Effect of Alendronate on the metabolism of cartilaginous and disc cells in vitro

双膦酸类药物已广泛用于治疗骨质疏松症。最近，阿伦磷酸钠可能具有软骨保护作用，这可能与骨关节炎有关。在软骨和椎间盘退变期间，聚集素浓度显著下降，造成不幸的生物力学和生理后果。然而，关于阿伦磷酸钠对椎间盘细胞的影响的许多问题仍然没有答案。本研究的目的是探讨细胞外渗透压和氧分压在调节阿伦磷酸钠合成GAG中的作用及其对软骨和间盘基质转化的影响。从跖骨关节软骨组织和牛尾盘髓核分离细胞，包裹在海藻酸珠中，在含6%胎牛血清的DMEM中培养，细胞密度为400万个/ml。然后在21%氧气中培养5d，细胞密度为10^&lt；-1；-10^&lt；-12&gt；L/阿仑磷酸钠；以未加阿伦磷酸钠培养的细胞为对照，比较渗透压和氧浓度对细胞生长的影响。用酶法测定乳酸产量，用DMB法检测糖胺聚糖(GAG)的积累。用标准的S-硫酸盐放射性比色法测定硫酸盐GAG合成速率。结果表明，浓度为10~(-10)~(-10)~(-10)~(-8)~(-8)mol/l的阿伦磷酸钠能促进软骨细胞和视盘细胞培养5天后每百万细胞的GAG产量。当浓度为10~(-8)mol/L时，增强作用最强。病理条件下(270mOmol，1%氧气)抑制细胞代谢可阻止阿伦磷酸钠(10mlt；-8；gt；L)诱导的每百万细胞GAG产量的增加，而健康条件下(400mOmol，5%氧气)阿伦磷酸钠可促进GAG产生，细胞代谢(乳酸产生)和GAG合成。综上所述，这些数据表明阿伦磷酸钠可能对健康细胞具有软骨保护作用。

Bisphosphonates have been widely used for treatment of osteoporosis. Recently, alendronate may have chondroprotective effects that could be of relevance in osteoarthritis. Aggrecan concentration falls markedly during cartilaginous and disc degeneration with unfortunate biomechanical and physiological consequences. However, many questions on the effect of alendronate to disc cells remain unanswered. The goals of this study were to determine the role of extracellular osmolality and oxygen tension in regulating production of GAG by alendronate and its effects on cartilahious and disc matrix turnover.Cells were isolated from the cartilaginous tissue of metatarsal joint and the nucleus pulposus of bovine caudal discs, encapsulated in alginate beads, and cultured in DMEM containing 6% foetal calf serum(FCS) at 400 mOsmol at cell densities of 4 million cells/ml. They were then cultured for 5 days under 21% oxygen with 10^<-1>-10^<-12> mol/l alendronate ; cell cultured without alendronate served as control and the effects of osmolarity and oxygen concentration were compared. Lactate production was measured enzymatically and glycosaminoglycan(GAG) accumulation was examined using a DMB assay. Rate of sulfate GAG synthesis was measured using a standard^<35> S-sulfate radioactive method. In the result, Alendronate at concentrations of 10^<-10>-10^<-8> mol/1 potentiated the GAG production per million cells by cartilaginous and disc cells in 5 days culture. The potentiated effect was maximal at 10^<-8> mol/l. Alendronate(10^<-8> mol/l)-induced increases in GAG production per million cells are prevented by inhibition of cell metabolism under pathologic conditions(270 mOsmol,<1% oxygen) as seen the osteoarthritis and degenerated disc ; but alendronate increased GAG production, cell metabolism(lactate production) and GAG synthesis in healthy conditions(400 mOsmol, 5% oxygen). In conclusion, this data suggest that alendronate may have chondroprotective effects on healthy cells.