Objective:To investigate the clinical value of peripheral blood T lymphocytes in the diagnosis and prognosis of patients with hepatocellular carcinoma (HCC) recurrence after liver transplantation.Methods:The clinical and laboratory data of 50 HCC patients, who received liver transplantation and were followed up in the Liver transplantation Center of Beijing Youan Hospital from January 2014 to December 2016, were retrospectively analyzed. The differences on clinical laboratory indicators and five-year survival were compared between HCC recurrence group ( n=29) and non-recurrence group ( n=21). Spearman correlate analysis was used to analyze the correlation between clinical laboratory indicators and HCC recurrence after liver transplantation. Receiver operator characteristic (ROC) curve was used to analyze the diagnostic value of CD4+T lymphocytes in HCC recurrence after liver transplantation. Kaplan-Meier survival curve was used to compare the survival time of patients with different CD4+T lymphocytes levels post liver transplantation. Results:Compared to non-recurrence group, the level of alanine aminotransferase, aspartate aminotransferase, γ-glutamyltransferase, albumin, lymphocytes, alpha-fetoprotein, protein induced by vitamin K deficiency or antagonist-Ⅱ, CD3+, CD4+and CD8+T lymphocytes were significantly different (all P<0.05). The median recurrence time after liver transplantation was 13.0 (6.0, 24.0) months, and the mortality rate was 100%. The 5-year mortality rate was 0 in the non-recurrence group. During 5-year follow-up, the median survival time of patients in the HCC recurrence group was 18.0 (9.0, 36.0) months, which was significantly lower than that of non-recurrence group [60.0 (60.0, 60.0) months, ( P<0.05)]. Compared with non-recurrence group, the CD3+, CD4+, and CD8+T lymphocytes were significantly lower in the recurrence group (all P<0.05). Spearman correlate analysis showed that HCC recurrence after liver transplantation was negatively correlated with the CD3+, CD8+and CD4+T lymphocytes ( r=-0.43, -0.38, -0.44, all P<0.05). ROC analysis showed that CD4+T lymphocytes at cutoff of≤265.50 cells/μl was valuable for the diagnosis of HCC recurrence after liver transplantation (specificity 100%, sensitivity 48.30%). Survival curve analysis showed that the survival time was significantly lower in the CD4≤265.50 cells/μl group [15.0 (10.0, 36.8) months] than that in the CD4>265.50 cells/μl group [53.0 (19.5, 60.0) months] ( P<0.05). Conclusion:There is a significant negative correlation between CD4+T lymphocytes and HCC recurrence after liver transplantation. CD4+T lymphocytes at cutoff value of≤265.50 cells/μl is valuable for the clinical diagnosis and prognosis evaluation of HCC recurrence after liver transplantation.

Objective:To investigate the efficacy and adverse reactions of ticagrelor combined with atorvastatin in the treatment of acute cerebral infarction (ACI).Methods:A total of 80 patients with ACI who were diagnosed and treated in Anhui Suixi County Hospital from October 2021 to October 2022 were selected retrospectively and randomly divided into the control group and observation group, each group with 40 cases. The patients in the control group were treated with routine basic treatment and atorvastatin for ACI. The patients in the observation group was treated with ticagrelor on the basis of the control group. The clinical efficacy, neurological function, daily living ability, platelet function (platelet count, platelet inhibition rate), inflammatory factors including high sensitivity C-reactive protein (hs-CRP), interleukin-6 (IL-6) and adverse reactions of the two groups were compared.Results:The total effective rate of the observation group was higher than that of the control group: 92.50%(37/40) vs. 72.50% (29/40), there was statistical differences ( P<0.05). After treatment, the score of National Institute of Health Stroke Scale of the observation group was lower than that of the control group: (9.37 ± 2.91) points vs. (14.20 ± 3.39) points, and the score of Barthel index scale (BI) was higher than that of the control group: (72.26 ± 13.27) points vs. (58.93 ± 9.43) points, there were statistical differences ( P<0.05). After treatment, the platelet count and platelet adenosine diphosphate (ADP) inhibition rate of the observation group were higher than those of the control group: (284.65 ± 41.58) × 10 9/L vs. (210.46 ± 36.12) × 10 9/L, (79.43 ± 16.42)% vs. (62.40 ± 13.95)%, there were statistical differences ( P<0.05). After treatment, the serum hs-CRP and IL-6 levels of the observation group were lower than those of the control group: (11.64 ± 2.96) mg/L vs. (19.75 ± 4.57) mg/L, (4.26 ± 0.93) ng/L vs. (8.95 ± 1.83) ng/L, there were statistical differences ( P<0.05). There was no significant difference in the incidence of adverse reactions between the two groups after treatment ( P>0.05). Conclusions:Ticagrelor combined with atorvastatin has a better therapeutic effect on ACI, which can effectively improve the neurological deficit and the ability of self-care.

Objective:To screen and identify differential proteins, analyze lipid metabolism-related proteins and pathways, and explore their functions and biological processes in liver tissue of patients with alcoholic liver disease using tandem mass tag (TMT) labeling technology.Methods:Liver tissues that met the inclusion criteria were collected. Eight samples from patients with alcoholic cirrhosis and three samples from the normal control group were screened out. The TMT technique was used to screen differential proteins, perform signaling pathway enrichment analysis, and analyze protein interaction networks to explore the biological processes involved in them.Results:Proteomic analysis identified 2 741 kinds of differentially expressed proteins in the two groups of data with statistical significance ( P < 0.05). The standard criteria of P < 0.05 and |log2(foldchange)| > 1 had screened out 106 kinds of differentially expressed proteins. Compared with the control group, the alcoholic liver disease group had 12 kinds of up-regulated proteins and 94 kinds of down-regulated proteins. Among them, there were 2 kinds of up-regulated differential proteins related to lipid metabolism and 14 kinds of down-regulated differential proteins. The results of bioinformatics analysis showed that these proteins were primarily involved in biological processes such as lipid transport, regulation of lipase activity, fatty acid binding, and cholesterol metabolism in lipid metabolism and also had a close link to signal pathways related to lipid metabolism such as peroxisome proliferator-activated receptor signaling pathways, cholesterol metabolism, triglyceride metabolism, and regulation of lipolysis in adipocytes. Conclusion:The 16 kinds of lipid metabolism-related differential proteins may be the key proteins in the pathogenesis of alcoholic liver disease.

The etiology and pathogenesis of autoimmune liver diseases has always been a hot area of research. Pathogen infections can elicit an autoimmune response and often become the key pathogenic factor of immune diseases. Based on the literature data and the author's clinical experience, this review will briefly introduce the role and influence of pathogen infections in the development and progression of autoimmune liver diseases from the aspects such as molecular mimicry mechanism, in order to further understand the pathogenesis of autoimmune liver diseases.

Background::The hepatitis B virus X (HBx) protein plays a critical role in the initiation and progression of hepatitis B virus (HBV)-associated hepatocellular carcinoma (HCC). In the early stage of the disease, HBx facilitates tumor onset by inactivating the tumor suppressor p53. The p53-encoding gene, however, is frequently mutated or deleted as the cancer progresses to the late stage and, under such circumstance, the p53 homolog TAp63 can harness HCC growth by transactivating several important p53-target genes.Methods::To determine whether HBx regulates TAp63, we performed co-immunoprecipitation assay, real-time quantitative polymerase chain reaction, immunoblotting, and flow cytometry analysis in p53-null cancer cell lines, Hep3B and H1299.Results::HBx interacts with the transactivation domain of TAp63, as HBx was co-immunoprecipitated with TAp63 but not with ΔNp63. The interaction between HBx and TAp63 abolished transcriptional activity of TAp63, as evidenced by the reduction of the levels of its target genes p21 and PUMA, consequently leading to restricted apoptosis and augmented proliferation of HCC cells. Conclusion::HBV induces progression of HCC that harbors defective p53 by inhibiting the tumor suppressor TAp63.

Objective:To investigate the molecular mechanism of sorafenib against hepatocellular carcinoma.Methods:Sorafenib efficacy was screened and verified by the hepatocellular carcinoma patient-derived tumor xenograft (PDX) model. Veterinary B-mode ultrasonography and in vivo confocal laser scanning microscopy were used to observe PDX angiogenesis. Immunohistochemistry was used to observe the expression of proliferation and angiogenesis-related proteins in PDX tissue. Real-time quantitative PCR technology was used to observe the RUNX3 gene in PDX tissues. SPSS 17.0 statistical software was used for statistical analysis.Results:Four cases of PDX were used to screen the efficacy of sorafenib. PDX1 had a significant response to sorafenib, with an inhibition rate of 68.07%. Compared with the control group, sorafenib had significantly inhibited PDX1 relative tumor volume (5.76±2.14 vs. 11.71±2.87, P<0.05). Cell division index (39.50±7.72 vs. 67.10±9.14, P<0.05) and Ki67 expression (288.6±43.40 vs. 531.70±55.60, P<0.05) were significantly decreased. Veterinary B-mode ultrasonography showed evident blood flow signals in PDX1 tumors. In vivo confocal laser scanning microscopy results showed that sorafenib had significantly reduced the total vessel length (1573.00±236.21 vs. 2675.03±162.00, P<0.05) and area (11 145.33±1931.97 vs. 20 105.37±885.93, P<0.05)) of PDX1 tumors. Immunohistochemical results showed that sorafenib had significantly down-regulated the protein expressions of CD34 (27.55±3.76 vs. 45.47±5.57, P<0.05), VEGF (16.33±2.86 vs. 22.77±3.20, P<0.05) and MVD (38.75±6.01 vs. 55.50±8.61, P<0.05). Real-time PCR results showed that sorafenib had significantly up-regulated RUNX3 gene expression (2.14±0.71 vs. 1.00±0.36, P<0.05). However, there was a negative correlation between the expression of RUNX3 gene and the ratio of VEGF-positive cells in sorafenib group ( R2=0.509 7). Conclusion:Sorafenib may inhibit the PDX angiogenesis and the growth of hepatocellular carcinoma by regulating the RUNX3-VEGF pathway.

ObjectiveTo investigate the differentially expressed proteins in the plasma exosome of acute-on-chronic liver failure (ACLF) patients with different prognoses, to analyze their functions and biological processes, and to provide a basis for clinical diagnosis. MethodsA prospective study was performed for 10 ACLF patients who were hospitalized and diagnosed in Beijing YouAn Hospital, Capital Medical University, from July 2019 to October 2019, and the patients were followed up for 90 days. The patients who died or received liver transplantation were enrolled as liver transplantation/death group (5 patients), and the patients who survived were enrolled as survival group (5 patients). The Mann-Whitney U test was used for comparison of general data between the two groups. The label-free quantitative proteomic method was used for identification and quantitative analysis of plasma exosome proteins to screen out differentially expressed proteins, and a functional enrichment analysis was performed. R-3.5.1 software was used to perform a hierarchical cluster analysis of differentially expressed proteins to analyze the biological processes involving these proteins. ResultsA total of 860 proteins were identified by the exosome proteomic analysis, and according to the criteria of upregulation ＞1.2 folds or downregulation ＞1.2 folds (P＜0.05), there were 116 differentially expressed proteins. Compared with the liver transplantation/death group, the survival group had 62 upregulated proteins and 54 downregulated proteins. The bioinformatics analysis showed that these differentially expressed proteins mainly participated in immune reaction, signal transduction, vesicle-mediated transport, cell death, and cell proliferation and were closely associated with the signaling pathways including inflammatory response, carbohydrate and amino acid metabolism, hepatocyte injury, and hepatocyte regeneration. ConclusionDifferentially expressed proteins screened out by the label-free quantitative proteomic method can be used as serological markers for the early diagnosis and prognostic evaluation of ACLF.

Objective To investigate the value of cytokines in bile combined with clinical indices in predicting the degree of liver injury after liver transplantation. MethodsA total of 16 patients undergoing liver transplantation who were hospitalized in Center of Organ Transplantation, The Affiliated Hospital of Qingdao University, from January to December 2018 were enrolled, and according to the level of alanine aminotransferase (ALT) on day 1 after surgery, the patients were divided into mild liver injury (ALT ＜500 U/L) group with 10 patients and severe liver injury (ALT ＞500 U/L) group with 6 patients. Bile samples were collected on days 1, 3, 5, and 7 after surgery, and MILLIPLEX assay was used to measure the levels of 17 cytokines. R software was used to perform principal component analysis (PCA) of bile cytokines and clinical indices and gene ontology (GO) enrichment analysis of bile cytokines. The two-independent-samples t-test was used for comparison of normally distributed continuous data between two groups; The Mann-Whitney U test was used for comparison of non-normally distributed continuous data between two groups. A Spearman correlation analysis was performed to evaluate the correlation between clinical indices and bile cytokines. ROC curve analysis was used to evaluate the predictive value of cytokines in bile and clinical indices for liver injury after liver transplantation. ResultsCompared with the mild liver injury group, the severe liver injury group had significantly higher expression levels of bile Fractalkine (Z=-2.828, P=0.003), soluble CD40 ligand (sCD40L) (Z=-2.850, P=0.008), interleukin-4 (Z=-2.398, P=0.017), CXCL10 (Z=-2.475, P=0.023), and macrophage inflammatory protein-1α (Z=-1844, P=0.043). The correlation analysis showed that on day 1 after liver transplantation, aspartate aminotransferase, ALT, and lactate dehydrogenase were positively correlated with the levels of several cytokines in bile (all P＜0.05). The area under the ROC curve of Fractalkine, sCD40L and AST were 0.933 (0.812-1.000), 0.833 (0.589-1.000) and 0.917 (0.779-1.000), respectively, suggesting that AST and Fractalkine and sCD40L in bile on the first day after liver transplantation have significant predictive value for liver injury. The results of PCA showed that bile cytokines combined with clinical indices on day 1 after liver transplantation could better distinguish the patients with mild liver injury from those with severe liver injury. GO analysis showed that bile cytokines were associated with positive feedback regulation of external stimulus, cell chemotaxis, receptor ligand activity, cytokine activity, and cytokine-cytokine receptor interaction. ConclusionFractalkine and sCD40L in bile can predict the degree of liver injury after liver transplantation.

Objective:To construct apoptosis-stimulating of p53 protein 2 (ASPP2) gene knockout mice using diethylnitrosamine (DEN)-induced liver cancer model to study the biological functions of ASPP2.Methods:The sgRNA oligonucleotides were constructed, and ASPP2 knockout mice were prepared with the CRISPR/Cas9 system. PCR and sequencing methods were used to identify the genotypes of F0 and F1 generations and their progeny. DEN was used to induce ASPP2+/- mice to establish liver cancer model.Results:PCR and sequencing results showed that ASPP2 gene was successfully knocked out in F0 generation mice. The genotype of F1 generation mice was accorded with ASPP2+/- and had obtained stable heredity. The success rate of DEN-induced liver cancer model (7/8 and 3 / 8) of ASPP2 + /-mice obtained by self-hybridization of F1 generation was significantly higher than that of wild-type mice.Conclusion:ASPP2 knockout mice were successfully constructed based on the CRISPR/Cas9 system. The success rate of DEN-induced liver cancer model of ASPP2 knockout mice was significantly higher than that of the wild-type mice.

Objective To discuss introperative value of laparoscopic splenectomy (LS) treatment assisted with preoperative splenic artery embolization (SAE) in the patients with the hypersplenism secondary to portal hypertension and splenomegaly. Methods The clinical data of 38 patients with the hypersplenism secondary to portal hypertension and splenomegaly admitted to the First People′s Hospital of Wenling from April 2015 to April 2018 were analyzed. Among them, 21 patients underwent LS alone (alone group) and 17 patients underwent LS assisted with preoperative SAE (combined group). Including length of the spleen and liver function Child-Pugh grade, operative time, blood transfusion rate, intraoperative blood loss, intraoperative blood transfusion volume and conversion rate were compared between two groups. Then the clinical value of LS treatment assisted with preoperative splenic artery embolization was discussed. Results The splenic volume of combined group was significantly reduced after SAE: (627 ± 195) cm3vs. (998 ± 251) cm3, P＜0.05. The conversion rate was 23.8%(5/21) in alone group, while no patient required open surgery in the combined group. Compared with that in alone group, operative time of the combined group was shorter [(143 ± 27) min vs. (189 ± 33) min], the blood loss volume was less [(155 ± 49) ml vs. (302 ± 76) ml)], and the differences were statistically significant (P＜ 0.05). The blood transfusion rates of combined group was lower [2/17 vs. 33.3% (7/21)], and intraoperative blood transfusion volume was less [(192 ± 42) ml vs. (399 ± 87) ml] . The differences were statistically significant (P＜0.05). Conclusions LS treatment assisted with preoperative SAE has some advantages, such as shorter operative time, lower surgical laparotomy rate, less intraoperative blood transfusion, less bleeding and shorter length of stay.