發(fā)布時(shí)間：2018-04-22 19:04

  本文選題：角膜上皮 + 神經(jīng)生長(zhǎng)因子　； 參考：《濟(jì)南大學(xué)》2017年碩士論文

【摘要】：目的通過(guò)正常小鼠和鏈脲佐菌素(STZ)誘導(dǎo)的I糖尿病小鼠的角膜上皮損傷模型以及小鼠角膜上皮干/祖細(xì)胞 三叉神經(jīng)節(jié)感覺(jué)神經(jīng)元(TKE2細(xì)胞 TG細(xì)胞)體外共培養(yǎng)模型,檢測(cè)小鼠角膜上皮細(xì)胞中神經(jīng)營(yíng)養(yǎng)因子(neurotrophic factors)的表達(dá)情況,研究其對(duì)小鼠角膜神經(jīng)再生的影響。方法用C57/BL6小鼠建立角膜上皮損傷模型,檢測(cè)并比較小鼠正常角膜(角膜上皮損傷未損傷)、再生期(角膜上皮損傷后2天)以及再生后(角膜上皮損傷后7天)角膜上皮細(xì)胞中神經(jīng)營(yíng)養(yǎng)因子的表達(dá)。體外培養(yǎng)小鼠角膜上皮干/祖細(xì)胞(TKE2細(xì)胞),利用劃痕模型檢測(cè)正常及損傷后TKE2細(xì)胞中神經(jīng)生長(zhǎng)因子(NGF)和膠質(zhì)細(xì)胞源性神經(jīng)生長(zhǎng)因子(GDNF)的表達(dá)變化。通過(guò)TKE2細(xì)胞 TG細(xì)胞體外共培養(yǎng)模型,檢測(cè)三叉神經(jīng)節(jié)感覺(jué)神經(jīng)元(TG細(xì)胞)軸突生長(zhǎng)變化。利用鏈脲佐菌素(STZ)誘導(dǎo)I型糖尿病小鼠,通過(guò)角膜上皮損傷修復(fù)模型,檢測(cè)糖尿病小鼠角膜上皮中NGF和GDNF表達(dá)變化;觀察糖尿病小鼠角膜上皮修復(fù)和神經(jīng)再生以及外源性NGF和GDNF對(duì)糖尿小鼠角膜上皮修復(fù)、神經(jīng)再生和角膜神經(jīng)敏感度恢復(fù)的影響。結(jié)果在角膜上皮修復(fù)和角膜神經(jīng)再生過(guò)程中,與正常角膜相比,角膜上皮中NGF和GDNF的表達(dá)在角膜上皮再生期(角膜上皮損傷后2天)顯著上升,再生后(損傷后第7天)部分恢復(fù);而NGF中和抗體或GDNF封閉多肽可顯著抑制角膜上皮損傷修復(fù)及神經(jīng)再生。與正常對(duì)照組相比,TKE2細(xì)胞損傷后(劃痕實(shí)驗(yàn)48h)NGF和GDNF的表達(dá)顯著上升,且劃痕后的條件培養(yǎng)上清可以顯著促進(jìn)TG細(xì)胞軸突的生長(zhǎng);而NGF中和抗體或GDNF封閉多肽可顯著抵消TKE2條件培養(yǎng)上清的促進(jìn)TG細(xì)胞軸突生長(zhǎng)的作用。微流體小室共培養(yǎng)實(shí)驗(yàn)結(jié)果顯示,TKE2細(xì)胞及其條件培養(yǎng)基可以顯著促進(jìn)小鼠TG細(xì)胞的軸突的生長(zhǎng),且呈現(xiàn)濃度依賴(lài)性。與正常小鼠相比,糖尿病小鼠角膜上皮損傷修復(fù)及神經(jīng)再生均顯著延遲;糖尿病小鼠再生期的角膜上皮NGF和GDNF的表達(dá)均顯著下降;而外源性給予NGF或GDNF均可顯著促進(jìn)糖尿病小鼠角膜上皮損傷愈合、神經(jīng)再生以及角膜神經(jīng)敏感度的恢復(fù)。結(jié)論角膜上皮細(xì)胞表達(dá)和分泌多種神經(jīng)營(yíng)養(yǎng)因子促進(jìn)角膜神經(jīng)再生,其中NGF和GDNF在角膜神經(jīng)再生中起著重要作用。
[Abstract]:Objective to study the model of corneal epithelial injury induced by streptozotocin (STZ) in normal mice and streptozotocin (STZ) induced by streptozotocin (STZ) and co-culture of TKE2 cells (TG cells) in rat corneal epithelial stem / progenitor cells. To detect the expression of neurotrophic factors in mouse corneal epithelial cells and to study the effect of neurotrophic factors on corneal regeneration in mice. Methods the corneal epithelial injury model was established in C57/BL6 mice. The expression of neurotrophic factor in normal cornea (no injury of corneal epithelium), regeneration period (2 days after corneal epithelium injury) and regeneration (7 days after corneal epithelial injury) were detected and compared. The expression of nerve growth factor (NGF) and glial cell derived nerve growth factor (GDF) in normal and injured TKE2 cells were detected by scratch model. The changes of axon growth in trigeminal ganglion sensory neurons (TG cells) were detected by co-culture of TKE2 cells and TG cells in vitro. Type I diabetic mice were induced by streptozotocin (STZ). The expression of NGF and GDNF in corneal epithelium of diabetic mice was detected by corneal epithelial injury repair model. The effects of exogenous NGF and GDNF on corneal epithelial repair, nerve regeneration and corneal nerve sensitivity recovery in diabetic mice were observed. Results in the process of corneal epithelium repair and corneal nerve regeneration, the expression of NGF and GDNF in corneal epithelium increased significantly during the period of corneal epithelium regeneration (2 days after corneal epithelial injury) compared with normal cornea. NGF neutralizing antibody or GDNF blocking polypeptide could significantly inhibit corneal epithelial injury repair and nerve regeneration after regeneration (7 days after injury). Compared with the normal control group, the expression of 48h)NGF and GDNF in TKE2 cells was significantly increased after injury (scratching test), and the supernatant of conditioned culture after scratch could significantly promote the growth of axons of TG cells. NGF neutralizing antibodies or GDNF blocking peptides could significantly counteract the role of TKE2 conditioned supernatants in promoting the axonal growth of TG cells. The results of microfluid chamber co-culture showed that TKE2 cells and their conditioned medium could significantly promote the axon growth of mouse TG cells in a concentration-dependent manner. Compared with normal mice, the corneal epithelial injury repair and nerve regeneration in diabetic mice were significantly delayed, and the expression of NGF and GDNF in corneal epithelium of diabetic mice decreased significantly during regeneration. Exogenous administration of NGF or GDNF could significantly promote corneal epithelial injury healing, nerve regeneration and recovery of corneal nerve sensitivity in diabetic mice. Conclusion the corneal epithelial cells express and secrete a variety of neurotrophic factors to promote corneal nerve regeneration. NGF and GDNF play an important role in corneal nerve regeneration.
【學(xué)位授予單位】：濟(jì)南大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2017
【分類(lèi)號(hào)】：R587.2;R772.2

【參考文獻(xiàn)】

相關(guān)期刊論文 前1條

1 王曄;周慶軍;謝立信;;糖尿病角膜病變發(fā)病機(jī)制的研究進(jìn)展[J];中華眼科雜志;2014年01期

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本文編號(hào)：1788512