钙磷对体外培养成骨细胞增殖、分化和矿化影响的研究

目的研究钙、磷对体外培养成骨细胞(OB)增殖、分化及矿化的作用,探讨钙、磷对骨代谢的影响。方法在体外培养OB的基础上,分别添加1、2、4mmol/L钙、1、2、4mmol/L磷及2mmol/L钙+1mmol/L磷(钙磷比2∶1)、1mmol/L钙+2mmol/L磷(钙磷比1∶2)。检测细胞增殖、碱性磷酸酶(ALP)活性、骨桥蛋白(OPN)和Ⅰ型胶原(Col-Ⅰ)含量及其mRNA表达情况。结果与对照组比,饲料中添加4mmol/L钙从D2开始促进OB增殖,差异显著(P<0.05),添加1、2mmol/L钙从D5开始差异显著(P<0.05),而钙磷比2:1与1:2到D8差异才显著(P<0.05)。除1mmol/L钙组外,各试验组从D2起均抑制细胞内ALP活性(P<0.05),在D5抑制其mRNA表达(P<0.01),在D2、D8促进其mRNA表达(P<0.05)。除1mmol/L钙组外,各试验组在D2均促进Col-Ⅰ分泌(P<0.01),但钙各组和1mmol/L磷组在D5、D8及2mmol/L磷组和钙磷比(1:2)组在D8均抑制其分泌(P<0.05)。各试验组,在D2、D8均促进Col-ⅠmRNA表达(P<0.01),在D5除钙磷比(1:2)组外均抑制其mRNA表达(P<0.01)。各试验组促进OPNmRNA表达(P<0.01),除1、2mmol/L钙组外,各实验组从D2起均促进其分泌(P<0.0)。结论添加钙及2种钙磷比(2:1,1:2)均能抑制成骨细胞早期分化,诱导其增殖及重叠生长,促进其基质成熟及矿化,而磷对其增殖作用不明显。

STUDY ON THE INFLUENCE OF CALCIUM,PHOSPHORUS ON PROLIFERATION, DIFFERENTIATION AND MINERALIZATION OF OSTEOBLASTS IN VITRO

Objective To clarify the influence of Ca and P on bone metabolism by investigating theeir effects on proliferation,differentiation and mineralization of osteoblasts(OBs)in vitro.Method OBs were cultured in media with Ca(1,2,4 mmol/L),P(1,2,4 mmol/L),Ca:P 1:2(1 mmol/L Ca~(+2) mmol/LP),and Ca:P 2:1(2 mmol/L Ca+1 mmol/L P).Cell proliferation,activity of alkaline phosphatase(ALP),content of osteopontin (OPN)and collagen type Ⅰ(Col-Ⅰ)of OBs,and their corresponding mRNA expression were measured.Results Compared with control group,Ca at 4 mmol/L promoted OBs proliferation significanty(P<0.05)from D2,while Ca at 1 and 2 mmoI/L from D5(P<0.05),and Ca:P(1:2,2:1)until D8.(P<0.05).All groups except 1 mmoI/LCa inhibited the activity of ALP(P<0.05)on D2,and inhibited its mRNA expression on D5(P<0.01),but enhanced its mRNA expression on D2 and D8(P<0.05).All groups except 1 mmol/L Ca promoted Col-Ⅰsecretion(P<0.01)on D2 but all Ca groups and 1 mmol/L P group on D5,D8,and 2 mmol/L P, Ca:P(1:2)group on D8 inhibited its secretion(P<0.05).All groups promoted Col-ⅠmRNA expression on D2、D8(P<0.01),but inhibited its mRNA expression on D5(P<0.01)except Ca:P(1:2)group.All groups enhanced OPN secretion(P<0.05)except 1,2 mmol/L Ca group on D2.All groups promoted OPN mRNA expression(P<0.01).Conclusion Adding Ca.and two Ca:P(2:1,1:2)Can inhibit early difierentiation and promote proliferation of OBs and induce matrix maturation,but P shows no significant effects on OBs proliferation.