Document Detail

Immunolocalization of NHE8 in rat kidney.
MedLine Citation:
PMID:  15522984     Owner:  NLM     Status:  MEDLINE    
In situ hybridization studies demonstrated that Na+/H+ exchanger NHE8 is expressed in kidney proximal tubules. Although membrane fractionation studies suggested apical brush-border localization, precise membrane localization could not be definitively established. The goal of the present study was to develop isoform-specific NHE8 antibodies as a tool to directly establish the localization of NHE8 protein in the kidney by immunocytochemistry. Toward this goal, two sets of antibodies that label different NHE8 epitopes were developed. Monoclonal antibody 7A11 and polyclonal antibody Rab65 both specifically labeled NHE8 by Western blotting as well as by immunofluorescence microscopy. The immunolocalization pattern in the kidney seen with both antibodies was the same, thereby validating NHE8 specificity. In particular, NHE8 expression was observed on the apical brush-border membrane of all proximal tubules from S1 to S3. The most intense staining was evident in proximal tubules in the deeper cortex and medulla with a significant but somewhat weaker staining in superficial proximal tubules. Colocalization studies with gamma-glutamyltranspeptidase and megalin indicated expression of NHE8 on both the microvillar surface membrane and the coated-pit region of proximal tubule cells, suggesting that NHE8 may be subject to endocytic retrieval and recycling. Although colocalizing in the proximal tubule with NHE3, no significant alteration in NHE8 protein expression was evident in NHE3-null mice. We conclude that NHE8 is expressed on the apical brush-border membrane of proximal tubule cells, where it may play a role in mediating or regulating ion transport in this nephron segment.
Sunita Goyal; SueAnn Mentone; Peter S Aronson
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't; Research Support, U.S. Gov't, P.H.S.     Date:  2004-11-02
Journal Detail:
Title:  American journal of physiology. Renal physiology     Volume:  288     ISSN:  1931-857X     ISO Abbreviation:  Am. J. Physiol. Renal Physiol.     Publication Date:  2005 Mar 
Date Detail:
Created Date:  2005-02-04     Completed Date:  2005-03-09     Revised Date:  2011-04-28    
Medline Journal Info:
Nlm Unique ID:  100901990     Medline TA:  Am J Physiol Renal Physiol     Country:  United States    
Other Details:
Languages:  eng     Pagination:  F530-8     Citation Subset:  IM    
Dept. of Internal Medicine, Yale School of Medicine, 1 Gilbert St., TAC S-255, PO Box 208029, New Haven, CT 06520-8029, USA.
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MeSH Terms
Amino Acid Sequence
Antibodies, Monoclonal / biosynthesis,  immunology
Antibody Specificity
Biotin / metabolism
COS Cells
Cercopithecus aethiops
Electrophoresis, Polyacrylamide Gel
Fluorescent Antibody Technique, Direct
Kidney / anatomy & histology*,  metabolism*
Kidney Tubules, Proximal / metabolism
Membranes / drug effects,  metabolism
Mice, Knockout
Microvilli / metabolism
Molecular Sequence Data
Rats, Sprague-Dawley
Sodium-Hydrogen Antiporter / genetics,  metabolism*
Grant Support
Reg. No./Substance:
0/Antibodies, Monoclonal; 0/Sodium-Hydrogen Antiporter; 58-85-5/Biotin

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