本文選題：角質(zhì)細(xì)胞生長(zhǎng)因子-2 + 克隆��； 參考：《吉林大學(xué)》2010年博士論文

【摘要】： 角質(zhì)細(xì)胞生長(zhǎng)因子-2(KGF-2)是成纖維細(xì)胞生長(zhǎng)因子家族的一員。KGF-2對(duì)上皮組織具有高度特異性,可加快上皮及基質(zhì)細(xì)胞的修復(fù),且不導(dǎo)致膠原的過(guò)度增生,對(duì)瘢痕的減少有積極的意義。雖然角膜細(xì)胞可分泌一定量的KGF-2,但及時(shí)修復(fù)仍需補(bǔ)充外源KGF-2,以促進(jìn)角膜上皮細(xì)胞及基質(zhì)細(xì)胞的增殖和移行。因此,開(kāi)發(fā)KGF-2產(chǎn)品,研究其對(duì)角膜損傷的作用及機(jī)制,對(duì)于拓展KGF-2臨床應(yīng)用具有重大意義。 本研究利用大腸桿菌原核表達(dá)系統(tǒng)成功構(gòu)建了KGF-2工程菌株,對(duì)KGF-2發(fā)酵及純化工藝進(jìn)行了研究,利用正交實(shí)驗(yàn)設(shè)計(jì)篩選到KGF-2滴眼液的保護(hù)劑配方,建立了KGF-2體外生物學(xué)活性的測(cè)定方法,以日本白兔堿燒傷模型和激光損傷模型研究了KGF-2體內(nèi)促角膜損傷修復(fù)作用及機(jī)制。動(dòng)物實(shí)驗(yàn)結(jié)果顯示,應(yīng)用不同濃度KGF-2進(jìn)行治療后,1周時(shí)各種損傷表現(xiàn)均較損傷對(duì)照組明顯減輕,角膜層厚度接近正常,水腫基本消退,且無(wú)明顯的炎癥反應(yīng);傷后2周無(wú)炎癥反應(yīng)復(fù)發(fā),角膜上皮細(xì)胞生長(zhǎng)良好,排列整齊,基質(zhì)層內(nèi)有較多膠原纖維,板層纖維排列整齊且與角膜上皮平行排列,與正常角膜結(jié)構(gòu)沒(méi)有明顯差別,實(shí)驗(yàn)結(jié)果提示KGF-2可明顯減輕角膜基質(zhì)水腫,抑制角膜基質(zhì)成纖維細(xì)胞過(guò)度增生,抑制炎性細(xì)胞浸潤(rùn),有利于角膜上皮細(xì)胞的再生與修復(fù)。陽(yáng)性對(duì)照藥bFGF治療組在治療2周后角膜上皮過(guò)度增生呈波浪狀,表面凹凸不平,基質(zhì)中有較多成纖維細(xì)胞增生,且板層纖維亦呈波浪狀。本實(shí)驗(yàn)結(jié)果提示,KGF-2可通過(guò)加速角膜上皮細(xì)胞的再生和遷移,促進(jìn)角膜基質(zhì)纖維修復(fù)等機(jī)制,促進(jìn)堿燒傷和激光燒傷后受損角膜的修復(fù),對(duì)角膜損失具有很好的治療作用。 本課題研究為KGF-2在眼科臨床的應(yīng)用奠定了基礎(chǔ)。
[Abstract]:Keratinocyte growth factor (KGF-2) is a member of fibroblast growth factor family. KGF-2 is highly specific to epithelial tissue, can accelerate the repair of epithelial and stromal cells, and does not cause excessive proliferation of collagen, which has positive significance for the reduction of scar. Although corneal cells can secrete a certain amount of KGF-2, it is necessary to supplement exogenous KGF-2 in order to promote the proliferation and migration of corneal epithelial cells and stromal cells. Therefore, it is of great significance to develop KGF-2 products and to study its effect and mechanism on corneal injury in order to expand the clinical application of KGF-2. In this study, KGF-2 engineering strain was successfully constructed by using E. coli prokaryotic expression system. The fermentation and purification of KGF-2 were studied. The protective agent formula of KGF-2 eye drops was screened by orthogonal design. A method for determining the biological activity of KGF-2 in vitro was established. The effects and mechanisms of KGF-2 on corneal injury in vivo were studied with alkali burn model and laser injury model of Japanese white rabbit. The results of animal experiments showed that all kinds of injuries were significantly alleviated at 1 week after treatment with different concentrations of KGF-2, corneal thickness was close to normal, edema basically disappeared, and there was no obvious inflammatory reaction. There was no recurrence of inflammatory reaction at 2 weeks after injury. Corneal epithelial cells grew well and arranged neatly. There were more collagen fibers in stromal layer and lamellar fibers were arranged neatly and parallel to corneal epithelium. There was no significant difference between corneal epithelial cells and normal corneal structure. The results suggest that KGF-2 can significantly reduce corneal stromal edema, inhibit the excessive proliferation of corneal stromal fibroblasts, inhibit the infiltration of inflammatory cells, and facilitate the regeneration and repair of corneal epithelial cells. After 2 weeks of treatment, the corneal epithelial hypertrophy in the bFGF group was wavy, the surface was uneven, there were more fibroblasts proliferation in the matrix, and the lamellar fibers were also wavy. The results suggest that KGF-2 can accelerate the regeneration and migration of corneal epithelial cells, promote corneal stromal fiber repair, promote the repair of damaged cornea after alkali burn and laser burn, and have a good therapeutic effect on corneal loss. This study lays a foundation for the clinical application of KGF-2 in ophthalmology.
【學(xué)位授予單位】：吉林大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2010
【分類號(hào)】：R772.21

【引證文獻(xiàn)】

相關(guān)博士學(xué)位論文 前2條

1 高榮蓮;成纖維細(xì)胞生長(zhǎng)因子-10對(duì)激光角膜燒傷的實(shí)驗(yàn)治療及分子機(jī)制研究[D];中國(guó)人民解放軍軍事醫(yī)學(xué)科學(xué)院;2011年

2 薛萍;角質(zhì)細(xì)胞生長(zhǎng)因子-1（KGF-1）在昆蟲(chóng)細(xì)胞中的表達(dá)及其活性研究[D];吉林農(nóng)業(yè)大學(xué);2012年

相關(guān)碩士學(xué)位論文 前1條

1 高麗昌;KGF-2對(duì)角膜基質(zhì)細(xì)胞的修復(fù)作用及其相關(guān)機(jī)制研究[D];吉林農(nóng)業(yè)大學(xué);2012年

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本文編號(hào)：1831937