García M E, Del Zotto H, Caride A J, Filoteo A G, Penniston J T, Rossi J P F C, Gagliardino J J
CENEXA, Centro de Endocrinología Experimental y Aplicada (UNLP-CONICET, PAHO/WHO Collaborating Center), Facultad de Ciencias Médicas, 60 y 120, 1900, La Plata, Argentina.
J Membr Biol. 2002 Jan 1;185(1):17-23. doi: 10.1007/s00232-001-0108-z. Epub 2002 Feb 5.
This work is aimed at identifying the presence and cellular distribution pattern of plasma membrane calcium pump (PMCA) isoforms in normal rat pancreatic islet. Microsomal fractions of isolated islets and exocrine tissue were analyzed to detect different PMCA isoforms. The cellular distribution pattern of these PMCAs in the islets was also studied in fixed pancreas sections incubated with antibodies against PMCAs and insulin. Antibody 5F10, which reacts with all PMCA variants, showed multiple bands in the blots in the 127-134 kDa region, indicating the presence of several isoforms. Microsomes also reacted positively with specific antibodies for individual PMCA isoforms, generating a band of the expected size. Antibody 5F10 immunocytochemically labeled the plasma cell membrane of both b- and non-b-cells, but predominantly the former. All islet cells were also labeled with antibodies against isoforms 1 and 4, while the antibody reacting with isoform 3 labeled exclusively b-cells. A few b- and non-b-cells were positively labeled with the antibody reacting with PMCA b variant. Negative results were obtained with the antibody against isoform 2. Further studies, together with previous reports on the modulatory effect of insulin secretagogues and blockers upon PMCA activity, may provide evidence of the importance of this particular PMCA expression for islet function under normal and pathological conditions.
这项工作旨在确定正常大鼠胰岛中质膜钙泵（PMCA）亚型的存在及其细胞分布模式。对分离的胰岛和外分泌组织的微粒体部分进行分析，以检测不同的PMCA亚型。还在与抗PMCA和胰岛素抗体孵育的固定胰腺切片中研究了这些PMCA在胰岛中的细胞分布模式。与所有PMCA变体反应的抗体5F10在印迹中127 - 134 kDa区域显示多条带，表明存在几种亚型。微粒体也与针对单个PMCA亚型的特异性抗体呈阳性反应，产生预期大小的条带。抗体5F10免疫细胞化学标记了β细胞和非β细胞的质膜，但主要是前者。所有胰岛细胞也用针对亚型1和4的抗体标记，而与亚型3反应的抗体仅标记β细胞。少数β细胞和非β细胞用与PMCAβ变体反应的抗体呈阳性标记。针对亚型2的抗体得到阴性结果。进一步的研究，连同先前关于胰岛素分泌促进剂和阻滞剂对PMCA活性调节作用的报道，可能为这种特定的PMCA表达在正常和病理条件下对胰岛功能的重要性提供证据。
