Objective: To investigate the relationship between the expression of integrin α 6 (ITGA6), miR-4484 and the pathologic stage of gastric cancer. Methods: Gastric cancer tissues and normal gastric mucosa tissues adjacent to cancer (>5 cm from tumor margin) of 30 patients with primary gastric cancer who underwent direct surgical resection without adjuvant therapy from June to September 2017 in West China Hospital of Sichuan University were selected. Real-time quantitative polymerase chain reaction (PCR) was used to detect the expression levels of miR-4484 and ITGA6, western blot was used to detect the expression level of ITGA6 protein, dual luciferase reporter gene was used to verify the relationship between ITGA6 and miR-4484. Spearman's correlation analysis was used to determine the relationship between miR-4484 and ITGA6 expression levels in gastric cancer tissues. Results: The expression level of ITGΑ6 in gastric cancer (32.30±13.47) was higher than that in matched normal gastric tissues (24.55±10.25, P=0.015), the area under the receiver operating characteristic (ROC) curve was 0.660 and the diagnostic sensitivity and specificity were 43.3% and 96.7%, respectively. The expression level of miR-4484 in gastric cancer (4.11±2.87) was lower than that of matched normal gastric tissues (5.75±2.80, P=0.029), the area under the ROC curve was 0.690 and the diagnostic sensitivity and specificity were 30.0% and 86.7%, respectively. The expression level of miR-4484 was negatively correlated with ITGA6 in gastric cancer tissues (r=-0.621, P<0.001). The expression level of ITGA6 protein in gastric cancer tissues (0.65±0.19) was higher than that in normal adjacent tissues (0.26±0.12, P<0.001). Compared with ITGA6 3'UTR wild-type+ miR-NC group, ITGA6 3'UTR wild-type+ miRNA mimics group had lower luciferase activity (50.69±5.10, 34.00±1.19, P<0.001), while the luciferase activity of ITGA6 3'UTR wild-type+ ASO miR-4484 group was higher than that of ITGA6 3'UTR wild-type+ miR-NC group (82.44±6.37, 50.69±5.10, P<0.001), indicated that ITGA6 was the direct target gene of miR-4484. The expression levels of miR-4484 in T1, T2, T3 and T4 (4a and 4b) gastric cancer tissues were 9.98±2.24, 5.28±2.03, 2.92±2.04 and 4.11±2.87, respectively, with statistical significance (P<0.001). The expression levels of ITGA6 in N0, N1, N2 and N3 gastric cancer tissues were 29.55±8.32, 21.71±3.75, 24.60±8.79 and 40.69±15.83, respectively, with statistical significance (P=0.022). The expression levels of miR-4484 in N0, N1, N2 and N3 gastric cancer tissues were 5.01±3.52, 5.48±2.76, 5.88±1.83 and 2.30±1.56, respectively, with statistical significance (P=0.032). The expression levels of ITGA6 in M0 and M1 gastric cancer tissues were 26.28±7.66 and 52.08±8.12, respectively, with statistical significance (P<0.001). The expression levels of miR-4484 in M0 and M1 gastric cancer tissues were 4.95±2.74 and 1.34±0.80, respectively, with statistical significance (P<0.001). Conclusions: ITGA6 is upregulated in gastric cancer tissues, while miR-4484 is downregulated in the gastric cancer group, and its expression level is related to the clinicopathological features of gastric cancer. ITGA6 is the direct target gene of miR-4484, implicates that miR-4484 may inhibit the invasion and metastasis of gastric cancer by regulating the expression of ITGA6. Both miR-4484 and ITGA6 may be the new prognostic markers and potential therapeutic targets of gastric cancer.
目的： 探讨整合素α6(ITGA6)蛋白、基因和miR-4484与胃癌病理分期的关系。 方法： 收集2017年6—9月于四川大学华西医院行手术治疗(未经辅助治疗)的30例原发性胃癌患者的胃癌及癌旁(距离瘤体>5 cm)正常胃黏膜组织。采用实时荧光定量PCR检测miR-4484和ITGΑ6的表达水平，采用Western blot法检测ITGA6蛋白的表达水平，采用双荧光素酶报告基因验证ITGΑ6与miR-4484的关系，采用Spearman相关分析明确miR-4484与ITGA6在胃癌组织中表达水平的相关性。 结果： 胃癌组织中ITGA6的表达水平(32.30±13.47)高于正常癌旁组织(24.55±10.25，P＝0.015)，受试者工作特征(ROC)曲线下面积为0.660，其诊断敏感度为43.3%，特异度为96.7%。胃癌组织中miR-4484的表达水平(4.11±2.87)低于正常癌旁组织(5.75±2.80，P＝0.029)，ROC曲线下面积为0.690，其敏感度为30.0%，特异度为86.7%。胃癌组织中miR-4484表达水平与ITGA6呈负相关(r＝-0.621，P<0.001)。ITGA6在胃癌组织中的表达水平(0.65±0.19)高于正常癌旁组织(0.26±0.12，P<0.001)。与ITGΑ6 3′UTR野生型+miR-NC组比较，ITGΑ6 3′UTR野生型+miRNA mimics组细胞荧光素酶活性降低(分别为50.69±5.10和34.00±1.19，P<0.001)，ITGΑ6 3′UTR野生型+ASO miR-4484组细胞荧光素酶活性高于ITGΑ6 3′UTR野生型+miR-NC组(分别为82.44±6.37和50.69±5.10，P<0.001)，ITGA6为miR-4484的直接靶基因。T1、T2、T3、T4(4a和4b)期胃癌组织中miR-4484的表达水平分别为9.98±2.24、5.28±2.03、2.92±2.04和4.11±2.87，差异有统计学意义(P<0.001)。N0、N1、N2、N3期胃癌组织中ITGA6的表达水平分别为29.55±8.32、21.71±3.75、24.60±8.79和40.69±15.83，差异有统计学意义(P＝0.022)。N0、N1、N2、N3期胃癌组织中miR-4484的表达水平分别为5.01±3.52、5.48±2.76、5.88±1.83和2.30±1.56，差异有统计学意义(P＝0.032)。M0、M1期胃癌组织中ITGA6的表达水平分别为26.28±7.66和52.08±8.12，差异有统计学意义(P<0.001)。M0、M1期胃癌组织中miR-4484的表达水平分别为4.95±2.74和1.34±0.80，差异有统计学意义(P<0.001)。 结论： ITGΑ6在胃癌组织中呈高表达，miR-4484在胃癌组织中呈低表达，且其表达水平与胃癌的临床病理特征有关。ITGΑ6为miR-4484的直接靶基因，miR-4484可能通过调控ITGΑ6发挥抑制胃癌侵袭转移的作用，miR-4484和ITGΑ6可能作为新的胃癌预后标志物及潜在的治疗靶点。.
Keywords: Gastric neoplasms; Integrin; Invasion and metastasis; microRNA.