Nodulisporic acid produces direct activation and positive allosteric modulation of AVR-14B, a glutamate-gated chloride channel from adult Brugia malayi.

Glutamate-gated chloride channels (GluCls) are unique to invertebrates and are targeted by macrocyclic lactones. In this study, we cloned an AVR-14B GluCl subunit from adult Brugia malayi, a causative agent of lymphatic filariasis in humans. To elucidate this channel's pharmacological properties, we used Xenopus laevis oocytes for expression and performed two-electrode voltage-clamp electrophysiology. The receptor was gated by the natural ligand L-glutamate (effective concentration, 50% [EC50] = 0.4 mM) and ivermectin (IVM; EC50 = 1.8 nM). We also characterized the effects of nodulisporic acid (NA) on Bma-AVR-14B and NA-produced dual effects on the receptor as an agonist and a type II positive allosteric modulator. Here we report characterization of the complex activity of NA on a nematode GluCl. Bma-AVR-14B demonstrated some unique pharmacological characteristics. IVM did not produce potentiation of L-glutamate-mediated responses but instead, reduced the channel's sensitivity for the ligand. Further electrophysiological exploration showed that IVM (at a moderate concentration of 0.1 nM) functioned as an inhibitor of both agonist and positive allosteric modulatory effects of NA. This suggests that IVM and NA share a complex interaction. The pharmacological properties of Bma-AVR-14B indicate that the channel is an important target of IVM and NA. In addition, the unique electrophysiological characteristics of Bma-AVR-14B could explain the observed variation in drug sensitivities of various nematode parasites. We have also shown the inhibitory effects of IVM and NA on adult worm motility using Worminator. RNA interference (RNAi) knockdown suggests that AVR-14 plays a role in influencing locomotion in B. malayi.

Immunometabolic impact of pancreastatin inhibitor PSTi8 in MCD induced mouse model of oxidative stress and steatohepatitis.

AIM: Pancreastatin, a dysglycemic hormone that encourages inflammation and steatosis in a variety of metabolic disorder animal models. The purpose of this study is to determine the effect of the pancreastatin inhibitor PSTi8 on immunometabolic changes in the liver of MCD-induced NASH mice. MAIN METHODS: Methionine and choline-deficient (MCD) diet was used for the development of NASH. Liver enzymes like SGOT, SGPT, and ALP and lipid profiles were also performed in the serum. Further, immunophenotyping study was performed in the liver through flowcytometer. Subsequently, Hematoxylin and Eosin, Picro Sirius Red and Masson's Trichrome staining were done to check the liver morphology and collagen staining, respectively. Inflammatory cytokines were measured through ELISA and gene expression through RT-PCR. The expression of α-SMA was examined using immunohistochemistry and immunofluorescence staining. KEY FINDINGS: PSTi8 inhibited the expression of lipogenic genes in the liver and attenuated bad cholesterol, SGOT, SGPT, and ALP in the serum. PSTi8 improved the liver morphology and attenuated collagen deposition. Subsequently, PSTi8 attenuated inflammatory M1-macrophages, CD8+T, CD4+T cells and increased anti-inflammatory M2 macrophages, T-reg and eosinophil populations in the liver. It also attenuated the expression of pro-inflammatory genes like Mcp1, Tnfα, and Il6. Apart from this, PSTi8 attenuated the oxidative stress marker, like ROS, and MDA and fibrosis marker α-SMA in the liver. It also decreased the apoptosis and ROS and MDA level in the liver. SIGNIFICANCE: Overall, these compressive studies revealed that PSTi8 exhibited beneficial effect on the liver of MCD-induced NASH mice by attenuating inflammation and oxidative stress.

Pancreastatin inhibitor PSTi8 ameliorates insulin resistance by decreasing fat accumulation and oxidative stress in high-fat diet-fed mice.

Abnormal fat accumulation, enhanced free fatty acids (FFA) release, and their metabolites cause insulin resistance (IR) in major glucose-lipid metabolic organs such as skeletal muscle and adipose tissue. However, excessive lipolysis and FFA release from adipose tissue elevate plasma FFA levels leading to oxidative stress and skeletal muscle IR. Indeed, in obese individuals, there is enhanced pro-inflammatory secretion from adipose tissue influencing insulin signaling in skeletal muscles. Here, we investigated the effect of PSTi8 on FFA-induced IR in both in vitro and in vivo models. Palmitate (Pal)-treated 3T3-L1 cells increased lipid accumulation as well as lipolysis, which reduced the insulin-stimulated glucose uptake. PSTi8 treatment significantly prevented Pal-induced lipid accumulation, and release and enhanced insulin-stimulated glucose uptake. It further reduced the release of pro-inflammatory cytokines from Pal-treated 3T3-L1 cells as well as from adipose tissue explants. In addition, PSTi8 treatment decreases M1 surface markers in Pal-treated bone marrow-derived monocytes (BMDM). PSTi8 treatment also significantly enhanced the Pal-mediated reduced skeletal muscle glucose disposal and reduced intracellular oxidative stress. In vitro effect of PSTi8 was consistent with in vivo HFD-fed mice IR model. PSTi8 treatment in HFD-fed mice significantly improved glucose metabolism and enhanced skeletal muscle insulin sensitivity with reduced adiposity and pro-inflammatory cytokines. Taken together, our results support that PSTi8 treatment can protect both adipose and skeletal muscles from FFA-induced IR.

The combined effect of Raspberry Ketone with Resveratrol against oxidative stress and steatohepatitis in rats: Pharmacokinetic and pharmacodynamic studies.

Raspberry Ketone (RK) and Resveratrol (RSV) are natural phenolic antioxidants and anti-inflammatory agents. However, its combined pharmacokinetic and pharmacodynamics potentials are not reported. The study aims to assess the combined effect of RK with RSV to protect rats from carbon-tetrachloride (CCl4) induced oxidative stress and NASH. The toxicant CCl4 was employed at a concentration of 1 mL/kg as a 1:1 (v/v) mixture with olive oil twice weekly for 6 weeks to induce liver toxicity. Animal treatment was followed for 2 weeks. Silymarin was used as a standard control drug to compare the hepatoprotective effect of RK and RSV. Hepatic histology, oxidative stress, MMP, reduced glutathione (GSH), plasma levels of SGOT, SGPT, and lipid profile (total cholesterol and triglycerides) were measured. Anti-inflammation genes (IL-10), and fibrotic genes (TGF-ß) were also examined in liver tissue. Oral administration of combined RK with RSV (50 + 50 mg/kg for 2 weeks) showed significantly more hepatoprotection by significantly decreasing elevated plasma markers and lipid profile than alone RK and RSV (100 mg/kg daily for 2 weeks). It also significantly alleviated the hepatic lipid peroxidation, restoring the activities of GSH levels in the liver. RT-PCR and Immunoblotting studies confirmed that significantly upregulation of anti-inflammation genes and protein expression (MMP-9) ameliorated the disease. Pharmacokinetic studies confirmed more synergistic stability in simulated gastric-intestinal fluids (FaSSGF, FaSSIF) and rat liver microsomes (CYP-450, NADPH oxidation & glucuronidation. Moreover, coadministration of drugs augmented the relative bioavailability, Vd/ F (L/Kg), and MRT0-∞( h), which leads to more efficacy. This pharmacokinetic and pharmacodynamic reveals a new adjuvant therapy for the treatment of steatohepatitis.

Augmented experimental design for bioavailability enhancement: a robust formulation of abiraterone acetate.

Abiraterone acetate (ABRTA) is clinically beneficial in management of metastatic castration-resistant prostate cancer (PC-3). With highlighted low solubility and permeability, orally hampered treatment of ABRTA necessitate high dose to achieve therapeutic efficacy. To triumph these challenges, we aimed to develop intestinal lymphatic transport facilitating lipid-based delivery to enhance bioavailability. ABRTA-containing self-nano emulsified drug delivery (ABRTA-SNEDDS) was statistically optimized by D-optimal design using design expert. Optimized formulation was characterized for particle size, thermodynamic stability, in vitro release, in vivo bioavailability, intestinal lymphatic transport, in vitro cytotoxic effect, anti-metastatic activity, and apoptosis study. Moreover, hemolysis and histopathology studies have been performed to assess pre-clinical safety. Nano-sized particles and successful saturated drug loading were obtained for optimized formulation. In vitro release upto 98.61 ± 3.20% reveal effective release of formulation at intestinal pH 6.8. ABRTA-SNEDDS formulation shows enhanced in vivo exposure of Abiraterone (2.5-fold) than ABRTA suspension in Sprague-Dawley rats. In vitro efficacy in PC-3 cell line indicates 3.69-fold higher therapeutic potential of nano drug delivery system. Hemolysis and histopathology study indicates no significant toxicities to red blood cells and tissues, respectively. Apparently, an opportunistic strategy to increasing bioavailability of ABRTA via intestinal lymphatic transport will create a viable platform in rapidly evolving chemotherapy. Enhanced translational utility of delivery was also supported through in vitro therapeutic efficacy and safety assessments. HighlightsAbiraterone acetate is a prostate cancer drug, impeded with low bioavailability.ABRTA loaded in self nano emulsifying drug delivery enhanced its bioavailability.Intestinal lymphatic transport played role in enhanced bioavailability of ABRTA.ABRTA-SNEDDS enhanced in vitro cytotoxic activity of ABRTA.ABRTA-SNEDDS found safe in preclinical safety evaluations.

EAT-18 is an essential auxiliary protein interacting with the non-alpha nAChR subunit EAT-2 to form a functional receptor.

Nematode parasites infect approximately 1.5 billion people globally and are a significant public health concern. There is an accepted need for new, more effective anthelmintic drugs. Nicotinic acetylcholine receptors on parasite nerve and somatic muscle are targets of the cholinomimetic anthelmintics, while glutamate-gated chloride channels in the pharynx of the nematode are affected by the avermectins. Here we describe a novel nicotinic acetylcholine receptor on the nematode pharynx that is a potential new drug target. This homomeric receptor is comprised of five non-α EAT-2 subunits and is not sensitive to existing cholinomimetic anthelmintics. We found that EAT-18, a novel auxiliary subunit protein, is essential for functional expression of the receptor. EAT-18 directly interacts with the mature receptor, and different homologs alter the pharmacological properties. Thus we have described not only a novel potential drug target but also a new type of obligate auxiliary protein for nAChRs.

PERIPAPILLARY PACHYCHOROID SYNDROME: New Insights.

PURPOSE: Available literature on peripapillary pachychoroid syndrome (PPS) is very limited. The purpose of this study is to evaluate features of PPS using multimodal imaging and shed further light on this entity. METHODS: This is a retrospective case series at a tertiary eye-care center. Patients with features of pachychoroid with preferential choroidal thickening in nasal macula along with pachyvessels were identified and included for analysis. The patients underwent fundus photography, fundus autofluorescence, optical coherence tomography, fundus fluorescein angiography, and indocyanine angiography. Images were separately analyzed by two different retina specialists. RESULTS: A total of 27 eyes of 14 patients with mean age of 52.2 years were included. Seven patients had bilateral PPS, whereas six had unilateral; one patient had only one seeing eye. A total of 21 eyes with PPS were studied. 52.4% of the eyes revealed retinal pigment epithelium gravitational tracks, outer retinal atrophy, and serous PEDs. None of the eyes showed choroidal folds. Fundus fluorescein angiography showed late leakage at macula in 33% eyes. Fundus autofluorescence showed features of PPS to be similar to central serous chorioretinopathy in most cases. CONCLUSION: Peripapillary pachychoroid syndrome shares common findings with central serous chorioretinopathy and seems to be a subset of central serous chorioretinopathy than a separate entity in pachychoroid disease spectrum.

PNEUMATIC VITREOLYSIS VERSUS PARS PLANA VITRECTOMY IN FOCAL SYMPTOMATIC VITREOMACULAR TRACTION SYNDROME: A Randomized Trial.

PURPOSE: To compare pneumatic vitreolysis and pars plana vitrectomy in the management of focal symptomatic vitreomacular traction (VMT). METHOD: Patients aged 18 years or older, with idiopathic focal symptomatic VMT and best-corrected visual acuity <20/40, without any other retinal pathology were randomized to undergo pneumatic vitreolysis (Group 1) or pars plana vitrectomy (Group 2). The primary outcome measure was resolution of traction confirmed with optical coherence tomography at 3 months. Secondary outcome measures were to compare changes in best-corrected visual acuity, central foveal thickness, and complications if any. RESULTS: A total of 30 eyes of 30 patients were included with 15 eyes in each group. Vitreomacular traction resolved successfully in 12 of 15 (80%) eyes in Group 1 and in all (100%) eyes in Group 2 (P = 0.224). The mean visual acuity improved from 0.80 ± 0.26 (20/126 Snellen's equivalent) to 0.70 ± 0.46 logMAR (20/100 Snellen's equivalent) in Group 1 (P = 0.71) and from 0.904 ± 0.44 (20/160 Snellen's equivalent) to 0.47 ± 0.26 logMAR (20/59 Snellen's equivalent) in Group 2 (P = 0.0016). Although 4 of 15 (26.66%) eyes in Group 1 had formation of full-thickness macular hole and 7 eyes required resurgery (4 for full-thickness macular hole and 3 for unresolved VMT), none in the pars plana vitrectomy group had any complications requiring resurgery (P = 0.0063). Two eyes in the pars plana vitrectomy group had intraoperative deroofing of the fovea leading to full-thickness macular hole. CONCLUSION: Pars plana vitrectomy is better than pneumatic vitreolysis as a single intervention in the management of focal symptomatic VMT.

Emerging nanotechnology based combination therapies of taxanes for multiple drug-resistant cancers.

'One drug- one target' to 'multiple drug- multiple targets' paradigm shifted to produce combination therapies, have found great outcomes to overcome multiple drug resistance (MDR). MDR is a significant barrier to the delivery of taxane-based anticancer medicines such as docetaxel, paclitaxel, and cabazitaxel. Due to MDR induced by drug efflux transporters, clinical application of these medications is impeded. To date, nanoformulations such as liposomes, micelles, polymeric nanoparticles, and gold nanoparticles have been investigated to deliver taxanes alone and in combination to reverse drug resistance. Despite the fact that various groups have already looked into taxane nano formulations in the literature, there isn't much in the way of polypharmacology and advanced nanoformulations with a focus on MDR. In this overview, we briefly covered the insights regarding MDR, difficulties related to current pharmaceutical products of taxanes, combination therapies of taxanes to combat MDR, all of which can be used to delve into cancer treatment.

Emodepside has sex-dependent immobilizing effects on adult Brugia malayi due to a differentially spliced binding pocket in the RCK1 region of the SLO-1 K channel.

Filariae are parasitic nematodes that are transmitted to their definitive host as third-stage larvae by arthropod vectors like mosquitoes. Filariae cause diseases including: lymphatic filariasis with distressing and disturbing symptoms like elephantiasis; and river blindness. Filarial diseases affect millions of people in 73 countries throughout the topics and sub-tropics. The drugs available for mass drug administration, (ivermectin, albendazole and diethylcarbamazine), are ineffective against adult filariae (macrofilariae) at the registered dosing regimen; this generates a real and urgent need to identify effective macrofilaricides. Emodepside, a veterinary anthelmintic registered for treatment of nematode infections in cats and dogs, is reported to have macrofilaricidal effects. Here, we explore the mode of action of emodepside using adult Brugia malayi, one of the species that causes lymphatic filariasis. Whole-parasite motility measurement with Worminator and patch-clamp of single muscle cells show that emodepside potently inhibits motility by activating voltage-gated potassium channels and that the male is more sensitive than the female. RNAi knock down suggests that emodepside targets SLO-1 K channels. We expressed slo-1 isoforms, with alternatively spliced exons at the RCK1 (Regulator of Conductance of Potassium) domain, heterologously in Xenopus laevis oocytes. We discovered that the slo-1f isoform, found in muscles of males, is more sensitive to emodepside than the slo-1a isoform found in muscles of females; and selective RNAi of the slo-1a isoform in female worms increased emodepside potency. In Onchocerca volvulus, that causes river blindness, we found two isoforms in adult females with homology to Bma-SLO-1A and Bma-SLO-1F at the RCK1 domain. In silico modeling identified an emodepside binding pocket in the same RCK1 region of different species of filaria that is affected by these splice variations. Our observations show that emodepside has potent macrofilaricidal effects and alternative splicing in the RCK1 binding pocket affects potency. Therefore, the evaluation of potential sex-dependent effects of an anthelmintic compound is of importance to prevent any under-dosing of one or the other gender of nematodes once given to patients.

miR159 Represses a Constitutive Pathogen Defense Response in Tobacco.

MicroR159 (miR159) regulation of GAMYB expression is highly conserved in terrestrial plants; however, its functional role remains poorly understood. In Arabidopsis (Arabidopsis thaliana), although GAMYB-like genes are constitutively transcribed during vegetative growth, their effects are suppressed by strong and constitutive silencing by miR159. GAMYB expression occurs only if miR159 function is inhibited, which results in detrimental pleiotropic defects, questioning the purpose of the miR159-GAMYB pathway. Here, miR159 function was inhibited in tobacco (Nicotiana tabacum) and rice (Oryza sativa) using miRNA MIM159 technology. Similar to observations in Arabidopsis, inhibition of miR159 in tobacco and rice resulted in pleiotropic defects including stunted growth, implying functional conservation of the miR159-GAMYB pathway among angiosperms. In MIM159 tobacco, transcriptome profiling revealed that genes associated with defense and programmed cell death were strongly activated, including a suite of 22 PATHOGENESIS-RELATED PROTEIN (PR) genes that were 100- to 1,000-fold upregulated. Constitutive expression of a miR159-resistant GAMYB transgene in tobacco resulted in phenotypes similar to that of MIM159 tobacco and activated PR gene expression, verifying the dependence of the above-mentioned changes on GAMYB expression. Consistent with the broad defense response, MIM159 tobacco appeared immune to Phytophthora infection. These findings suggest that the tobacco miR159-GAMYB pathway functions in the biotic defense response, which becomes activated upon miR159 inhibition. However, PR gene expression was not upregulated in Arabidopsis or rice when miR159 was inhibited, suggesting that miR159-GAMYB pathway functional differences exist between species, or factors in addition to miR159 inhibition are required in Arabidopsis and rice to activate this broad defense response.

Challenges of peptide and protein drug delivery by oral route: Current strategies to improve the bioavailability.

Advancement in biotechnology provided a notable expansion of peptide and protein therapeutics, used as antigens, vaccines, hormones. It has a prodigious potential to treat a broad spectrum of diseases such as cancer, metabolic disorders, bone disorders, and so forth. Protein and peptide therapeutics are administered parenterally due to their poor bioavailability and stability, restricting their use. Hence, research focuses on the oral delivery of peptides and proteins for the ease of self-administration. In the present review, we first address the main obstacles in the oral delivery system in addition to approaches used to enhance the stability and bioavailability of peptide/protein. We describe the physiochemical parameters of the peptides and proteins influencing bioavailability in the systemic circulation. It encounters, many barriers affecting its stability, such as poor cellular membrane permeability at the GIT site, enzymatic degradation (various proteases), and first-pass hepatic metabolism. Then describe the current approaches to overcome the challenges mentioned above by the use of absorption enhancers or carriers, structural modification, formulation and advance technology.

Role of 904 nm superpulsed laser-mediated photobiomodulation on nitroxidative stress and redox homeostasis in burn wound healing.

BACKGROUND: Burn wound healing is delayed due to several critical factors such as sustained inflammation, vascular disorder, neuropathy, enhanced proteolysis, infection, and oxidative stress. Burn wounds have limited oxygen supply owing to compromised blood circulation. Hypoxic burn milieu leads to free radicals overproduction incurring oxidative injury, which impedes repair process causing damage to cell membranes, proteins, lipids, and DNA. Photobiomodulation (PBM) with 904 nm superpulsed laser had shown potent healing efficacy via attenuating inflammation while enhancing proliferation, angiogenesis, collagen accumulation, and bioenergetic activation in burn wounds. METHODS: This study investigated the effects of 904 nm superpulsed laser at 0.4 mW/cm2 average power density, 0.2 J/cm2 total energy density, 100 Hz frequency, and 200 ns pulse width for 10 min daily for seven days postburn injury on nitroxidative stress, endogenous antioxidants status, and redox homeostasis. RESULTS: Photobiomodulation treatment significantly decreased reactive oxygen species, nitric oxide, and lipid peroxidation levels as compared to non-irradiated control. Further, protective action of PBM against protein oxidative damage was evidenced by reduced protein carbonylation and advanced oxidation protein product levels along with significantly enhanced endogenous antioxidants levels of SOD, catalase, GPx, GST, reduced glutathione, and thiol (T-SH, Np-SH, P-SH). Biochemical changes aid in reduction of oxidative stress and maintenance of redox homeostasis, which further well corroborated by significantly up-regulated protein expression of Nrf 2, hemeoxygenase (HO-1), and thioredoxin reductase 2 (Txnrd2). CONCLUSION: Photobiomodulation with 904 nm superpulsed laser led to reduction of nitroxidative stress, induction of endogenous antioxidants, and maintenance of redox homeostasis that could play a vital role in augmentation of burn wound healing.

Effects of Pulsed 810 nm Al-Ga-As Diode Laser on Wound Healing Under Immunosuppression: A Molecular Insight.

BACKGROUND AND OBJECTIVES: Dysregulated inflammation is one of the major contributing factors for the prevalence of non-healing chronic wound in immunosuppressed subjects. Photobiomodulation (PBM) has emerged as a potential non-thermal, light-based therapeutic healing intervention for the treatment of impaired wounds. STUDY DESIGN/MATERIALS AND METHODS: The present study delineates the underlying molecular mechanisms of PBM 810 nm laser-induced full-thickness cutaneous wound repair in immunosuppressed rats at continuous and pulsed wave-mode with power-density of 40 mW/cm 2 , fluence 22.6 J/cm 2 for 10 minutes daily for 7 post-wounding days. Molecular markers were assessed using biochemical, enzyme-linked immunosorbent assay quantification, enzyme kinetics and immunoblots analyses pertaining to inflammation, oxidative stress, cell survival, calcium signaling, and proliferation cascades. RESULTS: Results distinctly revealed that pulsed 810 nm (10 Hz) PBM potentially influenced the cell survival and proliferation signaling pathway by significantly upregulated phospho-protein kinase B(phospho-Akt), phospho-extracellular-signal-regulated kinase 1 (ERK1), transient receptor potential vanilloid-3 (TRPV3), Ca2+ , calmodulin, transforming growth factor-ß1 (TGF-ß1), TGF-ßR3, and Na + /K + -ATPase pump levels. PBM treatment resulted in reduction of exaggerated inflammatory responses evident by significantly repressed levels of interleukin-1ß (IL-1ß), IL-6, cyclooxygenase 2 (COX-2), and substance-P receptor (SPR), as well as inhibited apoptotic cell death by decreasing p53, cytochrome C, and caspase 3 levels (P < 0.05), which, in turn, effectively augment the wound repair in immunosuppressed rats. PBM treatment also lowered 4-hydroxynoneal (HNE) adduct level and NADP/NADPH ratio and upregulated the GRP78 expression, which might culminate into reduced oxidative stress and maintained the redox homeostasis. CONCLUSIONS: Taken together, these findings would be helpful in better understanding of the molecular aspects involved in pulsed 810 nm laser-mediated dermal wound healing in immunosuppressed rats through regulation of cell survival and proliferation via Ca2+ -calmodulin, Akt, ERK, and redox signaling. Lasers Surg. Med. © 2019 Wiley Periodicals, Inc.

Functional genomics in Brugia malayi reveal diverse muscle nAChRs and differences between cholinergic anthelmintics.

Many techniques for studying functional genomics of important target sites of anthelmintics have been restricted to Caenorhabditis elegans because they have failed when applied to animal parasites. To overcome these limitations, we have focused our research on the human nematode parasite Brugia malayi, which causes elephantiasis. Here, we combine single-cell PCR, whole muscle cell patch clamp, motility phenotyping (Worminator), and dsRNA for RNAi for functional genomic studies that have revealed, in vivo, four different muscle nAChRs (M-, L-, P-, and N-). The cholinergic anthelmintics had different selectivities for these receptors. We show that motility and patch-clamp responses to levamisole and pyrantel, but not morantel or nicotine, require the unc-38 and/or unc-29 genes. Derquantel behaved as a competitive antagonist and distinguished M-nAChRs activated by morantel (Kb 13.9 nM), P-nAChRs activated by pyrantel (Kb 126 nM), and L-nAChRs activated by levamisole (Kb 0.96 µM) and bephenium. Derquantel was a noncompetitive antagonist of nicotine, revealing N-type nAChRs. The presence of four diverse nAChRs on muscle is perhaps surprising and not predicted from the C. elegans model. The diverse nAChRs represent distinguishable drug targets with different functions: Knockdown of unc-38+unc-29 (L- and/or P-receptors) inhibited motility but knockdown of acr-16+acr-26 (M- and/or N-receptors) did not.

1400W, a highly selective inducible nitric oxide synthase inhibitor is a potential disease modifier in the rat kainate model of temporal lobe epilepsy.

Status epilepticus (SE) initiates epileptogenesis to transform normal brain to epileptic state which is characterized by spontaneous recurrent seizures (SRS). Prior to SRS, progressive changes occur in the brain soon after SE, for example, loss of blood-brain barrier (BBB) integrity, neuronal hyper-excitability (epileptiform spiking), neuroinflammation [reactive gliosis, high levels of reactive oxygen/nitrogen species (ROS/RNS)], neurodegeneration and synaptic re-organization. Our hypothesis was that modification of early epileptogenic events will alter the course of disease development and its progression. We tested the hypothesis in the rat kainate model of chronic epilepsy using a novel disease modifying drug, 1400W, a highly selective inhibitor of inducible nitric oxide synthase (iNOS/NOS-II). In an in vitro mouse brain slice model, using a multi-electrode array system, co-application of 1400W with kainate significantly suppressed kainate-induced epileptiform spiking. In the rats, in vivo, 4h after the induction of SE with kainate, 1400W (20mg/kg, i.p.) was administered twice daily for three days to target early events of epileptogenesis. The rats were subjected to continuous (24/7) video-EEG monitoring, remotely, for six months from epidurally implanted cortical electrodes. The 1400W treatment significantly reduced the epileptiform spike rate during the first 12-74h post-SE, which resulted in >90% reduction in SRS in long-term during the six month period when compared to the vehicle-treated control group (257±113 versus 19±10 episodes). Immunohistochemistry (IHC) of brain sections at seven days and six months revealed a significant reduction in; reactive astrogliosis and microgliosis (M1 type), extravascular serum albumin (a marker for BBB leakage) and neurodegeneration in the hippocampus, amygdala and entorhinal cortex in the 1400W-treated rats when compared to the vehicle control. In the seven day group, hippocampal Western blots revealed downregulation of inwardly-rectifying potassium (Kir 4.1) channels and glutamate transporter-1 (GLT-1) levels in the vehicle group, and 1400W treatment partially reversed Kir 4.1 levels, however, GLT-1 levels were unaffected. In the six month group, a significant reduction in mossy fiber staining intensity in the inner molecular layer of the dentate gyrus was observed in the 1400W-treated group. Overall these findings demonstrate that 1400W, by reducing the epileptiform spike rate during the first three days of post-insult, potentially modifies epileptogenesis and the severity of chronic epilepsy in the rat kainate model of TLE.

Study of single nucleotide polymorphisms of tumour necrosis factors and HSP genes in nasopharyngeal carcinoma in North East India.

Nasopharyngeal carcinoma (NPC) is an epithelial tumour with a distinctive racial and geographical distribution. High incidence of NPC has been reported from China, Southeast Asia, and northeast (NE) region of India. The immune mechanism plays incredibly role in pathogenesis of NPC. Tumour necrosis factors (TNFs) and heat shock protein 70 (HSP 70) constitute significant components of innate as well as adaptive host immunity. Multi-analytical approaches including logistic regression (LR), classification and regression tree (CART) and multifactor dimensionality reduction (MDR) were applied in 120 NPC cases and 100 controls to explore high order interactions among TNF-α (-308 G>A), TNF ß (+252 A>G), HSP 70-1 (+190 G>C), HSP 70-hom (+2437 T>C) genes and environmental risk factors. TNF ß was identified as the primary etiological factor by all three analytical approaches. Individual analysis of results showed protective effect of TNF ß GG genotype (adjusted odds ratio (OR2) = 0.27, 95 % CI = 0.125-0.611, P = 0.001), HSP 70 (+2437) CC genotype (OR2 = 0.17, 95 % CI = 0.0430.69, P = 0.013), while AG genotype of TNF ß was found significantly associated with risk of NPC (OR2 = 1.97, 95 % CI = 1.019-3.83, P = 0.04). Analysis of environmental factors demonstrated association of alcohol consumption, living in mud houses and use of firewood for cooking as major risk factors for NPC. Individual haplotype association analysis showed significant risk associated with GTGA haplotype (OR = 68.61, 95 % CI = 2.47-190.37, P = 0.013) while a protective effect with CCAA and GCGA haplotypes (OR = 0.19, 95 % CI = 0.05-0.75, P = 0.019; OR = 0.01 95 % CI = 0.05-0.30, P = 0.007). The multi-analytical approaches applied in this study helped in identification of distinct gene-gene and gene-environment interactions significant in risk assessment of NPC.

Immunoregulation of dendritic cells by the receptor T cell Ig and mucin protein-3 via Bruton's tyrosine kinase and c-Src.

The receptor T cell Ig and mucin protein-3 (TIM-3) has emerged as an important regulator of innate immune responses. However, whether TIM-3-induced signaling promotes or inhibits the activation and maturation of dendritic cells (DCs) still remains uncertain. In addition, the TIM-3 signaling events involved in this immunoregulatory function are yet to be established. In this article, we report that TIM-3 crosslinking by anti-TIM-3 Ab inhibited DC activation and maturation by blocking the NF-κB pathway. After Ab-mediated crosslinking, TIM-3 became tyrosine phosphorylated, which then sequentially bound and activated the nonreceptor tyrosine kinases Bruton's tyrosine kinase (Btk) and c-Src. Activation of Btk-c-Src signaling in turn triggered the secretion of some inhibitory factor (or factors) from DCs that inhibited the NF-κB pathway and subsequent activation and maturation of DCs. Silencing of Btk or c-Src abrogated the inhibitory effects of TIM-3 on DCs. These results demonstrate an essential role for Btk-c-Src signaling in TIM-3-induced DC suppression. Thus, in addition to demonstrating an inhibitory role for TIM-3 signaling in DC activation, we define the molecular mechanism by which TIM-3 mediates this effect.

Contribution of susceptibility locus at HLA class I region and environmental factors to occurrence of nasopharyngeal cancer in Northeast India.

High incidence of nasopharyngeal carcinoma (NPC) has been reported from China, Southeast Asia and Northeast (NE) region of India. Populations at geographic regions having higher incidence of NPC display human leukocyte antigen (HLA) distribution patterns different from areas having low incidence. The current study has investigated the contribution of environmental risk factors and ethnic variation of microsatellite markers in HLA region for the high incidence of NPC in NE India. Genotyping of HLA region using 33 microsatellite markers by fragment length analysis was done in 220 study subjects (120 NPC patients and 100 healthy controls). Association analysis showed two adjacent microsatellite markers HL003 (allele 121) and D6S2704 (allele 218) in the HLA class I region having association with high risk of NPC while allele 127 of HL003 and allele 255 of D6S2678 conferred a protective effect. The environmental factors mainly use of firewood (odds ratio (OR) = 3.797385, confidence interval (CI) = 1.97-7.30, P < 0), living in mud house (OR = 3.46, CI = 1.19-10.08, P = 0.022) and consumption of alcohol (OR = 2.11, CI = 1.02-4.37, P = 0.043) were found as major risk factors for NPC. Higher-order interaction showed combination of smoked food consumption and firewood use for cooking in multifactor dimensionality reduction (MDR) analysis and interaction of non-firewood users, non-ventilated houses and residence in mud houses in classification and regression tree (CART) analysis as the significant risk factors for NPC. Expression of Epstein-Barr virus (EBV) RNA was found in 92% (23/25) of NPC cases suggesting its significant role in NPC aetiopathogenesis. This study identified association of NPC with a susceptibility locus in the HLA class I region which has complex interaction with viral DNA and environmental factors.

Evidence for bactericidal activities of lipidic nanoemulsions against Pseudomonas aeruginosa.

Pseudomonas aeruginosa has been implicated in a broad range of infections and shown to acquire rapid resistance to anti-microbial agents. In the present, study we have used particular amalgamation of specific lipids that hold innate antibacterial activities, which can be transformed into cationized and non-cationized nanoemulsions. The anti-Pseudomonas activities were then elucidated by transmission/scanning electron microscopy, and atomic force microscopy. The microscopic studies revealed the cell lysis due to the formation of blebs, exudation of essential cellular contents and loss of characteristics contour of the cells. The microscopic studies were then corroborated by zone of inhibition, cytoplasmic release studies, time dependent killing and MIC determination. Conclusively, it can be inferred that the delivery issues of antibiotics could be reassessed by using certain excipients that possess inherent antibacterial properties. This will not only avoid unnecessary introduction of inactive excipients in the body, but will also reduce the dose of antibiotics because of synergistic effects of excipients and drug acting together.