Albrecht Elke, Kolisek Martin, Viergutz Torsten, Zitnan Rudolf, Schweigel Monika
Muscle Biology and Growth, Research Institute for the Biology of Farm Animals (FBN), Wilhelm-Stahl-Allee 2, 18196, Dummerstorf, Germany.
J Comp Physiol B. 2008 Mar;178(3):285-95. doi: 10.1007/s00360-007-0221-0. Epub 2007 Nov 8.
In this study, we have studied the expression, localization, and functionality of vacuolar-type H(+)-ATPase (vH(+)-ATPase) and Na(+)/K(+)-ATPase in the bovine rumen epithelium. Compared with the intracellular pH (pH(i)) of control rumen epithelial cells (REC; 7.06 +/- 0.07), application of inhibitors selective for vH(+)-ATPase (foliomycin) and Na(+)/K(+)-ATPase (ouabain) reduced pH(i) by 0.10 +/- 0.03 and 0.18 +/- 0.03 pH-units, respectively, thereby verifying the existence of both functional proteins. Results from qRT-PCR and immunoblotting clearly confirm the expression of vH(+)-ATPase B subunit in REC. However, the amount of Na(+)/K(+)-ATPase mRNA and protein is tenfold and 11-fold of those of vH(+)-ATPase subunit B, respectively, reflecting a lower overall abundance of the latter in REC. Na(+)/K(+)-ATPase immunostaining has revealed the protein in the plasma membrane of all REC from the stratum basale to stratum granulosum, with the highest abundance in basal cells. In contrast, the vH(+)-ATPase B subunit has been detected in groups of cells only, mainly localized in the stratum spinosum and stratum granulosum of the epithelium. Furthermore, vH(+)-ATPase has been detected in the cell membrane and in intracellular pools. Thus, functional vacuolar-type H(+) pumps are expressed in REC and probably play a role in the adaptation of epithelial transport processes.
在本研究中，我们研究了液泡型H(+)-ATP酶（vH(+)-ATP酶）和Na(+)/K(+)-ATP酶在牛瘤胃上皮中的表达、定位及功能。与对照瘤胃上皮细胞（REC；细胞内pH值（pH(i)）为7.06±0.07）相比，应用对vH(+)-ATP酶（叶霉素）和Na(+)/K(+)-ATP酶（哇巴因）具有选择性的抑制剂分别使pH(i)降低了0.10±0.03和0.18±0.03个pH单位，从而证实了这两种功能蛋白的存在。定量逆转录聚合酶链反应（qRT-PCR）和免疫印迹结果清楚地证实了REC中vH(+)-ATP酶B亚基的表达。然而，Na(+)/K(+)-ATP酶mRNA和蛋白的量分别是vH(+)-ATP酶亚基B的量的10倍和11倍，这反映出后者在REC中的总体丰度较低。Na(+)/K(+)-ATP酶免疫染色显示，从基底层到颗粒层的所有REC的质膜中均有该蛋白，在基底细胞中的丰度最高。相比之下，仅在成组细胞中检测到vH(+)-ATP酶B亚基，主要定位于上皮的棘层和颗粒层。此外，在细胞膜和细胞内池中也检测到了vH(+)-ATP酶。因此，功能性液泡型H(+)泵在REC中表达，可能在上皮转运过程的适应中发挥作用。
