MicroRNA-mediated interactions between host and hepatitis C virus

Micro RNAs(mi RNAs) are small noncoding RNAs. More than 2500 mature mi RNAs are detected in plants, animals and several types of viruses. Hepatitis C virus(HCV), which is a positive-sense, singlestranded RNA virus, does not encode viral mi RNA. However, HCV infection alters the expression of host mi RNAs, either in cell culture or in patients with liver disease progression, such as liver fibrosis, cirrhosis, and hepatocellular carcinoma. In turn, host mi RNAs regulate HCV life cycle through directly binding to HCV RNAs or indirectly targeting cellular m RNAs. Increasing evidence demonstrates that mi RNAs are one of the centered factors in the interaction network between virus and host. The competitive viral and host RNA hypothesis proposes a latent cross-regulation pattern between host m RNAs and HCV RNAs. High loads of HCV RNA sequester and de-repress host mi RNAs from their normal host targets and thus disturb host gene expression, indicating a means of adaptation for HCV to establish a persistent infection. Some special mi RNAs are closely correlated with liver-specific disease progression and the changed levels of mi RNAs are even higher sensitivity and specificity than those of traditional proteins. Therefore, some of them can serve as novel diagnostic/prognostic biomarkers in HCVinfected patients with liver diseases. They are also attractive therapeutic targets for development of new anti-HCV agents.     

Hepatitis C virus infection: virus/host interactions

Infection with the hepatitis C virus (HCV) is a leading cause of chronic liver disease world-wide. This paper examines our current understanding of the complex relationship between HCV and its host, especially potential mechanisms of viral persistence and resistance to interferon therapy, and the pathogenesis of liver injury in chronic HCV infection. HCV infection leads to viral persistence and chronic disease in a very high proportion of cases, despite broad humoral and cellular immunological responses to viral proteins. These responses may be thwarted by the high rate of mutation, which leads to the generation of a highly variable mixture of closely related genomes, referred to as a quasispecies, that persists and continuously evolves in infected individuals. Understanding this, and other mechanisms of viral persistence and immune escape, will be essential in developing effective future therapeutic and preventive strategies. As far as the pathogenesis of chronic hepatitis C is concerned, two non-mutually exclusive hypotheses have been raised: first, that HCV can be cytopathic and induce liver lesions by replicating in infected hepatocytes, and second, that liver lesions could be the result of specific or non-specific immune responses. In the absence of a cell-culture model, the direct cytopathogenicity of the virus cannot be assessed confidently. Recent data suggest that cytotoxic T cells and cytokines produced by both CD4 + (T helper) and cytotoxic T cells may be responsible for much of the damage that occurs in the livers of infected patients.     

Quantitative molecular monitoring of HIV-1 RNA during antiretroviral therapy

Summary Plasma HIV-1 RNA testing was used to monitor 43 HIV-1 infected patients newly placed on antiretroviral therapy or whose therapy had been recently changed. A polymerase chain reaction kit was used to measure HIV-1 RNA in clinical samples or frozen plasma. The cutoff of this test was 200 RNA copies/ml. The first group (11 patients) was stable on long-term zidovudine monotherapy when switched to stavudine. The HIV-1 RNA of three patients who had a regular decline in CD4⁺ T cell count did not change despite this switch, with a mean follow-up of 630 days. The HIV-1 RNA copy numbers of eight patients whose CD4⁺ T cell counts were stable declined an average of 0.53 log₁₀ between days 90 and 650. The second group (14 patients) was on long-term zidovudine monotherapy and had declining CD4⁺ T cell counts over the past 6 months. Lamivudine was added to this regimen on day 0. HIV-1 RNA copy number decreased rapidly within 30 d, reaching –0.86 log₁₀ on day 90, and this effect was maintained thereafter, with a mean follow-up of 161 days. There was a concomitant mean gain of +33 CD4⁺ T cells on day 90. The third group (nine patients) had never received anti-retroviral therapy and was given zidovudine + didanosine. HIV-1 RNA copy number decreased in all cases but one, reaching –1.31 log₁₀ on day 150. This decrease was transient in three cases. The last group (nine patients) had also not had previous anti-retroviral therapy and was given zidovudine + didanosine + lamivudine in combination. HIV-1 RNA copy numbers declined rapidly in all cases, to below the cutoff in eight cases within a mean period of 50.5 days. The CD4⁺ cell counts increased by 164 cells/µl on day 14 and by 201 cells/µl on day 180. The response to therapy of the total population of 43 patients varied according to cases. The relative changes in p24 antigen compared to HIV-1 RNA also differed between patients. Measurement of HIV-1 viremia appears to be a valuable tool in current practice for individualizing therapy.

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Quantitatives molekulares Monitoring der HIV-1 RNA unter Behandlung mit anti-retroviralen Substanzen

Zusammenfassung 43 HIV-1 infizierte Patienten, die erstmals auf eine anti-retrovirale Therapie eingestellt oder deren Behandlung umgestellt wurde, wurden einem Monitoring hinsichtlich der HIV-1 RNA im Plasma unterzogen. Zur quantitativen Bestimmung der HIV-1 RNA in klinischen Proben oder eingefrorenem Plasma wurde ein Polymerasekettenreaktions(PCR)-Kit verwendet. Die Nachweisgrenze dieses Tests lag bei 200 RNA-Kopien/ml. Die erste Gruppe von 11 Patienten war auf eine Langzeit-Zidovudin-Monotherapie eingestellt und wurde auf Stavudin umgestellt. Trotz der Therapieänderung nahm die HIV-1 RNA-Menge bei drei Patienten mit sinkenden CD4⁺ T-Zell-Zahlen bei einer Verlaufszeit von 630 Tagen nicht ab. Die acht Patienten mit stabilen CD4⁺ T-Zell-Zahlen ließen im Mittel eine Abnahme der HIV-1 RNA-Kopien um 0,53 Log₁₀ zwischen Behandlungstag 90 und 650 erkennen. Die 2. Gruppe von 14 Patienten hatte eine Langzeitbehandlung mit Zidovudin in Monotherapie erhalten, die CD4⁺-T-Zell-Zahlen nahmen in den vergangenen 6 Monaten ab. Sie erhielten zusätzlich Lamivudin vom Tag 0 an. Innerhalb 30 Tagen trat eine rasche Reduktion der HIV-1 RNA-Kopien ein, am Tag 90 betrug die Abnahme 0,86 Log₁₀. Der Effekt blieb über eine mittlere Verlaufsbeobachtungszeit von 161 Tagen erhalten. Zugleich hatten am Tag 90 die CD4⁺ T-Zellen um 33/µl zugenommen. In einer dritten Gruppe wurden neun Patienten erstmals auf eine antiretrovirale Therapie eingestellt. In allen Fällen nahm die Zahl der HIV-1 RNA-Kopien rasch ab, in acht Fällen unter den Grenzwert bei mittlerer Behandlungszeit von 50,5 Tagen. Die CD4⁺ T-Zellen erhöhten sich um 164/µl am Tag 180. Die gesamte Patientengruppe zeigte ein individuell stark variierendes Therapieansprechen. Im p24 Antigen-Spiegel-Verhalten waren verglichen mit den HIV-1 RNA-Werten ebenfalls Unterschiede zu finden. Die Bestimmung der HIV-1 Virämie erwies sich als wertvolle Methode für die individuelle Therapieeinstellung.

     

Influenza virus binds its host cell using multiple dynamic interactions

Influenza virus belongs to a wide range of enveloped viruses. The major spike protein hemagglutinin binds sialic acid residues of glycoproteins and glycolipids with dissociation constants in the millimolar range [Sauter NK, et al. (1992) Biochemistry 31:9609-9621], indicating a multivalent binding mode. Here, we characterized the attachment of influenza virus to host cell receptors using three independent approaches. Optical tweezers and atomic force microscopy-based single-molecule force spectroscopy revealed very low interaction forces. Further, the observation of sequential unbinding events strongly suggests a multivalent binding mode between virus and cell membrane. Molecular dynamics simulations reveal a variety of unbinding pathways that indicate a highly dynamic interaction between HA and its receptor, allowing rationalization of influenza virus-cell binding quantitatively at the molecular level.     

New insights on interactions between HIV-1 and HSV-2

Herpes simplex type 2 (HSV-2) infection is common and frequently asymptomatic. Concerns exist about the high prevalence of HSV-2, particularly in areas of high HIV prevalence, because of observations that HSV-2 is associated with an increased risk of HIV acquisition, transmission, and disease progression. Several randomized trials have tested or are testing whether HSV-2 treatment can limit the spread of HIV, with mixed results. Although treatment with acyclovir, 400 mg twice daily, does not reduce HIV incidence, suppressive acyclovir and valacyclovir reduce HIV levels in plasma and in the genital tract. Ongoing trials are evaluating whether HSV suppression will reduce HIV transmission and disease progression. Until a protective HSV-2 or HIV vaccine is available, effective interventions that reduce the effect of HSV-2 on HIV transmission are critically needed.     

Dynamics of HIV-1 recombination in its natural target cells

Genetic recombination is believed to assist HIV-1 diversification and escape from host immunity and antiviral therapies, yet this process remains largely unexamined within the natural target-cell populations. We developed a method for measuring HIV-1 recombination directly that employs reporter viruses bearing functional enhanced yellow fluorescent protein (YFP) and enhanced cyan fluorescent protein (CFP) genes in which recombination produces a modified GFP gene and GFP fluorescence in the infected cells. These reporter viruses allow simultaneous quantification of the dynamics of HIV-1 infection, coinfection, and recombination in cell culture and in animal models by flow-cytometric analysis. Multiround infection assays revealed that productive cellular coinfection was subject to little functional inhibition. As a result, generation of recombinants proceeded according to the square of the infection rate during HIV-1 replication in T lymphocytes and within human thymic grafts in severe combined immunodeficient (SCID)-hu (Thy/Liv) mice. These results suggest that increases in viral load may confer a compounding risk of virus escape by means of recombinational diversification. A single round of replication in T lymphocytes in culture generated an average of nine recombination events per virus, and infection of macrophages led to approximately 30 crossover events, making HIV-1 up to an order of magnitude more recombinogenic than recognized previously and demonstrating that the infected cell exerts a profound influence on the frequency of recombination.     

Lectin-carbohydrate interactions and infectivity of human immunodeficiency virus type 1 (HIV-1).

The aim of this study was to determine whether mannosyl-specific lectins, especially Concanavalin A (ConA), may bridge HIV-1 env glycoproteins to cell membranes to increase virus binding to its targets, and to what extent this lectin-carbohydrate interaction can modify HIV-1 infectivity for monocytic compared with lymphoid cells. Monocytic U937 and lymphoid CEM cells, which both express surface mannose, were utilized. Whether first incubated with env glycoprotein or with the cells, lectins bound both to the cells and to radiolabeled recombinant gp160 (rgp160). Thus, they enhanced rgp160 adsorption to the cells in a methyl-alpha-mannose inhibitable manner. ConA did not appear to bind to the V1 domain of CD4 at the U937 cell surface since Leu3a binding was not blocked in the presence of ConA, nor was recombinant CD4 retained on a ConA-agarose affinity matrix. Moreover, enhanced rgp160 binding to the cells was CD4 independent, since it was not modified by preincubating the cells with Leu3a. Finally, ConA did not inhibit the binding of CD4-IgG3 chimeric molecules to virions immobilized on nitrocellulose membrane, which argues against the possibility that it interferes with the interaction of gp120 and CD4. However, both when incubated with the virus or with the cells and despite mediating enhanced binding of env glycoprotein, ConA neutralized HIV-1 infectivity for monocytic U937 as well as for lymphoid CEM cells. In this respect, ConA behaves like neutralizing antibodies which do not interfere with CD4 binding of gp120 but rather with some later event that leads to virus entry. These findings obtained with plant lectins may be of relevance in vivo, inasmuch as endogenous mannosyl-binding proteins, which are known to function as opsonins, have been reported to inhibit in vitro infection by HIV-1.     

乙型和丙型肝炎病毒与肝细胞之间相互作用的分子生物学机制研究进展

乙型肝炎病毒(HBV)和丙型肝炎病毒(HCV)的感染目前还没有特效的治疗方法和治疗药物.造成这一局面的原因是多种多样的,其中关于HBV和HCV与肝细胞之间相互作用的分子生物学机制的研究进展相对滞后,是严重影响新型治疗技术和新型药物的研究与开发重要的原因之一.目前多数情况下的抗肝炎病毒治疗,如干扰素等都还是非特异性的治疗方法,因此其治疗效果不满意.肝炎病毒与肝细胞之间相互作用的分子生物学机制,如肝炎病毒的特异性受体?HBV DNA与肝细胞基因组整合的机制及后果?肝炎病毒蛋白与肝细胞蛋白之间的相互作用?肝炎病毒蛋白反式激活作…     

Hepatitis C virus infection and thyroid autoimmune disorders:A model of interactions between the host and the environment

The hepatitis C virus(HCV) infection is an important public health problem and it is associated with hepatic and extrahepatic manifestations. Autoimmune thyroid diseases are common in HCV infected patients and the standard interferon-based treatment is associated with an increase of the immune-mediated thyroid damage. Recent evidence in the literature analyzed critical points of the mechanisms of thyroid damage, focusing on the balance between the two sides of the interaction: The environment(virus infection with potential crossreaction) and the host(susceptibility genes with consistent immune response). The spectrum of antiviral treatment for chronic HCV infection is rapidly expanding for the development of dual o triple therapy. The availability of interferon-free combined treatment with direct antiviral agents for HCV is very promising, in order to ameliorate the patient compliance and to reduce the development of thyroid autoimmunity.     

Kinetics of human immunodeficiency virus type 1 (HIV-1) DNA and RNA synthesis during primary HIV-1 infection.

HIV-1 replication and viral burden in peripheral blood mononuclear cells (PBMC) have been reported to be high in primary infection but generally very low during the prolonged period of clinical latency. It is uncertain precisely when this transition occurs during the HIV-1 infection and what the relationship is between the changes in HIV-1 replication versus the clearance of infected cells in the overall control of viral replication. In the present study, the kinetics of viral burden (i.e., frequency of HIV-1-infected cells) and replication during primary and early-chronic infection were analyzed in PBMC of four acutely infected individuals. High frequencies of HIV-1-infected cells and high levels of virus replication were observed in PBMC after primary HIV-1 infection. Down-regulation of virus replication in PBMC was observed in all four patients coincident with the emergence of HIV-1-specific immune responses. Other parameters of virus replication, such as circulating plasma p24 antigen and plasma viremia showed similar kinetics. In contrast, a significant decline in viral burden in PBMC was observed in only one of four patients. These results indicate that the down-regulation in the levels of virus replication associated with the clinical transition from acute to chronic infection does not necessarily reflect a reduction in viral burden, thus suggesting the involvement of additional factors. Identification of these factors will be important in elucidating the host mechanisms involved in the early control of HIV-1 infection and disease.     

Identification of cellular proteins that bind to the human immunodeficiency virus type 1 trans-activation-responsive TAR element RNA.

Human immunodeficiency virus type 1 (HIV-1) trans-activator protein Tat activates the expression of its viral long terminal repeat (LTR) through a target transactivation-responsive element termed TAR. We have constructed cell lines that constitutively express the HIV-1 Tat protein. Analyses of nuclear proteins from these cells and from matched control cells that do not express Tat have identified three proteins that bind to a radiolabeled HIV-1 TAR RNA probe. These polypeptides are 100 kDa, 62 kDa, and 46 kDa in size. Competition experiments using a wild-type TAR RNA sequence, a biologically inactive mutant sequence of TAR, and an unrelated RNA species demonstrated that these proteins show higher binding affinity to wild-type TAR than to the other two non-trans-activatable sequences. We hypothesize that these cellular proteins may mediate a function necessary in Tat-dependent activation of the LTR. The fact that no differences were seen in the binding profiles of nuclear proteins to TAR RNA in Tat-producing and Tat-nonproducing cells suggests that Tat does not directly interact with TAR.     

Human immunodeficiency virus 1 tat protein binds trans-activation-responsive region (TAR) RNA in vitro.

tat, the trans-activator protein for human immunodeficiency virus 1 (HIV-1), has been expressed in Escherichia coli from synthetic genes. Purified tat binds specifically to HIV-1 trans-activation-responsive region (TAR) RNA in gel-retardation, filter-binding, and immunoprecipitation assays. tat does not bind detectably to antisense TAR RNA sequences, cellular mRNA sequences, variant TAR RNA sequences with altered stem-loop structures, or TAR DNA.