目的:观察大鼠颌下腺放射后导致的氧化应激对细胞凋亡的影响.方法:24只大鼠随机分为对照组及放射24 h、72 h及1周组.对照组未照射,放射组右侧颌下腺进行一次性20 Gy照射,检测各组大鼠唾液分泌流率.苏木精-伊红(hematoxylin-eosin,HE)染色观察颌下腺组织形态改变;酶联免疫吸附试验法(enzyme linked immu-nosorbent assay,ELISA)检测丙二醛(malondialdehyde,MDA)、还原型谷胱甘肽(glutathione,GSH)及超氧化物歧化酶(superoxide dismutase,SOD)水平.免疫组织化学染色(immunol histochemistry,IHC)观察8-羟基脱氧鸟苷(8-hydroxy-2 deoxyguanosine,8-OHdG)阳性表达;末端脱氧核苷酸转移酶dUTP缺口末端标记法(TdT-mediated dUTP nick-end labeling,TUNEL)检测颌下腺腺泡细胞凋亡情况;Western blot和实时定量聚合酶链反应(real-time fluorescence quantitative PCR,RT-qPCR)检测凋亡相关蛋白含半胱氨酸的天冬氨酸蛋白水解酶3(cysteinyl aspar-tate specific proteinase-3,Caspase-3)、Bcl-2 关联 X 的蛋白质(BCL2-associated X protein,BAX)及 B 淋巴细胞瘤-2基因(B-cell lymphoma-2,Bcl-2)的蛋白及mRNA表达情况.结果:放射后大鼠唾液分泌减少;细胞内空泡化、组织水肿及间隙增宽等病理变化逐渐加重;细胞出现明显的氧化应激反应,并伴随出现细胞的凋亡;凋亡相关蛋白BAX、Bcl-2及Caspase-3蛋白及mRNA表达量异常.结论:放射后氧化应激反应可能是颌下腺细胞凋亡及功能障碍的重要影响因素.
Effect of Oxidative Stress after Radiation on Apoptosis of Cells in Rats Submandibular Gland
Objective:To observe the effect of oxidative stress induced by submandibular gland(SMG)radiation on apoptosis of acinar cells in rats.Methods:Twenty-four rats were randomly divided into control group,24 h,72 h,and 1 w group.The control group was not irradiated,the right SMG of the radiation group rats was received one-time 20 Gy radiation,and the saliva secretion of each group was detected.The tissue morphology of SMG was ob-served by HE staining.The levels of malondialdehyde(MDA),reduced glutathione(GSH),and superoxide dismutase(SOD)were detected by ELISA.The positive expression of 8-OHdG was observed by IHC staining.The apoptosis of SMG cells was detected by TUNEL staining.The levels of apoptosis-related proteins BCL2-associated X protein(BAX),B-cell lymphoma-2(Bcl-2),and cysteinyl aspartate specific proteinase-3(Caspase-3)were detec-ted by western blot,and the mRNA expression was detected by RT-qPCR.Results:Salivary secretion was decreased after radiation.The pathological changes such as intracellular vacuolation,tissue edema,and gap widening were gradually increased.The cells showed obvious oxidative stress and apoptosis.The expression levels of apoptosis-related pro-teins BAX,Bcl-2,and Caspase-3 were abnormal.Conclu-sion:Post-radiation oxidative stress may be an important factor affecting apoptosis and dysfunction of SMG cells.
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