乳铁蛋白对原代大鼠成骨细胞IGFBP-3、IGFBP-4、IGFBP-5mRNA表达的影响

The influence of lactoferrin on the expression of m RNA of IGFBP-3,IGFBP-4 and IGFBP-5 in primary rat osteoblasts

目的研究乳铁蛋白对原代大鼠成骨细胞胰岛素生长因子结合蛋白-3(IGFBP-3)、胰岛素生长因子结合蛋白-4(IGFBP-4)、胰岛素生长因子结合蛋白-5(IGFBP-5)m RNA表达的影响。方法用混合酶消化法分离大鼠颅骨成骨细胞,进行原代培养。细胞以6×103/cm2接种于六孔板内,用不同浓度乳铁蛋白即0μg/ml(对照组)、0.1μg/ml(乳铁蛋白1组)、1μg/ml(乳铁蛋白2组)、10μg/ml(乳铁蛋白3组)、100μg/ml(乳铁蛋白4组)、1000μg/ml(乳铁蛋白5组)干预原代大鼠成骨细胞1、3、5、7 d后提取总RNA,采用荧光定量PCR技术分析IGFBP-3、IGFBP-4、IGFBP-5 m RNA的表达。结果与对照组比较,各浓度组及时间段对IGFBP-3 m RNA表达的影响不稳定,时高时低。与对照组比较,除了乳铁蛋白1组外,其他浓度对IGFBP-4 m RNA的表达呈浓度梯度的抑制(P<0.01)。与对照组比较,实验干预第1天,呈浓度梯度抑制IGFBP-5 m RNA的表达(P<0.01),乳铁蛋白4、5组在第5、7天表现为继续抑制(P<0.01),而其他时间及浓度对IGFBP-5 m RNA的表达调控不稳定。结论乳铁蛋白影响原代大鼠成骨细胞IGFBP-3、IGFBP-4、IGFBP-5 m RNA的表达,其中乳铁蛋白对IGFBP-3、IGFBP-5 m RNA表达的调控不稳定,而乳铁蛋白抑制IGFBP-4 m RNA的表达呈浓度依赖性,浓度越高抑制作用越强。

Objective To research the influence of lactoferrin on the expression of mRNA of insulin-like growth factor binding protein-3 （IGFBP-3）,insulin-like growth factor binding protein-4 （IGFBP-4） and insulin-like growth factor binding protein-5 （IGFBP-5） in primary rat osteoblasts. Methods The skull osteoblasts of rat was isolated by mixed en-zyme digestion to do primary culture.Cells were inoculated in six-well plates by 6×10^3/cm2,primary rat osteoblasts were intervented 1,3,5,7 days by different concentrations of lactoferrin protein [0μg/ml （control group）,0.1μg/ml （lactoferrin group 1）, 1μg/ml （lactoferrin group 2）,10μg/ml （lactoferrin group 3）,100μg/ml （lactoferrin group 4）,1000μg/ml （lactoferrin group 5）, then total RNA was extracted.IGFBP-3,IGFBP-4,IGFBP-5 mRNA expression was analyzed by using quantitative PCR. Results Compared with the control group,the expression of IGFBP-3 mRNA was unstable after invented by lactoferrin in different times.Compared with the control group,except the lactoferrin group 1,the expression of IGFBP-4 mRNA was inhibited and depended on concentration.Compared with the control group,the expression of IGFBP-5 mRNA was in-hibited depended on concentration on the first day,while it was inhibited in lactoferrin group 4 and lactoferrin group 5 on the fifth and seventh day （P〈0.01）.The expression and regulation of IGFBP-5 mRNA at other time and concentration was unstable. Conclusion Primary rat osteoblasts influence the expression of IGFBP-3,IGFBP-4,IGFBP-5 mRNA,of which,the expression regulation of IGFBP-3,IGFBP-5 mRNA is unstable,while lactoferrin inhibits the expression of IGFBP-4 mRNA shows a concentration-dependent manner,the higher the concentration,the stronger the inhibition.