静水压力刺激对人骨髓间充质干细胞成骨、成脂双向分化的影响

目的 研究短期和较长期的静水压力负荷对人骨髓间充质干细胞(BMSC)成骨或成脂的影响,并探讨细胞外信号调节激酶(ERK)1/2信号通路在其中作用.方法 对BMSC施加40或80 kPa的静水压强,分别持续1 h或4 h,然后加入10 mmol/L U0126阻断ERK1/2信号通路,通过半定量RTPCR和实时定量PCR方法检测成骨标志基因Cbfa1和骨钙素(OC)以及成脂标志性基因PPARγ2和Adipsin的mRNA表达水平.对BMSC施加40 kPa的静水压刺激,每天作用4 h,7 d和14 d后检测Cbfa1、OC、PPARγ2和Adipsin的mRNA表达水平,在3、7、14 d通过组织化学染色和定量测定方法检测碱性磷酸酶的活性和表达量.结果 40 kPa压强的静水压连续刺激4 h可以促进Cbfa1、OC的mRNA表达和抑制PPARγ2、Adipsin的mRNA表达.40 kPa的静水压连续作用7 d,仍表现为对BMSC的促成骨分化作用,然而连续作用14 d后,则表现为促成脂分化作用.成骨诱导条件并不协同静水压的促BMSC成骨分化作用.而U0126并不能完全抑制BMSC对静水压刺激的反应.结论 一定强度和作用时间的静水压刺激可以促进BMSC的成骨分化,但较长期的作用可能会促进成脂分化.成骨诱导培养条件与静水压的促BMSC成骨分化作用无协同作用.

The effect of hydrostatic compressive loading on the differentiation of osteogenic/adipogenic human bone marrow-derived mesenchymal stem cells

Objective To examine short-term and long-term effects of continuous hydrostatic compressive loading on the expression of osteogenic/adipogenic marker genes and alkaline phosphatase(ALP)activity of human bone marrow-derived mesenchymal stem cells fhBMSC);to assess the role of extracellular signal-regulated kinase(ERK)1/2 pathway in the response of hydrostatic compression.Methods In the short-term compressive loading study,BMSC maintained in osteogenic supplementation(OS)medium were subiected to hydrostatic pressure with a magnitude of 40 or 80 kPa,which were sustained for 1 or 4 h.Then U0126.the inhibitor of ERK1/2 pathway,was added to the culture media and cells were also subjected to the a same pressures.Expression of Cbfa1,osteocalcin(OC),PPARγ2 and Adipsin were investigated by semiquantitative and real-time quantitative-PCR.In the long-term compressive loading study, a sustained hydrostatic pressure with a magnitude of 40 kPa was applied on the BMSC for 4 h every day.ALP activity were assessed qualitatively by histochemical staining and expression of Cbfa1,OC, PPARγ2 and Adipsin were also investigated after 7 or 14 d.Levels of ALP and OC were compared after 3,7,14 d.Resuits The hydrostatic compression with a magnitude of 40 kPa and a stimulation time of 4 h promoted expression of the osteogenic marker genes and inhibited expression of adipogenic marker genes jn BMSC.When the cells were subjected to this condition for 7 d.the hydrostatic compression could induce osteogenic differentiation.But in the end of 14 d, the hydrostatic compression with the condition resulted in adipogenic differentiation.The hydrostatic compression enhanced the ALP expression and activity.The OS medium did not enhance,even reversed in some time,the positive effect of the hydrostatic compression on BMSC osteogenic differentiation.The U0126 also did not completely inhibit these responses to the hydrostatic compression.Concluston The hydrostatic compression can promote osteogenic differentiation,but the same loading advance adipogenic differnentiation in the long term.The OS medium has no synergistic effect with the hydrostatic compression on osteogenic differentiation of BMSC.