Inhibition of lung serine proteases in mice: a potentially new approach to control influenza infection

<p>Abstract</p> <p>Background</p> <p>Host serine proteases are essential for the influenza virus life cycle because the viral haemagglutinin is synthesized as a precursor which requires proteolytic maturation. Therefore, we studied the activity and expression of serine proteases in lungs from mice infected with influenza and evaluated the effect of serine protease inhibitors on virus replication both in cell culture and in infected mice.</p> <p>Results</p> <p>Two different inbred mouse strains were investigated: DBA/2J as a highly susceptible and C57Bl/6J as a more resistant strain to influenza virus infection. The serine proteases from lung homogenates of mice exhibited pH optima of 10.00. Using the substrate Bz-Val-Gly-Arg-<it>p</it>-nitroanilide or in zymograms, the intensities of proteolysis increased in homogenates from both mouse strains with time post infection (p.i.) with the mouse-adapted influenza virus A/Puerto Rico/8/34 (H1N1; PR8). In zymograms at day 7 p.i., proteolytic bands were stronger and numerous in lung homogenates from DBA/2J than C57Bl/6J mice. Real-time PCR results confirmed differential expression of several lung proteases before and after infecting mice with the H1N1 virus. The most strongly up-regulated proteases were <it>Gzma</it>, <it>Tmprss4</it>, <it>Elane</it>, <it>Ctrl</it>, <it>Gzmc </it>and <it>Gzmb</it>. Pretreatment of mouse and human lung cell lines with the serine protease inhibitors AEBSF or <it>p</it>AB or a cocktail of both prior to infection with the H1N1 or the A/Seal/Massachusetts/1/80 (H7N7; SC35M) virus resulted in a decrease in virus replication. Pretreatment of C57Bl/6J mice with either AEBSF or a cocktail of AEBSF and <it>p</it>AB prior to infection with the H1N1 virus significantly reduced weight loss and led to a faster recovery of treated versus untreated mice while <it>p</it>AB alone exerted a very poor effect. After infection with the H7N7 virus, the most significant reduction of weight loss was obtained upon pretreatment with either the protease inhibitor cocktail or <it>p</it>AB. Furthermore, pretreatment of C57BL/6J mice with AEBSF prior to infection resulted in a significant reduction in the levels of both the H1N1 and H7N7 nucleoproteins in mice lungs and also a significant reduction in the levels of the HA transcript in the lungs of the H1N1- but not the H7N7-infected mice.</p> <p>Conclusion</p> <p>Multiple serine protease activities might be implicated in mediating influenza infection. Blocking influenza A virus infection in cultured lung epithelia and in mice by the used serine protease inhibitors may provide an alternative approach for treatment of influenza infection.</p

Gene expression patterns during adaptation of a helminth parasite to different environmental niches

Using a genomic microarray, gene expression at three different developmental stages of the schistosome parasite were analyzed, resulting in the identification of 1154 developmentally enriched transcripts

Serodiagnosis of Schistosoma mansoni infections in an endemic area of Burkina Faso: performance of several immunological tests with different parasite antigens.

The performance of indirect haemagglutination assays (IHA), enzyme-linked immunosorbent assays (ELISA) and indirect immunofluorescent antibody tests (IFAT) were compared with 450 sera from a Schistosoma mansoni-endemic area in Burkina Faso. All participants in this survey provided at least one sample each of stool, urine and serum. From those with an egg-negative Kato-Katz thick smear, a second stool sample was examined. IHA was based on either extracts of adult S. mansoni worms (SmIHA) or S. japonicum egg antigen (SjIHA). For ELISA, three antigen preparations were used, namely: (i) soluble S. mansoni adult worm antigens (SWAP); (ii) soluble S. mansoni egg antigens (SEA); and (iii) a cationic exchange fraction of S. mansoni eggs (CEF6). IFAT was performed with S. mansoni male worm sections. Among the egg-excretors, the sensitivity of ELISA was high and egg antigens performed slightly better (SEA, 96%; CEF6, 97%) than worm antigen (94%). Sensitivity of IHA was satisfactory with homologous (Sm, >85%), but not heterologous (Sj, 56%) parasite antigen. In IFAT, the parenchyma-associated fluorescence showed high sensitivity (95%), but gut-associated fluorescence, which is known to be a sensitive diagnostic marker for schistosome-infected European travelers, was observed only in 76% of a sub-sample of 100 of the endemic sera. Among sera from egg-negative individuals, many gave positive reactions in several or all of the tests employed. These reactions (formally "false positive") are considered to represent true infections, since chemotherapy had not yet been delivered to this population. For the purpose of further surveys in Burkina Faso or other resource-poor settings, we suggest IHA as an accurate diagnostic test and propose to further improve its performance by including egg rather than worm antigens

Electrowinning of Non-Noble Metals with Simultaneous Hydrogen Evolution at Flow-Through Porous Electrodes II. Experimental

This paper presents an interpretation of the experimental results obtained on the electrowinning of zinc at a flow-through porous electrode in light of a mathematical model which was presented in Part I. The process is accompanied by simultaneous hydrogen evolution within the electrode, which increases the pore electrolyte resistivity and decreases the coulombic efficiency. We measured polarization curves, coulombic efficiencies, and current distributions under various conditions of zincate concentrations, flow rates, cell current, and electrode thickness. Reasonable agreement between the measured and predicted current distributions was obtained only under conditions of high electrolyte flow rates, low cell currents, and thinner electrodes. The deviations observed at low electrolyte flow rates and high cell currents are attributed to the agitating effects of the hydrogen gas bubbles, which enhance the local mass-transfer coefficient. This effect was not incorporated in the model due to the absence of adequate correlations

Immune responses to killed reassorted influenza virus supplemented with natural adjuvants

In this study, we investigated the immunomodulatory effects of a supplemented killed influenza virus (V) by Echinacea purpurea (E) and Nigella sativa (N) extracts and effect of changing the route of immunization from intramuscular (IM) to intraperitoneal (IP). At the 2nd-, 3rd- and 4th-week post-IM immunizations (WPIMI), the supplemented V with N (VN) induced the most significant IgM response unlike N alone. At the 2nd WPIMI, V or VN induced the highest significant IgG levels. At the 2nd-week post-IP immunization (WPIPI), E and VN induced the most significant IgG levels. Both at the 3rd and 4th WPIMI or WPIPI, various treatments induced significant increases in IgG. At the 4th WPIMI, E, V, and V with E (VE) induced significant increases in the CD4+ thymocytes while all IP treatments caused significant increase in their counts. V and VN induced the most significant IM induction of CD8+ thymocytes while their best IP stimulation was induced by N, VE, and VN. At the 4th WPIMI, various treatments caused significant increases in the mesenteric lymph node (MLN) CD4+, CD8+ counts. WPIPI with V or VE caused significant increases in both the CD4+- and CD8+ MLN cells, whereas VN significantly induced CD8+ MLN cells only. WPIPI with various treatments caused significant increases in the B-cell counts and the peak was obtained by VN

SmCL3, a Gastrodermal Cysteine Protease of the Human Blood Fluke Schistosoma mansoni

Parasitic infection caused by blood flukes of the genus Schistosoma is a major global health problem. More than 200 million people are infected. Identifying and characterizing the constituent enzymes of the parasite's biochemical pathways should reveal opportunities for developing new therapies (i.e., vaccines, drugs). Schistosomes feed on host blood, and a number of proteolytic enzymes (proteases) contribute to this process. We have identified and characterized a new protease, SmCL3 (for Schistosoma mansoni cathepsin L3), that is found within the gut tissue of the parasite. We have employed various biochemical and molecular biological methods and sequence similarity analyses to characterize SmCL3 and obtain insights into its possible functions in the parasite, as well as its evolutionary position among cathepsin L proteases in general. SmCL3 hydrolyzes major host blood proteins (serum albumin and hemoglobin) and is expressed in parasite life stages infecting the mammalian host. Enzyme substrate specificity detected by positional scanning-synthetic combinatorial library was confirmed by molecular modeling. A sequence analysis placed SmCL3 to the cluster of other cathepsins L in accordance with previous phylogenetic analyses

Interaction Between the Neglected Tropical Disease Human Schistosomiasis and HCV Infection in Egypt: a Puzzling Relationship

Abstract Egypt has the highest prevalence of chronic hepatitis C virus (HCV) infection and seropositivity worldwide, and it has been proposed that this enhanced susceptibility to HCV is related to coinfection with schistosomiasis. Although currently, there are no studies regarding the actual prevalence of both human schistosomiasis and schistosomiasis/HCV coinfection evidences strongly support that eliminating human schistosomiasis from Egypt is necessary to reduce both HCV prevalence and liver pathology. The present review highlights the significant impact of the neglected tropical disease human schistosomiasis on both susceptibility of Egyptians to HCV coinfection, severity of the resulting liver pathology, and poor response to antiviral therapy. The immune evasion mechanisms exerted by the HCV-NS3/4A protease domain, and the possible impact of immune evasion mechanisms exerted by proteases of larval, worm and egg stages of the parasite Schistosoma on human susceptibility to HCV infection are discussed. In addition, schistosome immune evasion mechanisms may include immunosuppression that in turn prevents clearance of HCV viremia and leads to relapsing HCV infection and severe liver pathology. I propose the generation of a replicon system from the most prevailing genotype (HCV4a) in Egypt and establishing its replication on hepatoplastoma or immune cells in presence of bilharzial antigens. Finally, the use of a humanized small animal model that can acquire both HCV and S. mansoni infections will be important to further understand in real time the impact of coinfection on both the immune system and liver pathology