CHARACTERIZATION OF THE Na+K+‐ATPase IN ISOLATED BOVINE ARTICULAR CHONDROCYTES; MOLECULAR EVIDENCE FOR MULTIPLE α AND β ISOFORMS
CHARACTERIZATION OF THE Na+K+‐ATPase IN ISOLATED BOVINE ARTICULAR CHONDROCYTES; MOLECULAR EVIDENCE FOR MULTIPLE α AND β ISOFORMS
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分离的牛关节软骨细胞中 Na+K+-ATP 酶的表征；多种 α 和 β 同种型的分子证据
作者：
A. Mobasheri;R. Errington;S. Golding;A. Hall;J. Urban
我们使用了针对Na+K+-ATP酶αα1、α2和α3）和ββ1和β2）亚基同种型的同种型特异性抗体，以确定其在分离的牛关节软骨细胞中的特异性定位。免疫印迹证实了α1和α3亚型的存在，尽管通过免疫荧光共聚焦激光扫描显微镜和PCR评估α1表达显著大于α3。类似的方法显示软骨细胞中存在β1和β2亚型，尽管质膜上的β2免疫染色比β1更点状，而β 1在亚细胞区室中占主导地位。发现Na+K+-ATP酶的质膜丰度对细胞外离子浓度敏感，并且细胞外Na+浓度的长期升高显著上调Na+K+-ATP酶密度，如通过特异性3 H-哇巴因结合测量的。我们的观察结果表明，α3和β2的表达并不局限于兴奋性组织，如以前报道的。软骨细胞中α3表达的生理相关性可能与其在细胞外环境中对细胞内Na+的低亲和力有关，其中Na+浓度（260- 350 mm）与其他细胞类型（140 mm）相比异常高。糖蛋白及其分支碳水化合物参与细胞识别事件，因此β2亚基糖蛋白可以通过与细胞骨架和基质大分子的组分物理相互作用来允许软骨细胞检测其细胞外环境的变化。
We have used isoform‐specific antibodies against the Na+K+‐ATPase αα1, α2 and α3) and ββ1 and β2) subunit isoforms in order to establish their specific localization in isolated bovine articular chondrocytes. Immunoblotting confirmed the presence of the α1 and α3 isoforms, although α1 expression was significantly greater than α3 as assessed by immunofluorescence confocal laser scanning microscopy and PCR. A similar approach revealed the presence of the β1 and β2 isoforms in chondrocytes, although β2 immunostaining on the plasma membrane was more punctate than β1 which in contrast predominated in a subcellular compartment. The plasma membrane abundance of the Na+K+‐ATPase was found to be sensitive to the extracellular ionic concentration and long‐term elevation of extracellular Na+concentration significantly upregulated Na+K+‐ATPase density as measured by specific3H‐ouabain binding. Our observations suggest that the expression of α3 and β2 is not restricted to excitable tissues as previously reported. The physiological relevance of α3 expression in chondrocytes may be related to its low affinity for intracellular Na+in an extracellular environment where Na+concentration is unusually high (260–350mm) compared to other cell types (140mm). Glycoproteins and their branched carbohydrates have been implicated in cell recognition events, thus the β2 subunit glycoprotein may allow the chondrocyte to detect changes in its extracellular environment by physically interacting with components of the cellular cytoskeleton and matrix macromolecules.
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DOI：
--
发表时间：
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期刊：
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影响因子：
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DOI：
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DOI：
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发表时间：
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期刊：
The Journal of biological chemistry
影响因子：
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DOI：
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The Journal of biological chemistry
影响因子：
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