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The coronavirus disease 2019 (COVID-19) has caused global public health and economic crises. Thus, new therapeutic strategies and effective vaccines are urgently needed to cope with this severe pandemic. The development of a broadly neutralizing antibody against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is one of the attractive treatment strategies for COVID-19. Currently, the receptor-binding domain (RBD) of the spike (S) protein is the main target of neutralizing antibodies when SARS-CoV-2 enters human cells through an interaction between the S protein and the angiotensin-converting enzyme 2 expressed on various human cells. A single monoclonal antibody (mAb) treatment is prone to selective pressure due to increased possibility of targeted epitope mutation, leading to viral escape. In addition, the antibody-dependent enhancement effect is a potential risk of enhancing the viral infection. These risks can be reduced using multiple mAbs that target nonoverlapping epitopes. Thus, a cocktail therapy combining two or more antibodies that recognize different regions of the viral surface may be the most effective therapeutic strategy.
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Humans , Antibodies, Monoclonal/therapeutic use , Antibodies, Neutralizing , Antibodies, Viral , COVID-19 , SARS-CoV-2 , Spike Glycoprotein, CoronavirusABSTRACT
Acute-on-chronic liver failure (ACLF) is a type of severe clinical syndrome which occurs on the basis of chronic liver diseases and has the main clinical features of acute liver decompensation, extrahepatic organ damage, and high short-term mortality rate. The underlying diseases of chronic liver diseases are mainly alcoholic hepatitis and chronic hepatitis C in Western countries, while chronic hepatitis B is the main underlying disease of chronic liver diseases in China and the Asia-Pacific region. Although there are differences in underlying liver diseases between the East and the West, the common pathological basis of most ACLF patients is usually liver fibrosis or cirrhosis caused by long-term chronic liver injury. At present, the research on ACLF has been widely carried out all over the world; however, due to the differences in region, population, and disease triggers, no consensus has been reached on the definition, diagnostic criteria, and disease management of ACLF between the East and the West. This article elaborates on the definition, pathogenesis, and management of ACLF, in order to provide clinicians with new therapeutic strategies that would improve the prognosis of ACLF.
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Objective To investigate the clinical features of hepatitis B core antibody (anti-HBc)positive patients with liver injury.Methods A total of 212 anti-HBc positive and HBsAg negative patients who were primarily diagnosed with liver injury from August 2013 to August 2014 at Ruijin Hospital were collected for this study.The patients were divided into cirrhosis group (n=60) and non-cirrhosis group (n =152) according to the status of cirrhosis.The 60 cases with cirrhosis were further compared with 60 cases with post-hepatitis B cirrhosis.The general information,biochemistry and immunology data were assessed.ANOVA was used to compare multiple groups of means,and Wilcoxon rank-sum test was used for non-parametric comparisons of the two groups.Results Only one case was positive for HBV DNA with the positivity rate of 0.5％.The causes for liver injury were as follows,60 cases with cryptogenic cirrhosis,which accounted for 28.3 ％;45 cases with drug-induced hepatitis,which accounted for 21.2 ％;33 cases with unexplained liver injury,which accounted for 15.6％;28 cases with acute hepatitis E,which accounted for 13.2％ and 15 cases with autoimmune hepatitis,which accounted for 7.1％.There were significant differences of T cell subpopulation,hepatitis B surface antibody (HBsAg) and hepatitis B e antibody (anti-HBe) quantitative level,red blood cells (RBC),platelet counts (PLT),prealbumin,albumin,alamine aminotransferase (ALT),aspartate transaminase (AST),international normalized ratio (INR),hyaluronic acid (HA),collagen Ⅲ (COL-Ⅲ) and collagen Ⅳ (COL-Ⅳ) between the cirrhosis group and non-cirrhosis group (all P＜0.05).The CD3+ CD4+ and CD3+ CD8+ counts,white blood cells (WBC),ALT,AST,total bilirubin (TBil) and albumin in anti-HBc-positive cirrhosis group were statistically different from those in post-hepatitis B cirrhosis group (all P＜0.05).Conclusions Some patients with positive anti-HBc still have HBV replication and infectivity.HBV anti-HBc positivity and HBsAg negativity may be associated with some cryptogenic cirrhosis and primary liver cancer.Patients with positive anti-HBc are prone to be complicated with drug-induced hepatitis,autoimmune hepatitis,and other liver damage related to immune mechanisms.Patients with cirrhosis have a higher risk to induce immune tolerance and progress to chronic disease than non-cirrhotic patients.Quantitative anti-HBc might be used as an indicator to predict disease progression after HBV infection.Disease condition in cirrhotic group with positive anti-HBc and negative HBsAg is less severe than that in post-hepatitis B cirrhosis group.
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Objective To construct lentivirus vectors carrying 16 short hairpin RNA (shRNA) expression cassettes targeting histone acetyltransferases and provide a powerful research approach to explore the mechanism of epigenetic genes in regulating hepatitis B virus (HBV).Methods Following the rule of short shRNA primer design,eight-pair primers (A ~ H )for each gene,which had stable interfering efficiency,were designed.The annealed primers were connected to the empty lentiviral vectors of shRNA for transformation.In order to confirm the positive clones,clones were analyzed by real-time polymerase chain reaction (RT-PCR ).Then, qualified plasmids were verified by enzyme digestion technology.Four shRNA lentivirus plasmids against the same gene were mixed to build lentivirus respectively.After the virus transfected into 293T cells for 48 and 72 hours,supernatants were collected to infect HepG2.2.15 cells.The percentage of fluorescent cells were observed and assessed by microscope 72 hours after infection.Results One hundred and twenty-eight lentiviral vectors of RNA interference (RNAi)library were constructed against 16 histone acetyltransferases and more than 80% of HepG2.2.15 cells were infected with lentivirus 72 hours after infection.Conclusions Sixteen shRNA lentivirus vectors against histone acetyltransferase are successfully constructed.Thus,a solid foundation for the study of the effect of human histone deacetylase on HBV replication is established.
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Objective To investigate whether a novel long-acting tumor necrotic factor (TNF) antagonist (soluble TNF receptor:IgG Fc [sTNFR:IgG-Fc]) can protect hepatocyte damage against liver failure caused by drugs in immunity-induced cirrhotic rats.Methods Wistar rats were repeatedly sensitized by human serum albumin (HSA) emulsified in complete freud adjuvant.The blood was collected at day 10 after the final sensitization.If anti-albumin antibody was positive,the rats were intravenously injected with HSA twice a week.After six weeks,liver cirrhosis was induced by immunity.All the model rats were divided into three groups with 15 each.Liver failure was induced with D-galactosamine/ lipopolysaccharide (LPS) intraperitoneal injection in the rats with liver cirrhosis in model group.The rats in pretreatment group were intraperitoneally injected with long-acting soluble TNF receptor p55 18 h before D-galactosamine/LPS injection.The control group were injected with 0.9％ sodium chloride.General condition,survival rate,liver function and pathological changes were all examined.Serum levels of interleukin (IL)-6,IL-22 and intrahepatic level of IL-6 were detected by enzyme linked immunosorbent assay (ELISA).The activity of Caspase 3 in hepatocyte lysis solution was measured by spectrophotography.Real-time polymerase chain reaction (PCR) was used to detect mRNA expressions of proliferating cell nuclear antigen (PCNA),bcl-2,bax and IL-22 receptor.Data were analyzed by variance analysis among groups.Results Rats in model group were dispirited with poor response after 12 hours and only 3 survived,compared with soluble TNF receptor p55 pre-treated group rats,in which all survived (P=0.029 8) with flexible response.Serum alanine aminotransferase levels in these two groups were (6 533± 360) and (105 ± 7) U/L,respectively.Hepatic regenerative nodule developed massive or submassive necrosis with septal fibrosis in model group,whereas soluble TNF receptor p55 alleviated the inflammatory and necrosis reaction of hepatic tissue.Serum IL-6 levels in model group and pretreatment group were (842.0±12.9) and (91.9±1.6) pg/mL,respectively (F=380.30,P＜0.01).Intrahepatic levels of IL-6 in these two groups were (26.2±1.2) and (11.1±0.8) pg/mL,respectively (F=176.90,P＜0.01),and serum IL-22 levels were (167.0±27.8) and (988.0±109.6) pg/mL,respectively (F=37.91,P＜0.01).Hepatic Caspase-3 activity was reduced by almost 60％ by soluble TNF receptor p55 pretreatment (F=303.70,P＜0.01) and bax expression reduced by 22％ (F=108.80,P＜0.01),while bcl-2 and PCNA expressions were up-regulated by 3.6-folds and 23.0-folds,respectively (F=115.60,P＜0.01; F=594.20,P＜0.01).Conclusions Long acting soluble TNF receptor p55 could improve survival rate,liver function and reduce inflammatory reaction of rats with liver failure induced by drugs on the basis of liver cirrhosis caused by immunity,which indicates that this drug may process a potential therapeutic value.
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Objective To measure the expression of circulating microRNA (miRNA)in patients with hepatitis B virus (HBV)-related liver failure and its relationship with disease prognosis.Methods The miRNA expressions in serum of 5 patients with HBV-related liver failure and 5 healthy control subjects were compared using Exiqon miRCURY LNATM miRNA microarray.The sera from 20 patients with chronic hepatitis B (CHB),20 patients hepatitis B related cirrhosis,50 patients with HBV-related liver failure and 40 healthy persons in Ruijin Hospital were collected.The relative expression of miRNA-595 was measured using quantitative real-time polymerase chain reaction (PCR).The relative expressions of miRNAs among groups were analyzed using student t test,the correlations were analyzed by Pearson and Spearman correlation.Results Microarray informed that 92 miRNAs changed significantly in patients with HBV-related liver failure,and miRNA-595 increased most significantly.The results of real-time PCR showed that the relative expressions of miRNA-595 ,miRNA-300 and miRNA-122 were 6.03 (t=3.134, P =0.003),3.12 (t=7.221 ,P <0.01)and 2.77 (t=2.671 ,P =0.021),which were higher compared to those in healthy control group.In the analysis of the relationship between miRNA-595 expression and disease prognosis in patients with HBV-related liver failure,the relative expressions of miRNA-595 in patients with CHB,hepatitis B related cirrhosis and HBV-related liver failure were 2.26 (t =3.780,P =0.001),3.32 (t = 6.111 ,P < 0.01)and 6.03 (t = 3.134,P = 0.003),respectively,which were all increased compared to that of the healthy control.The relative expression of miRNA-595 of patients with HBV-related liver failure was 2.66 times (t=2.450,P =0.043)higher than that of patients with CHB. When dividing patients according to prothrombin activity,miRNA-595 increased significantly in patients with early stage liver failure.When dividing patients according to model of end-stage liver disease (MELD) score,MELD score was positive correlated with the expression of miRNA-595 when MELD score was under 30 (r=0.673,P =0.004).The expression of serum miRNA-595 in survival group (11 .08,n=23) was higher than that in non-survival group (3.67,n = 27,t =4.309,P =0.041).Conclusions The expressions of miRNA595 ,miRNA-300 and miRNA-122 are all increased in patients with HBV-related liver failure,especially the expression of circulating miRNA-595 at early stage of the disease.The miRNA-595 may be used as a new serum biomarker for monitoring the severity of disease.
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Objective To explore the expressions of circulating microRNAs (miRNAs) in acute liver failure mice induced by D-galactosamine (GalN)/lipopolysaccharides (LPS) and the correlation with miRNAs in the liver.Methods Forty clean grade Balb/C mice,with 32 in the model group and 8 in the control group were enrolled in the study.Liver failure was induced by intraperitoneally injection of D-GalN and LPS in mice of the model group,while mice of the control group were intraperitoneally injected with 1 mL 0.9 ％ sodium chloride solution.Serum and liver samples were collected at 0,3,5,7 hours following administration,and eight mice should be supplied to each sample,and changes of alanine aminotransferase (ALT),aspartate aminotransferase (AST) and histopathology of the liver were observed.miRNA from both the serum and the liver was extracted,miRNA expression profile in the liver at 0,5,7 hours by locked nucleic acid (LNA)-miRNA microarray was analyzed and miRNA by quantitative real-time reverse transcription polymerase chain reaction (RT-PCR) was detected.Means of the two groups were compared using one-way ANOVA and correlation analyses were performed using Pearson and Spearman correlation.Results Expression of miRNAs in the liver tissue changed significantly over time with the occurrence of acute liver failure in the mice.Twenty-one miRNAs were up-regulated and 27 were down-regulated,among which miRNA-122 and miRNA-1187 were down-regulated while miRNA-146a and miRNA-155 were up-regulated.It was confirmed by the PCR assay that the expression of miRNA-122 and miRNA-1187 in the liver gradually decreased,while those in the serum were up-regulated over time.However,the expressions of inflammation associated miRNA-155 and miRNA-146a were up-regulated both in the serum and the liver after administration.The expressions of miRNA-122 and miRNA-1187 were negatively correlated between serum and liver (r=-0.477,P=0.0089,r=-0.420,P=0.231),while the expressions of miRNA-155 in serum and liver were positively correlated (r=0.678,P=0.0001).Moreover,the expressions of miRNA-122 (r=0.571,0.554) and miRNA-1187 (r=0.471,0.542) were also positively correlated with serum levels of ALT and AST (all P＜0.05).Liver and serum levels of miRNA-122 and miRNA-1187 changed significantly at 5 hours after administration,which preceded the changes of ALT/AST.Conclusions The expressions of miRNA-122 and miRNA-1187 in serum are well inversely correlated with the corresponding expressions in liver tissues during acute liver failure in mice.The changes of miRNA-122 and miRNA-1187 in the serum precede those of ALT/AST.These data suggest that serum miRNA-122 and miRNA-1187 might be the candidate serum biomarkers for early prediction of liver injury.
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Objective To observe the regulatory role of microRNA-1187(miR-1187)in hepatocyte apoptosis through miR-1187 targeting regulation of caspase-8 mRNA expression.Methods The acute liver failure model was established by injection of D-galactosamine plus lipopolysaccharides(LPS)in BALB/c mice.The liver tissues were collected for LNA-miRNA array analysis and functional analysis of genes targeted by miRNA.Embryonic murine hepatocyte cell line 2(BNL-CL2)was cultivated in vitro and treated with tumor necrosis factor(TNF)-α and D-galactosamine to induce the transfection of miR-1187 in transfected group or untransfected group.The expressions of miR-1187 and caspase-8 mRNA were detected by real-time polymeramse chain reaction(PCR)and caspase-8 protein was determined by Western blot.The apoptosis rate was detected by flow cytometry.The comparison of means between groups was done by t test.Results The miR-1187 signal was deceased with the development of acute liver failure.The 3'UTR of caspase-8 mRNA had direct binding sites with miR1187.In BNL-CL2 cell experiments,miR-1187 was down-regulated in untransfected group and decreased more slowly in transfected group(t=6.371,P<0.01).The expression of caspase-8 mRNA was up-regulated in untransfected group and increased less in transfected group(t=4.539,P<0.01).The apoptosis rate in transfected group was significantly lower than untransfected group(t=3.365,P<0.05).Conclusios miR-1187 is one of inhibitors of hepatocyte apoptosis.High expression of miR-1187 could regulate the expression of caspase-8 mRNA,thus inhibit the apoptosis of hepatocytes.
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ObjectiveTo investigate the histological features as well as the factors influencing liver disease progression in Chinese patients with chronic hepatitis C (CHC). MethodsA total of 102 CHC patients who underwent percutaneous liver biopsy between August 2007 and May 2010 were recruited. Age, gender, body mass index (BMI) and transmission route of recruited patients were recorded. Serum levels of alanine transaminase (ALT) and aspartate transaminase (AST), HCV genotypes, HCV viral load and liver histological changes were detected. Statistical analysis was done by t test and Logistic regression. ResultsThe serum levels of ALT and AST in CHC patients with histological activity index (HAI) ≥4 were much higher, while platelet (PLT) counts were lower than those with HAI ＜4(t=2.209, 2. 298 and 2. 565, respectively; all P＜0.05). Likewise, in patients with F≥3, the serum levels of ALT and AST as well as the mean age and the duration of infection were significantly elevated compared with F ＜ 3 group ( t = 3.497, 2. 758, 2. 340 and 2. 570,respectively; all P＜0. 05), while PLT counts were much lower (t = 2. 761, P=0. 007). The unvariate predictors for HAI≥4 were female, ALT＞1 × upper limits of normal (ULN), AST level,F≥3, HCV RNA≥6 lgIU/mL and PLT counts. By mutivariate analysis, the Ishak stage score was the only independent predictor for HAI≥4 (OR 3.098, 95％CI 1.884-5. 092; P＜0.01). Finally,the univariate predictors for F≥3 were age, BMI≥24 kg/m2 , ALT＞1 × ULN, AST level, HAI≥4,PLT counts and duration of infection≥ 15 years. Multivariate analysis revealed that age (OR 1. 074,95％CI 1.006-1. 146; P=0.033), ALT level (OR 1. 035, 95％CI 1.015－1.055; P＜0.01), ASTlevel (OR 0. 969, 95％CI 0. 948－0. 990; P=0. 005), the duration of infection ≥15 years (OR 37. 215, 95％CI 5. 816－238. 127; P＜0.01) and HAI≥4 (OR 1. 939, 95％CI 1. 426－2. 636; P＜0.01) were independent predictors for F≥ 3. ConclusionAge, ALT level, AST level, duration of infection≥15 years, HAI≥4 are independent predictors for liver fibrosis.
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Objective To analyze epidemiological and clinical characteristics of chronic hepatitis C (CHC) patients. Methods The clinical data of 323 CHC patients were collected. The transmission modes, clinical manifestations and virological features were recorded. The liver histological change was also analyzed in 39 cases whose liver biopsy samples were available. The comparison between two groups was performed by t test. Results Among the 323 CHC patients, 135 cases (41. 8%) had history of blood or blood products transfusion. Sixty-seven cases (20. 7%) had undergone surgery and trauma operation. Nineteen cases (5. 9%) had history of Chinese medicine acupuncture. Eighteen cases (5.6%) had undergone hemodialysis. Two patients (0.6%) were infected through vertical transmission. Twenty-one cases (6. 5%) had history of intravenous drug use and two cases (0. 6%) had history of unsafe sexual contact. The possible transmission routes for the other 78 cases (24.1%)were unknown. Fourteen patients (4. 3%) were co-infected with hepatitis B virus (HBV). The major prevalent genotypes were hepatitis C virus ( HCV) genotype lb and 2a, which were 145 cases (65. 3%) and 21 cases (9. 5%) respectively. HCV viral loads were as high as 1 × 105 IU/mL in 74 cases (26. 1%) and 1× 106 IU/mL in 103 cases (36. 4%). Twenty-three patients (7.1%) developed obvious clinical manifestations. Among 39 patients undergoing liver biopsy, 14 cases (35. 9%) had hepatic inflammation activity index (HAI)≥4, six cases (15. 4%) had fibrosis stage (F) ≥3, four cases (10. 3%) had HAI≥4 and F≥3. Conclusions The most common HCV transmission modes are blood transfusion and use of blood products. However, surgery and trauma operation should be paid more attention. Besides blood transfusion, the transmission modes of intravenous drug injection, hemodialysis and traditional Chinese medicine acupuncture are increasing. The major HCV genotypes are lb and 2a. The viral loads of most patients are relatively high. Most patients infected with HCV don't show any obvious hepatitis symptoms and physical signs. However, the liver biopsy results from 39 patients suggest that most patients develop liver histological changes.