近20年来药用植物成分体外抗HBV活性的研究进展

我国具有庞大的乙型肝炎患者群,同时又是药用植物资源十分丰富的国家.从药用植物中筛选具有抗乙肝病毒(HBV)活性的天然目标化合物,对以开发出抗病毒的高效价廉药物,对控制乙肝流行具有非常重要的意义.本文对近20a来药用植物的木脂素、多酚、生物碱、萜类、黄酮和香豆素类等70余个化学成分体外抗HBV活性的研究进展进行评述,以供参考.     

Advancements in in vitro hepatic models: application for drug screening and therapeutics

The liver is one of the most complex organs in the body, performing a multitude of functions. Liver tissue engineering is a combination of various strategies that aim at generating functional liver tissue that can help restore and/or support the ailing liver as it recuperates. Conventionally, in vitro culture has involved growing cells in different media compositions or layering them on matrices largely composed of native ECM components such as collagen or Matrigel. With recent advances in technology, more sophisticated techniques are being devised that are better equipped to capture distinct features of the liver in an in vivo microenvironment. Three-dimensional (3D) cultures of liver cells in 3D scaffolds, as spheroids or cell sheets, allow for a high degree of cell-cell and cell-matrix interaction and an in vivo-like architecture. More recently, decellularized matrices have been used as scaffolds that support ideal cell-matrix interactions. Microfabrication technologies initially used to pattern semiconductors in the integrated circuit industry have grown out of this field and now encompass a variety of methods to etch patterns onto both 2D and 3D scaffolds to allow incorporation of custom-made features resembling the fluid network and organization in native liver. This improvisation permits for enhanced vascularization and oxygen diffusion to the in vitro liver tissue. In this review, we discuss the various configurations that have been implemented in the in vitro culture of liver cells and their application in liver therapeutics in the form of implantable liver tissue constructs and tools for drug screening.     

抗病毒治疗在乙型肝炎相关性肝衰竭中的应用

乙型肝炎病毒(HBV)在HBV相关性肝衰竭的发病过程中起重要作用,其发病机制主要为免疫活化机制,免疫抑制者体内则可见肝细胞的直接损伤。为此,抗病毒治疗的作用日益受到重视。近年来,有关抗病毒治疗的时机及药物选择成为研究热点,提倡应尽早、及时采用强效低耐药的核苷(酸)类似物(NUCs)治疗,同时注意长期治疗的药物安全性。优化治疗或初始联合治疗是目前两种主要策略。     

X基因——抗乙型肝炎病毒治疗的新靶点

X基因是乙型肝炎病毒(hepatitis B virus,HBV)的四个基因之一.最近的研究表明,他不仅参与HBV感染宿主过程,还影响HBV病毒基因的复制和表达,直接或/和间接通过宿主的免疫反应或其他作用,导致HBV致病的系列病理过程,而且在HBV生命周期中具有非常重要的作用.降低或去除X基因转录,或者封闭X蛋白某些功能以及使X基因表达沉默均能显著抑制病毒复制水平.本文通过对X基因在HBV复制和基因表达中的作用以及针对X基因的干预对HBV复制和基因表达的影响的最新研究成果进行综述,提出X基因是乙肝抗病毒治疗的一个新靶点.     

新型抗乙型肝炎病毒药物的研究进展

目前HBV感染仍是全球性的公共卫生问题,全世界有高达4亿的慢性HBV感染者,我国约占10％.干扰素(IFN)及核苷(酸)类似物[nucleos(t)ide analogue,NUCs]的抗病毒疗效在大多数地区已得到认可.在抗病毒治疗方面虽有重大突破,但现有治疗方法却很少能达到病毒清除的效果,IFN不良反应大,而NUCs疗程长,易耐药.所以探索新型抗病毒药物是乙型肝炎防治领域的一大热点.随着对HBV生活史的深入了解及HBV感染系统的建立,在新型抗病毒药物研制方面取得很大进展.     

Naturally derived anti-hepatitis B virus agents and their mechanism of action

Despite that some approved drugs and genetically engineered vaccines against hepatitis B virus (HBV) are available for HBV patients, HBV infection is still a severe public health problem in the world. All the approved therapeutic drugs (including interferon-alpha and nucleoside analogues) have their limitations. No drugs or therapeutic methods can cure hepatitis B so far. Therefore, it is urgently needed to discover and develop new anti-HBV drugs, especially non-nucleoside agents. Naturally originated compounds with enormous molecular complexity and diversity offer a great opportunity to find novel anti-HBV lead compounds with specific antiviral mechanisms. In this review, the natural products against HBV are discussed according to their chemical classes such as terpenes, lignans, phenolic acids, polyphenols, lactones, alkaloids and flavonoids. Furthermore, novel mode of action or new targets of some representative anti-HBV natural products are also discussed. The aim of this review is to report new discoveries and updates pertaining to anti-HBV natural products in the last 20 years, especially novel skeletons and mode of action. Although many natural products with various skeletons have been reported to exhibit potent anti-HBV effects to date, scarcely any of them are found in the list of conventional anti-HBV drugs worldwide. Additionly, in anti-HBV mechanism of action, only a few references reported new targets or novel mode of action of anti-HBV natural products.     

Hepatitis flares and hepatitis B e antigen seroconversion: implication in anti-hepatitis B virus therapy

Hepatitis flares or acute exacerbations, defined as an abrupt elevation of serum alanine aminotransferase (ALT) over fivefold the upper limit of normal (ULN), of chronic hepatitis B virus (HBV) infection are the results of HLA-I restricted, cytotoxic T lymphocyte (CTL)-mediated immune response against HBV and its downstream mechanisms. Higher ALT levels reflect a more vigorous immune response and a more extensive hepatolysis that, in the extreme situation, may lead to decompensation and failure. In contrast, higher ALT also reflects a more robust immune clearance of HBV and, therefore, a higher chance of HBV-DNA loss and hepatitis B e antigen (HBeAg) seroconversion, both in the setting of natural course and drug therapy. Alanine aminotransferase of fivefold the ULN appears to be a significant cut-off level to categorize the patients in terms of endogenous immune response against HBV. Patients with ALT levels less than fivefold the ULN or those with a less vigorous immune response require immunomodulation to induce robust immune response to enhance HBV clearance. In contrast, those with a more vigorous immune response or those with ALT flare over fivefold the ULN should be monitored closely for spontaneous HBV clearance/HBeAg seroconversion or to start direct antiviral therapy in time to prevent the occurrence or deterioration of hepatic decompensation. In conclusion, a better understanding of the pathogenetic mechanisms and natural course of hepatitis flares, wiser selection of patients and the timing of drug therapy are crucial to achieve better treatment results.     

Vietnamese community screening for hepatitis B virus and hepatitis C virus

Summary. Asian Americans represent an important cohort at high risk for viral hepatitis. To determine the prevalence of Hepatitis B virus (HBV) and Hepatitis C virus (HCV) infection and HBV vaccination in a Vietnamese community, a total of 322 Vietnamese subjects from a local doctor’s office and annual Vietnamese Health Fair were included in this study. Demographic and clinical data were collected. 2.2% of the screened cohort tested positive for anti‐HCV and 9.3% tested positive for HBsAg. Unlike HBV‐positive subjects, HCV‐positive subjects had significantly higher liver enzymes (P = 0.0045 and P = 0.0332, respectively). The HBV‐positive group was more likely to report jaundice (P = 0.0138) and a family history of HBV (P = 0.0115) compared to HBV‐negative subjects. Forty‐eight patients (15.5%) reported a family history of liver disease (HBV, HCV, HCC, cirrhosis, other). Of this 48, 68.8% reported no personal history of HBV vaccination and 77.1% reported no family history of vaccination for HBV. Among the 183 subjects without a family history of liver disease, 156 (85.2%) reported no personal history of vaccination and 168 (91.8%) reported no family history of vaccination. HBV vaccination rates in those reporting a family history of liver disease were significantly higher (P = 0.020). There was a high prevalence of HBV infection in this community screening. Nevertheless, the rate for HBV vaccination was low. The low prevalence of abnormal liver enzymes in HBV‐positive subjects emphasizes the need for screening to be triggered by risk factors and not by abnormal liver enzymes.     

急性乙型肝炎病毒感染小鼠模型的建立

目的采用尾静脉液压法建立小鼠急性HBV感染的动物模型。方法以液压法将具有复制能力的HBV质粒pAAV-HBV1.2通过尾静脉注射到免疫功能正常的BALB/c小鼠体内,注射后第1、2、4、6、8d,分别采用改良赖氏法、时间分辨免疫荧光法、实时荧光定量PCR检测小鼠血清中谷丙转氨酶(ALT)、HBsAg、HBeAg、抗HBs、抗HBe、HBV DNA的水平,免疫组化检测肝组织HBsAg、HBcAg的表达。结果16只小鼠注射pAAV-HBV1.2后,有14只(85.7%)小鼠在注射后第1d血清中可检测到HBsAg,小鼠血清中HBsAg和HBeAg水平在第1d达高峰,之后逐渐下降,第8d均未能检测到。小鼠血清中HBV DNA在第2d达高峰,之后仍维持在较高水平,至第8d时为1.9×104copies/mL。至第8d肝组织中可见约5%的HBcAg阳性肝细胞和2%的HBsAg阳性肝细胞。结论采用尾静脉液压法成功的建立了小鼠急性HBV感染的动物模型。     

Lamivudine, adefovir and tenofovir exhibit long-lasting anti-hepatitis B virus activity in cell culture

In this work, we investigated the anti-hepatitis B virus (HBV) activity of lamivudine, adefovir, tenofovir, penciclovir and lobucavir after short-term (i.e. 24 or 48 h) or continuous (9 days) exposure of the HBV-containing cell line, HepG2 2.2.15, to these drugs. Lamivudine maintained significant anti-HBV activity when added for only 24 or 48 h to the cell cultures compared to when the drug was present for the whole period (9 days) on the cells, i.e. 50% effective concentration (EC₅₀) values for the inhibition of HBV DNA synthesis were 0.07 ± 0.02 μg ml⁻¹ after 24 h of incubation, 0.02 ± 0.01 μg ml⁻¹ after 48 h of incubation and 0.0016 ± 0.001 μg ml⁻¹ after 9 days of incubation. Similarly, the nucleoside phosphonate analogues, adefovir and tenofovir, retained significant anti-HBV activity when added for only a short period of time to the cells. The EC₅₀ values were 12 ± 1 μg ml⁻¹ (24 h) and 1.0 ± 0.2 μg ml⁻¹ (48 h) vs 0.003 ± 0.001 μg ml⁻¹ (9 days) for adefovir, and 6.5 ± 1.1 μg ml⁻¹ (24 h) and 0.8 ± 0.1 μg ml⁻¹ (48 h) vs 0.03 ± 0.02 μg ml⁻¹ (9 days) for tenofovir. In contrast, penciclovir and lobucavir lost most of their anti-viral activity when present on the cells for 48 h or less.     

Establishment and application of hepatic organoid model of hepatitis B virus infection

<正>Objective To establish the hepatic organoid of hepatitis B virus(HBV) infection on the basis of induced pluripotent stem cells (iPSC) and an inverted colloidal crystal polyethylene glycol scaffold (ICC),and to evaluate the antiviral effect of nucleoside drugs.Methods iPSC was differentiated into hepatocyte-like cells (HLC),and inoculated into ICC to construct a hepatic organoid.The relative mRNA expressions of Nanog homeobox (NANOG),     

The perils of relying on anti-hepatitis B total core antibody in screening individuals infected with HIV

Co-infection with HIV and hepatitis B virus (HBV) has serious long-term consequences. We describe a case of an HIV-infected heterosexual black African man with a delayed diagnosis of HBV infection. Baseline HBV screening was performed using a sequential testing algorithm starting with a total core antibody (anti-HBc) test, which was negative. He had no evidence of immunity against HBV and subsequently received three unsuccessful courses of HBV vaccination. He had mild but persistent elevation of liver enzymes over a five-year period despite maintaining full suppression of HIV replication on efavirenz, lamivudine and zidovudine; the latter was changed to abacavir due to lipoatrophy. Further testing revealed e-antibody positive chronic HBV infection with undetectable anti-HBc reactivity. High-grade HBV viraemia associated with L180M and M240V drug-resistance mutations was confirmed. He was subsequently switched to a tenofovir-based regimen, which achieved HBV suppression. Adopting effective HBV screening strategies in HIV-infected patients is recommended.     

Kinetics of anti-hepatitis C virus response during acute hepatitis in an immunosuppressed HCV chronically infected patient.

A hepatitis C virus (HCV) chronically infected patient developed an episode of acute hepatitis during the course of immunosuppressive therapy given for a lymphoproliferative disease. It was noted that anti-HCV antibody response, seen to be relatively stable during the follow-up, lowered dramatically in coincidence with the hepatocytolytic peak. A diagnostic liver biopsy taken at the time of the acute phase of hepatitis demonstrated a typical feature of lobular hepatitis with widespread lymphocytic infiltrates, the predominant type of which expressed CD8 immunophenotype. Cytotoxic and immunosuppressive drugs may interfere with hepatitis virus infections. However, at variance from hepatitis B virus infection in which acute liver decompensation develops after withdrawal of chemotherapy in our HCV chronically infected patient it appeared during the course of the treatment, suggesting a different hepatocytolytic mechanism. Although the actual frequency of the phenomenon is presently unknown, this observation seems to indicate that immunosuppressive and cytotoxic agents should be used with caution in HCV chronic infection in which cell-mediated immune response seems to play a major role in the production of the liver damage.     

Methylation of the viral genome in an in vitro model of herpes simplex virus latency.

An in vitro model of latency of herpes simplex virus type 1 (HSV-1) in a lymphoid cell line has been developed recently. CEM cells persistently infected with HSV-1 transiently ceased to produce virus for 24 days. This nonproductive state could either be reversed with phytohemagglutinin or maintained with concanavalin A. This system was used to study the relationship between DNA methylation and HSV-1 latency. DNA was probed for methylation by comparing the cleavage patterns generated by two pairs of restriction endonucleases (Sma I vs. Xma I and Hpa II vs. MspI); these enzymes show differential activity reflecting methylation of the recognition sequences. Viral DNA in the concanavalin A-treated cells (not producing virus) was found to be extensively methylated. By contrast, no methylated copies were detected in viral DNA from producer cells. About 800 days after the initial infection, the productive culture once again became nonproductive. Viral sequences in the latter cells were also methylated. Reconstitution experiments revealed 1-2 copies of viral DNA in cells from the latent stages and 40-80 copies in cells from productive stages. Most (if not all) of the viral genome is present in cells from various productive and latent stages. No differences in sequence arrangement were detected (although a terminal fragment of intracellular HSV-1 DNA appeared to be under-represented in latent cells). These results suggest a role for DNA methylation in the mechanism of HSV-1 latency in this system.