Basolateral and apical binding, internalization, and degradation of insulin by cultured kidney epithelial cells.
Basolateral and apical binding, internalization, and degradation of insulin by cultured kidney epithelial cells.
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培养的肾上皮细胞对胰岛素的基底外侧和顶端结合、内化和降解。
DOI：
10.1152/ajpendo.1989.257.6.e895
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发表时间：
1989
期刊：
影响因子：
--
通讯作者：
Frank,B
中科院分区：
文献类型：
--
作者：
Rabkin,R;Yagil,C;Frank,B
在体内，过滤后的胰岛素与根尖膜结合后，在近端小管中被吸收和降解。肾小管周围移除也会发生，涉及基底外侧受体的结合和降解。基侧降解是在细胞内进行还是在细胞表面进行尚不清楚。由于在活体内解决这一问题的困难，本研究以具有近端相似特征和胰岛素受体的培养的负鼠肾上皮细胞系为研究对象。细胞生长在聚碳酸酯过滤器上的隔开的井中，当融合时，单层有效地将培养良好地分离为顶端和基侧隔室。通过在根尖或基底室加入125I-胰岛素的单层孵育，分别研究了根尖和基底外侧的结合、内化和降解。在37℃时，胰岛素以时间依赖的方式与任一极相关联。这种相互作用是特异的，因为它被冷胰岛素竞争性抑制，但不被无关的多肽抑制。通过降低胞外pH实现了表面结合胰岛素与内化胰岛素的分离。在4℃时，添加到单层两侧的92%的放射性是表面结合的，而在37℃和1小时后，57%是表面结合的，43%是内化的。心尖部和基底侧部受体的亲和力相似(1-2 nM)，但基底侧部受体数量更多，因为在高胰岛素浓度(5×10(-8)M)时，基底侧部膜结合的细胞数是心尖部的5倍(250+/-81vs.56+/-11fm/10(6)细胞)。在暴露于细胞的任一极后退化。(摘要截短250字)
In vivo, filtered insulin is absorbed and degraded in proximal tubules after binding to the apical membrane. Peritubular removal also occurs and involves basolateral receptor binding and degradation. Whether basolateral degradation proceeds within the cell or on the cell surface is unknown. Because of the difficulties in addressing this question in vivo, this study was carried out with a cultured opossum kidney epithelium cell line with proximal-like features and insulin receptors. Cells were grown in partitioned wells on polycarbonate filters and, when confluent, the monolayer effectively separated the culture well into apical and basolateral compartments. Apical and basolateral binding, internalization, and degradation were studied separately by incubating monolayers with 125I-insulin added to either the apical or basal compartment. At 37 degrees C insulin associated with either pole in a time-dependent manner. This interaction was specific, for it was competitively inhibited by cold insulin but not by unrelated peptides. Separation of surface-bound from internalized insulin was achieved by lowering extracellular pH. At 4 degrees C, 92% of the radioactivity added to either side of the monolayer was surface-bound, whereas at 37 degrees C and after 1 h, 57% was surface-bound and 43% internalized. Affinity of apical and basolateral receptors were similar (1-2 nM), but basolateral receptor number was greater, for at high insulin concentrations (5 x 10(-8) M) basolateral membrane binding exceeded apical by fivefold (250 +/- 81 vs. 56 +/- 11 fm/10(6) cells). Degradation followed exposure to either pole of the cell.(ABSTRACT TRUNCATED AT 250 WORDS)