| Objectives:(1)To isolate and cultivate ADMSCs derived from Balb/c mice in vitro. Cell surface markers and multi-lineage differentiation potential were examined in vitro.(2) To observe the effect of intravenous transplantation of allogeneic ADMSCs on the survival rate, temperature, liver function and renal function, inflammatory cytokines such as TNF-α, IL-6and IL-8, as well as anti-inflammatroy cytokines such as IL-4and IL-10in septic mice, thus further to explore the therapy effect and mechanism of ADMSCs on septic mice induced by LPS.Methods:(1) Adipose tissues were obtained from Specific pathogen Free Balb/c mice and separated with collagenase digestion and attachment culture methods. Cells at passages3were used to examined the cell surface markers detected by Flow cytometry and multi-lineage differentiation potential capacity. The differentiation potential of the cells was proved by chodrogenic, osteogenic and adipogenic differentiation.(2) Sixty Balb/c mouse weighting22-28g (8-12week old) were randomly divided into one of three groups, Sham control group (Saline group), sepsis group (LPS group) and ADMSCs group,20mice for each group. The model of sepsis was induced by injection of LPS (15mg/kg,1mg/mL) via the tail vein. A sham intervention was performed using saline (15mg/kg) in sham control group mice. Saline (15mg/kg) was injected via the tail vein five minutes after administering LPS in Saline group and LPS group. ADMSCs (1×106,100uL) was injected in ADMSCs group. Mice were subsequently followed for48hours. Survival and temperature were recorded for each group every four hours.(3) Thirty Balb/c mouse weighting22-28g (8-12week old) were randomly divided into one of three groups, Sham control group (Saline group), sepsis group (LPS group) and ADMSCs group,10mice for each group. The model of sepsis was induced by injection of LPS (10mg/kg,1mg/mL) via the tail vein. Other treatment in each group were the same as above. Six hours after giving LPS, blood were collected to analyze the serum concentrations of biochemical markers (SCr, ALT and AST) and cytokines TNF-α, IL-4, IL-6, IL-8, IL-10.Results:(1) After isolation and culture, we obtained a large amount of ADMSCs. The flow cytometry showed CDllb-, CD45-, CD34+, CD44+, CD106+, CD29+. ADMSCs could be functionally induced into chodrogenic, adipocytes and osteoblasts in the presence of appropriate conditioned media.(2) There were no mice dead in Saline group, the survival rate in LPS group and ADMSCs group were declined as time. The survival rate at24h in LPS group was30%, which was significantly lower than ADMSCs group (65%). Survival analysis showed that overall survival was significantly improved in ADMSCs group as compared to LPS group (P<0.001).(3) The temperature at4h,8h,12h,16h in LPS group were under36℃and were significantly lower than that in Saline group (P<0.001), temperature at4h,8h and16h in ADMSCs group were significantly higher than that in LPS group (P<0.001).(4) The serum level of SCr, ALT and AST in LPS group at6h were significantly higher than that in Saline group (P<0.05), ALT and AST were significantly higher than that in ADMSCs group (P<0.05).(5) The serum levels of all cytokines tested were significantly higher in LPS group than that in Saline group (P<0.05). Serum levels of TNF-α, IL-4, IL-6, IL-8and IL-10in ADMSCs group were significantly lower than that in LPS group (P<0.05).Conclusion:(1) Obtained ADMSCs were characterized by adherent growth, high proliferation, stem cell phenotype and multipotent differentiation.(2) Hypothermia was occurred and the serum levels of SCr, ALT, AST, TNF-α, IL-4, IL-6, IL-8and IL-10were increased in model of sepsis.(3) Intravenous transplantation of allogeneic ADMSCs could decline serum levels of TNF-α, IL-4, IL-6, IL-8and IL-10in model of sepsis. ADMSCs showed bi-direction regulation role on inflammatory and it could protect the organ function, improve temperature change and improve survival rate in model of sepsis.6Figures,114references... |
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