本文選題：MPPa + 光動(dòng)力療法　； 參考：《重慶醫(yī)科大學(xué)》2017年碩士論文

【摘要】：目的:探討焦脫鎂葉綠酸-a甲酯(pyropheophorbide-a methyl ester,MPPa)介導(dǎo)的光動(dòng)力療法(photodynamic therapy,PDT)對(duì)人骨肉瘤MG63細(xì)胞凋亡的影響及其作用機(jī)制方法:取對(duì)數(shù)生長期人骨肉瘤MG63細(xì)胞分為4組,分別為空白對(duì)照組(對(duì)照組)、單純MPPa藥物組(MPPa組)、單純光照組(LED組)以及MPPa-PDT實(shí)驗(yàn)組(MPPa-PDT組)。MPPa-PDT組和MPPa組于避光條件下加入MPPa(0.75μmol/L),對(duì)照組及LED組加入等量完全培養(yǎng)基;避光孵育20 h后,LED組及MPPa-PDT組細(xì)胞接受集成LED特種光源照射120 s(劑量為4.8 J/cm2)。光照結(jié)束后,于避光條件下繼續(xù)培養(yǎng),倒置相差顯微鏡觀察細(xì)胞形態(tài)變化,透射電鏡觀察內(nèi)質(zhì)網(wǎng)形態(tài)變化,Hoechst33258染色觀察細(xì)胞凋亡,流式細(xì)胞術(shù)檢測細(xì)胞凋亡率及細(xì)胞內(nèi)鈣離子水平,Western blot檢測p-PERK、內(nèi)質(zhì)網(wǎng)源性轉(zhuǎn)錄因子(C/EBP homologous protein,CHOP)、活化半胱氨酸蛋白酶12(cleaved-Caspase-12)表達(dá)水平。結(jié)果:MPPa-PDT組倒置相差顯微鏡下見細(xì)胞明顯回縮變圓;Hoechst33258染色示細(xì)胞核固縮、碎裂等典型凋亡形態(tài)學(xué)改變;流式細(xì)胞術(shù)檢測細(xì)胞凋亡率為48.76%±3.54%,明顯高于對(duì)照組5.04%±0.41%、MPPa組5.33%±0.38%及LED組6.48%±0.46(P0.05);細(xì)胞內(nèi)鈣離子水平為485.29±58.77,顯著高于對(duì)照組(97.24±4.77)、MPPa組(97.95±6.30)、LED組(101.17±5.26)(P0.05);透射電鏡下見細(xì)胞內(nèi)質(zhì)網(wǎng)明顯腫脹;Western blot檢測示p-PERK、CHOP、cleaved-Caspase-12相對(duì)表達(dá)量分別于培養(yǎng)1、3、6 h后明顯高于其他3組(P0.05)。對(duì)照組、MPPa組及LED組細(xì)胞無顯著凋亡形態(tài)改變和內(nèi)質(zhì)網(wǎng)形態(tài)改變,上述檢測指標(biāo)組間比較,差異均無統(tǒng)計(jì)學(xué)意義(P0.05)。結(jié)論:MPPa-PDT可誘導(dǎo)人骨肉瘤MG63細(xì)胞發(fā)生凋亡,且內(nèi)質(zhì)網(wǎng)應(yīng)激反應(yīng)參與了MPPa-PDT誘導(dǎo)細(xì)胞凋亡的過程。
[Abstract]:Objective: to investigate the effect of photodynamic therapeutic therapy (photodynamic therapeutic therapy) on apoptosis of human osteosarcoma MG63 cells in logarithmic phase. Methods: human osteosarcoma MG63 cells in logarithmic phase were divided into 4 groups. They were blank control group (control group), MPPA drug group (MPPA group), light group (LED group), MPPa-PDT experimental group (MPPa-PDT group). MPPa-PDT group and MPPA group were added MPPA (0.75 渭 mol / L) in dark condition. The cells of LED group and MPPa-PDT group were exposed to integrated LED special light source for 120 s (dose 4.8 J / cm2). After illumination, cell morphology was observed by inverted phase contrast microscope, and cell apoptosis was observed by Hoechst33258 staining under transmission electron microscope. Apoptosis rate and intracellular calcium level were detected by flow cytometry. P-PERK, C / EBP homologous protein chop and cleaved-Caspase-12 were detected by Western blot. Results under the inverted phase contrast microscope, Hoechst33258 staining showed typical apoptotic morphological changes, such as nuclear pyknosis and fragmentation. The apoptotic rate detected by flow cytometry was 48.76% 鹵3.54, which was significantly higher than that in the control group (5.04% 鹵0.41g) MPPA group (5.33% 鹵0.38%) and LED group (6.48% 鹵0.46) (P0.05), and the intracellular calcium ion level was 485.29 鹵58.77, which was significantly higher than that in the control group (97.24 鹵4.77) MPPA group (97.95 鹵6.30) LED group (101.17 鹵5.26) (P0.05). The relative expression of p-PERKG CHOPOPcleaved-Caspase-12 was significantly higher than that of the other three groups after 6 h culture (P0.05). There were no significant changes in apoptotic morphology and endoplasmic reticulum (ER) morphology in MPPA group and LED group in control group. There was no significant difference between the two groups (P0.05). Conclusion the apoptosis of human osteosarcoma MG63 cells can be induced by 1: MPPa-PDT, and endoplasmic reticulum stress is involved in the process of MPPa-PDT induced apoptosis of human osteosarcoma MG63 cells.
【學(xué)位授予單位】：重慶醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類號(hào)】：R738

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