Cumulative evidence suggests that O-linkedβ-N-acetylglucosaminylation(OGlcNAcylation)plays an important regulatory role in pathophysiological processes.Although the regulatory mechanisms of O-GlcNAcylation in tumors ...Cumulative evidence suggests that O-linkedβ-N-acetylglucosaminylation(OGlcNAcylation)plays an important regulatory role in pathophysiological processes.Although the regulatory mechanisms of O-GlcNAcylation in tumors have been gradually elucidated,the potential mechanisms of O-GlcNAcylation in bone metabolism,particularly,in the osteogenic differentiation of bone marrow mesenchymal stromal cells(BMSCs)remains unexplored.In this study,the literature related to O-GlcNAcylation and BMSC osteogenic differentiation was reviewed,assuming that it could trigger more scholars to focus on research related to OGlcNAcylation and bone metabolism and provide insights into the development of novel therapeutic targets for bone metabolism disorders such as osteoporosis.展开更多
BACKGROUND Bone marrow mesenchymal stromal cells(BMSCs)are the commonly used seed cells in tissue engineering.Aryl hydrocarbon receptor(AhR)is a transcription factor involved in various cellular processes.However,the ...BACKGROUND Bone marrow mesenchymal stromal cells(BMSCs)are the commonly used seed cells in tissue engineering.Aryl hydrocarbon receptor(AhR)is a transcription factor involved in various cellular processes.However,the function of constitutive AhR in BMSCs remains unclear.AIM To investigate the role of AhR in the osteogenic and macrophage-modulating potential of mouse BMSCs(mBMSCs)and the underlying mechanism.METHODS Immunochemistry and immunofluorescent staining were used to observe the expression of AhR in mouse bone marrow tissue and mBMSCs.The overexpression or knockdown of AhR was achieved by lentivirus-mediated plasmid.The osteogenic potential was observed by alkaline phosphatase and alizarin red staining.The mRNA and protein levels of osteogenic markers were detected by quantitative polymerase chain reaction(qPCR)and western blot.After coculture with different mBMSCs,the cluster of differentiation(CD)86 and CD206 expressions levels in RAW 264.7 cells were analyzed by flow cytometry.To explore the underlying molecular mechanism,the interaction of AhR with signal transducer and activator of transcription 3(STAT3)was observed by co-immunoprecipitation and phosphorylation of STAT3 was detected by western blot.RESULTS AhR expressions in mouse bone marrow tissue and isolated mBMSCs were detected.AhR overexpression enhanced the osteogenic potential of mBMSCs while AhR knockdown suppressed it.The ratio of CD86+RAW 264.7 cells cocultured with AhR-overexpressed mBMSCs was reduced and that of CD206+cells was increased.AhR directly interacted with STAT3.AhR overexpression increased the phosphorylation of STAT3.After inhibition of STAT3 via stattic,the promotive effects of AhR overexpression on the osteogenic differentiation and macrophage-modulating were partially counteracted.CONCLUSION AhR plays a beneficial role in the regenerative potential of mBMSCs partially by increasing phosphorylation of STAT3.展开更多
Breast cancer is the predominant form of carcinoma among women worldwide,with 70%of advanced patients developing bone metastases,with a high mortality rate.In this sense,the bone marrow(BM)mesenchymal stem/stromal cel...Breast cancer is the predominant form of carcinoma among women worldwide,with 70%of advanced patients developing bone metastases,with a high mortality rate.In this sense,the bone marrow(BM)mesenchymal stem/stromal cells(MSCs)are critical for BM/bone homeostasis,and failures in their functionality,transform the BM into a premetastatic niche(PMN).We previously found that BM-MSCs from advanced breast cancer patients(BCPs,infiltrative ductal carcinoma,stage III-B)have an abnormal profile.This work aims to study some of the metabolic and molecular mechanisms underlying MSCs shift from a normal to an abnormal profile in this group of patients.A comparative analysis was undertaken,which included self-renewal capacity,morphology,proliferation capacity,cell cycle,reactive oxygen species(ROS)levels,and senescence-associatedβ‑galactosidase(SA‑β‑gal)staining of BMderived MSCs isolated from 14 BCPs and 9 healthy volunteers(HVs).Additionally,the expression and activity of the telomerase subunit TERT,as well as telomere length,were measured.Expression levels of pluripotency,osteogenic,and osteoclastogenic genes(OCT-4,SOX-2,M-CAM,RUNX-2,BMP-2,CCL-2,M-CSF,and IL-6)were also determined.The results showed that MSCs from BCPs had reduced,self-renewal and proliferation capacity.These cells also exhibited inhibited cell cycle progression and phenotypic changes,such as an enlarged and flattened appearance.Additionally,there was an increase in ROS and senescence levels and a decrease in the functional capacity of TERT to preserve telomere length.We also found an increase in pro-inflammatory/pro-osteoclastogenic gene expression and a decrease in pluripotency gene expression.We conclude that these changes could be responsible for the abnormal functional profile that MSCs show in this group of patients.展开更多
BACKGROUND The low survival rate of mesenchymal stem cells(MSCs)caused by anoikis,a form of apoptosis,limits the therapeutic efficacy of MSCs.As a proapoptotic molecule,mammalian Ste20-like kinase 1(Mst1)can increase ...BACKGROUND The low survival rate of mesenchymal stem cells(MSCs)caused by anoikis,a form of apoptosis,limits the therapeutic efficacy of MSCs.As a proapoptotic molecule,mammalian Ste20-like kinase 1(Mst1)can increase the production of reactive oxygen species(ROS),thereby promoting anoikis.Recently,we found that Mst1 inhibition could protect mouse bone marrow MSCs(mBMSCs)from H 2 O 2-induced cell apoptosis by inducing autophagy and reducing ROS production.However,the influence of Mst1 inhibition on anoikis in mBMSCs remains unclear.AIM To investigate the mechanisms by which Mst1 inhibition acts on anoikis in isolated mBMSCs.METHODS Poly-2-hydroxyethyl methacrylate-induced anoikis was used following the silencing of Mst1 expression by short hairpin RNA(shRNA)adenovirus transfection.Integrin(ITGs)were tested by flow cytometry.Autophagy and ITGα5β1 were inhibited using 3-methyladenine and small interfering RNA,respe-ctively.The alterations in anoikis were measured by Terminal-deoxynucleoitidyl Transferase Mediated Nick End Labeling and anoikis assays.The levels of the anoikis-related proteins ITGα5,ITGβ1,and phospho-focal adhesion kinase and the activation of caspase 3 and the autophagy-related proteins microtubules associated protein 1 light chain 3 II/I,Beclin1 and p62 were detected by Western blotting.RESULTS In isolated mBMSCs,Mst1 expression was upregulated,and Mst1 inhibition significantly reduced cell apoptosis,induced autophagy and decreased ROS levels.Mechanistically,we found that Mst1 inhibition could upregulate ITGα5 and ITGβ1 expression but not ITGα4,ITGαv,or ITGβ3 expression.Moreover,autophagy induced by upregulated ITGα5β1 expression following Mst1 inhibition played an essential role in the protective efficacy of Mst1 inhibition in averting anoikis.CONCLUSION Mst1 inhibition ameliorated autophagy formation,increased ITGα5β1 expression,and decreased the excessive production of ROS,thereby reducing cell apoptosis in isolated mBMSCs.Based on these results,Mst1 inhibition may provide a promising strategy to overcome anoikis of implanted MSCs.展开更多
In order to elucidate the action of La3+ on bone metabolism,effects of La3+ on the osteogenic and adipogenic differentiation of pri-mary mouse bone marrow stromal cells(BMSCs) were studied by 3-(4,5-dimethylthiazol-2-...In order to elucidate the action of La3+ on bone metabolism,effects of La3+ on the osteogenic and adipogenic differentiation of pri-mary mouse bone marrow stromal cells(BMSCs) were studied by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide(MTT) test,alkaline phosphatase(ALP) activity measurement,mineralized function,oil red O stain and measurement.The results showed that La3+ pro-moted the proliferation of BMSCs except at 1×10-10 and 1×10-6 mol/L.The effect of La3+ on the osteogenic differentiation depended on con-centrations at the 7th day,but the osteogenic differentiation was inhibited at any concentration at the 14th day.La3+ promoted the formation of mineralized matrix nodules except at 1×10-8 and 1×10-5 mol/L.La3+ inhibited adipogenic differentiation except at 1×10-10 and 1×10-7 mol/L at the 10th day,and inhibited adipogenic differentiation except at 1×10-9 mol/L at the 16th day.These findings suggested that La3+ might have protective effect on bone at appropriate dose and time.This would be valuable for better understanding the mechanism of the effect of La3+ on bone metabolism.展开更多
One of the main target organs for the lanthanides(Ln) is bone. Previous studies revealed that ytterbium(Yb) produced damage to the skeletal system in vivo. But the effects of Yb3+ on bone marrow stromal cells(BMSCs) i...One of the main target organs for the lanthanides(Ln) is bone. Previous studies revealed that ytterbium(Yb) produced damage to the skeletal system in vivo. But the effects of Yb3+ on bone marrow stromal cells(BMSCs) in vitro had not been reported. In this paper, cell viability, apoptosis, mitochondrial membrane potential(MMP), reactive oxygen species(ROS) and lactate dehydrogenase(LDH) were measured in order to study the effects of Yb3+ on BMSCs. The results indicated that Yb3+ displayed a slight positive effect on the BMSCs viability at concentrations of 1×10–6, 1×10–5, and 1×10–4 mol/L, but turned to decrease the viability of BMSCs at the highest concentration of 1×10–3 mol/L for 24, 48 and 72 h. Yb3+ at 1×10–3 mol/L promoted apoptosis of BMSCs, increased the levels of ROS and LDH, and decreased MMP in BMSCs. It suggested that the precipitate of Yb PO4 might decrease the viability of BMSCs. Yb3+ induced the apoptosis of BMSCs via mitochondrial pathway. The results might be useful for more rational application of Yb-based compounds in the future.展开更多
AIM:To investigate the malignant potential of hepatic stem cells derived from the bone marrow stromal cells (BMSCs) in a mouse model of chemical hepatocarcino- genesis. METHODS:BMSCs from male BALB/c mice were harvest...AIM:To investigate the malignant potential of hepatic stem cells derived from the bone marrow stromal cells (BMSCs) in a mouse model of chemical hepatocarcino- genesis. METHODS:BMSCs from male BALB/c mice were harvested and cultured, then transplanted into female syngenic BALB/ c mice via portal vein. Hepato-carcinogenesis was induced by 6 mo of treatment with diethylnitrosamine (DEN). Six months later, the liver was removed from each treated mouse and evaluated by immunohistochemistry and fluorescence in situ hybridization (FISH). RESULTS:Twenty-six percent of recipient mice survived and developed multiple hepatocellular carcinomas (HCCs). Immunohistochemically, HCC expressed placental form of glutathione-S-transferase (GST-P) and α-fetoprotein, but did not express cytokeratin 19. Y chromosome positive hepatocytes were detected by fluorescent in situ hybridization (FISH) in the liver of mice treated with DEN after BMSCs transplantation while no such hepatocytes were identified in the liver of mice not treated with DEN. No HCC was positive for the Y chromosome by FISH. CONCLUSION:Hepatic stem cells derived from the bone marrow stromal cells have a low malignant potential in our mouse model of chemical hepatocarcingenesis.展开更多
Transplantation of bone marrow stromal cells(BMSCs) enhanced the outgrowth of regenerating axons and promoted locomotor improvements of rats with spinal cord injury(SCI).BMSCs did not survive long-term,disappearing fr...Transplantation of bone marrow stromal cells(BMSCs) enhanced the outgrowth of regenerating axons and promoted locomotor improvements of rats with spinal cord injury(SCI).BMSCs did not survive long-term,disappearing from the spinal cord within 2–3 weeks after transplantation.Astrocyte-devoid areas,in which no astrocytes or oligodendrocytes were found,formed at the epicenter of the lesion.It was remarkable that numerous regenerating axons extended through such astrocyte-devoid areas.Regenerating axons were associated with Schwann cells embedded in extracellular matrices.Transplantation of choroid plexus epithelial cells(CPECs) also enhanced axonal regeneration and locomotor improvements in rats with SCI.Although CPECs disappeared from the spinal cord shortly after transplantation,an extensive outgrowth of regenerating axons occurred through astrocyte-devoid areas,as in the case of BMSC transplantation.These findings suggest that BMSCs and CPECs secret neurotrophic factors that promote tissue repair of the spinal cord,including axonal regeneration and reduced cavity formation.This means that transplantation of BMSCs and CPECs promotes "intrinsic" ability of the spinal cord to regenerate.The treatment to stimulate the intrinsic regeneration ability of the spinal cord is the safest method of clinical application for SCI.It should be emphasized that the generally anticipated long-term survival,proliferation and differentiation of transplanted cells are not necessarily desirable from the clinical point of view of safety.展开更多
BACKGROUND:Bone marrow cells can differentiate into hepatocytes in a suitable microenvironment.This study was undertaken to investigate the effects of transplanted bone marrow stromal cells (BMSCs) on liver fibrosis i...BACKGROUND:Bone marrow cells can differentiate into hepatocytes in a suitable microenvironment.This study was undertaken to investigate the effects of transplanted bone marrow stromal cells (BMSCs) on liver fibrosis in mice. METHODS:BMSCs were harvested and cultured from male BALB/c mice, then transplanted into female syngenic BALB/c mice via the portal vein. After partial hepatectomy, diethylnitrosamine (DEN) was administered to induce liver fibrosis. Controls received BMSCs and non-supplemented drinking water, the model group received DEN with their water, and the experimental group received BMSCs and DEN. Mice were killed after 3 months, and ALT, AST, hyaluronic acid (HA), and laminin (LN) in serum and hydroxyproline (Hyp) in the liver were assessed. Alpha-smooth muscle actin (α-SMA) in the liver was assessed by immunohistochemistry. Bone marrow- derived hepatocytes were identified by fluorescent in situ hybridization (FISH) in liver sections. RESULTS:BMSCs were shown to differentiate into hepatocyte-like phenotypes after hepatocyte growth factor treatment in vitro. Serum ALT, AST, HA, and LN were markedly reduced by transplanted BMSCs. Liver Hyp content and α-SMA staining in mice receiving BMSCs were lower than in the model group, consistent with altered liver pathology. FISH analysis revealed the presence of donor- derived hepatocytes in the injured liver after cross-gender mouse BMSC transplantation. After three months, about 10% of cells in the injured liver were bone marrow-derived. CONCLUSION:BMSCs transplanted via the portal vein can convert into hepatocytes to repair liver injury induced by DEN, restore liver function, and reduce liver fibrosis.展开更多
Objective To investigate the differentiation of bone marrow stromal cells (BMSC) into neuron-like cells and to explore their potential use for neural transplantation. Methods BMSC from rats and adult humans were cultu...Objective To investigate the differentiation of bone marrow stromal cells (BMSC) into neuron-like cells and to explore their potential use for neural transplantation. Methods BMSC from rats and adult humans were cultured in serum-containing media. Salvia miltiorrhiza was used to induce human BMSC (hBMSC) to differentiate. BMSC were identified with immunocytochemistry. Semi-quantitative RT-PCR was used to examine mRNA expression of neurofilament1 (NF1), nestin and neuron-specific enolase (NSE) in rat BMSC (rBMSC). Rat BMSC labelled by Hoschst33258 were transplanted into striatum of rats to trace migration and distribution. Results rBMSC expressed NSE, NF1 and nestin mRNA, and NF1 mRNA and expression was increased with induction of Salvia miltiorrhiza. A small number of hBMSC were stained by anti-nestin, anti-GFAP and anti-S100. Salvia miltiorrhiza could induce hBMSC to differentiate into neuron-like cells. Some differentiated neuron-like cells, that expressed NSE, beta-tubulin and NF-200, showed typical neuron morphology, but some neuron-like cells also expressed alpha smooth muscle protein, making their neuron identification complicated. rBMSC could migrate and adapted in the host brains after being transplanted. Conclusion Bone marrow stromal cells could express phenotypes of neurons, and Salvia miltiorrhiza could induce hBMSC to differentiate into neuron-like cells. If BMSC could be converted into neurons instead of mesenchymal derivatives, they would be an abundant and accessible cellular source to treat a variety of neurological diseases.展开更多
Objective To investigate the effects of Panax notoginseng saponins(PNS)on hydrogen peroxide(H2O2)-induced apoptosis in cultured rabbit bone marrow stromal cells(BMSCs).Methods BMSCs from 3-month-old New Zealand rabbit...Objective To investigate the effects of Panax notoginseng saponins(PNS)on hydrogen peroxide(H2O2)-induced apoptosis in cultured rabbit bone marrow stromal cells(BMSCs).Methods BMSCs from 3-month-old New Zealand rabbits were isolated and cultured by the density gradient centrifugation combined with adherent method.The cultured BMSCs were divided into three groups:normal control,H2O2 treatment(100μmol/L),and PNS pretreatment(0.1g/L).Intracellular reactive oxygen species(ROS)levels as the index of oxidative stress were measured by using 2'7'-dichlorodihydrofluorescein diacetate.Flow cytometry was used to observe the apoptosis of BMSCs by staining with annexinV-FITC/PI.The protein expression of Bax in BMSCs was analyzed by Western blotting.Activity of caspase-3 enzyme was measured by spectrofluorometry.Results Pretreatment with PNS significantly decreased intracellular ROS level induced by H2O2(P<0.01).PNS markedly attenuated H2O2-induced apoptosis rate from 38.68% to 19.24%(P<0.01).PNS reversed H2O2-induced augmentation of Bax expression.Furthermore,PNS markedly reduced the altered in activity of caspase-3 enzyme induced by H2O2(P<0.01).Conclusion PNS has a protective effect on hydrogen peroxide-induced apoptosis in cultured rabbit BMSCs by scavenging ROS and decreasing Bax expression and caspase-3 activity.展开更多
Adipose-derived stem cells and bone marrow-derived stromal stem cells were co-cultured with untreated or Aβ1-40-treated PC12 cells, or grown in supernatant derived from untreated or Aβ1-40-treated PC12 cells. Analys...Adipose-derived stem cells and bone marrow-derived stromal stem cells were co-cultured with untreated or Aβ1-40-treated PC12 cells, or grown in supernatant derived from untreated or Aβ1-40-treated PC12 cells. Analysis by western blot and quantitative real-time PCR showed that protein levels of Nanog, Oct4, and Sox2, and mRNA levels of miR/125a/3p were decreased, while expression of insulin-like growth factor-2 and neuron specific enolase was increased. In comparison, the generation of neuron specific enolase-positive cells was most successful when adipose-derived stem cells were co-cultured with Aβ1-40-treated PC12 cells. Our results demonstrate that adipose-derived stem cells and bone marrow-derived stromal stem cells exhibit trends of neuronal-like cell differentiation after co-culture with Aβ1-40-treated PC12 cells. This process may relate to a downregulation of miR-125a-3p mRNA expression and increased levels of insulin-like growth factor-2 expression.展开更多
We previously demonstrated that overexpression of tropomyosin receptor kinase A(TrkA)promotes the survival and Schwann celllike differentiation of bone marrow stromal stem cells in nerve grafts,thereby enhancing the r...We previously demonstrated that overexpression of tropomyosin receptor kinase A(TrkA)promotes the survival and Schwann celllike differentiation of bone marrow stromal stem cells in nerve grafts,thereby enhancing the regeneration and functional recovery of the peripheral nerve.In the present study,we investigated the molecular mechanisms underlying the neuroprotective effects of TrkA in bone marrow stromal stem cells seeded into nerve grafts.Bone marrow stromal stem cells from Sprague-Dawley rats were infected with recombinant lentivirus vector expressing rat TrkA,TrkA-shRNA or the respective control.The cells were then seeded into allogeneic rat acellular nerve allografts for bridging a 1-cm right sciatic nerve defect.Then,8 weeks after surgery,hematoxylin and eosin staining showed that compared with the control groups,the cells and fibers in the TrkA overexpressing group were more densely and uniformly arranged,whereas they were relatively sparse and arranged in a disordered manner in the TrkA-shRNA group.Western blot assay showed that compared with the control groups,the TrkA overexpressing group had higher expression of the myelin marker,myelin basic protein and the axonal marker neurofilament 200.The TrkA overexpressing group also had higher levels of various signaling molecules,including TrkA,pTrkA(Tyr490),extracellular signal-regulated kinases 1/2(Erkl/2),pErk1/2(Thr202/Tyr204),and the anti-apoptotic proteins Bcl-2 and Bcl-xL.In contrast,these proteins were downregulated,while the pro-apoptotic factors Bax and Bad were upregulated,in the TrkA-shRNA group.The levels of the TrkA effectors Akt and pAkt(Ser473)were not different among the groups.These results suggest that TrkA enhances the survival and regenerative capacity of bone marrow stromal stem cells through upregulation of the Erk/Bcl-2 pathway.All procedures were approved by the Animal Ethical and Welfare Committee of Shenzhen University,China in December 2014(approval No.AEWC-2014-001219).展开更多
AIM To investigate whether mesenchymal stem cells(MSCs) from adipose-derived stromal cells(ADSCs) and bone marrow stromal cells(BMSCs) have similar hepatic differentiation potential.METHODS Mouse ADSCs and BMSCs were ...AIM To investigate whether mesenchymal stem cells(MSCs) from adipose-derived stromal cells(ADSCs) and bone marrow stromal cells(BMSCs) have similar hepatic differentiation potential.METHODS Mouse ADSCs and BMSCs were isolated and cultured. Their morphological and phenotypic characteristics, as well as their multiple differentiation capacity were compared. A new culture system was established to induce ADSCs and BMSCs into functional hepatocytes. Reverse transcription polymerase chain reaction, Western blot, and immunofluorescence analyses were performed to identify the induced hepatocytelike cells. CM-Dil-labeled ADSCs and BMSCs were then transplanted into a mouse model of CCl4-induced acute liver failure. fluorescence microscopy was used to track the transplanted MSCs. Liver function was tested by an automatic biochemistry analyzer, and liver tissue histology was observed by hematoxylin and eosin(HE) staining.RESULTS ADSCs and BMSCs shared a similar morphology and multiple differentiation capacity, as well as a similar phenotype(with expression of CD29 and CD90 and no expression of CD11 b or CD45). Morphologically, ADSCs and BMSCs became round and epithelioid following hepatic induction. These two cell types differentiated into hepatocyte-like cells with similar expression of albumin, cytokeratin 18, cytokeratin 19, alpha fetoprotein, and cytochrome P450. fluorescence microscopy revealed that both ADSCs and BMSCs were observed in the mouse liver at different time points. Compared to the control group, both the function of the injured livers and HE staining showed significant improvement in the ADSC-and BMSC-transplanted mice. There was no significant difference between the two MSC groups.CONCLUSION ADSCs share a similar hepatic differentiation capacity and therapeutic effect with BMSCs in an acute liver failure model. ADSCs may represent an ideal seed cell type for cell transplantation or a bio-artificial liver support system.展开更多
This study investigated the ability of millimeter-wave(MMW)to promote the differen-tiation of bone marrow stromal cells(BMSCs)into cells with a neural phenotype.The BMSCs were primarily cultured.At passage 3,the cells...This study investigated the ability of millimeter-wave(MMW)to promote the differen-tiation of bone marrow stromal cells(BMSCs)into cells with a neural phenotype.The BMSCs were primarily cultured.At passage 3,the cells were induced byβ-mercaptoethanol(BME)in combination with MMW or BME alone.The expressions of nucleostemin(NS)and neuron-specific enolase(NSE) were detected by immunofluorescent staining and Western blotting respectively to identify the differentiation.The untreated BMSCs predominately expressed NS.After induced by BME and MMW,the BMSCs exhibited a dramatic decrease in NS expression and increase in NSE expression.The differentiation rate of the cells treated with BME and MMW in combination was significantly higher than that of the cells treated with BME alone(P<0.05).It was concluded that MMW exposure enhanced the inducing effect of BME on the differentiation of BMSCs into cells with a neural phenotype.展开更多
Objective To explore the possibilities of bone marrow stromal cells (MSCs) to adopt Schwann cell phenotype in vitro and in vivo in SD rats. Methods MSCs were obtained from tibia and femur bone marrow and cultured in c...Objective To explore the possibilities of bone marrow stromal cells (MSCs) to adopt Schwann cell phenotype in vitro and in vivo in SD rats. Methods MSCs were obtained from tibia and femur bone marrow and cultured in culture flasks. Beta-mercaptoethanol followed by retinoic acid, forskolin, basic-FGF, PDGF and heregulin were added to induce differentiation of MSCs’. Schwann cell markers, p75, S-100 and GFAP were used to discriminate induced properties of MSCs’ by immunofluorescent staining. PKH-67-labelled MSCs were transplanted into the mechanically injured rat sciatic nerve, and laser confocal microscopy was performed to localize the PKH67 labelled MSCs in the injured sciatic nerve two weeks after the operation. Fluorescence PKH67 attenuation rule was evaluated by flow cytometry in vitro. Results MSCs changed morphologically into cells resembling primary cultured Schwann cells after their induction in vitro. In vivo, a large number of MSCs were cumulated within the layer of epineurium around the injured nerve and expressed Schwann cell markers, p75, S-100, and GFAP. Conclusion MSCs are able to support nerve fiber regeneration and re-myelination by taking on Schwann cell function, and can be potentially used as possible substitutable cells for artificial nerve conduits to promote nerve regeneration.展开更多
Objective:To study the changes of gene expression profiles associated with osteoblasts differentiated from rat bone marrow stromal cells in vitro by gene chip technique.Methods:rat Rone marrow stromal cells were isola...Objective:To study the changes of gene expression profiles associated with osteoblasts differentiated from rat bone marrow stromal cells in vitro by gene chip technique.Methods:rat Rone marrow stromal cells were isolated and cultured,and differentiation was induced by dexamethasone,β-glycerol phosphate and vitamin C.Cellular mRNA was extracted and reverse transcribed into cDNA,thus related genes expression differences were detected by gene expression profile chip.Results:Calcifying nodules were visible in the induced cells.There were27.7%genes expressed differentially,three times more than the normal and induced cells,and some genes were related to transcription,translation,glycosylation modification.Extracellular matrix,signal molecules and metabolism were up—regulated.Conclusions:The gene chip technique can be used to detect the multi-gene different expression in the differentiationinduceed rat BMSCs,and these differentially expressed genes are necessary genes related to rat BMSCs proliferation and induction of osteoblastic differentiation.展开更多
To study the cartilage differentiation of mouse mesenchymal stem cells(MSCs) induced by cartilage-derived morphogenetic proteins-2 in vitro,the MSCs were isolated from mouse bone marrow and cultured in vitro.The cells...To study the cartilage differentiation of mouse mesenchymal stem cells(MSCs) induced by cartilage-derived morphogenetic proteins-2 in vitro,the MSCs were isolated from mouse bone marrow and cultured in vitro.The cells in passage 3 were induced into chondrogenic differentiation with different concentrations of recombinant human cartilage-derived morphogenetic proteins-2(0,10,20,50 and 100 ng/mL).After 14 days of induction,morphology of cells was observed under phase-contrast microscope.Collagen Ⅱ mRNA and protein were examined with RT-PCR,Western blotting and immunocytochemistry respectively and the sulfate glycosaminoglycan was measured by Alcian blue staining.RT-PCR showed that CDMP-2 could promote expression of collagen Ⅱ mRNA in an dose-dependant manner,especially at the concentration of 50 ng/mL and 100 ng/mL.Immunocytochemistry and Western blotting revealed a similar change.Alcian blue staining exhibited deposition of typical cartilage extracellular matrix.Our results suggest that mouse bone marrow mesencymal stem cells can differentiate into chondrogenic phonotype with the induction of CDMP-2 in vitro,which provides a basis for further research on the role of CDMP-2 in chondrogenesis.展开更多
BACKGROUND:Studies have demonstrated that bone marrow stromal cells (BMSCs) undergo neuronal differentiation under certain in vitro conditions.However,very few inducers of BMSC differentiation have been used in clinic...BACKGROUND:Studies have demonstrated that bone marrow stromal cells (BMSCs) undergo neuronal differentiation under certain in vitro conditions.However,very few inducers of BMSC differentiation have been used in clinical application.The effects of vascular endothelial growth factor (VEGF) on in vitro neuronal differentiation of BMSCs remain poorly understood.OBJECTIVE:To investigate the effect of VEGF on neuronal differentiation of BMSCs in vitro,and to determine the best VEGF concentration for experimental induction.DESIGN,TIME AND SETTING:In vitro comparative study was performed at the Central Laboratory and Laboratory of Male Reproductive Medicine,Shenzhen Hospital of Peking University from October 2008 to August 2009.MATERIALS:Recombinant human VEGF165 was purchased from Peprotech Asia,Rehovot,Israel.Neuron-specific enolase (NSE) was purchased from Beijing Biosynthesis Biotechnology,China.METHODS:BMSCs were harvested from adult Sprague Dawley rats.The passaged cells were pre-induced with 10 ng/mL basic fibroblast growth factor for 24 hours,followed by differentiation induction with 0,5,10,and 20 ng/mL VEGF,respectively.MAIN OUTCOME MEASURES:Morphological changes in BMSCs prior to and following VEGF induction.Expression of NSE following induction was determined by immunocytochemistry.RESULTS:Shrunken,round cells,with a strong refraction and thin bipolar or multipolar primary and secondary branches were observed 3 days after induction with 5,10,and 20 ng/mL VEGF.However,these changes were not observed in the control group.At 10 days after induction,the number of NSE-positive cells was greatest in the 10 ng/mL VEGF-treated group (P < 0.05).The number of NSE-positive cells was least in the control group at 3 and 10 days post-induction (P < 0.05).Moreover,the number of NSE-positive cells was greater at 10 days compared with at 3 days after induction (P < 0.05).CONCLUSION:Of the VEGF concentrations tested,10 ng/mL induced the greatest number of neuronal-like cells in vitro from BMSCs.展开更多
文摘Cumulative evidence suggests that O-linkedβ-N-acetylglucosaminylation(OGlcNAcylation)plays an important regulatory role in pathophysiological processes.Although the regulatory mechanisms of O-GlcNAcylation in tumors have been gradually elucidated,the potential mechanisms of O-GlcNAcylation in bone metabolism,particularly,in the osteogenic differentiation of bone marrow mesenchymal stromal cells(BMSCs)remains unexplored.In this study,the literature related to O-GlcNAcylation and BMSC osteogenic differentiation was reviewed,assuming that it could trigger more scholars to focus on research related to OGlcNAcylation and bone metabolism and provide insights into the development of novel therapeutic targets for bone metabolism disorders such as osteoporosis.
基金Supported by National Natural Science Foundation of China,No.82001014,No.82071090Hubei Provincial Natural Science Foundation of China,No.2022CFB115.
文摘BACKGROUND Bone marrow mesenchymal stromal cells(BMSCs)are the commonly used seed cells in tissue engineering.Aryl hydrocarbon receptor(AhR)is a transcription factor involved in various cellular processes.However,the function of constitutive AhR in BMSCs remains unclear.AIM To investigate the role of AhR in the osteogenic and macrophage-modulating potential of mouse BMSCs(mBMSCs)and the underlying mechanism.METHODS Immunochemistry and immunofluorescent staining were used to observe the expression of AhR in mouse bone marrow tissue and mBMSCs.The overexpression or knockdown of AhR was achieved by lentivirus-mediated plasmid.The osteogenic potential was observed by alkaline phosphatase and alizarin red staining.The mRNA and protein levels of osteogenic markers were detected by quantitative polymerase chain reaction(qPCR)and western blot.After coculture with different mBMSCs,the cluster of differentiation(CD)86 and CD206 expressions levels in RAW 264.7 cells were analyzed by flow cytometry.To explore the underlying molecular mechanism,the interaction of AhR with signal transducer and activator of transcription 3(STAT3)was observed by co-immunoprecipitation and phosphorylation of STAT3 was detected by western blot.RESULTS AhR expressions in mouse bone marrow tissue and isolated mBMSCs were detected.AhR overexpression enhanced the osteogenic potential of mBMSCs while AhR knockdown suppressed it.The ratio of CD86+RAW 264.7 cells cocultured with AhR-overexpressed mBMSCs was reduced and that of CD206+cells was increased.AhR directly interacted with STAT3.AhR overexpression increased the phosphorylation of STAT3.After inhibition of STAT3 via stattic,the promotive effects of AhR overexpression on the osteogenic differentiation and macrophage-modulating were partially counteracted.CONCLUSION AhR plays a beneficial role in the regenerative potential of mBMSCs partially by increasing phosphorylation of STAT3.
基金Supported by the FONCYT,Argentina(PICT 2016-#1093)CONICET,Argentina(PIP2014-2016,#300)Fundación Florencio Fiorini(Subsidio 2021-2022),Argentina.
文摘Breast cancer is the predominant form of carcinoma among women worldwide,with 70%of advanced patients developing bone metastases,with a high mortality rate.In this sense,the bone marrow(BM)mesenchymal stem/stromal cells(MSCs)are critical for BM/bone homeostasis,and failures in their functionality,transform the BM into a premetastatic niche(PMN).We previously found that BM-MSCs from advanced breast cancer patients(BCPs,infiltrative ductal carcinoma,stage III-B)have an abnormal profile.This work aims to study some of the metabolic and molecular mechanisms underlying MSCs shift from a normal to an abnormal profile in this group of patients.A comparative analysis was undertaken,which included self-renewal capacity,morphology,proliferation capacity,cell cycle,reactive oxygen species(ROS)levels,and senescence-associatedβ‑galactosidase(SA‑β‑gal)staining of BMderived MSCs isolated from 14 BCPs and 9 healthy volunteers(HVs).Additionally,the expression and activity of the telomerase subunit TERT,as well as telomere length,were measured.Expression levels of pluripotency,osteogenic,and osteoclastogenic genes(OCT-4,SOX-2,M-CAM,RUNX-2,BMP-2,CCL-2,M-CSF,and IL-6)were also determined.The results showed that MSCs from BCPs had reduced,self-renewal and proliferation capacity.These cells also exhibited inhibited cell cycle progression and phenotypic changes,such as an enlarged and flattened appearance.Additionally,there was an increase in ROS and senescence levels and a decrease in the functional capacity of TERT to preserve telomere length.We also found an increase in pro-inflammatory/pro-osteoclastogenic gene expression and a decrease in pluripotency gene expression.We conclude that these changes could be responsible for the abnormal functional profile that MSCs show in this group of patients.
基金Supported by Natural Science Foundation of Shandong Province,China,No.ZR2020MH014,No.ZR2021QH179 and No.ZR2021MH182.
文摘BACKGROUND The low survival rate of mesenchymal stem cells(MSCs)caused by anoikis,a form of apoptosis,limits the therapeutic efficacy of MSCs.As a proapoptotic molecule,mammalian Ste20-like kinase 1(Mst1)can increase the production of reactive oxygen species(ROS),thereby promoting anoikis.Recently,we found that Mst1 inhibition could protect mouse bone marrow MSCs(mBMSCs)from H 2 O 2-induced cell apoptosis by inducing autophagy and reducing ROS production.However,the influence of Mst1 inhibition on anoikis in mBMSCs remains unclear.AIM To investigate the mechanisms by which Mst1 inhibition acts on anoikis in isolated mBMSCs.METHODS Poly-2-hydroxyethyl methacrylate-induced anoikis was used following the silencing of Mst1 expression by short hairpin RNA(shRNA)adenovirus transfection.Integrin(ITGs)were tested by flow cytometry.Autophagy and ITGα5β1 were inhibited using 3-methyladenine and small interfering RNA,respe-ctively.The alterations in anoikis were measured by Terminal-deoxynucleoitidyl Transferase Mediated Nick End Labeling and anoikis assays.The levels of the anoikis-related proteins ITGα5,ITGβ1,and phospho-focal adhesion kinase and the activation of caspase 3 and the autophagy-related proteins microtubules associated protein 1 light chain 3 II/I,Beclin1 and p62 were detected by Western blotting.RESULTS In isolated mBMSCs,Mst1 expression was upregulated,and Mst1 inhibition significantly reduced cell apoptosis,induced autophagy and decreased ROS levels.Mechanistically,we found that Mst1 inhibition could upregulate ITGα5 and ITGβ1 expression but not ITGα4,ITGαv,or ITGβ3 expression.Moreover,autophagy induced by upregulated ITGα5β1 expression following Mst1 inhibition played an essential role in the protective efficacy of Mst1 inhibition in averting anoikis.CONCLUSION Mst1 inhibition ameliorated autophagy formation,increased ITGα5β1 expression,and decreased the excessive production of ROS,thereby reducing cell apoptosis in isolated mBMSCs.Based on these results,Mst1 inhibition may provide a promising strategy to overcome anoikis of implanted MSCs.
基金Project supported by the National Natural Science Foundation of China (20971034)Foundation for Key Program of Ministry of Education of China (208018)+2 种基金Returned Scholars of Hebei Province (207041)Natural Science Key Foundation of Hebei Province (B2009000161)Natural Science Foundation of Hebei University
文摘In order to elucidate the action of La3+ on bone metabolism,effects of La3+ on the osteogenic and adipogenic differentiation of pri-mary mouse bone marrow stromal cells(BMSCs) were studied by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide(MTT) test,alkaline phosphatase(ALP) activity measurement,mineralized function,oil red O stain and measurement.The results showed that La3+ pro-moted the proliferation of BMSCs except at 1×10-10 and 1×10-6 mol/L.The effect of La3+ on the osteogenic differentiation depended on con-centrations at the 7th day,but the osteogenic differentiation was inhibited at any concentration at the 14th day.La3+ promoted the formation of mineralized matrix nodules except at 1×10-8 and 1×10-5 mol/L.La3+ inhibited adipogenic differentiation except at 1×10-10 and 1×10-7 mol/L at the 10th day,and inhibited adipogenic differentiation except at 1×10-9 mol/L at the 16th day.These findings suggested that La3+ might have protective effect on bone at appropriate dose and time.This would be valuable for better understanding the mechanism of the effect of La3+ on bone metabolism.
基金Project supported by National Natural Science Foundation of China(21271059)Research Fund for the Doctoral Program of Higher Education of China(20111301110004)
文摘One of the main target organs for the lanthanides(Ln) is bone. Previous studies revealed that ytterbium(Yb) produced damage to the skeletal system in vivo. But the effects of Yb3+ on bone marrow stromal cells(BMSCs) in vitro had not been reported. In this paper, cell viability, apoptosis, mitochondrial membrane potential(MMP), reactive oxygen species(ROS) and lactate dehydrogenase(LDH) were measured in order to study the effects of Yb3+ on BMSCs. The results indicated that Yb3+ displayed a slight positive effect on the BMSCs viability at concentrations of 1×10–6, 1×10–5, and 1×10–4 mol/L, but turned to decrease the viability of BMSCs at the highest concentration of 1×10–3 mol/L for 24, 48 and 72 h. Yb3+ at 1×10–3 mol/L promoted apoptosis of BMSCs, increased the levels of ROS and LDH, and decreased MMP in BMSCs. It suggested that the precipitate of Yb PO4 might decrease the viability of BMSCs. Yb3+ induced the apoptosis of BMSCs via mitochondrial pathway. The results might be useful for more rational application of Yb-based compounds in the future.
基金The Natural Science Foundation of HainanProvince, No. 805107
文摘AIM:To investigate the malignant potential of hepatic stem cells derived from the bone marrow stromal cells (BMSCs) in a mouse model of chemical hepatocarcino- genesis. METHODS:BMSCs from male BALB/c mice were harvested and cultured, then transplanted into female syngenic BALB/ c mice via portal vein. Hepato-carcinogenesis was induced by 6 mo of treatment with diethylnitrosamine (DEN). Six months later, the liver was removed from each treated mouse and evaluated by immunohistochemistry and fluorescence in situ hybridization (FISH). RESULTS:Twenty-six percent of recipient mice survived and developed multiple hepatocellular carcinomas (HCCs). Immunohistochemically, HCC expressed placental form of glutathione-S-transferase (GST-P) and α-fetoprotein, but did not express cytokeratin 19. Y chromosome positive hepatocytes were detected by fluorescent in situ hybridization (FISH) in the liver of mice treated with DEN after BMSCs transplantation while no such hepatocytes were identified in the liver of mice not treated with DEN. No HCC was positive for the Y chromosome by FISH. CONCLUSION:Hepatic stem cells derived from the bone marrow stromal cells have a low malignant potential in our mouse model of chemical hepatocarcingenesis.
基金supported in part by grants from the Japanese Ministry of Education,Culture,Sports,Science,and Technology(No.2300125 to CI,No.15K10957 to NN,and No.26870744 to KK)
文摘Transplantation of bone marrow stromal cells(BMSCs) enhanced the outgrowth of regenerating axons and promoted locomotor improvements of rats with spinal cord injury(SCI).BMSCs did not survive long-term,disappearing from the spinal cord within 2–3 weeks after transplantation.Astrocyte-devoid areas,in which no astrocytes or oligodendrocytes were found,formed at the epicenter of the lesion.It was remarkable that numerous regenerating axons extended through such astrocyte-devoid areas.Regenerating axons were associated with Schwann cells embedded in extracellular matrices.Transplantation of choroid plexus epithelial cells(CPECs) also enhanced axonal regeneration and locomotor improvements in rats with SCI.Although CPECs disappeared from the spinal cord shortly after transplantation,an extensive outgrowth of regenerating axons occurred through astrocyte-devoid areas,as in the case of BMSC transplantation.These findings suggest that BMSCs and CPECs secret neurotrophic factors that promote tissue repair of the spinal cord,including axonal regeneration and reduced cavity formation.This means that transplantation of BMSCs and CPECs promotes "intrinsic" ability of the spinal cord to regenerate.The treatment to stimulate the intrinsic regeneration ability of the spinal cord is the safest method of clinical application for SCI.It should be emphasized that the generally anticipated long-term survival,proliferation and differentiation of transplanted cells are not necessarily desirable from the clinical point of view of safety.
文摘BACKGROUND:Bone marrow cells can differentiate into hepatocytes in a suitable microenvironment.This study was undertaken to investigate the effects of transplanted bone marrow stromal cells (BMSCs) on liver fibrosis in mice. METHODS:BMSCs were harvested and cultured from male BALB/c mice, then transplanted into female syngenic BALB/c mice via the portal vein. After partial hepatectomy, diethylnitrosamine (DEN) was administered to induce liver fibrosis. Controls received BMSCs and non-supplemented drinking water, the model group received DEN with their water, and the experimental group received BMSCs and DEN. Mice were killed after 3 months, and ALT, AST, hyaluronic acid (HA), and laminin (LN) in serum and hydroxyproline (Hyp) in the liver were assessed. Alpha-smooth muscle actin (α-SMA) in the liver was assessed by immunohistochemistry. Bone marrow- derived hepatocytes were identified by fluorescent in situ hybridization (FISH) in liver sections. RESULTS:BMSCs were shown to differentiate into hepatocyte-like phenotypes after hepatocyte growth factor treatment in vitro. Serum ALT, AST, HA, and LN were markedly reduced by transplanted BMSCs. Liver Hyp content and α-SMA staining in mice receiving BMSCs were lower than in the model group, consistent with altered liver pathology. FISH analysis revealed the presence of donor- derived hepatocytes in the injured liver after cross-gender mouse BMSC transplantation. After three months, about 10% of cells in the injured liver were bone marrow-derived. CONCLUSION:BMSCs transplanted via the portal vein can convert into hepatocytes to repair liver injury induced by DEN, restore liver function, and reduce liver fibrosis.
基金This work was supported by Natural Science Foundation of Guangdong Province (No. 012452, No. 020001).
文摘Objective To investigate the differentiation of bone marrow stromal cells (BMSC) into neuron-like cells and to explore their potential use for neural transplantation. Methods BMSC from rats and adult humans were cultured in serum-containing media. Salvia miltiorrhiza was used to induce human BMSC (hBMSC) to differentiate. BMSC were identified with immunocytochemistry. Semi-quantitative RT-PCR was used to examine mRNA expression of neurofilament1 (NF1), nestin and neuron-specific enolase (NSE) in rat BMSC (rBMSC). Rat BMSC labelled by Hoschst33258 were transplanted into striatum of rats to trace migration and distribution. Results rBMSC expressed NSE, NF1 and nestin mRNA, and NF1 mRNA and expression was increased with induction of Salvia miltiorrhiza. A small number of hBMSC were stained by anti-nestin, anti-GFAP and anti-S100. Salvia miltiorrhiza could induce hBMSC to differentiate into neuron-like cells. Some differentiated neuron-like cells, that expressed NSE, beta-tubulin and NF-200, showed typical neuron morphology, but some neuron-like cells also expressed alpha smooth muscle protein, making their neuron identification complicated. rBMSC could migrate and adapted in the host brains after being transplanted. Conclusion Bone marrow stromal cells could express phenotypes of neurons, and Salvia miltiorrhiza could induce hBMSC to differentiate into neuron-like cells. If BMSC could be converted into neurons instead of mesenchymal derivatives, they would be an abundant and accessible cellular source to treat a variety of neurological diseases.
基金supported by the National Natural Science Foundation of China(No.30600624)
文摘Objective To investigate the effects of Panax notoginseng saponins(PNS)on hydrogen peroxide(H2O2)-induced apoptosis in cultured rabbit bone marrow stromal cells(BMSCs).Methods BMSCs from 3-month-old New Zealand rabbits were isolated and cultured by the density gradient centrifugation combined with adherent method.The cultured BMSCs were divided into three groups:normal control,H2O2 treatment(100μmol/L),and PNS pretreatment(0.1g/L).Intracellular reactive oxygen species(ROS)levels as the index of oxidative stress were measured by using 2'7'-dichlorodihydrofluorescein diacetate.Flow cytometry was used to observe the apoptosis of BMSCs by staining with annexinV-FITC/PI.The protein expression of Bax in BMSCs was analyzed by Western blotting.Activity of caspase-3 enzyme was measured by spectrofluorometry.Results Pretreatment with PNS significantly decreased intracellular ROS level induced by H2O2(P<0.01).PNS markedly attenuated H2O2-induced apoptosis rate from 38.68% to 19.24%(P<0.01).PNS reversed H2O2-induced augmentation of Bax expression.Furthermore,PNS markedly reduced the altered in activity of caspase-3 enzyme induced by H2O2(P<0.01).Conclusion PNS has a protective effect on hydrogen peroxide-induced apoptosis in cultured rabbit BMSCs by scavenging ROS and decreasing Bax expression and caspase-3 activity.
基金the Plan Program of Shenyang Science and Technology Bureau, No. 1091161-0-00
文摘Adipose-derived stem cells and bone marrow-derived stromal stem cells were co-cultured with untreated or Aβ1-40-treated PC12 cells, or grown in supernatant derived from untreated or Aβ1-40-treated PC12 cells. Analysis by western blot and quantitative real-time PCR showed that protein levels of Nanog, Oct4, and Sox2, and mRNA levels of miR/125a/3p were decreased, while expression of insulin-like growth factor-2 and neuron specific enolase was increased. In comparison, the generation of neuron specific enolase-positive cells was most successful when adipose-derived stem cells were co-cultured with Aβ1-40-treated PC12 cells. Our results demonstrate that adipose-derived stem cells and bone marrow-derived stromal stem cells exhibit trends of neuronal-like cell differentiation after co-culture with Aβ1-40-treated PC12 cells. This process may relate to a downregulation of miR-125a-3p mRNA expression and increased levels of insulin-like growth factor-2 expression.
基金supported by the National Natural Science Foundation of China,No.81372041(to DW),and No.81801220(to MGZ)
文摘We previously demonstrated that overexpression of tropomyosin receptor kinase A(TrkA)promotes the survival and Schwann celllike differentiation of bone marrow stromal stem cells in nerve grafts,thereby enhancing the regeneration and functional recovery of the peripheral nerve.In the present study,we investigated the molecular mechanisms underlying the neuroprotective effects of TrkA in bone marrow stromal stem cells seeded into nerve grafts.Bone marrow stromal stem cells from Sprague-Dawley rats were infected with recombinant lentivirus vector expressing rat TrkA,TrkA-shRNA or the respective control.The cells were then seeded into allogeneic rat acellular nerve allografts for bridging a 1-cm right sciatic nerve defect.Then,8 weeks after surgery,hematoxylin and eosin staining showed that compared with the control groups,the cells and fibers in the TrkA overexpressing group were more densely and uniformly arranged,whereas they were relatively sparse and arranged in a disordered manner in the TrkA-shRNA group.Western blot assay showed that compared with the control groups,the TrkA overexpressing group had higher expression of the myelin marker,myelin basic protein and the axonal marker neurofilament 200.The TrkA overexpressing group also had higher levels of various signaling molecules,including TrkA,pTrkA(Tyr490),extracellular signal-regulated kinases 1/2(Erkl/2),pErk1/2(Thr202/Tyr204),and the anti-apoptotic proteins Bcl-2 and Bcl-xL.In contrast,these proteins were downregulated,while the pro-apoptotic factors Bax and Bad were upregulated,in the TrkA-shRNA group.The levels of the TrkA effectors Akt and pAkt(Ser473)were not different among the groups.These results suggest that TrkA enhances the survival and regenerative capacity of bone marrow stromal stem cells through upregulation of the Erk/Bcl-2 pathway.All procedures were approved by the Animal Ethical and Welfare Committee of Shenzhen University,China in December 2014(approval No.AEWC-2014-001219).
基金Supported by the National Natural Science foundation of China,No.30900669 and No.81473271Technology Nova Plan of Beijing City,No.2011117China Postdoctoral Science foundation,No.2016T90994
文摘AIM To investigate whether mesenchymal stem cells(MSCs) from adipose-derived stromal cells(ADSCs) and bone marrow stromal cells(BMSCs) have similar hepatic differentiation potential.METHODS Mouse ADSCs and BMSCs were isolated and cultured. Their morphological and phenotypic characteristics, as well as their multiple differentiation capacity were compared. A new culture system was established to induce ADSCs and BMSCs into functional hepatocytes. Reverse transcription polymerase chain reaction, Western blot, and immunofluorescence analyses were performed to identify the induced hepatocytelike cells. CM-Dil-labeled ADSCs and BMSCs were then transplanted into a mouse model of CCl4-induced acute liver failure. fluorescence microscopy was used to track the transplanted MSCs. Liver function was tested by an automatic biochemistry analyzer, and liver tissue histology was observed by hematoxylin and eosin(HE) staining.RESULTS ADSCs and BMSCs shared a similar morphology and multiple differentiation capacity, as well as a similar phenotype(with expression of CD29 and CD90 and no expression of CD11 b or CD45). Morphologically, ADSCs and BMSCs became round and epithelioid following hepatic induction. These two cell types differentiated into hepatocyte-like cells with similar expression of albumin, cytokeratin 18, cytokeratin 19, alpha fetoprotein, and cytochrome P450. fluorescence microscopy revealed that both ADSCs and BMSCs were observed in the mouse liver at different time points. Compared to the control group, both the function of the injured livers and HE staining showed significant improvement in the ADSC-and BMSC-transplanted mice. There was no significant difference between the two MSC groups.CONCLUSION ADSCs share a similar hepatic differentiation capacity and therapeutic effect with BMSCs in an acute liver failure model. ADSCs may represent an ideal seed cell type for cell transplantation or a bio-artificial liver support system.
文摘This study investigated the ability of millimeter-wave(MMW)to promote the differen-tiation of bone marrow stromal cells(BMSCs)into cells with a neural phenotype.The BMSCs were primarily cultured.At passage 3,the cells were induced byβ-mercaptoethanol(BME)in combination with MMW or BME alone.The expressions of nucleostemin(NS)and neuron-specific enolase(NSE) were detected by immunofluorescent staining and Western blotting respectively to identify the differentiation.The untreated BMSCs predominately expressed NS.After induced by BME and MMW,the BMSCs exhibited a dramatic decrease in NS expression and increase in NSE expression.The differentiation rate of the cells treated with BME and MMW in combination was significantly higher than that of the cells treated with BME alone(P<0.05).It was concluded that MMW exposure enhanced the inducing effect of BME on the differentiation of BMSCs into cells with a neural phenotype.
基金This research was supported by the 863 Program (2002AA205071) and National Natural Foundation of China (30271306)
文摘Objective To explore the possibilities of bone marrow stromal cells (MSCs) to adopt Schwann cell phenotype in vitro and in vivo in SD rats. Methods MSCs were obtained from tibia and femur bone marrow and cultured in culture flasks. Beta-mercaptoethanol followed by retinoic acid, forskolin, basic-FGF, PDGF and heregulin were added to induce differentiation of MSCs’. Schwann cell markers, p75, S-100 and GFAP were used to discriminate induced properties of MSCs’ by immunofluorescent staining. PKH-67-labelled MSCs were transplanted into the mechanically injured rat sciatic nerve, and laser confocal microscopy was performed to localize the PKH67 labelled MSCs in the injured sciatic nerve two weeks after the operation. Fluorescence PKH67 attenuation rule was evaluated by flow cytometry in vitro. Results MSCs changed morphologically into cells resembling primary cultured Schwann cells after their induction in vitro. In vivo, a large number of MSCs were cumulated within the layer of epineurium around the injured nerve and expressed Schwann cell markers, p75, S-100, and GFAP. Conclusion MSCs are able to support nerve fiber regeneration and re-myelination by taking on Schwann cell function, and can be potentially used as possible substitutable cells for artificial nerve conduits to promote nerve regeneration.
文摘Objective:To study the changes of gene expression profiles associated with osteoblasts differentiated from rat bone marrow stromal cells in vitro by gene chip technique.Methods:rat Rone marrow stromal cells were isolated and cultured,and differentiation was induced by dexamethasone,β-glycerol phosphate and vitamin C.Cellular mRNA was extracted and reverse transcribed into cDNA,thus related genes expression differences were detected by gene expression profile chip.Results:Calcifying nodules were visible in the induced cells.There were27.7%genes expressed differentially,three times more than the normal and induced cells,and some genes were related to transcription,translation,glycosylation modification.Extracellular matrix,signal molecules and metabolism were up—regulated.Conclusions:The gene chip technique can be used to detect the multi-gene different expression in the differentiationinduceed rat BMSCs,and these differentially expressed genes are necessary genes related to rat BMSCs proliferation and induction of osteoblastic differentiation.
基金This project was supported by a grant from the National Natural Sciences Foundation of China (No 30471753)
文摘To study the cartilage differentiation of mouse mesenchymal stem cells(MSCs) induced by cartilage-derived morphogenetic proteins-2 in vitro,the MSCs were isolated from mouse bone marrow and cultured in vitro.The cells in passage 3 were induced into chondrogenic differentiation with different concentrations of recombinant human cartilage-derived morphogenetic proteins-2(0,10,20,50 and 100 ng/mL).After 14 days of induction,morphology of cells was observed under phase-contrast microscope.Collagen Ⅱ mRNA and protein were examined with RT-PCR,Western blotting and immunocytochemistry respectively and the sulfate glycosaminoglycan was measured by Alcian blue staining.RT-PCR showed that CDMP-2 could promote expression of collagen Ⅱ mRNA in an dose-dependant manner,especially at the concentration of 50 ng/mL and 100 ng/mL.Immunocytochemistry and Western blotting revealed a similar change.Alcian blue staining exhibited deposition of typical cartilage extracellular matrix.Our results suggest that mouse bone marrow mesencymal stem cells can differentiate into chondrogenic phonotype with the induction of CDMP-2 in vitro,which provides a basis for further research on the role of CDMP-2 in chondrogenesis.
基金the Science and Technology Research Program of Shenzhen,No. 200802005
文摘BACKGROUND:Studies have demonstrated that bone marrow stromal cells (BMSCs) undergo neuronal differentiation under certain in vitro conditions.However,very few inducers of BMSC differentiation have been used in clinical application.The effects of vascular endothelial growth factor (VEGF) on in vitro neuronal differentiation of BMSCs remain poorly understood.OBJECTIVE:To investigate the effect of VEGF on neuronal differentiation of BMSCs in vitro,and to determine the best VEGF concentration for experimental induction.DESIGN,TIME AND SETTING:In vitro comparative study was performed at the Central Laboratory and Laboratory of Male Reproductive Medicine,Shenzhen Hospital of Peking University from October 2008 to August 2009.MATERIALS:Recombinant human VEGF165 was purchased from Peprotech Asia,Rehovot,Israel.Neuron-specific enolase (NSE) was purchased from Beijing Biosynthesis Biotechnology,China.METHODS:BMSCs were harvested from adult Sprague Dawley rats.The passaged cells were pre-induced with 10 ng/mL basic fibroblast growth factor for 24 hours,followed by differentiation induction with 0,5,10,and 20 ng/mL VEGF,respectively.MAIN OUTCOME MEASURES:Morphological changes in BMSCs prior to and following VEGF induction.Expression of NSE following induction was determined by immunocytochemistry.RESULTS:Shrunken,round cells,with a strong refraction and thin bipolar or multipolar primary and secondary branches were observed 3 days after induction with 5,10,and 20 ng/mL VEGF.However,these changes were not observed in the control group.At 10 days after induction,the number of NSE-positive cells was greatest in the 10 ng/mL VEGF-treated group (P < 0.05).The number of NSE-positive cells was least in the control group at 3 and 10 days post-induction (P < 0.05).Moreover,the number of NSE-positive cells was greater at 10 days compared with at 3 days after induction (P < 0.05).CONCLUSION:Of the VEGF concentrations tested,10 ng/mL induced the greatest number of neuronal-like cells in vitro from BMSCs.
