NHE-1核酶基因转染对大鼠肺动脉平滑肌细胞凋亡的影响

目的 探讨NHE-1核酶基因转染对大鼠肺动脉平滑肌细胞（PASMC）凋亡的影响。方法 构建NHE-1核酶基因逆转病毒载体，转染体外培养的大鼠PASMC，应用半定量RT-PCR法（sqRT-PCR）测定细胞内NHE-1 mRNA的表达、BCECF法测定pHi、流式细胞仪测细胞周期、电镜及原位细胞凋亡检测（TUNEL法）观察细胞凋亡情况。结果 转染NHE-1核酶基因的细胞，NHE-1 mRNA表达量及pHi较空载体转染细胞及未转染细胞显著降低，细胞凋亡率显著增加，电镜示细胞凋亡，凋亡小体形成。结论 NHE-1核酶基因转染可通过NHE-1抑制、细胞内酸化，显著诱导肺动脉平滑肌细胞凋亡。

Effects of transfected NHE-1 ribozyme gene on apoptosis of pulmonary artery smooth muscle cells of rats in vitro

Objective To explore the effects of transfected NHE 1 ribozyme gene on apoptosis of pulmonary artery smooth muscle cells(PASMC) in vitro . Methods NHE 1 ribozyme gene was designed, synthesized and cloned into plasmid PLXSN, the recombined plasmid was tansfered into cultured PASMC. Expression of NHE 1 mRNA was detected by sqRT PCR. Intracellular pH(pHi) was measured by fluorescence dye BCECF AM. Cell cycle was measured by flow cytometric DNA analysis. Cell apoptosis was observed with electron microscopy and terminal deoxynucleotidyl transferase medited dUTP nick end labeling(TUNEL) respectively. Results The NHE 1 mRNA expression and pHi of cells transfected with NHE 1 ribozyme gene decreased significantly than those of cells transfected with PLXSN and nontransfected control. Meanwhile, the apoptosis rate of cells transfected with NHE 1 ribozyme gene increased significantly. Electron microscopy showed apoptosis of cells transfected with NHE 1 ribozyme gene. Conclusion Transfected NHE 1 ribozyme gene can induce apoptosis of pulmonary artery smooth muscle cell by inhibiting NHE 1 and intracellular acidification.