In this paper, we investigate the effect and the possible mechanism of high glucose levels on the calcification of human aortic smooth muscle cells （HASMCs）. HASMCs were divided into four groups： normal glucose gro...In this paper, we investigate the effect and the possible mechanism of high glucose levels on the calcification of human aortic smooth muscle cells （HASMCs）. HASMCs were divided into four groups： normal glucose group （NG）, osmolality control group （OC）, high glucose group （HG, HASMCs culture medium containing 30 mmol/L glucose）, and high glucose plus recombinant human Noggin protein （bone morphogenetic protein-2 （BMP-2） antagonist） group （HN）. The mRNA levels and the protein expressions of BMP-2 and core binding factor alpha-1 （Cbfa-1） were measured by real-time quantitative polymerase chain reaction （PCR） and Western blot. After induced by 10 mmol/L β-glycerol phosphoric acid, cells were had＇vested for assessments of alkaline phosphatase （ALP） activities at Days 1,2, and 3, and intracellular calcium contents at Days 7 and 14, respectively. High glucose levels increased the mRNA levels and the protein expressions of BMP-2 and Cbfa-1 （P〈0.05）. The expression of Cbfa-1 was partially blocked by Noggin protein （P〈0.05）, while BMP-2 was not （P〉0.05）. After being induced by β-glycerol phosphoric acid, high glucose levels increased the ALP activity [（48.63±1.03） vs. （41.42±2.28） U/mg protein, Day 3; P〈0.05] and the intracellular calcium content [（2.76±0.09） vs. （1.75±0.07） μmol/mg protein, Day 14; P〈0.05] in a time-dependent manner when compared with the NG group, while the ALP activity could not be blocked by Noggin protein [（48.63±1.03） vs. （47.37±0.97） U/mg protein, Day 3; P〉0.05]. These results show that high glucose levels can evoke the calcification of HASMCs by inducing osteoblastic trans-differentiation and intracellular calcium deposition via the BMP-2/Cbfa-1 pathway, which can be partially blocked by Noggin protein.展开更多
文摘目的探讨巴戟天多糖含药血清对SD大鼠骨髓间充质干细胞(BMSCs)骨向分化过程中核心结合因子ɑ-1(Cbfɑ-1)m RNA表达的影响。方法全骨髓贴壁法分离培养SD大鼠BMSCs,采用p NPP法检测不同浓度r巴戟天多糖含药血清(高、中、低)对BMSCs碱性磷酸酶(ALP)活性的影响,并以其最佳作用浓度进行后续实验,后续实验分成空白组、成骨诱导组、巴戟天多糖含药血清组、联合组(巴戟天多糖含药血清组+成骨诱导组),各组用药14d,采用RT-PCR技术检测各组BMSCs Cbfa-1m RNA表达的情况。结果与对照组比较,不同剂量的巴戟天多糖含药血清均可提高BMSCs分泌ALP(F=126.278,P<0.05),其中高剂量组的作用强度高于其它剂量组;与空白组相比,成骨诱导组、巴戟天多糖含药血清组、联合组均能上调BMSCs中Cbfa-1 m RNA的表达(F=261.412,P<0.05);但与成骨诱导组相比,巴戟天多糖含药血清组不及成骨诱导组,两者差异有统计学意义(t=26.721,P<0.05)。结论巴戟天多糖含药血清能够促进BMSCs向成骨细胞分化,并能上调Cbfa-1 m RNA的表达,但其作用不及成骨诱导强。
文摘In this paper, we investigate the effect and the possible mechanism of high glucose levels on the calcification of human aortic smooth muscle cells （HASMCs）. HASMCs were divided into four groups： normal glucose group （NG）, osmolality control group （OC）, high glucose group （HG, HASMCs culture medium containing 30 mmol/L glucose）, and high glucose plus recombinant human Noggin protein （bone morphogenetic protein-2 （BMP-2） antagonist） group （HN）. The mRNA levels and the protein expressions of BMP-2 and core binding factor alpha-1 （Cbfa-1） were measured by real-time quantitative polymerase chain reaction （PCR） and Western blot. After induced by 10 mmol/L β-glycerol phosphoric acid, cells were had＇vested for assessments of alkaline phosphatase （ALP） activities at Days 1,2, and 3, and intracellular calcium contents at Days 7 and 14, respectively. High glucose levels increased the mRNA levels and the protein expressions of BMP-2 and Cbfa-1 （P〈0.05）. The expression of Cbfa-1 was partially blocked by Noggin protein （P〈0.05）, while BMP-2 was not （P〉0.05）. After being induced by β-glycerol phosphoric acid, high glucose levels increased the ALP activity [（48.63±1.03） vs. （41.42±2.28） U/mg protein, Day 3; P〈0.05] and the intracellular calcium content [（2.76±0.09） vs. （1.75±0.07） μmol/mg protein, Day 14; P〈0.05] in a time-dependent manner when compared with the NG group, while the ALP activity could not be blocked by Noggin protein [（48.63±1.03） vs. （47.37±0.97） U/mg protein, Day 3; P〉0.05]. These results show that high glucose levels can evoke the calcification of HASMCs by inducing osteoblastic trans-differentiation and intracellular calcium deposition via the BMP-2/Cbfa-1 pathway, which can be partially blocked by Noggin protein.
