Support of bone mineral deposition by regulation of pH

• Published in: American Journal of Physiology: Cell Physiology Vol. 315; no. 4; pp. C587 - C597
• Main Authors: Blair, Harry C, Larrouture, Quitterie C, Tourkova, Irina L, Liu, Li, Bian, Jing Hao, Stolz, Donna Beer, Nelson, Deborah J, Schlesinger, Paul H
• Format: Journal Article
• Language: English
• Published: United States American Physiological Society 01.10.2018
• Subjects: Acridine orange; Adenosine Triphosphate - metabolism; Alkaline phosphatase; Alkaline Phosphatase - genetics; Alkaline Phosphatase - metabolism; Bone density; Bone growth; Bone matrix; Bone Matrix - growth & development; Bone Matrix - metabolism; Calcification, Physiologic - genetics; Calcium - metabolism; Calcium phosphates; Cavitation; Cell Differentiation; Cell Membrane - genetics; Cell Membrane - metabolism; Cellular biology; Chloride Channels - genetics; Collagen; Collagen (type I); Collagen Type I - chemistry; Collagen Type I - genetics; Durapatite - metabolism; Extracellular matrix; Humans; Hydrogen; Hydrogen-Ion Concentration; Hydroxyapatite; Ion Transport - genetics; Levamisole; Levamisole - pharmacology; Membrane vesicles; Mesenchymal Stem Cells - metabolism; Mineralization; Na+/H+-exchanging ATPase; Osteoblasts; Osteoblasts - metabolism; Osteogenesis; pH effects; Phosphates; Phosphates - metabolism; Protons; Sodium; Sodium - metabolism; Sodium-Hydrogen Exchanger 1 - genetics; Surface Plasmon Resonance; Vacuolar Proton-Translocating ATPases - chemistry; Vacuolar Proton-Translocating ATPases - genetics; chloride-hydrogen antiport; sodium-hydrogen exchange; hydroxyapatite deposition; vacuolar H+-ATPase
• ISSN: 0363-6143; 1522-1563
• DOI: 10.1152/ajpcell.00056.2018

Osteoblasts secrete collagen and isolate bone matrix from extracellular space. In the matrix, alkaline phosphatase generates phosphate that combines with calcium to form mineral, liberating 8 H per 10 Ca deposited. However, pH-dependent hydroxyapatite deposition on bone collagen had not been shown. We studied the dependency of hydroxyapatite deposition on type I collagen on pH and phosphate by surface plasmon resonance in 0-5 mM phosphate at pH 6.8-7.4. Mineral deposition saturated at <1 mM Ca but was sensitive to phosphate. Mineral deposition was reversible, consistent with amorphous precipitation; stable deposition requiring EDTA removal appeared with time. At pH 6.8, little hydroxyapatite deposited on collagen; mineral accumulation increased 10-fold at pH 7.4. Previously, we showed high expression Na /H exchanger (NHE) and ClC transporters in osteoblasts. We hypothesized that, in combination, these move protons across osteoblasts to the general extracellular space. We made osteoblast membrane vesicles by nitrogen cavitation and used acridine orange quenching to characterize proton transport. We found H transport dependent on gradients of chloride or sodium, consistent with apical osteoblast ClC family Cl ,H antiporters and basolateral osteoblast NHE family Na /H exchangers. Little, if any, active H transport, supported by ATP, occurred. Major transporters include cariporide-sensitive NHE1 in basolateral membranes and ClC3 and ClC5 in apical osteoblast membranes. The mineralization inhibitor levamisole reduced bone formation and expression of alkaline phosphatase, NHE1, and ClC5. We conclude that mineral deposition in bone collagen is pH-dependent, in keeping with H removal by Cl ,H antiporters and Na /H -exchangers. Periodic orientation hydroxyapatite is organized on type I collagen-coiled coils.