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目的观察肾钙敏感受体(CaSR)在特发性高钙尿症(IH)发病中的作用。方法雄性遗传性高钙尿结石(GHS)大鼠和正常野生型SD大鼠各6只,相同的饮食和饲养条件下,检测各组大鼠血Ca~(2+)、P5~+、1,25(OH)_2D_3和连续2个24 h尿Ca~(2+)的水平,使用半定量逆转录-聚合酶链反应(RT-PCR)检测肾组织钙敏感受体mRNA表达,PCR产物纯化后进行DNA直接测序。结果两组大鼠血Ca~(2+)、P5~+、1,25-(OH)_2D_3水平相似,均在正常范围内,而GHS组平均24 h尿Ca~(2+)排泄量为正常对照(NC)组的3.36倍[分别为(2.66±0.64)、(0.79±0.06)mg/d],两组比较差异有统计学意义(t=5.82,P<0.01);GHS组CaSR mRNA平均相对吸光度(A)值为1.354±0.108,NC组为0.602±0.218,差异有统计学意义(t=6.19,P<0.01);CaSR基因第7外显子区域cDNA测序结果与Genebank(Accession:NM-016996)公布的碱基序列比较,GHS组有4只大鼠存在第3072位碱基G/A替换,使第1024位谷氨酸(Glu)变为赖氨酸(Lys),其余2只和NC组均与Genebank公布的碱基序列一致。结论肾组织CaSR mRNA高表达及其第7外显子Glu1024Lys替换可能是特发性高钙尿症发病的分子机制之一。
Objective To investigate the role of renal calcium-sensitive receptor (CaSR) in the pathogenesis of idiopathic hypercalciuria (IH). Methods Six male Wistar rats with high calcium calculus (GHS) and 6 normal wild-type (SD) rats were used to detect the changes of blood Ca 2+, P 5+, and 1 in rats under the same diet and feeding conditions , 25 (OH) _2D_3 and two 24 h urinary Ca 2+ levels. The expression of calcium sensitive receptor mRNA in renal tissue was detected by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) After direct DNA sequencing. Results The levels of Ca ~ (2 +), P5 ~ +, 1,25- (OH) _2D_3 in the two groups were all within the normal range, while the average 24 h urinary Ca ~ (2 +) excretion in the GHS group was (2.66 ± 0.64) and (0.79 ± 0.06) mg / d respectively in the NC group (t = 5.82, P <0.01). In the GHS group, the expression of CaSR mRNA The average relative absorbance (A) was 1.354 ± 0.108 in NC group and 0.602 ± 0.218 in NC group (t = 6.19, P <0.01). The results of cDNA sequencing of the exon 7 of CaSR gene and Genebank (Accession: NM-016996), there were 4 bases in GHS group with the base G / A substitution at position 3072, the 1024th glutamic acid (Glu) was changed to lysine (Lys), and the remaining 2 Only the NC group and Genebank published base sequence. Conclusion The high expression of CaSR mRNA and the substitution of Glu1024Lys in exon 7 may be one of the molecular mechanisms of idiopathic hypercalciuria.