【摘要】： 目的研究巨噬細(xì)胞吞噬凋亡細(xì)胞后自噬結(jié)構(gòu)的形態(tài)學(xué)特征,探討自噬對于巨噬細(xì)胞清除凋亡細(xì)胞的作用,以及自噬與異噬的相互關(guān)系。方法用環(huán)磷酰胺分別誘導(dǎo)淋巴細(xì)胞和Jurkat細(xì)胞凋亡。將凋亡的淋巴細(xì)胞和Jurkat細(xì)胞加入巨噬細(xì)胞培養(yǎng)皿,計算凋亡細(xì)胞清除率。通過Giesma染色和掃描電鏡觀察巨噬細(xì)胞吞噬凋亡細(xì)胞的表面結(jié)構(gòu)變化,透射電鏡下觀察巨噬細(xì)胞吞噬凋亡細(xì)胞后自噬前體、自噬體、自噬溶酶體和兩性體的分布和結(jié)構(gòu)特點,用圖像分析儀測量自噬結(jié)構(gòu)與細(xì)胞質(zhì)的斷面面積,求出面積之比。用MDC染色法標(biāo)記巨噬細(xì)胞的自噬體,在激光共聚焦掃描顯微鏡下觀察和計數(shù)。用3-MA抑制巨噬細(xì)胞的自噬,觀察凋亡細(xì)胞被清除變化。結(jié)果巨噬細(xì)胞活躍地吞噬凋亡細(xì)胞、凋亡細(xì)胞核、凋亡小體或其他細(xì)胞碎片,形成異噬體。與對照組比較,吞噬凋亡細(xì)胞組的巨噬細(xì)胞發(fā)生自噬的細(xì)胞增多,巨噬細(xì)胞內(nèi)自噬體數(shù)目增多,自噬前體、自噬體和自噬溶酶體與細(xì)胞質(zhì)的斷面面積之比增大。許多自噬體內(nèi)可見凋亡小體或其他細(xì)胞碎片,這些自噬體的體積較大,多位于細(xì)胞膜下。亦可見內(nèi)含細(xì)胞質(zhì)樣物質(zhì)、損傷線粒體或內(nèi)質(zhì)網(wǎng)等的自噬體,多位于線粒體和粗面內(nèi)質(zhì)網(wǎng)附近。未觀察到含有整個凋亡細(xì)胞或凋亡細(xì)胞核的自噬體。在部分細(xì)胞出現(xiàn)吞噬體和自噬體融合而成的兩性體。兩性體內(nèi)含凋亡細(xì)胞碎片和完整自噬體內(nèi)膜結(jié)構(gòu)。當(dāng)巨噬細(xì)胞自噬受抑制時,凋亡細(xì)胞清除率降低。結(jié)論巨噬細(xì)胞清除凋亡細(xì)胞時自噬功能顯著增強。自噬對于凋亡細(xì)胞的清除起著重要的直接和間接作用。在凋亡細(xì)胞清除時,自噬途徑和異噬途徑可通過兩性體的形成發(fā)生密切聯(lián)系。
[Abstract]:Objective to study the morphological characteristics of autophagy structure after phagocytosis of apoptotic cells, and to investigate the effect of autophagy on the clearance of apoptotic cells and the relationship between autophagy and heterophagy. Methods Cyclophosphamide was used to induce apoptosis of lymphocytes and Jurkat cells respectively. Apoptotic lymphocytes and Jurkat cells were added to macrophage culture dish to calculate the apoptotic cell clearance rate. The surface structure of phagocytic apoptotic cells of macrophages was observed by Giesma staining and scanning electron microscope. The distribution and structural characteristics of autophagy precursors autophagy lysosomes and amphoteric bodies after phagocytosis of apoptotic cells were observed under transmission electron microscope. The sectional area of autophagy structure and cytoplasm was measured by image analyzer and the area ratio was calculated. The autophagy of macrophages was labeled with MDC staining and observed and counted under confocal laser scanning microscope. 3-MA was used to inhibit macrophage autophagy and to observe the change of apoptotic cell scavenging. Results macrophages were active in phagocytosis of apoptotic cells, apoptotic nuclei, apoptotic bodies or other cell fragments. Compared with the control group, the number of autophagy and the ratio of autophagy precursor, autophagy and lysosome to cytoplasm were increased in the phagocytic apoptotic cell group. Apoptotic bodies or other cell fragments are found in many autophages. It was also found that autophagy containing cytoplasmic substance, damaged mitochondria or endoplasmic reticulum, mostly located near mitochondria and rough endoplasmic reticulum. No autophagy containing the whole apoptotic cell or apoptotic nucleus was observed. In some cells, phagocytosis and autophagy fused to form a hermaphrodite. Amphoteric body contains apoptotic cell fragments and intact autophagy membrane structure. When macrophage autophagy was inhibited, apoptotic cell clearance rate decreased. Conclusion the autophagy function of macrophages was significantly enhanced when the apoptotic cells were cleared. Autophagy plays an important direct and indirect role in the clearance of apoptotic cells. The autophagy pathway and heterophagocytic pathway are closely related to the formation of amphoteric body during the clearance of apoptotic cells.
【學(xué)位授予單位】：復(fù)旦大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2008
【分類號】：R329

【參考文獻(xiàn)】

相關(guān)期刊論文 前6條

1 王海杰,譚玉珍;激活巨噬細(xì)胞的肌動蛋白分布和鈣離子水平[J];解剖學(xué)報;2001年03期

2 王海杰,譚玉珍,李奇,李鴻帥;塵粒引起人支氣管肺淋巴結(jié)巨噬細(xì)胞的凋亡和bcl-2表達(dá)[J];解剖學(xué)報;2004年01期

3 王宇鯤;王海杰;譚玉珍;;CD44介導(dǎo)的透明質(zhì)酸促進巨噬細(xì)胞的吞噬功能[J];解剖學(xué)報;2006年03期

4 何韜;譚玉珍;王海杰;;自噬在巨噬細(xì)胞清除凋亡淋巴細(xì)胞中的作用[J];解剖學(xué)報;2008年04期

5 何韜;王海杰;譚玉珍;;自噬在細(xì)胞存活和死亡中的作用[J];生理科學(xué)進展;2008年01期

6 王海杰;譚玉珍;何韜;俞彰;;塵粒對巨噬細(xì)胞自噬的影響[J];中國病理生理雜志;2008年08期

，

本文編號：2324224