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探讨钙化的血管平滑肌细胞肾上腺髓质素生成和肾上腺髓质素受体系统一降钙素受体样受体和受体活性修饰蛋白基因表达的改变及其病理意义。采用β-甘油磷酸盐诱导培养的大鼠血管平滑肌细胞钙化;放射免疫法测定血管平滑肌细胞分泌的肾上腺髓质素含量;半定量逆转录聚合酶链反应测定细胞肾上腺髓质素、降钙素受体样受体和受体活性修饰蛋白的mRNA水平;原子吸收分光光度计测定细胞钙含量;碱性磷酸酶试剂盒测定血管平滑肌细胞碱性磷酸酶活性;β液体闪烁计数仪测定45Ca2+放射活性。结果发现,与非钙化血管平滑肌细胞比较,钙化血管平滑肌细胞内钙含量、45Ca2+摄入及碱性磷酸酶活性分别增加118％、174％和7倍(P<0.01);钙化细胞肾上腺髓质素分泌量增高99％(P<0.01),肾上腺髓质素、降钙素受体样受体、受体活性修饰蛋白2和3的mRNA水平分别增加78％、93.7％、91.8％和109.5％(P均<0.01)。肾上腺髓质素与降钙素受体样受体、受体活性修饰蛋白2和3的mRNA水平呈正相关,其相关系数分别为0.83、0.92和0.93(P均<0.01)。结果提示,血管平滑肌细胞肾上腺髓质素 旁/自分泌功能改变可能参与血管钙化的调节过程。
To investigate the changes of adrenomedullin production in calcified vascular smooth muscle cells and the expression of calcitonin receptor-like receptor and receptor activity-modifying protein in adrenomedullin receptor system and its pathological significance. Β-glycerophosphate was used to induce the calcification of cultured rat vascular smooth muscle cells; radioimmunoassay was used to measure the amount of adrenomedullin secreted by vascular smooth muscle cells; semi-quantitative reverse transcriptase-polymerase chain reaction was used to measure the expression of adrenomedullin, The mRNA level of the somatostatin receptor and receptor activity-modifying protein was determined. The calcium content of cells was determined by atomic absorption spectrophotometer. The alkaline phosphatase activity was measured by alkaline phosphatase kit. The radioactivity of 45Ca2 + was measured by the liquid scintillation counter. Compared with non-calcified vascular smooth muscle cells, calcium content, 45Ca2 + uptake and alkaline phosphatase activity in calcified vascular smooth muscle cells were increased by 118%, 174% and 7 times (P <0.01), respectively. Calcified cells adrenomedullin (P <0.01). The mRNA levels of adrenomedullin, calcitonin receptor-like receptor and receptor activity-modifying protein 2 and 3 increased by 78%, 93.7%, 91.8% and 109.5% P <0.01). Adrenomedullin was positively correlated with calcitonin receptor-like receptors and receptor activity-modifying proteins 2 and 3, with correlation coefficients of 0.83, 0.92 and 0.93, respectively (all P <0.01). The results suggest that changes in adrenomedullin / autocrine function of vascular smooth muscle cells may be involved in the regulation of vascular calcification.