降钙素基因相关肽受体成分蛋白在肝细胞癌中的表达及其与预后的关系

孙冠群; 周思蕾; 曾榃伦; 刘君宇; 梁喜俊; 程卓

doi:10.3969/j.issn.1001-5256.2022.06.021

降钙素基因相关肽受体成分蛋白在肝细胞癌中的表达及其与预后的关系

DOI: 10.3969/j.issn.1001-5256.2022.06.021

1.
海军军医大学转化医学研究中心临床肿瘤研究所，上海 200433
2.
海军军医大学附属东方肝胆外科医院肿瘤科，上海 200433

基金项目:

国家自然科学基金 (81801570);

上海市卫健委青年基金 (20194Y0269);

上海市科学技术委员会扬帆计划 (20YF1459600)

伦理学声明：本研究于2020年6月15日经由海军军医大学附属东方肝胆外科医院医学研究伦理委员会批准，所纳入患者均签署知情同意书。

利益冲突声明：本研究不存在研究者、伦理委员会成员、受试者监护人以及与公开研究成果有关的利益冲突。

作者贡献声明：孙冠群和周思蕾负责文章撰写及相关实验数据的获取；曾榃伦、刘君宇负责数据分析；梁喜俊负责文章撰写、修改指导；程卓负责关键性实验设计、文章撰写、修改指导及审定。

详细信息

通信作者:
程卓，chengzhuo1219@163.com

孙冠群、周思蕾对本文贡献等同，同为第一作者

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出版历程
- 收稿日期: 2021-11-01
- 录用日期: 2022-01-27
- 出版日期: 2022-06-20

Expression of calcitonin gene-related peptide-receptor component protein in hepatocellular carcinoma and its association with prognosis

1.
Clinical Cancer Institute, Center for Translational Medicine, Naval Medical University, Shanghai 200433, China
2.
Department of Oncology, Eastern Hepatobiliary Surgery Hospital, Naval Medical University, Shanghai 200433, China

Research funding:

National Natural Science Foundation of China (81801570);

Youth Fund Project of Shanghai Municipal Health Commission (20194Y0269);

Shanghai Sailing Project from the Science and Technology Commission of Shanghai Municipality (20YF1459600)

More Information

Corresponding author: CHENG Zhuo, chengzhuo1219@163.com (ORCID:0000-0002-2967-3167)

摘要

摘要: 目的检测降钙素基因相关肽受体成分蛋白(CRCP)在肝细胞癌及癌旁组织中的表达水平，探究其与肝细胞癌临床病理和患者预后的相关性。方法收集2003年6月—2009年9月在海军军医大学附属东方肝胆外科医院行手术切除治疗的79例肝细胞癌患者的肝癌组织及相应癌旁组织标本，制作组织芯片，免疫组化定量分析，提取蛋白进行Western Blot检测，比较CRCP在肝癌组织与癌旁组织中的表达差异。计数资料组间比较采用χ²检验，采用受试者工作特性曲线(ROC曲线)分析获得曲线下面积(AUC)，评估拟合优度，并使用Youden指数确定最佳截断值，利用Kaplan-Meier生存分析法分析CRCP表达与肝细胞癌预后的关系，两组间比较采用log-rank检验。结果 79例肝细胞癌患者中，男67例，女12例，年龄10~72岁；病理分级4级1例，3级61例，2级17例；20例合并门静脉癌栓；巴塞罗那分期0期5例，A期55例，B期11例，C期8例。免疫组化结果显示，75.9%的患者肝癌组织中CRCP表达水平明显低于癌旁组织。CRCP低表达肝细胞癌患者的CK19阳性、肿瘤包膜不完整、合并门静脉癌栓和肿瘤病理分级高的患者所占比例更高(χ²值分别为6.410、4.829、9.319、9.083，P值均＜0.05)。与CRCP低表达组相比，CRCP高表达组患者的总体生存期更长，复发率更低(P值分别为＜0.001、0.009)。另取6例患者的癌组织与癌旁组织进行Western blot检测，结果显示，4例患者肝癌组织中CRCP表达水平低于癌旁组织。结论肝癌组织中CRCP低表达相较于高表达患者预后更差，CRCP可能参与肝细胞癌的发生发展与转移过程，提示该分子或可作为肝细胞癌患者预后判定的生物标志物。
- 癌, 肝细胞 /
- 受体, 降钙素基因相关肽 /
- 肿瘤转移 /
- 预后
Abstract: Objective To investigate the expression level of calcitonin gene-related peptide-receptor component protein (CRCP) in hepatocellular carcinoma (HCC) tissue and adjacent tissue and its association with the clinicopathological features and prognosis of patients. Methods HCC and adjacent tissue samples were collected from 79 HCC patients who underwent surgical resection in Eastern Hepatobiliary Surgery Hospital, Navy Medical University, from June 2003 to September 2009. Tissue microarray was prepared, and immunohistochemistry was used for quantitative analysis. Related proteins were extracted and measured by Western blot, and the expression of CRCP was compared between HCC tissue and adjacent tissue. The chi-square test was used for comparison of categorical data between groups. The receiver operating characteristic (ROC) curve analysis was performed to obtain the area under the ROC curve (AUC), and goodness of fit was evaluated. Youden index was used to determine the optimal cut-off value. and the Kaplan-Meier survival analysis was used to analyze the association of CRCP expression with the recurrence and prognosis of HCC, and the log rank test was used for comparison between the two groups. Results Among the 79 HCC patients, there were 67 male patients and 12 female patients, with an age of 10-72 years, and 20 patients had portal vein tumor thrombus. As for pathological grade, 1 had grade 4 HCC, 61 had grade 3 HCC, and 17 had grade 2 HCC; as for BCLC stage, 5 had BCLC stage 0 HCC, 55 had BCLC stage A HCC, 11 had BCLC stage B HCC, and 8 had BCLC stage C HCC. Western blot showed that the expression level of CRCP in HCC tissue was lower than that in adjacent tissue in 4 patients, and immunohistochemistry showed that the expression level of CRCP in HCC tissue was significantly lower than that in adjacent tissue in 75.9% of the patients. Low CRCP expression was associated with CK19 positivity, incomplete tumor capsule, presence of portal vein tumor thrombus, and high pathological grade (χ²=6.410, 4.829, 9.319, and 9.083, all P < 0.05). Compared with the low CRCP expression group, the high CRCP expression group had a significantly longer overall survival time and a significantly lower recurrence rate (P < 0.001 and P=0.009). Conclusion Patients with low CRCP expression in HCC tissue tend to have a poorer prognosis than those with high CRCP expression, and CRCP may participate in the development, progression, and metastasis of HCC, suggesting that this molecule can be used as a potential biomarker to predict the prognosis of HCC patients.
- Carcinoma, Hepatocellular /
- Receptors, Calcitonin Gene-Related Peptide /
- Neoplasm Metastasis /
- Prognosis

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图 1 6例患者肝癌及癌旁组织Western Blot检测结果

注：N，癌旁组织；T，肿瘤组织。

Figure 1. Western Blot analysis of CRCP expression in the tumoral and peri-tumoral tissues from six HCC patients

下载: 全尺寸图片幻灯片

图 2 79例患者肝癌及癌旁组织免疫组化检测结果

注：a，2例肝癌患者癌组织及癌旁组织中的CRCP表达情况(×12.8)，CRCP在胞浆与胞核中均有表达，与癌组织相比，癌旁组织染色更深，提示癌旁组织中CRCP表达量更高。b，79例肝癌患者CRCP的相对表达，每位患者癌组织与癌旁组织的免疫组化评分(Positivity，简称P)相除后取log值[即log(P_癌组织/P_癌旁组织)]并作图，其中60例(75.9%)患者的癌组织CRCP表达量低于癌旁组织即为低表达部分，19例(24.1%)癌组织CRCP表达量高于癌旁组织即为高表达部分。

Figure 2. Immunohistochemistry analysis of CRCP expression in the tumoral and peri-tumoral tissues from 79 HCC patients

下载: 全尺寸图片幻灯片

图 3 肝癌组织中CRCP表达水平与患者预后的关系

Figure 3. Expression of CRCP in HCC tissues correlated with the prognosis

下载: 全尺寸图片幻灯片

表 1 CRCP分子表达与肝癌患者临床病理特征的关系

Table 1. The relationship between CRCP expression and clinicopathological characteristics in HCC patients

指标	例数	CRCP表达[例(%)]		χ²值	P值
指标	例数	高表达	低表达	χ²值	P值
性别				0.186	0.666
男	67	29(43.3)	38(56.7)
女	12	6(50.0)	6(50.0)
年龄				0.532	0.466
≤50岁	42	17(40.5)	25(59.5)
＞50岁	37	18(48.6)	19(51.4)
CK18				1.963	0.161
阳性	56	22(39.3)	34(60.7)
阴性	23	13(56.5)	10(43.5)
CK19				6.410	0.011
阳性	20	4(20.0)	16(80.0)
阴性	59	31(52.5)	28(47.5)
AFP				2.244	0.134
≤400 μg/L	54	27(50)	27(50)
＞400 μg/L	25	8(32)	17(68)
肿瘤大小				0.001	0.977
≤5 cm	45	20(44.4)	25(55.6)
＞5 cm	34	15(44.1)	19(55.9)
包膜				4.829	0.028
不完整	49	17(34.7)	32(65.3)
完整	30	18(60.0)	12(40.0)
肝硬化				1.332	0.722
无	35	17(48.6)	18(51.4)
大结节性	3	2(66.7)	1(33.3)
小	36	14(38.9)	22(61.1)
混合	5	2(40.0)	3(60.0)
门静脉癌栓				9.319	0.002
无	59	32(54.2)	27(45.8)
有	20	3(15.0)	17(85.0)
病理分级				9.083	0.003
1~2级	17	13(76.5)	4(23.5)
3~4级	62	22(35.5)	40(64.5)

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