First Department of Medicine, University of Szeged, 6720, Korányi fasor 8-10, Szeged, Hungary.
Pflugers Arch. 2012 Mar;463(3):459-75. doi: 10.1007/s00424-011-1059-6. Epub 2011 Dec 6.
Myofibroblasts play central roles in wound healing, deposition of the extracellular matrix and epithelial function. Their functions depend on migration and proliferation within the subepithelial matrix, which results in accelerated cellular metabolism. Upregulated metabolic pathways generate protons which need to be excreted to maintain intracellular pH (pH(i)). We isolated human gastric myofibroblasts (HGMs) from surgical specimens of five patients. Then we characterized, for the first time, the expression and functional activities of the Na(+)/H(+) exchanger (NHE) isoforms 1, 2 and 3, and the functional activities of the Na(+)/HCO(3)(-) cotransporter (NBC) and the anion exchanger (AE) in cultured HGMs using microfluorimetry, immunocytochemistry, reverse transcription polymerase chain reaction and immunoblot analysis. We showed that NHE1-3, NBC and AE activities are present in HGMs and that NHE1 is the most active of the NHEs. In scratch wound assays we also demonstrated (using the selective NHE inhibitor HOE-642) that carbachol and insulin like growth factor II (IGF-II) partly stimulate migration of HGMs in a NHE1-dependent manner. EdU incorporation assays revealed that IGF-II induces proliferation of HGMs which is inhibited by HOE-642. The results indicate that NHE1 is necessary for IGF-II-induced proliferation response of HGMs. Overall, we have characterized the pH(i) regulatory mechanisms of HGMs. In addition, we demonstrated that NHE1 activity contributes to both IGF-II- and carbachol-stimulated migration and that it is obligatory for IGF-II-induced proliferation of HGMs.
肌成纤维细胞在伤口愈合、细胞外基质的沉积和上皮功能中起着核心作用。它们的功能依赖于在黏膜下层基质中的迁移和增殖，这导致细胞代谢加速。上调的代谢途径产生质子，需要排出以维持细胞内 pH 值 (pH(i))。我们从五名患者的手术标本中分离出人胃肌成纤维细胞 (HGM)。然后，我们首次表征了培养的 HGM 中 Na(+)/H(+)交换器 (NHE) 亚型 1、2 和 3 的表达和功能活性，以及 Na(+)/HCO(3)(-)共转运体 (NBC) 和阴离子交换器 (AE) 的功能活性，使用微荧光法、免疫细胞化学、逆转录聚合酶链反应和免疫印迹分析。我们表明，NHE1-3、NBC 和 AE 活性存在于 HGM 中，并且 NHE1 是最活跃的 NHE。在划痕伤口测定中，我们还表明（使用选择性 NHE 抑制剂 HOE-642），乙酰胆碱和胰岛素样生长因子 II (IGF-II) 部分以 NHE1 依赖性方式刺激 HGM 的迁移。EdU 掺入测定表明，IGF-II 诱导 HGM 增殖，HOE-642 抑制其增殖。结果表明，NHE1 是 IGF-II 诱导的 HGM 增殖反应所必需的。总的来说，我们已经描述了 HGM 的 pH(i) 调节机制。此外，我们表明 NHE1 活性有助于 IGF-II 和乙酰胆碱刺激的迁移，并且它是 IGF-II 诱导的 HGM 增殖所必需的。
