注射催产素对小鼠骨密度和成骨细胞分化影响的实验研究

Effects of oxytocin injection on bone mineral density and osteoblast differentiation in mice

目的通过建立雌性小鼠骨质疏松模型并给予催产素( oxytocin,OT)腹腔注射来验证催产素对骨代谢的调控作用。方法选取同批次2 月龄雌性C57 /B6 小鼠15 只,随机分成3 组,每组5 只:①对照组( C),②造模组( OVX),③造模+催产素注射组( OVX+OT)。注射催产素前和注射8 周后分别测定各组小鼠左股骨和腰椎L4~ 6的骨密度,并提取小鼠骨髓细胞定向诱导分化,进行碱性磷酸酶( alkaline phosphatase,ALP)和Von Kossa 染色以鉴定成骨细胞分化水平,通过qPCR检测分化相关因子表达水平。结果OVX 组小鼠骨密度较C 组显著降低( P<0. 001),OVX+OT 组小鼠骨密度较OVX 组显著升高( P<0. 001)。ALP 染色显示,OVX 组成骨细胞分化较C 组差,细胞集落最少;OVX+OT 组成骨细胞分化情况较OVX 组好,染色面积增加了322. 5%。Von Kossa 染色发现OVX 组细胞集落和钙结节较C 组少且小,OVX+OT 组细胞集落和钙结节较OVX 组多,染色面积增加了62. 3%。而进一步添加催产素体外培养的成骨细胞其分化水平均比未加入催产素组有所提高。qPCR结果显示催产素能够促进成骨相关基因OPN、Runx2 和Osterix 的表达( P<0. 001)。结论催产素促进小鼠体内的骨合成代谢作用,提示其对骨质疏松症治疗具有潜在用途。

Objective To investigate and validate the regulation of oxytocin on bone metabolism by establishment of osteoporosis mouse model and injection of oxytocin in vivo. Methods Fifteen two-month-old C57/B6 mice were randomly divided into 3 groups,control group( C),OVX group( OVX),and OVX + oxytocin group( OVX+OT),with 5 mice in each group. After injecting oxytocin( 5 μg/mice/day,s. c.) for 8 weeks,bone mineral density( BMD) of L4-6 and the left femur of mice were measured. Mouse bone marrow cells were extracted and then stained with alkaline phosphatase( ALP) and Von Kossa staining to identify the level of osteoblast differentiation. Moreover,qPCR was used to identify the expression levels of differentiation-related genes. Results BMD of mice in OVX group was markedly lower than that of mice in C group( P<0. 0001). Oxytocin injection observably increased BMD of mice in OVX +OT group compared with mice in OVX group( P < 0. 0001). ALP staining result showed that the osteoblast differentiation in OVX group was worse than that in C group,and the cell colony was the least. The osteoblast differentiation in OVX +OT group was better than that in OVX group,with an increased dying area by 322. 5%. Von Kossa staining result showed that the cell colonies and calcium nodules in OVX group were smaller and less than that in C group.OVX+OT group had more cell colonies and calcium nodules compared with OVX group,with an increased dying area by 62. 3%.Furthermore,the differentiation of osteoblasts with addition of oxytocin in vitro was better than that without oxytocin. qPCR result indicated that oxytocin up-regulated the expression of osteogenesis-related genes such as RUNX2,OSTERIX,and OPN at the mRNA level( P<0. 0001). Conclusion Oxytocin enhances the anabolic action in regulating bone mass in mice,which suggests the potential use of this hormone for the treatment of osteoporosis.