Cellular localization of NKCC2 and its possible role in the Cl- absorption in the rat and human distal colonic epithelia.
Cellular localization of NKCC2 and its possible role in the Cl- absorption in the rat and human distal colonic epithelia.
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DOI：
10.1016/j.trsl.2011.04.003
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发表时间：
2011-09
期刊：
影响因子：
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通讯作者：
Jin-Xia Zhu;H. Xue;T. Ji;Ying Xing
中科院分区：
文献类型：
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作者：
Jin-Xia Zhu;H. Xue;T. Ji;Ying Xing
最近，我们证实了NKCC2在大鼠胃肠道（包括结肠远端粘膜）中的表达，NKCC2是NKCC在肾脏中特异性表达的一种吸收异构体。本研究旨在探讨其在结肠上皮中的定位及其在结肠离子运输中的可能作用。采用逆转录聚合酶链反应（RT-PCR）、Western blotting和免疫组织化学方法研究NKCC2的表达和定位。通过短路电流（ISC）监测研究NKCC2在结肠离子转运中的作用。结果表明，NKCC2在大鼠远端结肠和人乙状结肠上皮顶端区表达。NKCC1是一种隐秘的NKCC亚型，主要位于基底外侧膜，这已经得到了充分的证明。布美他尼是一种NKCC1和NKCC2的抑制剂，浆膜（基底侧）给予布美他尼，可抑制福斯克林诱导的浆膜isc升高66%，但可使管腔（根尖）福斯克林诱导的isc升高63%。此外，除Na+和K+阳离子参与ISCresponse外，在顶端添加amiloride （10 μmol/L）阻断上皮Na+通道、四乙基铵离子（TEA） （5 mmol/L）阻断K+通道或格列苯limide （0.1 mmol/L）阻断上皮Na+通道均不影响forskolin诱导的顶端isc升高。布美他尼预处理能显著增强福斯克林诱导的iscl浓度，使其降低114%和198%，而格列苯cliide （1 mmol/L）对其抑制作用大于60%。这一发现表明阴离子参与其中。此外，基底外侧HCO3−减少的根尖forskolin诱导的isc75%以上的去除表明，大鼠远端结肠根尖forskolin诱导的isc增加是由Cl−吸收和HCO3−分泌介导的。综上所述，NKCC2在大鼠远端结肠和人乙状结肠结肠上皮中广泛表达，尤其是在根尖膜中表达。它涉及结肠Cl -吸收和HCO3 -分泌的过程。
Recently, we demonstrated the expression of NKCC2, an absorptive isoform of NKCC specifically expressed in the kidney, in the rat gastrointestinal tract including the distal colonic mucosa. This study aims to investigate its localization in colonic epithelia and possible role in the colonic ion transport. Reverse transcription polymerase chain reaction (RT-PCR), Western blotting, and immunohistochemistry were used to investigate the expression and localization of NKCC2. The role of NKCC2 on the colonic ion transport was examined by mean of short-circuit current (ISC) monitoring. The results indicated that NKCC2 was expressed in the apical region of the epithelia in rat distal colon and human sigmoid colon. NKCC1, which is a secretive NKCC isoform, was localized predominantly in the basolateral membrane, which has been well documented. Serosal (basolateral) administration of bumetanide, an inhibitor of both NKCC1 and NKCC2, inhibited serosal forskolin-induced ISCincrease by 66% but enhanced the luminal (apical) forskolin-induced ISCresponse by 63%. Furthermore, the blocking of epithelial Na+channels by apical addition of amiloride (10 μmol/L), K+channels by tetraethylammoniumion (TEA) (5 mmol/L), or glibenclimide (0.1 mmol/L) did not affect apical forskolin-induced ISCincrease, excluding the involvement of cations, Na+and K+, in the ISCresponse. The luminal forskolin-induced ISCincrease was enhanced markedly by the apical pretreatment with bumetanide or the reduction of apical Cl−concentration by 114% and 198%, respectively, which were inhibited by apical addition of glibenclimide (1 mmol/L) by more than 60%. This finding suggests the involvement of an anion. Furthermore, the removal of basolateral HCO3−reduced apical forskolin-induced ISCby more than 75% indicated that the apical forskolin-induced ISCincrease in rat distal colon was mediated by Cl−absorption and HCO3−secretion. In conclusion, NKCC2 is expressed widely in the colonic epithelium in rat distal colon and human sigmoid colon, especially in the apical membrane. It involves the process of colonic Cl−absorption coupled with HCO3−secretion.