發(fā)布時(shí)間：2018-11-25 21:39

【摘要】：目的：了解肝臟自然殺傷細(xì)胞（natural killer cell，NK）在輪狀病毒（rhesusrotavirus，RRV）致小鼠膽道閉鎖（biliary atresia，BA）中的作用。 方法：實(shí)驗(yàn)動(dòng)物C57BL/6（B6）；TLR2基因敲除鼠[TLR2(-/-)]；C57BL/10（B10）；TLR4基因敲除鼠[TLR4(-/-)]。以上四種小鼠均購買自南京大學(xué)模式動(dòng)物研究所。其中B6和TLR2(-/-)背景相同，B10和TLR4(-/-)背景相同。 1.動(dòng)物實(shí)驗(yàn)：選擇生后1d、7d及成年（6-8w）三個(gè)時(shí)間的四種小鼠，腹腔注射RRV后24h，用磁珠分選技術(shù)分選肝臟NK細(xì)胞，，利用流式細(xì)胞術(shù)檢測(cè)肝NK細(xì)胞膜表面分子CD69表達(dá)水平，及NK細(xì)胞內(nèi)腫瘤壞死因子-α（TNF-α）和干擾素-γ（IFN-γ）的表達(dá)。分別進(jìn)行基因敲除小鼠和野生型小鼠的比較和生后1d、7d、成年小鼠的比較。 2.細(xì)胞實(shí)驗(yàn)：用非放射性細(xì)胞殺傷試劑盒檢測(cè)四種小鼠肝臟NK細(xì)胞對(duì)小鼠肝外膽管上皮細(xì)胞的殺傷活性。 結(jié)果：1.腹腔注射RRV后，野生型小鼠NK細(xì)胞活化表達(dá)水平比同齡的TLR敲除小鼠明顯較高； 2.腹腔注射RRV后新生小鼠NK細(xì)胞活化較低； 3.野生型小鼠肝臟NK細(xì)胞比同齡的TLR基因敲除小鼠殺傷肝外膽管上皮細(xì)胞的百分率高； 4.HMGB1活化后小鼠肝NK細(xì)胞對(duì)小鼠肝外膽管上皮細(xì)胞的殺傷百分率較正常組高；小鼠肝NK細(xì)胞對(duì)感染RRV后小鼠肝外膽管上皮細(xì)胞的殺傷百分率較正常組高。 結(jié)論：1.新生鼠肝臟NK細(xì)胞的功能不足，清除感染RRV的膽管上皮細(xì)胞能力低下； 2.HMGB1可以和NK細(xì)胞的表面TLR2和TLR4結(jié)合，并且激活NK細(xì)胞�；罨腘K對(duì)膽管上皮細(xì)胞有較強(qiáng)殺傷作用；RRV感染后的膽管上皮細(xì)胞容易被NK細(xì)胞殺傷。
[Abstract]:Aim: to investigate the role of hepatic natural killer cells (natural killer cell,NK) in rotavirus (rhesusrotavirus,RRV) -induced biliary atresia (biliary atresia,BA) in mice. Methods: C57BL/6 (B6); TLR2 knockout mice [TLR2 (- / -)]; C57BL/10 (B10); TLR4 gene knockout mice [TLR4 (- / -)]. The above four kinds of mice were purchased from the Institute of Model Animals, Nanjing University. Where B 6 and TLR2 (- / -) have the same background, and B 10 and TLR4 (- / -) have the same background. 1. Animal experiment: four kinds of mice were selected at 1 day after birth and 3 hours after birth (6-8 weeks). 24 hours after intraperitoneal injection of RRV, liver NK cells were sorted by magnetic bead sorting technique. Flow cytometry was used to detect the expression level of CD69 on the surface of liver NK cell membrane. And the expression of tumor necrosis factor- 偽 (TNF- 偽) and interferon- 緯 (IFN- 緯) in NK cells. The gene knockout mice and wild-type mice were compared respectively, and 1 day after birth and 7 days after birth, adult mice were compared. 2. Cell experiment: the cytotoxicity of four kinds of murine liver NK cells to mouse extrahepatic bile duct epithelial cells was detected by non-radioactive cell killing kit. Results: 1. After intraperitoneal injection of RRV, the activation and expression level of NK cells in wild type mice was significantly higher than that in TLR knockout mice of the same age; 2. The activation of NK cells in neonatal mice was lower after intraperitoneal injection of RRV. The rate of killing extrahepatic bile duct epithelial cells by NK cells of wild type mice was higher than that of TLR gene knockout mice of the same age, and the killing rate of NK cells on extrahepatic bile duct epithelial cells of mice after 4.HMGB1 activation was higher than that of normal mice. The killing rate of mouse liver NK cells on extrahepatic bile duct epithelial cells after RRV infection was higher than that of normal group. Conclusion: 1. The function of NK cells in the liver of neonatal rats was insufficient, and the ability of clearing bile duct epithelial cells infected with RRV was low, and 2.HMGB1 could bind to TLR2 and TLR4 on the surface of NK cells and activate NK cells. Activated NK has strong killing effect on bile duct epithelial cells, and bile duct epithelial cells infected with RRV are easy to be killed by NK cells.
【學(xué)位授予單位】：華中科技大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類號(hào)】：R725.7

【參考文獻(xiàn)】

相關(guān)期刊論文 前4條

1 孫世波,徐偉,孫鐵為,邰升,遲強(qiáng),韓德恩,曲曉菊;全腸外營養(yǎng)添加免疫調(diào)節(jié)劑黃芪對(duì)梗阻性黃疸病人術(shù)后的影響[J];腸外與腸內(nèi)營養(yǎng);2002年03期

2 鄭軍;胡國治;郭永忠;蔡國英;;梗阻性黃疸圍手術(shù)期ω-3脂肪酸對(duì)T細(xì)胞亞群及預(yù)后的影響[J];腸外與腸內(nèi)營養(yǎng);2005年06期

3 梁友玲,林超瓊,馮軍壇;口服維生素K_1在新生兒的安全性研究[J];兒科藥學(xué);2001年04期

4 陳煥偉,甄作均,潘文松;前白蛋白和視黃醇結(jié)合蛋白在腸外營養(yǎng)評(píng)估中的意義[J];腸外與腸內(nèi)營養(yǎng);2001年03期

本文編號(hào)：2357452