继发性右室肥厚患者钙调蛋白基因表达的研究

STUDY ON EXPRESSION OF CALMODULIN GENE IN THE PATIENTS WITH SECONDARY RIGHT VENTRICULAR HYPERTROPHY

目的 :探讨继发性右室肥厚 (RVH)患者心肌细胞内Ca2 + 超负荷及钙调蛋白mRNA和蛋白质表达的变化。方法 :采用三电极直流等离子体原子发射直读光电光谱法测定心肌细胞内Ca2 + 含量 ;采用RT -PCR和Westernblot法测定RVH患者和正常对照钙调蛋白mRNA和蛋白质表达的变化。结果 :与正常对照组相比 ,RVH患者心肌细胞内Ca2 + 含量提高 3倍以上〔(10 2 5 6 7± 4 14 71) μg/mlvs (2 70 86±10 9.2 4 ) μg/ml,P <0 .0 1〕 ;肌浆网 (SR)Ca2 + -ATPase和细胞膜L型Ca2 + 通道mRNA水平明显减低 ,分别减低 39 4 5%、33 33% (0 6 6± 0 2 8vs 1.0 9± 0 .11;0 .72± 0 .2 1vs 1.0 8± 0 .14,P均 <0 0 5) ,SR磷酸受纳蛋白和兰尼碱受体、肌集钙蛋白的mRNA转录差异无显著性 (P >0 0 5) ;Ca2 + -ATPase的蛋白质表达明显低于对照组 (0 .78± 0 .0 9vs 1.0 3± 0 .0 3,P <0 .0 5) ;磷酸受纳蛋白的相对含量差异无显著性 (P >0 .0 5)。结论 :RVH患者SRCa2 + -ATPasemRNA和蛋白质表达减低及细胞膜L型Ca2 + 通道mRNA转录减低是导致心肌细胞内Ca2 + 超负荷和RVH发生的主要分子生物学机制。

Objective:To elucidate molecular mechanisms of the changes of calmodulin gene expression when Ca 2+ is overload in cardic muscle cell of patients with secondary right ventricular hypertrophy(RVH).Methods:Ten patients with secondary RVH and 6 control subjects were randomly selected.The content of calcium was assayed by spectrocomparator.The mRNA amounts of these calmodulin genes including L type calcium channel,sarcoplasmic reticular(SR) Ca 2+ ATPase,ryanodine receptor,calsequestrin and phospholamban were detected by reverse transcription polymerase chain reaction(RT PCR) and normalized to the mRNA levels of actin.The protein levels of SR Ca 2+ ATPase and phospholamban were analyzed by Western blot analysis.Results:The content of calcium increased significantly in patients with RVH.RT PCR analyses showed that the steady state level of mRNA encoding the SR Ca 2+ ATPase and cardiac L type calcium channel decreased significantly in patients with RVH(0.66±0.28 vs 1.09±0.11,P<0.05,0.72±0.21 vs 1.08±0.14,P<0.05).In contrast,no alterations at the mRNA level for SR phospholamban,ryanodine receptor,calsequestrin were observed in patients with RVH compared with those in control subjects.Protein level of SR Ca 2+ ATPase was significantly reduced in patients with RVH(0.78±0.09 vs 1.03±0.03,P<0.05),whereas protein level of phospholamban was unchanged.Conclusion:The expressions of L type calcium channel and SR Ca 2+ ATPase were down regulated in patients with RVH.These changes may be a contributory factor for secondary RVH.