Wang Shu-guang, Chen Chang-hong, Li Da-jiang, Li Kun
Institute of Hepatobiliary Surgery, People's Liberation Army Southwest Hospital, Third Military Medical University, Chongqing 400038, China.
Zhonghua Yi Xue Za Zhi. 2004 Oct 2;84(19):1642-4.
To investigate the effect of antisense oligonucleotide (AS-OD) of laminin receptor (LNR) on urokinase-type plasminogen activator (u-PA) expression in human bile duct carcinoma cells.
Bile duct carcinoma cells of QBC939 line were cultured. Compounds of lipofectAMINE and AS-OD phosphorothioate of LNR of the concentrations of 3, 6, and 12 micromol/L respectively were prepared and then transfected into the QBC939 cells. Sense oligodeoxynucleotide (S-OD) of LNR and blank vector were used as controls. After culture of 48 hours, flow cytometry was used to detect the protein expression of LNR and the u-PA mRNA expression was detected by reverse-transcriptase polymerase chain reaction (RT-PCR). Cell invasion was examined by culture plate with Transwell microporous film before and after transfection.
Flow cytometry showed down-regulation of LNR protein expression in the cells transfected with AS-OD dose-dependently. The LNR protein expression of the cells transfected with AS-OD of the concentration of 6 micromol/L was down-regulated by over 25% in comparison with those of the blank vector group and S-OD group (both P < 0.05). RT-PCR showed that the u-PA mRNA expression of the AS-OD 6 micromol/L group was down-regulated by 30% in comparison with the 2 control groups. The number of migrating cells in the AS-OD group was 78 +/- 6, significantly lower than those in the S-OD group (105 +/- 11), and blank vector group (101 +/- 7), both P < 0.05.
The expression of laminin receptor in the bile duct carcinoma cells is related with the expression of u-PA gene expression. Inhibition of LNR expression decreases the u-PA gene expression, suggesting that LNR plays an important role in the invasion and metastasis of bile duct carcinoma.
探讨层粘连蛋白受体（LNR）反义寡核苷酸（AS-OD）对人胆管癌细胞尿激酶型纤溶酶原激活剂（u-PA）表达的影响。
培养QBC939系胆管癌细胞。分别配制浓度为3、6和12 μmol/L的脂质体转染试剂与LNR硫代磷酸酯反义寡核苷酸的复合物，然后转染至QBC939细胞。以LNR正义寡脱氧核苷酸（S-OD）和空载体作为对照。培养48小时后，采用流式细胞术检测LNR蛋白表达，采用逆转录聚合酶链反应（RT-PCR）检测u-PA mRNA表达。转染前后通过Transwell微孔膜培养板检测细胞侵袭能力。
流式细胞术显示，转染AS-OD的细胞中LNR蛋白表达呈剂量依赖性下调。与空载体组和S-OD组相比，转染6 μmol/L AS-OD的细胞LNR蛋白表达下调超过25%（均P < 0.05）。RT-PCR显示，6 μmol/L AS-OD组的u-PA mRNA表达较2个对照组下调30%。AS-OD组迁移细胞数为78±6，显著低于S-OD组（105±11）和空载体组（101±7），均P < 0.05。
胆管癌细胞中层粘连蛋白受体的表达与u-PA基因表达有关。抑制LNR表达可降低u-PA基因表达，提示LNR在胆管癌的侵袭和转移中起重要作用。
