Turner J R, Black E D, Ward J, Tse C M, Uchwat F A, Alli H A, Donowitz M, Madara J L, Angle J M
Department of Pathology, Wayne State University School of Medicine, Detroit, Michigan 48201, USA.
Am J Physiol Cell Physiol. 2000 Dec;279(6):C1918-24. doi: 10.1152/ajpcell.2000.279.6.C1918.
Initiation of intestinal Na(+)-glucose cotransport results in transient cell swelling and sustained increases in tight junction permeability. Since Na(+)/H(+) exchange has been implicated in volume regulation after physiological cell swelling, we hypothesized that Na(+)/H(+) exchange might also be required for Na(+)-glucose cotransport-dependent tight junction regulation. In Caco-2 monolayers with active Na(+)-glucose cotransport, inhibition of Na(+)/H(+) exchange with 200 microM 5-(N,N-dimethyl)- amiloride induced 36 +/- 2% increases in transepithelial resistance (TER). Evaluation using multiple Na(+)/H(+) exchange inhibitors showed that inhibition of the Na(+)/H(+) exchanger 3 (NHE3) isoform was most closely related to TER increases. TER increases due to NHE3 inhibition were related to cytoplasmic acidification because cytoplasmic alkalinization with 5 mM NH(4)Cl prevented both cytoplasmic acidification and TER increases. However, NHE3 inhibition did not affect TER when Na(+)-glucose cotransport was inhibited. Myosin II regulatory light chain (MLC) phosphorylation decreased up to 43 +/- 5% after inhibition of Na(+)/H(+) exchange, similar to previous studies that associate decreased MLC phosphorylation with increased TER after inhibition of Na(+)-glucose cotransport. However, NHE3 inhibitors did not diminish Na(+)-glucose cotransport. These data demonstrate that inhibition of NHE3 results in decreased MLC phosphorylation and increased TER and suggest that NHE3 may participate in the signaling pathway of Na(+)-glucose cotransport-dependent tight junction regulation.
肠道钠-葡萄糖协同转运的启动会导致短暂的细胞肿胀以及紧密连接通透性的持续增加。由于钠/氢交换已被认为在生理性细胞肿胀后的体积调节中发挥作用，我们推测钠-葡萄糖协同转运依赖性紧密连接调节可能也需要钠/氢交换。在具有活跃钠-葡萄糖协同转运的Caco-2单层细胞中，用200微摩尔5-(N,N-二甲基)-氨氯吡咪抑制钠/氢交换会导致跨上皮电阻（TER）增加36±2%。使用多种钠/氢交换抑制剂进行评估表明，钠/氢交换体3（NHE3）亚型的抑制与TER增加最为密切相关。由于NHE3抑制导致的TER增加与细胞质酸化有关，因为用5毫摩尔氯化铵进行细胞质碱化可防止细胞质酸化和TER增加。然而，当钠-葡萄糖协同转运被抑制时，NHE3抑制并不影响TER。抑制钠/氢交换后，肌球蛋白II调节轻链（MLC）磷酸化最多降低43±5%，这与之前关于抑制钠-葡萄糖协同转运后将MLC磷酸化降低与TER增加相关联的研究类似。然而，NHE3抑制剂并不减少钠-葡萄糖协同转运。这些数据表明，抑制NHE3会导致MLC磷酸化降低和TER增加，并提示NHE3可能参与钠-葡萄糖协同转运依赖性紧密连接调节的信号通路。
