自噬在巨噬细胞清除凋亡细胞中的作用
发布时间：2018-11-11 07:48
【摘要】： 目的研究巨噬细胞吞噬凋亡细胞后自噬结构的形态学特征,探讨自噬对于巨噬细胞清除凋亡细胞的作用,以及自噬与异噬的相互关系。方法用环磷酰胺分别诱导淋巴细胞和Jurkat细胞凋亡。将凋亡的淋巴细胞和Jurkat细胞加入巨噬细胞培养皿,计算凋亡细胞清除率。通过Giesma染色和扫描电镜观察巨噬细胞吞噬凋亡细胞的表面结构变化,透射电镜下观察巨噬细胞吞噬凋亡细胞后自噬前体、自噬体、自噬溶酶体和两性体的分布和结构特点,用图像分析仪测量自噬结构与细胞质的断面面积,求出面积之比。用MDC染色法标记巨噬细胞的自噬体,在激光共聚焦扫描显微镜下观察和计数。用3-MA抑制巨噬细胞的自噬,观察凋亡细胞被清除变化。结果巨噬细胞活跃地吞噬凋亡细胞、凋亡细胞核、凋亡小体或其他细胞碎片,形成异噬体。与对照组比较,吞噬凋亡细胞组的巨噬细胞发生自噬的细胞增多,巨噬细胞内自噬体数目增多,自噬前体、自噬体和自噬溶酶体与细胞质的断面面积之比增大。许多自噬体内可见凋亡小体或其他细胞碎片,这些自噬体的体积较大,多位于细胞膜下。亦可见内含细胞质样物质、损伤线粒体或内质网等的自噬体,多位于线粒体和粗面内质网附近。未观察到含有整个凋亡细胞或凋亡细胞核的自噬体。在部分细胞出现吞噬体和自噬体融合而成的两性体。两性体内含凋亡细胞碎片和完整自噬体内膜结构。当巨噬细胞自噬受抑制时,凋亡细胞清除率降低。结论巨噬细胞清除凋亡细胞时自噬功能显著增强。自噬对于凋亡细胞的清除起着重要的直接和间接作用。在凋亡细胞清除时,自噬途径和异噬途径可通过两性体的形成发生密切联系。
[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.
【学位授予单位】：复旦大学
【学位级别】：硕士
【学位授予年份】：2008
【分类号】：R329
本文编号：2324224
[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.
【学位授予单位】：复旦大学
【学位级别】：硕士
【学位授予年份】：2008
【分类号】：R329
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