Significance of Golgi Protein 73, Alpha-Fetoprotein and Vascular Endothelial Growth Factor Expression in Diagnosis of Primary Hepatic Cancer

Aim: We measured Golgi protein73 (GP73), alpha-fetoprotein (AFP), and vascular endothelial growth factor (VEGF) expression in primary hepatic cancer (PHC) and assessed their clinical significance. Methods: Forty-five PHC and tumor-adjacent specimens and 14 normal liver specimens were examined. GP73, AFP, and VEGF expression was measured via immunohistochemistry and a correlation of protein expression with clinical pathology of PHC was suggested. Results: GP73, AFP, and VEGF expression was significantly higher in PHC and tumor-adjacent tissue compared to tumor-adjacent tissue (0.143 ± 0.018 vs. 0.124 ± 0.027, 0.116 ± 0.026 vs. 0.098 ± 0.014, and 0.126 ± 0.027 vs. 0.092 ± 0.016, respectively;all p 0.088 ± 0.029, 0.098 ± 0.014 vs. 0.073 ± 0.011, and 0.092 ± 0.016 vs. 0.076 ± 0.018, respectively;all p < 0.05), respectively. GP73 expression was positively correlated with pathological grade and cirrhosis, but not with tumor size, nodules, clinical stage and serum AFP. VEGF expression was positively correlated with tumor size, nodules, portal vein tumor thrombus, and clinical stage, but not with the degree of tumor differentiation and serum AFP. Expression of GP73 and VEGF was greater than that of AFP in PHC (both p < 0.05). Conclusion: GP73 is highly expressed in PHC and may be a diagnostic marker. Combined detection of GP73, AFP, and VEGF is helpful for diagnosis of PHC.

Clinical evaluation of single or joint of golgi protein 73 and alpha-fetoprotein in hepatocellular carcinoma diagnosing

Objective: The aim of the present study was to further evaluate the clinical value of single or joint of golgi protein 73 (GP73) and alphafetoprotein (AFP) in diagnosis of hepatocellular (HCC). Methods: One hundred and eighteen, 94 and 47 serum samples from the patients with HCC, chronic liver disease (CLD) and liver cirrhosis (LC) were collected, respectively. Serum levels of AFP and GP73 were assayed with commercial kit according to the manufacturer's instructions. Results: Patients with HCC had higher serum concentration of AFP than that of the patients with CLD (P < 0.01), but was similar to that of the patients with LC. Serum GP73 levels in the patients with CLD or LC were significantly lower than that in the patients from HCC group (P < 0.01). Among 118 HCC patients, the positive rate of GP73 and AFP was 80.5% and 48.3%, respectively (P < 0.001). The ROC curve analysis showed that the AUC value of GP73 was higher than that of serum AFP. Moreover, the sensitivity and the accuracy of GP73 were 77.1% and 82.6%, respectively, which were greater more than that of AFP at 90% specificity (28.8% and 59.8%, respectively). The AUC, the sensitivity and the accuracy of GP73 in combination of AFP (AFP/GP73) were 0.855, 78.0% and 83.0%, respectively, which were similar to that of GP73 alone but were much higher than that of the single marker AFP. Conclusion: For HCC diagnosing, GP73 was more sensitive and specific than AFP. The diagnostic value of AFP/GP73 was similar to GP73 but was much higher than AFP.

血清GP73、HMGB1、FLP1、sST2与慢性乙型病毒性肝炎患者 HBV-DNA载量、肝功能及肝纤维化标志物的相关性分析

目的 研究血清高尔基体蛋白73(GP73)、高迁移率族蛋白B1(HMGB1)、纤维蛋白原样蛋白1(FLP1)、可溶性生长刺激表达基因2蛋白(sST2)与慢性乙型病毒性肝炎(CHB)患者乙型肝炎病毒-脱氧核糖核酸(HBV-DNA)载量、肝功能指标及肝纤维化标志物的相关性。方法 选择滨州市中医医院2021年2月至2022年2月收治的166例CHB患者作为研究对象。测定所有患者的HBV-DNA载量,并根据HBV-DNA载量的差异分为低载量组、中载量组、高载量组。另选取同期健康体检人员60例作为健康对照组。检测并比较各组肝功能指标水平、肝纤维化标志物水平,以及血清GP73、HMGB1、FLP1、sST2水平。以Spearman/Pearson相关分析血清GP73、HMGB1、FLP1、sST2与HBV-DNA载量、肝功能指标及肝纤维化标志物的相关性。结果 低载量组76例,中载量组50例,高载量组40例。低载量组、中载量组、高载量组血清GP73、HMGB1及sST2水平均高于健康对照组(P<0.05);且随着HBV-DNA载量的增加,GP73、HMGB1及sST2水平升高(P<0.05)。低载量组、中载量组、高载量组血清FLP1水平均低于健康对照组(P<0.05);且随着HBV-DNA载量的增加,FLP1水平下降(P<0.05)。中载量组、高载量组丙氨酸氨基转移酶(ALT)、天冬氨酸氨基转移酶(AST)及γ-谷氨酰转移酶(GGT)水平均高于低载量组(P<0.05),且高载量组ALT、AST及GGT水平高于中载量组(P<0.05)。中载量组、高载量组透明质酸(HA)、层粘连蛋白(LN)及Ⅳ型胶原(CⅣ)水平均高于低载量组(P<0.05),且高载量组HA、LN及CⅣ水平均高于中载量组(P<0.05)。血清GP73、HMGB1、sST2与CHB患者HBV-DNA载量、ALT、AST、GGT、HA、LN、CⅣ水平均呈正相关(r>0,P<0.05),而血清FLP1与CHB患者HBV-DNA载量、ALT、AST、GGT、HA、LN、CⅣ水平呈负相关(r<0,P<0.05)。结论 血清GP73、HMGB1、FLP1、sST2水平可有效反映CHB患者的HBV-DNA载量、肝功能和肝纤维化情况。

胆管细胞癌患者中miR-320a和GP73表达及预后研究

目的:观察胆管细胞癌患者中微小RNA-320a(miR-320a)、高尔基体膜蛋白73(GP73)协同作用及预后研究。方法:选取2014年6月至2018年12月本院收治的胆管细胞癌患者142例为胆管细胞癌组,并选取同时间段内本院健康体检者146例为健康组。以实时荧光定量法(qRT-PCR)检测两组血清miR-320a表达水平;以酶联免疫吸附法测定(enzyme-linked immunosorbent assay,ELISA)两组GP73表达水平;分析胆管细胞癌组患者血清中miR-320a、GP73表达水平与胆管细胞癌临床特征及miR-320a表达水平与GP73关系;分析胆管细胞癌预后的影响因素;分析血清miR-320a、GP73表达水平对胆管细胞癌的诊断价值。结果:胆管细胞癌组患者血清miR-320a表达水平明显低于健康组(P<0.05),GP73水平明显高于健康组(P<0.05);胆管细胞癌组患者血清miR-320a、GP73表达水平均与血管浸润、TNM分期、肝硬化有关(均P<0.05);胆管细胞癌患者血清中miR-320a表达水平与GP73呈负相关(P<0.05);血管浸润、TNM分期、肝硬化、GP73为胆管细胞癌预后的危险因素(P<0.05),miR-320a为胆管细胞癌预后的保护因素(P<0.05);血清miR-320a、GP73表达水平诊断胆管细胞癌的曲线下面积(area under curve,AUC)分别为0.875、0.878,对应灵敏度分别为84.5%、85.4%,特异度分别为79.5%、79.5%;血清miR-320c、GP73联合诊断胆管细胞癌的AUC为0.956,其灵敏度、特异度分别为90.8%、91.8%。结论:胆管细胞癌患者血清中miR-320a低表达,GP73高表达,miR-320a与GP73具有一定负相关性,两者可能通过相互作用,进而共同影响胆管细胞癌发生发展。

肝癌标志物高尔基体蛋白73基因在大肠杆菌中的表达

目的克隆、表达和鉴定肝癌标志物高尔基体蛋A73（G1773）基因序列（1028～1327bp）。方法应用Biosun生物学软件分析GP73全序列，预测其抗原表位，将GP73蛋白基因克隆到表达载体PGEX-4T-2上，构建重组表达质粒PGEX-4T-2／GP73,转化大肠杆菌BL21，异丙基-β—D-硫代半乳糖吡喃糖苷（WFG）诱导表达，利用GST亲和层析柱对重组蛋白进行纯化．并用蛋白印迹分析技术（Western blot）和酶联免疫吸附测定方法（ELISA）检测其抗原性．并与商品化的GP73抗原进行比较。结果本研究纯化的GP73蛋白在大肠杆菌中获得高效表达，SDS～聚丙烯酰胺凝胶电泳（SDS—PAGE）显示其相对分子质量为37kD，与预计大小一致。Western blot和ELISA实验证实，纯化的GP73蛋白具有良好的抗原性。结论本研究成功克隆和表达了GP73序列．为制备抗体和肝癌诊断试剂的研发奠定了实验基础。

丙型肝炎病毒对高尔基体蛋白73表达的影响

目的探讨丙型肝炎病毒(HCV)对高尔基体蛋白73(GP73)表达的影响。方法选取2015年3月至2016年12月期间上海市浦东新区公利医院收治的68例HCV患者为研究对象,以同期74例健康体检者为对照组,采用酶联免疫吸附试验(ELISA)检测两组受检者血清中GP73的含量,逆转录-聚合酶链式反应(RT-PCR)和免疫印迹法检测Huh7.5.1细胞中GP73 m RNA和蛋白的表达情况,ELISA检测细胞上清中GP73的含量,用SPSS19.0软件进行统计学分析。结果 HCV组患者的GP73血清含量为(146.5±12.8)ng/m L,明显高于健康对照组的(45.2±2.7)ng/m L,差异有统计学意义(P<0.05);感染HCV的Huh7.5.1细胞中GP73 m RNA和蛋白相对表达量分别为(1.204±0.143)和(1.174±0.136),明显高于对照细胞的(0.306±0.019)和(0.298±0.015),差异均具有统计学意义(P<0.05);HCV感染的细胞上清中GP73含量为(85.6±10.3)ng/m L,明显高于对照细胞的(39.5±8.3)ng/m L,差异具有统计学意义(P<0.05)。结论 HCV可能通过上调GP73的表达来参与HCC的形成,但HCV调节GP73表达的具体分子机制尚待进一步研究。

Golgi protein 73, hepatocellular carcinoma and other types of cancers

Hepatocellular carcinoma(HCC)is one of the most common malignant tumors with a low survival rate.The identification of mechanisms underlying the development of HCC helps uncover cellular and mo-lecular targets for the diagnosis,prevention,and treatment of HCC.Golgi protein 73(GP73)level is up-regulated in HCC patients and potentially can be a therapeutic target.Despite many studies devoted to GP73 as a marker for HCC early diagnosis,there is little discussion about the function of GP73 in HCC tumorigenesis.Given the poor response to currently available HCC therapies,a better understanding of the role of GP73 in HCC may provide a new therapeutic target for HCC.The current paper summarizes the role of GP73 as a diagnostic marker as well as its roles in liver carcinogenesis.Its roles in other types of cancer are also discussed.

Relationship between the Level of Serum Golgi Protein 73 and the Risk of Short-term Death in Patients with ALD-ACLF

Background and Aims:As a hepatocellular carcinoma biomarker,serum Golgi protein 73(GP73)is reportedly related to inflammation.Acute-on-chronic liver failure(ACLF)is characterized by severe systemic inflammation.In this study,we aimed to explore the association between the GP73 level and short-term mortality in patients with alcohol-associated liver disease-related ACLF(ALD-ACLF).Methods:This retrospective cohort study involved 126 Chinese adults with ALD-ACLF.Baseline serum GP73 level was measured using enzymelinked immunosorbent assay.Patients were followed-up for 90 d and outcomes were assessed.Data were analyzed using multivariate Cox regression and piecewise linear regression analyses.The predictive value of GP73 and classic models for the short-term prognosis of participants were evaluated and compared using receiver operating characteristic curves.Results:The serum GP73 level was independently associated with an increased mortality risk in patients with ALD-ACLF.Compared with the lowest tertile,the highest serum GP73 level predisposed patients with ALD-ACLF to a higher mortality risk in the fully adjusted model[at 28 days:hazard ratio(HR):4.29(0.99–18.54),p=0.0511;at 90 days:HR:3.52(1.15–10.79),p=0.0276].Further analysis revealed a positive linear association.GP73 significantly improved the accuracy of the Child-Turcotte-Pugh score,model for end-stage liver disease score,and model for end-stage liver diseasesodium score in predicting short-time prognosis of patients with ALD-ACLF.Conclusions:The serum GP73 level is a significant predictor of the subsequent risk of death in patients with ALD-ACLF.GP73 improved the predictive value of classic prognostic scores.

高尔基体糖蛋白73的表达特征及其对肝癌与肝硬化的鉴别诊断价值

目的研究血清高尔基体蛋白73（GP73）的表达特征及其在肝癌与肝硬化鉴别诊断中的临床价值。方法采用酶联免疫吸附试验（ELISA）检测80例原发性肝癌、65例肝硬化和50名健康体检者血清GP73含量，同时用电化学发光法平行测定血清AFP。多组比较采用Kruskal-Wallis检验，两组比较采用Mann-Whitney检验，ROC曲线分析检测效能并确定其临界值，敏感度、特异度比较用配对χ^2检验，GP73表达与临床参数的关系用Spearman等级相关分析。结果肝癌组GP73浓度中位数为282．0μg／L，高于肝硬化组（211．8μg／L）和健康对照组（58．3μg／L），差异有统计学意义（H=93．30，P〈0．01）。GP73、AFP鉴别肝癌与肝硬化患者的最佳临界值分别为318．1μg／L和13．4μg／L时，GP73的敏感度为45．0%（36／80），低于AFP的65．0％（52／80），χ^2=8．02，P〈0．05;GP73的特异度为83．1％（54／65），低于AFP的87．7％（57／65），χ^2=0．27，P〉0．05；GP73诊断肝癌的ROC曲线下面积为0．65（95％可信区间0．54～0．72），与AFP曲线下面积0．75（95％可信区间0．67～0．83）相比，差异无统计学意义（Z=1．88，P〉0．05）。血清GP73与肝硬化、血管浸润及肿瘤分期相关（r值分别为0．27、0．29、0．27，P值均〈0．05），与性别、年龄、AFP〉13．4μg／L、肿瘤大小和远处转移无相关关系（r值分别为0．13、0．10、0．03、0．18、0．04，P值均〉0．05）。结论在肝癌与肝硬化的鉴别中，血清GP73与AFP的诊断效能相似，但AFP的敏感度优于GP73。血清GP73的过表达可能与肿瘤的负荷和侵袭性有关。

高尔基体蛋白的基因克隆及原核表达

目的构建重组高尔基体蛋白的原核表达质粒并在大肠杆菌中诱导表达,为进一步研发肝癌诊断试剂奠定基础。方法通过RT-PCR获得GP73目的片段,将其克隆入表达载体p ET-28a,经酶切及测序鉴定后,在BL21菌株中经异丙基-β-D-硫代半乳糖苷(IPTG)诱导表达,用Western Blot鉴定。结果扩增的片段大小与预期一致;p ET-28a-GP73重组质粒通过酶切鉴定与DNA测序证实序列正确;诱导的蛋白用SDS-PAGE电泳和Western Blot检测,与预期相符可见一特异性条带。结论成功获得GP73目的基因并构建了p ET-28aGP73原核表达载体,实现了该融合蛋白的可溶性表达。