Antigen processing of glycoconjugate vaccines; the polysaccharide portion of the pneumococcal CRM197 conjugate vaccine co-localizes with MHC II on the antigen processing cell surface
Antigen processing of glycoconjugate vaccines; the polysaccharide portion of the pneumococcal CRM197 conjugate vaccine co-localizes with MHC II on the antigen processing cell surface
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DOI：
10.1016/j.vaccine.2009.03.064
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发表时间：
2009-05-21
期刊：
影响因子：
5.5
通讯作者：
Schreiber, John R.
中科院分区：
文献类型：
--
作者：
Lai, Zengzu;Schreiber, John R.
肺炎球菌（Pn）多糖（PS）是t非依赖性（TI）抗原，不会在婴儿中诱导免疫记忆或抗体。PnPS与载体蛋白CRM197结合可诱导婴儿产生ps特异性抗体，并具有与t依赖性（Td）抗原相似的记忆。通过抗原加工和MHC II载体蛋白的呈递以及T细胞的募集，缀合物提高了免疫原性，但PS附着在载体上的命运尚不清楚。为了确定PnPS-CRM197的PS成分在APC中的位置，我们分别对PS和蛋白进行了标记，并对其位置进行了跟踪。14-CRM197和19F-CRM197的PS用Alexa Fluor (R) 594肼（红色）特异性标记。在不标记PS的反应中，CRM197被单独标记为红色。标记的抗原与APC孵育，APC固定，渗透并与Alexa Fluor (R) 488标记为绿色的抗mhc II抗体孵育，然后共聚焦显微镜。标记的CRM197出现在APC表面，并与MHC II共定位（黄色）。标记的未偶联的14或19F PS没有到达APC表面，但标记的14- crm197和19F- crm197被内化并与MHC II共定位。14ps型单克隆抗体结合细胞内14ps型和PS- crm197。Brefeldin A和氯喹阻断了CRM197和PS标记的14-CRM197和19F-CRM197与MHC II共定位。这些数据表明，CRM197糖偶联物的PS组分进入核内体，与CRM197肽一起移动到APC表面，并与MHC II共定位。2009爱思唯尔有限公司版权所有。
Pneumococcal (Pn) polysaccharides (PS) are T-independent (TI) antigens and do not induce immunological memory or antibodies in infants. Conjugation of PnPS to the carrier protein CRM197 induces PS-specific antibody in infants, and memory similar to T-dependent (Td) antigens. Conjugates have improved immunogenicity via antigen processing and presentation of carrier protein with MHC II and recruitment of T cell help,but the fate of the PS attached to the carrier is Unknown. To determine the location of the PS component of PnPS-CRM197 in the APC, we separately labeled PS and protein and tracked their location. The PS of types 14-CRM197 and 19F-CRM197 was specifically labeled by Alexa Fluor (R) 594 hydrazide (red). The CRM197 was separately labeled red in a reaction that did not label PS. Labeled antigens were incubated with APC which were fixed, permeabilized and incubated with anti-MHC II antibody labeled green by Alexa Fluor (R) 488, followed by confocal microscopy. Labeled CRM197 was presented on APC surface and co-localized with MHC II (yellow). Labeled unconjugated 14 or 19F PS did not go to the APC Surface, but PS labeled 14-CRM197 and 19F-CRM197 was internalized and co-localized with MHC II. Monoclonal antibody to type 14 PS bound to intracellular type 14 PS and PS-CRM197. Brefeldin A and chloroquine blocked both CRM197 and PS labeled 14-CRM197 and 19F-CRM197 from co-localizing with MHC II. These data suggest that the PS component of the CRM197 glyconconjugate enters the endosome, travels with CRM197 peptides to the APC Surface and co-localizes with MHC II. (C) 2009 Elsevier Ltd. All rights reserved.