本文選題：組織工程　切入點：肝細胞　出處：《中國組織工程研究》2015年51期

【摘要】：背景:肝星狀細胞是肝內(nèi)分泌細胞外基質(zhì)的關(guān)鍵細胞類型,因此在肝纖維化研究中很重要。目的:實驗改良了肝星狀細胞原代培養(yǎng)方法,獲得充足的細胞量,用轉(zhuǎn)化生長因子β1刺激活化,研究致纖維化的相關(guān)因子表達情況。方法:氯胺酮麻醉小鼠后采用門靜脈穿刺原位灌注肝臟的方法分離肝臟,采用密度梯度離心獲得肝星狀細胞,利用形態(tài)學、免疫熒光染色等方法對原代肝星狀細胞進行鑒定;將培養(yǎng)24 h肝星狀細胞用質(zhì)量濃度10μg/L轉(zhuǎn)化生長因子β1刺激48 h,同時設(shè)PBS組進行對比。結(jié)果與結(jié)論:通過原位灌注肝臟并且酶消化的方法可以成功獲得小鼠原代肝星狀細胞,并且錐蟲藍染色活率為(97.2±0.8)%,免疫熒光染色純度為(90.4±1.2)%,細胞計數(shù)總量約2.5×106/只。實時熒光定量聚合酶鏈式反應(yīng)檢測顯示,與PBS組比較,轉(zhuǎn)化生長因子β1刺激組的肝星狀細胞平滑肌肌動蛋白α、Ⅰ型膠原蛋白、轉(zhuǎn)化生長因子β受體1、轉(zhuǎn)化生長因子β受體2 mR NA表達水平明顯升高(P0.05)。結(jié)果證實,實驗采用的改良的培養(yǎng)方法可更高效高質(zhì)量的獲得原代肝星狀細胞,轉(zhuǎn)化生長因子β1刺激可導致肝星狀細胞活化,分泌致纖維化因子。
[Abstract]:Background: hepatic stellate cells (HSCs) are the key cell types of extracellular matrix (ECM) of liver, so they are very important in the study of hepatic fibrosis.Aim: to improve the primary culture method of hepatic stellate cells (HSC), obtain sufficient cells, stimulate activation with transforming growth factor 尾 1 (TGF- 尾 1), and study the expression of fibrosis related factors.Methods: after ketamine anesthetized mice liver was isolated by portal vein puncture in situ perfusion. Hepatic stellate cells were obtained by density gradient centrifugation. Primary hepatic stellate cells were identified by means of morphology and immunofluorescence staining.Hepatic stellate cells were cultured for 24 h and stimulated with 10 渭 g / L TGF- 尾 1 for 48 h, and compared with PBS group.Results and conclusion: primary hepatic stellate cells of mice could be successfully obtained by in situ perfusion of liver and enzyme digestion. The viability of trypanosome blue staining was 97.2 鹵0.8. The purity of immunofluorescence staining was 90.4 鹵1.2. The total number of cells was about 2.5 脳 106 per mouse.Real-time fluorescence quantitative polymerase chain reaction showed that the hepatic stellate cell smooth muscle actin 偽, type I collagen were stimulated by transforming growth factor 尾 1 (TGF- 尾 1) compared with PBS group.The expression of transforming growth factor 尾 receptor 1 and transforming growth factor 尾 receptor 2mR na increased significantly (P 0.05).The results showed that the improved culture method could obtain the primary hepatic stellate cells more efficiently and qualitatively, and the stimulation of transforming growth factor 尾 1 could lead to the activation of hepatic stellate cells and the secretion of fibrogenic factors.
【作者單位】： 新疆醫(yī)科大學第一附屬醫(yī)院/臨床醫(yī)學研究院 新疆重大疾病醫(yī)學重點實驗室-省部共建國家重點實驗室培育基地 新疆包蟲病基礎(chǔ)醫(yī)學重點實驗室;新疆醫(yī)科大學基礎(chǔ)醫(yī)學院生物化學與分子生物學教研室;
【基金】：國家自然科學基金面上項目(81371838) 新疆維吾爾自治區(qū)自然科學基金青年項目(2015211C084)~~
【分類號】：R575.2

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