蛋白激酶C在腺苷A1受体调控缺氧心肌细胞线粒体通透性转换孔开放中的表达和作用

Role of protein kinase C in adenosine A1 receptor regulating opening of mitochondria permeability transition pore in hypoxic cardiac myocytes

目的观察腺苷A1受体的活化能否影响缺氧心肌细胞线粒体通透性转换孔的开放及蛋白激酶C(protein kinase C,PKC)在其中的作用。方法将原代培养的SD大鼠乳鼠心肌细胞分为5组:对照(常氧)组(A组),缺氧组(B组),缺氧+腺苷A1受体激动剂组(C组),缺氧+腺苷A1受体阻断剂组(D组),缺氧+腺苷A1受体激动剂+PKC阻断剂组(E组)。缺氧6h后以免疫荧光和免疫印迹法分析A、B、C、D4组PKC的表达变化,用酯化钙黄绿素和氯化钴共孵育法检测5组线粒体通透性转换孔(MPTP)开放,用CCK-8法检测细胞活力。结果缺氧6h后,B、D、E3组MPTP显著开放,细胞活力降低,而C组能明显减少MPTP的开放,减轻细胞损害,并能使PKC的表达量增加。结论缺氧可引起MPTP开放增加,腺苷A1受体激动剂可上调PKC表达,减少缺氧导致的MPTP开放,维持细胞活力。PKC通路可能是腺苷A1受体调控缺氧心肌细胞MPTP开放的重要途径。

Objective To investigate whether adenosine A1 receptor （A1 R） activation can affect the opening of mitochondria permeability transition pore （MPTP） in hypoxic cardiac myocytes and the effects of protein kinase C （PKC） in this process. Methods Neonatal rat cardiac myocytes in primary culture were randomized as normoxic group （A）, hypoxic group （B）, hypoxia and A1R agonist 2-chloro-N6-cyclopentyladenosine （CCPA, 500 nmol/L）（ C）, hypoxia and A1 R antagonist 8-cyclopentyl-1,3-dipropylxanthine （DPCPX, 500 nmol/L） （D）, hypoxia treated with CCPA （500 nmol/L） and PKC inhibitor chelerythine （CHE, 2 μmol/L） （E）. After hypoxia for 6 h, the expression of PKC, the opening of MPTP and the activity of cells were detected. Results The opening of MPTP increased and the activity of cardiac myocytes decreased in groups B, D and E. However, these phenomena in group C were opposite and PKC increased. Conclusion Hypoxia can increase the opening of MPTP and decrease the activity of cardiac cells, while A1R agonist abolishes these bad effects and increases the expression of PKC. Whereas, the PKC inhibitor prevents this protective role. PKC maybe is the key way that AIR regulates the opening of MPTP in hypoxic cardiac myocytes.