【摘要】：目的：通過氧化損傷標記物8-羥基脫氧鳥苷（8-OHDG）及細胞凋亡標記物半胱氨酸天冬氨酸蛋白酶-3（caspase-3）的檢測，分析心肌缺血缺氧、心肌缺血再灌注、心肌細胞氧化應激的病理變化及相關機制，在此基礎上研究氧化樟腦對氧化應激損傷后心肌細胞的保護作用。 方法：采用H9c2大鼠心肌細胞復制缺氧模型，隨機分為4組：正常組、氧化應激組、氧化應激+氧化樟腦A組、氧化應激+氧化樟腦B組。正常對照組：不作任何特殊處理；氧化應激組：將心肌細胞更換為無氧培養(yǎng)液，放入缺氧裝置內，繼續(xù)培養(yǎng)4h后，更換復氧液，再進行有氧培養(yǎng)4h，完成缺氧/復氧模型；氧化應激+氧化樟腦組：在心肌細胞缺氧/復氧完成后分別加入10ul/50ml（A組）、20ul/50ml（B組）濃度的氧化樟腦繼續(xù)培養(yǎng)6h。通過噻唑藍微量自動比色法（MTT）檢測各組心肌細胞活力。通過免疫化學染色觀察各組細胞氧化損傷標記物8-羥基脫氧鳥苷（8-OHDG）及細胞凋亡標記物半胱氨酸天冬氨酸蛋白酶-3（caspase-3）水平。統(tǒng)計學處理以Sigma Stat3.5軟件進行統(tǒng)計學處理，應用t-Test或one-wayANOVA分析，結果以Mean±SD表示，P0.05為有統(tǒng)計學差異。 結果： 1.氧化應激組H9c2心肌細胞繼續(xù)培養(yǎng)6h后進行MTT檢測。結果表明：氧化應激組吸光度值（A值）低于對照組（P0.05）。 2.氧化應激+氧化樟腦組H9c2心肌細胞繼續(xù)培養(yǎng)6h后進行MTT檢測。結果表明：氧化應激組吸光度值（A值）低于氧化應激+氧化樟腦組，氧化應激+氧化樟腦組吸光度值（A值）低于對照組（P0.05），氧化應激+氧化樟腦組較氧化應激組心肌細胞活力明顯升高。氧化應激+氧化樟腦B組心肌細胞活力高于氧化應激+氧化樟腦A組。 3.8-OhdG表達在對照組最低，氧化應激組最高，而氧化應激+氧化樟腦組較氧化應激組明顯降低，，氧化應激+氧化樟腦B組8-OhdG表達低于氧化應激+氧化樟腦A組。 4. caspase-3的蛋白表達在對照組最低，氧化應激組最高，而氧化應激+氧化樟腦組較氧化應激組明顯降低，氧化應激+氧化樟腦B組的caspase-3蛋白表達低于氧化應激+氧化樟腦A組。 結論：氧化樟腦對氧化應激損傷后心肌細胞具有保護作用，它可減輕氧化應激后心肌細胞的損傷，抑制氧化應激后心肌細胞的凋亡。
[Abstract]:Objective: to detect the oxidative damage marker 8-hydroxydeoxyguanosine (8-OHDG) and the apoptotic marker cysteine aspartate protease-3 (caspase-3) to analyze myocardial ischemia and hypoxia and myocardial ischemia-reperfusion. On the basis of pathological changes and related mechanisms of oxidative stress in cardiomyocytes, the protective effect of camphor on myocardial cells after oxidative stress was studied. Methods: hypoxia model was induced by H9c2 rat cardiomyocytes. The rats were randomly divided into 4 groups: normal group, oxidative stress oxidative camphor A group and oxidative stress oxidative camphor B group. Normal control group: without any special treatment, oxidative stress group: the cardiomyocytes were replaced with anaerobic culture medium, put into anoxic equipment, then cultured for 4 hours, then reoxygenated solution was changed, then aerobic culture was carried out for 4 hours to complete the hypoxia / reoxygenation model. Oxidative stress Oxycamphor group: after hypoxia / reoxygenation of cardiomyocytes were completed, 10ul/50ml (group A) 20 ulr / 50ml (group B) was added respectively to culture camphor for 6 h. The activity of myocardial cells in each group was detected by (MTT) with thiazolyl blue microcolorimetry. The levels of 8-hydroxydeoxyguanosine (8-OHDG) and cysteine aspartate protease -3 (caspase-3) were detected by immunocytochemical staining. Statistical processing was performed by Sigma Stat3.5 software, and t-Test or one-wayANOVA analysis was used. The result was that Mean 鹵SD was used to show statistical difference (P0.05). Results: 1. In oxidative stress group, H9c2 cardiomyocytes were cultured for 6 h and MTT was detected. The results showed that the absorbance (A value) of oxidative stress group was lower than that of control group (P0.05). H9c2 cardiomyocytes in oxidative stress group were cultured for 6 h and MTT was detected. The results showed that the absorbance value (A value) of oxidative stress group was lower than that of oxidative stress oxidative camphor group. The absorbance (A value) of oxidative stress camphor group was lower than that of control group (P0.05). The activity of myocardial cells in oxidative stress camphor group was significantly higher than that in oxidative stress group. The activity of cardiomyocytes in oxidative stress camphor B group was higher than that in oxidative stress camphor A group. The expression of 3.8-OhdG was the lowest in the control group and the highest in the oxidative stress group, while the oxidative stress oxidative camphor group was significantly lower than that in the oxidative stress group. The expression of 8-OhdG in oxidative stress camphor B group was lower than that in oxidative stress camphor A group. 4. The protein expression of caspase-3 was the lowest in the control group and the highest in the oxidative stress group, while the oxidative stress oxidative camphor group was significantly lower than that in the oxidative stress group. The expression of caspase-3 protein in oxidative stress camphor B group was lower than that in oxidative stress camphor A group. Conclusion: Oxycamphor has protective effect on cardiomyocytes after oxidative stress. It can reduce the injury of cardiomyocytes after oxidative stress and inhibit the apoptosis of cardiomyocytes after oxidative stress.
【學位授予單位】：吉林大學
【學位級別】：碩士
【學位授予年份】：2012
【分類號】：R363

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