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Regulation of apical membrane Na+/H+ exchangers NHE2 and NHE3 in intestinal epithelial cell line C2/bbe 总被引:3,自引:0,他引:3
McSwine Rebecca L.; Musch Mark W.; Bookstein Crescence; Xie Yue; Rao Mrinalini; Chang Eugene B. 《American journal of physiology. Cell physiology》1998,275(3):C693
We examined the regulation of theNa+/H+exchangers (NHEs) NHE2 and NHE3 by expressing them in human intestinalC2/bbe cells, which spontaneously differentiate and have little basalapical NHE activity. Unidirectional apical membrane22Na+influxes were measured in NHE2-transfected (C2N2) and NHE3-transfected (C2N3) cells under basal and stimulated conditions, and their activities were distinguished as the HOE-642-sensitive and -insensitive components of5-(N,N-dimethyl)amiloride-inhibitableflux. Both C2N2 and C2N3 cells exhibited increased apical membrane NHEactivity under non-acid-loaded conditions compared with nontransfected control cells. NHE2 was inhibited by 8-(4-chlorophenylthio)adenosine 3',5'-cyclic monophosphate and thapsigargin, was stimulatedby serum, and was unaffected by cGMP- and protein kinase C-dependent pathways. In contrast, NHE3 was inhibited by all regulatory pathways examined. Under acid-loaded conditions (which increase apical Na+ influx), NHE2 and NHE3exhibited similar patterns of regulation, suggesting that the secondmessenger effects observed were not secondary to effects on cell pH.Thus, in contrast to their expression in nonepithelial cells, NHE2 andNHE3 expressed in an epithelial cell line behave similarly toendogenously expressed intestinal apical membrane NHEs. We concludethat physiological regulation and function of epithelium-specific NHEsare dependent on tissue-specific factors and/or conditionalrequirements. 相似文献
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Tissue distribution of Na+/H+ exchanger isoforms NHE2 and NHE4 in rat intestine and kidney 总被引:2,自引:0,他引:2
Bookstein Crescence; Xie Yue; Rabenau Karen; Musch Mark W.; McSwine Rebbecca L.; Rao Mrinalini C.; Chang Eugene B. 《American journal of physiology. Cell physiology》1997,273(5):C1496
We present evidence that tissue distribution of two highlyconservedNa+/H+exchanger isoforms, NHE2 and NHE4, differs significantly from previously published reports. Riboprobes unique to each of these antiporters, from 5' (noncoding and coding) and 3' codingregions, were used to analyze mRNA from adult rat kidney and intestine by ribonuclease protection assay and in situ hybridization. In contrastto earlier work that concluded that both NHE2 and NHE4 were expressedthroughout the intestine and in the kidney, our data show that there isno NHE2 message in the kidney and NHE4 is not expressed in small orlarge intestine. Analyses of intestinal epithelial and kidney membraneproteins by an NHE2-specific antibody identified a doublet at <90 kDain intestine but not in kidney. NHE2 is highly expressed in theNa+-absorptive epithelium ofjejunum, ileum, and ascending and descending colon. NHE4 mRNA messageis found in the inner medulla of the kidney as previously reported (C. Bookstein, M. W. Musch, A. DePaoli, Y. Xie, M. Villereal, M. C. Rao,and E. B. Chang. J. Biol. Chem. 269:29704-29709, 1994) and not in the intestine. From these data, wespeculate that neither NHE2 nor NHE4 has a role in renalNa+ absorption. NHE2 is likelyinvolved in gut Na+ absorption,whereas NHE4 may have a specialized role in cell volume rectificationof inner medullary collecting duct cells. Knowledge of the correcttissue and cell-specific distribution of these two antiporters shouldhelp significantly in understanding their physiological roles. 相似文献
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C. Bookstein M.W. Musch P.K. Dudeja R.L. McSwine Y. Xie T.A. Brasitus M.C. Rao E.B. Chang 《The Journal of membrane biology》1997,160(3):183-192
This report presents a study of the effects of the membrane fluidizer, benzyl alcohol, on NHE isoforms 1 and 3. Using transfectants
of an NHE-deficient fibroblast, we analyzed each isoform separately. An increase in membrane fluidity resulted in a decrease
of ≈50% in the specific activities of both NHE1 and NHE3. Only V
max was affected; K
Na was unchanged. This effect was specific, as Na+, K+, ATPase activity was slightly stimulated. Inhibition of NHE1 and NHE3 was reversible and de novo protein synthesis was not required to restore NHE activity after washout of fluidizer. Inhibition kinetics of NHE1 by amiloride,
5-(N,N-dimethyl)amiloride (DMA), 5-(N-hexamethyl)amiloride (HMA) and 5-(N-ethyl-N-isopropyl)amiloride (EIPA) were largely
unchanged. Half-maximal inhibition of NHE3 was also reached at approximately the same concentrations of amiloride and analogues
in control and benzyl alcohol treated, suggesting that the amiloride binding site was unaffected. Inhibition of vesicular
transport by incubation at 4°C augmented the benzyl alcohol inhibition of NHE activity, suggesting that the fluidizer effect
does not solely involve vesicle trafficking. In summary, our data demonstrate that the physical state of membrane lipids (fluidity)
influences Na+/H+ exchange and may represent a physiological regulatory mechanism of NHE1 and NHE3 activity.
Received: 23 January 1997/Revised: 1 August 1997 相似文献
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