摘要
背景:胶原蛋白具有抗氧化作用,但既往在实验室主要将鼠尾胶原用于促进细胞贴壁和支架构建,对于其抗氧化作用目前尚无相关研究。目的:探讨鼠尾胶原对过氧化氢导致离体心肌细胞氧化损伤的保护作用。方法:将原代培养的SD大鼠乳鼠心肌细胞随机接种到铺有鼠尾胶原的培养皿(实验组)和普通培养皿(对照组)中,并且均用0,10,100μmol/L H2O2诱导。24 h后,以电子显微镜观察各组乳鼠心肌细胞的形态,应用MTT比色法检测心肌细胞存活率,TUNEL法检测心肌细胞凋亡形态,流式细胞技术检测心肌细胞凋亡率,黄嘌呤氧化酶法检测超氧化物歧化酶活力,硫代巴比妥比色法检测丙二醛水平,Western-Blot检测凋亡蛋白Bax、Bcl-2的表达。结果与结论:两组培养皿中,随着过氧化氢浓度的增高,均可见细胞凋亡状态明显加重,细胞存活率及细胞内过氧化氢活力明显下降,细胞凋亡率及细胞内丙二醛水平明显增高,Bcl-2/Bax比值显著降低,呈剂量依赖性。而在相同浓度过氧化氢诱导下,与对照组相比,实验组细胞凋亡状态明显减弱,细胞存活率、超氧化物歧化酶活力及Bcl-2/Bax比值增高,细胞凋亡率和丙二醛水平明显减低。表明鼠尾胶原对过氧化氢所致的心肌细胞氧化损伤具有保护作用,其机制可能与提高超氧化物歧化酶活力,减少丙二醛产生及提高Bcl-2的表达有关。
BACKGROUND： Although collagen has been approved to have antioxidant effect, there is no relevant research at the cell level. Previous experimental studies have focused on the effect of rat tail tendon collagens to promote cell adhesion and scaffold construction.
OBJECTIVE： To investigate the protective role of rat tail tendon collagen in oxidative stress of cardiomyocytes induced by hydrogen peroxide.
METHODS： Primary neonatal rat cardiomyocytes were seeded in culture dish with or without rat tail tendon collagen randomly, then both treated with various concentration of H2O2 （0, 10, 100 μmol/L） for 24 hours. Then, the morphology of cardiomyocytes was observed by inverted fluorescence microscope, the viability of cardiomyocytes wasmeasured by MTT assay, TUNEL was used to detect the apoptosis of cardiomyocytes, the percentage of apoptotic cells was determined by flow cytometry, superoxide dismutase activity was detected by xanthine oxidase method, glucosinolates barbitone colorimetry was used to detect the contents of malonaldehyde and western blot assay was used to detect the expression of Bax and Bcl-2.
RESULTS AND CONCLUSION： With the increasing of the concentration of H2O2, cardiomyocytes cultured with or without rat tail tendon collagen were observed severer in apoptotic state; the cell viability and superoxide dismutase activity were significantly reduced; the percentage of apoptotic cells and content of malonaldehyde were significantly increased; the Bcl-2/Bax ratio was reduced in dose-dependent manner. In the same concentration of H2O2, compared with the cells cultured without rat tail tendon collagen, the apoptotic state of cells cultured with rat tail tendon collagen was significantly relieved, the viability of cardiomyocytes, superoxide dismutase activity and Bcl-2/Bax ratio were higher, and the apoptosis rate and malonaldehyde content were reduced. These findings indicate that rat tail tendon collagen can protect cardiomyocytes against H2O2 induced oxidative stress injury through improvement of superoxide dismutase activity and Bcl-2 expression and reduction of malonaldehyde content.
出处
《中国组织工程研究》
CAS
CSCD
2014年第52期8463-8469,共7页
Chinese Journal of Tissue Engineering Research
基金
国家自然科学基金(81060021)~~
