Characterization of acid flux in osteoclasts from patients harboring a G215R mutation in ClC-7.

Abstract

The chloride-proton antiporter ClC-7 has been speculated to be involved in acidification of the lysosomes and the resorption lacunae in osteoclasts; however, neither direct measurements of chloride transport nor acidification have been performed. Human osteoclasts harboring a dominant negative mutation in ClC-7 (G215R) were isolated, and used these to investigate bone resorption measured by CTX-I, calcium release and pit scoring. The actin cytoskeleton of the osteoclasts was also investigated. ClC-7 enriched membranes from the osteoclasts were isolated, and used to test acidification rates in the presence of a V-ATPase and a chloride channel inhibitor, using a H(+) and Cl(-) driven approach. Finally, acidification rates in ClC-7 enriched membranes from ADOII osteoclasts and their corresponding controls were compared. Resorption by the G215R osteoclasts was reduced by 60% when measured by both CTX-I, calcium release, and pit area when comparing to age and sex matched controls. In addition, the ADOII osteoclasts showed no differences in actin ring formation. Finally, V-ATPase and chloride channel inhibitors completely abrogated the H(+) and Cl(-) driven acidification. Finally, the acid influx was reduced by maximally 50% in the ClC-7 deficient membrane fractions when comparing to controls. These data demonstrate that ClC-7 is essential for bone resorption, via its role in acidification of the lysosomes and resorption lacunae in osteoclasts.

• Authors: Kim Henriksen, Jeppe Gram, Anita Vibsig Neutzsky-Wulff, Vicki Kaiser Jensen, Morten H Dziegiel, Jens Bollerslev, Morten A Karsdal
• Journal: Biochemical and biophysical research communications (Biochem Biophys Res Commun) Vol. 378 Issue 4 Pg. 804-9 (Jan 23 2009) ISSN: 1090-2104 [Electronic] United States
• PMID: 19070589 (Publication Type: Journal Article)

Chemical References

• Acids
• CLCN7 protein, human
• Chloride Channels
• Arginine
• Vacuolar Proton-Translocating ATPases
• Calcium
• Glycine

Topics

• Acids (metabolism)
• Arginine (genetics)
• Bone Resorption (genetics, metabolism)
• Calcium (metabolism)
• Chloride Channels (antagonists & inhibitors, genetics, physiology)
• Genes, Dominant
• Glycine (genetics)
• Humans
• Hydrogen-Ion Concentration
• Lysosomes (metabolism)
• Mutation
• Osteoclasts (metabolism, ultrastructure)
• Vacuolar Proton-Translocating ATPases (antagonists & inhibitors, metabolism)