降钙素基因相关肽对乳腺癌细胞与成骨细胞共培养中骨代谢调控的研究

The regulatory effect of calcitonin gene-related peptide on bone metabolism of osteoblast cells co-cultured with breast cancer cells

目的:建立体外乳腺癌细胞与成骨细胞共培养体系,观察降钙素基因相关肽(calcitonin gene-related peptide,CGRP)对共培养条件下成骨细胞成熟和矿化能力的影响及其骨调节因子的变化。方法:乳腺癌细胞MDA-MB-231与成骨样细胞MG63共培养后,行CGRP干预,观察共培养条件下骨代谢调节因子护骨素(osteoprotegerin,OPG)、细胞核因子κB受体活化因子配体(receptor activator of NF-κB ligand,Rankl)和Runt相关转录因子2(Runt-related transcription factor2,Runx2)mRNA及蛋白的表达以及碱性磷酸酶的表达量和细胞矿化能力的变化。结果:与MG63细胞单独培养相比,MG63细胞与MDA-MB-231细胞共培养时Runx2和Rankl mRNA及蛋白的表达上调,同时伴有OPG mRNA及蛋白表达的下调(P<0.05);CGRP处理后,上述改变被逆转。此外,CGRP能明显增加MG63与MDA-MB-231细胞共培养后碱性磷酸酶的表达量,并促进细胞矿化结节的形成(P<0.05)。结论:乳腺癌细胞通过调节成骨细胞OPG-Rankl的表达促进破骨细胞的活性,通过调节Runx2的表达抑制成骨细胞的矿化活性。CGRP可逆转这一作用,进而促进骨修复。CGRP有望成为治疗乳腺癌骨转移的新制剂之一。

Objective：To construct co-culture system of breast cancer cells and osteoblasts in vitro and observe the effect of calcitonin gene-related peptide（CGRP） on the maturation and mineralizing activity of osteoblasts and the changes in bone regulating factors.Methods：The breast cancer cells MDA-MB-231 were co-cultured with osteoblast cells MG63,and then interfered with CGRP.The changes in the mRNA and protein expressions of bone metablism regulating factors osteoprotegerin（OPG）,receptor activator of NF-κB ligand（Rankl）,and Runt-related transcription factor 2（Runx2） were observed.The expression of alkaline phosphatase（ALP） was determined and the number of mineralized nodules was recorded.Results：The expressions of Rankl and Runx2 were up-regulated at both mRNA level and protein level,while the expression levels of OPG mRNA and protein were down-regulated in MG63 cells co-cultured with MDA-MB-231 cells（P〈0.05）.The changes could be reversed by CGRP treatment.CGRP significantly increased the production of ALP and stimulated the formation of the mineralized nodules when MG63 cells were co-cultured with MDA-MB-231 cells（P〈0.05）.Conclusion： Breast cancer cells MDA-MB-231 promotes the activity of osteoclast via regulating the OPG and Rank1 expressions in osteoblasts,and inhibits the mineralizing activity of osteoblast cells via the regulation of Runx2 expression.CGRP could reverse the effect of breast cancer cells on the osteoblast cells,thus prompting repair of bone tissues.CGRP may become a noval therapeutic agent for bone metastasis of breast cancer.