Luo Zi-Yin, Xiang Bing-Qian, Gao Hui, Qiu Xiao-Xiao, Hao Mao-Lin, Wang Wan-Tie
Ischemia/Reperfusion Injury Research Institute of Wenzhou Medical University, Wenzhou 325035, China.
Pathology Department of Hubei Provincial Hospital of Integrated Chinese & Western Medicine, Wuhan 430015, China.
Sheng Li Xue Bao. 2017 Aug 25;69(4):437-444.
To investigate the effects of dexmedetomidine (DEX) on hypoxia/reoxygenation (H/R) injury-induced cell apoptosis and caspase-12 expression, A549 cells were randomly divided into 4 groups: control group, DEX group, H/R group and DEX+H/R group. Cells of control and DEX groups were cultured in the normoxic incubator for 30 h. Cells of H/R and DEX+ H/R groups were incubated in the anoxic cultivation for 6 h, followed by normoxic culture for 24 h, and DEX (1 nmol/L) was added into the culture medium in DEX and DEX+H/R groups. Morphological changes were observed under the inverted microscope. Cell viability was detected by CCK-8. The apoptosis index (AI) of A549 cells was detected by TUNEL method. The activity of caspase-3 enzyme in cells was detected by using caspase-3 kit. The expressions of GRP78, caspase-12 protein and mRNA were determined by Western blot and RT-PCR respectively. Compared with control group, the morphological changes of the cultured cells were observed: some of the cell fusion occurred and the shape of the cells was multilateral; the cell viability was decreased significantly (P < 0.01), the number of apoptotic cells and the AI value, caspase-3 activity, and the expressions of GRP78, caspase-12 protein/mRNA were significantly increased (P < 0.01) in H/R group. While the administration of DEX alleviated the H/R injury-induced cell damage, obviously increased the cell viability (P < 0.01), significantly decreased the increment of apoptotic cells and the AI value induced by H/R injury (P < 0.01), and also dramatically decreased the H/R injury-induced high level of caspase-3 activity (P < 0.01) as well as high expression of caspase-12 protein and mRNA (P < 0.01). Taken together, the results suggest that DEX can effectively protect A549 cells from the H/R injury, which may be mediated by down-regulating the expression of caspase-12 and inhibiting cell apoptosis.
为研究右美托咪定（DEX）对缺氧/复氧（H/R）损伤诱导的细胞凋亡及半胱天冬酶-12表达的影响，将A549细胞随机分为4组：对照组、DEX组、H/R组和DEX+H/R组。对照组和DEX组细胞在常氧培养箱中培养30小时。H/R组和DEX+H/R组细胞先在缺氧条件下培养6小时，然后再进行24小时常氧培养，DEX组和DEX+H/R组的培养基中加入DEX（1纳摩尔/升）。在倒置显微镜下观察形态变化。采用CCK-8法检测细胞活力。采用TUNEL法检测A549细胞的凋亡指数（AI）。使用半胱天冬酶-3试剂盒检测细胞中半胱天冬酶-3的活性。分别通过蛋白质免疫印迹法和逆转录-聚合酶链反应（RT-PCR）检测葡萄糖调节蛋白78（GRP78）、半胱天冬酶-12蛋白及mRNA的表达。与对照组相比，观察到培养细胞的形态变化：部分细胞出现融合，细胞形状呈多边性；细胞活力显著降低（P<0.01），H/R组凋亡细胞数量、AI值以及半胱天冬酶-3活性和GRP78、半胱天冬酶-12蛋白/mRNA的表达均显著增加（P<0.01）。而给予DEX可减轻H/R损伤诱导的细胞损伤，显著提高细胞活力（P<0.01），显著降低H/R损伤诱导的凋亡细胞增加量和AI值（P<0.01），同时也显著降低H/R损伤诱导的高水平半胱天冬酶-3活性（P<0.01）以及半胱天冬酶-12蛋白和mRNA的高表达（P<0.01）。综上所述，结果表明DEX可有效保护A549细胞免受H/R损伤，这可能是通过下调半胱天冬酶-12的表达并抑制细胞凋亡来介导的。
