Osteoclast-stimulating activity of TIMP

由于TIMP-1和TIMP-2都被报道抑制骨吸收，我们研究了胎牛血清（FCS）中的TIMP-1和/或TIMP-2是否在体外影响骨细胞的骨吸收。首先，我们研究了抗TIMP单克隆抗体（α TIMP mAb）对破骨细胞骨吸收的影响。当α TIMP-1或α TIMP-2 mAb加入到含有10%FCS的培养基中时，与我们的预期相反，α TIMP-2 mAb显著抑制骨吸收，但α TIMP-1 mAb几乎不抑制骨吸收。然后检测从FCS中去除TIMP-1和/或TIMP-2对骨吸收的影响。无TIMP-2的FCS或无TIMP-1和TIMP-2的FCS均能显著抑制骨吸收，而无TIMP-1的FCS几乎不抑制骨吸收，这一结果与加入α TIMP单抗的结果一致。通过加入人重组TIMP，骨吸收活性几乎完全恢复。上述结果强烈地表明TIMP-2具有强有力的破骨细胞刺激活性。为了了解TIMP依赖性骨吸收的机制，我们计数并比较了含有10%FCS和不含TIMP-1和/或TIMP-2的FCS的每种培养基中抗酒石酸酸性磷酸酶（TRAP）阳性和多核细胞的数量。它们之间的数量基本上没有差异，这表明TIMP的作用似乎与成熟破骨细胞的功能表达有关，但不影响它们从祖细胞成熟。

As both TIMP-1 and TIMP-2 have been reported to inhibit bone resorption, we examined whether TIMP-1 and/or TIMP-2 in fetal calf serum (FCS), with which culture media were supplemented, affected osteoclastic bone resorption in vitro. At first we looked at the effect of anti-TIMP monoclonal antibodies (alpha TIMP mAbs) on the bone resorption by osteclasts. When alpha TIMP-1 or alpha TIMP-2 mAb was added to the culture medium containing 10% FCS,bone resorption was significantly suppressed with alpha TIMP-2 mAb against our expectation, but little with alpha TIMP-1 mAb. Then we checked the effect of TIMP-1 and/or TIMP-2 delection from FCS on the bone resorption. The bone resorption was significantly suppressed in either TIMP-2-free FCS or TIMP-1-and TIMP-2-free FCS,but little in TIMP-1-free FCS.These results are consistent with those obtained by the addition of alpha TIMP mAbs. The bone-resorbing activity was almost fully restored by the addition of human recombinant TIMPs. The above results strongly suggest that TIMP-2 has a potent osteoclast-stimulating activity. To get a clue to the mechanism of TIMP-dependent bone resorption, we counted and compared the number of tartrate-resistant acid phosphatase (TRAP)-positive and multinuclear cells in each culture medium containing either 10% FCS or TIMP-1-and/or TIMP-2-free FCS.There was essentially no difference in the number among them suggesting that TIMP's role seems to be related to the functional expression of matured osteoclasts, but not to their maturation from progenitor cells.