胶原蛋白肽含药血清促进成骨细胞增殖和分化

目的牛骨胶原蛋白肽(collagen peptides,CP)化合物能够显著增加股骨骨密度,并改善股骨的微结构,在骨质疏松症的预防和治疗中发挥重要的作用,它通过口服给药的形式吸收入肠。然而,此过程的分子机制尚不清楚。因此,本研究以阐明牛骨CP化合物血清对成骨细胞增殖与分化的影响。方法制备牛骨CP化合物大鼠血清,牛骨CP化合物处理的实验组和未经处理的对照组大鼠血清浓度为3%、6%、10%,加到无血清的DMEM溶液中,用MTT法和流式细胞术分别检测牛骨CP血清对MC3T3-E1细胞增殖和细胞周期的影响。茜素红矿化染色检测牛骨CP血清对成骨细胞矿化,用Pro Plus 6图像软件定量分析茜素红染色的含量。结果 MTT结果表明,3%、6%及10%的CP化合物含药血清与对照组相比,能够显著促进MC3T3-E1细胞增殖(P0.05)。然后将3%CP化合物含药血清,对照组(control,CN)血清分别作用于MC3T3成骨细胞,培养3 d,流式细胞术测定细胞周期。结果表明,与3%CN血清相比,3%CP血清组G1期比例显著减少,G2/S期比例显著增加(P0.01),表明牛骨CP通过促进G2/S期的百分含量而促进细胞的增殖。矿化染色结果表明CP血清处理21 d后能显著促进成骨细胞的矿化(P0.05)。结论牛骨CP血清在成骨细胞增殖与分化中起重要作用,为骨质疏松症的防治提供了理论依据。

Collagen peptides in serum promote the proliferation and differentiation of osteoblasts

Objective Bovine collagen peptides (CP) compounds could significantly elevate femoral bone mineral density and improve femoral bone microstructure, thus play an important role in the prevention and treatment of osteoporosis by oral administration and intestinal absorption. However, the molecular mechanisms of this process remain unclear. Therefore, we undertook experiments to clarify the effects of bovine CP compounds in serum on the proliferation and differentiation of osteoblast. Methods CP compounds in serum form were prepared, and rat serum containing bovine CP compounds (experimental groups) at the concentration of 3%, 6%, and 10% or control serum (control (CN) group) was added to DMEM without FBS. The effect of CP in serum on the proliferation and cell cycle of MC3T3 cells were examined by MTT and flow cytometry, respectively. Quantitative analysis of mineralization in osteoblasts using Alizarin red staining was measured by Image Software Pro Plus 6.0. Results We found that MTT values of MC3T3-E1 cells after treatment with bovine CP compounds at a concentration of 3%, 6% and 10% in serum were much higher than the control serum (P<0.05). MC3T3-E1 cells were harvested at 3d after treatment with 3% CP serum DMEM without FBS or CN serum. Cell cycle distribution analysis of MC3T3-E1 cells was conducted by flow cytometry. 3% CP compounds in serum led to increased cell cycle arrest in the G2/S phase at day 3 (P< 0.01), and G1 phase was significantly reduced compared with control serum.This suggested that CP compounds in serum might play an important role in osteoblast proliferation. Calcium deposit test showed that CP in serum significantly increased mineralization in osteoblasts at the 21st day (P<0.05). Conclusion Bovine CP compounds in serum played an important role in osteoblasts proliferation and differentiation, which provides a theoretical basis for the prevention and treatment of osteoporosis.