發(fā)布時(shí)間：2018-04-30 04:25

  本文選題：增生性玻璃體視網(wǎng)膜病變 + 視網(wǎng)膜色素上皮細(xì)胞　； 參考：《吉林大學(xué)》2012年碩士論文

【摘要】：研究表明,視網(wǎng)膜色素上皮(retinal pigment epithelium, RPE)細(xì)胞是增生性玻璃體視網(wǎng)膜病變(proliferation vitreoretinopathy, PVR)的增殖膜中的主要細(xì)胞來(lái)源,因此增殖性玻璃體視網(wǎng)膜病變的研究和防治多圍繞探討RPE細(xì)胞的異常行為和抑制RPE細(xì)胞及其他相關(guān)細(xì)胞的增殖。雷帕霉素(rapamycin, RAPA)是一種新型的大環(huán)內(nèi)酯類(lèi)免疫抑制劑,眼科方面已有研究應(yīng)用于動(dòng)物及體外細(xì)胞培養(yǎng)藥物實(shí)驗(yàn)。 目的： 本研究是在體外培養(yǎng)人視網(wǎng)膜色素上皮(human retinal pigment epithelium, hRPE)細(xì)胞的基礎(chǔ)之上,探討一定范圍濃度的RAPA對(duì)體外培養(yǎng)的hRPE細(xì)胞形態(tài)、增殖等生物學(xué)活性的影響。 方法： 復(fù)蘇并體外培養(yǎng)hRPE細(xì)胞并分為不同濃度雷帕霉素實(shí)驗(yàn)組(Ong/ml,5ng/ml,10ng/ml,20ng/ml,40ng/ml,80ng/ml)。顯微鏡下觀察各組細(xì)胞及藥物不同作用時(shí)間下,細(xì)胞的增殖狀態(tài)、倍增時(shí)間、細(xì)胞活力、生長(zhǎng)曲線及四唑鹽(MTT)比色法檢RAPA對(duì)hRPE細(xì)胞增殖的影響。 結(jié)果： 復(fù)蘇后細(xì)胞活力良好的hRPE細(xì)胞株,雷帕霉素在實(shí)驗(yàn)范圍內(nèi)抑制hRPE細(xì)胞的增殖。鏡下可見(jiàn)隨雷帕霉素濃度增加,細(xì)胞數(shù)目逐漸減少。經(jīng)80ng/ml雷帕霉素作用72小時(shí)后部分細(xì)胞變形崩解,偶見(jiàn)細(xì)胞浮起。對(duì)照組細(xì)胞分裂時(shí)間為1.83天,隨RAPA藥物濃度增加細(xì)胞分裂時(shí)間隨之延長(zhǎng)。生長(zhǎng)曲線顯示隨著雷帕霉素濃度的增加,作用時(shí)間的延長(zhǎng),hRPE細(xì)胞的生長(zhǎng)有明顯的抑制作用,并呈劑量時(shí)間效應(yīng)關(guān)系。通過(guò)生長(zhǎng)曲線可見(jiàn)對(duì)照組細(xì)胞增殖旺盛,各項(xiàng)生長(zhǎng)指標(biāo)均高于用藥組。臺(tái)盼蘭細(xì)胞活力測(cè)定結(jié)果為不同濃度雷帕霉素(Ong/ml,5ng/ml,1Ong/ml,20ng/ml,40ng/ml,80ng/ml)細(xì)胞活力分別為92.3%、89.3%、88.7%、87.7%、86.0%、82.6%。MTT比色法觀察細(xì)胞生長(zhǎng)基本規(guī)律結(jié)果示：各濃度藥物組作用24小時(shí)后與對(duì)照組比較計(jì)算得抑制率,分別為8.70%、22.06%、40.69%、42.31%、43.12%；72小時(shí)時(shí)分別為12.57%、26.35%、42.23%44.15%、47.12%。 結(jié)論： 1雷帕霉素對(duì)體外培養(yǎng)的hRPE細(xì)胞增殖具有有效抑制作用,其效應(yīng)在一定范圍內(nèi)呈濃度/時(shí)間依賴(lài)性。10～80ng/ml濃度范圍內(nèi),RAPA明顯抑制hRPE細(xì)胞增殖(P0.01)；但在20～80ng/ml濃度范圍內(nèi),各組間抑制作用無(wú)顯著性差異(P0.05)。 2雷帕霉素延長(zhǎng)hRPE細(xì)胞倍增時(shí)間,降低體外培養(yǎng)的hRPE細(xì)胞活性。 3雷帕霉素在80ng/ml時(shí)細(xì)胞抑制率近50%。 4雷帕霉素可能為臨床預(yù)防和治療PVR提供新思路,提升其在眼科的應(yīng)用價(jià)值。
[Abstract]:The results show that retinal pigment epithelium (RPE) cells are the main cell sources in proliferative membrane of proliferative vitreoretinopathy (PVR) of proliferative vitreoretinopathy (PVR). Therefore, the study and prevention of proliferative vitreoretinopathy focus on the study of abnormal behavior of RPE cells and inhibition of proliferation of RPE cells and other related cells. Rapamycin (rapamycin) is a new type of macrolide immunosuppressant, which has been used in animal and in vitro cell culture experiments in ophthalmology. Objective: Based on the culture of human retinal pigment epithelium (RPE) retinal pigment epithelium, hRPE) cells in vitro, the effects of certain concentration of RAPA on the morphology and proliferation of hRPE cells in vitro were investigated. Methods: HRPE cells were resuscitated and cultured in vitro and divided into different concentrations of rapamycin experimental group: 5 ng / ml 10 ng / ml 10 ng / ml 20 ng / ml 40 ng / ml / ml 80 ng / ml / ml. The effects of RAPA on the proliferation of hRPE cells were observed under microscope. The proliferation state, doubling time, cell viability, growth curve and tetrazolium trioxide (MTT) colorimetric assay were used to detect the effect of RAPA on the proliferation of hRPE cells. Results: After resuscitation, rapamycin inhibited the proliferation of hRPE cells. Microscopically, the number of cells decreased with the increase of rapamycin concentration. After 72 hours of treatment with 80ng/ml rapamycin, some of the cells were deformed and disintegrated, and occasionally the cells floated. The cell division time of the control group was 1.83 days, and the cell division time was prolonged with the increase of RAPA concentration. The growth curve showed that with the increase of rapamycin concentration, the growth of hRPE cells was significantly inhibited with the prolongation of the action time, and showed a dose-time effect. The growth curve showed that the growth index of the control group was higher than that of the drug group. Trypan blue cell viability assay showed that the cell viability of different concentrations of rapamycin Ong / ml 5 ng / ml 1 Ong / ml + 20 ng / ml 20 ng / ml + 20 ng / ml + 40ng / ml + 80ng / ml) was 92.3% and 89.3% respectively. The cell growth was observed by MTT colorimetric method. The results showed that the inhibition rate was calculated after 24 hours of treatment in each concentration group as compared with the control group. 42.31 and 43.12 for 72 hours, 12.57 and 26.35, 42.23D.15 and 47.12. respectively, with 8.70 and 22.06 and 40.69, and 42.31 and 43.12, respectively, with 12.57 and 26.35, and 42.23D.15 and 47.12, respectively. Conclusion: 1 rapamycin had an effective inhibitory effect on the proliferation of hRPE cells cultured in vitro, and its effect was in a dose-dependent / time-dependent manner. Rapa significantly inhibited the proliferation of hRPE cells in a concentration range of 80 ng / ml, but in the range of 20~80ng/ml concentration, rapamycin inhibited the proliferation of hRPE cells in a dose-dependent manner, but in the range of 20~80ng/ml concentration, rapamycin inhibited the proliferation of hRPE cells in a dose-dependent manner. There was no significant difference in the inhibitory effect among the three groups (P 0.05). 2 rapamycin prolonged the doubling time of hRPE cells and decreased the activity of hRPE cells cultured in vitro. 3 the inhibitory rate of rapamycin on 80ng/ml was nearly 50%. Rapamycin may provide a new idea for clinical prevention and treatment of PVR, and enhance its application value in ophthalmology.
【學(xué)位授予單位】：吉林大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類(lèi)號(hào)】：R774.1

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