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Am J Physiol Cell Physiol 275: C1330-C1341, 1998;
0363-6143/98 $5.00
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Vol. 275, Issue 5, C1330-C1341, November 1998

Na+-K+-Clminus cotransport in human fibroblasts is inhibited by cytomegalovirus infection

Lilia M. Maglova1, William E. Crowe1, Peter R. Smith1, Aníbal A. Altamirano2, and John M. Russell1

1 Department of Physiology, Allegheny University of the Health Sciences, Philadelphia, Pennsylvania 19129; and 2 Instituto de Investigaciones Cardiológicas, Facultad de Medicina, Universidad de Buenos Aires, 1122 Buenos Aires, Argentina

We examined the effects of human cytomegalovirus (HCMV) infection on the Na+-K+-Cl- cotransporter (NKCC) in a human fibroblast cell line. Using the Cl--sensitive dye MQAE, we showed that the mock-infected MRC-5 cells express a functional NKCC. 1) Intracellular Cl- concentration ([Cl-]i) was significantly reduced from 53.4 ± 3.4 mM to 35.1 ± 3.6 mM following bumetanide treatment. 2) Net Cl- efflux caused by replacement of external Cl- with gluconate was bumetanide sensitive. 3) In Cl--depleted mock-infected cells, the Cl- reuptake rate (in HCO-3-free media) was reduced in the absence of external Na+ and by treatment with bumetanide. After HCMV infection, we found that although [Cl-]i increased progressively [24 h postexposure (PE), 65.2 ± 4.5 mM; 72 h PE, 80.4 ± 5.0 mM], the bumetanide and Na+ sensitivities of [Cl-]i and net Cl- uptake and loss were reduced by 24 h PE and abolished by 72 h PE. Western blots using the NKCC-specific monoclonal antibody T4 showed an approximately ninefold decrease in the amount of NKCC protein after 72 h of infection. Thus HCMV infection resulted in the abolition of NKCC function coincident with the severe reduction in the amount of NKCC protein expressed.

bumetanide; intracellular chloride concentration, MRC-5 fibroblasts


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