体外培养人成骨细胞BGP、IGF-1基因表达的增龄改变

Influence of Donor Age on Gene BGP,IGF?1 Expression of Human Osteoblastic Cells Cultured in Vitro

目的 探讨体外培养不同年龄组人成骨细胞BGP、IGF 1基因表达的增龄改变规律。方法 选择≤ 5岁、3 0～ 4 0岁、5 0～ 60岁、≥ 70岁 4个年龄组的松质骨 ,用 1∶1DMEM HamF 12培养液进行培养。不同年龄组两代人成骨细胞培养至 14d抽提细胞内总RNA ,应用RT PCR方法半定量检测BGP、IGF 1基因表达情况 ,并进行各年龄组比较。结果 各年龄组体外培养至 14d的两代人成骨细胞所抽提的细胞内总RNA量随增龄而降低 (男r=- 0 .783 ,女r=- 0 .873 ,均P <0 .0 0 1) ;各年龄组成骨细胞均检测到BGP与IGF 1的mRNA表达。BGP与IGF 1mRNA表达量于幼年组较低 ,3 0～ 4 0岁最高 ,之后随年龄增长逐渐降低 ,其表达量和年龄明显相关 (BGP男r2 =0 .3 2 8,女r2 =0 .3 0 1;IGF 1男r2 =0 .3 0 3 ,女r2 =0 .3 3 8,均P <0 .0 5 ) ,性别之间未见明显差异。结论 体外培养人成骨细胞的总RNA量随增龄而降低 ,不同年龄组人成骨细胞均表达BGP、IGF 1mRNA 。

Purpose: To investigate the law of age-related changes of gene bone gla protein(BGP) and insulin-like growth factor-1 (IGF-1) expression in human osteoblastic cells of different donor age cultured in vitro. Methods: Human osteoblasts of different donor age (7 aged&le5 years, male 3, female 4; 10 aged 30 - 40 years, male 7, female 3; 12 aged 50 - 60 years, male 7, female 5; and 14 aged≥70 years, male 5, female 9) were isolated and cultured with the tissue-fragment-migrating-growth method. Total RNA was extracted with TRIzol reagent from each sample when the osteoblasts of different donor age in second passage were cultured for 14 days. RNA yield and purity were checked with ultraviolet spectrophotometer. Reverse transcriptase-polymerase chain reaction (RT-PCR) was performed with primers corresponding to BGP and IGF-1. mRNA expression of BGP and IGF-1 from each sample was measured with RT-PCR in semi-quantity and compared within different groups. Results: Quantity of total RNA in osteoblasts was decreased with donor aging when the human osteoblasts were cultured for 14 days (for male r = -0.783; for female, r = -0.873, P2 = 0.328, for female, r2 = 0. 301; IGF-1, for male, r2 = 0. 303, for female, r2 = 0. 338, P<0.05 respectively). There is no significant difference between sexes. Conclusions: Quantity of total RNA in human osteoblasts cultured in vitro was decreased with donor aging. Human osteoblasts from different donor age cultured in vitro can express BGP and IGF-1 mRNA. Their quantity by RT-PCR analyses was changed with donor aging.