Golgi protein 73 in liver fibrosis

Objective: This study aimed to evaluate the diagnostic value of serum Golgi protein 73(GP73) alone and GP73 combined with liver stiffness measurement (LSM), aspartate aminotransferase/platelet ratio index (APRI), and 4-factor-based fibrosis index (FIB4) in diagnosing liver fibrosis in patients with chronic liver disease of different etiologies. Methods: A diagnostic test. A total of 68 patients who underwent liver biopsy in the Department of Traditional and Western Medical Hepatology of the Third Hospital of Hebei Medical University from October 2019 to December 2020 were selected to detect serum GP73 levels. iLivTouch was used to assess liver stiffness measurement (LSM). In addition, serum alanine aminotransferase (ALT), aspartate aminotransferase (AST), gamma-glutamyl transferase (GGT), total bilirubin (TBil), direct bilirubin (DBil), triglyceride (TG), total cholesterol (TC), low-density lipoprotein (LDL) levels, and peripheral platelet (PLT) counts were assayed. The correlation between GP73 and the above indexes was assessed, and APRI and FIB-4 were calculated. SPSS 21.0 statistical software was used for statistical analysis. The area under the receiver operating characteristic curve was calculated to evaluate diagnostic efficacy of GP73 in identifying hepatic fibrosis stages. Furthermore, the difference between GP73 and liver stiffness, as well as APRI and FIB4 in diagnosing significant fibrosis was assessed. Results: Based on liver biopsy, 13, 18, 17, and 20 cases were diagnosed as stages S0-1, S2, S3, and S4, respectively. The AUC of GP73 diagnosing hepatic fibrosis stage S≥3 and S=4 were 0.806 and 0.844 at cut-off points of 2.06 and 3.27 μg/L, and the sensitivity and specificity were 93.5%, 61.5%, 90.0%, 70.3%, respectively. In addition, GP73 levels were positively correlated with the degree of liver fibrosis (r=0.547, P<0.001). Conclusions: The efficacy of serum GP73 level in diagnosing the degree of liver fibrosis in patients with chronic liver disease from different causes was significantly higher than that of APRI, FIB4, and LSM. The combination of GP73 and FIB4 can further improve the accuracy of diagnosis of liver fibrosis staging S≥3 and S=4, which is a reliable serological marker for the diagnosis of fibrosis in patients with chronic liver disease.

To observe the dynamic levels of serum Golgi protein 73(GP73) in patients prior to and after the onset of liver cancer, and to explore the related factors. From 2007 to 2012, a periodical screening program was carried out in a group of high risk population with positive Hepatitis B surface antigens (HBsAg) , twice a year. Their serum specimens from every screening time point were kept in Qidong Biobank until liver cancer was diagnosed. Thirty-nine patients with liver cancer were recruited for the study, each of them at least had three times of specimens collected as well as B ultrasound scan (BUS) exam results at onset of disease and within 30 months before diagnosed, amongst 6 time points. In total, there were 162 specimens collected to test GP73 by double-antibody sandwich enzyme-linked immuno-sorbent assay (ELISA). Statistical analyses of time series and differences among groups were performed by stata software 10. The average value of 39 patient's GP73 at the time point of liver cancer onset was (126.77 ± 73.73) µg/L, while the values at the other five time points prior to the onset were (128.32 ± 81.18) , (129.97 ± 83.62) , (127.38 ± 80.10) , (135.52 ± 97.88) and (138.24 ± 93.58) µg/L, respectively, with no significant difference (F = 0.07, P = 0.997). No obvious changing trends of GP73 were observed among the 39 liver cancer cases at the 6 time points. All 162 samples were divided into two groups: without hepatic cirrhosis (63 samples) and with cirrhosis (99 samples) according to findings of B-ultrasonic wave; whose average GP73 values were separately (97.16 ± 51.39) and (151.20 ± 91.68) µg/L. The difference showed statistical significance (F = 18.22, P < 0.01). Furthermore, if we grouped the samples by the average value of GP73 at 130.19 µg/L, then there were only 1/14 of the subjects without hepatic cirrhosis having higher GP73 values, but 12 of the 25 subjects with hepatic cirrhosis having higher GP73 values. The difference showed statistical significance (P = 0.013). The results of Linear regression model also showed that there was no correlation between GP73 and time series (t = 0.75, P = 0.455), but significant correlation between GP73 and hepatic cirrhosis (t = 4.30, P < 0.01). No significant changes of the dynamic levels of GP73 could be found among the liver cancer patients within 30 months prior to the onset of disease. GP73 values of the patients with liver cancer may depend on their background of hepatic diseases; and hepatic cirrhosis might be one of the main influencing factors or confounding factors.

Alanine aminotransferase (ALT) is the most commonly used marker of liver injury, but normal ALT levels are seen in a proportion of chronic hepatitis B virus (HBV)-infected patients with severe liver injury. Golgi protein 73 (GP73) is a promising alternative marker of liver injury. This study assessed the relation between GP73 levels and liver disease severity, monitored the kinetic changes in GP73 levels in chronic HBV patients receiving entecavir (ETV) therapy, and investigated the potential diagnostic and prognostic values of serum GP73 as a new liver injury biomarker in chronic HBV infections.This study enrolled 1150 patients with chronic HBV infections, 200 of whom were retrospectively enrolled in this study after receiving 1 year of ETV treatment. GP73 expression in liver tissue was detected by immunohistochemistry. GP73 levels in single or serial serum samples were measured by enzyme-linked immunosorbent assay.Immunohistochemical analysis indicated that GP73 protein expression in the liver increased progressively with pathologic progression from nonexistent or mild hepatitis to severe hepatitis and cirrhosis during chronic HBV infection. Serum GP73 levels were positively correlated with the disease severity of chronic HBV infections (r = 0.58, P < 0.001). In patients with normal ALT levels, serum GP73 concentrations were significantly higher in patients with prominent hepatic inflammatory injury and fibrosis than in patients without hepatic inflammatory injury or fibrosis. Serum GP73 concentrations and GP73 protein expression were decreased in the liver tissues of patients whose ALT levels normalized after 1 year of ETV antiviral therapy.Changes in serum GP73 levels were closely associated with changes in liver injury severity, and, therefore, GP73 may be an effective new liver inflammatory injury biomarker, and could be useful for monitoring the prognosis of chronic HBV infectious patients with normal ALT levels.

Objective To investigate the correlations of serum golgi protein 73 (GP73) levels with disease severity and liver pathology in patients with chronic hepatitis B (CHB). Methods A total of 430 patients including 107 mild CHB, 191 moderate CHB and 132 severe CHB were successively collected from the 180th Hospital of People's Liberation Army during January 2012 and October 2013, among whom 133 patients underwent liver biopsy. Thirty individuals were also enrolled as healthy controls. Serum GP73 levels were measured by enzyme linked immunosorbent assay (ELISA). Among 430 patients, 72 received antiviral, anti-inflammatory and hepatoprotective treatments, and serum levels of GP73 were observed before and after treatments. Variance analysis and SNK-q tests were performed to compare serum GP73 levels among different groups. Receiver operating characteristic (ROC) curve was used to evaluate serum GP73 levels in diagnosis of severe CHB. Pearson correlation coefficient and linear regression were used to evaluate the correlations of serum GP73 levels with pathological grading and staging of liver tissues. Results Serum levels of GP73 in CHB patients were higher than those in healthy controls (F=129.96, P<0.01), and it was positively correlated with disease severity in CHB patients (r=0.695, P<0.01). The area under curve (AUC) was 0.911 (95% confidence interval: 0.870-0.952) for diagnosis of severe CHB and with an optimal cut-off value of 163.00 ng/mL. The sensitivity and specificity was 90.00% and 81.95%, respectively. In 133 patients who underwent liver biopsy, the serum GP73 levels significantly increased with the elevation of hepatic inflammation activity grade and liver fibrosis stage (F=65.49 and 37.26, P<0.01), and positive correlations were observed (r=0.766 and 0.673, P<0.01). After antiviral, anti-inflammatory and hepatoprotective treatments, serum GP73 levels were decreased in 50 patients who achieved clinical remission [(178±87) ng/mL vs. (100±46) ng/mL, t=6.429, P<0.01], while GP73 levels were increased in 22 patients with disease progressed [(120±65) ng/mL vs. (174±64) ng/mL, t=-4.409, P<0.01]. Conclusion Serum GP73 levels are correlated with disease severity and liver pathology in CHB, and it can be used as an indicator of lever inflammation grade and fibrosis stage. Key words: Hepatitis B, chronic; Biopsy, needle; Immunohistochemisty; Golgi protein 73

mTORC1 Up-Regulates GP73 to Promote Proliferation and Migration of Hepatocellular Carcinoma Cells and Growth of Xenograft Tumors in Mice

Development of an alpha-fetoprotein and Golgi protein 73 multiplex detection assay using xMAP technology

Endoplasmic reticulum (ER) stress induced by hepatitis B virus (HBV) infection is associated with the development of liver fibrosis. Golgi protein 73 (GP73) is increased during HBV infection. Nevertheless, whether GP73 during HBV infection mediates ER stress in liver fibrosis is still poorly understood. TGF-β1 was used to induce HepG2.2.15 cells to establish liver fibrosis cells model. GP73 expression was evaluated using qRT-PCR analysis and Western blot. HepG2.2.15 cells viability and proliferation were assessed via CCK-8 assay and EdU assay, respectively. The protein levels of α-SMA, fibronectin, collagen I and collagen III for liver fibrosis, GRP78, p-PERK, p-eIF2α, ATF4 and CHOP for ER stress, as well as p-Smad2 and Smad2 were evaluated by Western blot. TGF-β1 incubation obviously elevated GP73 expression, while GP73 knockdown reduced the GP73 levels in HBV-transfected HepG2215 cells. GP73 knockdown reversed the effects of TGF-β1 exposure on HepG2.2.15 cells viability and proliferation. The protein levels of liver fibrosis marker, ERS marker and p-Smad2 were remarkably increased following TGF-β1 stimulation, which were counteracted by GP73 silence or the application of 4-phenylbutyric acid (4-PBA). However, these results were opposite after tunicamycin (TM) treatment. In conclusion, knockdown of GP73 potentially impeded the advancement of liver fibrosis via mediating ERs through Smad2 signal pathway.

The incidence of hepatocellular carcinoma (HCC) has increased over the past decades in China. Current screening methods of HCC such as detection of α-fetoprotein (AFP) combined with liver ultrasonography remain unsatisfactory. Many HCC patients have already missed the optimal treatment period when diagnosed. Our study aimed to evaluate the value of Glypican 3 (GPC3) and Golgi protein 73 (GP73) in the detection of HCC. Thirty-nine patients with HCC and 31 patients with liver cirrhosis were enrolled. The level of serum GPC3 and GP73 were determined by ELISA. The expression of GPC3 mRNA and GP73 mRNA in peripheral blood mononuclear cell (PBMC) and liver tissues were also measured with qRT-PCR. Then, receiving operating characteristic (ROC) curves were plotted to detect the sensitivity and specificity of serum GPC3 and GP73 in the diagnosis of HCC. The levels of serum GPC3 and GP73 in the HCC group were significantly higher than in the cirrhosis group (p < 0.0001). Patients with GPC3 > 9.3 μg/L and GP73 > 77.68 ng/mL had a risk of HCC of 92.31%. The HCC diagnosis ROC curve analysis indicated that when setting the GPC3 cutoff value > 9.3 μg/L, AUC = 0.956. The sensitivity and specificity of GPC3 were 89.74% and 96.77%, respectively, with a positive predictive value of 97.2%, negative predictive value of 88.2%, + LR of 27.82 and - LR of 0.11. When setting GP73 cutoff value > 77.68 ng/mL, AUC = 0.937. The sensitivity and specificity of GP73 were 92.31% and 83.87%, respectively, with positive predictive value of 87.8%, negative predictive value of 89.7%, + LR of 5.72 and - LR of 0.092. No significant difference (p > 0.05) was found between GPC3 and GP73 AUC in ROC curves, indicating that these two biomarkers were equivalent in the prediction of HCC. The expression of serum GPC3 and GP73 was significantly higher in the HCC patients compared with the cirrhosis patients. GPC3 and GP73 might be effective non-invasive diagnostic indicators of HCC.

Objective: To explore the diagnostic value and model of serum Golgi protein 73 (GP73) in patients with hepatitis C cirrhosis. Methods: 271 cases with chronic hepatitis C virus infection who were treated in the Fifth Medical Center of PLA General Hospital from January 2010 to December 2017 were retrospectively collected as the research objects, including 126 cases with hepatitis and 145 cases with liver cirrhosis. Serum GP73 and liver stiffness measurement (LSM) based on transient elastography test were performed in all patients. Simultaneously, blood routine, liver function, coagulation function and other related indicators were collected. GP73 diagnostic efficiency for liver cirrhosis was evaluated by receiver operating characteristic curve (ROC). GP73 diagnostic value was clarified after comparison with aspartate aminotransferase/platelet ratio index (APRI), FIB-4 index (FIB-4) and LSM. Compensated hepatitis C virus-related cirrhosis diagnostic model based on serological index was established by logistic regression analysis. Results: The area under the receiver operating characteristic curve (AUC) of GP73, LSM, FIB-4 and APRI in the diagnosis of compensated hepatitis C virus-related cirrhosis were 0.923, 0.839, 0.836 and 0.800 respectively, and GP73 had the best diagnostic efficiency (P <0.001). LSM and GP73 combined use had improved the diagnostic sensitivity of cirrhosis to 97.24%. Multivariate logistic regression analysis revealed that GP73, age, and platelets were independent predictors of cirrhosis.Compensated hepatitis C virus-related cirrhosis diagnostic model (GAP) was established based on the result: LogitP=1/[1+exp(6.145+0.013×platelet-0.059×age-0.059×GP73)].AUC model for diagnosing compensated liver cirrhosis was 0.944, and the optimal cut-off value was 0.56, with sensitivity and specificity of 84.03% and 92.06%, respectively, and the diagnostic efficiency of this model was better than that of APRI, FIB-4, LSM and GP73 alone (P<0.05). Conclusion: GP73 is a reliable serum biomarker for the diagnosis of compensated hepatitis C virus-related cirrhosis. The GAP diagnostic model based on GP73, platelet count, and age can further improve the diagnostic efficiency and help to diagnose patients with compensated hepatitis C virus-related cirrhosis.

Objective To investigate the expression and clinical significance of serum Golgi protein 73 (GP73) , Alpha-fetoproteins (AFP) and vascular endothelial growth factor (VEGF) in primary hepatic cancer (PHC) patients. Methods Serum AFP, GP73 and VEGF levels of forty-three PHC patients and 9 patients with benign hepatic tumor were detected by electrochemiluminescence and double antibody sandwich ELISA. Results The serum levels of GP73 in different ages, gender, tumor sizes was not significantly different. The serum level of GP73 in hepatitis B virus carriers was significantly higher than no hepatitis B virus carriers, and that in hepatic cirrhosis group was significantly higher than no cirrhosis group (P<0.05) . The serum level of GP73 in PHC group was significantly higher than benign liver tumor group[ (241.413±77.079) vs (101.866±74.192) μg/L, P<0.01]. The serum levels of GP73 , AFP and VEGF before the operation were significantly higher than those after the operation [ (247.231±77.009) μg/L vs (128.334±92.235)μg/L, (674.176±1090.083) μg/L vs(178.560±289.330)μg/L,(256.666±164.76) μg/L vs (149.072±158.643) μg/L, all P<0.01]. The area of AFP, GP73, VEGF under the ROC curve in the differential diagnosis of PHC were 0.894, 0.791, 0.612. The sensitivity and specificity value of GP73 were 88.4% and 77.7%, higher than those of AFP. The sensitivity and specificity value of AFP combined with GP73 reached 96.8% and 63.7%, higher than either single detection. Conclusion GP73 is a sensitivity and specificity PHC diagnostic serum markers. AFP combined with GP73 detection has higher accuracy for PHC diagnosis. Key words: Golgi protein 73; Alpha-fetoproteins; Vascular endothelial growth factor; Primary hepatic carcinoma

The aim of this study was to determine if Golgi protein-73 (GP73) is up-regulated in hepatocellular carcinoma (HCC), and to explore the possibility of using GP73 in diagnosis and treatment of HCC. Serum GP73 was detected by a quantitative ELISA assay. A total of 372 serum samples were included, among them 43 from healthy donors (Normal), 110 from either chronic hepatitis or cirrhosis (CH/LC), and 219 from HCC patients. The levels of GP73 were compared among the 3 groups. The received operating curve (ROC), sensitivity and specificity of GP73 for HCC patients were calculated. The average level of GP73 expression in normal, CH/LC and HCC groups were (22.1 ± 8.5) ng/ml, (81.4 ± 57.2) ng/ml and (271.5 ± 202.3) ng/ml, respectively. Serum GP73 levels were significantly higher in patients with HCC compared to those with CH/LC (P < 0.001). The GP73 area under ROC was 0.857. Put 100 ng/ml as the optimal cut-off point, GP73 had a sensitivity of 76.7% and a specifically of 73.2%. GP73 level had a significantly higher sensitivity than AFP (32.0%) in diagnosis of early HCC (P < 0.001). Moreover, GP73 level was elevated in the serum (72.5%, 108/149) of individuals with HCC who had serum AFP level less than 400 ng/ml. Following-up study of 4 HCC patients with low level AFP indicated that GP73 was associated with treatment and prognosis of HCC. Higher level of GP73 can be found in the serum of patients with HCC than those without. GP73 is better than AFP for the diagnosis of early HCC and in evaluating treatment result in patients with normal AFP. Further studies may help to validate both the role and mechanism of GP73 in diagnosis of HCC.

BackgroundHepatocellular carcinoma (HCC) is the most common liver cancer arising from the hepatocytes. The most prevalent noninvasive biological marker for diagnosis of HCC is alpha-fetoprotein (AFP). Golgi protein 73 (GP73) was considered as an impending biological marker for the early diagnosis of HCC.ObjectivesTo assess the value of GP73 in serum as a biological marker for HCC comparing its sensitivity and specificity with serum AFP.MethodsThis study included 90 participants (30 patients with hepatitis-C virus (HCV)-related HCC, 30 patients having liver cirrhosis on top of chronic HCV, and 30 healthy controls). The levels of AFP and GP73 in serum were measured by chemiluminescent immune-metric assay and enzyme-linked immunosorbent assay techniques, respectively.ResultsHCC and cirrhotic patients had considerably higher AFP (P < 0.001, <0.004) and GP73 (P < 0.001, 0.047) levels than controls, whereas HCC patients had significant higher values of AFP and GP73 than cirrhotic patients (P = 0.029, <0.001, respectively). Combined AFP + GP73 had the highest sensitivity and specificity (P < 0.001) for differentiating HCC patients from controls. In discrimination of HCC patients from cirrhotic patients, serum GP73 had the highest sensitivity and specificity (P < 0.001) than AFP (P = 0.009) and combined AFP + GP73 (P < 0.001). Moreover, in discrimination of metastatic patients from nonmetastatic ones, combined AFP + GP73 had the highest sensitivity (P < 0.001) than GP73 alone (P = 0.013), whereas AFP alone had the highest specificity (P = 0.058) than combined AFP + GP73.ConclusionGP73 is a reliable biological marker for early diagnosis and detection of distant metastasis of cancerous liver diseases in patients having liver cirrhosis on top of chronic HCV infection and it would yield better results when used combined with AFP.

Background: Hepatocellular Carcinoma (HCC) is one of the most common malignancy in the liver. Modalities of diagnostic are often an obstacle in HCC surveillance. Alpha fetoprotein (AFP) is one of protein that often used in the diagnostic of HCC in chronic liver disease. Golgi protein 73 (GP73), is one of the candidate biomarkers in early diagnostic of HCC and found in biliary epithelial cells but rarely expressed by normal hepatocytes. Expression of GP73 was reported to be increased in a large number of malignancies. Aims of this study to evaluate differences in Golgi protein 73 serum (sGP73) and AFP in diagnosing hepatocellular carcinoma in patients with liver cirrhosis. Materials and Methods: This cross-sectional study was conducted at Haji Adam Malik Hospital in 2020. Serum level of GP73 and others biomarker was detected using enzyme-like immunosorbent assay. Results: From 90 subjects, Liver cirrhosis and HCC group had significantly higher AFP than the control group. AFP was superior in determining HCC to GP73. At a cut off value of &gt; 394.5.00 ng/mL, AFP yielded a sensitivity of 83.3% and specificity of 67%, for discriminating liver cirrhosis and HCC (AUC 0.84), while GP73 with cut off value of &gt; 82.5 ng/mL, sensitivity of 70% and specificity of 57% (AUC 0.74). Conclusion:GP73 was significantly higher in HCC patients in comparison to non-HCC patients and healthy population. Compared with alpha fetoprotein, GP73 was superior in discriminating HCC in healthy population but inferior in group of liver cirrhosis. Keywords: Golgi Protein 73, Alpha Fetoprotein, Hepatocellular carcinoma.

ObjectiveTo evaluate the utility of Golgi protein 73 (GP73) in the diagnosis of non-alcoholic steatohepatitis (NASH) and hepatic fibrosis (HF) staging.MethodsNinety-one patients with non-alcoholic fatty liver disease (NAFLD) were allocated to NAFL (n = 46) and NASH (n = 45) groups according to their NAFLD activity score (NAS), and there were 30 healthy controls. Serum GP73 was measured by ELISA, GP73 protein expression was evaluated using immunohistochemistry, and FibroScan was used to determine liver hardness.ResultsThe serum GP73 concentrations of the NAFL and NASH groups were significantly higher than those of controls. GP73 expression in the liver of the patients gradually progressed from absent or low to moderate or high. Serum GP73 positively correlated with liver expression, and the serum and liver GP73 of the patients positively correlated with FibroScan value and HF stage. There was a strong positive correlation of the combination of alanine aminotransferase, gamma glutamyl transferase and GP73 with NASH. The combination of serum GP73 and FibroScan value was found to predict NASH (NAS > 4) and advanced HF (stage ≥2) in patients with NAFLD using receiver operating characteristic analysis.ConclusionSerum GP73 may be useful in the diagnosis of NASH and the staging of HF.

Hepatitis C virus (HCV) infection is a leading cause of chronic liver diseases and hepatocellular carcinoma (HCC) and Golgi protein 73 (GP73) is a serum biomarker for liver diseases and HCC. However, the mechanism underlying GP73 regulates HCV infection is largely unknown. Here, we revealed that GP73 acts as a novel negative regulator of host innate immunity to facilitate HCV infection. GP73 expression is activated and correlated with interferon-beta (IFN-β) production during HCV infection in patients’ serum, primary human hepatocytes (PHHs) and human hepatoma cells through mitochondrial antiviral signaling protein (MAVS), TNF receptor-associated factor 6 (TRAF6) and mitogen-activated protein kinase kinase/extracellular regulated protein kinase (MEK/ERK) pathway. Detailed studies revealed that HCV infection activates MAVS that in turn recruits TRAF6 via TRAF-interacting-motifs (TIMs), and TRAF6 subsequently directly recruits GP73 to MAVS via coiled-coil domain. After binding with MAVS and TRAF6, GP73 promotes MAVS and TRAF6 degradation through proteasome-dependent pathway. Moreover, GP73 attenuates IFN-β promoter, IFN-stimulated response element (ISRE) and nuclear factor κB (NF-κB) promoter and down-regulates IFN-β, IFN-λ1, interleukin-6 (IL-6) and IFN-stimulated gene 56 (ISG56), leading to the repression of host innate immunity. Finally, knock-down of GP73 down-regulates HCV infection and replication in Huh7-MAVSR cells and primary human hepatocytes (PHHs), but such repression is rescued by GP73m4 (a mutant GP73 resists to GP73-shRNA#4) in Huh7-MAVSR cells, suggesting that GP73 facilitates HCV infection. Taken together, we demonstrated that GP73 acts as a negative regulator of innate immunity to facilitate HCV infection by interacting with MAVS/TRAF6 and promoting MAVS/TRAF6 degradation. This study provides new insights into the mechanism of HCV infection and pathogenesis, and suggests that GP73 is a new potential antiviral target in the prevention and treatment of HCV associated diseases.