Background/Aims: Liver transplantation (LT) following total hepatectomy is a life-saving treatment for hepatocellular carcinoma (HCC). The HCC recurrence after LT hinders the effectiveness of the procedure. The objective of this study is to develop a pre-operative risk stratification model based on a liquid biopsy. Methods: We conducted a comprehensive multi-omics study of 260 HCC patients from three centers, including clinical data, low-coverage whole-genome sequencing of cell-free DNA (cfDNA) from plasma, as well as whole-exome, single-nucleus RNA, and spatial transcriptomics from matched tumor and non-tumor tissues. Results: We identified cfDNA-derived copy number alteration (CNA) signatures associated with post-transplant recurrence. By integrating cfDNA-derived CNA profiles with single-cell transcriptomic data, we traced recurrence-associated cfDNA to a distinct subpopulation of malignant cells within the primary tumor. These cells were embedded in a pro-metastatic microenvironment of specialized endothelial subtypes and cancer-associated fibroblasts. Notably, most recurrence-associated lesions were detectable in cfDNA prior to liver transplantation (LT). Building on these insights, we developed the ZJU Criteria based on CNA fragments and tumor markers, a pre-LT risk prediction tool that integrates conventional clinical factors with cfDNA-derived CNA signatures, and validated it using internal and independent external cohorts. Conclusion Our findings suggest that post-transplant recurrence commonly originates from advanced subclones that emerge late during tumor evolution. The ZJU Criteria provides an accurate, non-invasive strategy that significantly improves pre-LT risk stratification and clinical decision-making for patients with HCC.

Liver cancer (LC) remains one of the most prevalent tumors globally. Despite considerable advancements in its clinical treatment in recent years, the global burden of this disease is projected to increase. According to World Health Organization statistics, the number of LC deaths worldwide in 2022 was 760,000, ranking third among all cancers. Therefore, there is an urgent need to delve deeper into the realms of LC diagnosis, prognosis, and treatment. Proteomics, as an emerging technology leveraging mass spectrometry (MS)-based protein identification, liquid chromatography separation, and bioinformatics-driven data integration, enables the systematic characterization of proteome dynamics to address critical challenges in LC research. These challenges include the discovery of diagnostic and prognostic biomarkers as well as therapeutic targets. By scrutinizing the expression of specific proteins in both normal and cancerous tissues through high-throughput protein microarrays, researchers can uncover novel biomarkers. The use of these biomarkers alone or in combination with established ones holds the potential to significantly advance the fields of LC diagnosis, prognosis, and treatment. In general, the application of proteomics has significantly promoted the development of LC research. In this review, we summarized the latest advancements in the application of proteomics in LC, with a particular focus on diagnosis, prognosis, and treatment. We also concluded selected protein markers, aiming to offer guidance for further applications and present additional therapeutic targets for LC.

Compared with the widely used rodents,pigs are anatomically,physiologically,and genetically more similar to humans,making them high-quality models for the study of liver diseases.Here,we review the latest research progress on pigs as a model of human liver disease,including methods for establishing them and their advantages in studying cystic fibrosis liver disease,acute liver failure,liver regeneration,non-alcoholic fatty liver disease,liver tumors,and xenotransplantation.We also emphasize the impor-tance of genetic engineering techniques,mainly the CRISPR/Cas9 system,which has greatly enhanced the utility of porcine models as a tool for substantially advancing liver disease research.Genetic engineering is expected to propel the pig as one of the irreplaceable animal models for future biomedical research.

AIM:To explore the effect and mechanism of the interfering Gal-1 on epithelial-mesenchymal transition(EMT),migration and proliferation in MDA-MB-231 cells via transforming growth factor-β(TGF-β)pathway.METHODS:The stable cell lines(shGal-1)which Gal-1 expression were inhibited completely and their control cell lines were used as experimental cells.Western blot assay was used to detect the effects of shGal-1 on EMT process of MDA-MB-231 cells after TGF-β treatment;The effect of shGal-1 on cell migration and invasion after TGF-β treatment was verified by cell scratch and transwell test;The effect of shGal-1 on the TGF-β pathway related proteins were detected by western blot;Finally,the effect of shGal-1 on cell proliferation was detected by MTT and western blot.RESULTS:shGal-1 inhib-ited TGF-β-mediated EMT in MDA-MB-231 cells and regulated phosphorylation of pathway signaling molecules(ERK,AKT and GSK3β);shGal-1 could inhibit the proliferation of MDA-MB-231 cells.CONCLUSION:shGal-1 can inhibit the TGF-β-mediated EMT,migration and proliferation of MDA-MB-231 cells.

Background and aims:Inflammatory factors play significant roles in the development and occurrence of hepatocellular carcinoma(HCC).However,the tumor-protective functions of growth differentiation factors(GDFs)in HCC are yet to be clarified.In this study,we aimed to evaluate the expression levels of 10 GDFs in tumor and paratumor tissues from patients with HCC and perform in vitro and in vivo ex-periments to elucidate the role of GDF7 in regulating the proliferation and metastasis of HCC.Methods:The gene expression of 10 GDFs was compared between HCC and paratumors using The Cancer Genome Atlas dataset and patient-derived tissues.A tumor microarray containing 108 HCC tissue samples was used to explore the prognostic value of GDF7 expression.Loss-of-function experiments were also performed in vitro and in vivo to investigate the role of GDF7 in HCC.Results:The mRNA and protein levels of GDF7 were significantly lower in HCC tumors than in para-tumors(P＜0.001).Kaplan-Meier analysis showed that decreased GDF7 expression in HCC was asso-ciated with worse overall survival(5-year rate:61.8％vs.27.5％,P＜0.001)and increased recurrence risk(P＜0.001).Multivariate Cox regression analysis demonstrated that low GDF7 expression,the presence of microvascular invasion,and elevated alpha-fetoprotein(AFP)levels were independent risk factors for tumor recurrence and poor survival.Downregulation of GDF7 also increased the tumor growth in HCC cells and in an HCC xenograft model.GDF7 knockdown promoted migration and invasion via epithelial-mesenchymal transition.Meanwhile,a negative correlation between JunB proto-oncogene(JUNB)and GDF7 was observed in HCC tissues.Modulating JUNB levels altered GDF7 protein expression.Conclusions:GDF7 is a potential biomarker for predicting superior outcomes in patients with HCC.GDF7 amplification is a potential therapeutic option for HCC.

Tacrolimus (TAC), also called FK506, is one of the classical immunosuppressants to prevent allograft rejection after liver transplantation. However, it has been proved to be associated with post-transplant hyperlipemia. The mechanism behind this is unknown, and it is urgent to explore preventive strategies for hyperlipemia after transplantation. Therefore, we established a hyperlipemia mouse model to investigate the mechanism, by injecting TAC intraperitoneally for eight weeks. After TAC treatment, the mice developed hyperlipemia (manifested as elevated triglyceride (TG) and low-density lipoprotein cholesterol (LDL-c), as well as decreased high-density lipoprotein cholesterol (HDL-c)). Accumulation of lipid droplets was observed in the liver. In addition to lipid accumulation, TAC induced inhibition of the autophagy-lysosome pathway (microtubule-associated protein 1 light chain 3β (LC3B) II/I and LC3B II/actin ratios, transcription factor EB (TFEB), protein 62 (P62), and lysosomal-associated membrane protein 1 (LAMP1)) and downregulation of fibroblast growth factor 21 (FGF21) in vivo. Overexpression of FGF21 may reverse TAC-induced TG accumulation. In this mouse model, the recombinant FGF21 protein ameliorated hepatic lipid accumulation and hyperlipemia through repair of the autophagy-lysosome pathway. We conclude that TAC downregulates FGF21 and thus exacerbates lipid accumulation by impairing the autophagy-lysosome pathway. Recombinant FGF21 protein treatment could therefore reverse TAC-caused lipid accumulation and hypertriglyceridemia by enhancing autophagy.
Animals ; Mice ; Tacrolimus ; Liver ; Cholesterol, LDL ; Autophagy ; Disease Models, Animal

Objective:To explore the early and medium-long term outcomes of steatosis donor liver transplantation(LT)for an optimal clinical application.Methods:From January 2015 to December 2020, this retrospective cohort study was conducted jointly at Shulan (Hangzhou) Hospital, First Affiliated Hospital of Zhejiang University and First Hospital of Jilin University. The relevant clinicopathological and follow-up data were collected from 1535 LT recipients. For comparison, propensity score was utilized for case-control matching of steatosis and non-steatosis donor livers. According to presence or absence of liver steatosis, the recipients were divided into two groups of steatosis donor liver (n=243) and non-steatosis donor liver (n=1292). And 1∶1 propensity score matching was made for two groups. Then early and medium-long term outcomes of two groups were examined. Counts were described as absolute numbers. Kaplan-Meier method was employed for calculating survival time and plotting survival curve and Log-rank test for survival analysis. COX regression model was utilized for univariate and multivariate analyses. Based on basic metabolic disease pre-LT, steatosis donor liver recipients were divided into three subgroups: BMI ≥25 kg/m 2 with hypertension or diabetes (n=21), BMI<25 kg/m 2 and no hypertension or diabetes (n=130) and other recipients (n=92). A comparative study was performed for determining the prognosis of subgroups according to the different characteristics of recipient and donor liver. Results:No significant inter-group difference existed in 2-year survival post-LT ( P=0.174). However, significant inter-group difference in survival existed after 2 years post-LT ( P=0.004). And 3/5-year survival rate of steatosis donor liver was 66.4% and 44.2% respectively. Both were significantly lower than those of non-steatosis donor liver. Multivariate Cox regression analysis indicated that steatosis donor liver and male recipients were independent risk factors for prognosis >2 years survival post-LT( P=0.008, P=0.004). Subgroup analysis of steatosis liver donors showed that the prognosis of patients with BMI ≥25 kg/m 2 with hypertension or diabetes was significantly worse than other subgroups (BMI <25 kg/m 2 with no hypertension or diabetes and other recipients) <2 years survival post-LT ( P=0.029, P=0.043). Conclusions:Steatosis donor liver does not affect early survival of recipients, yet reduces medium-long term survival rate of recipients notably. In steatosis donor liver recipients, early survival rate declines markedly in recipients with preoperative BMI ≥25 kg/m 2 with hypertension or diabetes as compared with BMI <25 kg/m 2 with no hypertension or diabetes group.

Objective:To investigate the prognosis and influencing factors of liver transplantation (LT) for hepatocellular carcinoma (HCC) using steatotic donor liver.Methods:The retrospective cohort study was conducted. The clinicopathological data of 152 pairs of donors and the corresponding recipients undergoing LT for HCC in the two medical centers [89 pairs in Shulan (Hangzhou) Hospital and 63 pairs in the First Affiliated Hospital of Zhejiang University School of Medicine] from January 2015 to December 2019 were collected. Of 152 donors, there were 131 males and 21 females, aged (48±12)years, and there were 130 cases with liver mild steatosis and 22 cases with liver moderate steatosis. Of 152 recipients, there were 138 males and 14 females, aged (52±9)years. Observation indicators: (1) follow-up, overall survival and tumor recurrence free survival of recipients; (2) influencing factors for overall survival and tumor recurrence free survival of recipients; (3) construction and validation of nomogram prediction model for overall survival and tumor recurrence free survival of recipients. Follow-up was conducted using outpatient examination and telephone interview to detect survival and tumor recurrence of recipients up to December 2020. Measurement data with normal distribution were represented as Mean± SD, and measurement data with skewed distribution were represented as M( IQR). Count data were described as absolute numbers. The Kaplan-Meier method was used to calculate the survival time and draw survival curve, and the Log-Rank test was used for survival analysis. The COX regression model was used for univariate and multivariate analysis. The independent risk factors were brought into the R 3.6.2 software to construct nomogram prediction model and draw the receiver operating characteristic (ROC) curve. The accuracy and discrimination of the nomogram prediction model were evaluated using the area under curve (AUC) and the calibration curve. Results:(1) Follow-up, overall survival and tumor recurrence free survival of recipients. All the 152 recipients undergoing LT for HCC using steatotic donor liver were followed up for 45.8(27.6)months, with the overall survival time and tumor recurrence free survival time of 36.5(32.3)months and 30.4(34.6)months. The 1-year, 3-year overall survival rates and tumor recurrence free rates of the 152 recipients were 73.4%, 55.8% and 62.2%, 43.4%, respectively. (2) Influencing factors for overall survival and tumor recurrence free survival of recipients. Results of univariate analysis showed that the donor liver cold ischemia time (CIT), the donor liver warm ischemia time (WIT), graft-to-recipient weight ratio (GRWR), ABO compatibility, recipient body mass index (BMI), recipient tumor diameter, recipient tumor number, recipient tumor differentiation degree, recipient preoperative alpha fetoprotein (AFP) were related factors influencing the overall survival of recipients ( hazard ratio=6.26, 1.90, 2.47, 4.08, 0.55, 5.16, 3.62, 5.28, 2.65, 95% confidence interval as 3.01?13.03, 1.07?3.38, 1.36?4.49, 2.07?8.03, 0.31?0.98, 2.56?10.42, 1.95?6.72, 1.60?17.42, 1.48?5.01, P<0.05) and the donor liver CIT, GRWR, ABO compatibility, recipient tumor diameter, recipient tumor number, recipient tumor differentiation degree, recipient preoperative AFP were related factors influencing the tumor recurrence free survival of recipients ( hazard ratio=4.24, 2.53, 4.05, 3.39, 3.10, 5.19, 2.63, 95% confidence interval as 2.50?7.21, 1.54?4.17, 2.12?7.72, 2.04?5.62, 1.91?5.03, 2.04?13.18, 1.61?4.30, P<0.05). Results of multivariate analysis showed that donor liver CIT ≥8 hours, GRWR ≥2.5%, recipient tumor diameter ≥8 cm and recipient preoperative AFP ≥400 μg/L were independent risk factors influencing the overall survival of recipients ( hazard ratio=4.21, 2.58, 4.10, 2.27, 95% confidence interval as 1.98?8.96, 1.24?5.35, 1.35?12.43, 1.13?4.56, P<0.05) and donor liver CIT ≥8 hours, GRWR ≥2.5%, recipient tumor diameter ≥8 cm, recipient tumor number ≥3 and recipient preoperative AFP ≥400 μg/L were independent risk factors influencing the tumor recurrence free survival of recipients ( hazard ratio=3.37, 2.63, 2.42, 2.12, 2.22, 95% confidence interval as 1.70?6.67, 1.40?4.96, 1.04?5.66, 1.08?4.18, 1.26?3.90, P<0.05). (3) Construction and validation of nomogram prediction model for overall survival and tumor recurrence free survival of recipients. The donor live CIT, GRWR, recipient tumor diameter, recipient preoperative AFP were used to construct nomogram prediction model for overall survival of recipients and the donor liver CIT, GRWR, recipient tumor diameter, recipient tumor number, recipient preoperative AFP were used to construct nomogram prediction model for tumor recurrence free survival of recipients. The ROC curve showed that the AUC of the nomogram prediction model for overall survival of recipients was 0.84 (95% confidence interval as 0.76?0.92, P<0.05), with the optimal diagnostic value as 7.3 and the specificity and sensitivity as 87.6% and 70.0%. The AUC of the nomogram prediction model for tumor recurrence free survival of recipients was 0.79 (95% confidence interval as 0.71?0.87, P<0.05), with the optimal diagnostic value as 5.8 and the specificity and sensitivity as 97.4% and 52.5%. The calibration curve showed that the nomogram prediction model had good distinction for high risk recipients in overall survival and tumor recurrence free survival. Conclusion:Donor liver CIT ≥8 hours, GRWR ≥2.5%, recipient tumor diameter ≥8 cm and recipient preoperative AFP ≥400 μg/L are independent risk factors influencing the overall survival of recipients who underwent LT for HCC using steatotic donor liver and donor liver CIT ≥8 hours, GRWR ≥2.5%, recipient tumor diameter ≥8 cm, recipient tumor number ≥ 3 and recipient preoperative AFP ≥400 μg/L are independent risk factors influencing the tumor recurrence free survival of recipients.

Objective:To investigate the proper choice and improve the survival rate in neonates suffering from giant omphalocele (GO)with liver protrusion by analyzing the clinical characteristics, different treatments and outcomes.Methods:Neonates with GO admitted in Department of Neonatal Surgery in Wuhan Children′s Hospital were retrospected from January 2009 to May 2019.The treatment methods include: (1) repaired with similar silo-bag by staged operation; (2) one-stage repair with patch or not; (3) delayed repair by conservative treatment for some time (from 2 to 8 weeks) firstly.Data included gestational age, gender, birth weight, average diameter of omphalocele, contents of omphalocele, associated malformation, treatment method, duration of ventilator use and clinical outcome.These neonates were divided into 2 groups according to the contents of protrusion: group with liver protrusion and group with liver and other organs protrusion.Clinical data were summarized and compared between different groups.Results:Sixteen cases were collected, including 9 males and 7 females.The average age was (1.25±0.45) d (1-2 days), the average birth weight was (2.48±0.37) kg, and pregnant week was (36.23±1.17) weeks.The average diameter of the omphalocele was (9.88±3.30) cm, ranging from 5 cm to 15 cm.Seven cases were belonged to the group with liver protrusion, cases undergone one-stage repair, delayed repair were 6 cases and 1 case, respectively.There were 9 cases in group with liver and other organs protrusion, and the protruded organs included liver, intestine, colon or spleen.Among them, cases needed one-stage repair, delayed repair and silo-bag repair were 3 cases, 3 cases, 3 cases, respectively.Cases needed respiratory machine were 3 cases (42.8%) and 8 cases (88.89%) in group with liver protrusion and group with liver and other organs protrusion, respectively.Neonates had a longer time of needing respiratory machine in group with liver protrusion, comparing with the neonates in group with liver and other organs protrusion[(30.67±19.0) h vs.(106.25±69.36) h, P=0.021], and the risk rate to use respiratory machine was 5.143(95% CI: 0.727-36.368). There were no dead cases in group with liver protrusion.Three cases were dead in group with liver and other organs protrusion.All the patients were followed from 3 months to 4 years, and hernia in abdominal wall was found in 5 cases, including 1 case in group with liver protrusion and 4 cases in group with liver and other organs protrusion.If the dead cases had been excluded, the rate of hernia was 66.7% (4/6 cases) in group with liver and other organs protrusion. Conclusions:Individual treatment should be emphasized in neonates suffering from GO with liver protrusion.Neonates with single liver protrusion have a better prognosis than those with liver and other organs protrusion.One-stage repair is recommended in the former, and the delayed repair is highly recommended in the latter.The accessory liver should be considered when GO neonates presenting liver protrusion.