Objective： To explore the effects of bone marrow-derived mesenchymal stem cells （BMSCs）transfected with adenoviral vector carrying hepatocyte growth factor （HGF, Ad-HGF） on burn wound healing.Methods： BMSCs from...Objective： To explore the effects of bone marrow-derived mesenchymal stem cells （BMSCs）transfected with adenoviral vector carrying hepatocyte growth factor （HGF, Ad-HGF） on burn wound healing.Methods： BMSCs from male Wistar rats were separated and purified with Percoll separating medium by density gradient centrifugation and cultured with DMEM containing 20% fetal bovine serum （FBS）. Then BMSCs were transfected with Ad-HGF at the optimal gene transduction efficiency of 100 multiplicity of infection （MOI）. The efficiency of transfection and the expression of HGF in the suspension were detected by flow cytometry and enzyme linked immunosorbent assay （ELISA） respectively. Thirtytwo female rats were subjected to 90℃ water for 12 seconds to induce a partial thickness skin burn. The animals were randomly divided into mesenchymal stem cells （MSCs） treatment group （Group A）, Ad-HGF treatment group （Group B）,Ad-HGF-modified MSCs treatment group （Group C） and saline control group （Group D）. On days 3, 5, 7, 14 and 21 postburn, HE and Sirius red stain were performed to observe the burn wound healing and collagen content. The content of hydroxyproline in wounds was also detected.Transplanted cells and the expression of（sex-determining region Y） SRY gene were detected by in situ hybridization and polymerase chain reaction （PCR）, while the expression of HGF in wound tissues was detected by ELISA.Results： The result of flow cytometry showed that the transfection efficiency was 86.41% at 100 MOI. Compared with the control group, the content of HGF in the supernatant after transfection increased time-dependently and peaked at 48 h, showing significant differences at 24 h, 48 h,72 h and 96 h （P＜0.01 ）. Results of HE stain revealed that the range of re-epidermidalization in Group C was significantly larger than that in other groups in the first week. Three weeks postburn, the epidermis was significantly thicker in Group C than in other groups and the nails of dermis inserted into the derma of burn wounds. Sirius red stain showed that the content of collagen Ⅰ in Group C was much less compared with that in other groups 21 days postburn. In situ hybridization revealed an expression of SRY gene in burned female rats, consistent with the finding of PCR. Immunohistochemistry demonstrated the largest increase of HGF expression in Group C, whose contents of hydroxyproline,however, decreased on day 7 postburn. Compared with other groups, the content of HGF in the wounds of Group C increased obviously on day 14 after transfection （P＜0.05） and there was no significant difference among Groups A, B and D.Conclusion： Our study suggests that transplantation of MSCs modified with Ad-HGF has positive effects on the healing of burn wounds probably through differentiation and release of relevant cytokines.展开更多
OBJECTIVE:To investigate the influence and mechanism of salvianolic acid B(SalB) on apoptosis inhibition in rat bone marrow-derived mesenchymal stem cells(BMSCs) induced by hypoxia and serum deprivation(hypoxia/SD).ME...OBJECTIVE:To investigate the influence and mechanism of salvianolic acid B(SalB) on apoptosis inhibition in rat bone marrow-derived mesenchymal stem cells(BMSCs) induced by hypoxia and serum deprivation(hypoxia/SD).METHODS:SalB concentration of 0.1,1,10 or 100 mg/L(drug groups) were investigated for their ability to inhibit apoptosis in rat BMSCs.BMSCs in both the apoptosis model and drug groups were cultured under hypoxic conditions for 6 h,after which cell apoptosis and change in mitochondrial membrane potential(MMP) were detected using flow cytometry.Activation of caspase-3 was detected using western blot analysis.RESULTS:Hypoxia/SD induced apoptosis in rat BMSCs.The early apoptosis rate was lower in the drug groups compared to the apoptosis model group(P<0.05).SalB was found to inhibit the reduction in MMP and decrease the activation of caspase-3.CONCLUSION:0.1,1 and 10 mg/L of SalB inhibits activation of caspase-3 and early apoptosis of rat BMSCs induced by hypoxia/SD and could therefore enhance the survival rate of grafted stem cells.展开更多
基金This study was funded by a grant from the National Natural Science Foundation of China （No. 30772572）, two research grants from China postdoctoral Science Foundation （No. 20060390192 and No. 200801243）, and a research grant for science from Gansu Provincial Science ＆ Technology Department （No. 0708NKCA128）.
文摘Objective： To explore the effects of bone marrow-derived mesenchymal stem cells （BMSCs）transfected with adenoviral vector carrying hepatocyte growth factor （HGF, Ad-HGF） on burn wound healing.Methods： BMSCs from male Wistar rats were separated and purified with Percoll separating medium by density gradient centrifugation and cultured with DMEM containing 20% fetal bovine serum （FBS）. Then BMSCs were transfected with Ad-HGF at the optimal gene transduction efficiency of 100 multiplicity of infection （MOI）. The efficiency of transfection and the expression of HGF in the suspension were detected by flow cytometry and enzyme linked immunosorbent assay （ELISA） respectively. Thirtytwo female rats were subjected to 90℃ water for 12 seconds to induce a partial thickness skin burn. The animals were randomly divided into mesenchymal stem cells （MSCs） treatment group （Group A）, Ad-HGF treatment group （Group B）,Ad-HGF-modified MSCs treatment group （Group C） and saline control group （Group D）. On days 3, 5, 7, 14 and 21 postburn, HE and Sirius red stain were performed to observe the burn wound healing and collagen content. The content of hydroxyproline in wounds was also detected.Transplanted cells and the expression of（sex-determining region Y） SRY gene were detected by in situ hybridization and polymerase chain reaction （PCR）, while the expression of HGF in wound tissues was detected by ELISA.Results： The result of flow cytometry showed that the transfection efficiency was 86.41% at 100 MOI. Compared with the control group, the content of HGF in the supernatant after transfection increased time-dependently and peaked at 48 h, showing significant differences at 24 h, 48 h,72 h and 96 h （P＜0.01 ）. Results of HE stain revealed that the range of re-epidermidalization in Group C was significantly larger than that in other groups in the first week. Three weeks postburn, the epidermis was significantly thicker in Group C than in other groups and the nails of dermis inserted into the derma of burn wounds. Sirius red stain showed that the content of collagen Ⅰ in Group C was much less compared with that in other groups 21 days postburn. In situ hybridization revealed an expression of SRY gene in burned female rats, consistent with the finding of PCR. Immunohistochemistry demonstrated the largest increase of HGF expression in Group C, whose contents of hydroxyproline,however, decreased on day 7 postburn. Compared with other groups, the content of HGF in the wounds of Group C increased obviously on day 14 after transfection （P＜0.05） and there was no significant difference among Groups A, B and D.Conclusion： Our study suggests that transplantation of MSCs modified with Ad-HGF has positive effects on the healing of burn wounds probably through differentiation and release of relevant cytokines.
基金Supported by China Post-doctoral Scientific Fund (20090450542)
文摘OBJECTIVE:To investigate the influence and mechanism of salvianolic acid B(SalB) on apoptosis inhibition in rat bone marrow-derived mesenchymal stem cells(BMSCs) induced by hypoxia and serum deprivation(hypoxia/SD).METHODS:SalB concentration of 0.1,1,10 or 100 mg/L(drug groups) were investigated for their ability to inhibit apoptosis in rat BMSCs.BMSCs in both the apoptosis model and drug groups were cultured under hypoxic conditions for 6 h,after which cell apoptosis and change in mitochondrial membrane potential(MMP) were detected using flow cytometry.Activation of caspase-3 was detected using western blot analysis.RESULTS:Hypoxia/SD induced apoptosis in rat BMSCs.The early apoptosis rate was lower in the drug groups compared to the apoptosis model group(P<0.05).SalB was found to inhibit the reduction in MMP and decrease the activation of caspase-3.CONCLUSION:0.1,1 and 10 mg/L of SalB inhibits activation of caspase-3 and early apoptosis of rat BMSCs induced by hypoxia/SD and could therefore enhance the survival rate of grafted stem cells.
