Minocycline对rhHGF刺激ECV304细胞株表达MMP9的抑制作用

The supression function of Minocycline to the function that rhHGF stimulates ECV304 cell line to express MMP-9

目的探讨重组人类肝细胞增殖因子(recombinant human hepatocyte growth factor,rhHGF)刺激ECV304细胞株表达基质金属蛋白酶-9(matrjx metalloproteinase-9,MMP-9)的作用及米诺环素(minocycline)的影响作用.方法绘制正常ECV304细胞株生长曲线,明确最适生长浓度及对数生长期;明确rhHGF及minocycline的作用浓度;流式细胞仪(flow cytometer,FCM)检测ECV304细胞株的MMP-9水平.结果①ECV304细胞株的最适生长浓度为1.0E+4/mL,对数生长期为3～5d;rhHCF的作用浓度为8ng/mL,minocycline的作用浓度为1.0E-5mol/L.②对照组MMP-9的表达量为39.74%;培养基中加入rhHGF8ng/mL后,细胞存活率为1.388169±0.102033(P＜0.01),MMP-9的表达量为40.32%;培养基中加入minocycline1.0E5mol/L后,细胞存活率为0.294526±0.076829(P＜0.01),MMP-9的表达量为19.92%.③培养基中加入minocycline 1.0E5mol/L 15min后加入rhHGF 8ng/mL,细胞存活率为0.397291±0.099504(P＜0.01),MMP-9的表达量为22 28%.结论rhHGF可刺激ECV304细胞株增殖,使MMP-9的表达量增加;minocycline可抑制ECV304细胞株增殖,使MMP-9的表达量降低;minocycline可拮抗rhHGF所引起的ECV304细胞株增殖,从而使MMP-9的表达量降低.

OBJECTIVE To investigate the function th at rhHGF stimulated ECV304 cell line to express MMP-9 and the suppression functio n of minocycline.METHOD To draw normal growth curve of ECV304 cell line in order to make sure its optimal growth concentration and logarithm growth time; To mak e sure the valid concentration of rhHGF and minocycline; To test MMP-9 expressi on level by Flow cytometer.RESULTS ①The optimal growth concentration of ECV304 cell line was 1.0E+4/mL and logarithm growth time was 3～5days. The valid concentration of rhHGF was 8ng/mL and the valid concentration of minocycline was 1.0E-5mol/L. ②The expression level of MMP-9 was 39.74% in controls. Cell livability was 1.3 88169±0.102033(P<0.01) when acceded to culture medium with rhHGF 8ng/mL, an d the expression level of MMP-9 was 40.32%. Cell livability was 0.294526±0.076 829(P<0.01) when acceded to culture medium with minocycline 1.0E-5mol/L, an d the expression level of MMP-9 was 19.92%. ③Cell livability was 0.397291±0.0 99504(P<0.01) when acceded to culture medium with rhHGF 8ng/mL after 15minut es acceding minocycline 1.0E-5mol/L to culture medium, and the expression level of MMP-9 was 22.28%. CONCLUSION rhHGF could stimulate the proliferation of ECV304 ce ll line and increase the protein expression level of MMP-9. Minocycline could s uppress the proliferation of ECV304 cell line and decrease the protein expressio n level of MMP-9. Minocycline could interrupt the proliferation of ECV304 cell line caused by rhHGF and decrease MMP-9 protein level.