Inhibition of bovine viral diarrhea virus (BVDV) by mizoribine: synergistic effect of combination with interferon-alpha

Bovine viral diarrhea virus (BVDV) is a well-characterized member of the Flaviviridae family. BVDV may be a surrogate model for human hepatitis C virus (HCV), since HCV does not replicate efficiently in cell cultures and animals. Mizoribine, a nucleoside analog clinically used as an immunosuppressant, was found to be active against the replication of BVDV in cell culture. We further investigated the combination of mizoribine and interferon (IFN)-alpha for antiviral activity and found that the combination synergistically inhibited BVDV replication in bovine kidney cells, as monitored by the inhibition of virus-induced cytopathicity. The combination of ribavirin and IFN-alpha was also synergistic in inhibiting BVDV replication. Treatment of infected cells with a combination of mizoribine and IFN-alpha at the concentrations, at which the respective compounds proved to be inactive, markedly reduced viral infectivity in culture supernatants. These results indicate that mizoribine in combination with IFN-alpha may have potential for the treatment of HCV infection.     

Antiviral activity of 5'-O-carbonate-2',3'-dideoxy-3'-thiacytidine prodrugs against hepatitis B virus in HepG2 2.2.15 cells

The antiviral activities of lamivudine (3TC; 2',3'-dideoxy-3'-thiacytidine) and six 5'-O-carbonates of 3TC were determined by inhibition of hepatitis B virus (HBV) replication in HepG2 2.2.15 cells. HBV DNA in cell supernatants was quantified by real-time polymerase chain reaction (PCR). The results showed that 3TC-Etha was six times more active than 3TC and that 3TC-Buta, 3TC-Hexa and 3TC-Octa were approximately three times more active than 3TC. In contrast, 3TC-Penta and 3TC-Metha showed anti-HBV activity similar to that of the parent compound 3TC. In conclusion, 5'-O-carbonates of 3TC appear to be promising candidates as anti-HBV compounds. This modification could optimise the use of 3TC, a well-tolerated, effective and inexpensive drug, in monotherapy or combined therapy for chronic HBV infections as well as human immunodeficiency virus (HIV)/HBV co-infections.     

Evaluation of hepatitis B virus replication and proteomic analysis of HepG2.2.15 cell line after cyclosporine A treatment

Aim： The effect of cyclosporine A （CsA） on hepatitis B virus （HBV） replication was investigated, and proteomics expression differentiation after CsA treatment was studied in order to provide clues to explore the effect of CsA on HBV replication. Methods： Methyl thiazolyl tetrazolium （MTT） assay was used to evaluate the cytotoxicity of CsA. The HBV replication level in the HBV genomic DNA transfected HepG2.2.15 cell line was determined by an ELISA analysis of hepatitis B surface antigens （HBsAg） and Hepatitis B e antigens （HBeAg） in culture supernatant, while the intracellular HBV DNA replication level was analyzed by slot blot hybridization. Two-dimensional electrophoresis was used to investigate the alteration of protein expression in HepG2.2.15 after CsA treatment in vitro. The differentially-expressed proteins were identified by Matrix-assisted laser desorption/ionization-time of flight mass spectrometry combined with an online database search. Results： CsA was able to inhibit the expression of HBsAg, HBeAg, and HBV DNA replication in vitro in a dose-dependent manner. A proteomics analysis indicated that the expression of 17 proteins changed significantly in the CsA treatment group compared to the control group. Eleven of the 17 proteins were identified, including the overexpression of eukaryotic translation initiation factors （eIF） 3k, otubain 1, 14.3.3 protein, eIF2-1α, eIF5A, and the tyrosine 3/tryptophan 5-mono-oxygenase activation protein in CsA-treated HepG2.2.15 cells. The downregulation of the ferritin light subunit, erythrocyte cytosolic protein of 51 kDa （ECP-51）, stathmin 1/oncoprotein, adenine phosphoribosyl-transferase, and the position of a tumor protein, translationally- controlled 1, was shifted, suggesting it had undergone posttranslational modifications. Conclusion： Our study identified the inhibitory effect of CsA on HBV replication, and found that a group of proteins may be responsible for this inhibitory effect.     

乙肝病毒x蛋白结合蛋白抑制阿霉素诱导HepG2细胞凋亡的机制研究

目的探讨乙型肝炎病毒x蛋白结合蛋白(hepatitis Bvirus x-interacting protein,HBXIP)抑制阿霉素(doxorubicinhydrochloride,DOX,adriamycin,ADM)诱导HepG2肝癌细胞凋亡的作用及可能的分子机制,为研究肝细胞癌的临床耐药性奠定基础。方法以建立的稳定高表达HBXIP基因的HepG2细胞系为研究对象,分别用不同浓度的DOX处理高表达HBXIP组及对照组细胞,MTT法检测细胞的生存率,DAPI染色及Annexin V-FITC/PI染色检测细胞凋亡,免疫印迹法检测蛋白表达水平。结果 MTT结果表明,DOX能够抑制HepG2细胞的存活,但HBXIP对DOX诱导的HepG2细胞凋亡作用具有明显的拮抗效应,并抑制DOX诱导的caspase-3、caspase-9及其底物PARP的活化,同时促进Bcl-2蛋白的表达。结论 HBXIP蛋白能够抑制化疗药物DOX诱导的HepG2细胞凋亡,其机制可能与调节胱天蛋白酶家族关键因子的活性相关。     

晶珠肝泰舒在2.2.15细胞内对HBsAg和HBeAg的抑制作用

目的探讨晶珠肝泰舒胶囊的抗HBV作用.方法在乙型肝炎病毒基因转染的人肝癌细胞系2.2.15细胞中,研究晶珠肝泰舒对细胞的毒性和对HBsAg和HBeAg分泌的抑制效果.结果晶珠肝泰舒胶囊加入细胞培养8天,两批实验对细胞的半数中毒浓度平均为(3433.11±288)μg/ml,最大无毒浓度(TC0)平均为(1250±0)μg/ml.最大无毒浓度时抑制细胞分泌HBeAg率为(34.5±2.87)%,50%有效浓度为(2294.33±136.3)μg/ml,选择指数1.50±0.09.最大无毒浓度时抑制细胞分泌HBsAg率(31.1±3.1)%(P＜0.001),50%有效浓度为(2221.29±151.58)μg/ml,选择指数1.55±0.11.结论晶珠肝泰舒胶囊在2.2.15细胞培养中8天最大无毒浓度对HBeAg和HBsAg的分泌有抑制作用.     

乙型肝炎病毒基因型用干扰素α-2b联合苦参素治疗的临床研究

目的：研究乙型肝炎病毒基因型用干扰素α-2b联合苦参素治疗慢性乙型肝炎疗效的关系。方法：应用免疫荧光定量方法检测60例慢性乙型肝炎（慢乙肝）的乙肝病毒基因型，患者均在常规保肝治疗基础上应用干扰素α-2b和苦参素联合抗病毒治疗，疗程6个月。随访6个月。结果：治疗结束及随访6个月时血清HBeAg／HBeAb转换率、HBVDNA阴转率、HBVDNA降低2—5logL率为B基因型组分别为83.3％、83.3％、16．67％；66．67％、75．00％、25．00％；C基因型组分别为45．83％、47．92％、45．65％；35．42％、39．58％、41．67％。B基因型组与C基因型组比较．血清HBeAg／HBeAb转换率、HBVDNA阴转率、血清转氨酶及肝纤维化指标下降均有统计学意义（P〈0．05）。结论：干扰素α-2b联合苦参素治疗慢性乙型肝炎疗效及随访观察疗效B基因型优于C基因型。     

乙肝免疫球蛋白联合乙肝疫苗阻断乙肝病毒母婴垂直传播的临床研究

目的 研究乙肝免疫球蛋白(HBIG)联合乙肝疫苗应用于孕妇为乙肝病毒携带者时,对乙肝病毒母婴垂直传播的阻断效果.方法 A组83例,于孕28、32、36周分别注射HBIG 200 U,新生儿出生时及半月时分别注射200 U HBIG,并按0、1、6方案常规注射乙肝疫苗;B组87例仅常规产前检查及监护,新生儿行乙肝疫苗全程免疫.在1岁时检测血清抗-HBs.结果 A组较B组的婴儿1岁时外周血HBSAg阳性率显著降低,P＜0.05.结论 HBIG联合乙肝疫苗可以显著降低新生儿外周血HBSAg阳性率,但仍存在部分患儿的阻断失败.     

Enhancement of in vitro and in vivo anticancer activities of polysaccharide peptide from Grifola frondosa by chemical modifications

Context: Grifola frondosa (Polyporaceae), maitake, is a widely consumed edible mushroom in some Asian countries. The fruit bodies and mycelia of maitake have shown different bioactive compounds with anticancer and other therapeutic properties.

Objective: This study evaluated three chemically modified maitake polysaccharide-peptides’ (MPSP) adjuvant effect (in vivo) and anticancer activity (in vitro growth inhibitory effect) compared with crude MPSP from G. frondosa.

Materials and methods: We investigated the possibility of enhancing the adjuvant effect and anticancer effect of crude MPSP by using simple chemical modification methods to convert crude MPSP to phosphorylated, acetylated or esterified MPSPs. The adjuvant effect and growth inhibitory effect were evaluated by C6 cell inoculated rat model with cyclophosphamide (CPA) treatment and in vitro cell viability assay, respectively.

Results: All four tested MPSPs showed significant adjuvant effect to CPA treatment on rats inoculated with C6 cancer cells. In addition, an obvious growth inhibitory effect was observed in C6 cancer cells but not in normal brain cells treated with various forms of MPSPs. Only phosphorylation could significantly (p?<?0.05) improve the adjuvant effect (in vivo) and growth inhibitory effect. A same rank order (phosphorylated MPSP > esterified MPSP ≥ acetylated MPSP ≥ crude MPSP) of efficacy was observed in both the in vivo and in vitro assays.

Discussion and conclusion: This study showed chemical phosphorylation could markedly enhance both adjuvant effects and growth inhibitory effects. This study demonstrated the feasibility of enhancing the efficacy of MPSP by using a simple chemical modification method, and this provides a foundation for future study in this area.     

Changes in different regions of hepatitis B virus gene in hepatitis B 'e' antigen-negative patients with chronic hepatitis B: the effect of long-term lamivudine therapy

BACKGROUND: Lamivudine therapy for chronic hepatitis B has been associated with changes in different regions of the hepatitis B virus nucleotide sequence. AIM: To study changes in the sequences of polymerase and precore/core promoter regions of hepatitis B virus, before and during 5 years of therapy with lamivudine. METHODS: Eighty consecutive samples were taken from 10 chronic hepatitis B 'e' antigen-negative patients. RESULTS: Nine patients carried hepatitis B virus precore mutations during the study. Before therapy, wild type was replaced by A1896 in two (20%) cases. During treatment, A1896 reverted transitory to wild type in five cases (50%) and in one case wild type was replaced by A1896. The continuous detection of precore mutations during therapy was associated with a lower response rate. YMDD mutations were observed in nine cases and both, L180M and M204V/I mutations were simultaneously detected in six cases. About 75% of the patients with M204V mutations were responders and none with M204I or mixed pattern sustained response. CONCLUSION: Hepatitis B 'e' antigen-negative patients exhibit changes in the precore regions both spontaneously and under lamivudine therapy, the transitory reversion to wild type being most frequently witnessed. Patients carrying M204V mutations are more likely to respond to therapy. If, in further studies, these results are confirmed some patients with YMDD mutations could benefit from prolonging the duration of lamivudine therapy.     

Inhibition of hepatitis B virus (HBV) replication by pyrimidines bearing an acyclic moiety: effect on wild-type and mutant HBV

Chronic hepatitis B virus (HBV) infection remains a major health problem worldwide. The main clinical limitation of a current antiviral drug for HBV, lamivudine, is the emergence of drug-resistant viral strains upon prolonged therapy. A group of 5-, 6-, or 5,6-substituted acyclic pyrimidine nucleosides with a 1-[(2-hydroxyethoxy)methyl] moiety were synthesized and evaluated for antiviral activities. The target compounds were prepared by the reaction of silylated uracils possessing a variety of substituents at the C-5 or C-6 positions or both with 1,3-dioxolane in the presence of potassium iodide and chlorotrimethylsilane by a convenient and single-step synthesis. Among the compounds tested, 5-chloro and 5-bromo analogues possessing an acyclic glycosyl moiety were the most effective and selective antiviral agents in the in vitro assays against wild-type duck HBV (EC50=0.4-2.2 and 3.7-18.5 microM, respectively) and human HBV-containing 2.2.15 cells (EC50=4.5-45.4 and 18.5-37.7 microM, respectively). These compounds were also found to retain sensitivity against lamivudine-resistant HBV containing a single mutation (M204I) and double mutations (L180M/M204V). The compounds investigated did not show cytotoxicity to host HepG2 and Vero cells, up to the highest concentration tested. The results presented here confirm and accentuate the potential of acyclic pyrimidine nucleosides as anti-HBV agents and extend our previous observations. We herein report the capability of acyclic pyrimidine nucleosides to inhibit the replication of both wild-type and drug-resistant mutant HBV.     

Unfavourable effects of colchicine in combination with interferon-alpha in the treatment of chronic hepatitis C

BACKGROUND: The prognosis of chronic hepatitis depends on the progression of hepatic fibrosis. AIM: To investigate whether the antifibrotic drug colchicine, in combination with interferon-alpha has a role in the treatment of chronic hepatitis C. METHODS: Sixty-five HCV-RNA positive patients with chronic hepatitis were randomized to receive interferon-alpha, 6 MU t.i.w. for 6 months followed by 3 MU t.i.w. for further 6 months, with or without the adjunct of colchicine, 1 mg o.d., 6 days a week, for 3 years. We report an interim analysis after the first 18 months. RESULTS: Thirty-four patients received interferon-alpha and 31 received interferon-alpha and colchicine. The two groups were comparable for baseline data, including HCV-RNA levels, genotypes and histological grading/staging. Drop-outs and side-effects were similar. The proportion of patients who achieved alanine transaminase normalization or undetectable HCV-RNA at month 6 was higher in the interferon-alpha (68% and 47%, respectively) than in the interferon-alpha plus colchicine group (32% and 23%, P=0.004 and P=0. 04, respectively). End-of-treatment biochemical and virological response occurred in 41% and 29% of the interferon-alpha and 19% and 10% of the combination group, respectively (P=0.05 and P=0.05). Sustained biochemical response occurred in 26% of the interferon-alpha and 6% of the interferon-alpha plus colchicine group (P=0.03), corresponding percentages of sustained HCV-RNA loss being 21% and 3% (P=0.04). CONCLUSIONS: The combination of colchicine and interferon-alpha worsens the effectiveness of interferon-alpha alone in HCV chronic hepatitis. These alarming findings prompted us to interrupt the trial at this stage.     

Meta-analysis: combination therapy with interferon-alpha 2a/2b and ribavirin for patients with chronic hepatitis C previously non-responsive to interferon

BACKGROUND: The efficacy of interferon-alpha plus ribavirin treatment for patients not responding to interferon monotherapy is not well established. AIM: To assess the efficacy and safety of combination therapy with interferon-alpha 2a/2b plus ribavirin by performing a meta-analysis of randomized clinical trials. METHODS: A systematic search of electronic databases for randomized clinical trials of interferon-alpha 2a/2b plus ribavirin was conducted independently by two investigators. Data abstraction was performed. The primary end-point was a sustained virological response. Estimates of the common odds ratio were calculated using a random effects model. RESULTS: Of the 127 identified studies, 46 were considered for evaluation and 10 were included (1728 patients). The pooled sustained virological response was 12.6% (95% CI, 9.5-16.3%) for combination therapy vs. 2% (95% CI, 0.9-4.0%) for interferon monotherapy, with a common odds ratio of 5.49. Higher doses of interferon, a longer duration of therapy (48 weeks) and genotypes other than 1 and 4 were associated with an improvement in response. More side-effects and discontinuations were observed with combination therapy than with interferon monotherapy. CONCLUSIONS: Non-responders to interferon may benefit from re-treatment with combination therapy, especially from a 48-week regimen.     

Inhibition of hepatitis B virus polymerase-activity by various agents. Transient expression of hepatitis B virus DNA in hepatoma cells as novel system for evaluation of antiviral drugs

The effect of three putative antiviral drugs--aciclovir (acyclovir, CAS 59277-89-3), zidovudine (azidothymidine, CAS 30516-87-1) and sorangicin B (a macrocyclic lactone, CAS 100415-25-6)--on replication and gene expression of hepatitis B virus (HBV) was studied in HepG2 cells. Transfection of these cells with cloned circular HBV DNA resulted in the production and secretion of virions into the medium. When antiviral drugs were added in increasing concentrations (aciclovir at 0.5 microgram/ml to 150 micrograms/ml, zidovudine 0.1 microgram/ml to 30 micrograms/ml, sorangicin B 1 micrograms/ml to 30 micrograms/ml), the activity of the viral polymerase decreased in a dose-dependent manner. Production of viral proteins as measured by the secretion of HBsAg and HBeAg into the medium was unaffected, suggesting interference of these drugs with viral DNA/RNA synthesis. It is concluded that aciclovir, zidovudine and sorangicin B inhibit the replication of HBV. Furthermore, the cell system used in our study appears to be suitable for the rapid testing of antiviral drugs and their evaluation for possible studies in vivo.     

Verteporfin therapy in combination with triamcinolone: published studies investigating a potential synergistic effect

ABSTRACT

Evidence from randomized, placebo-controlled, double-masked studies has demonstrated that verteporfin (Visudyne) therapy is effective in reducing the risk of visual acuity loss in selected groups of patients with choroidal neovascularization (CNV) due to age-related macular degeneration (AMD). Pilot studies of intravitreal triamcinolone acetonide monotherapy revealed promising results in patients with CNV due to AMD, but the visual acuity outcomes in a randomized, controlled study were lower than anticipated. Recently, however, there has been growing interest in the adjunctive use of triamcinolone to improve visual acuity and reduce regrowth of CNV in patients receiving verteporfin therapy. This review article surveys the currently available evidence, based on a Medline search covering the years 1980–2005 and abstracts from recent scientific meetings. A number of small-scale, uncontrolled pilot studies have indicated that the combination of triamcinolone acetonide with verteporfin therapy may be beneficial. Evidence is now needed from randomized, controlled studies to determine whether the potential benefits of combination therapy outweigh the possible risks of increased intraocular pressure, endophthalmitis, and progression of cataract that have been observed in trials of triamcinolone monotherapy.     

拉米夫定联合香菇多糖治疗慢性乙型肝炎32例近期疗效观察

目的　观察和研究拉米夫定联合香菇多糖治疗慢性乙型肝炎的近期疗效。方法　选择慢性乙型肝炎患者 3 2例用拉米夫定 (10 0mg/d)和香菇菌多糖 (10mg/d)治疗为观察组 ,3 0例单用拉米夫定 (10 0mg/d)治疗作为对照组 ,治疗 6个月 ,观察症状体征、肝功能、乙肝病毒复制指标。结果　血清HBV -DNA阴转率与对照组之间无显著差异 (87 5 %对 83 3 % ,χ2 =0 2 17,P >0 0 5 ) ,观察组HBeAg阴转率显著高于对照组 (5 6 3 %对 2 6 7% ,χ2 =5 5 65 ,P <0 0 5 ) ,ALT的复常率亦显著高于对照组 (78 1%对 5 3 3 % ,χ2 =4 2 49,P<0 0 5 ) ,未发生不良反应。结论　拉米夫定联合香菇多糖治疗慢性乙型肝炎的近期疗效肯定 ,较单用拉米夫定疗效有明显提高 ,安全性良好 ,值得进一步研究推广     

基因乙肝疫苗联合乙肝免疫球蛋白阻断乙型肝炎母婴传播的研究

目的：探索阻断母婴乙肝病毒垂直传播的最佳免疫方案。方法：对本研究的所有乙肝病毒标记物阳性的孕妇于妊娠晚期每月肌注乙肝高价免疫球蛋白200IU,共3次,所有新生儿按预先设计方案给予不同的主动加被动免疫。结果：本研究的乙肝病毒母婴垂直传播阻断率显著高于以往文献报道的阻断率。但乙肝疫苗皮内注射组与肌肉注射组之间的HbsAg阳性率、HbsAb阳性率及抗体滴度差异无统计学意义。结论：孕妇妊娠晚期乙肝高价免疫球蛋白被动免疫,继之所生婴儿基因乙肝疫苗5μg/次（0、1、6方案）加大剂量乙肝高价免疫球蛋白200IU／次（0、1／2、1方案）联合免疫方案可明显提高乙肝病毒母婴垂直传播阻断率。