Nurse's A-Phase Material Enhance Adhesion, Growth and Differentiation of Human Bone Marrow-Derived Stromal Mesenchymal Stem Cells.
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ABSTRACT:
SUBMITTER: Rabadan-Ros R
PROVIDER: S-EPMC5506906 | biostudies-other | 2017 Mar
REPOSITORIES: biostudies-other
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Project description:The precise predictions of the differentiation direction and potential of mesenchymal stromal cells (MSCs) are an important key to the success of regenerative medicine. The expression levels of fate-determining genes may provide tools for predicting differentiation potential. The expression levels of 95 candidate marker genes and glycosaminoglycan (GAG) contents after chondrogenic induction in 10 undifferentiated ilium and 5 jaw MSC cultures were determined, and their correlations were analyzed. The expression levels of eight genes before the induction of chondrogenic MSC differentiation were significantly correlated with the GAG levels after induction. Based on correlation patterns, the eight genes were classified into two groups: group 1 genes (AURKB, E2F1, CDKN2D, LIF, and ACLY), related to cell cycle regulation, and group 2 genes (CD74, EFEMP1, and TGM2), involved in chondrogenesis. The expression levels of the group 2 genes were significantly correlated with the ages of the cell donors. The expression levels of CDKN2D, CD74, and TGM2 were >10-fold higher in highly potent MSCs (ilium MSCs) than in MSCs with limited potential (jaw MSCs). Three-dimensional (3D) scatter plot analyses of the expression levels of these genes showed reduced variability between donors and confirmed predictive potential. These data suggest that group 2 genes are involved in age-dependent decreases in the chondrogenic differentiation potential of MSCs, and combined 3D analyses of the expression profiles of three genes, including two group 2 genes, were predictive of MSC differentiation potential.
| S-EPMC6199884 | biostudies-literature
Project description:Purpose:This study is aimed at investigating the phenotype, differentiation potential, immunomodulatory properties, and responsiveness of saphenous vein vessel wall-derived mesenchymal stromal cells (SV-MSCs) to various TLR ligands and proinflammatory cytokines, as well as comparing their features to those of their bone marrow-derived counterparts (BM-MSCs). Methods:SV-MSCs were isolated by enzymatic digestion of the saphenous vein vessel wall. Phenotype analysis was carried out by flow cytometry and microscopy, whereas adipogenic, chondrogenic, and osteogenic differentiation potentials were tested in in vitro assays. For comparative analysis, the expression of different stemness, proliferation, and differentiation-related genes was determined by Affymetrix gene array. To compare the immunomodulatory properties of SV-MSCs and BM-MSCs, mixed lymphocyte reaction was applied. To investigate their responses to various activating stimuli, MSCs were treated with TLR ligands (LPS, PolyI:C) or proinflammatory cytokines (TNF?, IL-1?, IFN?), and the expression of various early innate immune response-related genes was assessed by qPCR, while secretion of selected cytokines and chemokines was measured by ELISA. Results:The isolated SV-MSCs were able to differentiate into bone, fat, and cartilage cells/direction in vitro. SV-MSCs expressed the most important MSC markers (CD29, CD44, CD73, CD90, and CD105) and shared almost identical phenotypic characteristics with BM-MSCs. Their gene expression pattern and activation pathways were close to those of BM-MSCs. SV-MSCs showed better immunosuppressive activity inhibiting phytohemagglutinin-induced T lymphocyte proliferation in vitro than BM-MSCs. Cellular responses to treatments mimicking inflammatory conditions were comparable in the bone marrow- and saphenous vein-derived MSCs. Namely, similar to BM-MSCs, SV-MSCs secreted increased amount of IL-6 and IL-8 after 12- or 24-hour treatment with LPS, PolyI:C, TNF?, or IL-1?, compared to untreated controls. Interestingly, a different CXCL-10/IP-10 secretion pattern could be observed under inflammatory conditions in the two types of MSCs. Conclusion:Based on our results, cells isolated from saphenous vein vessel wall fulfilled the ISCT's (International Society for Cellular Therapy) criteria for multipotent mesenchymal stromal cells, and no significant differences in the phenotype, gene expression pattern, and responsiveness to inflammatory stimuli could be observed between BM-MSCs and SV-MSCs, while the latter cells have more potent immunosuppressive activity in vitro. Further functional assays have to be performed to reveal whether SV-MSCs could be useful for certain regenerative therapeutic applications or tissue engineering purposes.
| S-EPMC7596426 | biostudies-literature
Project description:BackgroundThe skeletal muscle reconstruction occurs thanks to unipotent stem cells, i.e., satellite cells. The satellite cells remain quiescent and localized between myofiber sarcolemma and basal lamina. They are activated in response to muscle injury, proliferate, differentiate into myoblasts, and recreate myofibers. The stem and progenitor cells support skeletal muscle regeneration, which could be disturbed by extensive damage, sarcopenia, cachexia, or genetic diseases like dystrophy. Many lines of evidence showed that the level of oxygen regulates the course of cell proliferation and differentiation.MethodsIn the present study, we analyzed hypoxia impact on human and pig bone marrow-derived mesenchymal stromal cell (MSC) and mouse myoblast proliferation, differentiation, and fusion. Moreover, the influence of the transplantation of human bone marrow-derived MSCs cultured under hypoxic conditions on skeletal muscle regeneration was studied.ResultsWe showed that bone marrow-derived MSCs increased VEGF expression and improved myogenesis under hypoxic conditions in vitro. Transplantation of hypoxia preconditioned bone marrow-derived MSCs into injured muscles resulted in the improved cell engraftment and formation of new vessels.ConclusionsWe suggested that SDF-1 and VEGF secreted by hypoxia preconditioned bone marrow-derived MSCs played an essential role in cell engraftment and angiogenesis. Importantly, hypoxia preconditioned bone marrow-derived MSCs more efficiently engrafted injured muscles; however, they did not undergo myogenic differentiation.
| S-EPMC8351116 | biostudies-literature
Project description:BACKGROUND AND PURPOSE: Caffeine is consumed extensively in Europe and North America. As a risk factor for osteoporosis, epidemiological studies have observed that caffeine can decrease bone mineral density, adversely affect calcium absorption and increase the risk of bone fracture. However, the exact mechanisms have not been fully investigated. Here, we examined the effects of caffeine on the viability and osteogenesis of rat bone marrow-derived mesenchymal stromal cells (rBMSCs). EXPERIMENTAL APPROACH: Cell viability, apoptosis and necrosis were quantified using thymidine incorporation and flow cytometry. Sequential gene expressions in osteogenic process were measured by real-time PCR. cAMP, alkaline phosphatase and osteocalcin were assessed by immunoassay, spectrophotometry and radioimmunoassay, respectively. Mineralization was determined by calcium deposition. KEY RESULTS: After treating BMSCs with high caffeine concentrations (0.1-1mM), their viability decreased in a concentration-dependent manner. This cell death was primarily due to necrosis and, to a small extent, apoptosis. Genes and protein sequentially expressed in osteogenesis, including Cbfa1/Runx2, collagen I, alkaline phosphatase and its protein, were significantly downregulated except for osteocalcin and its protein. Moreover, caffeine inhibited calcium deposition in a concentration- and time-dependent manner, but increased intracellular cAMP in a concentration-dependent manner. CONCLUSIONS AND IMPLICATIONS: By suppressing the commitment of BMSCs to the osteogenic lineage and selectively inhibiting gene expression, caffeine downregulated some important events in osteogenesis and ultimately affected bone mass.
| S-EPMC3010566 | biostudies-literature
Project description:Guanylate Binding Proteins (GBPs) are a group of cytokine-inducible large guanosine triphosphatase. Previous studies have shown high expression of GBP1 in circulating monocytes of premenopausal subjects was correlated to extremely low peak bone mass, which is considered as an important determinant of osteoporosis. However, whether GBPs play a role in regulation of osteogenesis of mesenchymal stromal cells (MSCs) remains largely unknown. In the present study, we found that mRNA expression of GBP1 was highest among all the GBPs, and it was dramatically downregulated during osteogenic differentiation of human MSCs derived from bone marrow (hBM-MSCs). While siRNA-mediated knockdown of GBP1 promoted osteogenesis, overexpression of GBP1 suppressed osteogenesis of hBM-MSCs. Furthermore, we found GBP1 is required for expression of indoleamine 2,3 dioxygenase (IDO), Interleukin 6 (IL-6) and IL-8 induced by treatment with Interferon-? (IFN-?). Depletion of GBP1 rescued the inhibited osteogenesis induced by IFN-? treatment, at least in part. Collectively, our findings indicate GBP1 inhibits osteogenic differentiation of MSCs, and inhibition of GBP1 expression may prevent development of osteoporosis and facilitate MSC-based bone regeneration.
| S-EPMC5773562 | biostudies-literature
Project description:The authors' previous study demonstrated that miR?128 may exert an inhibitory effect on the osteogenic differentiation of bone marrow?derived mesenchymal stem cells (BM?MSCs), but its downstream mechanisms remain to be elucidated. The aim of the present study was to investigate the microRNA (miRNA/miR) and mRNA profiles of differentiated and undifferentiated BM?MSCs and explore new downstream targets for miR?128. The sequencing datasets of GSE107279 (miRNA) and GSE112318 (mRNA) were downloaded from the Gene Expression Omnibus database. The differentially expressed miRNAs (DEMs) and genes (DEGs) were identified using the DESeq2 method. The target genes of DEMs were predicted by the miRwalk 2.0 database. The hub target genes of miR?128 were screened by constructing the protein?protein interaction (PPI) network and module analysis. The expression levels of miR?128 and crucial target genes were validated by reverse transcription?quantitative (RT?q) PCR before or after transfection of miR?128 mimics to BM?MSCs. The miRNA expression profile analysis identified miR?128 as one of the significantly downregulated DEMs (total 338) in differentiated BM?MSCs compared with the undifferentiated control. A total of 103 predicted target genes of miR?128?3p were overlapped with upregulated DEGs. By calculating the topological properties of each protein in the PPI network, 6 upregulated genes (KIT, NTRK2, YWHAB, GAB1, AXIN1 and RUNX1; fold change was the highest for NTRK2) were considered to be hub genes. Of these, 4 were enriched in module 4 (RUNX1, KIT, GAB1 and AXIN1; RUNX1 was particularly crucial as it can interact with the others), while one was enriched in module 7 (YWHAB). The expression levels of miR?128 and these 6 target genes during the osteogenic differentiation were experimentally confirmed by RT?qPCR. In addition, the expression levels of these 6 genes were significantly reversed after transfection of miR?128?3p mimics into rat BM?MSCs compared with the miR?control group. These findings indicated that miR?128?3p may inhibit the osteoblast differentiation of BM?MSCs by downregulation of these 6 genes, particularly RUNX1, YWHAB and NTRK2.
| S-EPMC7646956 | biostudies-literature
Project description:BackgroundOsteogenesis Imperfecta (OI) (OMIM %259450) is a heterogeneous group of inherited disorders characterized by increased bone fragility, with clinical severity ranging from mild to lethal. The majority of OI cases are caused by mutations in COL1A1 or COL1A2. Bruck Syndrome (BS) is a further recessively-inherited OI-like phenotype in which bone fragility is associated with the unusual finding of pterygia and contractures of the large joints. Notably, several studies have failed to show any abnormalities in the biosynthesis of collagen 1 in BS patientes. Evidence was obtained for a specific defect of the procollagen telopeptide lysine hydroxylation in BS, whereas mutations in the gene PLOD2 have been identified. Recently, several studies described FKBP10 mutations in OI-like and BS patients, suggesting that FKBP10 is a bonafide BS locus.MethodsWe analyzed the coding region and intron/exon boundaries of COL1A1, COL1A2, PLOD2 and FKBP10 genes by sequence analysis using an ABI PRISM 3130 automated sequencer and Big Dye Terminator Sequencing protocol. Mononuclear cells obtained from the bone marrow of BS, OI patients and healthy donors were cultured and osteogenic differentiation was induced. The gene expression of osteoblast specific markers were also evaluated during the osteoblastic differentiation of mesenchymal stem cell (MSC) by qRT-PCR using an ABI7500 Sequence Detection System.ResultsNo mutations in COL1A1, COL1A2 or PLOD2 were found in BS patient. We found a homozygous 1-base-pair duplication (c.831dupC) that is predicted to produce a translational frameshift mutation and a premature protein truncation 17 aminoacids downstream (p.Gly278ArgfsX95). The gene expression of osteoblast specific markers BGLAP, COL1A1, MSX2, SPARC and VDR was evaluated by Real Time RT-PCR during differentiation into osteoblasts and results showed similar patterns of osteoblast markers expression in BS and healthy controls. On the other hand, when compared with OI patients, the expression pattern of these genes was found to be different.ConclusionsOur work suggests that the gene expression profiles observed during mesenchymal stromal cell differentiation into osteoblast are distinct in BS patients as compared to OI patients. The present study shows for the first time that genes involved in osteogenesis are differentially expressed in BS and OI patients.
| S-EPMC4857408 | biostudies-literature
Project description:Bone marrow-derived mesenchymal stromal cells (hMSCs) are capable of differentiating into the osteogenic lineage, and for osteogenic differentiation, mechanical loading is a relevant stimulus. Mechanotransduction leads to the formation of second messengers such as cAMP, cGMP, or Ca2+ influx resulting in the activation of transcription factors mediating gene regulation. The second messengers cAMP and cGMP are degraded by phosphodiesterase isoenzymes (PDE), but the role of these enzymes during osteogenic differentiation or mechanotransduction remains unclear. Here, we focused on the isoenzyme phosphodiesterase 10A (PDE10A) and its role during osteogenic commitment and mechanotransduction. We observed a time-dependent decrease of PDE10A expression in hMSC undergoing differentiation towards the osteogenic lineage. PDE10A inhibition by papaverine diminished osteogenic differentiation. While applying mechanical strain via cyclic stretching of hMSCs led to an upregulation of PDE10A gene expression, inhibition of PDE10A using the drug papaverine repressed expression of mechanoresponsive genes. We conclude that PDE10A is a modulator of osteogenic differentiation as well as mechanotransduction in hMSCs. Our data further suggests that the relative increase of cAMP, rather than the absolute cAMP level, is a key driver of the observed effects.
| S-EPMC7294361 | biostudies-literature
Project description:UnlabelledStem cell therapy is an emerging alternative therapeutic or disease-modifying strategy for amyotrophic lateral sclerosis (ALS). The aim of this open-label phase I clinical trial was to evaluate the safety of two repeated intrathecal injections of autologous bone marrow (BM)-derived mesenchymal stromal cells (MSCs) in ALS patients. Eight patients with definite or probable ALS were enrolled. After a 3-month lead-in period, autologous MSCs were isolated two times from the BM at an interval of 26 days and were then expanded in vitro for 28 days and suspended in autologous cerebrospinal fluid. Of the 8 patients, 7 received 2 intrathecal injections of autologous MSCs (1 × 10(6) cells per kg) 26 days apart. Clinical or laboratory measurements were recorded to evaluate the safety 12 months after the first MSC injection. The ALS Functional Rating Scale-Revised (ALSFRS-R), the Appel ALS score, and forced vital capacity were used to evaluate the patients' disease status. One patient died before treatment and was withdrawn from the study. With the exception of that patient, no serious adverse events were observed during the 12-month follow-up period. Most of the adverse events were self-limited or subsided after supportive treatment within 4 days. Decline in the ALSFRS-R score was not accelerated during the 6-month follow-up period. Two repeated intrathecal injections of autologous MSCs were safe and feasible throughout the duration of the 12-month follow-up period.SignificanceStem cell therapy is an emerging alternative therapeutic or disease-modifying strategy for amyotrophic lateral sclerosis (ALS). To the authors' best knowledge, there are no clinical trials to evaluate the safety of repeated intrathecal injections of autologous bone marrow mesenchymal stromal cells in ALS. After the clinical trial (phase I/II) was conducted, the stem cell (HYNR-CS, NEURONATA-R) was included in the revision of the regulations on orphan drug designation (number 160; December 31, 2013) and approved as a New Drug Application (Department of Cell and Gene Therapy 233; July 30, 2014) by the Korean Food and Drug Administration. The phase II trial is expected to be reported later.
| S-EPMC4449093 | biostudies-literature
Project description:There has been considerable interest in the generation of functional mesenchymal stromal cell (MSC) preparations from induced pluripotent stem cells (iPSCs) and this is now regarded as a potential source of unlimited, standardized, high-quality cells for therapeutic applications in regenerative medicine. Although iMSCs meet minimal criteria for defining MSCs in terms of marker expression, there are substantial differences in terms of trilineage potential, specifically a marked reduction in chondrogenic and adipogenic propensity in iMSCs compared with bone marrow-derived (BM) MSCs. To reveal the cellular basis underlying these differences, we conducted phenotypic, functional, and genetic comparisons between iMSCs and BM-MSCs. We found that iMSCs express very high levels of both KDR and MSX2 compared with BM-MSCs. In addition, BM-MSCs had significantly higher levels of PDGFRα. These distinct gene expression profiles were maintained during culture expansion, suggesting that prepared iMSCs are more closely related to vascular progenitor cells (VPCs). Although VPCs can differentiate along the chondrogenic, osteogenic, and adipogenic pathways, they require different inductive conditions compared with BM-MSCs. These observations suggest to us that iMSCs, based on current widely used preparation protocols, do not represent a true alternative to primary MSCs isolated from BM. Furthermore, this study highlights the fact that high levels of expression of typical MSC markers such as CD73, CD90, and CD105 are insufficient to distinguish MSCs from other mesodermal progenitors in differentiated induced pluripotent stem cell cultures. Stem Cells 2019;37:754-765.
| S-EPMC6591688 | biostudies-literature