STAT3 Promotes Schistosome-Induced Liver Injury by Inflammation, Oxidative Stress, Proliferation, and Apoptosis Signal Pathway.

Schistosomiasis is a parasitic helminth disease that can cause organ lesions leading to health damage. During a schistosome infection, schistosome eggs can flow into the liver along the portal vein. Numerous inflammatory cells gather around the eggs, causing granulomas and fibrosis in the liver. In this process, many molecules are involved in the initiation and regulation of the fibrous scar formation. However, the precise molecular mechanisms responsible for the progression of granuloma formation and fibrosis initiation caused by schistosome infection have not been extensively studied. In this study, C57BL/6 wild-type mice and Stat3flox/flox Alb-Cre mice were infected with cercariae of Schistosoma japonicum Liver injury, effector molecule levels, and RNA transcriptome resequencing of liver tissue were detected at 4, 5, and 6 weeks postinfection. We investigated the role of STAT3 (signal transducer and activator of transcription 3) in Schistosoma-induced liver injury in mice. After 6 weeks postinfection, there was obvious liver fibrosis. A sustained pathological process (inflammation, oxidative stress, proliferation, and apoptosis) occurred in S. japonicum-induced liver fibrosis initiation. Meanwhile, we observed activation of the STAT3 pathway in hepatic injury during S. japonicum infection by RNA transcriptome resequencing. Liver deficiency of phospho-STAT3 alleviated infection-induced liver dysfunction, hepatic granuloma formation, and fibrosis initiation. It also promoted STAT3-dependent apoptosis and reduced liver inflammation, oxidative stress, and proliferation. Our results suggest that STAT3 signal pathway and its mediating inflammation, oxidative stress, proliferation, and apoptosis are involved in S. japonicum-induced liver injury and may be a new potential guideline for the treatment of schistosomiasis.

Roadmap to functional cure of chronic hepatitis B: An expert consensus.

Hepatitis B virus (HBV) infection continues to be a major public health issue worldwide. HBsAg loss is associated with functional remission and improved long-term outcome, and is considered to be a 'functional cure' (also referred to as clinical or immunologic cure) for chronic hepatitis B. This ideal goal of therapy can be achieved using optimized combination regimens with direct-acting antivirals [eg nucleos(t)ide analogues (NAs)] and immunomodulators [eg pegylated interferon alpha2a (Peg-IFN)] in selected patients with chronic hepatitis B. Among different combination therapies currently available, those with NA lead-in followed by Peg-IFN in virally suppressed patients has been demonstrated to be effective. This review provides an updated overview of the evidence supporting the use of combination therapies and summarizes expert consensus on the roadmap to attain functional cure for chronic hepatitis B patients.

Study on the relationship between insulin growth factor 1 and liver fibrosis in patients with chronic hepatitis C with type 2 diabetes mellitus.

OBJECTIVE: To investigate the correlation between serum protein level of insulin growth factor 1 (IGF-1) and the degree of liver fibrosis in patients with chronic hepatitis C (CHC) combined with type 2 diabetes mellitus (T2DM). METHODS: The cases are divided into four groups. Then serum levels of IFG-1, alanine aminotransferase (ALT), aspartate aminotransferase (AST), hepatitis C virus (HCV) RNA, and HCV genotypes were detected simultaneously in patients with hepatitis C, liver stiffness measurement (LSM) was measured by transient elastography, and aspartate aminotransferase platelet ratio (APRI) score was determined. RESULTS: There was no significant difference between CHC with T2DM group and CHC group in diabetes family history (P > 0.05), but the difference between the two groups were significantly lower than that of T2DM group ( P < 0.05). The levels of fasting insulin and homeostatic model assessment of insulin resistance (HOMA-IR) in CHC group with T2DM group were significantly higher than those in the other two groups ( P < 0.05), while the IGF-1 RNA and the serum protein level in the two groups were significantly lower than those in the CHC group, and were lower than those in the control group ( P < 0.05). The level of serum IGF-1 was negatively correlated with HOMA-IR, LSM, and APRI score in CHC with T2DM group ( r = -0.71, -0.75, and -0.69; P < 0.01). CONCLUSION: The degree of hepatic fibrosis in patients with CHC combined with T2DM was higher than that in non-T2DM patients with CHC, which was mainly related to insulin resistance (IR) induced by 1b genotype HCV infection. IR can lead to impaired synthesis of IGF-1, and the degree of damage has a corresponding relationship with hepatic fibrosis.

[Cloning of the Helicobacter pylori thioredoxin-1 gene and characterization].

OBJECTIVE: To clone the Helicobacter pylori (Hp) thioredoxin-1 (Trx1) gene and construct the recombinant expression vector containing the target gene, then to express and purify the protein, and detect its activity. METHODS: The cDNA gene of the Hp Trx1 was amplified by RT-PCR from the international standard strain 26695, using the specific primers containing double endonuclease digesting sites. The Hp Trx1 cDNA was then inserted into the pEASY-T1 vector to construct the pEASY-T1-Hp Trx1 recombinant vector. The next step was to double digest the pEASY-T1-Hp Trx1 recombinant vector and insert the target gene into pET-30a to construct the pET-30a-Hp Trx1 recombinant vector, which was transferred to E.coli BL21 plys S to express the Hp Trx1 protein. The recombinant protein was purified by Ni affinity chromatography, and then its activity of disulfide reductase was detected. RESULTS: By DNA sequencing, the Hp Trx1 cDNA was successfully inserted into the pET-30a vector and was in accordance with GenBank (HP0824). The E.coli containing pET-30a-Hp Trx1 recombinant vector successfully expressed Hp Trx1 protein. Through the detection of the activity, the recombinant Hp Trx1 protein was identified to have the activity of disulfide reductase. CONCLUSION: The prokaryotic expression vector pET-30a-Hp Trx1 was successfully constructed. The recombinant protein Hp Trx1 was successfully expressed and purified, which had the activity of disulfide reductase. This study lays the foundation for further research on the biological activity of Hp Trx1 and the mechanism of its function in tumor genesis.

Transmission of hepatitis E virus from rabbits to cynomolgus macaques.

The recent discovery of hepatitis E virus (HEV) strains in rabbits in the People's Republic of China and the United States revealed that rabbits are another noteworthy reservoir of HEV. However, whether HEV from rabbits can infect humans is unclear. To study the zoonotic potential for and pathogenesis of rabbit HEV, we infected 2 cynomolgus macaques and 2 rabbits with an HEV strain from rabbits in China. Typical hepatitis developed in both monkeys; they exhibited elevated liver enzymes, viremia, virus shedding in fecal specimens, and seroconversion. Comparison of the complete genome sequence of HEV passed in the macaques with that of the inoculum showed 99.8% nucleotide identity. Rabbit HEV RNA (positive- and negative-stranded) was detectable in various tissues from the experimentally infected rabbits, indicating that extrahepatic replication may be common. Thus, HEV is transmissible from rabbits to cynomolgus macaques, which suggests that rabbits may be a new source of human HEV infection.

Better performance of PIVKA-II for detecting hepatocellular carcinoma in patients with chronic liver disease with normal total bilirubin.

BACKGROUND: Serum protein induced by vitamin K absence or antagonist-II (PIVKA-II) is a promising biomarker for hepatocellular carcinoma (HCC) surveillance. AIM: To identify the contributing factors related to the abnormal elevation of PIVKA-II level and assess their potential influence on the performance of PIVKA-II in detecting HCC. METHODS: This study retrospectively enrolled in 784 chronic liver disease (CLD) patients and 267 HCC patients in Mengchao Hepatobiliary Hospital of Fujian Medical University from April 2016 to December 2019. Logistic regression and the area under the receiver operating characteristic curve (AUC) were used to evaluate the influencing factors and diagnostic performance of PIVKA-II for HCC, respectively. RESULTS: Elevated PIVKA-II levels were independently positively associated with alcohol-related liver disease, serum alkaline phosphatase (ALP), and total bilirubin (TBIL) for CLD patients and aspartate aminotransferase (AST) and tumor size for HCC patients (all P < 0.05). Serum PIVKA-II were significantly lower in patients with viral etiology, ALP ≤ 1 × upper limit of normal (ULN), TBIL ≤ 1 × ULN, and AST ≤ 1 × ULN than in those with nonviral disease and abnormal ALP, TBIL, or AST (all P < 0.05), but the differences disappeared in patients with early-stage HCC. For patients with TBIL ≤ 1 × ULN, the AUC of PIVKA-II was significantly higher compared to that in patients with TBIL > 1 × ULN (0.817 vs 0.669, P = 0.015), while the difference between ALP ≤ 1 × ULN and ALP > 1 × ULN was not statistically significant (0.783 vs 0.729, P = 0.398). These trends were then more prominently perceived in subgroups of patients with viral etiology and HBV alone. CONCLUSION: Serum PIVKA-II has better performance in detecting HCC at an early stage for CLD patients with normal serum TBIL.

Enterovirus 71 viral capsid protein linear epitopes: identification and characterization.

BACKGROUND: To characterize the human humoral immune response against enterovirus 71 (EV71) infection and map human epitopes on the viral capsid proteins. METHODS: A series of 256 peptides spanning the capsid proteins (VP1, VP2, VP3) of BJ08 strain (genomic C4) were synthesized. An indirect enzyme-linked immunosorbent assay (ELISA) was carried out to detect anti-EV71 IgM and IgG in sera of infected children in acute or recovery phase. The partially overlapped peptides contained 12 amino acids and were coated in the plate as antigen (0.1 µg/µl). Sera from rabbits immunized with inactivated BJ08 virus were also used to screen the peptide panel. RESULTS: A total of 10 human anti-EV71 IgM epitopes (vp1-14 in VP1; vp2-6, 21, 40 and 50 in VP2 and vp3-10, 12, 15, 24 and 75 in VP3) were identified in acute phase sera. In contrast, only one anti-EV71 IgG epitope in VP1 (vp1-15) was identified in sera of recovery stage. Four rabbit anti-EV71 IgG epitopes (vp1-14, 31, 54 and 71) were identified and mapped to VP1. CONCLUSION: These data suggested that human IgM epitopes were mainly mapped to VP2 and VP3 with multi-epitope responses occurred at acute infection, while the only IgG epitope located on protein VP1 was activated in recovery phase sera. The dynamic changes of humoral immune response at different stages of infection may have public health significance in evaluation of EV71 vaccine immunogenicity and the clinical application of diagnostic reagents.

A type-specific nested PCR assay established and applied for investigation of HBV genotype and subgenotype in Chinese patients with chronic HBV infection.

BACKGROUND: Many studies have suggested that hepatitis B virus (HBV) genotypes show not only geographical distribution and race specificity, but also are associated with disease progression and response to interferon treatment. The objective of this study was to develop a nested polymerase chain reaction (nPCR) assay for genotypes A-D and subgenotypes B1, B2, C1 and C2 of hepatitis B virus (HBV) and to investigate the distribution characteristics of HBV genotypes/subgenotype in China. METHODS: After redesigning the primers and optimizing the reaction conditions using common Taq polymerase, the sensitivity, specificity and reproducibility of the method were evaluated using plasmids and serum samples. In total, 642 serum samples from patients with chronic HBV infection were applied to investigate the distribution of HBV genotype and subgenotype in China. RESULTS: The genotype and subgenotype could be identified when the HBV DNA load of a sample was ≥10(2.3) IU/mL. For the 639 successfully genotyped samples, the sequencing results of 130 randomly selected samples (20.3%, 130/639) were consistent with those of the nPCR method. The present study showed that HBV genotype B (11.2%, 72/642), C (68.2%, 438/642) and D (7.2%, 46/642) were circulating in China, while genotype C was the dominant strain except for western region where genotype D was the prevalent strain. The main subgenotypes of genotypes B and C were B2 (87.5%, 63/72) and C2 (92.9%, 407/438), respectively. CONCLUSIONS: The low-cost nPCR method would be a useful tool for clinical and epidemiological investigation in the regions where genotypes A-D are predominant.

Higher dN/dS ratios in the HCV core gene, but not in the E1/HVR1 gene, are associated with human immunodeficiency virus-associated immunosuppression.

Coinfection with HCV and HIV is prevalent among former commercial blood donors in some rural areas in China. Genetic variability of the HCV core and E1/HVR1 was investigated in 23 patients chronically infected with HCV-1b, with or without concomitant HIV infection. Genetic variability in the core sequence was higher under HIV-associated immunocompromised conditions. Both the Shannon entropy values at each nucleotide position and the dN/dS values at each codon were statistically higher in HIV/HCV-coinfected patients with lower CD4+ T cell counts (p-values were <0.0001 and equal to 0.0372, respectively). The more significant difference of dN/dS value occurred in a specific subregion of the core gene that is enriched in CTL/Th epitopes (p = 0.0083). The dN/dS values of full-length core antigen were found to be negatively correlated with the S/CO ratio of plasma anti-HCV antibodies. By contrast, no significant difference in genetic diversity/complexity and dN/dS values in the E1/HVR1 region was found between those two groups. These results suggest that the dN/dS ratio in the core gene, but not in the E1/HVR1 gene, is influenced more by host CD4+ T cell-mediated cellular immunity.

[Impact of proton pump inhibitor omeprazole on the antiplatelet effect of clopidogrel in individuals with various CYP2C19*2 genotypes].

OBJECTIVE: To investigate the impact of omeprazole on platelet response to clopidogrel and the effect of polymorphisms of CYP2C19 on the antiplatelet effect of clopidogrel. METHODS: Platelet aggregation (PA) was assessed before 300 mg aspirin plus 300 mg loading dose of clopidogrel and after 300 mg aspirin plus 75 mg maintenance dose of clopidogrel 7 days later in 414 patients with acute coronary syndrome who have undergone percutaneous coronary intervention (PCI). Thereafter, gastric mucosal protective drugs were given (omeprazolem 20 mg, n=224 or cimetidine 800 mg, n=190). Fourteen days later, PA was measured again. Genotypes of CYP2C19*2 were analyzed with polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). RESULTS: After taken aspirin and clopidogrel, PA has decreased significantly in both groups. Compared with cimetidine, omeprazole had no significant impact on PA on 7 and 21 days post PCI. Compared with homozygotes or heterozygotes for the wild-type CYP2C19*2, patients with CYP2C19*2 AA genotype had significantly higher PA on 7 and 21 days post PCI (P<0.05). CONCLUSION: No attenuating effect on platelet response to clopidogrel has been observed for Omeprazole. The variant of CYP2C19*2 AA genotype is significantly associated with attenuated response to clopidogrel.

Coexistence of 2 types of atrial tachycardias and right ventricular outflow tract tachycardia.

Double tachycardia is a relatively uncommon type of tachycardia. In this report, we discuss a 68-year-old woman with history of frequent palpitations. Electrophysiologic study revealed that narrow QRS tachycardias from 2 origins and 1 wide QRS tachycardia were induced and each of the tachycardias was induced by the other. We found that 2 focal atrial tachycardias and 1 ventricular tachycardia originated from right ventricular outflow tract. All of these tachycardias were successfully ablated during one session, and no recurrence appeared during 10 months of follow-up.

[Upgrading to resynchronization therapy in patients with heart failure after chronic right ventricular apical pacing].

OBJECTIVE: To investigate the clinical efficacy of cardiac resynchronization therapy (CRT) through biventricular pacing in chronically right ventricular apical paced patients with heart failure. METHODS: Ten chronically right ventricular apical paced patients with left ventricular ejection fraction (EF) ≤ 35% underwent CRT upgrading. And the follow-up period was over 12 months. Seven of them reported a significant improvement in their symptoms. Two patients died and one patient had no response. As compared with pre-CRT, CRT significantly improved NYHA classification, decreased left atrium diameter [(43 ± 5) mm vs (46 ± 7) mm], pulmonary arterial pressure [(42 ± 6) mm Hg vs (54 ± 13) mm Hg] and BNP [(184 ± 73) ng/L vs (545 ± 286) ng/L] (P < 0.05), improved left ventricular EF [(35 ± 5)% vs (32 ± 4)%]. Tissue Doppler imaging revealed the maximal difference of time to peak myocardial systolic contraction of 12 left ventricular segment shortened [(136 ± 28) ms vs (97 ± 18) ms], interventricular mechanical delay shortened [(52 ± 5) ms vs (31 ± 6) ms)] after upgrading. CONCLUSION: CRT upgrading from right ventricular apical pacing may improve left ventricular function in patients with heart failure.

[Quantitative analyses of intrahepatic HBV cccDNA and serum HBsAg in 54 patients with chronic hepatitis B].

OBJECTIVE: To quantitatively detect intrahepatic HBV covalently closed circular DNA (cccDNA) and serum HBsAg; and to analyze the relationship between the two parameters and with serum HBV DNA level. METHODS: Intrahepatic cccDNA (copies/cell) was quantitated by plasmid-safe ATP-dependent Danes (PSAD) digestion in combination with rolling circle amplification and gap-spanning selective real-time PCR assay using formalin fixed paraffin-embedded liver biopsy samples. HBsAg was measured by chemiluminescence's reagent manufactured by Abbott Company using sera sampled at time-point of liver biopsy. RESULTS: Intrahepatic cccDNA level was positively correlated with serum HBsAg level (r = 0.459, P < 0.001), but not correlated with serum HBV DNA level. Serum HBsAg level was positively correlated with serum HBV DNA level (r = 0.328, P = 0.015), and reversely correlated with HBV replicative efficiency defined as the ratio of serum HBV DNA to cccDNA (r = -0.373, P = 0.005). CONCLUSION: In patients with chronic hepatitis B, intrahepatic cccDNA level is correlated with serum HBsAg level. The two parameters combined with serum HBV DNA may comprehensively reflect HBV replication activity and help evaluation of antiviral therapeutic efficacy.

[Sequence analysis of two different subtypes of thioredoxin gene of Helicobacter pylori].

OBJECTIVE: To analyze the gene sequences of Trx1 and Trx2 of Helicobacter pylori (HP) from different gastric diseases, define the amino acid sequences and investigate the relationship with these diseases. METHODS: The HP strains were isolated from gastric mucosa of 25 patients with chronic gastritis, peptic ulcer and gastric cancer respectively and cultured on solid blood agar medium. The primers of Trx1 and Trx2 were designed according to the sequences of GenBank. The Trx1 and Trx2 were respectively amplified by PCR (polymerase chain reaction) and were sequenced by the bioinformatics method. The sequences were compared with those of the international standard HP strains. RESULTS: The whole sequence of Trx1 and the first 295 bp of Trx2 were successfully amplified to allow for sequence comparison by BioEdit. The Trx1 from different HP strains contained 321 bp encoding 106 amino acids. The mutation ratio was 13.4% (43/321). All amino acids contained the same active motif Cys-Gly-Pro-Cys. The homology of Trx1 amino acids from 25 strains was 97.2% (103/106). The mutation ratio for first 295 bp of Trx2 was 18.6% (55/295). All amino acids encoded by Trx2 contained one CXXC zone while the Cys-Gly-Pro-Cys motif was not found. The homology of Trx2 amino acids was 84.7% (83/98). CONCLUSION: Two different subtypes of Trx from clinically isolated Hp strains have mutation sites. But the homology of encoded amino acids is relatively high because of invalid mutations. Trx1 of HP strains from different diseases all contains a redox active motif Cys-Gly-Pro-Cys while Trx2 contains only a CXXC zone. The above results will provide valuable rationales for future studies of the biological function and pathogenic mechanisms of HP Trx1 and Trx2.

T3098C and T53C mutations of HBV genotype C is associated with HBV infection progress.

OBJECTIVE: To analyze the association between mutation(s) in preS region of HBV and hepatitis B disease progress in Chinese patients with genotype C chronic HBV infection. METHODS: Ninety-three patients with chronic genotype C HBV infection, including 24 asymptomatic carriers (ASC), 26 patients with chronic hepatitis B (CHB), 22 patients with liver cirrhosis (LC) and 21 HCC patients were investigated. Levels of HBV DNA, HBeAg, alanine aminotransferase (ALT), asparate transaminase (AST) were measured. HBV preS region was analyzed by PCR direct sequencing. RESULTS: The prevalence of preS T3098C and T53C mutations of genotype C HBV was significantly higher in LC and HCC patients than ASC and CHB patients. The rate of T3098C mutation in ASC, CHB, LC, and HCC patients were 0.00% (0/24), 3.85% (1/26), 9.09% (2/22), and 30.77% (8/22), respectively (P=0.0015), while the rate of T53C mutation was 12.50% (3/24), 3.85% (1/26), 40.91% (9/22), and 42.31% (11/26), respectively (P=0.0012). CONCLUSION: The frequency of genotype C HBV preS T3098C and T53C mutations is associated with hepatitis B infection progression

Puerarin inhibits apoptosis and inflammation in myocardial cells via PPARα expression in rats with chronic heart failure.

Chronic heart failure affects myocardial energy metabolism and cardiac function. Puerarin has been reported to improve cardiac function through regulation of energy metabolism in mice with myocardial infarction. The aim of the current study was to determine whether puerarin can improve body weight and reduce inflammation and apoptosis in rats with chronic heart failure. Rats were divided into three groups: Puerarin, PBS and sham group. Transverse aortic constriction was performed to induce chronic heart failure in the puerarin an PBS groups. Cardiac function, apoptosis and inflammation were evaluated following a 4-week treatment in rats with chronic heart failure. The results demonstrated that puerarin significantly increased the survival rate of rats and improved cardiac function compared with the PBS group. In addition, puerarin decreased lactate dehydrogenase and succinate dehydrogenase activity compared with the PBS group. Puerarin treatment increased the expression levels of glucose transporter type 4 and decreased the expression levels of CD36. Additionally, puerarin decreased the levels inflammatory factors, including tumor necrosis factor α, interleukin (IL)-1ß and IL-6 in serum and myocardial tissue compared with the PBS group. Puerarin upregulated peroxisome proliferator-activated receptor α (PPARα) and its downstream target genes nuclear respiratory factor 1, FOS proto-oncogene, YY1 transcription factor, acetyl-coenzyme A carboxylase a, Fas cell surface death receptor, L-type pyruvate kinase and acetyl-coenzyme A dehydrogenase medium chain in myocardial cells from rats with chronic heart failure. These results demonstrated that puerarin inhibited apoptosis and inflammation in myocardial cells via the PPARα pathway. In conclusion, the present study indicated that puerarin may exhibit antiapoptotic and anti-inflammatory activity through the PPARα pathway in rats with chronic heart failure.

The type VI secretion system protein AsaA in Acinetobacter baumannii is a periplasmic protein physically interacting with TssM and required for T6SS assembly.

Type VI secretion system (T6SS) is described as a macromolecular secretion machine that is utilized for bacterial competition. The gene clusters encoding T6SS are composed of core tss genes and tag genes. However, the clusters differ greatly in different pathogens due to the great changes accumulated during the long-term evolution. In this work, we identified a novel hypothetical periplasmic protein designated as AsaA which is encoded by the first gene of the T6SS cluster in the genus Acinetobacter. By constructing asaA mutant, we delineated its relative contributions to bacterial competition and secretion of T6SS effector Hcp. Subsequently, we studied the localization of AsaA and potential proteins that may have interactions with AsaA. Our results showed that AsaA in Acinetobacter baumannii (A. baumannii) localized in the bacterial periplasmic space. Results based on bacterial two-hybrid system and protein pull-down assays indicated that it was most likely to affect the assembly or stability of T6SS by interacting with the T6SS core protein TssM. Collectively, our findings of AsaA is most likely a key step in understanding of the T6SS functions in A. baumannii.

Mutations in preS genes of genotype C hepatitis B virus in patients with chronic hepatitis B and hepatocellular carcinoma.

BACKGROUND: Hepatitis B virus (HBV) preS mutations are frequently isolated from patients with severe forms of liver disease. Meanwhile, genotype C has been shown to cause more serious liver disease than genotype B. This study assesses the frequency of preS mutation in Chinese patients with genotype C chronic HBV infection and its relation to liver damage. METHODS: Seventy-nine persistently infected patients (25 asymptomatic carriers, 28 with chronic hepatitis, and 26 with hepatocellular carcinoma) with genotype C HBV were analyzed. Levels of HBV DNA, hepatitis B e antigen (HBeAg), alanine aminotransferase, and aspartate transaminase and mutations in the preS region were determined. RESULTS: The correlations of preS deletion with disease progression were distinct: preS deletion mutations were more commonly found in the hepatocellular carcinoma (HCC) group than in the chronic hepatitis B (CHB) or asymptomatic carrier (ASC) groups, with the frequencies of 38.46% (10/26) in the HCC, 7.14% (2/28) in the CHB, and 4.00% (1/25) in the ASC (P = 0.001) groups. The HBeAg-positive rate and HBV DNA levels were comparable between patients with the preS mutation and those without. CONCLUSIONS: PreS deletion mutations of genotype C HBV might play a role in HBV-related hepatocarcinogenesis.