G蛋白耦联受体49在胰腺癌组织中的表达及其临床意义

Expression and its clinical significance of G-protein coupled receptor 49 in pancreatic carcinoma

目的探讨G蛋白耦联受体49（GPR49）在胰腺癌组织中表达的临床意义及其生物学作用。方法采用组织芯片和免疫组织化学染色技术比较77例胰腺癌患者癌组织和癌旁组织中GPR49的表达，并分析其在不同病理分级和临床分期患者癌组织中的表达水平。以GPR49阳性胰腺癌细胞株CFPAC-1为细胞模型，GPR49的配体R-脊椎蛋白1进行体外刺激，采用细胞计数方法分析其对CFPAC-1细胞的促增殖作用，并采用流式细胞术分析R-脊椎蛋白1对CFPAC-1细胞表达细胞膜CD44水平的影响。将R-脊椎蛋白1孵育后的CFPAD1细胞皮下接种于裸鼠，观察小鼠成瘤时间和肿瘤大小。统计学分析分别采用t检验和单因素方差分析。结果77份胰腺癌组织中普遍表达GPR49。胰腺癌组织中GPR49的免疫组织化学染色综合评分为（9．0±2．4）分，高于癌旁组织的（5．7±2．4）分，差异有统计学意义（t=8．995，P〈0．01）。GPR49的表达与患者肿瘤大小、病理分级、淋巴结转移、临床分期等无明显相关性（P均〉0．05）。体外实验表明，R-脊椎蛋白1可促进CFPAC-1细胞增殖，上调CFPAC-1表达CD44。体内实验表明，R-脊椎蛋白1刺激组小鼠在接种CFPAC-1细胞后30d的肿瘤体积为（606．0±188．0）mm3，大于PBS刺激组的（364．2±83．7）mm3，差异有统计学意义（t=2．616，P=0．031）。结论GPR49普遍表达于胰腺癌细胞，R-脊椎蛋白1／GPR49途径对胰腺癌细胞具有促增殖作用，提示其可能是干预胰腺癌生长的潜在靶点。

Objective To investigate the clinical significance and biological role of G-protein coupled receptor 49 （GPR49） expression in pancreatic carcinoma. Methods GPR49 expression in tumor and adjacent normal tissues of 77 patients with pancreatic cancer was compared by tissue microarray and immunohistochemistry. And then, the GPR49 expression levels in the tumor tissues of patients with different pathological grades and clinical stages were analyzed. GPR49 positive pancreatic cancer cell line CFPAC-1 was taken as cellular model. CFPAC-1 ceils were stimulated with roof plate-specific spondin （RSPO）I, the ligand of GPR49, in vitro. The effect of RSPO1 on CFPAC-1 cells proliferation was evaluated with cell counting. The effect of RSPO1 on the expression of membrane molecular CD44 in CFPAC-1 cells was detected by flow cytometry. CFPAC-1 cells incubated with RSPO1 were subcutaneously implanted into nude mice. And then, the time of tumor formation and tumor size were observed. T test and single factor analysis of variance were performed for statistical analysis. Results GPR49 was widely expressed in all 77 pancreatic cancer tissues. By immunohistochemistry, the score of GPR49 expression in pancreatic cancer tissues was 9.0±2.4, which was higher than that of adjacent normal tissues （5.7±2.4）, and the difference was statistically significant （t= 8. 995, P〈0.01）. There was no correlation between GPR49expression and tumor sizes, pathological grades, lymph node metastasis, and clinical stages （all P 〉0.05）. The results of experiments in vitro indicated that RSPO1 could promote CFPAC-1 cells proliferation and up-regulate CD44 expression in CFPAC-1 cells. Experiments in vivo demonstrated that after 30 days the tumor volume of mice implanted with RSPOl-pretreated CFPAC-1 cells was （606.0 ± 188.0） mm3, which was larger than that of PBS-pretreated group （（364. 2 ± 83. 7） mm3）, and the difference was statistically significant （t=2. 616, P〈0. 031）. Conclusion GPR49 is widely expressed in pancreatic cancer cells and RSPO1/GPR49 pathway has play a role in promoting the proliferation of pancreatic cancer cells, which might be a potential target for interfering pancreatic cancer.