【摘要】：目的研究血小板源性生長因子-α受體沉默對人晶狀體上皮細(xì)胞增殖及凋亡的影響,為治療后發(fā)性白內(nèi)障提供實驗依據(jù)。 方法1、體外培養(yǎng)人晶狀體上皮細(xì)胞株SRA01/04,使用陽離子脂質(zhì)體將合成的血小板源性生長因子-α受體反義寡核苷酸(PDGFR-α ASODN)轉(zhuǎn)染入細(xì)胞株SRA01/04,倒置顯微鏡下觀察SRA01/04的形態(tài)學(xué)改變。2、按照實驗分組(對照組、錯義寡核苷酸組、反義寡核苷酸組)PDGFR-α ASODN作用于SRA01/04細(xì)胞24h、48h、72h后,MTT法檢測細(xì)胞的增殖。3、根據(jù)實驗分組將PDGFR-α ASODN作用于SRA01/04細(xì)胞48h后,流式細(xì)胞儀檢測PDGFR-α ASODN對SRA01/04細(xì)胞周期及細(xì)胞凋亡的影響；Hoechst33258熒光染料染色法分析細(xì)胞凋亡；4、RT-PCR檢測PDGFR-α ASODN作用48h后PDGF-α受體表達(dá)的變化情況。 結(jié)果1、PDGFR-α ASODN轉(zhuǎn)染SRA01/04細(xì)胞48h后,實驗組大部分細(xì)胞皺縮、變圓、甚至碎裂,死亡細(xì)胞漂浮于培養(yǎng)液中,部分活細(xì)胞分裂相變少、貼壁不牢、胞體邊界模糊,有的甚至失去了原來的形狀；與低濃度藥物組比較,高濃度藥物組對細(xì)胞的抑制作用增強。2、PDGFR-α ASODN作用于SRA01/04細(xì)胞24h～72h, SRA01/04細(xì)胞增殖受抑制,且隨時間的延長抑制作用增強,實驗組較對照組和錯義寡核苷酸組比較對細(xì)胞的抑制作用增強,差異有統(tǒng)計學(xué)意義(P㩳0.05)。3、PDGFR-α ASODN能誘導(dǎo)SA.R01/04細(xì)胞凋亡,PDGFR-α ASODN作用于SRA01/04細(xì)胞48h后實驗組細(xì)胞凋亡率分別為(3.22±0.25)%、(5.29±0.27)%,與對照組(0.75±0.67)%和錯義寡核苷酸組(1.46±0.60)%比較,差異有統(tǒng)計學(xué)意義(P0.05)；實驗組G1期細(xì)胞分布率分別為(53.31±1.30)%、(59.98±0.95)%,與對照組(47.73±1.18)%和錯義寡核苷酸組(49.48±1.09)%比較,差異有統(tǒng)計學(xué)意義(P0.05)。4、PDGFR-α ASODN作用于SRA01/04細(xì)胞48h后,Hochest33258染色結(jié)果表明：實驗組細(xì)胞核固縮、核濃染、致密凋亡細(xì)胞與對照組比較增多。5、PDGFR-α ASODN作用于SRA01/04細(xì)胞48h后,PDGF-α受體在SRA01/04中表達(dá)下調(diào)。 結(jié)論1、PDGF-α受體反義寡核苷酸能抑制人晶狀體上皮細(xì)胞的增殖,誘導(dǎo)其凋亡。2、PDGF-α受體反義寡核苷酸沉默SRA01/04中PDGF-α受體基因的表達(dá)。3、PDGF-α受體在人晶狀體上皮細(xì)胞增殖過程中起重要作用。
[Abstract]:Objective to study the effect of platelet-derived growth factor-偽 receptor silencing on the proliferation and apoptosis of human lens epithelial cells. Methods 1. Human lens epithelial cell line SRA01/04, was transfected into SRA01/04, cell line by using cationic liposome to transfect the synthesized antisense oligodeoxynucleotide of platelet-derived growth factor- 偽 receptor (PDGFR- 偽 ASODN) into the cell line SRA01/04, in vitro. The morphologic changes of SRA01/04 were observed under inverted microscope. (2) PDGFR- 偽 ASODN (control group, missense oligonucleotide group, antisense oligonucleotide group) was treated with PDGFR- 偽 ASODN for 24 h, 48 h and 72 h, respectively. MTT assay was used to detect the proliferation of SRA01/04 cells. 3. After treated with PDGFR- 偽 ASODN for 48 h, the effects of PDGFR- 偽 ASODN on SRA01/04 cell cycle and apoptosis were detected by flow cytometry. Hoechst33258 fluorescent dye staining was used to detect apoptosis, and RT-PCR was used to detect the expression of PDGF- 偽 receptor after 48 h of PDGFR- 偽 ASODN treatment. Results 1After transfection of PDGFR- 偽 ASODN into SRA01/04 cells for 48 h, most of the cells in the experimental group were shrinked, rounded or even broken, the dead cells were floating in the culture medium, some of the living cells had less mitotic phase change, the adherent was not firm, and the boundaries of the cells were blurred. Some even lost their original shape; Compared with the low concentration drug group, the inhibitory effect of high concentration drug group on SRA01/04 cells was enhanced. 2 PDGFR- 偽 ASODN inhibited the proliferation of SRA01/04 cells for 24 h and 72 h, and the inhibitory effect was enhanced with the prolongation of time. Compared with the control group and the missense oligonucleotide group, the inhibitory effect of the experimental group on SA.R01/04 cells was significantly increased (P0. 05). 3 PDGFR- 偽 ASODN could induce the apoptosis of SA.R01/04 cells. After treated with PDGFR- 偽 ASODN for 48 h, the apoptotic rates of SRA01/04 cells in the experimental group were (3.22 鹵0.25)% and (5.29 鹵0.27)%, respectively, compared with those in the control group (0.75 鹵0.67)% and missense oligonucleotide group (1.46 鹵0.60)%. The difference was statistically significant (P0.05). The distribution rates of G1 phase cells in the experimental group were (53.31 鹵1.30)% and (59.98 鹵0.95)%, respectively, compared with those in the control group (47.73 鹵1.18)% and missense oligonucleotide group (49.48 鹵1.09)%. The difference was statistically significant (P0.05). (4) after treated with PDGFR- 偽 ASODN for 48 h, the results of Hochest33258 staining showed that the nuclei of the experimental group were pyknosis, dense staining and dense apoptotic cells were more than those of the control group. After PDGFR- 偽 ASODN was treated with SRA01/04 cells for 48 h, the expression of PDGF- 偽 receptor was down-regulated in SRA01/04. Conclusion 1 the antisense oligonucleotide of PDGF- 偽 receptor can inhibit the proliferation of human lens epithelial cells and induce its apoptosis. 2 the antisense oligonucleotide of PDGF- 偽 receptor silenced the expression of PDGF- 偽 receptor gene in SRA01/04. PDGF- 偽 receptor plays an important role in the proliferation of human lens epithelial cells.
【學(xué)位授予單位】：桂林醫(yī)學(xué)院
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2012
【分類號】：R776.1

【參考文獻(xiàn)】

相關(guān)期刊論文 前10條

1 宋艷萍,丁琴,劉瓊,趙霞;Nd:YAG激光切除兒童后發(fā)性白內(nèi)障50例[J];國際眼科雜志;2005年03期

2 陶津華;安小玲;孔郡;張瑞君;張勁松;;EGF體外對人晶狀體上皮細(xì)胞遷移的影響[J];國際眼科雜志;2006年01期

3 王建明;孫乃學(xué);馮海曉;熊蕾;惠娜;范雅稚;;Nd∶YAG激光兩種切開方法治療兒童后發(fā)性白內(nèi)障的臨床研究[J];國際眼科雜志;2008年08期

4 嚴(yán)浩;邱慶華;王艷麗;;EGFR反義寡核苷酸對人視網(wǎng)膜神經(jīng)膠質(zhì)細(xì)胞遷移的影響[J];國際眼科雜志;2010年08期

5 邱梅園;彭燕一;黃嵐珍;張玉明;向秋;;PDGF-α受體反義寡核苷酸對體外視網(wǎng)膜色素上皮細(xì)胞增殖和凋亡的影響[J];國際眼科雜志;2011年02期

6 柯治生;蔡小軍;鄧小艷;;白細(xì)胞介素-1β反義寡核苷酸脂質(zhì)體抑制后發(fā)性白內(nèi)障的實驗研究[J];武漢大學(xué)學(xué)報(醫(yī)學(xué)版);2006年02期

7 倪琦;曾思恩;譚寧;郭芳;張美艷;;姜黃素對肝癌Hep1細(xì)胞增殖及凋亡的影響[J];山東醫(yī)藥;2010年07期

8 孔s，

本文編號：2379974