摘要
目的建立测定肾移植术后患者外周血淋巴细胞中次黄嘌呤核苷酸脱氢酶(IMPDH)Ⅰ型和Ⅱ型基因表达水平的Real-time PCR方法,研究吗替麦考酚酯(MMF)药效个体差异。方法取肾移植后门诊复查患者外周血分离淋巴细胞,提取总RNA,进行Real-time PCR反应,选用β-actin作为内参,靶基因的相对定量按公式得出:靶基因=2-(△△Ct),高效液相色谱法测定吗替麦考酚酸(MPA)浓度。结果IMPDH表达个体间差异大(Ⅰ型0.118～3.313;Ⅱ型0.185～2.928),MPA对于个体IMPDH两种亚型的抑制程度不同,MPA的谷浓度>1.0mg·L-1时,IMPDHⅠ型和Ⅱ型的基因表达水平表现出明显的抑制作用。结论本法实用性强,结果可靠、重现性好,可用于肾移植患者服用MMF后的药效个体化研究。
OBJECTIVE To develop a real-time PCR method for the determination of IMP dehydrogenase （IMPDH） isoforms Ⅰ and Ⅱ gene expression level in order to evaluate the mycophenolate mofetil（MMF） therapeutic effect in renal transplant recipients. METHODS The leukomonocytes were separated from renal transplant recipients peripheral blood. RNA was isolated before RT-PCR reaction. IMPDH Ⅰ and Ⅱ gene expression was normalized using β-actin. Mocophenolic acid（MPA） trough concentration（C0） was determined by HPLC. RESULTS A wide interpatient variability of the gene expression （IMPDH Ⅰ ： 0.118-3.313; IMPDH Ⅱ： 0.185-2.928） was observed. The level of MPA inhibition was significantly different between the two isoforms. MPA showed the strong inhibition on IMPDH gene expression when its concentration reached 1.0 mg.L^-1. CONCLUSION Real-time PCR method to deterimine IMPDH gene expression level is convenient, reliable and reproducible, and suitable for monitoring therapeutic effect of mycophenolate mofetil in renal transplantation patients.
出处
《中国药学杂志》
CAS
CSCD
北大核心
2009年第9期649-653,共5页
Chinese Pharmaceutical Journal
