Arkema J M, Schadee-Eestermans I L, Broekhuis-Fluitsma D M, Hoefsmit E C
Department of Cell Biology, Medical Faculty, Vrije University, Amsterdam, The Netherlands.
Immunobiology. 1991 Nov;183(5):396-407. doi: 10.1016/S0171-2985(11)80524-3.
Class II molecules are a prerequisite for antigen presentation. We studied whether class II molecules can be found in the endocytic and/or lysosomal route of dendritic cells (DC), which are very potent antigen-presenting cells. Therefore first immunolabelling for HLA-DR alpha chain was applied on ultrathin cryosections of cells of which plasma membrane HLA-DR/DQ molecules were labelled in suspension, followed by incubation with the endocytic marker BSA-gold. Second, immunolabelling for HLA-DR alpha chains was applied on ultrathin cryosections of cells on which enzyme cytochemistry for acid phosphatase (APh) was performed to see whether the class II positive vesicles belong to the lysosomal compartment. Third, this immunolabelling was applied on cryosections of cells pretreated with the protein synthesis inhibitor cycloheximide (CHX) to see whether the class II positive vesicles are derived from biosynthesis. We found limited uptake of BSA-gold into endosomes and lysosomes, some of which also contained endocytozed HLA-DR/DQ. APh and HLA-DR were observed in the same vesicles but also vesicles containing either HLA-DR or APh were found. However, many class II positive vesicles were found, which were apparently not accessible to exogenous molecules. Moreover, the amount of class II positive vesicles decreased after treatment of the cells with CHX, suggesting that these vesicles form part of the biosynthetic route. These results imply that there is a cluster of class II positive vesicles, probably a storage compartment, that has connections with the lysosomal system. The concentration of lysosomes and class II positive vesicles in the juxtanuclear area of DC is probably of crucial importance in the processing of antigens.
II类分子是抗原呈递的前提条件。我们研究了在树突状细胞（DC）的内吞和/或溶酶体途径中是否能发现II类分子，树突状细胞是非常有效的抗原呈递细胞。因此，首先对悬浮状态下细胞膜HLA-DR/DQ分子被标记的细胞超薄冰冻切片进行HLA-DRα链的免疫标记，然后与内吞标记物牛血清白蛋白-金孵育。其次，对进行了酸性磷酸酶（APh）酶细胞化学检测的细胞超薄冰冻切片进行HLA-DRα链的免疫标记，以观察II类阳性囊泡是否属于溶酶体区室。第三，将这种免疫标记应用于用蛋白质合成抑制剂环己酰亚胺（CHX）预处理的细胞冰冻切片上，以观察II类阳性囊泡是否来源于生物合成。我们发现牛血清白蛋白-金进入内体和溶酶体的摄取有限，其中一些内体和溶酶体也含有内吞的HLA-DR/DQ。在同一囊泡中观察到了APh和HLA-DR，但也发现了只含有HLA-DR或APh的囊泡。然而，发现了许多II类阳性囊泡，外源性分子显然无法进入这些囊泡。此外，用CHX处理细胞后，II类阳性囊泡的数量减少，这表明这些囊泡是生物合成途径的一部分。这些结果意味着存在一群II类阳性囊泡，可能是一个储存区室，它与溶酶体系统有联系。树突状细胞近核区域中溶酶体和II类阳性囊泡的聚集可能在抗原加工过程中至关重要。
