骨康灵联合^(99)Tc-MDP治疗糖皮质激素诱发骨质疏松细胞分子机制研究

The study of cellular and molecular mechanism of the combination therapy using Gukangling decoction and technetium [^(99)Tc]methylene diphosphonate for glucocorticoid-induced osteoporosis

目的观察骨康灵和99Tc-MDP联合治疗糖皮质激素诱发的骨质疏松优于单独治疗的细胞分子机制。方法用SD大鼠随机分为DEX(地塞米松)、DEX-MDP(地塞米松+99 Tc-MDP 2mg/kg)、DEX-Z(地塞米松+骨康灵2ml/kg)和DEX-MDP-Z(地塞米松+99 Tc-MDP 2mg/kg+骨康灵2ml/kg)组。给药后颈动脉放血,制备含药血清,用MTT和PNPP法检测其对成骨细胞增殖与分化的影响;用TRAP染色和骨片吸收陷窝法观察其对破骨细胞的数目和功能影响;取大鼠股骨骨髓细胞,用Real-time PCR方法检测骨髓细胞RUNX2和PPARγ的表达。结果显示与对照组及单独治疗组相比联合用药组对成骨细胞有微弱的促分化作用,对破骨细胞有较强的抑制其数量和骨吸收的功能;同时联合用药能上调RUNX2与PPARγ的比值。结论中药骨康灵联合99Tc-MDP能有效抑制破骨细胞的数目和功能,同时促进骨髓间充质干细胞向成骨细胞前体分化,这可能是其优于单独治疗的细胞分子机制之一。

Objective To observe the cellular and molecular mechanism of the better effect of the combination therapy using Gukangling and technetium [ 99 Tc ] methylene diphosphonate （99 Tc-MDP） for glucocorticoid-induced osteoporosis than that of signal therapy. Methods SD Rats were randomly divided into DEX group, DEX-MDP group （2 mg/kg MDP）, DEX-Z group （2 mg/kg Gukangling）, and DEX- MDP-Z group. At the end of drug administration, blood from carotids of all rats was collected to produce drug containing serum. The effects on the proliferation and differentiation of osteoblasts were detected using MTT and PNPP methods. The effects on the number and function of osteoclasts were detected using the TRAP methods and observation of lacuna on bone slips. Rat bone marrow cells were collected from the femur. The expressions of RUNX2 and PPAR~, of the bone marrow cells were detected using real-time PCR method. Results Compared with the control group and the single therapy group, the combination therapy slightly promoted osteoblast differentiation, and strongly inhibited osteoclast number and osteoclastic bone absorption. The combination therapy also enhanced the ratio of RUNX2 and PPARγ. Conclusion The combination therapy using Gukangling and 99Tc-MDP can effectively inhibit osteoclast number and osteoclast function, and can promote differentiation of bone marrow mesenchymal cells into osteoblast precursors. This may be one of the reasons for the superior of combination therapy over single therapy.