本文關(guān)鍵詞： 角膜膨隆 角膜成纖維細(xì)胞 力學(xué)牽拉 炎性因子 基質(zhì)金屬蛋白酶 膠原　出處：《太原理工大學(xué)》2016年博士論文　論文類型：學(xué)位論文

【摘要】：擴(kuò)張性角膜疾病是需要進(jìn)行角膜移植的第二大適應(yīng)癥,包括圓錐角膜和屈光手術(shù)后的角膜膨隆(醫(yī)源性角膜膨隆)。角膜屈光手術(shù)后角膜變薄,在眼內(nèi)壓、用力揉眼等情況下使角膜的應(yīng)變?cè)龃?同時(shí),角膜屈光手術(shù)后上皮細(xì)胞分泌炎性因子IL-1β參與傷口愈合,尤其是術(shù)后干眼癥會(huì)導(dǎo)致淚液中IL-1β的升高,使基質(zhì)層的細(xì)胞處于復(fù)雜的力學(xué)和生物學(xué)環(huán)境中。當(dāng)角膜基質(zhì)層受損后,角膜基質(zhì)細(xì)胞會(huì)分化為角膜成纖維細(xì)胞參與角膜的損傷修復(fù)和傷口愈合。圓錐角膜患者淚液中和上皮細(xì)胞TNF-α表達(dá)的升高,以及圓錐角膜患者常伴有異常揉眼的習(xí)慣,使得圓錐角膜成纖維細(xì)胞也處于復(fù)雜的力學(xué)和生物學(xué)環(huán)境。為了研究擴(kuò)張性角膜疾病的發(fā)生機(jī)制,本文對(duì)角膜成纖維細(xì)胞在IL-1β存在的條件下施加了不同幅度的力學(xué)牽拉,研究角膜成纖維細(xì)胞形態(tài)和細(xì)胞外基質(zhì)的變化。同時(shí),提取了圓錐角膜成纖維細(xì)胞,研究炎性因子TNF-α對(duì)圓錐角膜成纖維細(xì)胞基質(zhì)金屬蛋白酶及其抑制劑表達(dá)的影響。主要工作及研究結(jié)果如下:(1)體外對(duì)角膜成纖維細(xì)胞施加了不同幅度(5%、10%、15%)的力學(xué)牽拉,研究力學(xué)刺激對(duì)角膜成纖維細(xì)胞形態(tài)的影響,發(fā)現(xiàn)力學(xué)牽拉使角膜成纖維細(xì)胞的F-actin發(fā)生收縮,牽拉幅度越大F-actin收縮程度越大,使角膜成纖維細(xì)胞的遷移受到抑制。(2)探討了在炎性因子IL-1β存在時(shí),不同幅度(5%、10%)、不同持續(xù)時(shí)間(12h、24h、36h)的力學(xué)牽拉作用對(duì)角膜成纖維細(xì)胞i型膠原、lumican表達(dá)的影響。同時(shí),用明膠酶譜法和westernblot對(duì)不同幅度作用36h時(shí)角膜成纖維細(xì)胞上清液中mmp2、mmp9的表達(dá)進(jìn)行了檢測(cè)。研究發(fā)現(xiàn):力學(xué)牽拉、IL-1β短時(shí)間(12h)單獨(dú)作用能降低角膜成纖維細(xì)胞Ⅰ型膠原的合成,這可能是角膜成纖維細(xì)胞對(duì)其做出的的應(yīng)急反應(yīng)。低幅度(5%)牽拉單獨(dú)作用較長(zhǎng)時(shí)間(24h、36h)能促進(jìn)角膜成纖維細(xì)胞Ⅰ型膠原的合成,高幅度(10%)牽拉則抑制Ⅰ型膠原的合成。IL-1β與力學(xué)牽拉共同作用使角膜成纖維細(xì)胞Ⅰ型膠原合成降低。力學(xué)牽拉短時(shí)間單獨(dú)作用時(shí),低幅度能促進(jìn)lumican的表達(dá),高幅度卻抑制其表達(dá);IL-1β與力學(xué)牽拉共同作用能促進(jìn)lumican的表達(dá)。IL-1β能誘導(dǎo)角膜成纖維細(xì)胞mmp9的表達(dá),并促進(jìn)mmp2的表達(dá);力學(xué)牽拉能進(jìn)一步促進(jìn)IL-1β引起的mmp2表達(dá)的升高。說明炎性因子與力學(xué)牽拉共同作用使角膜成纖維細(xì)胞膠原合成減少、基質(zhì)降解增多,可能會(huì)引起醫(yī)源性角膜膨隆的發(fā)生。(3)圓錐角膜患者淚液中TNF-α表達(dá)的升高以及上皮細(xì)胞TNF-α的高表達(dá),使圓錐角膜成纖維細(xì)胞處于炎癥環(huán)境中,通過對(duì)圓錐角膜成纖維細(xì)胞施加不同濃度的TNF-α作用,發(fā)現(xiàn)TNF-α能促進(jìn)圓錐角膜成纖維細(xì)胞mmps的表達(dá),而抑制其timps的表達(dá)。MMPs/TIMPs的失衡將導(dǎo)致圓錐角膜成纖維細(xì)胞外基質(zhì)的降解增多,進(jìn)而可能導(dǎo)致角膜組織結(jié)構(gòu)的完整性遭受破壞。(4)對(duì)圓錐角膜成纖維施加TNF-α和力學(xué)牽拉的作用,研究發(fā)現(xiàn)兩者共同作用使圓錐角膜成纖維細(xì)胞的增殖受到抑制,細(xì)胞凋亡不受影響。角膜成纖維細(xì)胞數(shù)目的減少可能會(huì)使細(xì)胞外基質(zhì)的合成降低,角膜的生物力學(xué)性能發(fā)生改變。綜上所述:炎性因子和力學(xué)刺激通過調(diào)節(jié)細(xì)胞外基質(zhì)的代謝、角膜成纖維細(xì)胞的增殖在擴(kuò)張性角膜疾病的發(fā)生中起著重要的作用。
[Abstract]:The expansion of corneal diseases is the need for corneal transplantation second indications, including keratoconus after refractive surgery and corneal ectasia (iatrogenic corneal ectasia). After corneal refractive surgery and corneal thinning in intraocular pressure, eye rubbing force under the condition that the corneal strain increases; at the same time, corneal refractive surgery after the epithelial cells secrete inflammatory factor IL-1 is involved in wound healing, especially postoperative dry eye would lead to an increase in tear IL-1 beta, the stroma cells in mechanical and biological environment in the complex. When the cornea is damaged, corneal stromal cells can differentiate into corneal fibroblasts in corneal damage and repair wound healing. Tears of patients with keratoconus and expression of epithelial cells of TNF- alpha increased, and keratoconus patients often accompanied by abnormal Rouyan habits, the keratoconus fibroblasts are complex and Mechanical Biological environment. In order to study the pathogenesis of corneal expansion diseases, the corneal fibroblasts in the presence of beta IL-1 applied mechanics of different amplitude changes of corneal stretch, morphology and cell extracellular matrix fibers. At the same time, the extraction of keratoconus fibroblasts on inflammatory factor TNF- alpha effect of expression of matrix metalloproteinase and its inhibitor of keratoconus. The main work and research results are as follows: (1) in vitro on corneal fibroblasts was applied into different ranges (5%, 10%, 15%) of the mechanical stretch of mechanical stimulation effect of fibroblast morphology on the cornea, and found that mechanical stretch. Corneal fibroblasts F-actin shrink, stretch the bigger the contraction of F-actin is greater, the corneal fibroblast migration was inhibited. (2) discussed in the presence of inflammatory factor IL-1 beta, different The amplitude (5%, 10%), different duration (12h, 24h, 36h) the mechanical stretch effect on corneal fibroblast collagen type I, lumican expression. At the same time, and Westernblot of different amplitude 36h corneal fibroblast MMP2 in the supernatant by gelatin zymography, and the expression of MMP9 was detected study found that: the mechanical stretch, IL-1 beta short time (12h) alone can reduce corneal collagen synthesis in fibroblasts, which may be the response of corneal fibroblasts to make its low amplitude. (5%) stretch alone a longer period of time (24h, 36h) can promote corneal type I collagen synthesis in fibroblasts, high amplitude (10%) led by the synthesis of.IL-1 beta and mechanical pull inhibited collagen stretch to make corneal fibroblasts collagen synthesis decreased. Mechanical stretch short time alone, low amplitude can promote the expression of lumican, high amplitude But the degree of inhibition of its expression; IL-1 beta and mechanical pull together to.IL-1 beta can expression of fibroblast induced corneal MMP9 promote the expression of lumican, and promote MMP2 expression; mechanical stretch can increase further promote IL-1 beta induced MMP2 expression. The inflammatory factor and mechanical stretch joint action of the cornea fibroblasts reduced collagen synthesis, extracellular matrix degradation increased, may cause iatrogenic corneal ectasia. (3) increased and the high expression of epithelial cells of TNF- alpha TNF- alpha expression in tears of patients with keratoconus, the cone corneal fibroblasts in the inflammatory environment, the effects of different concentrations of alpha TNF- applied by the cone corneal fibroblasts, TNF- alpha keratoconus can promote the expression of MMPs, expression of.MMPs/TIMPs and inhibit the imbalance of TIMPs will lead to keratoconus fibroblast extracellular matrix degradation. Much, which may lead to the integrity of the corneal tissue structure damage. (4) applied to a fiber TNF- alpha and mechanical stretch of keratoconus, the study found that both of them make the corneal fibroblast proliferation was inhibited, apoptosis is not affected. Corneal fibroblasts may lead to the reduction in the number of the synthesis of extracellular matrix decreased and the corneal biomechanical properties change. Conclusion: inflammatory factors and mechanical stimulation by regulating the metabolism of extracellular matrix, corneal fibroblast proliferation plays an important role in the expansion of corneal disease occurrence.

【學(xué)位授予單位】：太原理工大學(xué)
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2016
【分類號(hào)】：R772.2;R318.01

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