CAJ | 학술논문

目的用蛋白质组学方法分析去甲肾上腺素(NE)刺激人肺动脉阻力血管平滑肌细胞(PASMC)增殖前后细胞蛋白表达谱的变化。方法通过组织贴块法建立人PASMC细胞株；细胞经同步化后，用10-5 mol/L NE处理细胞24h，对照组未作处理，采用双向凝胶电泳、质谱和生物信息学蛋白质组方法比较NE处理前后细胞蛋白表达谱的变化，并用质谱分析差异在5倍以上的蛋白质分子；进一步使用实时RT-PCR和Western blot法验证差异蛋白的表达变化。结果原代培养的人PASMC细胞纯度＞99％;PASMC经NE刺激后，多种细胞蛋白表达发生改变，其功能涉及细胞骨架相关蛋白、代谢及信号转导等多方面；实时RT-PCR和Western blot结果显示，NE刺激后在细胞内转录和翻译水平，α-烯醇化酶基因表达上调，与二维电泳结果一致。结论NE刺激人PASMC后，可引起细胞内多种不同功能的蛋白表达发生改变，并部分通过α-烯醇化酶基因调控细胞生物学功能。
목적용단백질조학방법분석거갑신상선소(NE)자격인폐동맥조력혈관평활기세포(PASMC)증식전후세포단백표체보적변화。방법통과조직첩괴법건립인PASMC세포주；세포경동보화후，용10-5 mol/L NE처리세포24h，대조조미작처리，채용쌍향응효전영、질보화생물신식학단백질조방법비교NE처리전후세포단백표체보적변화，병용질보분석차이재5배이상적단백질분자；진일보사용실시RT-PCR화Western blot법험증차이단백적표체변화。결과 원대배양적인PASMC세포순도＞99％;PASMC경NE자격후，다충세포단백표체발생개변，기공능섭급세포골가상관단백、대사급신호전도등다방면；실시RT-PCR화Western blot결과현시，NE자격후재세포내전록화번역수평，α-희순화매기인표체상조，여이유전영결과일치。결론NE자격인PASMC후，가인기세포내다충불동공능적단백표체발생개변，병부분통과α-희순화매기인조공세포생물학공능。
ObjectiveTo observe the effect of noradrenalin(NE) on human pulmonary arterial smooth muscle cells (PASMC) by using a proteomic approach.Methods Human PASMC were cultured primarily in vitro.Experiments were performed in the 3rd to 5th passages of the cells.The human PASMC were cultured in serum-free medium for 24 h prior to treatment with either NE (10-5 mol/L, the test group)or completed-serum culture medium (the control group) for 24 h. And then analysis via 2-DE gel electrophoresis and MALDI-TOF-MS was performed to display the different protein profiles of whole cell protein from cultures of the control and the NE-treatment group. Real-time RT-PCR and Western blot analysis were used to confirm the proteomic analysis.Results The purity of the primary culture cells was about 99％.When the human PASMC were treated by NE, the expression of different groups of cellular proteins was changed, including cell cytoskeleton-associated proteins, cell signal-associated proteins, and glycolytic and metabolism-associated proteins. The results were confirmed using real-time RT-PCR and Western blot. NE enhanced the proliferation of human PASMC partly by affecting the expression of αenolase.Conclusion The data suggest that a wide range of signaling pathways may be involved in NEinduced proliferation of human PASMC, and α-enolase associated pathway may be an important one.