ADS)

Montaser, Laila M.; Fawzy, Sherin M.

2015-08-01

Tissue engineering and regenerative medicine have been constantly developing of late due to the major progress in cell and organ transplantation, as well as advances in materials science and engineering. Although stem cells hold great potential for the treatment of many injuries and degenerative diseases, several obstacles must be overcome before their therapeutic application can be realized. These include the development of advanced techniques to understand and control functions of micro environmental signals and novel methods to track and guide transplanted stem cells. A major complication encountered with stem cell therapies has been the failure of injected cells to engraft to target tissues. The application of nanotechnology to stem cell biology would be able to address those challenges. Combinations of stem cell therapy and nanotechnology in tissue engineering and regenerative medicine have achieved significant advances. These combinations allow nanotechnology to engineer scaffolds with various features to control stem cell fate decisions. Fabrication of Nano fiber cell scaffolds onto which stem cells can adhere and spread, forming a niche-like microenvironment which can guide stem cells to proceed to heal damaged tissues. In this paper, current and emergent approach based on stem cells in the field of liver tissue engineering is presented for specific application. The combination of stem cells and tissue engineering opens new perspectives in tissue regeneration for stem cell therapy because of the potential to control stem cell behavior with the physical and chemical characteristics of the engineered scaffold environment.

Stem-Cell-Based Tumorigenesis in Adult Drosophila.

PubMed

Hou, S X; Singh, S R

2017-01-01

Recent studies suggest that a small subset of cells within a tumor, the so-called cancer stem cells (CSCs), are responsible for tumor propagation, relapse, and the eventual death of most cancer patients. CSCs may derive from a few tumor-initiating cells, which are either transformed normal stem cells or reprogrammed differentiated cells after acquiring initial cancer-causing mutations. CSCs and normal stem cells share some properties, but CSCs differ from normal stem cells in their tumorigenic ability. Notably, CSCs are usually resistant to chemo- and radiation therapies. Despite the apparent roles of CSCs in human cancers, the biology underlying their behaviors remains poorly understood. Over the past few years, studies in Drosophila have significantly contributed to this new frontier of cancer research. Here, we first review how stem-cell tumors are initiated and propagated in Drosophila, through niche appropriation in the posterior midgut and through stem-cell competition for niche occupancy in the testis. We then discuss the differences between normal and tumorigenic stem cells, revealed by studying Ras V12 -transformed stem-cell tumors in the Drosophila kidney. Finally, we review the biology behind therapy resistance, which has been elucidated through studies of stem-cell resistance and sensitivity to death inducers using female germline stem cells and intestinal stem cells of the posterior midgut. We expect that screens using adult Drosophila neoplastic stem-cell tumor models will be valuable for identifying novel and effective compounds for treating human cancers. © 2017 Elsevier Inc. All rights reserved.

Stem cells with potential to generate insulin producing cells in man.

PubMed

Zulewski, Henryk

2006-10-14

Replacement of insulin-producing cells represents an almost ideal treatment for patients with diabetes mellitus type 1. Transplantation of pancreatic islets of Langerhans--although successful in experienced centres--is limited by the lack of donor organs. Generation of insulin-producing cells from stem cells represents an attractive alternative. Stem cells with the potential to differentiate into insulin-producing cells include embryonic stem cells (ESC) as well as adult stem cells from various tissues including the pancreas, liver, central nervous system, bone marrow and adipose tissue. The use of human ESC is hampered by ethical concerns and the inability to create patient specific ESC with therapeutic cloning. Among adult stem cells mesenchymal stem cells appear to have a particular developmental plasticity ex vivo that include their ability to adopt a pancreatic endocrine phenotype. The present review summarises the current knowledge on the development of insulin-producing cells from stem cells with special emphasis on human mesenchymal stem cells isolated from the pancreas and adipose tissue.

Stem cells with potential to generate insulin-producing cells in man.

PubMed

Zulewski, Henryk

2007-03-02

Replacement of insulin-producing cells represents an almost ideal treatment for patients with diabetes mellitus type 1. Transplantation of pancreatic islets of Langerhans--although successful in experienced centres--is limited by the lack of donor organs. Generation of insulin-producing cells from stem cells represents an attractive alternative. Stem cells with the potential to differentiate into insulin-producing cells include embryonic stem cells (ESC) as well as adult stem cells from various tissues including the pancreas, liver, central nervous system, bone marrow and adipose tissue. The use of human ESC is hampered by ethical concerns and the inability to create patient specific ESC with therapeutic cloning. Among adult stem cells mesenchymal stem cells appear to have a particular developmental plasticity ex vivo that include their ability to adopt a pancreatic endocrine phenotype. The present review summarises the current knowledge on the development of insulin-producing cells from stem cells with special emphasis on human mesenchymal stem cells isolated from the pancreas and adipose tissue.

Mechanical forces direct stem cell behaviour in development and regeneration

PubMed Central

Vining, Kyle H.; Mooney, David J.

2018-01-01

Stem cells and their local microenvironment, or niche, communicate through mechanical, cues to regulate cell fate and cell behaviour, and to guide developmental processes. During embryonic development, mechanical forces are involved in patterning and organogenesis. The physical environment of pluripotent stem cells regulates their differentiation and self-renewal. Mechanical and physical cues are also important in adult tissues, where adult stem cells require physical interactions with the extracellular matrix to maintain their potency. In vitro, synthetic models of the stem cell niche can be used to precisely control and manipulate the biophysical and biochemical properties of the stem cell microenvironment and examine how the mode and magnitude of mechanical cues, such as matrix stiffness or applied forces, direct stem cell differentiation and function. Fundamental insights on the mechanobiology of stem cells also inform the design of artificial niches to support stem cells for regenerative therapies. PMID:29115301

Recent Advances towards the Clinical Application of Stem Cells for Retinal Regeneration

PubMed Central

Becker, Silke; Jayaram, Hari; Limb, G. Astrid

2012-01-01

Retinal degenerative diseases constitute a major cause of irreversible blindness in the world. Stem cell-based therapies offer hope for these patients at risk of or suffering from blindness due to the deterioration of the neural retina. Various sources of stem cells are currently being investigated, ranging from human embryonic stem cells to adult-derived induced pluripotent stem cells as well as human Müller stem cells, with the first clinical trials to investigate the safety and tolerability of human embryonic stem cell-derived retinal pigment epithelium cells having recently commenced. This review aims to summarize the latest advances in the development of stem cell strategies for the replacement of retinal neurons and their supportive cells, the retinal pigment epithelium (RPE) affected by retinal degenerative conditions. Particular emphasis will be given to the advances in stem cell transplantation and the challenges associated with their translation into clinical practice. PMID:24710533

Stem-Cell Therapy Advances in China.

PubMed

Hu, Lei; Zhao, Bin; Wang, Songlin

2018-02-01

Stem-cell therapy is a promising method for treating patients with a wide range of diseases and injuries. Increasing government funding of scientific research has promoted rapid developments in stem-cell research in China, as evidenced by the substantial increase in the number and quality of publications in the past 5 years. Multiple high-quality studies have been performed in China that concern cell reprogramming, stem-cell homeostasis, gene modifications, and immunomodulation. The number of translation studies, including basic and preclinical investigations, has also increased. Around 100 stem-cell banks have been established in China, 10 stem-cell drugs are currently in the approval process, and >400 stem cell-based clinical trials are currently registered in China. With continued state funding, advanced biotechnical support, and the development of regulatory standards for the clinical application of stem cells, further innovations are expected that will lead to a boom in stem-cell therapies. This review highlights recent achievements in stem-cell research in China and discusses future prospects.

New insights into mechanisms of stem cell daughter fate determination in regenerative tissues.

PubMed

Sada, Aiko; Tumbar, Tudorita

2013-01-01

Stem cells can self-renew and differentiate over extended periods of time. Understanding how stem cells acquire their fates is a central question in stem cell biology. Early work in Drosophila germ line and neuroblast showed that fate choice is achieved by strict asymmetric divisions that can generate each time one stem and one differentiated cell. More recent work suggests that during homeostasis, some stem cells can divide symmetrically to generate two differentiated cells or two identical stem cells to compensate for stem cell loss that occurred by direct differentiation or apoptosis. The interplay of all these factors ensures constant tissue regeneration and the maintenance of stem cell pool size. This interplay can be modeled as a population-deterministic dynamics that, at least in some systems, may be described as stochastic behavior. Here, we overview recent progress made on the characterization of stem cell dynamics in regenerative tissues. Copyright © 2013 Elsevier Inc. All rights reserved.

Selection of Phage Display Peptides Targeting Human Pluripotent Stem Cell-Derived Progenitor Cell Lines.

PubMed

Bignone, Paola A; Krupa, Rachel A; West, Michael D; Larocca, David

2016-01-01

The ability of human pluripotent stem cells (hPS) to both self-renew and differentiate into virtually any cell type makes them a promising source of cells for cell-based regenerative therapies. However, stem cell identity, purity, and scalability remain formidable challenges that need to be overcome for translation of pluripotent stem cell research into clinical applications. Directed differentiation from hPS cells is inefficient and residual contamination with pluripotent cells that have the potential to form tumors remains problematic. The derivation of scalable (self-renewing) embryonic progenitor stem cell lines offers a solution because they are well defined and clonally pure. Clonally pure progenitor stem cell lines also provide a means for identifying cell surface targeting reagents that are useful for identification, tracking, and repeated derivation of the corresponding progenitor stem cell types from additional hPS cell sources. Such stem cell targeting reagents can then be applied to the manufacture of genetically diverse banks of human embryonic progenitor cell lines for drug screening, disease modeling, and cell therapy. Here we present methods to identify human embryonic progenitor stem cell targeting peptides by selection of phage display libraries on clonal embryonic progenitor cell lines and demonstrate their use for targeting quantum dots (Qdots) for stem cell labeling.

Extinction models for cancer stem cell therapy

PubMed Central

Sehl, Mary; Zhou, Hua; Sinsheimer, Janet S.; Lange, Kenneth L.

2012-01-01

Cells with stem cell-like properties are now viewed as initiating and sustaining many cancers. This suggests that cancer can be cured by driving these cancer stem cells to extinction. The problem with this strategy is that ordinary stem cells are apt to be killed in the process. This paper sets bounds on the killing differential (difference between death rates of cancer stem cells and normal stem cells) that must exist for the survival of an adequate number of normal stem cells. Our main tools are birth–death Markov chains in continuous time. In this framework, we investigate the extinction times of cancer stem cells and normal stem cells. Application of extreme value theory from mathematical statistics yields an accurate asymptotic distribution and corresponding moments for both extinction times. We compare these distributions for the two cell populations as a function of the killing rates. Perhaps a more telling comparison involves the number of normal stem cells NH at the extinction time of the cancer stem cells. Conditioning on the asymptotic time to extinction of the cancer stem cells allows us to calculate the asymptotic mean and variance of NH. The full distribution of NH can be retrieved by the finite Fourier transform and, in some parameter regimes, by an eigenfunction expansion. Finally, we discuss the impact of quiescence (the resting state) on stem cell dynamics. Quiescence can act as a sanctuary for cancer stem cells and imperils the proposed therapy. We approach the complication of quiescence via multitype branching process models and stochastic simulation. Improvements to the τ-leaping method of stochastic simulation make it a versatile tool in this context. We conclude that the proposed therapy must target quiescent cancer stem cells as well as actively dividing cancer stem cells. The current cancer models demonstrate the virtue of attacking the same quantitative questions from a variety of modeling, mathematical, and computational perspectives. PMID:22001354

Extinction models for cancer stem cell therapy.

PubMed

Sehl, Mary; Zhou, Hua; Sinsheimer, Janet S; Lange, Kenneth L

2011-12-01

Cells with stem cell-like properties are now viewed as initiating and sustaining many cancers. This suggests that cancer can be cured by driving these cancer stem cells to extinction. The problem with this strategy is that ordinary stem cells are apt to be killed in the process. This paper sets bounds on the killing differential (difference between death rates of cancer stem cells and normal stem cells) that must exist for the survival of an adequate number of normal stem cells. Our main tools are birth-death Markov chains in continuous time. In this framework, we investigate the extinction times of cancer stem cells and normal stem cells. Application of extreme value theory from mathematical statistics yields an accurate asymptotic distribution and corresponding moments for both extinction times. We compare these distributions for the two cell populations as a function of the killing rates. Perhaps a more telling comparison involves the number of normal stem cells NH at the extinction time of the cancer stem cells. Conditioning on the asymptotic time to extinction of the cancer stem cells allows us to calculate the asymptotic mean and variance of NH. The full distribution of NH can be retrieved by the finite Fourier transform and, in some parameter regimes, by an eigenfunction expansion. Finally, we discuss the impact of quiescence (the resting state) on stem cell dynamics. Quiescence can act as a sanctuary for cancer stem cells and imperils the proposed therapy. We approach the complication of quiescence via multitype branching process models and stochastic simulation. Improvements to the τ-leaping method of stochastic simulation make it a versatile tool in this context. We conclude that the proposed therapy must target quiescent cancer stem cells as well as actively dividing cancer stem cells. The current cancer models demonstrate the virtue of attacking the same quantitative questions from a variety of modeling, mathematical, and computational perspectives. Copyright © 2011 Elsevier Inc. All rights reserved.

Aging and stem cell therapy: AMPK as an applicable pharmacological target for rejuvenation of aged stem cells and achieving higher efficacy in stem cell therapy.

PubMed

Khorraminejad-Shirazi, Mohammadhossein; Farahmandnia, Mohammad; Kardeh, Bahareh; Estedlal, Alireza; Kardeh, Sina; Monabati, Ahmad

2017-10-19

In recent years, tissue regeneration has become a promising field for developing stem cell-based transplantation therapies for human patients. Adult stem cells are affected by the same aging mechanisms that involve somatic cells. One of the mechanisms involved in cellular aging is hyperactivation of mechanistic target of rapamycin complex 1 (mTORC1) and disruption of 5' adenosine monophosphate-activated protein kinase (AMPK). Aging of stem cells results in their impaired regenerative capacity and depletion of stem cell pools in adult tissue, which results in lower efficacy of stem cell therapy. By utilizing an effective therapeutic intervention for aged stem cells, stem cell therapy can become more promising for future application. mTORC1 inhibition is a practical approach to preserve the stem cell pool. In this article, we review the dynamic interaction between sirtuin (silent mating type information regulation 2 homolog) 1, AMPK, and mTORC1. We propose that using AMPK activators such as 5-aminoimidazole-4-carboxamide ribonucleotide, A769662, metformin, and oxidized nicotinamide adenine dinucleotide (NAD + ) are practical ways to be employed for achieving better optimized results in stem cell-based transplantation therapies. Copyright © 2017 King Faisal Specialist Hospital & Research Centre. Published by Elsevier B.V. All rights reserved.

Stemness of spermatogonial stem cells encapsulated in alginate hydrogel during cryopreservation.

PubMed

Pirnia, A; Parivar, K; Hemadi, M; Yaghmaei, P; Gholami, M

2017-06-01

This study investigated the effect of spermatogonial stem cell encapsulated in alginate hydrogel during cryopreservation, as cells were protected against damage during cryopreservation within the hydrogel. Spermatogonial stem cells were isolated from the testes of Balb/c mice pups (6 days old), purified in laminin-coated dishes and CD90.1 microbeads, encapsulated in alginate hydrogel and then cryopreserved. After thawing, cell viability and Spermatogonial stem cell (SSC) colony diameter were evaluated. After RNA was isolated and cDNA was synthesised, the expression of stemness genes was considered using RT real-time PCR. Finally, spermatogonial stem cells labelled with BrdU were transplanted to busulfan azoospermic mouse models. Lin28a and Sall4 genes were significantly upregulated after cryopreservation in alginate hydrogel. However, cell viability was significantly decreased. The diameter of colonies consisting of spermatogonial stem cells freeze-thawed in alginate microbeads showed no significant difference with fresh spermatogonial stem cells and the control group. The injection of freeze-thawed spermatogonial stem cells encapsulated in alginate hydrogel resulted in spermatogenesis recovery. Alginate mimics the extracellular matrices (ECM) for spermatogonial stem cells; therefore, it can support stemness potential during the cell cryopreservation process and restart spermatogenesis after transplantation. © 2016 Blackwell Verlag GmbH.

Clinical, laboratory and molecular signs of immunodeficiency in patients with partial oculo-cutaneous albinism

PubMed Central

2013-01-01

Hypopigmentation disorders that are associated with immunodeficiency feature both partial albinism of hair, skin and eyes together with leukocyte defects. These disorders include Chediak Higashi (CHS), Griscelli (GS), Hermansky-Pudlak (HPS) and MAPBP-interacting protein deficiency syndromes. These are heterogeneous autosomal recessive conditions in which the causal genes encode proteins with specific roles in the biogenesis, function and trafficking of secretory lysosomes. In certain specialized cells, these organelles serve as a storage compartment. Impaired secretion of specific effector proteins from that intracellular compartment affects biological activities. In particular, these intracellular granules are essential constituents of melanocytes, platelets, granulocytes, cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells. Thus, abnormalities affect pigmentation, primary hemostasis, blood cell counts and lymphocyte cytotoxic activity against microbial pathogens. Among eight genetically distinct types of HPS, only type 2 is characterized by immunodeficiency. Recently, a new subtype, HPS9, was defined in patients presenting with immunodeficiency and oculocutaneous albinism, associated with mutations in the pallidin-encoding gene, PLDN. Hypopigmentation together with recurrent childhood bacterial or viral infections suggests syndromic albinism. T and NK cell cytotoxicity are generally impaired in patients with these disorders. Specific clinical and biochemical phenotypes can allow differential diagnoses among these disorders before molecular testing. Ocular symptoms, including nystagmus, that are usually evident at birth, are common in patients with HPS2 or CHS. Albinism with short stature is unique to MAPBP-interacting protein (MAPBPIP) deficiency, while hemophagocytic lymphohistiocytosis (HLH) mainly suggests a diagnosis of CHS or GS type 2 (GS2). Neurological disease is a long-term complication of CHS, but is uncommon in other syndromic albinism. Chronic neutropenia is a feature of HPS2 and MAPBPIP-deficiency syndrome, whereas it is usually transient in CHS and GS2. In every patient, an accurate diagnosis is required for prompt and appropriate treatment, particularly in patients who develop HLH or in whom bone marrow transplant is required. This review describes the molecular and pathogenetic mechanisms of these diseases, focusing on clinical and biochemical aspects that allow early differential diagnosis. PMID:24134793

Normal and cancer mammary stem cells evade interferon-induced constraint through the miR-199a-LCOR Axis

PubMed Central

Celià-Terrassa, Toni; Liu, Daniel; Choudhury, Abrar; Hang, Xiang; Wei, Yong; Zamalloa, Jose; Alfaro-Aco, Raymundo; Chakrabarti, Rumela; Jiang, Yi-Zhou; Koh, Bong Ihn; Smith, Heath; DeCoste, Christina; Li, Jun-Jing; Shao, Zhi-Ming; Kang, Yibin

2017-01-01

Tumor-initiating cells (TICs), or cancer stem cells (CSC), possess stem cell-like properties observed in normal adult tissue stem cells. Normal and cancerous stem cells may therefore share regulatory mechanisms for maintaining self-renewing capacity and resisting differentiation elicited by cell-intrinsic or microenvironmental cues. Here, we show that miR-199a promotes stem cell properties in mammary stem cells (MaSCs) and breast CSCs by directly repressing nuclear receptor corepressor LCOR, which primes interferon (IFN) responses. Elevated miR-199a expression in stem cell-enriched populations protects normal and malignant stem-like cells from differentiation and senescence induced by IFNs that are produced by epithelial and immune cells in the mammary gland. Importantly, the miR-199a-LCOR-IFN axis is activated in poorly differentiated ER− breast tumors, functionally promotes tumor initiation and metastasis, and is associated with poor clinical outcome. Our study therefore reveals a common mechanism shared by normal and malignant stem cells to protect them from suppressive immune cytokine signaling. PMID:28530657

The Role of Epithelial-Mesenchymal Transition in the Formation of Normal and Neoplastic Mammary Epithelial Stem Cells

DTIC Science & Technology

2011-09-01

separating stem cell and non- stem cell populations of normal and breast cancer cells and identified EMT transcription factors most likely involved in... stem cell biology. Preliminary results directly demonstrate that transient induction of EMT increases the number of mammary epithelial stem cells...EMT and entrance into a stem - cell state. The outcome of these experiments holds important implications for the mechanisms controlling the formation of

Role of the Stem Cell Niche in Hormone-Induced Tumorigenesis in Fetal Mouse Mammary Epithelium

DTIC Science & Technology

2005-08-01

responsive, self renewing and pluripotent. A structure specialized to contain and regulate stem cell activity has been structurally and molecularly...described in Drosophila and some mammalian tissues. The structure, the stem cell niche, functions to 1) shield the stem cell from the burden of incoming...directing stem cell renewal and maturation, 3) prevent stem cells from wandering through the tissue and producing new cells inappropriately, 4) prevent

The Effect of Laser Irradiation on Adipose Derived Stem Cell Proliferation and Differentiation

NASA Astrophysics Data System (ADS)

Abrahamse, H.; de Villiers, J.; Mvula, B.

2009-06-01

There are two fundamental types of stem cells: Embryonic Stem cells and Adult Stem cells. Adult Stem cells have a more restricted potential and can usually differentiate into a few different cell types. In the body these cells facilitate the replacement or repair of damaged or diseased cells in organs. Low intensity laser irradiation was shown to increase stem cell migration and stimulate proliferation and it is thought that treatment of these cells with laser irradiation may increase the stem cell harvest and have a positive effect on the viability and proliferation. Our research is aimed at determining the effect of laser irradiation on differentiation of Adipose Derived Stem Cells (ADSCs) into different cell types using a diode laser with a wavelength of 636 nm and at 5 J/cm2. Confirmation of stem cell characteristics and well as subsequent differentiation were assessed using Western blot analysis and cellular morphology supported by fluorescent live cell imaging. Functionality of subsequent differentiated cells was confirmed by measuring adenosine triphosphate (ATP) production and cell viability.

Alpha-fetoprotein, stem cells and cancer: how study of the production of alpha-fetoprotein during chemical hepatocarcinogenesis led to reaffirmation of the stem cell theory of cancer.

PubMed

Sell, Stewart

2008-01-01

Identification of the cells in the liver that produce alpha-fetoprotein during development, in response to liver injury and during the early stages of chemical hepatocarcinogenesis led to the conclusion that maturation arrest of liver-determined tissue stem cells was the cellular process that gives rise to hepatocellular carcinomas. When the cellular changes in these processes were compared to that of the formation of teratocarcinomas, the hypothesis arose that all cancers arise from maturation arrest of tissue-determined stem cells. This was essentially a reinterpretation of the embryonal rest theory of cancer whereby tissue stem cells take the role of embryonal rests. A corollary of the stem cell theory of the origin of cancer is that cancers contain the same functional cell populations as normal tissues: stem cells, transit-amplifying cells and mature cells. Cancer stem cells retain the essential feature of normal stem cells: the ability to self-renew. Growth of cancers is due to continued proliferation of cancer transit-amplifying cells that do not differentiate to mature cells (maturation arrest). On the other hand, cancer stem cells generally divide very rarely and contribute little to tumor growth. However, the presence of cancer stem cells in tumors is believed to be responsible for the properties of immortalization, transplantability and resistance to therapy characteristic of cancers. Current therapies for cancer (chemotherapy, radiotherapy, antiangiogenesis and differentiation therapy) are directed against the cancer transit-amplifying cells. When these therapies are discontinued, the cancer reforms from the cancer stem cells. Therapy directed toward interruption of the cell signaling pathways that maintain cancer stem cells could lead to new modalities to the prevention of regrowth of the cancer. Copyright 2008 S. Karger AG, Basel.

ALPHA-FETOPROTEIN (AFP), STEM CELLS, AND CANCER: HOW STUDY OF THE PRODUCTION OF AFP DURING CHEMICAL HEPATOCARCINOGENESIS LED TO REAFFIRMATION OF THE STEM CELL THEORY OF CANCER

PubMed Central

Sell, Stewart

2008-01-01

Identification of the cells in the liver that produce alpha-fetoprotein (AFP) during development, in response to liver injury, and during the early stages of chemical hepatocarcinogenesis led to the conclusion that maturation arrest of liver-determined tissue stem cells was the cellular process that gives rise to hepatocellular carcinomas (HCC). When the cellular changes in these processes were compared that of the formation of teratocarcinomas, the hypothesis arose that all cancers arise from maturation arrest of tissue determined stem cells. This was essentially a reinterpretation of the embryonal rest theory of cancer whereby tissue stem cells take the role of embryonal rests. A corollary of the stem cell theory of the origin of cancer is that cancers contain the same functional cell populations as do normal tissues: stem cells, transit-amplifying cells, and mature cells. Cancer stem cells retain the essential feature of normal stem cells: the ability to self-renew. Growth of cancers is due to continued proliferation of cancer transit-amplifying cells that do not differentiate to mature cells (maturation arrest). On the other hand, cancer stem cells generally divide very rarely and contribute little to tumor growth. However, the presence of cancer stem cells in tumors is believed to be responsible for the properties of immortalization, transplantability and resistance to therapy characteristic of cancers. Current therapies for cancer (chemotherapy, radiotherapy, anti-angiogenesis and differentiation therapy) are directed against the cancer transit amplifying cells. When these therapies are discontinued, the cancer re-forms from the cancer stem cells. Therapy directed toward interruption of the cell-signaling pathways that maintain cancer stem cells could lead to new modalities to the prevention of re-growth of the cancer. PMID:18612221

A Comparative Transcriptomic Analysis Reveals Conserved Features of Stem Cell Pluripotency in Planarians and Mammals

PubMed Central

Labbé, Roselyne M.; Irimia, Manuel; Currie, Ko W.; Lin, Alexander; Zhu, Shu Jun; Brown, David D.R.; Ross, Eric J.; Voisin, Veronique; Bader, Gary D.; Blencowe, Benjamin J.; Pearson, Bret J.

2014-01-01

Many long-lived species of animals require the function of adult stem cells throughout their lives. However, the transcriptomes of stem cells in invertebrates and vertebrates have not been compared, and consequently, ancestral regulatory circuits that control stem cell populations remain poorly defined. In this study, we have used data from high-throughput RNA sequencing to compare the transcriptomes of pluripotent adult stem cells from planarians with the transcriptomes of human and mouse pluripotent embryonic stem cells. From a stringently defined set of 4,432 orthologs shared between planarians, mice and humans, we identified 123 conserved genes that are ≥5-fold differentially expressed in stem cells from all three species. Guided by this gene set, we used RNAi screening in adult planarians to discover novel stem cell regulators, which we found to affect the stem cell-associated functions of tissue homeostasis, regeneration, and stem cell maintenance. Examples of genes that disrupted these processes included the orthologs of TBL3, PSD12, TTC27, and RACK1. From these analyses, we concluded that by comparing stem cell transcriptomes from diverse species, it is possible to uncover conserved factors that function in stem cell biology. These results provide insights into which genes comprised the ancestral circuitry underlying the control of stem cell self-renewal and pluripotency. PMID:22696458

Optimization of Swine Breeding Programs Using Genomic Selection with ZPLAN+

PubMed Central

Lopez, B. M.; Kang, H. S.; Kim, T. H.; Viterbo, V. S.; Kim, H. S.; Na, C. S.; Seo, K. S.

2016-01-01

The objective of this study was to evaluate the present conventional selection program of a swine nucleus farm and compare it with a new selection strategy employing genomic enhanced breeding value (GEBV) as the selection criteria. The ZPLAN+ software was employed to calculate and compare the genetic gain, total cost, return and profit of each selection strategy. The first strategy reflected the current conventional breeding program, which was a progeny test system (CS). The second strategy was a selection scheme based strictly on genomic information (GS1). The third scenario was the same as GS1, but the selection by GEBV was further supplemented by the performance test (GS2). The last scenario was a mixture of genomic information and progeny tests (GS3). The results showed that the accuracy of the selection index of young boars of GS1 was 26% higher than that of CS. On the other hand, both GS2 and GS3 gave 31% higher accuracy than CS for young boars. The annual monetary genetic gain of GS1, GS2 and GS3 was 10%, 12%, and 11% higher, respectively, than that of CS. As expected, the discounted costs of genomic selection strategies were higher than those of CS. The costs of GS1, GS2 and GS3 were 35%, 73%, and 89% higher than those of CS, respectively, assuming a genotyping cost of $120. As a result, the discounted profit per animal of GS1 and GS2 was 8% and 2% higher, respectively, than that of CS while GS3 was 6% lower. Comparison among genomic breeding scenarios revealed that GS1 was more profitable than GS2 and GS3. The genomic selection schemes, especially GS1 and GS2, were clearly superior to the conventional scheme in terms of monetary genetic gain and profit. PMID:26954222

Investigating the mincing method for isolation of adipose-derived stem cells from pregnant women fat.

PubMed

Li, Yuan-Sheng; Chen, Pao-Jen; Wu, Li-Wei; Chou, Pei-Wen; Sun, Li-Yi; Chiou, Tzyy-Wen

2018-02-01

The success of stem cell application in regenerative medicine, usually require a stable source of stem or progenitor cells. Fat tissue represents a good source of stem cells because it is rich in stem cells and there are fewer ethical issues related to the use of such stem cells, unlike embryonic stem cells. Therefore, there has been increased interest in adipose-derived stem cells (ADSCs) for tissue engineering applications. Here, we aim to provide an easy processing method for isolating adult stem cells from human adipose tissue harvested from the subcutaneous fat of the abdominal wall during gynecologic surgery. We used a homogenizer to mince fat and compared the results with those obtained from the traditional cut method involving a sterile scalpel and forceps. Our results showed that our method provides another stable and quality source of stem cells that could be used in cases with a large quantity of fat. Furthermore, we found that pregnancy adipose-derived stem cells (P-ADSCs) could be maintained in vitro for extended periods with a stable population doubling and low senescence levels. P-ADSCs could also differentiate in vitro into adipogenic, osteogenic, chondrogenic, and insulin-producing cells in the presence of lineage-specific induction factors. In conclusion, like human lipoaspirates, adipose tissues obtained from pregnant women contain multipotent cells with better proliferation and showed great promise for use in both stem cell banking studies as well as in stem cell therapy.

Gremlin 1 Identifies a Skeletal Stem Cell with Bone, Cartilage, and Reticular Stromal Potential

PubMed Central

Worthley, Daniel L.; Churchill, Michael; Compton, Jocelyn T.; Tailor, Yagnesh; Rao, Meenakshi; Si, Yiling; Levin, Daniel; Schwartz, Matthew G.; Uygur, Aysu; Hayakawa, Yoku; Gross, Stefanie; Renz, Bernhard W.; Setlik, Wanda; Martinez, Ashley N.; Chen, Xiaowei; Nizami, Saqib; Lee, Heon Goo; Kang, H. Paco; Caldwell, Jon-Michael; Asfaha, Samuel; Westphalen, C. Benedikt; Graham, Trevor; Jin, Guangchun; Nagar, Karan; Wang, Hongshan; Kheirbek, Mazen A.; Kolhe, Alka; Carpenter, Jared; Glaire, Mark; Nair, Abhinav; Renders, Simon; Manieri, Nicholas; Muthupalani, Sureshkumar; Fox, James G.; Reichert, Maximilian; Giraud, Andrew S.; Schwabe, Robert F.; Pradere, Jean-Phillipe; Walton, Katherine; Prakash, Ajay; Gumucio, Deborah; Rustgi, Anil K.; Stappenbeck, Thaddeus S.; Friedman, Richard A.; Gershon, Michael D.; Sims, Peter; Grikscheit, Tracy; Lee, Francis Y.; Karsenty, Gerard; Mukherjee, Siddhartha; Wang, Timothy C.

2014-01-01

The stem cells that maintain and repair the postnatal skeleton remain undefined. One model suggests that perisinusoidal mesenchymal stem cells (MSCs) give rise to osteoblasts, chondrocytes, marrow stromal cells, and adipocytes, although the existence of these cells has not been proven through fate-mapping experiments. We demonstrate here that expression of the bone morphogenetic protein (BMP) antagonist gremlin 1 defines a population of osteochondroreticular (OCR) stem cells in the bone marrow. OCR stem cells self-renew and generate osteoblasts, chondrocytes, and reticular marrow stromal cells, but not adipocytes. OCR stem cells are concentrated within the metaphysis of long bones not in the perisinusoidal space and are needed for bone development, bone remodeling, and fracture repair. Grem1 expression also identifies intestinal reticular stem cells (iRSCs) that are cells of origin for the periepithelial intestinal mesenchymal sheath. Grem1 expression identifies distinct connective tissue stem cells in both the bone (OCR stem cells) and the intestine (iRSCs). PMID:25594183

Biochemistry of epidermal stem cells.

PubMed

Eckert, Richard L; Adhikary, Gautam; Balasubramanian, Sivaprakasam; Rorke, Ellen A; Vemuri, Mohan C; Boucher, Shayne E; Bickenbach, Jackie R; Kerr, Candace

2013-02-01

The epidermis is an important protective barrier that is essential for maintenance of life. Maintaining this barrier requires continuous cell proliferation and differentiation. Moreover, these processes must be balanced to produce a normal epidermis. The stem cells of the epidermis reside in specific locations in the basal epidermis, hair follicle and sebaceous glands and these cells are responsible for replenishment of this tissue. A great deal of effort has gone into identifying protein epitopes that mark stem cells, in identifying stem cell niche locations, and in understanding how stem cell populations are related. We discuss these studies as they apply to understanding normal epidermal homeostasis and skin cancer. An assortment of stem cell markers have been identified that permit assignment of stem cells to specific regions of the epidermis, and progress has been made in understanding the role of these cells in normal epidermal homeostasis and in conditions of tissue stress. A key finding is the multiple stem cell populations exist in epidermis that give rise to different structures, and that multiple stem cell types may contribute to repair in damaged epidermis. Understanding epidermal stem cell biology is likely to lead to important therapies for treating skin diseases and cancer, and will also contribute to our understanding of stem cells in other systems. This article is part of a Special Issue entitled Biochemistry of Stem Cells. Copyright © 2012 Elsevier B.V. All rights reserved.

Basics and applications of stem cells in the pancreas.

PubMed

Sekine, Keisuke; Taniguchi, Hideki

2012-11-01

Enormous efforts have been made to establish pancreatic stem/progenitor cells as a source for regenerative medicine for the treatment of diabetes mellitus. In recent years, it has been recognized that the self-renewal of beta cells is the dominant process involved in postnatal beta-cell regeneration and expansion. Nevertheless, several in-vitro studies have suggested that ductal or as yet unidentified cells are candidates for pancreatic stem/progenitor cells that can differentiate into multilineage cells, including insulin(+) cells. The question remains as to whether beta cells are generated postnatally from stem/progenitor cells other than pre-existing beta cells. Furthermore, mutated pancreatic stem cells are considered to be prospective candidates for cancer stem cells or tumor-initiating cells. This review highlights recent progress in pancreatic stem/progenitor cell research.

Ablation of proliferating neural stem cells during early life is sufficient to reduce adult hippocampal neurogenesis.

PubMed

Youssef, Mary; Krish, Varsha S; Kirshenbaum, Greer S; Atsak, Piray; Lass, Tamara J; Lieberman, Sophie R; Leonardo, E David; Dranovsky, Alex

2018-05-09

Environmental exposures during early life, but not during adolescence or adulthood, lead to persistent reductions in neurogenesis in the adult hippocampal dentate gyrus (DG). The mechanisms by which early life exposures lead to long-term deficits in neurogenesis remain unclear. Here, we investigated whether targeted ablation of dividing neural stem cells during early life is sufficient to produce long-term decreases in DG neurogenesis. Having previously found that the stem cell lineage is resistant to long-term effects of transient ablation of dividing stem cells during adolescence or adulthood (Kirshenbaum et al., 2014), we used a similar pharmacogenetic approach to target dividing neural stem cells for elimination during early life periods sensitive to environmental insults. We then assessed the Nestin stem cell lineage in adulthood. We found that the adult neural stem cell reservoir was depleted following ablation during the first postnatal week, when stem cells were highly proliferative, but not during the third postnatal week, when stem cells were more quiescent. Remarkably, ablating proliferating stem cells during either the first or third postnatal week led to reduced adult neurogenesis out of proportion to the changes in the stem cell pool, indicating a disruption of the stem cell function or niche following stem cell ablation in early life. These results highlight the first three postnatal weeks as a series of sensitive periods during which elimination of dividing stem cells leads to lasting alterations in adult DG neurogenesis and stem cell function. These findings contribute to our understanding of the relationship between DG development and adult neurogenesis, as well as suggest a possible mechanism by which early life experiences may lead to lasting deficits in adult hippocampal neurogenesis. This article is protected by copyright. All rights reserved. © 2018 Wiley Periodicals, Inc.

State performance in pluripotent and adult stem cell research, 2009-2016.

PubMed

Surani, Sana H; Levine, Aaron D

2018-04-01

To examine how the geographic distribution of pluripotent and adult stem cell research publications within the USA differs from other areas of biomedical research. Publication count data for pluripotent stem cell research, adult stem cell research and a comparison group representative of biomedical research more broadly were collected and analyzed for each US state from 2009 to 2016. The distribution of pluripotent stem cell research differed from the other fields with overperformance in pluripotent stem cell research observed in California, as well as Wisconsin, Massachusetts, Maryland and Connecticut. Our analysis suggests that permissive state stem cell policy may be one of the several factors contributing to strong state performance in pluripotent stem cell research.

Stem cell clinics online: the direct-to-consumer portrayal of stem cell medicine.

PubMed

Lau, Darren; Ogbogu, Ubaka; Taylor, Benjamin; Stafinski, Tania; Menon, Devidas; Caulfield, Timothy

2008-12-04

Despite the immature state of stem cell medicine, patients are seeking and accessing putative stem cell therapies in an "early market" in which direct-to-consumer advertising via the internet likely plays an important role. We analyzed stem cell clinic websites and appraised the relevant published clinical evidence of stem cell therapies to address three questions about the direct-to-consumer portrayal of stem cell medicine in this early market: What sorts of therapies are being offered? How are they portrayed? Is there clinical evidence to support the use of these therapies? We found that the portrayal of stem cell medicine on provider websites is optimistic and unsubstantiated by peer-reviewed literature.

Effects of Telomerase and Telomere Length on Epidermal Stem Cell Behavior

NASA Astrophysics Data System (ADS)

Flores, Ignacio; Cayuela, María L.; Blasco, María A.

2005-08-01

A key process in organ homeostasis is the mobilization of stem cells out of their niches. We show through analysis of mouse models that telomere length, as well as the catalytic component of telomerase, Tert, are critical determinants in the mobilization of epidermal stem cells. Telomere shortening inhibited mobilization of stem cells out of their niche, impaired hair growth, and resulted in suppression of stem cell proliferative capacity in vitro. In contrast, Tert overexpression in the absence of changes in telomere length promoted stem cell mobilization, hair growth, and stem cell proliferation in vitro. The effects of telomeres and telomerase on stem cell biology anticipate their role in cancer and aging.

Antimicrobial efficiency of PAA/(PVP/CHI) erodible polysaccharide multilayer through loading and controlled release of antibiotics.

PubMed

Xu, Qingwen; Liu, Huihua; Ye, Zi; Nan, Kaihui; Lin, Sen; Chen, Hao; Wang, Bailiang

2017-04-01

The adhesion of bacteria and subsequent formation of biofilm on the surface of implants greatly affect the long-term use of the implants. The low molar mass gentamicin (GS) cations could hardly be directly incorporated into the multilayer films through alternately deposition with a polyanion. Herein, we have designed and constructed a (poly(acrylic acid)/(polyvinylpyrrolidone/chitosan)) n ((PAA/(PVP/CHI)) n ) multilayer films through layer-by-layer self-assembly method. Through increasing the pH to destroy hydrogen bonding between PAA and PVP, PVP released into the solution and GS simultaneously combined with PAA through electrostatic interactions. The loading dosage of GS into the (PAA/(PVP/CHI)) 10 multilayer film was up to 153.84±18.64μg/cm 2 and could be precisely tuned through changing the thickness of the films. The release behaviour of GS in phosphate buffer saline could also be regulated through thermal cross-linking of the films. The drug-loaded multilayer films displayed efficient against three kinds of Gram-positive and three kinds of Gram-negative bacteria and one kind of fungi, and good biocompatibility towards human lens epithelial cells. GS-loaded multilayer films-coated polydimethylsiloxane (PDMS) were compared with pristine PDMS in the rabbit subcutaneous S. aureus infection model. The antimicrobial-coated implants yielded a much lower degree of infections than pristine implants at day seven. Copyright © 2016 Elsevier Ltd. All rights reserved.

Microarray-Based Analysis of Methylation of 1st Trimester Trisomic Placentas from Down Syndrome, Edwards Syndrome and Patau Syndrome.

PubMed

Hatt, Lotte; Aagaard, Mads M; Bach, Cathrine; Graakjaer, Jesper; Sommer, Steffen; Agerholm, Inge E; Kølvraa, Steen; Bojesen, Anders

2016-01-01

Methylation-based non-invasive prenatal testing of fetal aneuploidies is an alternative method that could possibly improve fetal aneuploidy diagnosis, especially for trisomy 13(T13) and trisomy 18(T18). Our aim was to study the methylation landscape in placenta DNA from trisomy 13, 18 and 21 pregnancies in an attempt to find trisomy-specific methylation differences better suited for non-invasive prenatal diagnosis. We have conducted high-resolution methylation specific bead chip microarray analyses assessing more than 450,000 CpGs analyzing placentas from 12 T21 pregnancies, 12 T18 pregnancies and 6 T13 pregnancies. We have compared the methylation landscape of the trisomic placentas to the methylation landscape from normal placental DNA and to maternal blood cell DNA. Comparing trisomic placentas to normal placentas we identified 217 and 219 differentially methylated CpGs for CVS T18 and CVS T13, respectively (delta β>0.2, FDR<0.05), but only three differentially methylated CpGs for T21. However, the methylation differences was only modest (delta β<0.4), making them less suitable as diagnostic markers. Gene ontology enrichment analysis revealed that the gene set connected to theT18 differentially methylated CpGs was highly enriched for GO terms related to"DNA binding" and "transcription factor binding" coupled to the RNA polymerase II transcription. In the gene set connected to the T13 differentially methylated CpGs we found no significant enrichments.

Microarray-Based Analysis of Methylation of 1st Trimester Trisomic Placentas from Down Syndrome, Edwards Syndrome and Patau Syndrome

PubMed Central

Hatt, Lotte; Aagaard, Mads M.; Bach, Cathrine; Graakjaer, Jesper; Sommer, Steffen; Agerholm, Inge E.; Bojesen, Anders

2016-01-01

Methylation-based non-invasive prenatal testing of fetal aneuploidies is an alternative method that could possibly improve fetal aneuploidy diagnosis, especially for trisomy 13(T13) and trisomy 18(T18). Our aim was to study the methylation landscape in placenta DNA from trisomy 13, 18 and 21 pregnancies in an attempt to find trisomy–specific methylation differences better suited for non-invasive prenatal diagnosis. We have conducted high-resolution methylation specific bead chip microarray analyses assessing more than 450,000 CpGs analyzing placentas from 12 T21 pregnancies, 12 T18 pregnancies and 6 T13 pregnancies. We have compared the methylation landscape of the trisomic placentas to the methylation landscape from normal placental DNA and to maternal blood cell DNA. Comparing trisomic placentas to normal placentas we identified 217 and 219 differentially methylated CpGs for CVS T18 and CVS T13, respectively (delta β>0.2, FDR<0.05), but only three differentially methylated CpGs for T21. However, the methylation differences was only modest (delta β<0.4), making them less suitable as diagnostic markers. Gene ontology enrichment analysis revealed that the gene set connected to theT18 differentially methylated CpGs was highly enriched for GO terms related to”DNA binding” and “transcription factor binding” coupled to the RNA polymerase II transcription. In the gene set connected to the T13 differentially methylated CpGs we found no significant enrichments. PMID:27490343

Effector gene birth in plant parasitic nematodes: Neofunctionalization of a housekeeping glutathione synthetase gene

PubMed Central

Lilley, Catherine J.; Maqbool, Abbas; Wu, Duqing; Yusup, Hazijah B.; Jones, Laura M.; Birch, Paul R. J.; Urwin, Peter E.

2018-01-01

Plant pathogens and parasites are a major threat to global food security. Plant parasitism has arisen four times independently within the phylum Nematoda, resulting in at least one parasite of every major food crop in the world. Some species within the most economically important order (Tylenchida) secrete proteins termed effectors into their host during infection to re-programme host development and immunity. The precise detail of how nematodes evolve new effectors is not clear. Here we reconstruct the evolutionary history of a novel effector gene family. We show that during the evolution of plant parasitism in the Tylenchida, the housekeeping glutathione synthetase (GS) gene was extensively replicated. New GS paralogues acquired multiple dorsal gland promoter elements, altered spatial expression to the secretory dorsal gland, altered temporal expression to primarily parasitic stages, and gained a signal peptide for secretion. The gene products are delivered into the host plant cell during infection, giving rise to “GS-like effectors”. Remarkably, by solving the structure of GS-like effectors we show that during this process they have also diversified in biochemical activity, and likely represent the founding members of a novel class of GS-like enzyme. Our results demonstrate the re-purposing of an endogenous housekeeping gene to form a family of effectors with modified functions. We anticipate that our discovery will be a blueprint to understand the evolution of other plant-parasitic nematode effectors, and the foundation to uncover a novel enzymatic function. PMID:29641602

Prospects for neural stem cell-based therapies for neurological diseases.

PubMed

Imitola, Jaime

2007-10-01

Neural stem and progenitor cells have great potential for the treatment of neurological disorders. However, many obstacles remain to translate this field to the patient's bedside, including rationales for using neural stem cells in individual neurological disorders; the challenges of neural stem cell biology; and the caveats of current strategies of isolation and culturing neural precursors. Addressing these challenges is critical for the translation of neural stem cell biology to the clinic. Recent work using neural stem cells has yielded novel biologic concepts such as the importance of the reciprocal interaction between neural stem cells and the neurodegenerative environment. The prospect of using transplants of neural stem cells and progenitors to treat neurological diseases requires a better understanding of the molecular mechanisms of both neural stem cell behavior in experimental models and the intrinsic repair capacity of the injured brain.

Impact of genomic damage and ageing on stem cell function

PubMed Central

Behrens, Axel; van Deursen, Jan M.; Rudolph, K. Lenhard; Schumacher, Björn

2014-01-01

Impairment of stem cell function contributes to the progressive deterioration of tissue maintenance and repair with ageing. Evidence is mounting that age-dependent accumulation of DNA damage in both stem cells and cells that comprise the stem cell microenvironment are partly responsible for stem cell dysfunction with ageing. Here, we review the impact of the various types of DNA damage that accumulate with ageing on stem cell functionality, as well as the development of cancer. We discuss DNA-damage-induced cell intrinsic and extrinsic alterations that influence these processes, and review recent advances in understanding systemic adjustments to DNA damage and how they affect stem cells. PMID:24576896

Lgr proteins in epithelial stem cell biology.

PubMed

Barker, Nick; Tan, Shawna; Clevers, Hans

2013-06-01

The ultimate success of global efforts to exploit adult stem cells for regenerative medicine will depend heavily on the availability of robust, highly selective stem cell surface markers that facilitate the isolation of stem cells from human tissues. Any subsequent expansion or manipulation of isolated stem cells will also require an intimate knowledge of the mechanisms that regulate these cells, to ensure maintenance of their regenerative capacities and to minimize the risk of introducing undesirable growth traits that could pose health risks for patients. A subclass of leucine-rich repeat-containing G-protein-coupled receptor (Lgr) proteins has recently gained prominence as adult stem cell markers with crucial roles in maintaining stem cell functions. Here, we discuss the major impact that their discovery has had on our understanding of adult stem cell biology in various self-renewing tissues and in accelerating progress towards the development of effective stem cell therapies.

Nanotechnology in the regulation of stem cell behavior

NASA Astrophysics Data System (ADS)

Wu, King-Chuen; Tseng, Ching-Li; Wu, Chi-Chang; Kao, Feng-Chen; Tu, Yuan-Kun; So, Edmund C.; Wang, Yang-Kao

2013-10-01

Stem cells are known for their potential to repair damaged tissues. The adhesion, growth and differentiation of stem cells are likely controlled by the surrounding microenvironment which contains both chemical and physical cues. Physical cues in the microenvironment, for example, nanotopography, were shown to play important roles in stem cell fate decisions. Thus, controlling stem cell behavior by nanoscale topography has become an important issue in stem cell biology. Nanotechnology has emerged as a new exciting field and research from this field has greatly advanced. Nanotechnology allows the manipulation of sophisticated surfaces/scaffolds which can mimic the cellular environment for regulating cellular behaviors. Thus, we summarize recent studies on nanotechnology with applications to stem cell biology, including the regulation of stem cell adhesion, growth, differentiation, tracking and imaging. Understanding the interactions of nanomaterials with stem cells may provide the knowledge to apply to cell-scaffold combinations in tissue engineering and regenerative medicine.

Characterization of Amniotic Stem Cells

PubMed Central

Koike, Chika; Zhou, Kaixuan; Takeda, Yuji; Fathy, Moustafa; Okabe, Motonori; Yoshida, Toshiko; Nakamura, Yukio; Kato, Yukio

2014-01-01

Abstract The amnion membrane is developed from embryo-derived cells, and amniotic cells have been shown to exhibit multidifferentiation potential. These cells represent a desirable source for stem cells for a variety of reasons. However, to date very few molecular analyses of amnion-derived cells have been reported, and efficient markers for isolating the stem cells remain unclear. This paper assesses the characterization of amnion-derived cells as stem cells by examining stemness marker expressions for amnion-derived epithelial cells and mesenchymal cells by flow cytometry, immunocytochemistry, and quantitative PCR. Flow cytometry revealed that amnion epithelial cells expressed CD133, CD 271, and TRA-1-60, whereas mecenchymal cells expressed CD44, CD73, CD90, and CD105. Immunohistochemistry showed that both cells expressed the stemness markers Oct3/4, Sox2, Klf4, and SSEA4. Stemness genes' expression in amnion epithelial cells, mesenchymal cells, fibroblast, bone marrow–derived mesenchymal stem cells (MSCs), and induced pluripotent stem cells (iPSCs) was compared by quantitative reverse-transcription polymerase chain reaction (RT-PCR). Amnion-derived epithelial cells and mesenchymal cells expressed Oct3/4, Nanog, and Klf4 more than bone marrow–derived MSCs. The sorted TRA1-60–positive cells expressed Oct3/4, Nanog, and Klf4 more than unsorted cells or TRA1-60–negative cells. TRA1-60 can be a marker for isolating amnion epithelial stem cells. PMID:25068631

Current applications of human pluripotent stem cells: possibilities and challenges.

PubMed

Ho, Pai-Jiun; Yen, Men-Luh; Yet, Shaw-Fang; Yen, B Linju

2012-01-01

Stem cells are self-renewable cells with the differentiation capacity to develop into somatic cells with biological functions. This ability to sustain a renewable source of multi- and/or pluripotential differentiation has brought new hope to the field of regenerative medicine in terms of cell therapy and tissue engineering. Moreover, stem cells are invaluable tools as in vitro models for studying diverse fields, from basic scientific questions such as developmental processes and lineage commitment, to practical application including drug screening and testing. The stem cells with widest differentiation potential are pluripotent stem cells (PSCs), which are rare cells with the ability to generate somatic cells from all three germ layers. PSCs are considered the most optimal choice for therapeutic potential of stem cells, bringing new impetus to the field of regenerative medicine. In this article, we discuss the therapeutic potential of human PSCs (hPSCs) including human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs), reviewing the current preclinical and clinical data using these stem cells. We describe the classification of different sources of hPSCs, ongoing research, and currently encountered clinical obstacles of these novel and versatile human stem cells.

Monolayered mesenchymal stem cells repair scarred myocardium after myocardial infarction.

PubMed

Miyahara, Yoshinori; Nagaya, Noritoshi; Kataoka, Masaharu; Yanagawa, Bobby; Tanaka, Koichi; Hao, Hiroyuki; Ishino, Kozo; Ishida, Hideyuki; Shimizu, Tatsuya; Kangawa, Kenji; Sano, Shunji; Okano, Teruo; Kitamura, Soichiro; Mori, Hidezo

2006-04-01

Mesenchymal stem cells are multipotent cells that can differentiate into cardiomyocytes and vascular endothelial cells. Here we show, using cell sheet technology, that monolayered mesenchymal stem cells have multipotent and self-propagating properties after transplantation into infarcted rat hearts. We cultured adipose tissue-derived mesenchymal stem cells characterized by flow cytometry using temperature-responsive culture dishes. Four weeks after coronary ligation, we transplanted the monolayered mesenchymal stem cells onto the scarred myocardium. After transplantation, the engrafted sheet gradually grew to form a thick stratum that included newly formed vessels, undifferentiated cells and few cardiomyocytes. The mesenchymal stem cell sheet also acted through paracrine pathways to trigger angiogenesis. Unlike a fibroblast cell sheet, the monolayered mesenchymal stem cells reversed wall thinning in the scar area and improved cardiac function in rats with myocardial infarction. Thus, transplantation of monolayered mesenchymal stem cells may be a new therapeutic strategy for cardiac tissue regeneration.

Genetic and epigenetic instability of stem cells.

PubMed

Rajamani, Karthyayani; Li, Yuan-Sheng; Hsieh, Dean-Kuo; Lin, Shinn-Zong; Harn, Horng-Jyh; Chiou, Tzyy-Wen

2014-01-01

Recently, research on stem cells has been receiving an increasing amount of attention, both for its advantages and disadvantages. Genetic and epigenetic instabilities among stem cells have been a recurring obstacle to progress in regenerative medicine using stem cells. Various reports have stated that these instabilities can transform stem cells when transferred in vivo and thus have the potential to develop tumors. Previous research has shown that various extrinsic and intrinsic factors can contribute to the stability of stem cells. The extrinsic factors include growth supplements, growth factors, oxygen tension, passage technique, and cryopreservation. Controlling these factors based on previous reports may assist researchers in developing strategies for the production and clinical application of "safe" stem cells. On the other hand, the intrinsic factors can be unpredictable and uncontrollable; therefore, to ensure the successful use of stem cells in regenerative medicine, it is imperative to develop and implement appropriate strategies and technique for culturing stem cells and to confirm the genetic and epigenetic safety of these stem cells before employing them in clinical trials.

Dipeptide species regulate p38MAPK–Smad3 signalling to maintain chronic myelogenous leukaemia stem cells

PubMed Central

Naka, Kazuhito; Jomen, Yoshie; Ishihara, Kaori; Kim, Junil; Ishimoto, Takahiro; Bae, Eun-Jin; Mohney, Robert P.; Stirdivant, Steven M.; Oshima, Hiroko; Oshima, Masanobu; Kim, Dong-Wook; Nakauchi, Hiromitsu; Takihara, Yoshihiro; Kato, Yukio; Ooshima, Akira; Kim, Seong-Jin

2015-01-01

Understanding the specific survival of the rare chronic myelogenous leukaemia (CML) stem cell population could provide a target for therapeutics aimed at eradicating these cells. However, little is known about how survival signalling is regulated in CML stem cells. In this study, we survey global metabolic differences between murine normal haematopoietic stem cells (HSCs) and CML stem cells using metabolomics techniques. Strikingly, we show that CML stem cells accumulate significantly higher levels of certain dipeptide species than normal HSCs. Once internalized, these dipeptide species activate amino-acid signalling via a pathway involving p38MAPK and the stemness transcription factor Smad3, which promotes CML stem cell maintenance. Importantly, pharmacological inhibition of dipeptide uptake inhibits CML stem cell activity in vivo. Our results demonstrate that dipeptide species support CML stem cell maintenance by activating p38MAPK–Smad3 signalling in vivo, and thus point towards a potential therapeutic target for CML treatment. PMID:26289811

Engineering Stem Cells for Biomedical Applications.

PubMed

Yin, Perry T; Han, Edward; Lee, Ki-Bum

2016-01-07

Stem cells are characterized by a number of useful properties, including their ability to migrate, differentiate, and secrete a variety of therapeutic molecules such as immunomodulatory factors. As such, numerous pre-clinical and clinical studies have utilized stem cell-based therapies and demonstrated their tremendous potential for the treatment of various human diseases and disorders. Recently, efforts have focused on engineering stem cells in order to further enhance their innate abilities as well as to confer them with new functionalities, which can then be used in various biomedical applications. These engineered stem cells can take on a number of forms. For instance, engineered stem cells encompass the genetic modification of stem cells as well as the use of stem cells for gene delivery, nanoparticle loading and delivery, and even small molecule drug delivery. The present Review gives an in-depth account of the current status of engineered stem cells, including potential cell sources, the most common methods used to engineer stem cells, and the utilization of engineered stem cells in various biomedical applications, with a particular focus on tissue regeneration, the treatment of immunodeficiency diseases, and cancer. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

The dopamine D1 receptor is expressed and facilitates relaxation in airway smooth muscle.

PubMed

Mizuta, Kentaro; Zhang, Yi; Xu, Dingbang; Mizuta, Fumiko; D'Ovidio, Frank; Masaki, Eiji; Emala, Charles W

2013-09-02

Dopamine signaling is mediated by Gs protein-coupled "D1-like" receptors (D1 and D5) and Gi-coupled "D2-like" receptors (D2-4). In asthmatic patients, inhaled dopamine induces bronchodilation. Although the Gi-coupled dopamine D2 receptor is expressed and sensitizes adenylyl cyclase activity in airway smooth muscle (ASM) cells, the Gs-coupled dopamine D1-like receptor subtypes have never been identified on these cells. Activation of Gs-coupled receptors stimulates cyclic AMP (cAMP) production through the stimulation of adenylyl cyclase, which promotes ASM relaxation. We questioned whether the dopamine D1-like receptor is expressed on ASM, and modulates its function through Gs-coupling. The mRNA and protein expression of dopamine D1-like receptor subtypes in both native human and guinea pig ASM tissue and cultured human ASM (HASM) cells was measured. To characterize the stimulation of cAMP through the dopamine D1 receptor, HASM cells were treated with dopamine or the dopamine D1-like receptor agonists (A68930 or SKF38393) before cAMP measurements. To evaluate whether the activation of dopamine D1 receptor induces ASM relaxation, guinea pig tracheal rings suspended under isometric tension in organ baths were treated with cumulatively increasing concentrations of dopamine or A68930, following an acetylcholine-induced contraction with or without the cAMP-dependent protein kinase (PKA) inhibitor Rp-cAMPS, the large-conductance calcium-activated potassium (BKCa) channel blocker iberiotoxin, or the exchange proteins directly activated by cAMP (Epac) antagonist NSC45576. Messenger RNA encoding the dopamine D1 and D5 receptors were detected in native human ASM tissue and cultured HASM cells. Immunoblots confirmed the protein expression of the dopamine D1 receptor in both native human and guinea pig ASM tissue and cultured HASM cells. The dopamine D1 receptor was also immunohistochemically localized to both human and guinea pig ASM. The dopamine D1-like receptor agonists stimulated cAMP production in HASM cells, which was reversed by the selective dopamine D1-like receptor antagonists SCH23390 or SCH39166. A68930 relaxed acetylcholine-contracted guinea pig tracheal rings, which was attenuated by Rp-cAMPS but not by iberiotoxin or NSC45576. These results demonstrate that the dopamine D1 receptors are expressed on ASM and regulate smooth muscle force via cAMP activation of PKA, and offer a novel target for therapeutic relaxation of ASM.

The dopamine D1 receptor is expressed and facilitates relaxation in airway smooth muscle

PubMed Central

2013-01-01

Background Dopamine signaling is mediated by Gs protein-coupled “D1-like” receptors (D1 and D5) and Gi-coupled “D2-like” receptors (D2-4). In asthmatic patients, inhaled dopamine induces bronchodilation. Although the Gi-coupled dopamine D2 receptor is expressed and sensitizes adenylyl cyclase activity in airway smooth muscle (ASM) cells, the Gs-coupled dopamine D1-like receptor subtypes have never been identified on these cells. Activation of Gs-coupled receptors stimulates cyclic AMP (cAMP) production through the stimulation of adenylyl cyclase, which promotes ASM relaxation. We questioned whether the dopamine D1-like receptor is expressed on ASM, and modulates its function through Gs-coupling. Methods The mRNA and protein expression of dopamine D1-like receptor subtypes in both native human and guinea pig ASM tissue and cultured human ASM (HASM) cells was measured. To characterize the stimulation of cAMP through the dopamine D1 receptor, HASM cells were treated with dopamine or the dopamine D1-like receptor agonists (A68930 or SKF38393) before cAMP measurements. To evaluate whether the activation of dopamine D1 receptor induces ASM relaxation, guinea pig tracheal rings suspended under isometric tension in organ baths were treated with cumulatively increasing concentrations of dopamine or A68930, following an acetylcholine-induced contraction with or without the cAMP-dependent protein kinase (PKA) inhibitor Rp-cAMPS, the large-conductance calcium-activated potassium (BKCa) channel blocker iberiotoxin, or the exchange proteins directly activated by cAMP (Epac) antagonist NSC45576. Results Messenger RNA encoding the dopamine D1 and D5 receptors were detected in native human ASM tissue and cultured HASM cells. Immunoblots confirmed the protein expression of the dopamine D1 receptor in both native human and guinea pig ASM tissue and cultured HASM cells. The dopamine D1 receptor was also immunohistochemically localized to both human and guinea pig ASM. The dopamine D1-like receptor agonists stimulated cAMP production in HASM cells, which was reversed by the selective dopamine D1-like receptor antagonists SCH23390 or SCH39166. A68930 relaxed acetylcholine-contracted guinea pig tracheal rings, which was attenuated by Rp-cAMPS but not by iberiotoxin or NSC45576. Conclusions These results demonstrate that the dopamine D1 receptors are expressed on ASM and regulate smooth muscle force via cAMP activation of PKA, and offer a novel target for therapeutic relaxation of ASM. PMID:24004608

Modeling TSC and LAM Using Patient Derived Induced Pluripotent Stem Cells

DTIC Science & Technology

2016-10-01

lentiviral knockdown, and CRISPR /Cas9 genome editing in embryonic stem cells (ESCs). We have characterized the iPSCs extensively and found that they display...induced pluripotent stem cells (iPSCs) embryonic stem cells (ESCs) reprogramming CRISPR /Cas9 genome editing neural stem cells (NSCs) neural crest... CRISPR /cas9 in two additional human pluripotent stem cell lines (WA07 (H7) – female cell line registry #0061; and a control male iPSC lines generated

Biochemistry of epidermal stem cells☆

PubMed Central

Eckert, Richard L.; Adhikary, Gautam; Balasubramanian, Sivaprakasam; Rorke, Ellen A.; Vemuri, Mohan C.; Boucher, Shayne E.; Bickenbach, Jackie R.; Kerr, Candace

2014-01-01

Background The epidermis is an important protective barrier that is essential for maintenance of life. Maintaining this barrier requires continuous cell proliferation and differentiation. Moreover, these processes must be balanced to produce a normal epidermis. The stem cells of the epidermis reside in specific locations in the basal epidermis, hair follicle and sebaceous glands and these cells are responsible for replenishment of this tissue. Scope of review A great deal of effort has gone into identifying protein epitopes that mark stem cells, in identifying stem cell niche locations, and in understanding how stem cell populations are related. We discuss these studies as they apply to understanding normal epidermal homeostasis and skin cancer. Major conclusions An assortment of stem cell markers have been identified that permit assignment of stem cells to specific regions of the epidermis, and progress has been made in understanding the role of these cells in normal epidermal homeostasis and in conditions of tissue stress. A key finding is the multiple stem cell populations exist in epidermis that give rise to different structures, and that multiple stem cell types may contribute to repair in damaged epidermis. General significance Understanding epidermal stem cell biology is likely to lead to important therapies for treating skin diseases and cancer, and will also contribute to our understanding of stem cells in other systems. This article is part of a Special Issue entitled Biochemistry of Stem Cells. PMID:22820019

Cryopreservation of Human Stem Cells for Clinical Application: A Review

PubMed Central

Hunt, Charles J.

2011-01-01

Summary Stem cells have been used in a clinical setting for many years. Haematopoietic stem cells have been used for the treatment of both haematological and non-haematological disease; while more recently mesenchymal stem cells derived from bone marrow have been the subject of both laboratory and early clinical studies. Whilst these cells show both multipotency and expansion potential, they nonetheless do not form stable cell lines in culture which is likely to limit the breadth of their application in the field of regenerative medicine. Human embryonic stem cells are pluripotent cells, capable of forming stable cell lines which retain the capacity to differentiate into cells from all three germ layers. This makes them of special significance in both regenerative medicine and toxicology. Induced pluripotent stem (iPS) cells may also provide a similar breadth of utility without some of the confounding ethical issues surrounding embryonic stem cells. An essential pre-requisite to the commercial and clinical application of stem cells are suitable cryopreservation protocols for long-term storage. Whilst effective methods for cryopreservation and storage have been developed for haematopoietic and mesenchymal stem cells, embryonic cells and iPS cells have proved more refractory. This paper reviews the current state of cryopreservation as it pertains to stem cells and in particular the embryonic and iPS cell. PMID:21566712

Cryopreservation of Human Stem Cells for Clinical Application: A Review.

PubMed

Hunt, Charles J

2011-01-01

SUMMARY: Stem cells have been used in a clinical setting for many years. Haematopoietic stem cells have been used for the treatment of both haematological and non-haematological disease; while more recently mesenchymal stem cells derived from bone marrow have been the subject of both laboratory and early clinical studies. Whilst these cells show both multipotency and expansion potential, they nonetheless do not form stable cell lines in culture which is likely to limit the breadth of their application in the field of regenerative medicine. Human embryonic stem cells are pluripotent cells, capable of forming stable cell lines which retain the capacity to differentiate into cells from all three germ layers. This makes them of special significance in both regenerative medicine and toxicology. Induced pluripotent stem (iPS) cells may also provide a similar breadth of utility without some of the confounding ethical issues surrounding embryonic stem cells. An essential pre-requisite to the commercial and clinical application of stem cells are suitable cryopreservation protocols for long-term storage. Whilst effective methods for cryopreservation and storage have been developed for haematopoietic and mesenchymal stem cells, embryonic cells and iPS cells have proved more refractory. This paper reviews the current state of cryopreservation as it pertains to stem cells and in particular the embryonic and iPS cell.

YAP/TAZ enhance mammalian embryonic neural stem cell characteristics in a Tead-dependent manner

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

Han, Dasol; Byun, Sung-Hyun; Park, Soojeong

Mammalian brain development is regulated by multiple signaling pathways controlling cell proliferation, migration and differentiation. Here we show that YAP/TAZ enhance embryonic neural stem cell characteristics in a cell autonomous fashion using diverse experimental approaches. Introduction of retroviral vectors expressing YAP or TAZ into the mouse embryonic brain induced cell localization in the ventricular zone (VZ), which is the embryonic neural stem cell niche. This change in cell distribution in the cortical layer is due to the increased stemness of infected cells; YAP-expressing cells were colabeled with Sox2, a neural stem cell marker, and YAP/TAZ increased the frequency and sizemore » of neurospheres, indicating enhanced self-renewal- and proliferative ability of neural stem cells. These effects appear to be TEA domain family transcription factor (Tead)–dependent; a Tead binding-defective YAP mutant lost the ability to promote neural stem cell characteristics. Consistently, in utero gene transfer of a constitutively active form of Tead2 (Tead2-VP16) recapitulated all the features of YAP/TAZ overexpression, and dominant negative Tead2-EnR resulted in marked cell exit from the VZ toward outer cortical layers. Taken together, these results indicate that the Tead-dependent YAP/TAZ signaling pathway plays important roles in neural stem cell maintenance by enhancing stemness of neural stem cells during mammalian brain development. - Highlights: • Roles of YAP and Tead in vivo during mammalian brain development are clarified. • Expression of YAP promotes embryonic neural stem cell characteristics in vivo in a cell autonomous fashion. • Enhancement of neural stem cell characteristics by YAP depends on Tead. • Transcriptionally active form of Tead alone can recapitulate the effects of YAP. • Transcriptionally repressive form of Tead severely reduces stem cell characteristics.« less

Recent progress in stem cell differentiation directed by material and mechanical cues.

PubMed

Lin, Xunxun; Shi, Yuan; Cao, Yilin; Liu, Wei

2016-02-02

Stem cells play essential roles in tissue regeneration in vivo via specific lineage differentiation induced by environmental factors. In the past, biochemical signals were the focus of induced stem cell differentiation. As reported by Engler et al (2006 Cell 126 677-89), biophysical signal mediated stem cell differentiation could also serve as an important inducer. With the advancement of material science, it becomes a possible strategy to generate active biophysical signals for directing stem cell fate through specially designed material microstructures. In the past five years, significant progress has been made in this field, and these designed biophysical signals include material elasticity/rigidity, micropatterned structure, extracellular matrix (ECM) coated materials, material transmitted extracellular mechanical force etc. A large number of investigations involved material directed differentiation of mesenchymal stem cells, neural stem/progenitor cells, adipose derived stem cells, hematopoietic stem/progenitor cells, embryonic stem cells and other cells. Hydrogel based materials were commonly used to create varied mechanical properties via modifying the ratio of different components, crosslinking levels, matrix concentration and conjugation with other components. Among them, polyacrylamide (PAM) and polydimethylsiloxane (PDMS) hydrogels remained the major types of material. Specially designed micropatterning was not only able to create a unique topographical surface to control cell shape, alignment, cell-cell and cell-matrix contact for basic stem cell biology study, but also could be integrated with 3D bioprinting to generate micropattered 3D structure and thus to induce stem cell based tissue regeneration. ECM coating on a specific topographical structure was capable of inducing even more specific and potent stem cell differentiation along with soluble factors and mechanical force. The article overviews the progress of the past five years in this particular field.

Engineering Hydrogel Microenvironments to Recapitulate the Stem Cell Niche.

PubMed

Madl, Christopher M; Heilshorn, Sarah C

2018-06-04

Stem cells are a powerful resource for many applications including regenerative medicine, patient-specific disease modeling, and toxicology screening. However, eliciting the desired behavior from stem cells, such as expansion in a naïve state or differentiation into a particular mature lineage, remains challenging. Drawing inspiration from the native stem cell niche, hydrogel platforms have been developed to regulate stem cell fate by controlling microenvironmental parameters including matrix mechanics, degradability, cell-adhesive ligand presentation, local microstructure, and cell-cell interactions. We survey techniques for modulating hydrogel properties and review the effects of microenvironmental parameters on maintaining stemness and controlling differentiation for a variety of stem cell types. Looking forward, we envision future hydrogel designs spanning a spectrum of complexity, ranging from simple, fully defined materials for industrial expansion of stem cells to complex, biomimetic systems for organotypic cell culture models.

Incorporation of Biomaterials in Multicellular Aggregates Modulates Pluripotent Stem Cell Differentiation

PubMed Central

Bratt-Leal, Andrés M.; Carpenedo, Richard L.; Ungrin, Mark; Zandstra, Peter W.; McDevitt, Todd C.

2010-01-01

Biomaterials are increasingly being used to engineer the biochemical and biophysical properties of the extracellular stem cell microenvironment in order to tailor niche characteristics and direct cell phenotype. To date, stem cell-biomaterial interactions have largely been studied by introducing stem cells into artificial environments, such as 2D cell culture on biomaterial surfaces, encapsulation of cell suspensions within hydrogel materials, or cell seeding on 3D polymeric scaffolds. In this study, microparticles fabricated from different materials, such as agarose, PLGA and gelatin, were stably integrated, in a dose-dependent manner, within aggregates of pluripotent stem cells (PSCs) prior to differentiation as a means to directly examine stem cell-biomaterial interactions in 3D. Interestingly, the presence of the materials within the stem cell aggregates differentially modulated the gene and protein expression patterns of several differentiation markers without adversely affecting cell viability. Microparticle incorporation within 3D stem cell aggregates can control the spatial presentation of extracellular environmental cues (i.e. soluble factors, extracellular matrix and intercellular adhesion molecules) as a means to direct the differentiation of stem cells for tissue engineering and regenerative medicine applications. In addition, these results suggest that the physical presence of microparticles within stem cell aggregates does not compromise PSC differentiation, but in fact the choice of biomaterials can impact the propensity of stem cells to adopt particular differentiated cell phenotypes. PMID:20864164

Markers for the identification of tendon-derived stem cells in vitro and tendon stem cells in situ - update and future development.

PubMed

Lui, Pauline Po Yee

2015-06-02

The efficacy of tendon-derived stem cells (TDSCs) for the promotion of tendon and tendon-bone junction repair has been reported in animal studies. Modulation of the tendon stem cell niche in vivo has also been reported to influence tendon structure. There is a need to have specific and reliable markers that can define TDSCs in vitro and tendon stem cells in situ for several reasons: to understand the basic biology of TDSCs and their subpopulations in vitro; to understand the identity, niches and functions of tendon/progenitor stem cells in vivo; to meet the governmental regulatory requirements for quality of TDSCs when translating the exciting preclinical findings into clinical trial/practice; and to develop new treatment strategies for mobilizing endogenous stem/progenitor cells in tendon. TDSCs were reported to express the common mesenchymal stem cell (MSC) markers and some embryonic stem cell (ESC) markers, and there were attempts to use these markers to label tendon stem cells in situ. Are these stem cell markers useful for the identification of TDSCs in vitro and tracking of tendon stem cells in situ? This review aims to discuss the values of the panel of MSC, ESC and tendon-related markers for the identification of TDSCs in vitro. Important factors influencing marker expression by TDSCs are discussed. The usefulness and limitations of the panel of MSC, ESC and tendon-related markers for tracking stem cells in tendon, especially tendon stem cells, in situ are then reviewed. Future research directions are proposed.

Haematopoietic stem and progenitor cells from human pluripotent stem cells

PubMed Central

Sugimura, Ryohichi; Jha, Deepak Kumar; Han, Areum; Soria-Valles, Clara; da Rocha, Edroaldo Lummertz; Lu, Yi-Fen; Goettel, Jeremy A.; Serrao, Erik; Rowe, R. Grant; Malleshaiah, Mohan; Wong, Irene; Sousa, Patricia; Zhu, Ted N.; Ditadi, Andrea; Keller, Gordon; Engelman, Alan N.; Snapper, Scott B.; Doulatov, Sergei; Daley, George Q.

2018-01-01

A variety of tissue lineages can be differentiated from pluripotent stem cells by mimicking embryonic development through stepwise exposure to morphogens, or by conversion of one differentiated cell type into another by enforced expression of master transcription factors. Here, to yield functional human haematopoietic stem cells, we perform morphogen-directed differentiation of human pluripotent stem cells into haemogenic endothelium followed by screening of 26 candidate haematopoietic stem-cell-specifying transcription factors for their capacity to promote multi-lineage haematopoietic engraftment in mouse hosts. We recover seven transcription factors (ERG, HOXA5, HOXA9, HOXA10, LCOR, RUNX1 and SPI1) that are sufficient to convert haemogenic endothelium into haematopoietic stem and progenitor cells that engraft myeloid, B and T cells in primary and secondary mouse recipients. Our combined approach of morphogen-driven differentiation and transcription-factor-mediated cell fate conversion produces haematopoietic stem and progenitor cells from pluripotent stem cells and holds promise for modelling haematopoietic disease in humanized mice and for therapeutic strategies in genetic blood disorders. PMID:28514439

Stem Cells in the Trabecular Meshwork for Regulating Intraocular Pressure.

PubMed

Yun, Hongmin; Zhou, Yi; Wills, Andrew; Du, Yiqin

2016-06-01

Intraocular pressure (IOP) is still the main treatment target for glaucoma. Outflow resistance mainly exists at the trabecular meshwork (TM) outflow pathway, which is responsible for IOP regulation. Changes of TM cellularity and TM extracellular matrix turnover may play important roles in IOP regulation. In this article, we review basic anatomy and physiology of the outflow pathway and TM stem cell characteristics regarding the location, isolation, identification and function. TM stem cells are localized at the insert region of the TM and are label-retaining in vivo. They can be isolated by side-population cell sorting, cloning culture, or sphere culture. TM stem cells are multipotent with the ability to home to the TM region and differentiate into TM cells in vivo. Other stem cell types, such as adipose-derived stem cells, mesenchymal stem cells and induced pluripotent stem cells have been discovered for TM cell differentiation and TM regeneration. We also review glaucomatous animal models, which are suitable to study stem cell-based therapies for TM regeneration.

Stem Cells in the Trabecular Meshwork for Regulating Intraocular Pressure

PubMed Central

Yun, Hongmin; Zhou, Yi; Wills, Andrew

2016-01-01

Abstract Intraocular pressure (IOP) is still the main treatment target for glaucoma. Outflow resistance mainly exists at the trabecular meshwork (TM) outflow pathway, which is responsible for IOP regulation. Changes of TM cellularity and TM extracellular matrix turnover may play important roles in IOP regulation. In this article, we review basic anatomy and physiology of the outflow pathway and TM stem cell characteristics regarding the location, isolation, identification and function. TM stem cells are localized at the insert region of the TM and are label-retaining in vivo. They can be isolated by side-population cell sorting, cloning culture, or sphere culture. TM stem cells are multipotent with the ability to home to the TM region and differentiate into TM cells in vivo. Other stem cell types, such as adipose-derived stem cells, mesenchymal stem cells and induced pluripotent stem cells have been discovered for TM cell differentiation and TM regeneration. We also review glaucomatous animal models, which are suitable to study stem cell-based therapies for TM regeneration. PMID:27183473

New perspectives in human stem cell therapeutic research.

PubMed

Trounson, Alan

2009-06-11

Human stem cells are in evaluation in clinical stem cell trials, primarily as autologous bone marrow studies, autologous and allogenic mesenchymal stem cell trials, and some allogenic neural stem cell transplantation projects. Safety and efficacy are being addressed for a number of disease state applications. There is considerable data supporting safety of bone marrow and mesenchymal stem cell transplants but the efficacy data are variable and of mixed benefit. Mechanisms of action of many of these cells are unknown and this raises the concern of unpredictable results in the future. Nevertheless there is considerable optimism that immune suppression and anti-inflammatory properties of mesenchymal stem cells will be of benefit for many conditions such as graft versus host disease, solid organ transplants and pulmonary fibrosis. Where bone marrow and mesenchymal stem cells are being studied for heart disease, stroke and other neurodegenerative disorders, again progress is mixed and mostly without significant benefit. However, correction of multiple sclerosis, at least in the short term is encouraging. Clinical trials on the use of embryonic stem cell derivatives for spinal injury and macular degeneration are beginning and a raft of other clinical trials can be expected soon, for example, the use of neural stem cells for killing inoperable glioma and embryonic stem cells for regenerating beta islet cells for diabetes. The change in attitude to embryonic stem cell research with the incoming Obama administration heralds a new co-operative environment for study and evaluation of stem cell therapies. The Californian stem cell initiative (California Institute for Regenerative Medicine) has engendered global collaboration for this new medicine that will now also be supported by the US Federal Government. The active participation of governments, academia, biotechnology, pharmaceutical companies, and private investment is a powerful consortium for advances in health.

Invincible, but not invisible: imaging approaches toward in vivo detection of cancer stem cells.

PubMed

Hart, Lori S; El-Deiry, Wafik S

2008-06-10

With evidence emerging in support of a cancer stem-cell model of carcinogenesis, it is of paramount importance to identify and image these elusive cells in their natural environment. The cancer stem-cell hypothesis has the potential to explain unresolved questions of tumorigenesis, tumor heterogeneity, chemotherapeutic and radiation resistance, and even the metastatic phenotype. Intravital imaging of cancer stem cells could be of great value for determining prognosis, as well as monitoring therapeutic efficacy and influencing therapeutic protocols. Cancer stem cells represent a rare population of cells, as low as 0.1% of cells within a human tumor, and the phenotype of isolated cancer stem cells is easily altered when placed under in vitro conditions. This represents a challenge in studying cancer stem cells without manipulation or extraction from their natural environment. Advanced imaging techniques allow for the in vivo observation of physiological events at cellular resolution. Cancer stem-cell studies must take advantage of such technology to promote a better understanding of the cancer stem-cell model in relation to tumor growth and metastasis, as well as to potentially improve on the principles by which cancers are treated. This review examines the opportunities for in vivo imaging of putative cancer stem cells with regard to currently accepted cancer stem-cell characteristics and advanced imaging technologies.

Neural stem cell-based treatment for neurodegenerative diseases.

PubMed

Kim, Seung U; Lee, Hong J; Kim, Yun B

2013-10-01

Human neurodegenerative diseases such as Parkinson's disease (PD), Huntington's disease (HD), amyotrophic lateral sclerosis (ALS) and Alzheimer's disease (AD) are caused by a loss of neurons and glia in the brain or spinal cord. Neurons and glial cells have successfully been generated from stem cells such as embryonic stem cells (ESCs), mesenchymal stem cells (MSCs) and neural stem cells (NSCs), and stem cell-based cell therapies for neurodegenerative diseases have been developed. A recent advance in generation of a new class of pluripotent stem cells, induced pluripotent stem cells (iPSCs), derived from patients' own skin fibroblasts, opens doors for a totally new field of personalized medicine. Transplantation of NSCs, neurons or glia generated from stem cells in animal models of neurodegenerative diseases, including PD, HD, ALS and AD, demonstrates clinical improvement and also life extension of these animals. Additional therapeutic benefits in these animals can be provided by stem cell-mediated gene transfer of therapeutic genes such as neurotrophic factors and enzymes. Although further research is still needed, cell and gene therapy based on stem cells, particularly using neurons and glia derived from iPSCs, ESCs or NSCs, will become a routine treatment for patients suffering from neurodegenerative diseases and also stroke and spinal cord injury. © 2013 Japanese Society of Neuropathology.

Constitutive Proteasomal Degradation of TWIST-1 in Epithelial Ovarian Cancer Stem Cells Impacts Differentiation and Metastatic Potential

PubMed Central

Yin, Gang; Alvero, Ayesha B.; Craveiro, Vinicius; Holmberg, Jennie C.; Fu, Han-Hsuan; Montagna, Michele K.; Yang, Yang; Chefetz-Menaker, Ilana; Nuti, Sudhakar; Rossi, Michael; Silasi, Dan-Arin; Rutherford, Thomas; Mor, Gil

2013-01-01

Epithelial-mesenchymal transition (EMT) is a critical process for embryogenesis but is abnormally activated during cancer metastasis and recurrence. This process enables epithelial cancer cells to acquire mobility and traits associated with stemness. It is unknown whether epithelial stem cells or epithelial cancer stem cells are able to undergo EMT, and what molecular mechanism regulates this process in these specific cell types. We found that Epithelial Ovarian Cancer Stem cells (EOC stem cells) are the source of metastatic progenitor cells through a differentiation process involving EMT and Mesenchymal-Epithelial Transition (MET). We demonstrate both in vivo and in vitro the differentiation of EOC stem cells into mesenchymal spheroid-forming cells (MSFCs) and their capacity to initiate an active carcinomatosis. Furthermore, we demonstrate that human EOC stem cells injected i.p in mice are able to form ovarian tumors, suggesting that the EOC stem cells have the ability to “home” to the ovaries and establish tumors. Most interestingly, we found that TWIST1 is constitutively degraded in EOC stem cells, and that the acquisition of TWIST1 requires additional signals that will trigger the differentiation process. These findings are relevant for understanding the differentiation and metastasis process in EOC stem cells. PMID:22349827

Hepatic differentiation of pluripotent stem cells.

PubMed

Loya, Komal; Eggenschwiler, Reto; Ko, Kinarm; Sgodda, Malte; André, Francoise; Bleidissel, Martina; Schöler, Hans R; Cantz, Tobias

2009-10-01

In regenerative medicine pluripotent stem cells are considered to be a valuable self-renewing source for therapeutic cell transplantations, given that a functional organ-specific phenotype can be acquired by in vitro differentiation protocols. Furthermore, derivatives of pluripotent stem cells that mimic fetal progenitor stages could serve as an important tool to analyze organ development with in vitro approaches. Because of ethical issues regarding the generation of human embryonic stem (ES) cells, other sources for pluripotent stem cells are intensively studied. Like in less developed vertebrates, pluripotent stem cells can be generated from the female germline even in mammals, via parthenogenetic activation of oocytes. Recently, testis-derived pluripotent stem cells were derived from the male germline. Therefore, we compared two different hepatic differentiation approaches and analyzed the generation of definitive endoderm progenitor cells and their further maturation into a hepatic phenotype using murine parthenogenetic ES cells, germline-derived pluripotent stem cells, and ES cells. Applying quantitative RT-PCR, both germline-derived pluripotent cell lines show similar differentiation capabilities as normal murine ES cells and can be considered an alternative source for pluripotent stem cells in regenerative medicine.

An overview on ethical considerations in stem cell research in Iran and ethical recommendations: A review.

PubMed

Farajkhoda, Tahmineh

2017-02-01

Conducting research on the stem cell lines might bring some worthy good to public. Human Stem Cells (hSCs) research has provided opportunities for scientific progresses and new therapies, but some complex ethical matters should be noticed to ensure that stem cell research is carried out in an ethically appropriate manner. The aim of this review article is to discuss the importance of stem cell research, code of ethics for stem cell research in Iran and ethical recommendation. Generation of stem cells for research from human embryo or adult stem cells, saving, maintenance and using of them are the main ethical, legal and jurisprudence concerns in Iran. Concerns regarding human reproduction or human cloning, breach of human dignity, genetic manipulation and probability of tumorogenisity are observed in adult/somatic stem cells. Destruction of embryo to generate stem cell is an important matter in Iran. In this regards, obtaining stem cell from donated frozen embryos through infertility treatment that would be discarded is an acceptable solution in Iran for generation of embryo for research. Ethical, legal, and jurisprudence strategies for using adult/somatic stem cells are determination of ownership of stem cells, trade prohibition of human body, supervision on bio banks and information of Oversight Committee on Stem Cell Research. Recommendations to handle ethical issues for conducting stem cell research are well-designed studies, compliance codes of ethics in biomedical research (specifically codes of ethics on stem cell research, codes of ethics on clinical trials studies and codes of ethics on animals studies), appropriate collaboration with ethics committees and respecting of rights of participants (including both of human and animal rights) in research. In addition, there is a necessity for extending global networks of bioethics for strengthening communications within organizations at both the regional and international level, strengthening legislation systems, designing and establishing convenient collaborative educational courses at different levels.

An overview on ethical considerations in stem cell research in Iran and ethical recommendations: A review

PubMed Central

Farajkhoda, Tahmineh

2017-01-01

Conducting research on the stem cell lines might bring some worthy good to public. Human Stem Cells (hSCs) research has provided opportunities for scientific progresses and new therapies, but some complex ethical matters should be noticed to ensure that stem cell research is carried out in an ethically appropriate manner. The aim of this review article is to discuss the importance of stem cell research, code of ethics for stem cell research in Iran and ethical recommendation. Generation of stem cells for research from human embryo or adult stem cells, saving, maintenance and using of them are the main ethical, legal and jurisprudence concerns in Iran. Concerns regarding human reproduction or human cloning, breach of human dignity, genetic manipulation and probability of tumorogenisity are observed in adult/somatic stem cells. Destruction of embryo to generate stem cell is an important matter in Iran. In this regards, obtaining stem cell from donated frozen embryos through infertility treatment that would be discarded is an acceptable solution in Iran for generation of embryo for research. Ethical, legal, and jurisprudence strategies for using adult/somatic stem cells are determination of ownership of stem cells, trade prohibition of human body, supervision on bio banks and information of Oversight Committee on Stem Cell Research. Recommendations to handle ethical issues for conducting stem cell research are well-designed studies, compliance codes of ethics in biomedical research (specifically codes of ethics on stem cell research, codes of ethics on clinical trials studies and codes of ethics on animals studies), appropriate collaboration with ethics committees and respecting of rights of participants (including both of human and animal rights) in research. In addition, there is a necessity for extending global networks of bioethics for strengthening communications within organizations at both the regional and international level, strengthening legislation systems, designing and establishing convenient collaborative educational courses at different levels. PMID:28462397

Effect of Dedifferentiation on Time to Mutation Acquisition in Stem Cell-Driven Cancers

PubMed Central

Jilkine, Alexandra; Gutenkunst, Ryan N.

2014-01-01

Accumulating evidence suggests that many tumors have a hierarchical organization, with the bulk of the tumor composed of relatively differentiated short-lived progenitor cells that are maintained by a small population of undifferentiated long-lived cancer stem cells. It is unclear, however, whether cancer stem cells originate from normal stem cells or from dedifferentiated progenitor cells. To address this, we mathematically modeled the effect of dedifferentiation on carcinogenesis. We considered a hybrid stochastic-deterministic model of mutation accumulation in both stem cells and progenitors, including dedifferentiation of progenitor cells to a stem cell-like state. We performed exact computer simulations of the emergence of tumor subpopulations with two mutations, and we derived semi-analytical estimates for the waiting time distribution to fixation. Our results suggest that dedifferentiation may play an important role in carcinogenesis, depending on how stem cell homeostasis is maintained. If the stem cell population size is held strictly constant (due to all divisions being asymmetric), we found that dedifferentiation acts like a positive selective force in the stem cell population and thus speeds carcinogenesis. If the stem cell population size is allowed to vary stochastically with density-dependent reproduction rates (allowing both symmetric and asymmetric divisions), we found that dedifferentiation beyond a critical threshold leads to exponential growth of the stem cell population. Thus, dedifferentiation may play a crucial role, the common modeling assumption of constant stem cell population size may not be adequate, and further progress in understanding carcinogenesis demands a more detailed mechanistic understanding of stem cell homeostasis. PMID:24603301

Differential diagnosis of Bartter syndrome, Gitelman syndrome, and pseudo-Bartter/Gitelman syndrome based on clinical characteristics.

PubMed

Matsunoshita, Natsuki; Nozu, Kandai; Shono, Akemi; Nozu, Yoshimi; Fu, Xue Jun; Morisada, Naoya; Kamiyoshi, Naohiro; Ohtsubo, Hiromi; Ninchoji, Takeshi; Minamikawa, Shogo; Yamamura, Tomohiko; Nakanishi, Koichi; Yoshikawa, Norishige; Shima, Yuko; Kaito, Hiroshi; Iijima, Kazumoto

2016-02-01

Phenotypic overlap exists among type III Bartter syndrome (BS), Gitelman syndrome (GS), and pseudo-BS/GS (p-BS/GS), which are clinically difficult to distinguish. We aimed to clarify the differences between these diseases, allowing accurate diagnosis based on their clinical features. A total of 163 patients with genetically defined type III BS (n = 30), GS (n = 90), and p-BS/GS (n = 43) were included. Age at diagnosis, sex, body mass index, estimated glomerular filtration rate, and serum and urine electrolyte concentrations were determined. Patients with p-BS/GS were significantly older at diagnosis than those with type III BS and GS. Patients with p-BS/GS included a significantly higher percentage of women and had a lower body mass index and estimated glomerular filtration rate than did patients with GS. Although hypomagnesemia and hypocalciuria were predominant biochemical findings in patients with GS, 17 and 23% of patients with type III BS and p-BS/GS, respectively, also showed these abnormalities. Of patients with type III BS, GS, and p-BS/GS, 40, 12, and 63%, respectively, presented with chronic kidney disease. This study clarified the clinical differences between BS, GS, and p-BS/GS for the first time, which will help clinicians establish differential diagnoses for these three conditions.

Clinical significance of prospectively assigned Gleason tertiary pattern 4 in contemporary Gleason score 3+3=6 prostate cancer.

PubMed

Doshi, Chirag; Vacchio, Michael; Attwood, Kristopher; Murekeyisoni, Christine; Mehedint, Diana C; Badkhshan, Shervin; Azabdaftari, Gissou; Sule, Norbert; Guru, Khurshid A; Mohler, James L; Kauffman, Eric C

2016-06-01

To determine the oncologic impact of prospectively assigned tertiary pattern 4 in contemporary Gleason score (GS) 3 + 3 = 6 radical prostatectomy (RP) specimens. Oncologic outcomes were retrospectively reviewed for 720 consecutive patients from a single National Comprehensive Cancer Network (NCCN) center with at least 6 months follow-up after RP for GS3 + 3 = 6 (GS6, N = 222), GS6 with tertiary pattern 4 (GS6t4, N = 62), or GS3 + 4 = 7 (N = 436) prostate cancer, as prospectively graded since 2006 using the 2005 International Society of Urologic Pathologists criteria. Preoperative NCCN risk category, RP pathology, progression-free survival (PFS) and metastasis-free survival (MFS) were compared among the GS6, GS6t4, and GS3 + 4 = 7 groups using χ(2) , Kaplan-Meier, and log-rank analyses. The incidence of low NCCN preoperative risk classification for GS6t4 patients (63%) was less than that for GS6 patients (77%) while greater than that for GS3 + 4 = 7 patients (30%, P < 0.001). GS6t4 patients had RP pathologic features which were intermediate in risk between that of GS6 and GS3 + 4 = 7 based on extraprostatic extension (27% vs. 6% vs. 31%, respectively, P < 0.001) and mean percentage of prostate gland involvement (13% vs. 10% vs. 16%, respectively, P < 0.001). With a mean overall follow-up of 42 months, PFS for GS6t4 patients (5-year 85%) was intermediate between that of GS6 (5-year 93%) and GS3 + 4 = 7 (5-year 76%) patients (P < 0.001). The 5-year MFS rate was 100% for GS6 and GS6t4 patients compared to 97% for GS3 + 4 = 7 patients (P = 0.07). This study provides the longest follow-up to date for RP patients with prospectively assigned GS6t4 and supports a risk for adverse RP pathology and postoperative disease progression that is intermediate between GS6 and GS3 + 4 = 7. Whether a tertiary pattern 4 in GS6 disease increases the risk of metastasis is uncertain and requires longer term study. Given favorable oncologic outcomes, less stringent postoperative surveillance for both GS6 and GS6t4 patients may be warranted. © 2016 Wiley Periodicals, Inc.

PGE2 /EP4 Signaling Controls the Transfer of the Mammary Stem Cell State by Lipid Rafts in Extracellular Vesicles.

PubMed

Lin, Meng-Chieh; Chen, Shih-Yin; Tsai, Ho-Min; He, Pei-Lin; Lin, Yen-Chun; Herschman, Harvey; Li, Hua-Jung

2017-02-01

Prostaglandin E 2 (PGE 2 )-initiated signaling contributes to stem cell homeostasis and regeneration. However, it is unclear how PGE 2 signaling controls cell stemness. This study identifies a previously unknown mechanism by which PGE 2 /prostaglandin E receptor 4 (EP 4 ) signaling regulates multiple signaling pathways (e.g., PI3K/Akt signaling, TGFβ signaling, Wnt signaling, EGFR signaling) which maintain the basal mammary stem cell phenotype. A shift of basal mammary epithelial stem cells (MaSCs) from a mesenchymal/stem cell state to a non-basal-MaSC state occurs in response to prostaglandin E receptor 4 (EP 4 ) antagonism. EP 4 antagonists elicit release of signaling components, by controlling their trafficking into extracellular vesicles/exosomes in a lipid raft/caveolae-dependent manner. Consequently, EP 4 antagonism indirectly inactivates, through induced extracellular vesicle/exosome release, pathways required for mammary epithelial stem cell homeostasis, e.g. canonical/noncanonical Wnt, TGFβ and PI3K/Akt pathways. EP 4 antagonism causes signaling receptors and signaling components to shift from non-lipid raft fractions to lipid raft fractions, and to then be released in EP 4 antagonist-induced extracellular vesicles/exosomes, resulting in the loss of the stem cell state by mammary epithelial stem cells. In contrast, luminal mammary epithelial cells can acquire basal stem cell properties following ingestion of EP 4 antagonist-induced stem cell extracellular vesicles/exosomes, and can then form mammary glands. These findings demonstrate that PGE 2 /EP 4 signaling controls homeostasis of mammary epithelial stem cells through regulating extracellular vesicle/exosome release. Reprogramming of mammary epithelial cells can result from EP 4 -mediated stem cell property transfer by extracellular vesicles/exosomes containing caveolae-associated proteins, between mammary basal and luminal epithelial cells. Stem Cells 2017;35:425-444. © 2016 The Authors STEM CELLS published by Wiley Periodicals, Inc. on behalf of AlphaMed Press.

The Drosophila ovarian and testis stem cell niches: similar somatic stem cells and signals.

PubMed

Decotto, Eva; Spradling, Allan C

2005-10-01

The stem cell niches at the apex of Drosophila ovaries and testes have been viewed as distinct in two major respects. While both contain germline stem cells, the testis niche also contains "cyst progenitor" stem cells, which divide to produce somatic cells that encase developing germ cells. Moreover, while both niches utilize BMP signaling, the testis niche requires a key JAK/STAT signal. We now show, by lineage marking, that the ovarian niche also contains a second type of stem cell. These "escort stem cells" morphologically resemble testis cyst progenitor cells and their daughters encase developing cysts before undergoing apoptosis at the time of follicle formation. In addition, we show that JAK/STAT signaling also plays a critical role in ovarian niche function, and acts within escort cells. These observations reveal striking similarities in the stem cell niches of male and female gonads, and suggest that they are largely governed by common mechanisms.

Placenta-an alternative source of stem cells

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

Matikainen, Tiina; Laine, Jarmo

2005-09-01

The two most promising practical applications of human stem cells are cellular replacement therapies in human disease and toxicological screening of candidate drug molecules. Both require a source of human stem cells that can be isolated, purified, expanded in number and differentiated into the cell type of choice in a controlled manner. Currently, uses of both embryonic and adult stem cells are investigated. While embryonic stem cells are pluripotent and can differentiate into any specialised cell type, their use requires establishment of embryonic stem cell lines using the inner cell mass of an early pre-implantation embryo. As the blastocyst ismore » destroyed during the process, ethical issues need to be carefully considered. The use of embryonic stem cells is also limited by the difficulties in growing large numbers of the cells without inducing spontaneous differentiation, and the problems in controlling directed differentiation of the cells. The use of adult stem cells, typically derived from bone marrow, but also from other tissues, is ethically non-controversial but their differentiation potential is more limited than that of the embryonic stem cells. Since human cord blood, umbilical cord, placenta and amnion are normally discarded at birth, they provide an easily accessible alternative source of stem cells. We review the potential and current status of the use of adult stem cells derived from the placenta or umbilical cord in therapeutic and toxicological applications.« less

Pluripotent stem cells and reprogrammed cells in farm animals.

PubMed

Nowak-Imialek, Monika; Kues, Wilfried; Carnwath, Joseph W; Niemann, Heiner

2011-08-01

Pluripotent cells are unique because of their ability to differentiate into the cell lineages forming the entire organism. True pluripotent stem cells with germ line contribution have been reported for mice and rats. Human pluripotent cells share numerous features of pluripotentiality, but confirmation of their in vivo capacity for germ line contribution is impossible due to ethical and legal restrictions. Progress toward derivation of embryonic stem cells from domestic species has been made, but the derived cells were not able to produce germ line chimeras and thus are termed embryonic stem-like cells. However, domestic animals, in particular the domestic pig (Sus scrofa), are excellent large animals models, in which the clinical potential of stem cell therapies can be studied. Reprogramming technologies for somatic cells, including somatic cell nuclear transfer, cell fusion, in vitro culture in the presence of cell extracts, in vitro conversion of adult unipotent spermatogonial stem cells into germ line derived pluripotent stem cells, and transduction with reprogramming factors have been developed with the goal of obtaining pluripotent, germ line competent stem cells from domestic animals. This review summarizes the present state of the art in the derivation and maintenance of pluripotent stem cells in domestic animals.

Information on Stem Cell Research

MedlinePlus

... of stem cells share similar properties there are differences as well. For example, ES cells and iPS cells are able to differentiate into any type of cell, whereas adult stem cells are more restricted in their potential. The promise of all stem cells for use ...

The Development of Stem Cell-Based Treatment for Liver Failure.

PubMed

Zhu, Tiantian; Li, Yuwen; Guo, Yusheng; Zhu, Chuanlong

2017-01-01

Liver failure is a devastating clinical syndrome with a persistently mortality rate despite advanced care. Orthotopic liver transplantation protected patients from hepatic failure. Yet, limitations including postoperative complications, high costs, and shortages of donor organs defect its application. The development of stem cell therapy complements the deficiencies of liver transplantation, due to the inherent ability of stem cells to proliferate and differentiate. Understand the source of stem cells, as well as the advantages and disadvantages of stem cell therapy. Based on published papers, we discussed the cell sources and therapeutic effect of stem cells. We also summarized the pros and cons, as well as optimization of stem cell-based treatment. Finally outlook future prospects of stem cell therapy. Stem cells may be harvested from a variety of human tissues, and then used to promote the convalescence of hepatocellular function. The emergence of the co-cultured system, tissueengineered technology and genetic modfication has further enhanced the functionality of stem cells. However, the tumorigenicity, the low survival rate and the scarcity of long-term treatment effect are obstacles for the further development of stem cell therapy. In this review, we highlight current research findings and present the future prospects in the area of stem cell-based treatment for liver failure. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

76 FR 11491 - Advisory Council on Blood Stem Cell Transplantation; Request for Nominations for Voting Members

Federal Register 2010, 2011, 2012, 2013, 2014

2011-03-02

... Council on Blood Stem Cell Transplantation; Request for Nominations for Voting Members AGENCY: Health... on Blood Stem Cell Transplantation. The Advisory Council on Blood Stem Cell Transplantation was...: Nominations should be submitted to the Executive Secretary, Advisory Council on Blood Stem Cell...

3 CFR - Guidelines for Human Stem Cell Research

Code of Federal Regulations, 2010 CFR

2010-01-01

... 3 The President 1 2010-01-01 2010-01-01 false Guidelines for Human Stem Cell Research Presidential Documents Other Presidential Documents Memorandum of July 30, 2009 Guidelines for Human Stem Cell Research..., scientifically worthy human stem cell research, including human embryonic stem cell research, to the extent...

Aging, metabolism and stem cells: Spotlight on muscle stem cells.

PubMed

García-Prat, Laura; Muñoz-Cánoves, Pura

2017-04-15

All tissues and organs undergo a progressive regenerative decline as they age. This decline has been mainly attributed to loss of stem cell number and/or function, and both stem cell-intrinsic changes and alterations in local niches and/or systemic environment over time are known to contribute to the stem cell aging phenotype. Advancing in the molecular understanding of the deterioration of stem cell cells with aging is key for targeting the specific causes of tissue regenerative dysfunction at advanced stages of life. Here, we revise exciting recent findings on why stem cells age and the consequences on tissue regeneration, with a special focus on regeneration of skeletal muscle. We also highlight newly identified common molecular pathways affecting diverse types of aging stem cells, such as altered proteostasis, metabolism, or senescence entry, and discuss the questions raised by these findings. Finally, we comment on emerging stem cell rejuvenation strategies, principally emanating from studies on muscle stem cells, which will surely burst tissue regeneration research for future benefit of the increasing human aging population. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

In vitro spatially organizing the differentiation in individual multicellular stem cell aggregates.

PubMed

Qi, Hao; Huang, Guoyou; Han, Yu Long; Lin, Wang; Li, Xiujun; Wang, Shuqi; Lu, Tian Jian; Xu, Feng

2016-01-01

With significant potential as a robust source to produce specific somatic cells for regenerative medicine, stem cells have attracted increasing attention from both academia and government. In vivo, stem cell differentiation is a process under complicated regulations to precisely build tissue with unique spatial structures. Since multicellular spheroidal aggregates of stem cells, commonly called as embryoid bodies (EBs), are considered to be capable of recapitulating the events in early stage of embryonic development, a variety of methods have been developed to form EBs in vitro for studying differentiation of embryonic stem cells. The regulation of stem cell differentiation is crucial in directing stem cells to build tissue with the correct spatial architecture for specific functions. However, stem cells within the three-dimensional multicellular aggregates undergo differentiation in a less unpredictable and spatially controlled manner in vitro than in vivo. Recently, various microengineering technologies have been developed to manipulate stem cells in vitro in a spatially controlled manner. Herein, we take the spotlight on these technologies and researches that bring us the new potential for manipulation of stem cells for specific purposes.

Stem cells in nephrology: present status and future.

PubMed

Watorek, Ewa; Klinger, Marian

2006-01-01

Stem cell biology is currently developing rapidly because of the potential therapeutic utility of stem cells. The ability to acquire any desired phenotype raises hope for regenerative therapies. Manipulation of these cells is a potentially valuable tool; however, the mechanisms of stem cell differentiation and plasticity are currently beyond our control. In the field of nephrology, the presence of adult kidney stem cells has been debated. Renal adult stem cells may be descendants of some early kidney progenitors, or may be derived from bone marrow. Evidence of a hematopoietic stem-cell contribution to renal repair encourages the possibility of bone marrow or stem cell transplantation as a means of treating autoimmune glomerulopathies. The transplantation of fetal kidney tissue containing renal progenitors, which then develop into functional nephrons, is a step towards renal regeneration. According to recent reports, the development of functional nephrons from human mesenchymal stem cells in rodent whole-embryo culture is possible. Establishing in vitro self organs from autologous stem cells would be a promising therapeutic solution in light of the shortage of allogenic organs and the unresolved problem of chronic allograft rejection.

Socializing with the neighbors: stem cells and their niche.

PubMed

Fuchs, Elaine; Tumbar, Tudorita; Guasch, Geraldine

2004-03-19

The potential of stem cells in regenerative medicine relies upon removing them from their natural habitat, propagating them in culture, and placing them into a foreign tissue environment. To do so, it is essential to understand how stem cells interact with their microenvironment, the so-called stem cell niche, to establish and maintain their properties. In this review, we examine adult stem cell niches and their impact on stem cell biology.

Stem Cells News Update: A Personal Perspective

PubMed Central

Wong, SC

2013-01-01

This article is a follow-up to a previous Commentary published in 2011. It updates some of the events mentioned in that Commentary and continues with more interesting and exciting news on stem cell research and the emerging field of Regenerative Medicine. Some of the news includes: 1) the 2012 Nobel Prize for Medicine awarded to John B. Gurdon and Shinya Yamanaka; 2) the cloning of human embryonic stem cells; 3) the continued search for truly pluripotent adult stem cells via in vitro and in vivo protocols; 4) the breakthrough in organ replacements; 5) the global stem cell race; 6) the global stem cell cryo-preservation business; 7) the worldwide stem cell donor registries, and 8) the issue of government regulation on stem cell therapy. PMID:24778557

Stem cells news update: a personal perspective.

PubMed

Wong, Sc

2013-12-01

This article is a follow-up to a previous Commentary published in 2011. It updates some of the events mentioned in that Commentary and continues with more interesting and exciting news on stem cell research and the emerging field of Regenerative Medicine. Some of the news includes: 1) the 2012 Nobel Prize for Medicine awarded to John B. Gurdon and Shinya Yamanaka; 2) the cloning of human embryonic stem cells; 3) the continued search for truly pluripotent adult stem cells via in vitro and in vivo protocols; 4) the breakthrough in organ replacements; 5) the global stem cell race; 6) the global stem cell cryo-preservation business; 7) the worldwide stem cell donor registries, and 8) the issue of government regulation on stem cell therapy.

Platelet-rich plasma derived growth factors contribute to stem cell differentiation in musculoskeletal regeneration

NASA Astrophysics Data System (ADS)

Qian, Yun; Han, Qixin; Chen, Wei; Song, Jialin; Zhao, Xiaotian; Ouyang, Yuanming; Yuan, Weien; Fan, Cunyi

2017-10-01

Stem cell treatment and platelet-rich plasma (PRP) therapy are two significant issues in regenerative medicine. Stem cells such as bone marrow mesenchymal stem cells, adipose-derived stem cells and periodontal ligament stem cells can be successfully applied in the field of tissue regeneration. PRP, a natural product isolated from whole blood, can secrete multiple growth factors (GFs) for regulating physiological activities. These GFs can stimulate proliferation and differentiation of different stem cells in injury models. Therefore, combination of both agents receives wide expectations in regenerative medicine, especially in bone, cartilage and tendon repair. In this review, we thoroughly discussed the interaction and underlying mechanisms of platelet-rich plasma derived growth factors with stem cells, and assessed their functions in cell differentiation for musculoskeletal regeneration.

Elements of the niche for adult stem cell expansion

PubMed Central

Redondo, Patricia A; Pavlou, Marina; Loizidou, Marilena; Cheema, Umber

2017-01-01

Adult stem cells are crucial for tissue homeostasis. These cells reside within exclusive locations in tissues, termed niches, which protect adult stem cell fidelity and regulate their many functions through biophysical-, biochemical- and cellular-mediated mechanisms. There is a growing understanding of how these mechanisms and their components contribute towards maintaining stem cell quiescence, self-renewal, expansion and differentiation patterns. In vitro expansion of adult stem cells is a powerful tool for understanding stem cell biology, and for tissue engineering and regenerative medicine applications. However, it is technically challenging, since adult stem cell removal from their native microenvironment has negative repercussions on their sustainability. In this review, we overview specific elements of the biomimetic niche and how recreating such elements can help in vitro propagation of adult stem cells. PMID:28890779

Platelet-Rich Plasma Derived Growth Factors Contribute to Stem Cell Differentiation in Musculoskeletal Regeneration.

PubMed

Qian, Yun; Han, Qixin; Chen, Wei; Song, Jialin; Zhao, Xiaotian; Ouyang, Yuanming; Yuan, Weien; Fan, Cunyi

2017-01-01

Stem cell treatment and platelet-rich plasma (PRP) therapy are two significant issues in regenerative medicine. Stem cells such as bone marrow mesenchymal stem cells, adipose-derived stem cells and periodontal ligament stem cells can be successfully applied in the field of tissue regeneration. PRP, a natural product isolated from whole blood, can secrete multiple growth factors (GFs) for regulating physiological activities. These GFs can stimulate proliferation and differentiation of different stem cells in injury models. Therefore, combination of both agents receives wide expectations in regenerative medicine, especially in bone, cartilage and tendon repair. In this review, we thoroughly discussed the interaction and underlying mechanisms of PRP derived GFs with stem cells, and assessed their functions in cell differentiation for musculoskeletal regeneration.

Elements of the niche for adult stem cell expansion.

PubMed

Redondo, Patricia A; Pavlou, Marina; Loizidou, Marilena; Cheema, Umber

2017-01-01

Adult stem cells are crucial for tissue homeostasis. These cells reside within exclusive locations in tissues, termed niches, which protect adult stem cell fidelity and regulate their many functions through biophysical-, biochemical- and cellular-mediated mechanisms. There is a growing understanding of how these mechanisms and their components contribute towards maintaining stem cell quiescence, self-renewal, expansion and differentiation patterns. In vitro expansion of adult stem cells is a powerful tool for understanding stem cell biology, and for tissue engineering and regenerative medicine applications. However, it is technically challenging, since adult stem cell removal from their native microenvironment has negative repercussions on their sustainability. In this review, we overview specific elements of the biomimetic niche and how recreating such elements can help in vitro propagation of adult stem cells.

Stem cell maintenance by manipulating signaling pathways: past, current and future

PubMed Central

Chen, Xi; Ye, Shoudong; Ying, Qi-Long

2015-01-01

Pluripotent stem cells only exist in a narrow window during early embryonic development, whereas multipotent stem cells are abundant throughout embryonic development and are retainedin various adult tissues and organs. While pluripotent stem cell lines have been established from several species, including mouse, rat, and human, it is still challenging to establish stable multipotent stem cell lines from embryonic or adult tissues. Based on current knowledge, we anticipate that by manipulating extrinsic and intrinsic signaling pathways, most if not all types of stem cells can be maintained in a long-term culture. In this article, we summarize current culture conditions established for the long-term maintenance of authentic pluripotent and multipotent stem cells and the signaling pathways involved. We also discuss the general principles of stem cell maintenance and propose several strategies on the establishment of novel stem cell lines through manipulation of signaling pathways. [BMB Reports 2015; 48(12): 668-676] PMID:26497581

Stem cells and female reproduction.

PubMed

Du, Hongling; Taylor, Hugh S

2009-02-01

Several recent findings in stem cell biology have resulted in new opportunities for the treatment of reproductive disease. Endometrial regeneration can be driven by bone marrow derived stem cells. This finding has potential implications for the treatment of uterine disorders. It also supports a new theory for the etiology of endometriosis. The ovaries have been shown to contain stem cells that form oocytes in adults and can be cultured in vitro to develop mature oocytes. Stem cells from the fetus have been demonstrated to lead to microchimerism in the mother and implicated in several maternal diseases. Additionally the placenta may be another source of hematopoietic stem cell. Finally endometrial derived stem cells have been demonstrated to differentiate into non-reproductive tissues. While we are just beginning to understand stem cells and many key questions remain, the potential advantages of stem cells in reproductive biology and medicine are apparent.

The potential of nanofibers in tissue engineering and stem cell therapy.

PubMed

Gholizadeh-Ghaleh Aziz, Shiva; Gholizadeh-Ghaleh Aziz, Sara; Akbarzadeh, Abolfazl

2016-08-01

Electrospinning is a technique in which materials in solution are shaped into continuous nano- and micro-sized fibers. Combining stem cells with biomaterial scaffolds and nanofibers affords a favorable approach for bone tissue engineering, stem cell growth and transfer, ocular surface reconstruction, and treatment of congenital corneal diseases. This review seeks to describe the current examples of the use of scaffolds in stem cell therapy. Stem cells are classified as adult or embryonic stem (ES) cells, and the advantages and drawbacks of each group are detailed. The nanofibers and scaffolds are further classified in Tables I and II , which describe specific examples from the literature. Finally, the current applications of biomaterial scaffolds containing stem cells for tissue engineering applications are presented. Overall, this review seeks to give an overview of the biomaterials available for use in combination with stem cells, and the application of nanofibers in stem cell therapy.

The stem cell patent landscape as relevant to cancer vaccines.

PubMed

Wang, Shyh-Jen

2011-10-01

Cancer vaccine targeting cancer stem cells is proposed to serve as a potent immunotherapy. Thus, it would be useful to examine the main trends in stem cell patenting activity as a guide for those seeking to develop such cancer vaccines. We found that a substantial number of stem cell patents were granted up to the end of 2010, including ~2000 issued in the US. Many of these have been filed since 2001, including 7,551 applications in the US. Stem cell development, as evidenced by the numbers of PubMed articles, has matured steadily in recent years. However, the other metrics, such as the number of patent applications, the technology-science linkage and the number of patent assignees, have been stagnant. Moreover, the ownership of stem cell patents is still quiet fragmented across multiple organizations, and the number of stem cell patent assignees from the business sector has not increased significantly. Academic and nonprofit institutions not only account for a large share of stem cell patents but also apply for patents continually. Based on this analysis, the strength of stem cell resources seems to remain stagnant in recent years due to the ban on government funding of embryonic stem cell research. Furthermore, the patent prosecution or technical barriers in the field of stem cells would be another main reason that the number of US-issued stem cell patents for each application have been in gradual decline since 2000. Therefore, we consider stem cell technology to still be under development.

Fraudsters operate and officialdom turns a blind eye: a proposal for controlling stem cell therapy in China.

PubMed

Jiang, Li; Dong, Bing He

2016-09-01

Stem cell tourism-the flow of patients from home countries to destination countries to obtain stem cell treatment-is a growing business in China. Many concerns have been raised regarding fraudsters that operate unsafe stem cell therapies and an officialdom that turns a blind eye to the questionable technology. The Chinese regulatory approach to stem cell research is based on Guidelines and Administrative Measures, rather than legislation, and may have no binding force on certain institutions, such as military hospitals. There is no liability and traceability system and no visible set of penalties for non-compliance in the stem cell legal framework. In addition to the lack of safety and efficacy systems in the regulations, no specific expert authority has been established to monitor stem cell therapy to date. Recognizing the global nature of stem cell tourism, this article argues that resolving stem cell tourism issues may require not only the Chinese government but also an international mechanism for transparency and ethical oversight. A stringent set of international regulations that govern stem cell therapies can encourage China to improve stem cell regulation and enforcement to fulfill its obligations. Through an international consensus, a minimum standard for clinical stem cell research and a central enforcement system will be provided. As a result, rogue clinics that conduct unauthorized stem cell therapies can be penalized, and countries that are reluctant to implement the reconciled regulations should be sanctioned.

Comparative Transcriptome Analyses Reveal a Special Glucosinolate Metabolism Mechanism in Brassica alboglabra Sprouts

PubMed Central

Guo, Rongfang; Huang, Zhongkai; Deng, Yanping; Chen, Xiaodong; XuHan, Xu; Lai, Zhongxiong

2016-01-01

Brassica sprouts contain abundant phytochemicals, especially glucosinolates (GSs). Various methods have been used to enhance GS content in sprouts. However, the molecular basis of GS metabolism in sprouts remains an open question. Here we employed RNA-seq analysis to compare the transcriptomes of high-GS (JL-08) and low-GS (JL-09) Brassica alboglabra sprouts. Paired-end Illumina RNA-seq reads were generated and mapped to the Brassica oleracea reference genome. The differentially expressed genes were analyzed between JL-08 and JL-09. Among these, 1477 genes were up-regulated and 1239 down-regulated in JL-09 compared with JL-08. Enrichment analysis of these differentially expressed genes showed that the GS biosynthesis had the smallest enrichment factor and the highest Q-value of all metabolic pathways in Kyoto Encyclopedia of Genes and Genomes database, indicating the main metabolic difference between JL-08 and JL-09 is the GS biosynthetic pathway. Thirty-seven genes of the sequenced data were annotated as putatively involved in GS biosynthesis, degradation, and regulation, of which 11 were differentially expressed in JL-08 and JL-09. The expression level of GS degradation enzyme myrosinase in high-GS JL-08 was lower compared with low-GS JL-09. Surprisingly, in high-GS JL-08, the expression levels of GS biosynthesis genes were also lower than those in low-GS JL-09. As the GS contents in sprouts are determined by dynamic equilibrium of seed stored GS mobilization, de novo synthesis, degradation, and extra transport, the result of this study leads us to suggest that efforts to increase GS content should focus on either raising GS content in seeds or decreasing myrosinase activity, rather than improving the expression level of GS biosynthesis genes in sprouts. PMID:27757119

Drug-polymer interaction between glucosamine sulfate and alginate nanoparticles: FTIR, DSC and dielectric spectroscopy studies

NASA Astrophysics Data System (ADS)

El-Houssiny, A. S.; Ward, A. A.; Mostafa, D. M.; Abd-El-Messieh, S. L.; Abdel-Nour, K. N.; Darwish, M. M.; Khalil, W. A.

2016-06-01

This work involves the preparation and characterization of alginate nanoparticles (Alg NPs) as a new transdermal carrier for site particular transport of glucosamine sulfate (GS). The GS-Alg NPs were examined through transmission electron microscopy (TEM), Fourier transform infrared spectroscopy (FTIR), differential scanning calorimetry (DSC) and dielectric spectroscopy. GS-Alg NPs was efficiently prepared via ionic gelation method which generates favorable conditions for the entrapment of hydrophilic drugs. The TEM studies revealed that GS-Alg NPs are discrete and have spherical shapes. FTIR studies showed a spectral change of the characteristic absorptions bands of Alg NPs after encapsulation with GS because of the amine groups of GS and the carboxylic acid groups of Alg. The DSC data showed changes in the thermal behavior of GS-Alg NPs after the addition of GS indicating signs of main chemical interaction among the drug (GS) and the polymer (Alg). The absence of the drug melting endothermic peak within the DSC thermogram of GS-Alg NPs indicating that GS is molecularly dispersed in the NPs and not crystallize. From the dielectric study, it was found modifications within the dielectric loss (ɛ″) and conductivity (σ) values after the addition of GS. The ɛ″ and σ values of Alg NPs decreased after the addition of GS which indicated the successful encapsulation of GS within Alg NPs. Furthermore, the dielectric study indicated an increase of the activation energy and the relaxation time for the first process in the GS-Alg NPs as compared to Alg NPs. Consequently, the existing observations indicated an initiation of electrostatic interaction among the amine group of GS and carboxyl group of Alg indicating the successful encapsulation of GS inside Alg NPs which could provide favorable circumstance for the encapsulation of GS for topical management.

Epigenetic Control of Stem Cell Potential During Homeostasis, Aging, and Disease

PubMed Central

Beerman, Isabel; Rossi, Derrick J.

2015-01-01

Stem cell decline is an important cellular driver of aging-associated pathophysiology in multiple tissues. Epigenetic regulation is central to establishing and maintaining stem cell function, and emerging evidence indicates that epigenetic dysregulation contributes to the altered potential of stem cells during aging. Unlike terminally differentiated cells, the impact of epigenetic dysregulation in stem cells is propagated beyond self; alterations can be heritably transmitted to differentiated progeny, in addition to being perpetuated and amplified within the stem cell pool through self-renewal divisions. This review focuses on recent studies examining epigenetic regulation of tissue-specific stem cells in homeostasis, aging, and aging-related disease. PMID:26046761

Attitude of A Sample of Iranian Researchers toward The Future of Stem Cell Research.

PubMed

Lotfipanah, Mahdi; Azadeh, Fereydoon; Totonchi, Mehdi; Omani-Samani, Reza

2018-10-01

Stem cells that have unlimited proliferation potential as well as differentiation potency are considered to be a promising future treatment method for incurable diseases. The aim of the present study is to evaluate the future trend of stem cell researches from researchers' viewpoints. This was a cross-sectional descriptive study on researchers involved in stem cell research at Royan Institute. We designed a questionnaire using a qualitative study based on expert opinion and a literature review. Content validity was performed using three rounds of the Delphi method with experts. Face validity was undertaken by a Persian literature expert and a graphics designer. The questionnaire was distributed among 150 researchers involved in stem cell studies in Royan Institute biology laboratories. We collected 138 completed questionnaires. The mean age of participants was 31.13 ± 5.8 years; most (60.9%) were females. Participants (76.1%) considered the budget to be the most important issue in stem cell research, 79.7% needed financial support from the government, and 77.5% felt that charities could contribute substantially to stem cell research. A total of 90.6% of participants stated that stem cells should lead to commercial usage which could support future researches (86.2%). The aim of stem cell research was stipulated as increasing health status of the society according to 92.8% of the participants. At present, among cell types, importance was attached to cord blood and adult stem cells. Researchers emphasized the importance of mesenchymal stem cells (MSCs) rather than hematopoietic stem cells (HSCs, 57.73%). The prime priorities were given to cancer so that stem cell research could be directed to sphere stem cell research whereas the least preference was given to skin research. Regenerative medicine is considered the future of stem cell research with emphasis on application of these cells, especially in cancer treatment. Copyright© by Royan Institute. All rights reserved.

Translating stem cell research: challenges at the research frontier.

PubMed

Magnus, David

2010-01-01

This paper will address the translation of basic stem cell research into clinical research. While "stem cell" trials are sometimes used to describe established practices of bone marrow transplantation or transplantation of primary cells derived from bone marrow, for the purposes of this paper, I am primarily focusing on stem cell trials which are far less established, including use of hESC derived stem cells. The central ethical challenges in stem cell clinical trials arise in frontier research, not in standard, well-established areas of research.

Murine hepatocellular carcinoma derived stem cells reveal epithelial-to-mesenchymal plasticity.

PubMed

Jayachandran, Aparna; Shrestha, Ritu; Dhungel, Bijay; Huang, I-Tao; Vasconcelos, Marianna Yumi Kawashima; Morrison, Brian J; Ramlogan-Steel, Charmaine A; Steel, Jason C

2017-09-26

To establish a model to enrich and characterize stem-like cells from murine normal liver and hepatocellular carcinoma (HCC) cell lines and to further investigate stem-like cell association with epithelial-to-mesenchymal transition (EMT). In this study, we utilized a stem cell conditioned serum-free medium to enrich stem-like cells from mouse HCC and normal liver cell lines, Hepa 1-6 and AML12, respectively. We isolated the 3-dimensional spheres and assessed their stemness characteristics by evaluating the RNA levels of stemness genes and a cell surface stem cell marker by quantitative reverse transcriptase-PCR (qRT-PCR). Next, we examined the relationship between stem cells and EMT using qRT-PCR. Three-dimensional spheres were enriched by culturing murine HCC and normal hepatocyte cell lines in stem cell conditioned serum-free medium supplemented with epidermal growth factor, basic fibroblast growth factor and heparin sulfate. The 3-dimensional spheres had enhanced stemness markers such as Klf4 and Bmi1 and hepatic cancer stem cell (CSC) marker Cd44 compared to parental cells grown as adherent cultures. We report that epithelial markers E-cadherin and ZO-1 were downregulated, while mesenchymal markers Vimentin and Fibronectin were upregulated in 3-dimensional spheres. The 3-dimensional spheres also exhibited changes in expression of Snai , Zeb and Twist family of EMT transcription factors. Our novel method successfully enriched stem-like cells which possessed an EMT phenotype. The isolation and characterization of murine hepatic CSCs could establish a precise target for the development of more effective therapies for HCC.

Wnt/β-Catenin Signaling Determines the Vasculogenic Fate of Postnatal Mesenchymal Stem Cells.

PubMed

Zhang, Zhaocheng; Nör, Felipe; Oh, Min; Cucco, Carolina; Shi, Songtao; Nör, Jacques E

2016-06-01

Vasculogenesis is the process of de novo blood vessel formation observed primarily during embryonic development. Emerging evidence suggest that postnatal mesenchymal stem cells are capable of recapitulating vasculogenesis when these cells are engaged in tissue regeneration. However, the mechanisms underlining the vasculogenic differentiation of mesenchymal stem cells remain unclear. Here, we used stem cells from human permanent teeth (dental pulp stem cells [DPSC]) or deciduous teeth (stem cells from human exfoliated deciduous teeth [SHED]) as models of postnatal primary human mesenchymal stem cells to understand mechanisms regulating their vasculogenic fate. GFP-tagged mesenchymal stem cells seeded in human tooth slice/scaffolds and transplanted into immunodeficient mice differentiate into human blood vessels that anastomize with the mouse vasculature. In vitro, vascular endothelial growth factor (VEGF) induced the vasculogenic differentiation of DPSC and SHED via potent activation of Wnt/β-catenin signaling. Further, activation of Wnt signaling is sufficient to induce the vasculogenic differentiation of postnatal mesenchymal stem cells, while Wnt inhibition blocked this process. Notably, β-catenin-silenced DPSC no longer differentiate into endothelial cells in vitro, and showed impaired vasculogenesis in vivo. Collectively, these data demonstrate that VEGF signaling through the canonical Wnt/β-catenin pathway defines the vasculogenic fate of postnatal mesenchymal stem cells. Stem Cells 2016;34:1576-1587. © 2016 AlphaMed Press.

MiR-7 inhibited peripheral nerve injury repair by affecting neural stem cells migration and proliferation through cdc42.

PubMed

Zhou, Nan; Hao, Shuang; Huang, Zongqiang; Wang, Weiwei; Yan, Penghui; Zhou, Wei; Zhu, Qihang; Liu, Xiaokang

2018-01-01

Objective Neural stem cells play an important role in the recovery and regeneration of peripheral nerve injury, and the microRNA-7 (miR-7) regulates differentiation of neural stem cells. This study aimed to explore the role of miR-7 in neural stem cells homing and proliferation and its influence on peripheral nerve injury repair. Methods The mice model of peripheral nerve injury was created by segmental sciatic nerve defect (sciatic nerve injury), and neural stem cells treatment was performed with a gelatin hydrogel conduit containing neural stem cells inserted into the sciatic nerve injury mice. The Sciatic Function Index was used to quantify sciatic nerve functional recovery in the mice. The messenger RNA and protein expression were detected by reverse transcription polymerase chain reaction and Western blot, respectively. Luciferase reporter assay was used to confirm the binding between miR-7 and the 3'UTR of cell division cycle protein 42 (cdc42). The neural stem cells migration and proliferation were analyzed by transwell assay and a Cell-LightTM EdU DNA Cell Proliferation kit, respectively. Results Neural stem cells treatment significantly promoted nerve repair in sciatic nerve injury mice. MiR-7 expression was decreased in sciatic nerve injury mice with neural stem cells treatment, and miR-7 mimic transfected into neural stem cells suppressed migration and proliferation, while miR-7 inhibitor promoted migration and proliferation. The expression level and effect of cdc42 on neural stem cells migration and proliferation were opposite to miR-7, and the luciferase reporter assay proved that cdc42 was a target of miR-7. Using co-transfection into neural stem cells, we found pcDNA3.1-cdc42 and si-cdc42 could reverse respectively the role of miR-7 mimic and miR-7 inhibitor on neural stem cells migration and proliferation. In addition, miR-7 mimic-transfected neural stem cells could abolish the protective role of neural stem cells on peripheral nerve injury. Conclusion MiR-7 inhibited peripheral nerve injury repair by affecting neural stem cells migration and proliferation through cdc42.

International Society for Stem Cell Research

MedlinePlus

... cell and regenerative medicine community. More stem cell research Take a closer look Recent Blogs View All ... nonprofit organization & the voice of the stem cell research community The International Society for Stem Cell Research ( ...

Stem cell regenerative potential combined with nanotechnology and tissue engineering for myocardial regeneration.

PubMed

Calin, Manuela; Stan, Daniela; Simion, Viorel

2013-07-01

The stem cell-based therapy for post-infarction myocardial regeneration has been introduced more than a decade ago, but the functional improvement obtained is limited due to the poor retention and short survival rate of transplanted cells into the damaged myocardium. More recently, the emerging nanotechnology concepts for advanced diagnostics and therapy provide promising opportunities of using stem cells for myocardial regeneration. In this paper will be provided an overview of the use of nanotechnology approaches in stem cell research for: 1) cell labeling to track the distribution of stem cells after transplantation, 2) nanoparticle-mediated gene delivery to stem cells to promote their homing, engraftment, survival and differentiation in the ischemic myocardium and 3) obtaining of bio-inspired materials to provide suitable myocardial scaffolds for delivery of stem cells or stem cell-derived factors.

A Reading List for A- and S-level Biology--Part VII

ERIC Educational Resources Information Center

Shaw, G. W.

1975-01-01

Presents a list of articles published from January to December, 1974, on genetics, cells, nutrition, respiration, excretion, general physiology, conservation, ecology, reproduction, locomotion, disease, growth, response, behavior, and classification. (GS)

Advances in Bone Marrow Stem Cell Therapy for Retinal Dysfunction

PubMed Central

Park, Susanna S.; Moisseiev, Elad; Bauer, Gerhard; Anderson, Johnathon D.; Grant, Maria B.; Zam, Azhar; Zawadzki, Robert J.; Werner, John S.; Nolta, Jan A.

2016-01-01

The most common cause of untreatable vision loss is dysfunction of the retina. Conditions, such as age-related macular degeneration, diabetic retinopathy and glaucoma remain leading causes of untreatable blindness worldwide. Various stem cell approaches are being explored for treatment of retinal regeneration. The rationale for using bone marrow stem cells to treat retinal dysfunction is based on preclinical evidence showing that bone marrow stem cells can rescue degenerating and ischemic retina. These stem cells have primarily paracrine trophic effects although some cells can directly incorporate into damaged tissue. Since the paracrine trophic effects can have regenerative effects on multiple cells in the retina, the use of this cell therapy is not limited to a particular retinal condition. Autologous bone marrow-derived stem cells are being explored in early clinical trials as therapy for various retinal conditions. These bone marrow stem cells include mesenchymal stem cells, mononuclear cells and CD34+ cells. Autologous therapy requires no systemic immunosuppression or donor matching. Intravitreal delivery of CD34+ cells and mononuclear cells appears to be tolerated and is being explored since some of these cells can home into the damaged retina after intravitreal administration. The safety of intravitreal delivery of mesenchymal stem cells has not been well established. This review provides an update of the current evidence in support of the use of bone marrow stem cells as treatment for retinal dysfunction. The potential limitations and complications of using certain forms of bone marrow stem cells as therapy are discussed. Future directions of research include methods to optimize the therapeutic potential of these stem cells, non-cellular alternatives using extracellular vesicles, and in vivo high-resolution retinal imaging to detect cellular changes in the retina following cell therapy. PMID:27784628

Role of stem cell derived exosomes in tumor biology.

PubMed

Sharma, Aman

2018-03-15

Exosomes are nano-scale messengers loaded with bio-molecular cargo of RNA, DNA, and Proteins. As a master regulator of cellular signaling, stem cell (both normal, and cancer stem cells) secreted exosome orchestrate various autocrine and paracrine functions which alter tumor micro-environment, growth and progression. Exosomes secreted by one of the two important stem cell phenotypes in cancers a) Mesenchymal stem cells, and b) Cancer stem cells not only promote cancerous growth but also impart therapy resistance in cancer cells. In tumors, normal or mesenchymal stem cell (MSCs) derived exosomes (MSC-exo) modulate tumor hallmarks by delivering unique miRNA species to neighboring cells and help in tumor progression. Apart from regulating tumor cell fate, MSC-exo are also capable of inducing physiological processes, for example, angiogenesis, metastasis and so forth. Similarly, cancer stem cells (CSCs) derived exosomes (CSC-exo) contain stemness-specific proteins, self-renewal promoting regulatory miRNAs, and survival factors. CSC-exo specific cargo maintains tumor heterogeneity and alters tumor progression. In this review we critically discuss the importance of stem cell specific exosomes in tumor cell signaling pathways with their role in tumor biology. © 2017 UICC.

Stochasticity and Spatial Interaction Govern Stem Cell Differentiation Dynamics

NASA Astrophysics Data System (ADS)

Smith, Quinton; Stukalin, Evgeny; Kusuma, Sravanti; Gerecht, Sharon; Sun, Sean X.

2015-07-01

Stem cell differentiation underlies many fundamental processes such as development, tissue growth and regeneration, as well as disease progression. Understanding how stem cell differentiation is controlled in mixed cell populations is an important step in developing quantitative models of cell population dynamics. Here we focus on quantifying the role of cell-cell interactions in determining stem cell fate. Toward this, we monitor stem cell differentiation in adherent cultures on micropatterns and collect statistical cell fate data. Results show high cell fate variability and a bimodal probability distribution of stem cell fraction on small (80-140 μm diameter) micropatterns. On larger (225-500 μm diameter) micropatterns, the variability is also high but the distribution of the stem cell fraction becomes unimodal. Using a stochastic model, we analyze the differentiation dynamics and quantitatively determine the differentiation probability as a function of stem cell fraction. Results indicate that stem cells can interact and sense cellular composition in their immediate neighborhood and adjust their differentiation probability accordingly. Blocking epithelial cadherin (E-cadherin) can diminish this cell-cell contact mediated sensing. For larger micropatterns, cell motility adds a spatial dimension to the picture. Taken together, we find stochasticity and cell-cell interactions are important factors in determining cell fate in mixed cell populations.

NANOS2 acts downstream of glial cell line-derived neurotrophic factor signaling to suppress differentiation of spermatogonial stem cells.

PubMed

Sada, Aiko; Hasegawa, Kazuteru; Pin, Pui Han; Saga, Yumiko

2012-02-01

Stem cells are maintained by both stem cell-extrinsic niche signals and stem cell-intrinsic factors. During murine spermatogenesis, glial cell line-derived neurotrophic factor (GDNF) signal emanated from Sertoli cells and germ cell-intrinsic factor NANOS2 represent key regulators for the maintenance of spermatogonial stem cells. However, it remains unclear how these factors intersect in stem cells to control their cellular state. Here, we show that GDNF signaling is essential to maintain NANOS2 expression, and overexpression of Nanos2 can alleviate the stem cell loss phenotype caused by the depletion of Gfra1, a receptor for GDNF. By using an inducible Cre-loxP system, we show that NANOS2 expression is downregulated upon the conditional knockout (cKO) of Gfra1, while ectopic expression of Nanos2 in GFRA1-negative spermatogonia does not induce de novo GFRA1 expression. Furthermore, overexpression of Nanos2 in the Gfra1-cKO testes prevents precocious differentiation of the Gfra1-knockout stem cells and partially rescues the stem cell loss phenotypes of Gfra1-deficient mice, indicating that the stem cell differentiation can be suppressed by NANOS2 even in the absence of GDNF signaling. Taken together, we suggest that NANOS2 acts downstream of GDNF signaling to maintain undifferentiated state of spermatogonial stem cells. Copyright © 2011 AlphaMed Press.

Isolation and functional interrogation of adult human prostate epithelial stem cells at single cell resolution.

PubMed

Hu, Wen-Yang; Hu, Dan-Ping; Xie, Lishi; Li, Ye; Majumdar, Shyama; Nonn, Larisa; Hu, Hong; Shioda, Toshi; Prins, Gail S

2017-08-01

Using primary cultures of normal human prostate epithelial cells, we developed a novel prostasphere-based, label-retention assay that permits identification and isolation of stem cells at a single cell level. Their bona fide stem cell nature was corroborated using in vitro and in vivo regenerative assays and documentation of symmetric/asymmetric division. Robust WNT10B and KRT13 levels without E-cadherin or KRT14 staining distinguished individual stem cells from daughter progenitors in spheroids. Following FACS to isolate label-retaining stem cells from label-free progenitors, RNA-seq identified unique gene signatures for the separate populations which may serve as useful biomarkers. Knockdown of KRT13 or PRAC1 reduced sphere formation and symmetric self-renewal highlighting their role in stem cell maintenance. Pathways analysis identified ribosome biogenesis and membrane estrogen-receptor signaling enriched in stem cells with NF-ĸB signaling enriched in progenitors; activities that were biologically confirmed. Further, bioassays identified heightened autophagy flux and reduced metabolism in stem cells relative to progenitors. These approaches similarly identified stem-like cells from prostate cancer specimens and prostate, breast and colon cancer cell lines suggesting wide applicability. Together, the present studies isolate and identify unique characteristics of normal human prostate stem cells and uncover processes that maintain stem cell homeostasis in the prostate gland. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

Materials as stem cell regulators

PubMed Central

Murphy, William L.; McDevitt, Todd C.; Engler, Adam J.

2014-01-01

The stem cell/material interface is a complex, dynamic microenvironment in which the cell and the material cooperatively dictate one another's fate: the cell by remodelling its surroundings, and the material through its inherent properties (such as adhesivity, stiffness, nanostructure or degradability). Stem cells in contact with materials are able to sense their properties, integrate cues via signal propagation and ultimately translate parallel signalling information into cell fate decisions. However, discovering the mechanisms by which stem cells respond to inherent material characteristics is challenging because of the highly complex, multicomponent signalling milieu present in the stem cell environment. In this Review, we discuss recent evidence that shows that inherent material properties may be engineered to dictate stem cell fate decisions, and overview a subset of the operative signal transduction mechanisms that have begun to emerge. Further developments in stem cell engineering and mechanotransduction are poised to have substantial implications for stem cell biology and regenerative medicine. PMID:24845994

Stem cells in genetically-engineered mouse models of prostate cancer

PubMed Central

Shibata, Maho; Shen, Michael M.

2015-01-01

The cancer stem cell model proposes that tumors have a hierarchical organization in which tumorigenic cells give rise to non-tumorigenic cells, with only a subset of stem-like cells able to propagate the tumor. In the case of prostate cancer, recent analyses of genetically engineered mouse (GEM) models have provided evidence supporting the existence of cancer stem cells in vivo. These studies suggest that cancer stem cells capable of tumor propagation exist at various stages of tumor progression from prostatic intraepithelial neoplasia (PIN) to advanced metastatic and castration-resistant disease. However, studies of stem cells in prostate cancer have been limited by available approaches for evaluating their functional properties in cell culture and transplantation assays. Given the role of the tumor microenvironment and the putative cancer stem cell niche, future studies using GEM models to analyze cancer stem cells in their native tissue microenvironment are likely to be highly informative. PMID:26341780

The evolution of chicken stem cell culture methods.

PubMed

Farzaneh, M; Attari, F; Mozdziak, P E; Khoshnam, S E

2017-12-01

1. The avian embryo is an excellent model for studying embryology and the production of pharmaceutical proteins in transgenic chickens. Furthermore, chicken stem cells have the potential for proliferation and differentiation and emerged as an attractive tool for various cell-based technologies. 2. The objective of these studies is the derivation and culture of these stem cells is the production of transgenic birds for recombinant biomaterials and vaccine manufacture, drug and cytotoxicity testing, as well as to gain insight into basic science, including cell tracking. 3. Despite similarities among the established chicken stem cell lines, fundamental differences have been reported between their culture conditions and applications. Recent conventional protocols used for expansion and culture of chicken stem cells mostly depend on feeder cells, serum-containing media and static culture. 4. Utilising chicken stem cells for generation of cell-based transgenic birds and a variety of vaccines requires large-scale cell production. However, scaling up the conventional adherent chicken stem cells is challenging and labour intensive. Development of a suspension cell culture process for chicken embryonic stem cells (cESCs), chicken primordial germ cells (PGCs) and chicken induced pluripotent stem cells (ciPSCs) will be an important advance for increasing the growth kinetics of these cells. 6. This review describes various approaches and suggestions to achieve optimal cell growth for defined chicken stem cells cultures and use in future manufacturing applications.

Development of hematopoietic stem and progenitor cells from human pluripotent stem cells.

PubMed

Chen, Tong; Wang, Fen; Wu, Mengyao; Wang, Zack Z

2015-07-01

Human pluripotent stem cells (hPSCs), including human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs), provide a new cell source for regenerative medicine, disease modeling, drug discovery, and preclinical toxicity screening. Understanding of the onset and the sequential process of hematopoietic cells from differentiated hPSCs will enable the achievement of personalized medicine and provide an in vitro platform for studying of human hematopoietic development and disease. During embryogenesis, hemogenic endothelial cells, a specified subset of endothelial cells in embryonic endothelium, are the primary source of multipotent hematopoietic stem cells. In this review, we discuss current status in the generation of multipotent hematopoietic stem and progenitor cells from hPSCs via hemogenic endothelial cells. We also review the achievements in direct reprogramming from non-hematopoietic cells to hematopoietic stem and progenitor cells. Further characterization of hematopoietic differentiation in hPSCs will improve our understanding of blood development and expedite the development of hPSC-derived blood products for therapeutic purpose. © 2015 Wiley Periodicals, Inc.

Therapeutic microparticles functionalized with biomimetic cardiac stem cell membranes and secretome

PubMed Central

Tang, Junnan; Shen, Deliang; Caranasos, Thomas George; Wang, Zegen; Vandergriff, Adam C.; Allen, Tyler A.; Hensley, Michael Taylor; Dinh, Phuong-Uyen; Cores, Jhon; Li, Tao-Sheng; Zhang, Jinying; Kan, Quancheng; Cheng, Ke

2017-01-01

Stem cell therapy represents a promising strategy in regenerative medicine. However, cells need to be carefully preserved and processed before usage. In addition, cell transplantation carries immunogenicity and/or tumourigenicity risks. Mounting lines of evidence indicate that stem cells exert their beneficial effects mainly through secretion (of regenerative factors) and membrane-based cell–cell interaction with the injured cells. Here, we fabricate a synthetic cell-mimicking microparticle (CMMP) that recapitulates stem cell functions in tissue repair. CMMPs carry similar secreted proteins and membranes as genuine cardiac stem cells do. In a mouse model of myocardial infarction, injection of CMMPs leads to the preservation of viable myocardium and augmentation of cardiac functions similar to cardiac stem cell therapy. CMMPs (derived from human cells) do not stimulate T-cell infiltration in immuno-competent mice. In conclusion, CMMPs act as ‘synthetic stem cells’ which mimic the paracrine and biointerfacing activities of natural stem cells in therapeutic cardiac regeneration. PMID:28045024

Stem cell plasticity enables hair regeneration following Lgr5+ cell loss.

PubMed

Hoeck, Joerg D; Biehs, Brian; Kurtova, Antonina V; Kljavin, Noelyn M; de Sousa E Melo, Felipe; Alicke, Bruno; Koeppen, Hartmut; Modrusan, Zora; Piskol, Robert; de Sauvage, Frederic J

2017-06-01

Under injury conditions, dedicated stem cell populations govern tissue regeneration. However, the molecular mechanisms that induce stem cell regeneration and enable plasticity are poorly understood. Here, we investigate stem cell recovery in the context of the hair follicle to understand how two molecularly distinct stem cell populations are integrated. Utilizing diphtheria-toxin-mediated cell ablation of Lgr5 + (leucine-rich repeat-containing G-protein-coupled receptor 5) stem cells, we show that killing of Lgr5 + cells in mice abrogates hair regeneration but this is reversible. During recovery, CD34 + (CD34 antigen) stem cells activate inflammatory response programs and start dividing. Pharmacological attenuation of inflammation inhibits CD34 + cell proliferation. Subsequently, the Wnt pathway controls the recovery of Lgr5 + cells and inhibition of Wnt signalling prevents Lgr5 + cell and hair germ recovery. Thus, our study uncovers a compensatory relationship between two stem cell populations and the underlying molecular mechanisms that enable hair follicle regeneration.

SILAC proteomics of planarians identifies Ncoa5 as a conserved component of pluripotent stem cells.

PubMed

Böser, Alexander; Drexler, Hannes C A; Reuter, Hanna; Schmitz, Henning; Wu, Guangming; Schöler, Hans R; Gentile, Luca; Bartscherer, Kerstin

2013-11-27

Planarian regeneration depends on the presence of pluripotent stem cells in the adult. We developed an in vivo stable isotope labeling by amino acids in cell culture (SILAC) protocol in planarians to identify proteins that are enriched in planarian stem cells. Through a comparison of SILAC proteomes of normal and stem cell-depleted planarians and of a stem cell-enriched population of sorted cells, we identified hundreds of stem cell proteins. One of these is an ortholog of nuclear receptor coactivator-5 (Ncoa5/CIA), which is known to regulate estrogen-receptor-mediated transcription in human cells. We show that Ncoa5 is essential for the maintenance of the pluripotent stem cell population in planarians and that a putative mouse ortholog is expressed in pluripotent cells of the embryo. Our study thus identifies a conserved component of pluripotent stem cells, demonstrating that planarians, in particular, when combined with in vivo SILAC, are a powerful model in stem cell research. Copyright © 2013 The Authors. Published by Elsevier Inc. All rights reserved.

Stem cells in reproductive medicine: ready for the patient?

PubMed

Vassena, R; Eguizabal, C; Heindryckx, B; Sermon, K; Simon, C; van Pelt, A M M; Veiga, A; Zambelli, F

2015-09-01

Are there effective and clinically validated stem cell-based therapies for reproductive diseases? At the moment, clinically validated stem cell treatments for reproductive diseases and alterations are not available. Research in stem cells and regenerative medicine is growing in scope, and its translation to the clinic is heralded by the recent initiation of controlled clinical trials with pluripotent derived cells. Unfortunately, stem cell 'treatments' are currently offered to patients outside of the controlled framework of scientifically sound research and regulated clinical trials. Both physicians and patients in reproductive medicine are often unsure about stem cells therapeutic options. An international working group was assembled to review critically the available scientific literature in both the human species and animal models. This review includes work published in English until December 2014, and available through Pubmed. A few areas of research in stem cell and reproductive medicine were identified: in vitro gamete production, endometrial regeneration, erectile dysfunction amelioration, vaginal reconstruction. The stem cells studied range from pluripotent (embryonic stem cells and induced pluripotent stem cells) to monopotent stem cells, such as spermatogonial stem cells or mesenchymal stem cells. The vast majority of studies have been carried out in animal models, with data that are preliminary at best. This review was not conducted in a systematic fashion, and reports in publications not indexed in Pubmed were not analyzed. A much broader clinical knowledge will have to be acquired before translation to the clinic of stem cell therapies in reproductive medicine; patients and physicians should be wary of unfounded claims of improvement of existing medical conditions; at the moment, effective stem cell treatment for reproductive diseases and alterations is not available. None. NA. © The Author 2015. 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.

I.V. infusion of brain-derived neurotrophic factor gene-modified human mesenchymal stem cells protects against injury in a cerebral ischemia model in adult rat.

PubMed

Nomura, T; Honmou, O; Harada, K; Houkin, K; Hamada, H; Kocsis, J D

2005-01-01

I.V. delivery of mesenchymal stem cells prepared from adult bone marrow reduces infarction size and ameliorates functional deficits in rat cerebral ischemia models. Administration of the brain-derived neurotrophic factor to the infarction site has also been demonstrated to be neuroprotective. To test the hypothesis that brain-derived neurotrophic factor contributes to the therapeutic benefits of mesenchymal stem cell delivery, we compared the efficacy of systemic delivery of human mesenchymal stem cells and human mesenchymal stem cells transfected with a fiber-mutant F/RGD adenovirus vector with a brain-derived neurotrophic factor gene (brain-derived neurotrophic factor-human mesenchymal stem cells). A permanent middle cerebral artery occlusion was induced by intraluminal vascular occlusion with a microfilament. Human mesenchymal stem cells and brain-derived neurotrophic factor-human mesenchymal stem cells were i.v. injected into the rats 6 h after middle cerebral artery occlusion. Lesion size was assessed at 6 h, 1, 3 and 7 days using MR imaging, and histological methods. Functional outcome was assessed using the treadmill stress test. Both human mesenchymal stem cells and brain-derived neurotrophic factor-human mesenchymal stem cells reduced lesion volume and elicited functional improvement compared with the control sham group, but the effect was greater in the brain-derived neurotrophic factor-human mesenchymal stem cell group. ELISA analysis of the infarcted hemisphere revealed an increase in brain-derived neurotrophic factor in the human mesenchymal stem cell groups, but a greater increase in the brain-derived neurotrophic factor-human mesenchymal stem cell group. These data support the hypothesis that brain-derived neurotrophic factor contributes to neuroprotection in cerebral ischemia and cellular delivery of brain-derived neurotrophic factor can be achieved by i.v. delivery of human mesenchymal stem cells.

Distinct roles of neuroepithelial-like and radial glia-like progenitor cells in cerebellar regeneration.

PubMed

Kaslin, Jan; Kroehne, Volker; Ganz, Julia; Hans, Stefan; Brand, Michael

2017-04-15

Zebrafish can regenerate after brain injury, and the regenerative process is driven by resident stem cells. Stem cells are heterogeneous in the vertebrate brain, but the significance of having heterogeneous stem cells in regeneration is not understood. Limited availability of specific stem cells might impair the regeneration of particular cell lineages. We studied regeneration of the adult zebrafish cerebellum, which contains two major stem and progenitor cell types: ventricular zone and neuroepithelial cells. Using conditional lineage tracing we demonstrate that cerebellar regeneration depends on the availability of specific stem cells. Radial glia-like cells are thought to be the predominant stem cell type in homeostasis and after injury. However, we find that radial glia-like cells play a minor role in adult cerebellar neurogenesis and in recovery after injury. Instead, we find that neuroepithelial cells are the predominant stem cell type supporting cerebellar regeneration after injury. Zebrafish are able to regenerate many, but not all, cell types in the cerebellum, which emphasizes the need to understand the contribution of different adult neural stem and progenitor cell subtypes in the vertebrate central nervous system. © 2017. Published by The Company of Biologists Ltd.

Effects of serum immunoglobulins from patients with complex regional pain syndrome (CRPS) on depolarisation-induced calcium transients in isolated dorsal root ganglion (DRG) neurons.

PubMed

Reilly, Joanne M; Dharmalingam, Backialakshmi; Marsh, Stephen J; Thompson, Victoria; Goebel, Andreas; Brown, David A

2016-03-01

Complex regional pain syndrome (CRPS) is thought to have an auto-immune component. One such target recently proposed from the effects of auto-immune IgGs on Ca(2+) transients in cardiac myocytes and cell lines is the α1-adrenoceptor. We have tested whether such IgGs exerted comparable effects on nociceptive sensory neurons isolated from rat dorsal root ganglia. Depolarisation-induced [Ca(2+)]i transients were generated by applying 30 mM KCl for 2 min and monitored by Fura-2 fluorescence imaging. No IgGs tested (including 3 from CRPS patients) had any significant effect on these [Ca(2+)]i transients. However, IgG from one CRPS patient consistently and significantly reduced the K(+)-induced response of cells that had been pre-incubated for 24h with a mixture of inflammatory mediators (1 μM histamine, 5-hydroxytryptamine, bradykinin and PGE2). Since this pre-incubation also appeared to induce a comparable inhibitory response to the α1-agonist phenylephrine, this is compatible with the α1-adrenoceptor as a target for CRPS auto-immunity. A mechanism whereby this might enhance pain is suggested. Copyright © 2015. Published by Elsevier Inc.

Role of the Stem Cell Niche in Hormone-induced Tumorigenesis in Fetal Mouse Mammary Epithelium

DTIC Science & Technology

2006-08-01

Develop an immunohistochemical method for identifying stem cells and stem cell niches, and to use this to determine if in utero estrogenic...overstimulation causes changes in the number of stem cells or their niches. To extend the power of ex vivo stem cell isolation and enumeration by providing a...marginal success due primarily to 1) most antibodies previously reputed to be stem cell specific turned out to be present in differentiated mammary

Wnt and BMP Signaling Crosstalk in Regulating Dental Stem Cells: Implications in Dental Tissue Engineering

PubMed Central

Zhang, Fugui; Song, Jinglin; Zhang, Hongmei; Huang, Enyi; Song, Dongzhe; Tollemar, Viktor; Wang, Jing; Wang, Jinhua; Mohammed, Maryam; Wei, Qiang; Fan, Jiaming; Liao, Junyi; Zou, Yulong; Liu, Feng; Hu, Xue; Qu, Xiangyang; Chen, Liqun; Yu, Xinyi; Luu, Hue H.; Lee, Michael J.; He, Tong-Chuan; Ji, Ping

2016-01-01

Tooth is a complex hard tissue organ and consists of multiple cell types that are regulated by important signaling pathways such as Wnt and BMP signaling. Serious injuries and/or loss of tooth or periodontal tissues may significantly impact aesthetic appearance, essential oral functions and the quality of life. Regenerative dentistry holds great promise in treating oral/dental disorders. The past decade has witnessed a rapid expansion of our understanding of the biological features of dental stem cells, along with the signaling mechanisms governing stem cell self-renewal and differentiation. In this review, we first summarize the biological characteristics of seven types of dental stem cells, including dental pulp stem cells, stem cells from apical papilla, stem cells from human exfoliated deciduous teeth, dental follicle precursor cells, periodontal ligament stem cells, alveolar bone-derived mesenchymal stem cells (MSCs), and MSCs from gingiva. We then focus on how these stem cells are regulated by bone morphogenetic protein (BMP) and/or Wnt signaling by examining the interplays between these pathways. Lastly, we analyze the current status of dental tissue engineering strategies that utilize oral/dental stem cells by harnessing the interplays between BMP and Wnt pathways. We also highlight the challenges that must be addressed before the dental stem cells may reach any clinical applications. Thus, we can expect to witness significant progresses to be made in regenerative dentistry in the coming decade. PMID:28491933

Extracellular Matrix as a Regulator of Epidermal Stem Cell Fate.

PubMed

Chermnykh, Elina; Kalabusheva, Ekaterina; Vorotelyak, Ekaterina

2018-03-27

Epidermal stem cells reside within the specific anatomic location, called niche, which is a microenvironment that interacts with stem cells to regulate their fate. Regulation of many important processes, including maintenance of stem cell quiescence, self-renewal, and homeostasis, as well as the regulation of division and differentiation, are common functions of the stem cell niche. As it was shown in multiple studies, extracellular matrix (ECM) contributes a lot to stem cell niches in various tissues, including that of skin. In epidermis, ECM is represented, primarily, by a highly specialized ECM structure, basement membrane (BM), which separates the epidermal and dermal compartments. Epidermal stem cells contact with BM, but when they lose the contact and migrate to the overlying layers, they undergo terminal differentiation. When considering all of these factors, ECM is of fundamental importance in regulating epidermal stem cells maintenance, proper mobilization, and differentiation. Here, we summarize the remarkable progress that has recently been made in the research of ECM role in regulating epidermal stem cell fate, paying special attention to the hair follicle stem cell niche. We show that the destruction of ECM components impairs epidermal stem cell morphogenesis and homeostasis. A deep understanding of ECM molecular structure as well as the development of in vitro system for stem cell maintaining by ECM proteins may bring us to developing new approaches for regenerative medicine.

Knowledge and Attitude about Stem Cells and Their Application in Medicine among Nursing Students in Universiti Sains Malaysia, Malaysia

PubMed Central

LYE, Jee Leng; SOON, Lean Keng; WAN AHMAD, Wan Amir Nizam; TAN, Suat Cheng

2015-01-01

Background: Stem cell research has been extensively explored worldwide to enhance human health in medical setting. Nevertheless, there is currently no full understanding of the stem cell knowledge and attitude levels among student nurses in Malaysia. This study aimed to assess the level of stem cell knowledge, attitude toward stem cell application in medicine, and its association with years of education, among Universiti Sains Malaysia (USM) undergraduate nursing students. Methods: A cross-sectional study (n = 88) was conducted using self-administered questionnaire consisted of demographic information, stem cells knowledge and attitude statements. Data was analysed using Statistical Package Social Software 20.0. Results: The majority of participants (92%) had moderate knowledge score about stem cells. Many students (33%) worried that stem cell application might cause a harm to humanity yet had a positive (76.1%) attitude towards its therapeutic potential (45.5%). Poor correlation between knowledge and attitude (r = 0.08) indicated that acceptance towards stem cell is not solely based on the knowledge level but also on other factors including religion and culture. Conclusion: Therefore, this study suggests that various educational programs on stem cell should be implemented considering the religion, cultural, social, and behavioural determinants in the population to improve stem cell knowledge and encourage a more positive attitude towards stem cells in medicine among these nursing students. PMID:26715905

Knowledge and Attitude about Stem Cells and Their Application in Medicine among Nursing Students in Universiti Sains Malaysia, Malaysia.

PubMed

Lye, Jee Leng; Soon, Lean Keng; Wan Ahmad, Wan Amir Nizam; Tan, Suat Cheng

2015-01-01

Stem cell research has been extensively explored worldwide to enhance human health in medical setting. Nevertheless, there is currently no full understanding of the stem cell knowledge and attitude levels among student nurses in Malaysia. This study aimed to assess the level of stem cell knowledge, attitude toward stem cell application in medicine, and its association with years of education, among Universiti Sains Malaysia (USM) undergraduate nursing students. A cross-sectional study (n = 88) was conducted using self-administered questionnaire consisted of demographic information, stem cells knowledge and attitude statements. Data was analysed using Statistical Package Social Software 20.0. The majority of participants (92%) had moderate knowledge score about stem cells. Many students (33%) worried that stem cell application might cause a harm to humanity yet had a positive (76.1%) attitude towards its therapeutic potential (45.5%). Poor correlation between knowledge and attitude (r = 0.08) indicated that acceptance towards stem cell is not solely based on the knowledge level but also on other factors including religion and culture. Therefore, this study suggests that various educational programs on stem cell should be implemented considering the religion, cultural, social, and behavioural determinants in the population to improve stem cell knowledge and encourage a more positive attitude towards stem cells in medicine among these nursing students.

Stem cell motility enables a density-dependent rate of fate commitment during scaled resizing of adult organs

NASA Astrophysics Data System (ADS)

Du, Xinxin; O'Brien, Lucy; Riedel-Kruse, Ingmar

Many adult organs grow or shrink to accommodate fluctuating levels of physiological demand. Specifically, the intestine of the fruit fly (the midgut) expands four-fold in the number of mature cells and, proportionally, the number of stem cells when the fly eats. However, the cellular behaviors that give rise to this stem scaling are not well-understood. Here we present a biophysical model of the adult fly midgut. A set of differential equations can recapitulate the physiological kinetics of cells during midgut growth and shrinkage as long as the rate of stem cell fate commitment depends on the stem cell number density in the tissue. To elucidate the source of this dependence, we model the tissue in a 2D simulation with soft spheres, where stem cells choose fate commitment through Delta-Notch pathway interactions with other stem cells, a known process in fly midguts. We find that as long as stem cells exhibit a large enough amplitude of random motion through the tissue (`stem cell motility'), and explore a large enough `territory' in their lifetime, stem cell scaling can occur. These model observations are confirmed through in vivo live-imaging, where we indeed see that stem cells are motile in the fly midgut.

Ethics and Policy Issues for Stem Cell Research and Pulmonary Medicine

PubMed Central

Lowenthal, Justin

2015-01-01

Stem cell research and related initiatives in regenerative medicine, cell-based therapy, and tissue engineering have generated considerable scientific and public interest. Researchers are applying stem cell technologies to chest medicine in a variety of ways: using stem cells as models for drug discovery, testing stem cell-based therapies for conditions as diverse as COPD and cystic fibrosis, and producing functional lung and tracheal tissue for physiologic modeling and potential transplantation. Although significant scientific obstacles remain, it is likely that stem cell-based regenerative medicine will have a significant clinical impact in chest medicine. However, stem cell research has also generated substantial controversy, posing a variety of ethical and regulatory challenges for research and clinical practice. Some of the most prominent ethical questions related to the use of stem cell technologies in chest medicine include (1) implications for donors, (2) scientific prerequisites for clinical testing and use, (3) stem cell tourism, (4) innovation and clinical use of emerging stem cell-based interventions, (5) responsible translation of stem cell-based therapies to clinical use, and (6) appropriate and equitable access to emerging therapies. Having a sense of these issues should help to put emerging scientific advances into appropriate context and to ensure the responsible clinical translation of promising therapeutics. PMID:25732448

A Survey of Italian Physicians' Opinion about Stem Cells Research: What Doctors Prefer and What the Law Requires

PubMed Central

Frati, Paola; Pacchiarotti, Arianna; D'Errico, Stefano

2014-01-01

To evaluate the Italian physicians' knowledge/information level about the therapeutic potential of stem cells, the research choice between embryonic and cordonal stem cells, and the preference between autologous and heterologous storage of cordonal stem cells, we performed a national survey. The questionnaire—distributed to 3361 physicians—involved physicians of different religious orientations and of different medical specialities. Most of the physicians involved (67%) were Catholics, and the majority were gynaecologists and paediatricians (43%) who are mainly in charge to inform future mothers about the possibility of cordonal stem cells conservation. The majority of the physicians interviewed do not have specific knowledge about stem cells (59%), most of them having only generic information (92%). The largest part of physicians prefer to use umbilical cord blood cells rather than embryonic stem cells. Nevertheless, a large percentage of physicians were in favour of embryo research, especially when embryos are supernumerary (44% versus 34%). Eighty-seven % of the physicians interviewed proved to have a general knowledge about stem cells and believe in their therapeutic potential. They prefer research on cordonal stem cells rather than on embryo stem cells. Although they are in favour of heterologous stem cells donation, they still prefer cryopreservation for personal use. PMID:24877099

Ethics and policy issues for stem cell research and pulmonary medicine.

PubMed

Lowenthal, Justin; Sugarman, Jeremy

2015-03-01

Stem cell research and related initiatives in regenerative medicine, cell-based therapy, and tissue engineering have generated considerable scientific and public interest. Researchers are applying stem cell technologies to chest medicine in a variety of ways: using stem cells as models for drug discovery, testing stem cell-based therapies for conditions as diverse as COPD and cystic fibrosis, and producing functional lung and tracheal tissue for physiologic modeling and potential transplantation. Although significant scientific obstacles remain, it is likely that stem cell-based regenerative medicine will have a significant clinical impact in chest medicine. However, stem cell research has also generated substantial controversy, posing a variety of ethical and regulatory challenges for research and clinical practice. Some of the most prominent ethical questions related to the use of stem cell technologies in chest medicine include (1) implications for donors, (2) scientific prerequisites for clinical testing and use, (3) stem cell tourism, (4) innovation and clinical use of emerging stem cell-based interventions, (5) responsible translation of stem cell-based therapies to clinical use, and (6) appropriate and equitable access to emerging therapies. Having a sense of these issues should help to put emerging scientific advances into appropriate context and to ensure the responsible clinical translation of promising therapeutics.

Propagation of human spermatogonial stem cells in vitro.

PubMed

Sadri-Ardekani, Hooman; Mizrak, Sefika C; van Daalen, Saskia K M; Korver, Cindy M; Roepers-Gajadien, Hermien L; Koruji, Morteza; Hovingh, Suzanne; de Reijke, Theo M; de la Rosette, Jean J M C H; van der Veen, Fulco; de Rooij, Dirk G; Repping, Sjoerd; van Pelt, Ans M M

2009-11-18

Young boys treated with high-dose chemotherapy are often confronted with infertility once they reach adulthood. Cryopreserving testicular tissue before chemotherapy and autotransplantation of spermatogonial stem cells at a later stage could theoretically allow for restoration of fertility. To establish in vitro propagation of human spermatogonial stem cells from small testicular biopsies to obtain an adequate number of cells for successful transplantation. Study performed from April 2007 to July 2009 using testis material donated by 6 adult men who underwent orchidectomy as part of prostate cancer treatment. Testicular cells were isolated and cultured in supplemented StemPro medium; germline stem cell clusters that arose were subcultured on human placental laminin-coated dishes in the same medium. Presence of spermatogonia was determined by reverse transcriptase polymerase chain reaction and immunofluorescence for spermatogonial markers. To test for the presence of functional spermatogonial stem cells in culture, xenotransplantation to testes of immunodeficient mice was performed, and migrated human spermatogonial stem cells after transplantation were detected by COT-1 fluorescence in situ hybridization. The number of colonized spermatogonial stem cells transplanted at early and later points during culture were counted to determine propagation. Propagation of spermatogonial stem cells over time. Testicular cells could be cultured and propagated up to 15 weeks. Germline stem cell clusters arose in the testicular cell cultures from all 6 men and could be subcultured and propagated up to 28 weeks. Expression of spermatogonial markers on both the RNA and protein level was maintained throughout the entire culture period. In 4 of 6 men, xenotransplantation to mice demonstrated the presence of functional spermatogonial stem cells, even after prolonged in vitro culture. Spermatogonial stem cell numbers increased 53-fold within 19 days in the testicular cell culture and increased 18,450-fold within 64 days in the germline stem cell subculture. Long-term culture and propagation of human spermatogonial stem cells in vitro is achievable.

Genetic and epigenetic profiling of CLL disease progression reveals limited somatic evolution and suggests a relationship to memory-cell development.

PubMed

Smith, E N; Ghia, E M; DeBoever, C M; Rassenti, L Z; Jepsen, K; Yoon, K-A; Matsui, H; Rozenzhak, S; Alakus, H; Shepard, P J; Dai, Y; Khosroheidari, M; Bina, M; Gunderson, K L; Messer, K; Muthuswamy, L; Hudson, T J; Harismendy, O; Barrett, C L; Jamieson, C H M; Carson, D A; Kipps, T J; Frazer, K A

2015-04-10

We examined genetic and epigenetic changes that occur during disease progression from indolent to aggressive forms of chronic lymphocytic leukemia (CLL) using serial samples from 27 patients. Analysis of DNA mutations grouped the leukemia cases into three categories: evolving (26%), expanding (26%) and static (47%). Thus, approximately three-quarters of the CLL cases had little to no genetic subclonal evolution. However, we identified significant recurrent DNA methylation changes during progression at 4752 CpGs enriched for regions near Polycomb 2 repressive complex (PRC2) targets. Progression-associated CpGs near the PRC2 targets undergo methylation changes in the same direction during disease progression as during normal development from naive to memory B cells. Our study shows that CLL progression does not typically occur via subclonal evolution, but that certain CpG sites undergo recurrent methylation changes. Our results suggest CLL progression may involve developmental processes shared in common with the generation of normal memory B cells.

“Inclonals”

PubMed Central

Hakim, Rahely

2009-01-01

Full-length antibodies and antibodies that ferry a cargo to target cells are desired biopharmaceuticals. We describe the production of full-length IgGs and IgG-toxin fusion proteins in E. coli. In the presented examples of anti CD30 and anti EGF-receptor antibodies, the antibody heavy and light chains or toxin fusions thereof were expressed in separate bacterial cultures, where they accumulated as insoluble inclusion bodies. Following refolding and purification, high yields (up to 50 mg/L of shake flask culture) of highly purified (>90%) full-length antibodies and antibody-toxin fusions were obtained. The bacterially produced antibodies, named “Inclonals,” equaled the performance of the same IgGs that were produced using conventional mammalian cell culture in binding properties as well as in cell killing potency. The rapid and cost effective IgG production process and the high quality of the resultant product may make the bacterial production of full-length IgG and IgG-drug fusion proteins an attractive option for antibody production and a significant contribution to recombinant antibody technology. PMID:20065645