本文選題：晶狀體上皮細(xì)胞 + 后發(fā)性白內(nèi)障　； 參考：《山西醫(yī)科大學(xué)》2010年碩士論文

【摘要】： 目的：通過(guò)體外細(xì)胞培養(yǎng)研究蛋白酶體抑制劑MG132對(duì)HLEC增殖、移行和分化的抑制作用,初步探索MG132防治后發(fā)性白內(nèi)障的機(jī)制。 方法： 1、采用組織塊培養(yǎng)法,對(duì)人晶狀體前囊膜進(jìn)行培養(yǎng),并利用形態(tài)學(xué)觀察和體外培養(yǎng)HLEC特異性的“晶狀體小體”進(jìn)行細(xì)胞鑒定。 2、MTT比色法：傳代的HLEC分別加入FGF-2(10ng/ml); MG132(10nM); MG132 +FGF-2(FGF-2, 10ng/ml; MG132, 10μM)并作對(duì)照,培養(yǎng)24h后以MTT法測(cè)定吸光度(OD)值,計(jì)算其增生率。 3、移行能力測(cè)定：傳代的HLEC分別加入FGF-2(10ng/ml); MG132(10μM); MG132 +FGF-2(FGF-2, 10ng/ml; MG132, 10μM)并作對(duì)照,用無(wú)菌棉棒在細(xì)胞層中劃出細(xì)胞裸露區(qū),培養(yǎng)24h后對(duì)行至裸露區(qū)的細(xì)胞計(jì)數(shù),計(jì)算移行能力。 4、細(xì)胞免疫組化法：收集傳代的HLEC分別加入TGF-β2(10ng/ml); MG132(10μM); MG132+TGF-β2(TGF-β2, 10ng/ml; MG132,10μM)并作對(duì)照,培養(yǎng)24h后免疫組化檢測(cè)胞漿中FN的表達(dá)。 結(jié)果： 1、HLEC原代培養(yǎng)中,細(xì)胞接種后24h內(nèi),即有部分上皮細(xì)胞自囊膜片生長(zhǎng)移出,貼壁生長(zhǎng)。長(zhǎng)期培養(yǎng)的HLEC中,可觀察到細(xì)胞呈同心圓環(huán)繞的小球體狀結(jié)構(gòu),即體外培養(yǎng)LEC特征性的“晶狀體小體”。 2、10ng/ml FGF-2能誘導(dǎo)HLEC增殖24.2%；10μM的MG132能夠有效抑制HLEC增殖,增殖率為-25.4%且能夠有效抑制FGF-2誘導(dǎo)的HLEC增殖,增殖率為-20.1%。MG132+FGF-2組和FGF-2組對(duì)HLEC的作用有顯著性差異(P0.05)。 3、10ng/ml FGF-2能誘導(dǎo)HLEC移行,移行能力為33.6%,10μM的MG132能夠強(qiáng)有效地抑制FGF-2誘導(dǎo)HLEC的移行作用。MG132+FGF-2組和FGF-2組對(duì)HLEC的作用有顯著性差異(P0.05)。 4、TGF-β2組可見(jiàn)HLEC胞漿中多量黃褐色沉積物；而MG132+TGF-β2組FN的表達(dá)明顯下降,與對(duì)照組相比無(wú)明顯差異；MG132組接近對(duì)照組。 結(jié)論：1.本實(shí)驗(yàn)成功地進(jìn)行了HLEC的體外培養(yǎng),獲得了高純度的HLEC細(xì)胞,可以作為后發(fā)障的細(xì)胞學(xué)研究的實(shí)驗(yàn)對(duì)象。2.無(wú)論FGF-2、TGF-β2存在與否,MG132能強(qiáng)有效抑制HLEC增殖、移行和分化。
[Abstract]:Aim: to investigate the inhibitory effect of proteasome inhibitor MG132 on the proliferation, migration and differentiation of HLEC in vitro, and to explore the mechanism of MG132 in preventing and treating post-cataract. Methods: 1. The anterior capsule of human lens was cultured by tissue mass culture, and the HLEC specific "lens bodies" were identified by morphological observation and in vitro culture. 2MTT colorimetric method: the HLEC was added to FGF-2G / ml; MG132N / 10nMN; MG132 FGF-2FGF-2FGF-2FGF-2FGF-2, 10 渭 M) and compared with the control. After 24 hours of culture, the absorbance of HLEC was determined by MTT method, and the proliferation rate was calculated. 3. Translocation ability: HLEC was added into FGF-2G / ml; MG132(10 渭 Mel; MG132 FGF-2FGF-2FGF-2,10ng / ml; MG132,10 渭 Mrespectively. The bare areas of cells were marked out in the cell layer with sterile cotton rods. After 24 hours of culture, the number of cells moving to the exposed area was counted and the transference ability was calculated. (4) Immunohistochemical method: the cultured HLEC was added to TGF- 尾 _ 2G / ml; MG132(10 渭 M; MG132 TGF- 尾 _ 2 to TGF- 尾 _ 2, 10 ng / ml; MG132C _ (10 渭 M), respectively. The expression of FN in the cytoplasm was detected by immunohistochemistry for 24 hours. Results: 1 in primary culture of HLEC, within 24 hours after inoculation, some epithelial cells grew out of the capsule and grew on the wall. In HLEC cultured for a long time, it was observed that the cells were concentric round small globular structure, that is, the characteristic "lens bodies" of LEC cultured in vitro. 210 ng / ml FGF-2 could induce HLEC proliferation 24.2um 10 渭 M MG132 could effectively inhibit the proliferation of HLEC, the proliferation rate was -25.4% and could effectively inhibit the HLEC proliferation induced by FGF-2. The proliferation rate was -20.1. MG132 FGF-2 group and FGF-2 group had significant effect on HLEC (P 0.05). 310 ng / ml FGF-2 could induce HLEC migration. MG132 with a migration capacity of 33.6 渭 M and 10 渭 M could strongly and effectively inhibit HLEC migration induced by FGF-2. There was significant difference between MG132 FGF-2 group and FGF-2 group in HLEC. 4TGF- 尾 2 group showed a large amount of yellow brown sediment in the cytoplasm of HLEC, while the expression of FN in MG132 TGF- 尾 2 group was significantly decreased, and there was no significant difference between MG132 group and control group. Conclusion 1. In this experiment, HLEC cells were successfully cultured in vitro, and high purity HLEC cells were obtained. Whether FGF-2TGF- 尾 2 exists or not, MG132 can effectively inhibit the proliferation, migration and differentiation of HLEC.
【學(xué)位授予單位】：山西醫(yī)科大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2010
【分類(lèi)號(hào)】：R776.1

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