Genomic Epidemiology of Imported Cases of COVID-19 in Guangdong Province, China, October 2020 - May 2021.

Objective: The pandemic of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has been engendering enormous hazards to the world. We obtained the complete genome sequences of SARS-CoV-2 from imported cases admitted to the Guangzhou Eighth People's Hospital, which was appointed by the Guangdong provincial government to treat coronavirus disease 2019 (COVID-19). The SARS-CoV-2 diversity was analyzed, and the mutation characteristics, time, and regional trend of variant emergence were evaluated. Methods: In total, 177 throat swab samples were obtained from COVID-19 patients (from October 2020 to May 2021). High-throughput sequencing technology was used to detect the viral sequences of patients infected with SARS-CoV-2. Phylogenetic and molecular evolutionary analyses were used to evaluate the mutation characteristics and the time and regional trends of variants. Results: We observed that the imported cases mainly occurred after January 2021, peaking in May 2021, with the highest proportion observed from cases originating from the United States. The main lineages were found in Europe, Africa, and North America, and B.1.1.7 and B.1.351 were the two major sublineages. Sublineage B.1.618 was the Asian lineage (Indian) found in this study, and B.1.1.228 was not included in the lineage list of the Pangolin web. A reasonably high homology was observed among all samples. The total frequency of mutations showed that the open reading frame 1a (ORF1a) protein had the highest mutation density at the nucleotide level, and the D614G mutation in the spike protein was the commonest at the amino acid level. Most importantly, we identified some amino acid mutations in positions S, ORF7b, and ORF9b, and they have neither been reported on the Global Initiative of Sharing All Influenza Data nor published in PubMed among all missense mutations. Conclusion: These results suggested the diversity of lineages and sublineages and the high homology at the amino acid level among imported cases infected with SARS-CoV-2 in Guangdong Province, China.

Efficacy and safety of AnluoHuaxian pills on chronic hepatitis B with normal or minimally elevated alanine transaminase and early liver fibrosis: A randomized controlled trial.

ETHNOPHARMACOLOGICAL RELEVANCE: The AnluoHuaxian pill (AHP) is a widely used patented medicine for chronic hepatitis B (CHB) patients with advanced fibrosis or cirrhosis that has been used in China for more than 15 years. However, data are lacking on whether monotherapy with AHP can be effective in CHB patients with alanine aminotransferase (ALT) levels less than 2 times the upper limit of normal (ALT<2ULN) and early liver fibrosis (F ≤ 2). AIM OF THE STUDY: We aimed to investigate whether monotherapy with AHP improves liver histology in these patients. MATERIALS AND METHODS: In this double-blind, randomized, placebo-controlled trial, 270 CHB patients with ALT<2ULN and F ≤ 2 were treated in 12 hospitals in China. The patients were randomly assigned to an intervention (AHP) group and a placebo group at a ratio of 2:1. Of these 270 enrolled patients, 147 had paired liver biopsies. The primary end point was histological change after 48 weeks of treatment. RESULTS: Per-protocol analysis revealed that the rate of histologic improvement in liver fibrosis patients in the AHP group was significantly higher than that in the placebo group (37.7% vs. 19.5%, P = 0.035) after 48 weeks of treatment, which was consistent with results from intention-to-treat and sensitivity analyses. Moreover, after adjusting for baseline characteristics, AHP was superior to placebo with respect to improving liver fibrosis (odds ratio [OR] = 2.58, 95% confidence interval [CI]: (1.01, 6.63),P = 0.049) and liver histology (OR = 3.62, 95% CI: (1.42, 9.20),P = 0.007). In noninvasive measurement of liver fibrosis (FibroScan®), the level of liver stiffness measurement (LSM) had decreased significantly at 48 weeks (5.1 kPa) compared with that at baseline (5.7 kPa) (P = 0.008) in the AHP group, whereas it did not decrease significantly in the placebo group. Cirrhosis developed in one patient in the placebo group but in no patients in the AHP group. No serious side effects occurred in the AHP-treated patients. CONCLUSIONS: Treatment of CHB patients who had ALT<2ULN and F ≤ 2 with the traditional Chinese medicine AHP for 48 weeks improves liver fibrosis. However, due to the short duration of treatment and the limited sample size of liver pathology, the long-term benefits of AHP in reducing fibrosis and the risk of cirrhosis and hepatocellular carcinoma in these patients need to be further studied in the future.

Effect of Recombinant Human Granulocyte Colony-Stimulating Factor for Patients With Coronavirus Disease 2019 (COVID-19) and Lymphopenia: A Randomized Clinical Trial.

Importance: Lymphopenia is common and correlates with poor clinical outcomes in patients with coronavirus disease 2019 (COVID-19). Objective: To determine whether a therapy that increases peripheral blood leukocyte and lymphocyte cell counts leads to clinical improvement in patients with COVID-19. Design, Setting and Participants: Between February 18 and April 10, 2020, we conducted an open-label, multicenter, randomized clinical trial at 3 participating centers in China. The main eligibility criteria were pneumonia, a blood lymphocyte cell count of 800 per µL (to convert to ×109/L, multiply by 0.001) or lower, and no comorbidities. Severe acute respiratory syndrome coronavirus 2 infection was confirmed with reverse-transcription polymerase chain reaction testing. Exposures: Usual care alone, or usual care plus 3 doses of recombinant human granulocyte colony-stimulating factor (rhG-CSF, 5 µg/kg, subcutaneously at days 0-2). Main Outcomes and Measures: The primary end point was the time from randomization to improvement of at least 1 point on a 7-category disease severity score. Results: Of 200 participants, 112 (56%) were men and the median (interquartile range [IQR]) age was 45 (40-55) years. There was random assignment of 100 patients (50%) to the rhG-CSF group and 100 (50%) to the usual care group. Time to clinical improvement was similar between groups (rhG-CSF group median of 12 days (IQR, 10-16 days) vs usual care group median of 13 days (IQR, 11-17 days); hazard ratio, 1.28; 95% CI, 0.95-1.71; P = .06). For secondary end points, the proportion of patients progressing to acute respiratory distress syndrome, sepsis, or septic shock was lower in the rhG-CSF group (rhG-CSF group, 2% vs usual care group, 15%; difference, -13%; 95%CI, -21.4% to -5.4%). At 21 days, 2 patients (2%) had died in the rhG-CSF group compared with 10 patients (10%) in the usual care group (hazard ratio, 0.19; 95%CI, 0.04-0.88). At day 5, the lymphocyte cell count was higher in the rhG-CSF group (rhG-CSF group median of 1050/µL vs usual care group median of 620/µL; Hodges-Lehmann estimate of the difference in medians, 440; 95% CI, 380-490). Serious adverse events, such as sepsis or septic shock, respiratory failure, and acute respiratory distress syndrome, occurred in 29 patients (14.5%) in the rhG-CSF group and 42 patients (21%) in the usual care group. Conclusion and Relevance: In preliminary findings from a randomized clinical trial, rhG-CSF treatment for patients with COVID-19 with lymphopenia but no comorbidities did not accelerate clinical improvement, but the number of patients developing critical illness or dying may have been reduced. Larger studies that include a broader range of patients with COVID-19 should be conducted. Trial Registration: Chinese Clinical Trial Registry: ChiCTR2000030007.

Clinical characteristics and outcomes of hospitalised patients with COVID-19 treated in Hubei (epicentre) and outside Hubei (non-epicentre): a nationwide analysis of China.

BACKGROUND: During the outbreak of coronavirus disease 2019 (COVID-19), consistent and considerable differences in disease severity and mortality rate of patients treated in Hubei province compared to those in other parts of China have been observed. We sought to compare the clinical characteristics and outcomes of patients being treated inside and outside Hubei province, and explore the factors underlying these differences. METHODS: Collaborating with the National Health Commission, we established a retrospective cohort to study hospitalised COVID-19 cases in China. Clinical characteristics, the rate of severe events and deaths, and the time to critical illness (invasive ventilation or intensive care unit admission or death) were compared between patients within and outside Hubei. The impact of Wuhan-related exposure (a presumed key factor that drove the severe situation in Hubei, as Wuhan is the epicentre as well the administrative centre of Hubei province) and the duration between symptom onset and admission on prognosis were also determined. RESULTS: At the data cut-off (31 January 2020), 1590 cases from 575 hospitals in 31 provincial administrative regions were collected (core cohort). The overall rate of severe cases and mortality was 16.0% and 3.2%, respectively. Patients in Hubei (predominantly with Wuhan-related exposure, 597 (92.3%) out of 647) were older (mean age 49.7 versus 44.9 years), had more cases with comorbidity (32.9% versus 19.7%), higher symptomatic burden, abnormal radiologic manifestations and, especially, a longer waiting time between symptom onset and admission (5.7 versus 4.5 days) compared with patients outside Hubei. Patients in Hubei (severe event rate 23.0% versus 11.1%, death rate 7.3% versus 0.3%, HR (95% CI) for critical illness 1.59 (1.05-2.41)) have a poorer prognosis compared with patients outside Hubei after adjusting for age and comorbidity. However, among patients outside Hubei, the duration from symptom onset to hospitalisation (mean 4.4 versus 4.7 days) and prognosis (HR (95%) 0.84 (0.40-1.80)) were similar between patients with or without Wuhan-related exposure. In the overall population, the waiting time, but neither treated in Hubei nor Wuhan-related exposure, remained an independent prognostic factor (HR (95%) 1.05 (1.01-1.08)). CONCLUSION: There were more severe cases and poorer outcomes for COVID-19 patients treated in Hubei, which might be attributed to the prolonged duration of symptom onset to hospitalisation in the epicentre. Future studies to determine the reason for delaying hospitalisation are warranted.

Comorbidity and its impact on 1590 patients with COVID-19 in China: a nationwide analysis.

BACKGROUND: The coronavirus disease 2019 (COVID-19) outbreak is evolving rapidly worldwide. OBJECTIVE: To evaluate the risk of serious adverse outcomes in patients with COVID-19 by stratifying the comorbidity status. METHODS: We analysed data from 1590 laboratory confirmed hospitalised patients from 575 hospitals in 31 provinces/autonomous regions/provincial municipalities across mainland China between 11 December 2019 and 31 January 2020. We analysed the composite end-points, which consisted of admission to an intensive care unit, invasive ventilation or death. The risk of reaching the composite end-points was compared according to the presence and number of comorbidities. RESULTS: The mean age was 48.9 years and 686 (42.7%) patients were female. Severe cases accounted for 16.0% of the study population. 131 (8.2%) patients reached the composite end-points. 399 (25.1%) reported having at least one comorbidity. The most prevalent comorbidity was hypertension (16.9%), followed by diabetes (8.2%). 130 (8.2%) patients reported having two or more comorbidities. After adjusting for age and smoking status, COPD (HR (95% CI) 2.681 (1.424-5.048)), diabetes (1.59 (1.03-2.45)), hypertension (1.58 (1.07-2.32)) and malignancy (3.50 (1.60-7.64)) were risk factors of reaching the composite end-points. The hazard ratio (95% CI) was 1.79 (1.16-2.77) among patients with at least one comorbidity and 2.59 (1.61-4.17) among patients with two or more comorbidities. CONCLUSION: Among laboratory confirmed cases of COVID-19, patients with any comorbidity yielded poorer clinical outcomes than those without. A greater number of comorbidities also correlated with poorer clinical outcomes.

Clinical Characteristics of Coronavirus Disease 2019 in China.

BACKGROUND: Since December 2019, when coronavirus disease 2019 (Covid-19) emerged in Wuhan city and rapidly spread throughout China, data have been needed on the clinical characteristics of the affected patients. METHODS: We extracted data regarding 1099 patients with laboratory-confirmed Covid-19 from 552 hospitals in 30 provinces, autonomous regions, and municipalities in mainland China through January 29, 2020. The primary composite end point was admission to an intensive care unit (ICU), the use of mechanical ventilation, or death. RESULTS: The median age of the patients was 47 years; 41.9% of the patients were female. The primary composite end point occurred in 67 patients (6.1%), including 5.0% who were admitted to the ICU, 2.3% who underwent invasive mechanical ventilation, and 1.4% who died. Only 1.9% of the patients had a history of direct contact with wildlife. Among nonresidents of Wuhan, 72.3% had contact with residents of Wuhan, including 31.3% who had visited the city. The most common symptoms were fever (43.8% on admission and 88.7% during hospitalization) and cough (67.8%). Diarrhea was uncommon (3.8%). The median incubation period was 4 days (interquartile range, 2 to 7). On admission, ground-glass opacity was the most common radiologic finding on chest computed tomography (CT) (56.4%). No radiographic or CT abnormality was found in 157 of 877 patients (17.9%) with nonsevere disease and in 5 of 173 patients (2.9%) with severe disease. Lymphocytopenia was present in 83.2% of the patients on admission. CONCLUSIONS: During the first 2 months of the current outbreak, Covid-19 spread rapidly throughout China and caused varying degrees of illness. Patients often presented without fever, and many did not have abnormal radiologic findings. (Funded by the National Health Commission of China and others.).

Anesthetic agent etiomidate induces apoptosis in N2a brain tumor cell line.

The present study identified the cytotoxic effects of etomidate on the N2a neuroblastoma cell line. Etomidate induced apoptosis in N2a cells in a concentration­dependent manner, which was confirmed by western blotting and flow cytometry. Phase contrast microscopy was used to analyze the effect of etomidate on morphological characteristics. The number of the apoptotic cells was increased and confirmed by DAPI and PI staining, which served as a characteristic hallmark of apoptosis. Additionally, etomidate led to loss of mitochondrial membrane potential and resulted in the generation of reactive oxygen species in N2a cells. The western blot analysis revealed that N2a cells treated with etomidate had a significant modulation of pro­apoptotic proteins, includingpoly ADP­ribose polymerase (PARP), cleaved PARP, caspase­9 and procaspase­3. In conclusion, the present study determined that etomidate induced cytotoxic and apoptotic effects in N2a brain tumor cells in vitro.

IFIT1 polymorphisms predict interferon-α treatment efficiency for hepatitis B virus infection.

AIM: To investigate the association between interferon-induced protein with tetratricopeptide repeats 1 (IFIT1) polymorphisms and interferon-α (IFNα) treatment efficiency among Chinese hepatitis B virus (HBV) infection patients. METHODS: Two hundred and twenty five newly diagnosed chronic hepatitis B (CHB) patients were enrolled in the study. All of these patients received IFNα treatment for a course of 48 wk, and were followed up for 24 wk after the treatment was end. Clinical information about virological response, hepatitis B e antigen (HBeAg) seroconversion rate and combined response at the end of the treatment, as well as the sustained response by the time of following up 24 wk after the treatment, was collected. Four tag-single nucleotide polymorphisms (SNPs) of IFIT1 were selected and assessed for their association with these clinical outcomes. RESULTS: At the end of the treatment, HBeAg seroconversion was observed in 27.1% patients. Thirty-six point nine percent patients achieved virological response, and 15.6% patients exhibited combined response. Sustained response was obtained in 26.2% patients. The main HBV genotype of the study was genotype B. Patients who infected with HBV genotype B or C showed better treatment efficiency, no matter which clinical outcome was considered. Among the four SNPs assessed, rs303218 (A > G) was found to be significantly associated with the end point virological response when assuming additive model [OR = 0.64 (95%CI: 0.42-0.96), P = 0.032]. Patients who carried rs303218 GG genotype had a rather higher rate of achieving virological response (response rate: 52%, OR = 0.40, 95%CI: 0.18-0.91; P = 0.028) when compared to those had AA genotype (response rate: 27%). The most significant interaction was observed in patients who had relative lower baseline aspartate transaminase. No association between SNPs and HBeAg seroconversion, combined response or sustained response was observed. CONCLUSION: IFIT1 involves in the regulation of IFNα treatment for CHB and its polymorphism rs303218 can predict the end point virological response. The finding requires further validation.

[Analysis of death causes of 345 cases with HIV/AIDS in Guangdong area].

OBJECTIVE: To analyze the death causes of 345 cases with HIV/AIDS in Guangdong area. METHODS: The situations of 345 hospitalized death cases with HIV/AIDS were conducted by retrospective analysis. RESULTS: (1)There were total 3406 hospitalized cases with HIV/AIDS in a hospital from January 2001 to December 2011 and 345 cases died, the fatality rate was 10. 13%. Since 2005 the introduction of free anti-viral treatment, the fatality rate of HIV/AIDS declined. The fatality rate of the patients whose CD4+ T lymphocyte counts <200 cells/microl was 14.61% (299/2046) and it was significantly higher than that of patients whose CD4 T lymphocyte counts >or=200 cells/microl (P <0.01). (2) 99.42% of the death cases had more than one kind of opportunistic infections (OI) and there were 924 cases of OI totally. 84. 64% of OI related to the death directly. Fungal infection was the most common in OI, followed by bacterial infection. Most OI occurred in the lungs, mouth, other systemic disseminated diseases, gastrointestine, central nerver system, septicemia, skin. The AIDS defining opportunistic infections such as several pneumonia, disseminated penicilliosis marneffei and CNS infections accounted for 29.65%. Other factors that caused HIV/AIDS death included opportunistic tumors, HIV related disease and non AIDS-related disease accounted for 15.36%. No accepted effective highly active antiretroviral therapy (HARRT) also constituted factors of death. Among cases which accepted HARRT treatment, only 6.96% had the period of treatment over three months. CONCLUSION: The fatality rate of end-stage AIDS patients was high and the opportunistic infections was the most important cause of death. Early diagnosis and treatment for opportunistic infections, timely effective HARRT were the key to improve the quality of life of AIDS patients.

[Hepatitis B virus (HBV) mutation in enhancer I (HBV Enh I)/X-promoter and the relationship between chronic HBV-related disease spectrum].

OBJECTIVE: To investigate the hepatitis B virus (HBV) mutation in the Enhancer I (HBV Enh I)/X-promoter and to analysis the relationship between chronic HBV-related disease spectrum. METHODS: 275 patients were enrolled in this study, including 100 cases of chronic hepatitis B (CHB), 74 cases of liver cirrhosis (LC), 101 cases of hepatocellular carcinoma (HCC), grouping by different HBV genotypes, using semi-nested PCR amplification of HBV Enh I/X-promoter and sequencing DNA, the mutations were determined by alignment to HBV reference sequence, the data was compared by chi2 test and analyzed by multivariate logistic regression. RESULTS: (1) Genotyping results: 61.48% (158/257) were infected with HBV genotype B, including 70 cases of CHB, 36 cases of LC and 52 cases of HCC; 38.52% (117/257) were infected with HBV genotype C, including 30 cases of CHB, 38 cases of LC and 49 cases of HCC. (2) In the patients were infected with HBV genotype B, A1123Y mutation in LC was significantly higher than in CHB (30.56% vs. 8.58%, chi2 = 8.533, P = 0.005, A = 4.693, 95% CI [1.567-14.056]), HCC was significantly higher than in CHB (28.85% vs. 8.58%, chi2 = 8.607, P = 0.003, A = 4.324,95% CI [1.544-2.109]); A1317G mutation in HCC was significantly higher than in CHB (30.77% vs. 7.14%, chi2 = 11.687, P = 0.001, A = 5.778, 95% CI [1.955-17.076]). In the patients were infected with HBV genotype C, T1323C mutation in HCC was significantly higher than in CHB (30.61% vs. 6.67%, chi2 = 6.318, P = 0.12, A = 6.176, 95% CI [1.301-29.331]). (3) Multivariate regression analyses showed that A1317G (OR = 5.706, 95% CI [1.770-18.837], P = 0.004) and T1323C (A = 5.810, 95% CI [1.114-30.306], P = 0.037) mutation were risk factors for HCC. CONCLUSION: HBV Enh I/X-promoter mutations were associated with the development of LC and HCC, the mutations can help to predict the occurrence of LC and HCC.

[An analysis of opportunistic infection in 762 inpatients with human immunodeficiency virus infection in Guangdong areas].

OBJECTIVE: To analyze the characteristics of opportunistic infection (OI) in patients with HIV/AIDS in Guangdong and the relationship between OI and the change in blood CD4+ T lymphocyte count (CD4+). METHODS: Seven hundred and sixty two patients with HIV/AIDS admitted were analyzed. RESULTS: Among all the 762 patients, 704 (92.39%) had more than one kind of OI, with 1428 episodes totally. Etiologically, fungus infection (38.38%) was most common, followed by bacteria (36.20%), and virus (7.77%) infection. Most OI occurred in the lungs (33.05%), mouth (26.89%), skin (10.29%) and gastro-intestine (8.96%). Septicemia and other systemic disseminated diseases accounted for 6.58% and 9.94% respectively. The incidence of OI in patients with CD4+≥200/µl (103/136, 75.74%) was significantly lower than that in patients with CD4+<200/µl (601/626, 96.01%), P<0.01. All the AIDS defining OI were found in patients with CD4+<200/µl. Among them, 81.97% of patients with pneumonia carinii pneumonia (PCP), 71.43% of patients with cytomegalovirus retinitis and all the patients with cryptococcal meningitis, disseminated cryptococcosis, disseminated histoplasmosis, mycobacterium avium intracellular complex (MAC), disseminated penicilliosis marneffei and toxoplasma cerebritis had the CD4+ less than 50/µl. CONCLUSIONS: The most common OI in patients with AIDS in Guangdong area are fungi, bacterial and viral infections. Lung, mouth, skin, gastro-intestine and systemic disseminated infections are the most prevalent infections. As the CD4+ decreased, the incidence of OI especially AIDS defining OI increased. Dynamic detection of CD4+ will be of great help for the prediction, prevention, early diagnosis and treatment of OI in patients with AIDS.

[Inhibitory effect of anluohuaxianwan on experimental hepatic fibrosis in rats].

OBJECTIVE: To investigate the effects of anluohuaqianwan on experimental hepatic fibrosis induced by dimethyl nitrosamine (DMN) in rats. METHODS: 36 male SD rats were randomly dividied into three groups: model group, normal group, anluohuaqianwan group. The rats in the three groups were treated with DMN daily for 4 weeks. The liver function was detected using auto biochemistry analyzer, the serum HA, LN, IV-C, PIIIP were detected by immunoradiometry, the histopathology was observed in the left liver lobe after HE staining, the expression of matrix metalloproteinase-2 (MMP-2) in liver tissue were detected by immunohistochemistry. RESULTS: The serum levels of ALT, AST, ALP, TP, ALB and the contents of HA, LN, IV-C in model group were significantly increased compared to these in the normal group (P less than 0.01). The serum levels of ALT, AST and the contents of HA in anluohuaqianwan group were significantly lower than those in the model group (P less than 0.01). The liver MMP-2 in the model group was significantly increased compared to that in the normal group (P less than 0.05). The expression of MMP-2 in liver tissue of model group was lower than that in the anluohuaqianwan group (P less than 0.05). CONCLUSION: Anluohuaqianwan can inhibit liver fibrosis in rats induced by DMN.

[The infection of dendritic cells by recombinant adenoviral vector carrying HBsAg-HSP70 chimeric gene and its biological characteristics observations].

OBJECTIVE: To test the infeciton efficiency of recombinant adenoviral vector carrying HBsAg-HSP70 chimeric gene and to abserve its biological characteristics. METHODS: Peripheral blood mononuclear cells (PBMC) were separated from healthy blood donor and they were infected by Ad-HSP70-HBsAg on the first day of isolation. DCs were induced in medium with cytokines IL-4, GM-CSF and TNF-alpha in vitro. The biological characteristics of DC induced were analyzed by inverted fluorecent microscope, RT-PCR, flow cytometer (FACS), and mixed lymphocyte reaction (MLR). RESULTS: The traced gene-GFP were abserved in DCs by inverted fluorecent microscope and HSP70-HBsAg gene mRNA expression was detected by RT-PCR after the Ad-HSP70-HBsAg infection. FACS analysis shown that the expression of CD1a, CD80, CD86 and HLA-DR on surfece of two groups of DCs were similar. MLR showed that there are not a statitic difference of stimulated index (SI) between two groups. CONCLUSION: Results indicated that Ad-HSP70-HBsAg can effectively infected DCs without affecting its biological characteristics.

[Case-controlled study of entecavir treatment for chronic severe hepatitis B].

OBJECTIVE: To evaluate the efficacy and safety of entecavir (ETV) treatment for chronic severe hepatitis B. METHODS: 78 patients with chronic severe hepatitis B and positive HBV DNA were divided into ETV group and control group, each group had 39 patients. ETV group was given the same conventional therapy as control group, and was treated with ETV. The change of liver function, PTA, HBV DNA level were observed, and adverse events were recorded. The effective rate of treatment between ETV group and control group, the baseline characteristics between the effective cases and non-responsive cases after ETV treatment were compared at week 12. RESULTS: The baseline characteristics were well balanced between ETV group and control group. The effective rate of ETV group was 56.41% versus 33.33% of control group at week 12 (P = 0.0405). The effective rate of ETV group was higher than that of control group, in the early stage of chronic severe hepatitis B (P = 0.0275), but there was no statistically significant in the middle or late stage (P = 0.4687). The comparison result of baseline characteristics between the effective and non-responsive cases after ETV treatment showed: there were statistically different in age, bilirubin level, HBV DNA level and stage of the severe hepatitis, proportion of cirrhosis, but no statistically different in cholinesterase level, alpha-fetoprotein level and sex ratio, the proportion of ascites, positive HBeAg (P > 0.05). No serious adverse events occurred. CONCLUSIONS: ETV improves the curative effect when used in the early stage of chronic severe hepatitis B, and may not in the middle and late stage. The curative effect of ETV may be affected by age, bilirubin level, HBV DNA level and stage of the severe hepatitis, cirrhosis. ETV has good security in the treatment for chronic severe hepatitis B.

[Construction of replication-deficient recombinant adenoviral vector carrying HBsAg and HSP70 chimeric gene and its expression in vitro].

OBJECTIVE: To construct a recombinant adenoviral vector carrying HBcAg-HSP70 chimeric gene by homologous recombination in bacteria and to detect its expression in vitro. METHODS: Heat shock protein 70 gene from Mycobacterium tuberculosis were amplified by PCR and were cloned to adenoviral shuttle plasmid pAdTrack-CMV-HBsAg. Then the resultant pAdTrack-CMV-HBsAg-HSP70 was cotransfected into BJ5183 bacteria with the plasmid pAdeasy-1. The adenoviral plasmid carrying HBsAg-HSP70 gene (pAd-HBsAg-HSP70) was generated with homologous recombination in bacteria and the adenoviruses were produced in 293 cells. Several kinds of mammal cells (293 cells and Vero cells) were infected with adenoviruses and the expression of HBsAg-HSP70 was detected by RT-PCR and ELISA in vitro. RESULTS: The adenoviral plasmids pAd-HBsAg-HSP70 were obtained by selection for kanamycin resistance and confirmed by restriction endonuclease Pac analyses. The recombinant adenoviruses Ad-HBsAg-HSP70 were packaged successfully in 293 cells. The titer of Ad-HBsAg-HSP70 was up to 2 x 10(12) pfu/L after the second passage of proliferation in 293 cells. HBsAg and HSP70 were expressed efficiently in mammal cells after infection. CONCLUSION: The recombinant adenoviruses expressing HBsAg and HSP70 were constructed successfully which can be used further in study of gene therapy for HBV.

[Diammonium glycyrrhizinate does not affect efficacy of adefovir dipivoxil therapy in patients with HBeAg-positive chronic hepatitis B].

OBJECTIVE: To evaluate the effect of diammonium glycyrrhizinate on the antiviral therapy with adefovir dipivoxil (ADV) in patients with HBeAg-positive chronic hepatitis B. METHODS: Patients with HBeAg-positive chronic hepatitis B enrolled in this study were randomized to receive either ADV 10 mg/d fir 48 weeks or placebo for 24 weeks followed by ADV 10 mg/d for 24 weeks. Antiviral activities of diammonium glycyrrhizinate administered during the trial were studied with respect to virological and serologic response, and ALT normalization. RESULTS: Twenty-one of 142 patients in ADV group vs. 11 of 68 patients in placebo group were treated with diammonium glycyrrhizinate. There was no significant difference in virological, serological and biochemical responses between patients with or without diammonium glycyrrhizinate in both therapy groups. During double-blind period, virological response was significantly worse in patients only receiving diammonium glycyrrhizinate than those combined with ADV. CONCLUSION: Diammonium glycyrrhizinate had no antiviral activity and exerts no influence on the efficacy of ADV treatment in patients with HBeAg-positive chronic hepatitis B.

[Efficacy of adefovir dipivoxil was not related to genotypes B and C of hepatitis B virus: a randomized, double-blind, multicenter clinical study].

OBJECTIVE: To determine the relationship between the response to adefovir dipivoxil (ADV) treatment in patients with HBV genotypes B and C of HBeAg positive chronic hepatitis B. METHODS: This clinical trial was a randomized, double-blind, placebo-controlled, multicenter study. A total of 226 eligible patients with HBeAg positive chronic hepatitis B were randomized (in a ratio of 2:1) receiving ADV 10 mg/d for 48 weeks (ADV+ADV group) or placebo for 24 weeks followed by ADV 10 mg/d for 24 weeks (PLB+ADV group). The primary efficacy was virologic response. The genotypes of HBV were determined by PCR-restricted fragment length polymorphism (RFLP) method using serum samples before therapy. rtN236T and rtA181V mutations were confirmed by sequencing. RESULTS: In this study, HBV genotype C was 66.7%, genotype B was 25.2%. Genotype B was more common in Guangzhou. Patients with genotype B were much younger than those with the genotype C. Patients with genotype B previously received less anti-HBV therapy. There were no significant difference in virological response (including mean reduction in HBV DNA level from baseline, serum HBV DNA load after treatment and HBV DNA undetectable rate) and serological response (the rate of HBeAg loss and HBeAg seroconversion) between patients infected with genotypes B and C in both treatment arms. CONCLUSION: There were no significant difference in virological and serological response to ADV therapy between patients infected with HBV genotype B and C.

[Induction of specific cytotoxic T lymphocyte response against hepatitis B virus by hepatitis B virus S gene-modified dendritic cells].

OBJECTIVE: To explore whether hepatitis B virus (HBV) S gene-modified dendritic cells (DCs) might induce a specific cytotoxic T lymphocyte (CTL) response. METHODS: The recombinant adenoviruses carrying HBsAg genes were prepared and used to transfect DCs generated from cord blood. The efficacy of transfection was observed through the expression of enhanced green fluorescent protein (EGFP) in DCs and the expression of HBsAg was detected by ELISA. HBV S gene-modified DCs were co-cultured with T cells from cord blood and T cells stimulating activities were detected using mixed lymphocyte reaction (MLR). The CTL assay was carried out to assess the ability of CTL lines to lyse target cells of HepG(2)22.1.5 by measuring lactate dehydrogenase (LDH) release. RESULTS: The results showed that HBV S genes were expressed in DCs with high efficacy by recombinant adenoviral vector. DCs had a normal shape after transfection. The result of MLR showed that HBV S gene-modified DCs could effectively stimulate naive T cells to proliferate. The induced specific CTL lines could lyse target cells of HepG(2)22.1.5. CONCLUSIONS: HBV S gene-modified DCs enhanced the function to induce a specific CTL effect, showing its promise for developing anti-viral vaccine in future.