Hepatitis E： From behind the scenes to the front stage / 临床肝胆病杂志

‍Hepatitis E is a type of viral hepatitis caused by hepatitis E virus （HEV） infection. There are four main genotypes of HEV， namely genotypes 1‍ — ‍4. Compared with hepatitis B and hepatitis C， previous research on hepatitis E has made slow progress； however， in recent years， hepatitis E has gradually attracted more attention in China， achieving the transition from behind the scenes to the front stage. First， China has a large population base， a large number of people infected with hepatitis E， and a heavy disease burden， and thus it is necessary to attach great importance to the prevention， diagnosis， and treatment of hepatitis E. Second， the World Health Organization （WHO） has put forward the ambitious goal of eliminating viral hepatitis by 2030， and China has made remarkable achievements in the prevention and treatment of hepatitis B and hepatitis C. At present， the incidence and mortality rates of hepatitis E in China have exceeded those of hepatitis A， and the prevention and treatment of hepatitis E in China is of great significance to actively respond to the call of the WHO. Finally， Chinese researchers in the field of hepatitis E have formed Chinese Consortium for the Study of Hepatitis E， formulated related expert consensus， and established international cooperation platforms， which further promotes the research on hepatitis E in China.

Targeting TRMT5 suppresses hepatocellular carcinoma progression via inhibiting the HIF-1α pathways / 浙江大学学报（英文版）（B辑：生物医学和生物技术）

Accumulating evidence has confirmed the links between transfer RNA (tRNA) modifications and tumor progression. The present study is the first to explore the role of tRNA methyltransferase 5 (TRMT5), which catalyzes the m1G37 modification of mitochondrial tRNAs in hepatocellular carcinoma (HCC) progression. Here, based on bioinformatics and clinical analyses, we identified that TRMT5 expression was upregulated in HCC, which correlated with poor prognosis. Silencing TRMT5 attenuated HCC proliferation and metastasis both in vivo and in vitro, which may be partially explained by declined extracellular acidification rate (ECAR) and oxygen consumption rate (OCR). Mechanistically, we discovered that knockdown of TRMT5 inactivated the hypoxia-inducible factor-1 (HIF-1) signaling pathway by preventing HIF-1α stability through the enhancement of cellular oxygen content. Moreover, our data indicated that inhibition of TRMT5 sensitized HCC to doxorubicin by adjusting HIF-‍1α. In conclusion, our study revealed that targeting TRMT5 could inhibit HCC progression and increase the susceptibility of tumor cells to chemotherapy drugs. Thus, TRMT5 might be a carcinogenesis candidate gene that could serve as a potential target for HCC therapy.

Prognostic value of serum free triiodothyronine in patients with hepatitis E-related acute liver failure / 中华临床感染病杂志

Objective:To investigate the prognostic value of serum free triiodothyronine (FT3) in patients with hepatitis E-related acute liver failure (HEV-ALF).Methods:Clinical data of 88 patients with HEV-ALF and 86 patients with acute hepatitis E (AHE) were collected from the member hospitals of Chinese Consortium for the Study of Hepatitis E between January 2016 and December 2021; the data of 100 health subjects who underwent health check-up in Suzhou Municipal Hospital were also collected as healthy control (HC) group. Serum FT3 levels were analyzed in all subjects. HEV-ALF patients were divided into survival group ( n=73) and death group ( n=15) according to their 30 day survival. Correlation between serum FT3 level and prognosis of HEV-ALF patients were analyzed by Cox regression and orthogonal partial least squares discriminant analysis (OPLS-DA). The receiver operating characteristic (ROC) curve and decision curve analysis (DCA) were used to assess the predictive value of serum FT3 levels for predicting the prognosis of patients, and its prediction efficacy was compared with conventional Model for End-Stage Liver Disease (MELD), King’s College Hospital criteria (KCH) and Child-Pugh models. Results:The levels of serum FT3 in HEV-ALF patients were significantly lower than those in AHE patients and HC group ( P=0.006 or <0.001). Cox regression analysis showed that international standardized ratio ( HR=17.984, 95% CI 2.804-115.362), hepatic encephalopathy ( HR=12.895, 95% CI 2.386-69.695) and total cholesterol ( HR=2.448, 95% CI 1.108-5.409) were independent risk factors for death in HEV-ALF patients, and serum FT3 level ( HR=0.323, 95% CI 0.119-0.876) was a protective factor. OPLS-DA results showed serum FT3 levels had high predictive value. ROC curve analysis results showed that the area under the curve was 0.828 (95% CI 0.733-0.900, P<0.001), the sensitivity was 80.00%, and the specificity was 78.08%. DCA showed that FT3 has good prediction ability and decision-making level serum FT3 levels in patients with improvement and fluctuation were significantly higher than those in the patients with deterioration ( P<0.05 or <0.01). Conclusion:Serum FT3 levels are closely related to the prognosis of HEV-ALF patients and it may be used as a biomarker for the prognosis of patients with HEV-ALF.

Comparison of Sonazoid contrast-enhanced ultrasound and contrast-enhanced MRI in the diagnosis of focal liver lesions / 中华超声影像学杂志

Objective:To compare the diagnostic efficacies of Sonazoid contrast-enhanced ultrasound (CEUS) and contrast-enhanced magnetic resonance imaging (CE-MRI) in the diagnosis of focal liver lesions (FLLs), and to evaluate the clinical value of Sonazoid.Methods:A total of 58 FLLs in 50 patients who underwent Sonazoid-CEUS and CE-MRI examinations from July 2019 to January 2021 in the First Affiliated Hospital of Zhejiang University School of Medicine were enrolled in this study according to the inclusion criteria. The final diagnostic reference standard was decided by surgical pathology or ultrasound-guided biopsy pathology. Sonazoid-CEUS and CE-MRI features of benign and malignant FLLs were analyzed, and the sensitivity, specificity, positive predictive value, negative predictive value, and diagnostic coincidence rate of the two tests were calculated respectively.Results:There was a statistically significant difference between benign and malignant FLLs in the imaging pattern of homogeneous or heterogeneous intratumoral enhancement in the artery-dominant phase and washout images in the late phase( P<0.001).9.8%(4/41) of the malignant lesions did not decrease until the late phase but decreased in the post-vascular phase. The sensitivity, specificity, positive predictive value, negative predictive value, and diagnostic coincidence rate of the two tests were 97.6%, 52.9%, 83.3%, 90.0%, 84.5%(Sonazoid-CEUS) and 85.4%, 64.7%, 85.4%, 64.7%, 79.3%(CE-MRI), the differences of sensitivity and specificity were not statistically significant ( P=0.125, P=0.687). Conclusions:The vascular phase in Sonazoid-CEUS is still an important diagnostic sign of FLLs, and the unique Kupffer phase can provide additional information for the diagnosis. Sonazoid-CEUS has the same important value as CE-MRI in the diagnosis of FLLs.

Clinical features and risk factors of invasive pulmonary aspergillosis in patients with chronic obstructive pulmonary disease / 中华临床感染病杂志

Objective:To analyze the clinical features and risk factors of invasive pulmonary aspergillosis (IPA) in patients with chronic obstructive pulmonary disease (COPD).Methods:Ninety COPD patients with IPA admitted in the First Affiliated Hospital, Zhejiang University School of Medicine and Zhejiang Provincial People’s Hospital from January 2015 to September 2019 were enrolled, and 180 COPD patients without IPA admitted in the same period were selected as control group. The clinical data of the patients in both groups were analyzed retrospectively. Chi square test was used to compare the imaging characteristics of patients in two groups, and multivariate conditional Logistic regression analysis was used to explore the risk factors of IPA in COPD patients. Results:Among 90 cases of COPD with IPA, the culture of lower respiratory tract samples identified Aspergillus fumigatus in 78 cases, Aspergillus flavus in 6 cases, Aspergillus fumigatus with Aspergillus flavus in 1 case, Aspergillu sterrus in 1 case, and Aspergillus nigerwere in 1 case; 1 case of Aspergillus mycelium was found by sputum exfoliation cytology and 2 cases were positive for serum galactomannan. Chest CT images showed patchy infiltrating shadow (87.8%), pleural effusion (36.7%), nodules (33.3%), cavity (18.9%), consolidation shadow (17.8%), halo sign (14.4%) and air crescent sign (2.2%). The incidence of patchy infiltrating shadow, consolidation shadow, halo sign and cavity were higher in COPD patients with IPA compared to control group ( P<0.05 or P<0.01). Multivariate conditional Logistic regression analysis showed that prolonged hospital stay ( OR=1.183, 95% CI 1.047-1.336), combination of two antibiotics ( OR=5.391, 95% CI 1.241-23.421), duration of antibiotic treatment ≥14 days ( OR=5.275, 95% CI 1.586-17.541), cumulative use of antibiotics ( OR=2.270, 95% CI 1.406-3.664) were the risk factors of COPD with IPA ( P<0.05 or P<0.01). Conclusion:The risk factors of IPA in COPD patients include long duration of hospital stay, combination of two kinds of antibiotics, more than 14 days of antibiotic treatment, and more varieties of antibiotics. If the above risk factors exist in patients with COPD, etiology and serology examination and dynamic monitoring of chest CT scan should be performed for early diagnosis and improved prognosis.

Study on the serotype, drug sensitivity and virulence genes of clinical isolates of Salmonella in Zhejiang Province / 中华传染病杂志

Objective:To investigate the serotype distribution, drug resistance and the virulence gene profiles of clinical isolates of Salmonella in Zhejiang Province. Methods:A retrospective study of 463 clinical isolates of Salmonella in nine regions of Zhejiang Province from 2010 to 2017 was conducted. Their serotypes were detected using the Kauffmann-White scheme based on serological detection. Antibiotics susceptibility tests were carried out using K-B disk diffusion method. Eleven virulence genes were detected by polymerase chain reaction. Results:A total of 35 serotypes were identified among the 463 strains of Salmonella. The dominant serotypes were Salmonella typhimurium (41.90%(194/463)) and Salmonella enteritis (22.25%(103/463)). The resistance rate of Salmonella to ampicillin was the highest (66.52%(308/463)), followed by ampicillin/sulbactam (46.87%(217/463)), while low resistance rate to piperacillin/tazobactam was observed (3.24%(15/463)). All strains were sensitive to imipenem and meropenem. Finally, 188 strains (40.60%) of multi-drug resistance were found. The virulence genes hilA, ssrB, marT, siiD, sopB and pagN were found in all Salmonella strains. The virulence gene vexA was found only in Salmonella typhi and Salmonella Dublin; virulence gene icmF was mainly found in Salmonella enteritis. The virulence plasmid gene spvB and pefA were mainly found in Salmonella enteritidis and Salmonella typhimurium and invariably appeared in pairs. The virulence gene cdtB was mainly found in Salmonella typhi and Salmonella paratyphi A. Conclusions:Salmonella typhimurium and Salmonella enteritis are the main clinically isolated Salmonella strains in Zhejiang Province. The situation of multi-drug resistance is severe and a variety of virulence genes are highly detected.

Pregnant women complicated with COVID-19: a clinical analysis of 3 cases / 浙江大学学报·医学版

OBJECTIVE@#To analyze the clinical characteristics and pregnancy outcomes of pregnant women complicated with coronavirus disease 2019 (COVID-19).@*METHODS@#The clinical data of 3 pregnant women with COVID-19 admitted to the First Affiliated Hospital of Zhejiang University School of Medicine from January 19 to February 10, 2020 were retrospectively analyzed.@*RESULTS@#There was one case in the first-trimester pregnancy (case 1), one in the second-trimester pregnancy (case 2) and one in third-trimester pregnancy (case 3). Cough, fever, fatigue, lung imaging changes were the main manifestations. The white cell count, lymphocyte percentage had no significantly changes in case 1 and case 3, while the levels of C-reactive protein (CRP), erythrocyte sedimentation rate (ESR), IL-6 and IL-10 elevated. The lymphocyte count and lymphocyte percentage decreased and the inflammatory indicators significantly increased in case 2. All patients were treated with antiviral, antitussive, oxygen inhalation; case 3 received glucocorticoids, case 2 with severe illness received glucocorticoids and additionally gamma globulin. All three cases were cured and discharged. Case 1 with early pregnancy chose to terminate pregnancy after discharge; case 2 chose to continue pregnancy without obstetric complications; and case 3 had cesarean section delivery due to abnormal fetal heart monitoring.@*CONCLUSIONS@#The report shows that COVID-19 in pregnancy women could be cured with active treatment, and the maternal and fetal outcomes can be satisfactory.

Effect of shift rotation culture on formation and activity of encapsulated hepatocytes aggregates / 浙江大学学报·医学版

To observe the effect of uniform and shift rotation culture on the formation and activity of the alginate-chitosan (AC) microencapsulated HepLL immortalized human hepatocytes and HepG2 cells aggregates.AC microcapsulated HepG2 and HepLL cells were randomly divided into two groups. Each group was divided into 3 subgroups according to uniform and shift rotation culture.The size and number of aggregates were observed and measured under laser confocal microscopy and inverted microscope dynamically. The amount of albumin synthesis was detected by ELISA, the clearance of ammonia was detected by colorimetry, and diazepam conversion function was detected by high performance liquid chromatography (HPLC).On day 6, 8, 10, 12, 14 and 16, the number and size of the aggregates, albumin synthesis, diazepam clearance and ammonium clearance increased significantly in shift rotation culture group than in uniform group (all<0.01). The albumin synthesis, diazepam clearance, and ammonium clearance in the microencapsulated HepLL groups were significantly higher than those of HepG2 cells at any time (all<0.01).Shift rotation culture can significantly promote the formation and increase the activity of AC microencapsulated HepLL and HepG2 aggregates, and HepLL cells may be more suitable for bioartificial liver than HepG2.

Establishment and further identification of immortalization human hepatocyte lines / 中华传染病杂志

Objective To establish immortalized human hepatocyte lines for studies of bioartificial liver,hepatocyte transplantation,and drug metabolism in vitro.Methods Primary human hepatocytes were isolated by 4-step perfusion technique with collagenase and transfected with recombinant retrovirus containing Simian virus 40 large T antigen(SV40 LT).Subsequently,immortalized human hepatocytes were evaluated by analysis of gene expression and functional characteristics in vitro.Results Two immortalized human hepatocyte lines,HepLi2 and HepLi3,were obtained after primary human hepatocytes being infected by SV40 LT containing recombinant retrovirus for 3-4 weeks.The immortalized human hepatocytes showed classical appearance of hepatocyte observed by phase contrast microscope.The protein expression of SV40 LT in HepLi2 and HepLi3 cells were detected by Western blotting.The mRNA expressions of albumin(Alb),glutathione S-transferase(GST-p),human blood coagulation factor X(HBCF-X)and β-actin in HepLi2 and HepLi3 cells were detected by reverse transcription polymerase chain reaction(RT-PCR),and the mRNA expressions of cytochrome(CY)450 subtypes(CYP3A5,CYP2E1,CYP2C8-19 and CYP3A4)in HepLi2 and HepLi3 cells were also observed by RT-PCR.Levels of alanine transaminase (ALT),lactate dehydrogenase(LDH)and Alb were detected in the supernatant of immortalized human hepatoeyte culture.Conclusions The immortalized human hepatocyte lines have the biological characteristics of primary human hepatocytes and have the CYP450 functions of hepatocytes,which may be heIDful for the studies of bioartificial liver,heoatocvte transplantation and drug metabolism in vitro.

Immortalization of porcine hepatocyte lines by transfection with simian virus 40 large T antigen and human telomerase reverse transcriptase / 中华传染病杂志

Objective To establish normally immortalized porcine hepatocyte lines by ectopic expression of simian virus 40 large T (SV40LT) antigen and the human telomerase reverse transcriptase(hTERT). Methods Primary porcine Hepatoeyte cells were transfeeted with recombinant retrovirus containing SV40LT or hTERT respectively. Subsequently drug resistant cell clones were screened and expanded for further studies. Immortalized porcine hepatocyte was confirmed by examination. Results The morphological phenotype of the transfected cells was similar to the primary porcine hepatocyte. One clone, HepLP, has been maintained in cultue for half year, and expanded by more than 60 passages. SV40 LT and hTERT could be detected in transfected porcine hepatocyte. Pig albumin mRNA was also detected by RT-PCR. No tumor formation occurred when HepLP cells were injected into Balb/c nude mice. Conclusions The immortalized, nontumorigenic, porcine hepatoeytes maintained the properties of porcine primary hepatocytes such as the albumin secretion. This generation of immortalized porcine hepatocyte may be helpful for bioartifical liver support system, hepatocytes transplantation, drug/toxicological studies, and liver biologic studies.

Efficacy of adefovir dipivoxil in treatment of chronic hepatitis B patients with lamivudine resistance / 中华临床感染病杂志

Objective To investigate the efficacy and safety of adefovir dipivoxil (ADV) in treatment of chronic hepatitis B (CHB) patients with lamivudine (LAM) resistance. Methods There were treatment group (32 CHB patients with LAM resistance) and historical control group (24 CHB patients with LAM resistance) in this study. The treatment group received ADV 10 mg/d and LAM 100 mg/d for 48 weeks; the historical control group continued to use LAM monotherapy. During the treatment causes, serum HBV DNA levels, liver function and HBV serology were monitored regularly, and safety assessments were also conducted. Results In treatment group, mean HBV DNA levels decreased by 2.56 log10 eopies/ml and 2.93 log10 copies/ml, virus response rates were 50. 0% and 75.0%, ALT normalization rates were 53.1% and 68.8% after 24 and 48 weeks of treatment, respectively. The histological improvement rate was 65.6% after 48 weeks. Comparing with those in control group, the differences were statistically significant ( P <0. 05), while there was no significant statistical differences in HBeAg loss rate and HBeAg seroconversion rate between two groups. There was no severe adverse event during the treatment. Conclusion ADV is effective and safe in treatment of lamivudine-resistant CHB.

Establishment of a hepatocyte line from human with SV40LT antigen / 中华传染病杂志

Objective To establish a hepatocyte line from human for bioartificial liver support system and hepatocyte transplantation. Methods Hepatocytes isolated from donor liver of a 25 year old brain death male were transfected with pcDNA3.1(-) recombined vector containing the genes encoding Simian Virus 40 large T antigen. Characteristics of the immortalized hepatocyte line were evaluated by morphologic and functional methods. Results One of the hepatocyte clones displayed highly differentiated liver function and immortalized characteristics has been screened after 42 day's selection of 700～300 ?g/ml G418. The immortalized hepatocyte appeared epithelial and displayed morphologic characteristics of liver parenchymal cells. ALB, AST, ALT and LDH secreted from hepatocyte line were detected. Transmission electron microscope examination revealed that immortalized cells contained a round nucleus with nucleoli, moderate numbers of mitochondria, numerous rough endoplasmic reticulums, and some vacuoles. Adjacent cells were joined with junctional complexes. RT PCR and Western blot detection revealed that immortalized hepatocyte could express ALB mRNA and showed immunoreactive of cytochrome P450 2E1. Conclusions The new established hepatocyte line from human retained some specificbiologic functions and morphological cha racteristics of primary cultured hepatocytes.