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 resorp tion by osteoclasts. We have studied expression of NHE-1, an isoform o f the NHE, in chicken bone marrow mononuclear phagocyte precursors dur ing differentiation into the osteoclast phenotype in culture, A monocl onal antibody raised against the carboxy-terminus of NHE-1 detected th e presence of a similar to 100 kDa protein (similar to the mammalian f orm of NHE-1) in the osteoclasts, Laser scanning confocal microscopy r evealed association with the alpha(v) beta(3) integrin and focal adhes ion kinase (pp(125)FAK) at the basolateral membrane (BLM) of the osteo clast in addition to a more generalized cellular distribution. A fragm ent of avian NHE-1 cDNA was obtained by polymerase chain reaction clon ing, and it was used to characterize expression of NHE-1 transcripts i n 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 i ntracellular pH (pH(i)) from 6.87 to 7.10 in the absence of bicarbonat e and was inhibited by ethylisopropylamiloride, an inhibitor of Na-H e xchange, Using ribonuclease protection assays, we found that NHE-1 tra nscripts are induced as cells differentiate in vitro and in response t o 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.