RECK基因在胰腺癌中的表达及其与预后的关系

Expression of RECK in pancreatic carcinoma and its relationship with prognosis

目的观察人胰腺癌组织及细胞株中RECK基因的表达，探讨重组慢病毒LV-RECK对胰腺癌动物模型的治疗效果以及该基因与胰腺癌预后的关系。方法免疫组化法检测42例胰腺癌及相应正常胰腺组织中RECK的表达，分析RECK表达与胰腺癌临床病理特征及预后的关系。采用Western Blotting法检测三个胰腺癌细胞株（PANC-1，MIAPa Ca-2，AsPC1）中RECK的表达。统计分析RECK基因表达与临床病理特征及预后的关系。建立胰腺癌动物模型，应用LV—RECK进行治疗，观察抑瘤效果，并进行生存分析。结果RECK在胰腺癌细胞株中不表达。胰腺癌组织的RECK表达率（45．2％）较正常胰腺组织（88．1％）明显降低（P〈0．01）。RECK表达与胰腺癌的TNM分期、淋巴结转移、局部浸润存在相关性（P〈0．05）。Kaplan-Meier生存分析显示，RECK阳性表达组患者的生存期较阴性组明显延长（P=0．000）。单因素Cox分析显示，RECK表达、TNM分期、淋巴结转移及局部浸润与预后有关（P〈0．05）。多因素Cox分析显示，只有RECK表达具有独立的预后意义（P=0．000）。动物实验中，LVRECK治疗组移植瘤明显缩小（P〈0．05），微血管密度明显降低（P〈0．05），凋亡指数显著升高（P〈O．05），生存期显著延长（P〈0．05）。结论RECK表达与胰腺癌的侵袭转移及预后密切相关，可作为胰腺癌的独立预后指标。RECK基因过表达可以抑制肿瘤血管新生，诱导肿瘤细胞凋亡，从而抑制移植瘤的生长，改善荷瘤鼠的预后，为胰腺癌治疗提供了一个新的途径。

Objective To study the expression of reversion-inducing-cysteine-rich protein with Kazal motifs （RECK） in human pancreatic carcinoma tissues and pancreatic carcinoma cell lines; the effects of recombinant lentiviruses carrying RECK gene（LV-RECK） therapy on human pancreatic car cinoma xenograft in nude mice; and to find out the relationship between the expression of RECK and the prognosis of pancreatic carcinoma. Methods Immunohistochemical method was used to detect the expression of RECK in the resected specimens of pancreatic carcinoma and their corresponding normal pancreatic tissues in 42 patients. Western blotting was used to examine the expression of RECK in human pancreatic carcinoma cell lines （PANC-1, MIAPaCa-2, AsPC-1）. Statistical analyses were performed to determine the relationship between RECK expression and the clinicopathological characteristics and prognosis in pancreatic carcinoma. Subcutaneous xenograft tumor models of human pancreatic carcinoma were established in nude mice. These nude mice were then divided into the experimental group, the negative control group and the blank control group randomly. The three groups of nude mice were intratumorally injected with LV-RECK, LV-EGFP and normal saline （NS） respectively. The antitumor effect was studied. Immunohistocbemical method was used to detect the expression of RECK and microvessel density （MVD）. Terminal deoxynucleotidyl transferase mediated dUTP-DIG nick end labeling （TUNEL） was used to detect the apoptosis of tumor cells. Survival analysis was performed. Results All three pancreatic carcinoma cell lines did not express RECK. The overall positive rate of RECK expression was 45.2% （19/42） in pancreatic carcinoma, and 88.1% （37/42） in normal pancreatic tissue. The expression level of RECK was significantly lower in the tumor tissues than inthe normal tissues （P〈0.01）. The expression of RECK was significantly associated with TNM stage, lymph node metastasis and local infiltration of pancreatic carcinoma （P〈0.05）. Kap]an Meier survival analysis revealed that the survival time was significantly longer in the RECK positive patient group than in the RECK negative patient group. Univariate Cox regression analysis revealed that RECK expression, TNM stage, lymph node metastasis and local infihration were significantly related with prognosis for pancreatic carcinoma （P〈0.05）. Multivariate Cox regression analysis revealed that only RECK expression remained as an independent significant factor in predicting the prognosis of pan- creatic carcinoma （P〈0.001）. When compared with the negative control and the blank control groups, the volume of subcutaneous xenograft tumor in the experimental group was significantly de creased （P〈0.05）. RECK protein in the experimental group was re-expressed. MVD of the expert mental group was significantly less than those of the control groups （P〈0.05）. Apoptoticindex （AI） of the experimental group was significantly higher than those of the control groups （P〈0.05）. The survival time of nude mice in the experimental group was significantly longer than those in the control groups （P〈0.05）. Conclusions RECK expression was closely related to invasion, metastasis and prognosis of pancreatic carcinoma and it was an independent prognostic marker. RECK gene over-ex pression inhibited neovascularization of pancreatic carcinoma, induced apoptosis of tumor cells, inhibi ted the growth of tumor xenograft and improved the prognosis of tumor bearing mice. These results suggest a possible new treatment for pancreatic carcinoma.