Immunization with recombinant adenovirus synthesizing the secretory form of Japanese encephalitis virus envelope protein protects adenovirus-exposed mice against lethal encephalitis

Replication-defective recombinant adenoviruses (RAds) were constructed that synthesized the pre-membrane and envelope (E) proteins of Japanese encephalitis virus (JEV). Recombinant virus RAdEa synthesized Ea, the membrane-anchored E protein, and RAdEs synthesized Es, the secretory E protein. Compared with RAdEs, RAdEa replicated poorly in HEK 293A cells and synthesized lower amounts of E protein. Oral immunization of mice with RAds generated low titers of anti-JEV antibodies that had little JEV neutralizing activity. Intra-muscular (IM) immunization of mice with either RAd generated high titers of anti-JEV antibodies. Interestingly, RAdEa induced only low titers of JEV neutralizing antibodies. Titers were significantly higher in case of RAdEs immunization. Splenocytes from mice immunized IM with RAds secreted large amounts of interferon-γ and moderate amounts of interleukin-5 in the presence of JEV and showed cytotoxic activity against JEV-infected cells. Naïve mice immunized IM with RAdEs showed complete protection against a lethal dose of JEV given intra-cerebrally. In order to study the effect of the pre-existing adenovirus 5 (Ad5) immunity on the outcome of the RAdEs immunization, mice were exposed to Ad5 through IM or intra-nasal (IN) routes before immunization with RAdEs. Mice exposed to Ad5 through the IN route, when immunized with RAdEs given IM, or those exposed to Ad5 through the IM route, when immunized with RAdEs given IN, were completely protected against lethal JEV challenge.     

Divergence of larval resource acquisition for water conservation and starvation resistance in Drosophila melanogaster

Laboratory selection experiments have evidenced storage of energy metabolites in adult flies of desiccation and starvation resistant strains of D. melanogaster but resource acquisition during larval stages has received lesser attention. For wild populations of D. melanogaster, it is not clear whether larvae acquire similar or different energy metabolites for desiccation and starvation resistance. We tested the hypothesis whether larval acquisition of energy metabolites is consistent with divergence of desiccation and starvation resistance in darker and lighter isofemale lines of D. melanogaster. Our results are interesting in several respects. First, we found contrasting patterns of larval resource acquisition, i.e., accumulation of higher carbohydrates during 3rd instar larval stage of darker flies versus higher levels of triglycerides in 1st and 2nd larval instars of lighter flies. Second, 3rd instar larvae of darker flies showed ~40?h longer duration of development at 21°C; and greater accumulation of carbohydrates (trehalose and glycogen) in fed larvae as compared with larvae non-fed after 150?h of egg laying. Third, darker isofemale lines have shown significant increase in total water content (18%); hemolymph (86%) and dehydration tolerance (11%) as compared to lighter isofemale lines. Loss of hemolymph water under desiccation stress until death was significantly higher in darker as compared to lighter isofemale lines but tissue water loss was similar. Fourth, for larvae of darker flies, about 65% energy content is contributed by carbohydrates for conferring greater desiccation resistance while the larvae of lighter flies acquire 2/3 energy from lipids for sustaining starvation resistance; and such energy differences persist in the newly eclosed flies. Thus, larval stages of wild-caught darker and lighter flies have evolved independent physiological processes for the accumulation of energy metabolites to cope with desiccation or starvation stress.     

Highly enhanced adsorption for decontamination of lead ions from battery wastewaters using chitosan functionalized with xanthate

Decontamination of lead ions from aqueous media has been investigated using cross linked xanthated chitosan (CMC) as an adsorbent. Various physico-chemical parameters such as contact time, amount of adsorbent, concentration of adsorbate were optimized to simulate the best conditions which can be used to decontaminate lead from aqueous media using CMC as an adsorbent. The atomic absorption spectrometric technique was used to determine the distribution of lead. Maximum adsorption was observed at both pH 4 and 5. The adsorption data followed both Freundlich and Langmuir isotherms. Langmuir isotherm gave a saturated capacity of 322.6+/-1.2mg/g at pH 4. From the FTIR spectra analysis, it was concluded that xanthate and amino group participate in the adsorption process. The developed procedure was successfully applied for the removal of lead ions from real battery wastewater samples.     

Molecular and computational approaches to understand resistance of New Delhi metallo β-lactamase variants (NDM-1, NDM-4, NDM-5, NDM-6, NDM-7)-producing strains against carbapenems

The discovery of NDM-1 and its variants has caused the emergence of antibiotic resistance in the community and hospital setting, causing major concern for health care across the globe. New Delhi Metallo-β-lactamase is known to hydrolyse almost all β-lactam antibiotics. Studies have shown the hydrolytic activates of NDM-1 and some of its variants, however a comparative study of these NDM variants has not been explored in detail. Hence, we proposed to check their catalytic activity by performing a comparative study between NDM-1 and its variants. The study was initiated to clone NDM variants (NDM-1, NDM-4, NDM-5, NDM-6 and NDM-7) followed by overexpression of the recombinant proteins to check their hydrolytic properties against β-lactam antibiotics. The minimum inhibitory concentration of carbapenems antibiotics for bla_NDM-5 clone was found four fold increased, whereas no change was observed in the clones having other variants. The hydrolytic activity of carbapenem with NDM-5 variant was found to be augmented as per the kinetics parameter where Km was decreased and kcat, kcat/Km values increased as compared to the NDM-1. Molecular docking studies were employed to identify the variations in the binding ability among all NDM variants with imipenem or meropenem. Simulation studies at 100?ns showed a good stability of NDM-5 with imipenem and meropenem as compared to NDM-1. CD spectroscopy data revealed significant changes in the secondary structure of NDM variants. We conclude that NDM-5 showed higher hydrolytic activity as compared to other variants. This study provides a comparative analysis of the severity of NDM producing strains.     

Nonsteroidal Anti-inflammatory Drugs Diclofenac and Celecoxib Attenuates Wnt/β-Catenin/Tcf Signaling Pathway in Human Glioblastoma Cells

Glioblastoma, the most common and aggressive primary brain tumors, carry a bleak prognosis and often recur even after standard treatment modalities. Emerging evidence suggests that deregulation of the Wnt/β-catenin/Tcf signaling pathway contributes to glioblastoma progression. Nonsteroidal anti-inflammatory drugs (NSAIDs) inhibit tumor cell proliferation by suppressing Wnt/β-catenin/Tcf signaling in various human malignancies. In this study, we sought to inhibit Wnt/β-catenin/Tcf signaling in glioblastoma cells by the NSAIDs diclofenac and celecoxib. Both diclofenac and celecoxib significantly reduced the proliferation, colony formation and migration of human glioblastoma cells. Diclofenac and celecoxib downregulated β-catenin/Tcf reporter activity. Western and qRT-PCR analysis showed that diclofenac and celecoxib reduced the expression of β-catenin target genes Axin2, cyclin D1 and c-Myc. In addition, the cytoplasmic accumulation and nuclear translocation of β-catenin was significantly reduced following diclofenac and celecoxib treatment. Furthermore, diclofenac and celecoxib significantly increased phosphorylation of β-catenin and reduced the phosphorylation of GSK3β. These results clearly indicated that diclofenac and celecoxib are potential therapeutic agents against glioblastoma cells that act by suppressing the activation of Wnt/β-catenin/Tcf signaling.     

Removing the threat of diclofenac to critically endangered Asian vultures

Veterinary use of the nonsteroidal anti-inflammatory (NSAID) drug diclofenac in South Asia has resulted in the collapse of populations of three vulture species of the genusGyps to the most severe category of global extinction risk. Vultures are exposed to diclofenac when scavenging on livestock treated with the drug shortly before death. Diclofenac causes kidney damage, increased serum uric acid concentrations, visceral gout, and death. Concern about this issue led the Indian Government to announce its intention to ban the veterinary use of diclofenac by September 2005. Implementation of a ban is still in progress late in 2005, and to facilitate this we sought potential alternative NSAIDs by obtaining information from captive bird collections worldwide. We found that the NSAID meloxicam had been administered to 35 captiveGyps vultures with no apparent ill effects. We then undertook a phased programme of safety testing of meloxicam on the African white-backed vultureGyps africanus, which we had previously established to be as susceptible to diclofenac poisoning as the endangered AsianGyps vultures. We estimated the likely maximum level of exposure (MLE) of wild vultures and dosed birds by gavage (oral administration) with increasing quantities of the drug until the likely MLE was exceeded in a sample of 40G. africanus. Subsequently, sixG. africanus were fed tissues from cattle which had been treated with a higher than standard veterinary course of meloxicam prior to death. In the final phase, ten Asian vultures of two of the endangered species(Gyps bengalensis,Gyps indicus) were dosed with meloxicam by gavage; five of them at more than the likely MLE dosage. All meloxicam-treated birds survived all treatments, and none suffered any obvious clinical effects. Serum uric acid concentrations remained within the normal limits throughout, and were significantly lower than those from birds treated with diclofenac in other studies. We conclude that meloxicam is of low toxicity toGyps vultures and that its use in place of diclofenac would reduce vulture mortality substantially in the Indian subcontinent. Meloxicam is already available for veterinary use in India.     

Pleiotrophin Gene Therapy for Peripheral Ischemia: Evaluation of Full-Length and Truncated Gene Variants

Pleiotrophin (PTN) is a growth factor with both pro-angiogenic and limited pro-tumorigenic activity. We evaluated the potential for PTN to be used for safe angiogenic gene therapy using the full length gene and a truncated gene variant lacking the domain implicated in tumorigenesis. Mouse myoblasts were transduced to express full length or truncated PTN (PTN or T-PTN), along with a LacZ reporter gene, and injected into mouse limb muscle and myocardium. In cultured myoblasts, PTN was expressed and secreted via the Golgi apparatus, but T-PTN was not properly secreted. Nonetheless, no evidence of uncontrolled growth was observed in cells expressing either form of PTN. PTN gene delivery to myocardium, and non-ischemic skeletal muscle, did not result in a detectable change in vascularity or function. In ischemic hindlimb at 14 days post-implantation, intramuscular injection with PTN-expressing myoblasts led to a significant increase in skin perfusion and muscle arteriole density. We conclude that (1) delivery of the full length PTN gene to muscle can be accomplished without tumorigenesis, (2) the truncated PTN gene may be difficult to use in a gene therapy context due to inefficient secretion, (3) PTN gene delivery leads to functional benefit in the mouse acute ischemic hindlimb model.     

Characterization and Cytotoxic Evaluation of Bacteriocins Possessing Antibiofilm Activity Produced by Lactobacillus plantarum SJ33

International Journal of Peptide Research and Therapeutics - Biofilm forming pathogens are among the major causes of hospital-acquired infections and are not much affected by antibiotic treatment....