ASM/Cer通路介导模拟失重大鼠股动脉平滑肌细胞增殖与凋亡改变及表型转换

Acid sphingomyelinase/ceramide pathway regulates proliferation,apoptosis and phenotype switch of femoral artery in simulated weightless rats

目的探究模拟失重(simulated weightlessness,SW)大鼠股动脉(femoral artery,FA)酸性鞘磷脂酶(acid sphingomyelinase,ASM)/神经酰胺(ceramide,Cer)通路改变及在FA平滑肌细胞增殖与凋亡改变及表型转换中的作用。方法采用后肢不荷重尾部悬吊(hindlimb unloaded tail suspended,HU)大鼠模拟微重力状态下血流动力学变化,通过蛋白免疫印迹分析、免疫组织化学染色等检测模拟失重大鼠ASM表达量、Cer含量以及血管平滑肌细胞增殖与凋亡及表型的改变。结果与对照(CON)组相比,HU大鼠FA血管平滑肌细胞中增殖细胞核抗原(proliferating cell nuclear antigen,PCNA)、半胱氨酸-天冬氨酸蛋白酶3(cysteinyl aspartate specific proteinase 3,Caspase3)、B淋巴细胞瘤-2(B-cell lymphoma-2,Bcl-2)和Bcl-2相关X蛋白(Bcl-2 associated X protein,Bax)的表达量,合成表型标记物(波形蛋白,Vimentin)/收缩表型标记物(α平滑肌肌动蛋白,α-smooth muscle actin,α-SMA)的比值以及ASM和Cer水平均显著升高(P<0.05)。此外,ASM抑制剂地昔帕明(desipramine,Dpm)孵育或盐酸多塞平(doxepin hydrochloride,DOX)灌胃后,HU大鼠FA中Bax的表达水平和Vimentin/α-SMA比值均显著降低(P<0.05)。Dpm孵育后,HU大鼠FA中Bcl-2表达呈上升趋势;DOX灌胃可显著增高HU组大鼠FA中Bcl-2的水平(P<0.05)。结论模拟失重可导致大鼠FA平滑肌细胞的增殖和凋亡水平均显著升高,并由收缩表型向合成表型转换,该重塑过程涉及ASM/Cer通路的过度激活。

AIM To investigate the role of acid sphingomyelinase(ASM)/ceramide(Cer)in proliferation,apoptosis and phenotype switch of vascular smooth muscle cells(VSMCs)in the femoral artery(FA)of simulated weightless rats.METHODS Rats were subject to hindlimb-unloaded tailsuspension(HU)to simulate the hemodynamic effect of weightlessness on FA.The expressions of ASM and Cer,and representative markers of proliferation,apoptosis and phenotype switch of VSMCs were examined by Western blotting and immunohistochemistry(IHC).RESULTS The protein expressions of proliferating cell nuclear antigen(PCNA),cysteinyl aspartate specific proteinase 3(Caspase3),B-cell lymphoma-2(Bcl-2)and Bcl-2 associated X protein(Bax)along with the ratio of Vimentin toα-smooth muscle actin(α-SMA)in FA were increased significantly by HU(P<0.05).Meanwhile,the ASM protein and Cer abundance in FA of HU rats were higher than those in CON(P<0.05).Furthermore,desipramine(Dpm)or doxepin hydrochloride(DOX),the two inhibitors of ASM,reversed Bax expression and the vimentin/α-SMA ratio in FA of HU rats substantially(P<0.05).Notably,the expression of Bcl-2 in FA of HU rats showed a tendency to be increased after Dpm treatment and was increased significantly after DOX gavage.CONCLUSION Simulated weightlessness modulates the proliferation and apoptosis of VSMCs in FA,promoting the phenotype switch from a contractile to synthetic state,which is attributable to the over-activation of the ASM/Cer pathway.