On the inhibition of plant virus multiplication by ribavirin

The inhibition of the replication of potato virus X (PVX), belladonna mottle virus, tobacco mosaic virus, potato virus Y (PVY), and tobacco necrosis virus by ribavirin and pyrazofurin is described with emphasis on the inhibition of PVX by ribavirin. Ribavirin inhibits an early step of PVX replication. The inhibition is reversed to different degrees by all ribo- and deoxyribonucleosides, most strongly by thymidine. In tobacco leaves, nucleosides compete with ribavirin for phosphorylation to monophosphate by a nucleoside phosphotransferase. However, the final and main phosphorylation product of ribavirin is triphosphate. It is suggested that ribavirin triphosphate is the antiviral form and that it acts by inhibiting the capping of viral RNAs. This mode of action cannot be applied to the inhibition of PVY, the RNA of which is probably covalently linked to a protein at the 5'-terminus.     

Selective inhibition of virus multiplication by new acylated 1,2,4-triazole derivatives

Six out of 99 new acylated 1,2,4-triazole derivatives specifically inhibited rubella virus replication in RK 13 cell cultures. These are the following: 3-methylthio-5-(2-chlorobenzamido)-1H-1,2,4-triazole; 3-methylthio-5-(2-bromobenzamido)-1H-1,2,4-triazole; 3-methylthio-5-(2-methylbenzamido)-1H-1,2,4-triazole; 3-methylthio-5-(2-nitrobenzamido)-1H-1,2,4-triazole; 3-methylthio-5-(2-methylthiobenzamido)-1H-1,2,4-triazole and 3-ethylthio-5-(2-methylbenzamido)-1H-1,2,4-triazole. The compounds did not directly interfere with the infectivity of the rubella virus particles and the antiviral effect was demonstrable only within cells infected with rubella virus. The active compounds did not inhibit the replication of herpes simplex virus type 1, influenza virus and adenovirus in cell culture systems. Structure-activity relationships are discussed.     

LL-37 restored glucocorticoid sensitivity impaired by virus dsRNA in lung

Glucocorticoids play a key role in treatment of inflammatory lung diseases including both airway and parenchymal lung diseases. RNA viral infections are major causes of chronic lung disease exacerbations and can determine glucocorticoid resistance. The antibacterial peptide LL-37, the only member of human cathelicidin family, also functions as antiviral-activity enhancer. However, whether it can alleviate the glucocorticoid resistance caused by RNA viruses remains unclear. Here, we used type I (BEAS-2B) and type II (A549) lung epithelial cells to assess the effect of LL-37 on dsRNA-induced glucocorticoid resistance. We verified that LL-37 and polyinosinic-polycytidylic acid (poly I:C, a mimic of viral dsRNA) interact and enter both cell lines. Co-treatment with LL-37 and poly I:C increased glucocorticoid-induced expression of promyelocytic leukemia zinc finger (PLZF), an anti-inflammatory protein, compared to poly I:C alone. Pre-treatment with LL-37 also restored transactivation of the glucocorticoid response element (GRE). Moreover, LL-37 rescued poly I:C-induced glucocorticoid resistance by increasing phosphorylation and nuclear translocation of glucocorticoid receptor. Importantly, LL-37 downregulated poly I:C-induced Erk and Akt signaling pathways in lung epithelial cells. Finally, we verified our data in vivo, showing that mCRAMP, the mouse LL-37 ortholog, can alleviate poly I:C-induced glucocorticoid insensitivity in a murine asthma model. In summary, this study showed that LL-37 restored glucocorticoid sensitivity impaired by dsRNA possibly by inhibiting Akt pathway, in addition to Erk1/2 pathway. These findings suggest LL-37 as a therapeutic agent for treatment of viral infections in inflammatory pulmonary diseases.     

Bovine leukemia virus inhibition in vitro by ribavirin

Ribavirin treatment of an established monolayer of fetal lamb kidney (FLK) cells chronically infected with bovine leukemia virus (BLV) resulted in inhibition of the extra- and intracellular internal viral polypeptide antigen, as measured by CF test, at concentrations of 10 and 32 μg/ml, respectively. Similar treatment of F-81 cells newly infected with BLV caused significant reduction in viral syncytia formation at ribavirin levels as low as 3.2 μg/ml. Attempts to eliminate or reduce the BLV infection in FLK cells by 8 passages of the cells in the continual presence of 3.2 or 1.0 μg/ml of ribavirin were unsuccessful. Multiple passages of FLK cells in the presence of higher concentrations of ribavirin substantially retarded cell growth, although short-term treatment of established cell monolayers appeared to be well tolerated as evidenced by cell appearance. Biochemical cytostatic studies of resting F-81 cell monolayers showed cytostatic effects at the same dosage levels where antiviral effects occurred.     

Therapeutic developments in matrix metalloproteinase inhibition

A family of zinc-dependent endopeptidases called matrix metalloproteinases (MMPs) have the capacity to degrade all elements of the extracellular matrix (ECM) and are required for homeostatic maintenance of the ECM. However, interest in MMPs predominantly arises from the accumulating evidence implicating that dysregulated MMP expression plays a role in mediating or accompanying a diverse array of pathologies. These include tumour invasion and metastasis and inflammatory diseases characterised by excessive tissue destruction, such as arthritis, periodontal disease, atherosclerosis, plaque rupture, arterial aneurysms, postmyocardial infarction, ventricular remodelling and cardiac rupture. Several patents representing therapeutic drugs and strategies to treat the associated conditions have been claimed, some resulting in clinical drug trials. This review will: i) summarise the current status of our understanding of MMPs and how they participate in normal and functional ECM degradation; ii) review therapeutic efforts to favourably alter the balance between MMP proteolysis and ECM sythesis; and iii) critically evaluate recent studies that have importantly advanced our understanding of the complexities of MMP function and propose areas where future efforts to develop therapeutic strategies might be most beneficial and productive.     

Lead exposure enhances virus multiplication and pathogenesis in mice

Suppression of the immune system by environmental xenobiotics may cause increased susceptibility of the host towards a variety of microbial pathogens an result in a life-threatening state. We investigated whether lead exposure would enhance susceptibility to Semliki Forest Virus (SFV). Mice orally exposed to lead acetate (62.5, 125, 250 or 500 mg/kg bw) exhibited increased mortality and decreased mean survival time compared to untreated animals on challenge with SFV. The mortality was associated with enhancement of high virus titer and earlier appearance of virus in lead-exposed mice. Histopathological studies observed enhancement of viral pathogenesis in a dose dependent pattern in the lead-dosed group challenged with SFV. The results indicate that exposure to lead enhanced susceptibility to viral infection. Environmental metal contamination and subsequent infection by pathogenic microbes points necessitate studies on the interaction of environmental pollutants on the immune system.     

Some particularities of Sendai virus multiplication in chorioallantoic membrane fragments

Marked differences were recorded in the kinetics of infectant and hemagglutinating (HA) activities of parinfluenza type 1 (Sendai) virus grown in chorioallantoic membrane (CAM) fragments in roller tubes with or without daily changes of the culture medium. The differences were dependent on cultivation conditions, the time interval post inoculation and the state of the CAM cells. Taking into account these conditions, it is possible to obtain either highly infectant virus or virus with low infectivity but high HA activity.     

Therapeutic inhibition of leukocyte recruitment in inflammatory diseases

The ingress of leukocytes into sites of inflammation is crucial for the pathogenesis of arthritis and other inflammatory conditions. Chemokines, their receptors and cellular adhesion molecules (CAMs) are involved in this process. Here, the roles of the most relevant chemokines, chemokine receptors and CAMs are briefly reviewed. There have been several attempts to target chemokine- and CAM-mediated pathways in preclinical studies using animal models of arthritis, and in a limited number of human studies. In this review, the most recent advances in anti-chemokine and anti-adhesion therapeutic strategies are summarized.     

Therapeutic inhibition of CXCR2 by Reparixin attenuates acute lung injury in mice

Background and purpose:

Acute lung injury (ALI) remains a major challenge in critical care medicine. Both neutrophils and chemokines have been proposed as key components in the development of ALI. The main chemokine receptor on neutrophils is CXCR2, which regulates neutrophil recruitment and vascular permeability, but no small molecule CXCR2 inhibitor has been demonstrated to be effective in ALI or animal models of ALI. To investigate the functional relevance of the CXCR2 inhibitor Reparixin in vivo, we determined its effects in two models of ALI, induced by either lipopolysaccharide (LPS) inhalation or acid instillation.

Experimental approach:

In two ALI models in mice, we measured vascular permeability by Evans blue and evaluated neutrophil recruitment into the lung vasculature, interstitium and airspace by flow cytometry.

Key results:

Pharmacological inhibition of CXCR2 by Reparixin reduced CXCL1-induced leukocyte arrest in the microcirculation of the cremaster muscle, but did not influence arrest in response to leukotriene B₄ (LTB₄) demonstrating specificity. Reparixin (15 μg g⁻¹) reduced neutrophil recruitment in the lung by approximately 50% in a model of LPS-induced ALI. A higher dose did not provide additional reduction of neutrophil recruitment. This dose also reduced accumulation of neutrophils in the interstitial compartment and vascular permeability in LPS-induced ALI. Furthermore, both prophylactic and therapeutic application of Reparixin improved gas exchange, and reduced neutrophil recruitment and vascular permeability in a clinically relevant model of acid-induced ALI.

Conclusions and implications:

Reparixin, a non-competitive allosteric CXCR2 inhibitor attenuates ALI by reducing neutrophil recruitment and vascular permeability.     

Polymer-bound 6' sialyl-N-acetyllactosamine protects mice infected by influenza virus

To develop a mouse model for testing receptor attachment inhibitors of human influenza viruses, the human clinical virus isolate in MDCK cells A/NIB/23/89M (H1N1) was adapted to mice by serial passaging through mouse lungs. The adaptation enhanced the viral pathogenicity for mice, but preserved the virus receptor binding phenotype, preferential binding to 2-6-linked sialic acid receptors and low affinity for 2-3-linked receptors. Sequencing of the HA gene of the mouse-adapted virus A/NIB/23/89-MA revealed a loss of the glycosylation sites in positions 94 and 163 of HA1 and substitutions 275Asp-->Gly in HA1 and 145Asn-->Asp in HA2. The four mouse strains tested differed significantly in their sensitivity to A/NIB/23/89-MA with the sensitivity increasing in the order of BALB/cJCitMoise, C57BL/6LacSto, CBA/CaLacSto and A/SnJCitMoise strains. Testing of protective efficacy of the polyacrylamide conjugate bearing Neu5Acalpha2-6Galbeta1-4GlcNAc trisaccharide under conditions of lethal or sublethal virus infection demonstrated a strong protective effect of this preparation. In particular, aerosol treatment of mice with the polymeric attachment inhibitor on 24-110 h after infection completely prevented mortality in sensitive animals and lessened disease symptoms in more resistant mouse strains.     

呼吸道合胞病毒感染豚鼠动物模型的建立

目的 建立呼吸道合胞病毒（respiratory syncytial virus,RSV)感染豚鼠致细支气管炎、肺炎的动物模型，观察肺部的病理变化。方法 1个月大小的豚鼠，分为病毒接种组（30只）和对照组（10只），病毒接种组豚鼠鼻腔接种RSV病毒悬液，对照组接种PBS液，分别于接种后6、14天取肺组织进行光镜与电镜检查，并用组织培养法分离病毒，阳性结果通过免疫荧光法鉴定。结果 病毒接种组豚鼠肺组织均出现细支气管炎、肺炎的病理改变，接种后14天有3例为阳性。结论 豚鼠鼻腔接种RSV可以建立细支气管炎，肺炎的动物模型，研究其发病机理及RSV感染在气道高反应性产生中的作用。     

Effect of ceruloplasmin on Sendai virus multiplication in chorioallantoic membrane fragments.

Three stages can be distinguished in the evolution of HA titer during Sendai virus multiplication in chorioallantoic membrane (CAM) fragments in relation to the age of CAM and conditions of inoculation and maintenance. Ceruloplasmin has an inhibitory effect on virus multiplication, more marked in the case of stationarily maintained CAM fragments and dependent on the relationship between the moment of ceruloplasmin addition and of virus inoculation. Maximum inhibition is achieved by inoculation of Sendai virus (1 Ha u) after 4 1/2-hour preincubation with ceruloplasmin (5 x 10(-7)M). The experimental data plead for the formation of a virus-ceruloplasmin complex able to inhibit both penetration and release of virus synthesized in the cell.     

Potent antiviral activity of amprenavir in primary macrophages infected by human immunodeficiency virus

Objective of the present study was then to assess the antiviral activity of the protease inhibitor amprenavir in macrophages (M/M), and to compare it with its efficacy in peripheral blood lymphocytes (PBL). M/M were obtained from blood of sero-negative healthy donors and infected with M-tropic HIV-1 strain (HIV-1(Ba-L)). The stabilized infection was assessed by monitoring the HIV-1 p24 gag antigen production in the supernatants of M/M cultures. In the setting of acute infection (treatment before HIV-1 challenge), amprenavir showed substantial activity both in M/M and PBL at similar concentrations (EC(50): 0.011 and 0.031 microM, respectively); complete inhibition of HIV-1 replication was achieved in both cell types at concentration of about 2 microM. In the setting of chronical infection (i.e. antiviral treatment several days after established infection), an antiviral effect of amprenavir was achieved in M/M, but at concentrations higher than those active in acutely infected M/M (EC(50): 0.72 microM, EC(90): 18.2 microM). The antiviral effect in chronically infected M/M was sustained for at least 2 weeks of continuous treatment. These findings suggest that amprenavir (at relatively high concentrations) has a clinically relevant antiviral effect in persistently infected reservoirs of HIV.     

对虾白斑综合征病毒胸腺嘧啶核苷酸合成酶基因的系统进化分析

目的为了对对虾白斑综合征病毒 (whitespotsyndromevirus,WSSV)中最为保守的胸腺嘧啶核苷酸合成酶 (WSV TS)基因 ,进行该病毒的系统发育学研究。方法通过GenBankTM 数据库和DNAMAN分析系统 ,对不同生物来源的胸腺嘧啶核苷酸合成酶进行同源性比较并构建了该基因的系统发育进化树。结果 :WSV TS的氨基酸序列与来自高等生物 ,特别是同哺乳动物有着很高的同源性 ,而与多种病毒的同源性较低。结论WSV ts基因是WSSV基因组中最为保守的基因 ,其进化树的构建对于理解WSSV的进化地位有重要的帮助。     

Neutropenia in patients infected with human immunodeficiency virus

The possible mechanisms of neutropenia associated with both human immunodeficiency virus (HIV) infection and drug treatment in adults are examined, and the current and investigational strategies for managing neutropenia are reviewed. Neutropenia associated with HIV arises from diverse mechanisms, including cellular immune dysfunction, direct effects on progenitor cells, humoral immune dysfunction, and vitamin deficiencies. Drug-induced neutropenia may be related to direct cytotoxic effects, immunologic mediators, and the effects of vitamin depletion on the bone marrow. Bone marrow toxicity in patients receiving zidovudine appears to be more frequent in those patients with advanced disease, low CD4 cell counts, a pretreatment anemia, low serum vitamin B12 levels, and low or low normal serum folic acid levels. Patients with AIDS also are at increased risk for adverse events associated with folate antagonists and sulfonamides compared with other patient populations. Lithium therapy has improved neutrophil counts in patients receiving zidovudine; however, the toxicities associated with use of lithium, combined with the lower dosages of zidovudine now recommended, may obviate its use. The use of colony-stimulating factors appears promising for increasing the number and function of circulating neutrophils. Although concomitant use of interferon alfa and zidovudine may result in a strong synergistic anti-HIV effect, dose-limiting neutropenia has been reported in patients receiving the combination. There are currently no controlled data assessing the effectiveness of intravenous immune globulin in the treatment of HIV-related or drug-related neutropenia. In evaluating neutropenia, the clinician must attempt to discern whether the neutropenia is more likely related to disease state(s) or drug therapies. Potential management strategies include modulation of the disease state, discontinuation or dose reduction of the offending agent, or administration of exogenous immune enhancer.     

Therapeutic inhibition of the renin angiotensin aldosterone system

Background: Inhibitors of the renin angiotensin aldosterone system (RAAS) represent some of the most widely prescribed, successful and well-tolerated therapeutics of recent times and are of proven worth in the management/prevention of cardiovascular disease and diabetic nephropathy. However, as knowledge has grown about the RAAS and its manifold alternate pathways, loci of action and dynamic response to inhibition, so has the clinical debate as how to best use existing therapeutics as well as how best to conceptualise and design RAAS inhibitors of the future. Objective: To provide an overview of the several points of therapeutic anti-RAAS intervention, many of which have already been exploited from ‘upstream’ renin inhibition to ‘midstream’ ACE inhibition to ‘downstream’ angiotensin AT1 receptor blockade. Methods: A search of patents for RAAS inhibitors recorded in 2008 was conducted along with a relevant literature search. Results/conclusion: Each intervention has merits and demerits with implications for RAAS ‘escape’ phenomena, ‘dual inhibition’ therapy, long-term clinical efficacy and adverse drug reactions. Renin inhibitors offer the most complete RAAS inhibition, but more downstream interventions are likely to recruit supplementary anti-RAAS mediators and receptor signalling pathways. Furthermore, managing hyperkalaemia-stimulated aldosterone escape during combined ACE inhibitor and angiotensin receptor blocker treatment may realise the full clinical potential of dual inhibition therapy.     

Use of the yellow fever virus vaccine strain 17D for the study of strategies for the treatment of yellow fever virus infections

We have employed the attenuated vaccine strain 17D of yellow fever virus (YFV) to evaluate the inhibitory effect of a selected series of compounds on YFV in Vero cells. Use of the vaccine strain does not require high-level microbiological containment facilities and should allow extensive screening. In addition, YFV may serve as a model for other flaviviruses including hepatitis C virus (HCV), and thus strategies for the treatment of YFV infections may apply to flavivirus infections in general. In the present study, several compounds belonging to different classes of nucleoside analogues and polyanions were evaluated for their inhibitory effect on the replication of YFV. Compounds that are targeted at: (i) IMP dehydrogenase (ribavirin, EICAR, tiazofurin, selenazofurin and mycophenolic acid), (ii) OMP decarboxylase (pyrazofurin and 6-azauridine), (iii) CTP synthetase (carbodine and cyclopentenyl cytosine), (iv) dihydrofolate reductase (methotrexate) and the (v) sulfated polymers (dextran sulfate and PAVAS) proved inhibitory to the replication of YFV. Mycophenolic acid (EC₅₀: 0.08 μg/ml), EICAR (EC₅₀: 0.8 μ/ml) and methotrexate (EC₅₀: 0.07 μg/ml) were the most effective. The finding that EICAR and mycophenolic acid, despite their potent anti-YFV activity, had little or no effect on the replication of the bunyavirus Punta Toro or herpes simplex virus in Vero cells, indicates that their anti-YFV activity is rather specific and does not merely result from cytotoxicity. Inhibitors of S-adenosylhomocysteine hydrolase (SAH hydrolase) and thymidylate synthase were found to be devoid of anti-YFV activity.