Cellular distribution and regulation of NHE-1 isoform of the Na-H exchanger in the avian osteoclast

The sodium-hydrogen exchanger (NHE) has been implicated in bone resorption by osteoclasts. We have studied expression of NHE-1, an isoform of the NHE, in chicken bone marrow mononuclear phagocyte precursors during differentiation into the osteoclast phenotype in culture, A monoclonal antibody raised against the carboxy-terminus of NHE-1 detected the presence of a similar to 100 kDa protein (similar to the mammalian form of NHE-1) in the osteoclasts, Laser scanning confocal microscopy revealed association with the alpha(v) beta(3) integrin and focal adhesion kinase (pp(125)FAK) at the basolateral membrane (BLM) of the osteoclast in addition to a more generalized cellular distribution. A fragment of avian NHE-1 cDNA was obtained by polymerase chain reaction cloning, and it was used to characterize expression of NHE-1 transcripts in cultured chicken osteoclast precursors. The avian NHE-1 message was a 3.9 kB band on Northern analysis, which differed from the mammalian message, Retinoic acid (RA) elicited an increase in the steady-state intracellular pH (pH(i)) from 6.87 to 7.10 in the absence of bicarbonate and was inhibited by ethylisopropylamiloride, an inhibitor of Na-H exchange, Using ribonuclease protection assays, we found that NHE-1 transcripts are induced as cells differentiate in vitro and in response to 13-cis-RA. Western blot analysis indicated that protein levels also increased in response to 13-cis-RA, Our results demonstrate expression of NHE-1 in avian osteoclasts with a complex cellular distribution in culture, and NHE-1 expression is induced as tells differentiate into mature osteoclasts in response to 13-cis-RA.