Effects Of POLY(I:C) And LPS On The Proliferation And Secretion Function Of Human Mesenchemal Stem Cells On Which TLR3 And TLR4 Were Expressed

Objective The aim of this study was to dectect the expression of Toll-like receptors of mesenchemal stem cells derived from bone marrow of healthy donor(BM-MSC) and to study effects of TLR3 and TLR 4 activator POLY(I:C) and LPS on proliferation of BM-MSC and on cytokines secretion by BM-MSC respectively.Methods BM-MSC was isolated from bone marrow of healthy donor by FicoU method.Expression of CD34,CD45,HLA-DR,CD44 and CD71 in BM-MSC were shown by flow cytometry.CD71 was detected in BM-MSC by immunocytochemistry.TLR1-10 mRNA levels in BM-MSC were evaluated by semiquantitative RT-PCR.Effects of POLY(I:C) and LPS on proliferation of BM-MSC were studied by MTF method.Effects of POLY(I:C) and LPS on secretion of IL-10,IL-12 and IL-6 by BM-MSC were studied by ELISA method.Results The Results showed that expression of CD34,CD45 and HLA-DR in BM-MSC was negative while expression of CD44 and CD71 was positive by FCM.CD71 was localized in BM-MSC by immunocytochemistry.All of TLR1-10 mRNA was found in BM-MSC with high expression level of TLR2,TLR3,TLR4,TLR7,TLR8,TLR9 and low expression level of TLR1,TLRS,TLR6,TLR10. Viable counts of different concentration teams of POLY(I:C) were less than that of 10%FBS team (p=0.000,respectively),but more than that of DMEM team(p＜0.05,respectively) 5 days after incubation while a semilogarithm linear correlation was found between concentration and viable count(r=-0.998,p =0.039).However,viable counts of moderate and high concentration teams of LPS were less than that of 10%FBS team(p=0.013,p=0.003),but more than that of DMEM team(p=0.001,p=0.002) 7 days after incubation.Expression of CD34,CD45 and HLA-DR in BM-MSC was negative while expression of CD44 and CD71 was positive by FCM.CD71 was localized in BM-MSC by immunocytochemistry.High levels of TLR3 and 4 mRNA was found to be expressed in BM-MSC by RT-PCR.Compared with the control,the amout of IL-6 secreted by BM-MSC incubated with different doses of TLR3 activator POLY(I:C) was increased significantly(P=0.000,P=0.021),and the amount of IL-6 secreted was more in 25ug/mL group than that in 2.5ug/mL group(P=0.000).But when compared with the control,the amout of IL-10 secreted incubated with different doses of TLR3 activator POLY(I:C) was decreased significantly (P=0.000) and the amount of IL-10 secreted was less in 250ug/mL group than that in 25ug/mL group (P=0.000) while the amount of IL-12 secreted was not affected.Compared with the control,the amout of IL-6 secreted by BM-MSC incubated with different doses of TLR4 activator LPS was increased significantly(P=0.000),and there was no difference among the three groups(P＞0.05) while the amount of IL-10 an IL-12 secreted were not affected.:Conclusion Different levels of TLR1-10 mRNA were expressed in BM-MSC of healthy donor.It is suggested that expression of TLR3 and 4 on BM-MSC be functional.