Regulation of Na-K-2Cl cotransport in osteoblasts.
Regulation of Na-K-2Cl cotransport in osteoblasts.
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成骨细胞中 Na-K-2Cl 共转运的调节。
DOI：
10.1152/ajpcell.1991.261.3.c433
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发表时间：
1991
期刊：
影响因子：
--
通讯作者：
Muallem,S
中科院分区：
文献类型：
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作者：
Whisenant,N;Zhang,BX;Khademazad,M;Loessberg,P;Muallem,S
采用~（86）Rb摄取来跟踪骨肉瘤细胞系UMR-106-01中Na-K-2Cl共转运的活性。86 Rb摄取的哇巴因耐药部分对布美他尼和呋塞米敏感。呋塞米敏感的86 Rb摄取需要存在的Na+，K+，和Cl-在孵育介质中。这些观察结果表明在成骨细胞中存在Na-K-2Cl共转运系统。协同转运蛋白活性被激动剂刺激，激动剂增加腺苷3 '，5'-环一磷酸（cAMP）、胞质游离Ca 2+（[Ca 2 +]i）和蛋白激酶C（PKC）活性，如甲状旁腺激素（PTH）和前列腺素E2（PGE 2）。然而，内皮素，增加[Ca 2 +]i和PKC活性，而不影响细胞水平的cAMP，是无效的刺激协同转运。因此，用毛喉素增加细胞cAMP与PTH和PGE 2刺激协同转运蛋白一样有效。用TPA刺激PKC以时间和浓度依赖性方式抑制协同转运蛋白。在任何12-O-十四烷酰基-佛波醇-13-乙酸酯（TPA）浓度或孵育时间下均未显示共转运刺激。Na-K-2Cl协同转运蛋白受细胞收缩刺激。在低渗介质中溶胀细胞并随后在等渗介质中收缩后观察到最大刺激。通过细胞收缩的刺激可以在对照、激动剂、cAMP和TPA处理的细胞中证明。这些观察结果表明：1）成骨细胞Na-K-2Cl协同转运蛋白主要通过增加细胞内cAMP被促钙激素激活，2）在成骨细胞中，协同转运蛋白受不同生化途径的独立调节。
Uptake of 86Rb was used to follow the activity of Na-K-2Cl cotransport in the osteosarcoma cell line UMR-106-01. The ouabain-resistant fraction of 86Rb uptake was sensitive to bumetanide and furosemide. Furosemide-sensitive 86Rb uptake required the presence of Na+, K+, and Cl- in the incubation medium. These observations indicate the presence of a Na-K-2Cl cotransport system in osteoblasts. Cotransporter activity was stimulated by agonists which increase adenosine 3',5'-cyclic monophosphate (cAMP), cytosolic free Ca2+ ([Ca2+]i), and protein kinase C (PKC) activity such as parathyroid hormone (PTH) and prostaglandin E2 (PGE2). However, endothelin, which increases [Ca2+]i and PKC activity without affecting cellular levels of cAMP, was ineffective in stimulating the cotransporter. Accordingly, increasing cellular cAMP with forskolin was as effective as PTH and PGE2 in stimulating the cotransporter. Stimulation of PKC with TPA inhibited the cotransporter in a time- and concentration-dependent manner. No stimulation of cotransport could be demonstrated at any 12-O-tetradecanoyl-phorbol-13-acetate (TPA) concentration or incubation time. The Na-K-2Cl cotransporter was stimulated by cell shrinkage. Maximal stimulation was observed after swelling the cells in hypotonic medium and subsequent shrinkage in isotonic medium. Stimulation by cell shrinkage can be demonstrated in control, agonist-, cAMP-, and TPA-treated cells. These observations suggest that 1) the osteoblastic Na-K-2Cl cotransporter is activated by calciotropic hormones predominantly through an increase in cellular cAMP, and 2) in osteoblasts, the cotransporter is independently regulated by different biochemical pathways.