Bone marrow mesenchymal stem cells protect myocardial ischemia-reperfusion injury by inducing regulatory T cells in mice / 中华急诊医学杂志

Objective:To investigate the protective effect and mechanism of bone marrow-derived mesenchymal stem cell (BM-MSC) on myocardial ischemia-reperfusion injury (MIRI) in mice.Methods:Twenty four C57 MIRI mice were randomly(random number) divided into four groups: SO group, RI group, MSC+RI group, and MSC + RI+ PC61 group. The ratio of Treg were detected by flow cytometry. Serum levels of CK, TNI, BNP, IL-10 and TGF-β were measured by ELISA. The histological changes of myocardium were observed by HE staining. The number of cardiomyocyte apoptosis was measured by TUNEL staining, and the area ratio of myocardial infarction were determined by TTC staining. One-way ANOVA was used to analyze the data.Results:In the MSC + RI group, the ratio of Treg and the levels of IL-10 and TGF-β were the highest, while CK, TNI and BNP were the lowest ( P<0.01) .The number of myocardial apoptotic cells, infarct size and tissue fibrosis were the least ( P<0.01) . Conclusions:MSC can induce the production of Treg, increase the release of anti-inflammatory cytokines IL-10 and TGF-β, and reduce the inflammatory injury after myocardial ischemia-reperfusion.

Clinical Safety and Efficacy of Autologous Bone Marrow-Derived Mesenchymal Stem Cell Transplantation in Sensorineural Hearing Loss Patients

Stem cell transplantation represents a promising therapy for several degenerating and necrotic diseases. In several animal studies, we could find hearing restoration after inoculation of the mesenchymal stem cells' as well as mesenchymal stem cells' differentiation of hair cells and spiral ganglion. But until now, no clinical study has been reported directly for the human being. In this pilot studies, we applied mesenchymal stem cells to human beings trans-venously. Although we verified the safety of the autologous bone marrow stem cell transplantation in sensorineural hearing loss patients but we could not achieve significant improvement in hearing.

Safety of the bone marrow derived mesenchymal stem cells after HLA A 2 gene silencing / 西安交通大学学报(医学版)

Objective To explore the safety of the human bone marrow‐derived mesenchymal stem cells (hBMSCs) after silencing of human leukocyte antigen A2 expression .Methods We divided the cells into three groups:normal cultured cells of the 8th passage served as control group , and hBMSCs after HLA A2 silencing expression of the 5th and 15th passage as experimental groups 1 and 2 ,respectively .The hBMSCs were recultured by sterile methods .The growth curve ,telomerase activation ,and expressions of P27 ,cyclin D2 and cyclin‐dependent kinase 4 (CDK4) were utilized to explore the safety of the hBMSCs induced by LV‐siRNA‐HLA A2 .The BMSCs were transplanted to the subcutaneous layer of nude mice .Tissue types were detected 24 weeks after transplantation . Results The cell curves had no obvious left or right shift in all the groups . The telomerease activation in experimental groups 1 and 2 did not significantly differ from those in control group . The expressions of anti‐oncogene P27 ,cyclin D2 and CDK4 had no obvious difference between the two experimental groups and control group , either . There was only ectopic osteogenesis 24 weeks after the BMSCs (HLA A2 gene silenced ) were transplanted to the subcutaneous layer of the nude mice .Conclusion There was no obvious evidence to support that hBMSCs had undergone change in safety after the silencing of HLA A 2 expression .

Bone marrow-derived mesenchymal stem cells improved prognosis in rats with extended liver resection / 中华肝胆外科杂志

Objective To study the role of mesenchymal stem cells (MSC) in an animal model combining ischemia-reperfusion with 85％ liver resection.Methods Eight-week-old male SD rats received BM-MSC by tail vein and then underwent 30-min ischemia followed by 85％ liver resection.The survival rate was monitored for 7 days after surgery.Liver regeneration was assessed on day 2 after hepatectomy.Liver damage,liver cell apoptosis,and cytokine expression in the first 24 h after hepatectomy were also assessed.Results BM-MSC mostly homed to the spleen.Transplantation significantly inhibited myeloperoxidase [(19.9 ± 6.0) mg/g vs.(41.4 ± 10.2) mg/g] and downregulated proinflammatory cytokines.BM-MSC significantly reduced the ALT and AST levels [AST (1 475 ± 275) IU/L vs.(2 550 ± 441) IU/L,P ＜ 0.05;ALT (738 ± 101) IU/L vs.(1 113 ± 268) IU/L,P ＜ 0.05].The attenuation of liver injury was also verified histologically 24 h after surgery.Liver cell apoptosis was markedly reduced.Moreover,BM-MSC infusion significantly promoted remnant liver regeneration.As a result,the survival rate was improved by BM-MSC treatment in this model (95％ vs 70％,P ＜ 0.05).Conclusion In an animal model combining ischemia-reperfusion with 85％ liver resection,BM-MSC infusion attenuated liver injury and promoted hepatocyte regeneration,resulting in improved survival rate.

Inhibition of hepatic stellate cells by bone marrow-derived mesenchymal stem cells in hepatic fibrosis

BACKGROUND/AIMS: Therapies involving bone-marrow-derived mesenchymal stem cells (BM-MSCs) have considerable potential in the management of hepatic disease. BM-MSCs have been investigated in regenerative medicine due to their ability to secrete various growth factors and cytokines that regress hepatic fibrosis and enhance hepatocyte functionality. The aim of this study was to determine the antifibrosis effect of BM-MSCs on activated hepatic stellate cells (HSCs) and the mechanism underlying how BM-MSCs modulate the function of activated HSCs. METHODS: We used HSCs in both direct and indirect co-culture systems with BM-MSCs to evaluate the antifibrosis effect of BM-MSCs. The cell viability and apoptosis were evaluated by a direct co-culture system of activated HSCs with BM-MSCs. The activations of both HSCs alone and HSCs with BM-MSCs in the direct co-culture system were observed by immunocytochemistry for alpha-smooth muscle actin (alpha-SMA). The levels of growth factors and cytokines were evaluated by an indirect co-culture system of activated HSCs with BM-MSCs. RESULTS: The BM-MSCs in the direct co-culture system significantly decreased the production of alpha-SMA and the viability of activated HSCs, whereas they induced the apoptosis of activated HSCs. The BM-MSCs in the indirect co-culture system decreased the production of transforming growth factor-beta1 and interleukin (IL)-6, whereas they increased the production of hepatocyte growth factor and IL-10. These results confirmed that the juxtacrine and paracrine effects of BM-MSCs can inhibit the proliferative, fibrogenic function of activated HSCs and have the potential to reverse the fibrotic process by inhibiting the production of alpha-SMA and inducing the apoptosis of HSCs. CONCLUSIONS: These results have demonstrated that BM-MSCs may exert an antifibrosis effect by modulating the function of activated HSCs.

Synergistic Effect of Bone Marrow-Derived Mesenchymal Stem Cells and Platelet-Rich Plasma in Streptozotocin-Induced Diabetic Rats

BACKGROUND: Diabetic wounds are a major clinical challenge, because minor skin wounds can lead to chronic, unhealed ulcers and ultimately result in infection, gangrene, or even amputation. Studies on bone marrow derived mesenchymal stem cells (BMSCs) and a series of growth factors have revealed their many benefits for wound healing and regeneration. Platelet-rich plasma (PRP) may improve the environment for BMSC development and differentiation. However, whether combined use of BMSCs and PRP may be more effective for accelerating diabetic ulcer healing remains unclear. OBJECTIVE: We investigated the efficacy of BMSCs and PRP for the repair of refractory wound healing in a diabetic rat model. METHODS: Forty-eight rats with diabetes mellitus induced by streptozotocin were divided into four groups: treatment with BMSCs plus PRP, BMSCs alone, PRP alone, phosphate buffered saline. The rate of wound closure was quantified. A histopathological study was conducted regarding wound depth and the skin edge at 7, 14, and 28 days after surgery. RESULTS: Wound healing rates were significantly higher in the BMSC plus PRP group than in the other groups. The immunohistochemistry results showed that the expression of platelet/endothelial cell adhesion molecule 1, proliferating cell nuclear antigen, and transforming growth factor-beta1 increased significantly in the BMSC plus PRP group compared to the other treatment groups. On day 7, CD68 expression increased significantly in the wounds of the BMSC plus PRP group, but decreased markedly at day 14 compared to the controls. CONCLUSION: The combination of BMSCs and PRP aids diabetic wound repair and regeneration.

The impact of the bone marrow-derived mesenchymal stem cell on the microenvironment of the human limbal stem cell / 中华实验眼科杂志

Background When limbal stem cell deficiency (LSCD) occurs,not only the limbal stem cells (LSCs) were damaged,but also the LSCs matrix microenvironment was under destruction.The treatment of LSCD include both replenishing of stem cells and restoration of microenvironment.So far,the method to improve the microenvironment of LSCD still exist limitation and urgently need to establish more appropriate microenvironment for the LSCs growth in vitro.Objective This study was to investigate whether the human bone marrow-derived mesenchymal stem cells (BMSCs) can be used as the ideal cells to repair limbal microenvironment and its possible mechanism of repairing limbal microenvironment during the human LSCs amplification in vitro as feeder cells.Methods BMSCs were cultured and passaged in vitro,and flow cytometry was used to assay the expressions of CD45,CD71,CD90,CD105 and HLA-DR and directionally induced BMSCs to the osteoblasts and adipocytes.BMSCs were treated using mitomycin C (MMC) to use as the feeder cells.LSCs were separately co-cultured with BMSCs,Swiss-3T3 feeder cells and free-feeder cells,and colony-forming efficiency (CFE) of the LSCs was compared among different co-cultured groups.LSCs were then cultured sequentially and identified by flow cytometry.Expression of cytokines in BMSCs was confirmed by reverse transcriptional polymerase chain reaction (RT-PCR).Results Cultured BMSCs showed a good homogeneity,with a lot of expressions of interstitial cell markers such as CD71,CD90,CD105 and less expressions of hematopoietic cell markers including CD45 and HLA-DR.After separately cocultured with feeder cells for 12 days,the CFE of the LSCs co-cultured with BMSCs,Swiss-3T3 and no feeder cells was 3.67％ ±0.58％,4.30％ ± 1.54％ and 0.20％ ±0.10％,showing a statistical significant difference among the three groups(F =15.420,P =0.040).There was no statistically significant difference in the C FE of the LSCs between the BMSCs feeder group and the Swiss-3T3 feeder cells group(P =0.456),between the BMSCs feeder group and the free-feeder cells group or the Swiss-3T3 co-culture group and the free-feeder cells group (P =0.005,0.002).LSCs presented with a positive response for ABCG2 antigen in the co-cultured with BMSCs group.Basic fibroblast growth factor(bFGF),stem cell factor (SCF) and N-cadherin(N-cad) were positively expressed in the BMSCs as feeder cells.Conclusions Human BMSCs-derived feeder cells can improve the growth of the stromal microenvironment of the LSCs and enhance their proliferation ability.Human BMSCs are suitable for engineering of epithelial sheets.

The research on the immuno-modulatory abilities of myelodysplastic syndromes patients bone marrow derived mesenchymal stem cells / 中华微生物学和免疫学杂志

Objective To investigate that whether the immuno-modulatory capacity is intact in myelodysplastic syndrome-refractory anemia (MDS-RA) derived mesenchymal stem cell (MSCs) and what are the differences compare with those from normal donors.Methods Isolating MSC from MDS and normal donors respectively and exam their differences on T lymphocyte activation,proliferation and suppression.Results The results showed that the capacity of suppressing T cell proliferation and activation is weakened.Conclusion We propose that MSCs from pathological environment might be abnormal and should not be used for autologous transplantation.

Biological activity of the cultured rabbit bone marrow-derived mesenchymal stem cells in vitro / 西安交通大学学报(医学版)

Objective To observe the effect of the in vitro isolation, culture and identification of rabbit bone marrow-derived mesenchymal stem cells (BMSCs) and explore the differentiation potentials of BMSCs. Methods BMSCs were isolated and purified by density gradient centrifugation combined with attachment culture method. The biological characteristics of BMSCs were observed under an inverted microscope. The BMSCs were identified with HE staining and CD34/CD44 immunohistochemical staining. The biological activity of BMSCs was detected by osteogenic induction and steatogenic induction. Results BMSCs were isolated successfully in vitro and had good homogenicity. Immunohistochemical staining of CD34/CD44 showed strong positive expression of CD44 and negative expression of CD34. Fortis osteogenic ability and positive staining of ALP could be detected by osteogenic induction, and fortis steatogenic ability and positive staining of axungia could be detected by steatogenic induction. Conclusion BMSCs have the capability of multi-directional differentiation in vitro.

Efficacy and Safety of Autologous Bone Marrow-derived Mesenchymal Stem Cell Treatment in Patients With Amyotrophic Lateral Sclerosis

BACKGROUND: This study was performed to investigate the efficacy and safety of intrathecal autologous bone marrowderived mesenchymal stem cells (MSCs) treatment for patients with ALS. METHODS: After a lead-in period for 3 months, 22 patients were treated with MSCs twice at an interval of 1 month. After initial MSCs injection, all patients were followed up for 3 months and their disease course, clinical characteristics were assessed. Disease status of patients were analyzed with ALS functional rating scale-revised (ALSFRS-R) for primary outcome measure, and additional clinical findings after treatment were all collected for secondary outcome measure and safety. Age and disease-duration matched patients with ALS were selected as a control group. RESULTS: During the follow-up period, MSCs treatment yielded a significant lesser change of ALSFRS-R score, compared to control group (1.54 vs 3.56, p<0.01). Moreover, the slop of decline of ALSFRS-R was significantly lower during the follow-up period, compared to the lead-in period in MSCs treatment group (2.68 vs 1.54, p=0.04), whereas the slopes during the two periods were not different in the control group (3.15 vs 3.56, p=0.37). MSCs treatment was well tolerated except for occurrences of transient headache, low back pain, and myalgia. CONCLUSIONS: Our results show that intrathecal MSCs injection can slow disease progression and might be used as a disease modifying modality as an alternative treatment choice in patients with ALS.

Evaluation of Spinal Fusion Using Bone Marrow Derived Mesenchymal Stem Cells with or without Fibroblast Growth Factor-4

OBJECTIVE: In this study, the authors assessed the ability of rat bone marrow derived mesenchymal stem cells (BMDMSCs), in the presence of a growth factor, fibroblast growth factor-4 (FGF-4) and hydroxyapatite, to act as a scaffold for posterolateral spinal fusion in a rat model. METHODS: Using a rat posterolateral spine fusion model, the experimental study comprised 3 groups. Group 1 was composed of 6 animals that were implanted with 0.08 gram hydroxyapatite only. Group 2 was composed of 6 animals that were implanted with 0.08 gram hydroxyapatite containing 1 x 10(6)/ 60 microliter rat of BMDMSCs. Group 3 was composed of 6 animals that were implanted with 0.08 gram hydroxyapatite containing 1 x 10(6)/ 60 microliter of rat BMDMSCs and FGF-4 1 microgram to induce the bony differentiation of the BMDMSCs. Rats were assessed using radiographs obtained at 4, 6, and 8 weeks postoperatively. After sacrifice, spines were explanted and assessed by manual palpation, high-resolution microcomputerized tomography, and histological analysis. RESULTS: Radiographic, high-resolution microcomputerized tomographic, and manual palpation revealed spinal fusion in five rats (83%) in Group 2 at 8 weeks. However, in Group 1, three (60%) rats developed fusion at L4-L5 by radiography and two (40%) by manual palpation in radiographic examination. In addition, in Group 3, bone fusion was observed in only 50% of rats by manual palpation and radiographic examination at this time. CONCLUSION: The present study demonstrates that 0.08 gram of hydroxyapatite with 1 x 10(6)/ 60 microliter rat of BMDMSCs induced bone fusion. FGF-4, added to differentiate primitive 1 x 10(6)/ 60 microliter rat of BMDMSCs did not induce fusion. Based on histologic data, FGF-4 appears to induce fibrotic change rather than differentiation to bone by 1 x 10(6)/ 60 microliter rat of BMDMSCs.

The Effect of Human Joint Fluid in the Surviviorship and Proliferation of Bone-Marrow Derived Progenitor Cell / 대한정형외과연구학회지

PURPOSE: To investigate the effect of human joint fluid media on the survival and proliferation of bone marrow derived precusor cell, and to provide the basic data of the intraarticular injection with scaffold-free progenitor cell in advanced degenerative osteoarthritis (OA). MATERIALS AND METHODS: We obtained the joint fluid and bone marrow from 15 patients who had total knee arthroplasty due to degenerative OA, and isolated the mesenchymal progenitor cells (MPCs) from bone marrow by washing and ten times subculture. We devided the control and experiment groups according to the addition of joint fluid at various ratios (1/100, 1/10, 1, 10, 100, 1000 microliter), and statistically analyzed the numbers of mesenchymal progenitor cell proliferated according to the culture period. RESULTS: The experiments using joint fluid without centrifuge showed the increase of MPCs as the culture poriod was extended and was independent to the existence of fetal bovine serum, the dose dependent pattern in the increase of MPCs in proportion to the dose of joint fluid, and statistically significant increase MPCs in 10, 100, 1000 microliter of serum contained groups, 1000 microliter of serum-free groups (p=0.039, p=0.017, p=0.077, p=0.004). The experiments using joint fluid with centrifuge showed the increase of MPCs as the culture period was extended and was independent of fetal bovine serum, and the dose dependent pattern in the increase of MPCs in proportion to the dose of joint fluid, and statistically significant increase MPCs in 1000 microliter of both serum contained and serum-free groups (p=0.006, p=0.024). CONCLUSION: MSCs not only can survive, but also proliferate in human joint fluid. The rate of proliferaton is increased faster by the adding of joint fluid than only using common media in cell culture. And the experiment shows the dose dependent pattern in the increase of MPCs in proportion to the dose of joint fluid.