Nordström T, Shrode L D, Rotstein O D, Romanek R, Goto T, Heersche J N, Manolson M F, Brisseau G F, Grinstein S
Division of Cell Biology, Hospital for Sick Children, Toronto, Ontario, Canada M5G 1X8.
J Biol Chem. 1997 Mar 7;272(10):6354-60. doi: 10.1074/jbc.272.10.6354.
Proton extrusion into an extracellular resorption compartment is an essential component of bone degradation by osteoclasts. Chronic metabolic acidosis is known to induce negative calcium balance and bone loss by stimulating osteoclastic bone resorption, but the underlying mechanism is not known. The present studies were undertaken to evaluate whether chronic acidosis affects proton extrusion mechanisms in osteoclasts cultured on glass coverslips. Acidosis, mimicked experimentally by maintaining the cells at extracellular pH 6.5, rapidly lowered intracellular pH to 6.8. However, after 2 hours, a proportion of cells demonstrated the capacity to restore intracellular pH to near normal levels. To define the mechanism responsible for this recovery, the activity of individual H+ transport pathways was analyzed. We found that chronic acid treatment for up to 6 h did not significantly affect the cellular buffering power or Na+/H+ antiport activity. In contrast, chronic acidosis activated vacuolar H+ pumps in the osteoclasts. Although only approximately 5% of the control cells displayed proton pump activity, about 40% of cells kept at extracellular pH 6. 5 for 4-6 h were able to recover from the acute acid load by means of bafilomycin A1-sensitive proton extrusion. Conversely, the H+-selective conductance recently described in the plasma membrane of osteoclasts was clearly inhibited in the cells exposed to chronic acidosis. Following acid treatment, the activation threshold of the H+ conductance was shifted to more positive potentials, and the current density was significantly reduced. Considered together, these results suggest that induction of plasmalemmal vacuolar type ATPase activity by chronic acidosis, generated either systemically due to metabolic disease or locally at sites of inflammation, is likely to stimulate osteoclastic bone resorption and thus to promote bone loss.
质子排泌到细胞外吸收区室是破骨细胞降解骨组织的一个重要组成部分。已知慢性代谢性酸中毒通过刺激破骨细胞骨吸收诱导负钙平衡和骨质流失，但其潜在机制尚不清楚。本研究旨在评估慢性酸中毒是否会影响在玻璃盖玻片上培养的破骨细胞的质子排泌机制。通过将细胞维持在细胞外pH 6.5来模拟酸中毒，可迅速将细胞内pH降至6.8。然而，2小时后，一部分细胞表现出将细胞内pH恢复到接近正常水平的能力。为了确定这种恢复的机制，分析了各个H⁺转运途径的活性。我们发现，长达6小时的慢性酸处理对细胞缓冲能力或Na⁺/H⁺反向转运活性没有显著影响。相反，慢性酸中毒激活了破骨细胞中的液泡H⁺泵。虽然只有约5%的对照细胞显示出质子泵活性，但在细胞外pH 6.5下维持4 - 6小时的细胞中，约40%能够通过巴弗洛霉素A1敏感的质子排泌从急性酸负荷中恢复。相反，最近在破骨细胞质膜中描述的H⁺选择性电导在暴露于慢性酸中毒的细胞中明显受到抑制。酸处理后，H⁺电导的激活阈值向更正的电位移动，电流密度显著降低。综合考虑，这些结果表明，由于代谢疾病全身产生或在炎症部位局部产生的慢性酸中毒诱导质膜液泡型ATP酶活性，可能会刺激破骨细胞骨吸收，从而促进骨质流失。
