Oral bioavailability studies of niosomal formulations of Cyclosporine A in albino rabbits.

Cyclosporine A (CsA) is an immunosuppressant agent. Two niosomal formulations of CsA, FTS and FSB were formulated. Both formulations were studied in terms of size, polydispersity index (PDI), morphology and entrapment efficacy etc. Niosomal formulations FTS and FSB and plain aqueous dispersion were given to three assemblies of Albino rabbits (n=8 per group). CsA levels in plasma were determined at appropriate time intervals and pharmacokinetic parameters were evaluated. The percentage entrapment efficiencies of FTS and FSB were found to be 77.29 and 89.31% for respectively. Transmission electron microscopy results indicated spherical nature of niosomes. In vivo studies demonstrated that the value of Cmax for the FSB formulation was 1968.419 ng/ml and it was 1498.951 ng/ml and 1073.87 ng/ml for FTS and aqueous dispersion of CsA (control) respectively. It was found that both niosomal formulation FTS & FSB presented significantly high (p<0.05) Cmax, AUC0-t, MRT 0-inf and half-life (t1/2) as associated to plain drug dispersion. However niosomal formulation FSB exhibited better in-vivo performance as compared to FTS. It was established that CsA can be successfully entrapped in niosomes. So niosomes are promising vehicle for CsA oral delivery.

Duck IFIT5 differentially regulates Tembusu virus replication and inhibits virus-triggered innate immune response.

Mammalian interferon-induced protein with tetratricopeptide repeats family proteins (IFITs) play important roles in host innate immune response to viruses. Recently, studies have shown that IFIT from poultry also plays a crucial part in antiviral function. This study first reports the regulation of duck Tembusu virus (DTMUV) replication by IFIT5 and the effect of duck IFIT5 (duIFIT5) on the innate immune response after DTMUV infection. Firstly, duIFIT5 was obviously increased in duck embryo fibroblast cells (DEFs) infected with DTMUV. Compared to the negative control, we found that in the duIFIT5-overexpressing group, the DTMUV titer at 24 h post infection (hpi) was significantly reduced, but the viral titer was strikingly increased at 48 hpi. Moreover, overexpression of duIFIT5 could significantly inhibit IFN-ß transcription and IFN-ß promoter activation at indicated time points after DTMUV infection. Further, in DTMUV-infected or poly(I:C)-stimulated DEFs, overexpression of duIFIT5 also significantly inhibited the activation of NF-κB and IRF7 promoters, as well as the activation of downstream IFN induced the interferon-stimulated response element (ISRE) promoter. Meanwhile, the transcription level of antiviral protein Mx, but not OASL, was obviously decreased at various time points. The opposite results were obtained by knockdown of duIFIT5 in DTMUV-infected or poly(I:C)-stimulated DEFs. Compared to the negative control, knockdown of duIFIT5 promoted DTMUV titer and DTMUV envelope (E) protein expression at 24 hpi, but DTMUV titer and E protein expression was markedly decreased at 48 hpi. Additionally, the promoters of IFN-ß, NF-κB, IRF7 and ISRE were significantly activated in the duIFIT5 knockdown group. Collectively, duIFIT5 differentially regulates DTMUV replication and inhibits virus-triggered innate immune response.

New Perspectives on Galleria mellonella Larvae as a Host Model Using Riemerella anatipestifer as a Proof of Concept.

Galleria mellonella larvae have been used as a host model to study interactions between pathogens and hosts for several years. However, whether the model is useful to interrogate Riemerella anatipestifer infection biology remained unknown. This study aimed to exploit the potential of G. mellonella larvae and reveal their limitations as a host model for R. anatipestifer infection. G. mellonella larvae were shown to be effective for virulence evaluations of different R. anatipestifer strains. Furthermore, the virulent strain R. anatipestifer CH-1 had a stronger ability to proliferate than the attenuated strain R. anatipestifer ATCC 11845 in both G. mellonella larvae and ducklings. Unconventionally it was shown that G. mellonella larvae cannot be used to evaluate the efficacy of antimicrobials and their combinations. Additionally, it was shown that certain virulence factors, such as OmpA (B739_0861), B739_1208, B739_1343, and Wza (B739_1124), were specific only for ducklings, suggesting that G. mellonella larvae must be cautiously used to identify virulence factors of R. anatipestifer Evaluation of heme uptake-related virulence genes, such as tonB1 and tonB2, required preincubating the strains with hemoglobin before infection of G. mellonella larvae since R. anatipestifer cannot obtain a heme source from G. mellonella larvae. In conclusion, this study revealed the applicability and limitations of G. mellonella as a model with which to study the pathogen-host interaction, particularly in the context of R. anatipestifer infection.

Anticancer potential of quercetin: A comprehensive review.

Diet plays a key role to maintaining healthy life. Many natural products present in our diet, such as flavonoids, can prevent the progression of cancer. Quercetin, a distinctive bioactive flavonoid, is a dietary component that has attracted the attention of dietitians and medicinal chemists due to its numerous health-promoting effects. It is an outstanding antioxidant that has a well-documented role in reducing different human cancers. Quercetin exhibits direct proapoptotic effects on tumor cells and thus can inhibit the progress of numerous human cancers. The anticancer effect of quercetin has been documented in numerous in vitro and in vivo studies that involved several cell lines and animal models. On the other hand, the high toxic effect of quercetin against cancer cells is accompanied with little or no side effects or harm to normal cells. Accordingly, this review presents an overview of recent developments on the use of quercetin against different types of cancer along with mechanisms of action. In addition, the present review summarizes the literature pertaining to quercetin as an anticancer agent and provides an assessment of the potential utilization of this natural compound as a complimentary or alternative medicine for preventing and treating cancer.

A review on epidemiology, global prevalence and economical losses of fasciolosis in ruminants.

Fasciolosis is an important plant borne trematode zoonosis in ruminants caused by the Fasciola hepatica and Fasciola gigentica, It is classified as a neglected tropical disease and found in more than 50 countries especially where sheep and cattle are reared. Fasciolosis is a serious animal health problems in many rural and urban areas of world, causing significant financial losses due to decrease in production and viscera condemnation in animals. Accurate diagnosis of fasciolosis is always remained a challenging task for the field practitioners. There is no comprehensive summary on the occurrence and distribution of the infection at international level. Therefore, we intended to provide a complete overview on the prevalence and epidemiology of fasciolosis in farm animals from a global prospective. It includes to map the global distribution of fasciolosis in different areas of the world to identify the endemic regions which may be a source of potential disease outbreak. The financial liability related to fasciolosis on the livestock production has also been addressed. For this purpose, the published data during 2000-2015 (15 years) on fasciolosis was reviewed and collected by electronic literature search of four databases including Google, PubMed, Science Direct, and Web of Science. Data presented are contemplated to enhance our current understanding of the parasite's geographical distribution, host range, and economic losses. Information provided would be useful for the application of more effective control strategies against fasciolosis in different geo-economics regions of the world.

Evidence of JEV in Culex tritaeniorhynchus and pigs from high altitude regions of Tibet, China.

BACKGROUND & OBJECTIVES: Culex tritaeniorhynchus is the primary vector of Japanese encephalitis virus (JEV) which is a major threat to animals and humans health. This virus has been reported earlier from low altitude regions of Tibet, in mosquitoes, Tibetan pigs and local Tibetans, but no reports are available for the probable propagation of JE to high altitude regions (3100 m) of Tibet. This study aimed to find the evidence of JEV in Cx. tritaeniorhynchus and pigs from high altitude regions of Tibet, China. METHODS: In total, 102 porcine serum samples and eight pools of Cx. tritaeniorhynchus were subjected to real-time PCR (RT-PCR) for the amplification of a fragment (~ 420 bp) of the NS1 gene. The resultant amplicons of the genes were subsequently analyzed and sequenced. RESULTS: Overall seroprevalence of JE in Tibetan pigs was 6.86%, while five pools of Cx. tritaeniorhynchus were found positive for JEV. Results showed genotype III as the most prevalent (100%) among JEV positive isolates. Furthermore, phylogenetic analysis of the JEV positive strains showed 98.8-99% similarity to four reference strains from China (JEV-Hubei, Whe, HYZ and CQ11-66). INTERPRETATION & CONCLUSION: The present study revealed that JEV has extended its geographic range to high altitude regions of Tibet. The factors like increased tourism and transportation might play key role in the transmission of JE that comprises a potential health risk for humans and animals.

Modulation of release and pharmacokinetics from nanoscale lipid prodrugs of dexamethasone with variable linkage chemistry.

In an attempt to tune drug release and subsequent pharmacokinetics once administered intravenously, we have synthesized three lipid-drug conjugates (LDCs) of dexamethasone (DXM) each possessing a different lipid-drug chemical linkage: namely ester, carbamate and carbonate. These LDCs were thoroughly characterized before being turned into nanoscale particles by an emulsion-evaporation process using DSPE-PEG2000 (Distearoyl-sn-Glycero-3-Phosphoethanolamine-N-(methoxy(polyethylene glycol)-2000) as the only excipient. Spherical nanoparticles (NPs) of about 140-170 nm, with a negative zeta potential, were obtained for each LDC and exhibited good stability upon storage at 4 °C for 45 days with no recrystallization of LDCs observed. LDC encapsulation efficacy was above 95% for the three LDCs, leading to a LDC loading of about 90% and an equivalent DXM loading above 50%. Although the ester and carbonate NPs did not exhibit any toxicity up to an equivalent DXM concentration of 100 µg/mL, the carbamate LDC NPs appeared very toxic towards RAW 264.7 macrophages and were discarded. Both ester and carbonate LDC NPs were shown to exert anti-inflammatory activity on LPS-activated macrophages. DXM release from LDC NPs in murine plasma was faster from ester than from carbonate NPs. Finally, pharmacokinetics and biodistribution were conducted, showing a lower exposure to DXM from carbonate LDC NPs than from ester LDC NPs, correlated with the slower DXM release from carbonate LDC NPs. These results outline the need for extended studies to find the best prodrug system for extended drug release.

Identification of potential inhibitors targeting yellow fever virus helicase through ligand and structure-based computational studies.

Yellow fever is a flavivirus having plus-sensed RNA which encodes a single polyprotein. Host proteases cut this polyprotein into seven nonstructural proteins including a vital NS3 protein. The present study aims to identify the most effective inhibitor against the helicase (NS3) using different advanced ligand and structure-based computational studies. A set of 300 ligands was selected against helicase by chemical structural similarity model, which are similar to S-adenosyl-l-cysteine using infiniSee. This tool screens billions of compounds through a similarity search from in-built chemical spaces (CHEMriya, Galaxi, KnowledgeSpace and REALSpace). The pharmacophore was designed from ligands in the library that showed same features. According to the sequence of ligands, six compounds (29, 87, 99, 116, 148, and 208) were taken for pharmacophore designing against helicase protein. Subsequently, compounds from the library which showed the best pharmacophore shared-features were docked using FlexX functionality of SeeSAR and their optibrium properties were analyzed. Afterward, their ADME was improved by replacing the unfavorable fragments, which resulted in the generation of new compounds. The selected best compounds (301, 302, 303 and 304) were docked using SeeSAR and their pharmacokinetics and toxicological properties were evaluated using SwissADME. The optimal inhibitor for yellow fever helicase was 2-amino-N-(4-(dimethylamino)thiazol-2-yl)-4-methyloxazole-5-carboxamide (302), which exhibits promising potential for drug development.Communicated by Ramaswamy H. Sarma.

Diclofenac prodrugs nanoparticles: An alternative and efficient treatment for rheumatoid arthritis?

We have synthesized new lipidic prodrugs of diclofenac by grafting aliphatic chains (C10, C12, C16 and C18) to diclofenac through an ester bond. Their molecular formulas were confirmed through HR-MS and the formation of ester bond by FTIR and NMR spectroscopy. Nanoparticles of the different prodrugs were successfully formulated using emulsion evaporation method and DSPE-PEG2000 as the only excipient. All nanoparticles were spherical and had a size between 110 and 150 nm, PdI ≤ 0.2 and negative Zeta potential values from -30 to -50 mV. In addition, they were stable upon storage at 4 °C up to 30-35 days. The encapsulation efficiency of the prodrug was above 90 % independently of the aliphatic chain length grafted. Nanoparticles did not induce any toxicity on LPS-activated THP-1 cells up to a concentration of 100 µg/mL (equivalent diclofenac) whereas diclofenac sodium salt IC50 was around 20 µg/mL. Following incubation of nanoparticles with LPS-activated THP-1 cells, a dose dependent inhibition of TNF-α was observed comparable to standard diclofenac sodium. Based on in vitro studies representative nanoparticles, Prodrug 3 NPs (C16 aliphatic chain) were selected for further in vitro and in vivo studies. Upon incubation in murine plasma, Prodrug 3 NPs underwent an enzymatic cleavage and almost 70 % of diclofenac was released from nanoparticles in 8 h. In vivo studies on a collagen induced arthritis murine model showed contrasted results: on one hand Prodrug 3 NPs led to a significant decrease of arthritis score and of paw volume compared to PBS after the second injection, on the other hand the third injection induced an important hepatic toxicity with the death of half of the mice from the NP group. To promote the reduction of inflammation while avoiding hepatic toxicity using NPs would require to precisely study the No Observable Adverse Effect Level and the schedule of administration in the future.

Comparative genomics and metabolomics analysis of Riemerella anatipestifer strain CH-1 and CH-2.

Riemerella anatipestifer is a major pathogenic microorganism in poultry causing serositis with significant mortality. Serotype 1 and 2 were most pathogenic, prevalent, and liable over the world. In this study, the intracellular metabolites in R. anatipestifer strains RA-CH-1 (serotype 1) and RA-CH-2 (serotype 2) were identified by gas chromatography-mass spectrometer (GC-MS). The metabolic profiles were performed using hierarchical clustering and partial least squares discriminant analysis (PLS-DA). The results of hierarchical cluster analysis showed that the amounts of the detected metabolites were more abundant in RA-CH-2. RA-CH-1 and RA-CH-2 were separated by the PLS-DA model. 24 potential biomarkers participated in nine metabolisms were contributed predominantly to the separation. Based on the complete genome sequence database and metabolite data, the first large-scale metabolic models of iJL463 (RA-CH-1) and iDZ470 (RA-CH-2) were reconstructed. In addition, we explained the change of purine metabolism combined with the transcriptome and metabolomics data. The study showed that it is possible to detect and differentiate between these two organisms based on their intracellular metabolites using GC-MS. The present research fills a gap in the metabolomics characteristics of R. anatipestifer.

In vitro Characterization and Release Studies of Combined Nonionic Surfactant-Based Vesicles for the Prolonged Delivery of an Immunosuppressant Model Drug.

BACKGROUND: Cyclosporine A (CsA) is an exceptional immunosuppressant used for the treatment of immune disorders. Niosomal vesicles are promising drug carriers that are formed by self-association of nonionic surfactants and cholesterol in an aqueous phase. The objective of the study was to formulate combined nonionic surfactant based vesicles and to evaluate their in vitro characterization, release studies and in vivo studies. MATERIALS AND METHODS: Five niosomal formulations (F7 to F11) were prepared using the thin film hydration method. The molar ratio of cholesterol and non-ionic surfactant taken was 1:1. In formulation F10, the combination of surfactants Span 20 and Brij 35 was used. The niosomes were characterized by zeta sizer and SEM for particle size analysis, in vitro drug release and stability studies. The pharmacokinetic studies were conducted on healthy albino rabbits. RESULTS: The size of niosome was found in the range of 427.1 nm to 972.3 nm. SEM image of optimized formulations F10 exhibit the spherical nature of niosomal vesicles. DSC thermograms of niosomal formulations exhibited a broadened endothermic peak. The stability study exhibited that all formulations are stable and negligible change of vesicle size and entrapment was observed with time. The percentage drug release was significantly higher as compared to CsA plain dispersion for all niosomal formulations at pH 1.2 and 7.4. The release kinetic behavior showed that all preparations were best described by zero order and can release active ingredient in a sustained manner. The pharmacokinetic data showed the test formulation (F10) possessed greater bioavailability as compared to the reference formulation (CsA aqueous dispersion). CONCLUSION: The formulation F10 demonstrated a comparatively more delayed rate of release with enhanced dissolution as compared to a single surfactant scheme. The F10 formulation can be a remarkable nanotechnology for prolonged delivery of CsA orally with improved dissolution profile and bioavailability.

Dexamethasone palmitate nanoparticles: An efficient treatment for rheumatoid arthritis.

Rheumatoid arthritis (RA) is a prevalent autoimmune disease characterized by joint inflammation, bone and cartilage erosion. The use of glucocorticoids in the treatment of RA is hampered by significant side effects induced by their unfavorable pharmacokinetics. Delivering glucocorticoids by means of nanotechnologies is promising but the encapsulation of highly crystalline and poorly water-soluble drugs results in poor loading and low stability. We report here the design of 130 nm nanoparticles made of solely dexamethasone palmitate, stabilized by polyethylene glycol-linked phospholipids displaying a negative zeta potential (-55 mV), high entrapment efficiency and stability over 21 days under storage at 4 °C. X ray diffraction showed no crystallization of the drug. When incubated in serum, nanoparticles released free dexamethasone which explains the in vitro anti-inflammatory effect on LPS-activated RAW 264.7 macrophages. Moreover, we demonstrate in a murine collagen-induced arthritis model the improved therapeutic efficacy of these nanoparticles. Their passive accumulation in arthritic joints leads to disease remission and recovery of the joint structure at a dose of 1 mg/kg dexamethasone, without any adverse effects. Dexamethasone palmitate nanoparticles are promising in the treatment of inflammation in rheumatoid arthritis with a very significant difference occurring at the late stage of inflammation allowing to prevent the progression of the disease.

Therapeutic effects of duck Tembusu virus capsid protein fused with staphylococcal nuclease protein to target Tembusu infection in vitro.

Tembusu virus (TMUV), a member of the genus flavivirus, primarily causes egg-drop syndrome in ducks and is associated with low disease mortality but high morbidity. The commercially available live vaccines for treating TMUV currently include the main WF100, HB, and FX2010-180P strains, and efficient treatment and/or preventative measures are still urgently needed. Capsid-targeted viral inactivation (CTVI) is a conceptually powerful new antiviral strategy that is based on two proteins from the capsid protein of a virus and a crucial effector molecule. The effector molecule can destroy the viral DNA/RNA or interfere with the proper folding of key viral proteins, while the capsid protein mainly plays a role in viral integration and assembly; the fusion proteins are incorporated into virions during packaging. This study aimed to explore the potential use of this strategy in duck TMUV. Our results revealed that these fusion proteins can be expressed in susceptible BHK21 cells without cytotoxicity and possess excellent Ca2+-dependent nuclease activity, and their expression is also detectable in DF-1 cells. Compared to those in the negative controls (BHK21 and BHK21/pcDNA3.1(+) cells), the numbers of viral RNA copies in TMUV-infected BHK21/Cap-SNase and BHK21/Cap-Linker-SNase cells were reduced by 48 h, and the effect of Cap-Linker-SNase was superior to that of Cap-SNase. As anticipated, these results suggest that these fusion proteins contribute to viral resistance to treatment. Thus, CTVI might be applicable for TMUV inhibition as a novel antiviral therapeutic candidate during viral infection.

The VP3 protein of duck hepatitis A virus mediates host cell adsorption and apoptosis.

Duck hepatitis A virus (DHAV) causes an infectious disease that mainly affects 1- to 4-week-old ducklings, resulting in considerable loss to the duck industry. Although there have been many studies on DHAV in recent years, the effects on host infection and pathogenesis of DHAV-1 remain largely unknown. This study investigated the effects of the DHAV-1 structural protein VP3 on DHAV-1 virus adsorption and apoptosis to explore the role of VP3 in the viral life cycle. The effects of DHAV-1 VP3 and an antibody against the protein on virion adsorption was analyzed by qRT-PCR. The results showed that the virus copy number for the rabbit anti-VP3 IgG-treated group was significantly lower than that for the negative control group but higher than that for the rabbit anti-DHAV-1 IgG-treated group. This result indicates that VP3 mediates DHAV-1 virus adsorption but that it is not the only protein that involved in this process. In addition, a eukaryotic recombinant plasmid, pCAGGS/VP3, was transfected into duck embryo fibroblasts (DEFs), and the apoptotic rate was determined by DAPI staining, the TUNEL assay and flow cytometry. DAPI staining showed nucleus fragmentation and nuclear edge shifting. TUNEL assay results revealed yellow nuclei, and flow cytometry indicated a significant increase in the apoptotic rate. In addition, qRT-PCR revealed increased in the transcriptional levels of the apoptotic caspase-3, -8 and -9, with the largest increase for caspase-3, followed by caspase-9 and caspase-8. Enzyme activity analysis confirmed these results. Furthermore, the VP3 protein decreased the mitochondrial membrane potential, and the transcriptional levels of the proapoptotic factors Bak, Cyt c and Apaf-1 in the mitochondrial apoptotic pathway were significantly upregulated. These data suggest that expression of VP3 in DEFs induces apoptosis and may primarily activate caspase-3-induced apoptosis through mitochondrion-mediated intrinsic pathways. The findings provide scientific data to clarify DHAV-1 infection and pathogenesis.