【摘要】：目的光動(dòng)力治療是近年來(lái)新興起的一種腫瘤治療方式,光敏劑是光動(dòng)力治療的關(guān)鍵。本研究旨在明確葉綠酸f光動(dòng)力學(xué)體外殺傷膀胱癌細(xì)胞的效果及其可能機(jī)制。方法體外培養(yǎng)膀胱癌5637和T24細(xì)胞,CCK-8法檢測(cè)葉綠酸f結(jié)合650nm激光對(duì)膀胱癌細(xì)胞的生長(zhǎng)抑制作用。葉綠酸f和特異性線粒體熒光探針同細(xì)胞共孵育4h后,激光共聚焦顯微鏡觀察其亞細(xì)胞分布;DAPI染色PDT后12h的癌細(xì)胞,熒光顯微鏡下觀察細(xì)胞形態(tài)學(xué)改變;采用流式細(xì)胞儀分析凋亡率。結(jié)果 4J/cm2激光照射后,1、2和4μg/mL的葉綠酸f對(duì)5637細(xì)胞生長(zhǎng)抑制率分別為33.92%、57.38%和84.88%,在2J/cm2激光照射后分別為25.4%、51.21%和70.09%;對(duì)T24細(xì)胞,2.5、5和10μg/mL濃度藥物4J/cm2生長(zhǎng)抑制率分別為34.03%、60.11%和86.51%,2J/cm2分別為31.79%、53.83%和72.89%。激光共聚焦顯微鏡觀察到葉綠酸f的紅色熒光與線粒體探針的綠色熒光分布范圍一致,都在細(xì)胞質(zhì)中,細(xì)胞核中無(wú)分布。DAPI染色后熒光顯微鏡下可見處理組細(xì)胞均出現(xiàn)胞核邊緣不規(guī)則,核固縮,核溶解、碎裂,核小體碎片增加等現(xiàn)象,空白對(duì)照組細(xì)胞則未見到此類凋亡表現(xiàn)。流式細(xì)胞術(shù)檢測(cè)結(jié)果顯示,4μg/mL濃度結(jié)合4J/cm2激光處理組5637細(xì)胞凋亡率為(50.61±1.66)%,對(duì)照組凋亡率為(4.05±0.12)%,差異有統(tǒng)計(jì)學(xué)意義,U=6.21,P=0.042;10μg/mL濃度結(jié)合4J/cm2組T24細(xì)胞凋亡率為(54.3±1.32)%,對(duì)照組凋亡率為(11.31±0.71)%。差異有顯著性意義,U=7.35,P=0.030。結(jié)論葉綠酸f結(jié)合650nm激光,能夠高效殺滅膀胱癌細(xì)胞,該殺滅效應(yīng)隨藥物劑量及激光能量的增加明顯升高。殺滅機(jī)制可能是藥物進(jìn)入癌細(xì)胞后定位于線粒體,誘導(dǎo)腫瘤細(xì)胞凋亡。
[Abstract]:Objective photodynamic therapy is a new method of tumor therapy in recent years. Guang Min is the key of photodynamic therapy. The aim of this study was to determine the photodynamic effect and possible mechanism of chloric acid f on bladder cancer cells in vitro. Methods 5637 and T24 cells of bladder cancer were cultured in vitro. The growth inhibitory effect of folate f combined with 650nm laser on bladder cancer cells was detected by CCK-8 method. The distribution of subcells was observed by laser confocal microscopy after incubating with chloric acid f and specific mitochondrial fluorescence probe for 4 h, and the morphological changes of cancer cells 12 h after PDT staining by DAPI were observed under fluorescence microscope. Apoptosis rate was analyzed by flow cytometry. Results after 4J/cm2 laser irradiation, the growth inhibition rates of 5637 cells were 33.92% and 84.88%, respectively, which were 51.21% and 70.09% after 2J/cm2 laser irradiation, and 34.0360.11% and 86.51J / cm 2 for T24 cells with 2.55 渭 g/mL and 10 渭 g/mL concentrations, respectively, and 72.89% for T24 cells, and 86.51J / cm2 and 72.89cm ~ 2 for T24 cells, respectively.The inhibitory rates were 57.38% and 84.88%, respectively, for T24 cells, and for T24 cells, the inhibitory rates of 2.55 渭 g/mL and 10 渭 g/mL concentrations were 34.0360.11% and 86.51J / cm ~ 2, respectively, and 72.89%, respectively. Laser confocal microscopy showed that the red fluorescence of chloric acid f was consistent with the green fluorescence of mitochondrial probe, and both of them were in the cytoplasm. After DAPI staining, the cells in the treated group showed irregular nuclear edge, pyknosis, nucleolysis, fragmentation and increase of nucleosome fragments, but no such apoptosis was observed in the blank control group. The apoptosis rate of 5637 cells was (50.61 鹵1.66)% in 4 渭 g/mL and (4.05 鹵0.12)% in control group. The apoptosis rate of T24 cells was (54.3 鹵1.32)% and (11.31 鹵0.71)% in 4J/cm2 group. There was a significant difference between the two groups. Conclusion folic acid f combined with 650nm laser can effectively kill bladder cancer cells, and the killing effect is obviously increased with the increase of drug dose and laser energy. The mechanism of killing may be that the drug is located in mitochondria after entering cancer cells and induces apoptosis of tumor cells.
【作者單位】： 上海市浦東新區(qū)公利醫(yī)院泌尿外科;
【基金】：國(guó)家自然科學(xué)基金(81272845) 上海市浦東新區(qū)科技發(fā)展基金(PKJ2015-Y21) 上海市衛(wèi)計(jì)委重點(diǎn)學(xué)科資助(ZK2015A11)
【分類號(hào)】：R737.14

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