Search

Lead-like drugs, or drugs below molecular weight 300, are an important and sometimes overlooked component of the current pharmacopeia and contemporary medicinal chemistry practice. To examine the recent state-of-the-art in lead-like drug discovery, we surveyed recent drug approvals from 2011 to 2017 and top 200 prescribed medications, as well as provide case studies on recently approved lead-like drugs. Many of these recent drugs are close analogs of previously known drugs or natural substrates, with a key focus of their medicinal chemistry optimization being the choice of a low molecular weight starting point and maintaining low molecular weight during the optimization. However, the identification of low molecular weight starting points may be limited by the availability of suitable low molecular weight screening sets. To increase the discovery rate of lead-like drugs, we suggest an increased focus on inclusion and prosecution of lead-like starting points in screening libraries.

We investigated whether in-utero Cd(II) chloride exposure of the dams between 14th to 21st day of gestation affects memory and learning, oxidative stress, antioxidant enzyme activity and their gene expression in brain of the pups in their adulthood. In the Morris water maze, cadmium (Cd) exposure impaired spatial memory which was reversed following co-treatment with quercetin (100 mg/kg). In the passive avoidance paradigm, retention memory was adversely affected but was significantly reversed by co treatment with quercetin (25, 50, 100 mg/kg). The malondialdehyde and catalase (CAT) levels and glutathione-S-transferase (GST) activity were increased significantly in Cd-treated group, but were reversed by quercetin (all doses). The gene expression for CAT and GST in brain tissue of Cd treated animals also increased many folds as compared to the control, and this effect was decreased on co-treatment with quercetin (all doses), thus matching with the respective enzyme activities. Quercetin (25 mg/kg) when co-treated with Cd caused a decrease in GST activity compared to control, which points towards a complex interplay with oxidative free radicals and promoters and transcription factors. Thus, Cd exposure during late gestation causes impaired spatial and retention memory in the next generation which may be due to alteration of activity as well as gene expression of the antioxidant enzymes, CAT and GST. Quercetin may offer some protection of memory impairment probably by modulating these effects.

Stable, catalytically active palladium nanoparticles of various average diameters (1.9-7.4 nm) have been synthesized and characterized by X-ray diffraction, spectroscopy, and microscopy techniques to demonstrate remarkable size-dependent and renewed catalytic activity toward the Suzuki-Miyaura coupling reaction in green protocol. The catalytic activity is found to depend on the amount of the reducing agent, stabilizer-precursor ratio, solvent composition, and aryl halides used. The product obtained by this reaction is characterized by 1 H NMR, 13 C NMR, and IR spectroscopy analyses. A newly developed kinetic equation illustrates that while the catalyst particles of the lowest dimension are gradually exposed to the reactants and hence activated due to partial removal of capping polymer from the catalyst surface, others are deactivated due to agglomeration during the progress of the reaction, as conformed by the microscopic profiles of the used and unused catalysts., Competing Interests: The authors declare no competing financial interest.

Purpose: To determine whether there are identifiable, reproducible findings in the trabecular meshwork (TM) of patients with primary open-angle glaucoma (POAG) who underwent Kahook Dual Blade (KDB) goniotomy., Design: Noncomparative retrospective case series., Methods: Tertiary academic referral center, Veterans Affairs Medical Center. Thirteen patients (14 eyes) with POAG (100%) were treated with KDB goniotomy from May to December 2017. Isolated TM tissue was collected from 9 patients (10 eyes) and submitted for histologic analysis. Hematoxylin-eosin, periodic acid-Schiff, and elastin Van Gieson stains were completed, in addition to immunohistochemistry for collagen IV., Results: Mean age of patients was 74.2 ± 6.7 years. Trabecular beams were identified in all 10 specimens, although distorted in 4 samples, of which 3 had a history of laser trabeculoplasty. Collagen IV staining was present in 10 of 10 samples, coating the trabecular beams. Elastin was present in 8 of 10 samples along the trabecular beams. Intraocular pressure and number of glaucoma medications decreased significantly in all cases postoperatively (P < .0001, P = .035, respectively)., Conclusions: This pilot study demonstrates that tissue obtained during KDB goniotomy has a high yield of containing TM compared to reported yield of TM in specimens collected from traditional ab externo trabeculectomy (71% vs 20%, respectively). These goniotomy specimens possess sufficient anatomic preservation to be studied histologically. Trabecular meshwork obtained with this procedure may provide a novel modality to study TM dysfunction in open-angle glaucomas., (Published by Elsevier Inc.)

Background: Emergence of multi-drug-resistant tuberculosis is a serious challenge for successful global tuberculosis control. Early diagnosis of drug-resistant tuberculosis by direct nitrate reductase assay (NRA) aids in appropriate treatment and reduction in disease transmission, particularly in countries with high tuberculosis burden. The aim of this study was to evaluate the performance of NRA for direct detection of resistance to rifampicin and isoniazid in Mycobacterium tuberculosis in laboratories with limited resources., Methods: Fifty-eight new smear-positive sputum samples were processed as per the guidelines of revised national tuberculosis control program, India. The performance of NRA on middlebrook 7H11 agar was evaluated for detection of rifampicin and isoniazid resistance directly on smear-positive sputum specimens, and the results were compared with conventional proportion method. Sensitivity and specificity of the test were compared with the gold standard proportion method. Mc Nemar chi-square test was used to find out the significant difference between two methods., Results: Direct NRA for detection of rifampicin resistance was 85.7% sensitive and 100% specific, whereas sensitivity and specificity of isoniazid resistance were 87.5% and 100%, respectively. Agreement between NRA and proportion method was 98% for both the drugs. The mean days of drug susceptibility testing results were 19.3 days for NRA and 72 days for conventional proportion method. The results of NRA were available in 21 days for 83% of the samples., Conclusions: Direct NRA on middlebrook 7H11 medium is a highly sensitive, reliable, and significantly faster method to perform drug susceptibility testing. It has the potential to be implemented for rapid detection of multi-drug-resistant tuberculosis against insufficient resources., Competing Interests: The authors declare that they have no competing interests.This study was approved by B.P Koirala Institute of Health Sciences Institutional Ethical Review Board (reference number Acd. 1576/070/071). Informed written consent to participate in the study was obtained from participants.Not applicable.Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Aspergillosis is a fungal disease due to Aspergillus molds that can affect both humans and animals. As routine diagnosis remains difficult, improvement of basic knowledge with respect to its pathophysiology is critical to search for new biomarkers of infection and new therapeutic targets. Large-scale proteomics allows assessment of protein changes during various disease processes. In the present study, mass spectrometry iTRAQ® (isobaric tags for relative and absolute quantitation) protocol was used for direct identification and relative quantitation of host proteins in diseased fluids and tissues collected from an experimental rat model challenged with Aspergillus, as well as in blood obtained from naturally-infected penguins. In all, mass spectrometry analysis revealed that proteome during aspergillosis was mostly represented by proteins that usually express role in metabolic processes and biological process regulation. Ten and 17 proteins were significantly ≥4.0-fold overrepresented in blood of Aspergillus-diseased rats and penguins, respectively, while five and 39 were negatively ≥4.0-fold depleted within the same samples. In rat lungs, 33 proteins were identified with positive or negative relative changes versus controls and were quite different from those identified in the blood. Except for some zinc finger proteins, kinases, and histone transferases, and while three pathways were common (Wnt, cadherin and FGF), great inter-species variabilities were observed regarding the identity of the differentially-represented proteins. Thus, this finding confirmed how difficult it is to define a unique biomarker of infection. iTRAQ® protocol appears as a convenient proteomic tool that is greatly suited to ex vivo exploratory studies and should be considered as preliminary step before validation of new diagnostic markers and new therapeutic targets in humans., Competing Interests: The authors have declared that no competing interests exist.

Nano particles of a few α/β Bi 2 O 3 hetero-junctions of various compositions synthesized by one- pot hydrothermal method, exhibit exceptional and synergistic photo-catalytic activity for the degradation of Rhodamine-B in aqueous solution under natural sunlight. Pure α and pure β Bi 2 O 3 are also synthesized by control post heating of synthesized hetero-junction. The nano-materials were characterized by diffraction (XRD), microscopic and spectroscopic techniques. The XRD reveals α-β phase hetero-junctions of Bi 2 O 3 are made of α-Bi 2 O 3 and β-Bi 2 O 3 with average dimensions within 13-113 and 5-71 nm respectively and having band gap range of 2.4- 2.9 eV. The spectrophotometrically determined % degradation of the dye and associated rate constant on the best hetero-junction are increased by 4.5(/2.1) and 3.3(/1.2) times than these on pure α (/β). The effects of operational parameters and trapping agents have been analyzed. The maximum removal of the dye was achieved up to 99.6% in 3 h using 0.5 g/L photo-catalyst at pH 3. The reusability test shows that the photo-catalytic activity is retained excellently due to change in chemical nature of the catalyst from α -Bi 2 O 3 to β-Bi 2 O 3, Bi 2 O 2 CO 3 and BiOCl. A suitable mechanism is proposed., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Cultured trabecular meshwork (TM) cells are a valuable model system to study the cellular mechanisms involved in the regulation of conventional outflow resistance and thus intraocular pressure; and their dysfunction resulting in ocular hypertension. In this review, we describe the standard procedures used for the isolation of TM cells from several animal species including humans, and the methods used to validate their identity. Having a set of standard practices for TM cells will increase the scientific rigor when used as a model, and enable other researchers to replicate and build upon previous findings., (Copyright © 2018. Published by Elsevier Ltd.)

Studies have linked the serine-threonine kinase MAP4K4 to the regulation of a number of biological processes and/or diseases, including diabetes, cancer, inflammation, and angiogenesis. With a majority of the members of our lead series (e.g., 1) suffering from time-dependent inhibition (TDI) of CYP3A4, we sought design avenues that would eliminate this risk. One such approach arose from the observation that carboxylic acid-based intermediates employed in our discovery efforts retained high MAP4K4 inhibitory potency and were devoid of the TDI risk. The medicinal chemistry effort that led to the discovery of this central nervous system-impaired inhibitor together with its preclinical safety profile is described.

Sodium-phosphate cotransporter 2a, or NaPi2a (SLC34A1), is a solute-carrier (SLC) transporter located in the kidney proximal tubule that reabsorbs glomerular-filtered phosphate. Inhibition of NaPi2a may enhance urinary phosphate excretion and correct maladaptive mineral and hormonal derangements associated with increased cardiovascular risk in chronic kidney disease-mineral and bone disorder (CKD-MBD). To date, only nonselective NaPi inhibitors have been described. Herein, we detail the discovery of the first series of selective NaPi2a inhibitors, resulting from optimization of a high-throughput screening hit. The oral PK profile of inhibitor PF-06869206 ( 6f ) in rodents allows for the exploration of the pharmacology of selective NaPi2a inhibition., Competing Interests: The authors declare no competing financial interest.

Optimization of the pharmacokinetic (PK) properties of a series of activators of adenosine monophosphate-activated protein kinase (AMPK) is described. Derivatives of the previously described 5-aryl-indole-3-carboxylic acid clinical candidate (1) were examined with the goal of reducing glucuronidation rate and minimizing renal excretion. Compounds 10 (PF-06679142) and 14 (PF-06685249) exhibited robust activation of AMPK in rat kidneys as well as desirable oral absorption, low plasma clearance, and negligible renal clearance in preclinical species. A correlation of in vivo renal clearance in rats with in vitro uptake by human and rat renal organic anion transporters (human OAT/rat Oat) was identified. Variation of polar functional groups was critical to mitigate active renal clearance mediated by the Oat3 transporter. Modification of either the 6-chloroindole core to a 4,6-difluoroindole or the 5-phenyl substituent to a substituted 5-(3-pyridyl) group provided improved metabolic stability while minimizing propensity for active transport by OAT3.

Homeostasis of intraocular pressure (IOP) is important for the maintenance of anterior eye anatomic integrity, minimizing pressure-associated damage to the optic nerve, and maintaining a pressure gradient for blood flow to the eye. IOP is regulated by equilibrium between aqueous humor (AH) production and its outflow. The ciliary body (CB) is thought to actively secrete AH. However, whether AH composition and in particular, its phospholipids are entirely due to CB secretion remains uncertain. Comparison of phospholipids released by cultured CB, phospholipids present within CB tissue, within AH, and within blood and serum are consistent with release of most phospholipids into the AH by the CB. Treatment of CB in culture with timolol, a non-specific beta-adrenergic antagonist, alters the release of phospholipids by CB into the media. However, dorzalamide, a carbonic anhydrase inhibitor that reduces production of AH, does not affect phospholipid release thereby suggesting timolol, which also decreases IOP through decreased AH outflow, affects other physiological homeostatic mechanisms regulating aqueous outflow. These outflow changes also affect the composition of secreted phospholipids. We present evidence that release of lipids by the CB has a prolonged survival effect on cultured primary TM cells and TM tissue., (© 2017 Wiley Periodicals, Inc.)

This protocol provides a step-by-step guide to shotgun sphingolipid analysis of aqueous humor. We describe the Bligh and Dyer crude lipid extraction method and electrospray ionization tandem mass spectrometry (ESI-MS/MS) coupled with MZmine 2.21 data processing for identification and ratiometric quantitation of sphingosine, sphingosine-1-phosphate, sphingomyelin, and ceramide.

iTRAQ 4plex method enables multiplexing of up to four samples for simultaneous quantitation to improve sensitivity and scope of proteomic analysis. Here, we describe iTRAQ 4plex labeling of human aqueous humor specimens followed by ultra-high performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) and data analysis for peptide identification and quantitation using Proteome Discoverer software. The protocol can be applied for other animals as well; however, pooling of specimens may be required to obtain sufficient amount of protein for labeling.

Background: Cholera is known to be transmitted from person to person, and inactivated oral cholera vaccines (OCVs) have been shown to confer herd protection via interruption of this transmission. However, the geographic dimensions of chains of person-to-person transmission of cholera are uncertain. The ability of OCVs to confer herd protection was used to define these dimensions in two cholera-endemic settings, one in rural Bangladesh and the other in urban India., Methods: Two large randomized, placebo-controlled trials of inactivated OCVs, one in rural Matlab, Bangladesh and the other in urban Kolkata, India, were reanalyzed. Vaccine herd protection was evaluated by relating the risk of cholera in placebo recipients to vaccine coverage of surrounding residents residing within concentric rings. In Matlab, concentric rings in 100-m increments up to 700m were evaluated; in Kolkata, 50-m increments up to 350m were evaluated., Results: One hundred and eight cholera cases among 24667 placebo recipients were detected during 1year of post-vaccination follow-up at Matlab; 128 cholera cases among 34968 placebo recipients were detected during 3 years of follow-up in Kolkata. Consistent inverse relationships were observed between vaccine coverage of the ring and the risk of cholera in the central placebo recipient for rings with radii up to 500m in Matlab and up to 150m in Kolkata., Conclusions: These results suggest that the dimensions of chains of person-to-person transmission in endemic settings can be quite large and may differ substantially from setting to setting. Using OCVs as 'probes' to define these dimensions can inform geographical targeting strategies for the deployment of these vaccines in endemic settings., (Copyright © 2017 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

We present our ab initio molecular dynamics (MD) study of the effect of Si on the oxidation of α-Ti(0   0   0   1) surfaces. We varied the Si concentration in the first layer of the surface from 0 to 25 at.% and the oxygen coverage ( θ ) on the surface was varied up to 1 monolayer (ML). The MD was performed at 300, 600 and 973 K. For θ  = 0.5 ML, oxygen penetration into the slab was not observed after 16 ps of MD at 973 K while for θ  > 0.5 ML, oxygen penetration into the Ti slab was observed even at 300 K. From Bader charge analysis, we confirmed the formation of the oxide layer on the surface of the Ti slab. At higher temperatures, the Si atoms diffused from the first layer to the interior of the slab, while the Ti atoms moved from second layer to the first layer. The pair correlation function shows the formation of a disordered Ti-O network during the initial stage of oxidation. Si was found to have a strong influence on the penetration of oxygen in the Ti slab at high temperatures.

Background: Besides environmental risk factors, genetic factors play a crucial role in the pathogenesis of primary hypertension. The current study is to unravel whether hypertensive phenotypes vary in mice with different genetic background., Methods: Hypertension was induced in C57BL/6J (B6), DBA/2J (D2), and 25 BXD strains by administrating angiotensin (Ang)II (2.5 mg/kg/day infused by osmotic minipump) for 4 weeks. Systolic blood pressure was monitored before (baseline) and after 4 weeks of AngII treatment by tail cuff. Cardiac and renal fibrosis was evaluated by picrosirius red staining and collagen volume fraction (CVF) was quantitated using imaging analyzing system; cardiac transforming growth factor (TGF)-β gene expression was monitored by RT-PCR, and inflammatory response was detected by immunohistochemical ED-1 staining., Results: AngII infusion caused hypertension in all strains. However, blood pressure elevation was more evident in the D2 strain than the B6 group, while it was widely variable among BXD strains. Furthermore, chronic AngII treatment lead to development of hypertensive cardiac and renal diseases. Cardiac and renal CVF levels in the D2 strain was significantly higher than the B6 cohort, whereas these varied vastly across BXD strains. Moreover, cardiac TGF-β mRNA levels were markedly diverse among various mouse strains., Conclusion: Our study unequivocally demonstrates that in response to AngII, BXDs with different genetic background expressed hypertension phenotypes with varied degree in severity. It implicates that genomics contribute to pathogenesis of primary hypertension. Building upon the genotype and hypertensive phenotypes, the BXD cohort can be further exploited experimentally to identify genes that influence blood pressure., (© American Journal of Hypertension, Ltd 2017. All rights reserved. For Permissions, please email: journals.permissions@oup.com)

Climatic droplet keratopathy (CDK) is an acquired degenerative disease predominantly affecting males over 40 years old. It results in progressive corneal opacities usually affecting both eyes. CDK is multifactorial and its etiology remains unknown. Our recent findings are consistent with CDK pathology being driven by environmental factors with oxidative stress playing an important role (e.g.,, contributing to lipid peroxidation) rather than climate factors. The changes in corneal lipid composition affected by environmental factors remain understudied. The purpose of this study was to systematically investigate phospholipids profile (phosphatidylcholine [PC] and phosphatidylserine [PS]) in corneas from CDK patients using tandem mass spectrometry. Samples from CDK areas and from non-affected areas were obtained from patients diagnosed with CDK who underwent cataract surgery, were subjected to lipid extraction using a modified Bligh and Dyer method; protein concentrations were determined using the Bradford's method. Lipids were identified and subjected to ratiometric quantification using TSQ Quantum Access Max triple quadrupole mass spectrometer, using appropriate class specific lipid standards. All phospholipid classes showed lower total amounts in affected areas compared to control areas from CDK's corneas. Comparative profiles of two phospholipid classes (PC, PS) between CDK areas and control areas showed several common species between them. We also found a few unique lipids that were absent in CDK areas compared to controls and vice versa. Lower amount of phospholipids in CDK areas compared to control areas could be attributed to the lipid peroxidation in the affected corneal regions as a consequence of increased oxidative stress. J. Cell. Biochem. 118: 3920-3931, 2017. © 2017 Wiley Periodicals, Inc., (© 2017 Wiley Periodicals, Inc.)

Photoreceptor death and retinal gliosis underlie the majority of vision threatening retinal diseases including retinal detachment (RD). Although the underlying pathobiology of vision limiting processes in RD is not fully understood, inflammation is known to play a critical role. We conducted an iTRAQ proteomic screen of up- and down-regulated proteins in a murine model of RD to identify potential targetable candidates. Macrophage migration inhibitory factor (MIF) was identified and evaluated for neurotoxic and pro-gliotic effects during RD. Systemic administration of the MIF inhibitor ISO-1 significantly blocked photoreceptor apoptosis, outer nuclear layer (ONL) thinning, and retinal gliosis. ISO-1 and MIF knockout (MIFKO) had greater accumulation of Müller glia pERK expression in the detached retina, suggesting that Müller survival pathways might underlie the neuroprotective response. Our data show the feasibility of the MIF-inhibitor ISO-1 to block pathological damage responses in retinal detachment and provide a rationale to explore MIF inhibition as a potential therapeutic option for RD.

Background & Objectives: Data on serial cervical length (CL) measurements in pregnancy at term to predict spontaneous labour onset are scarce and conflicting. This study was conducted to observe CL changes preceding spontaneous onset of labour, by serial transvaginal sonography (TVS) and transabdominal sonography (TAS), in nulliparous Indian women near term., Methods: Only nulliparous women with a singleton foetus in cephalic presentation and who confirmed their gestational age were recruited. Sonographic CL measurements were taken at weekly intervals from 36 wk gestation onwards by a single ultrasonologist. Transabdominal and transvaginal measurements were undertaken using the suitable transducer probes with the women in the supine position., Results: A total of 104 women with spontaneous onset of labour were evaluated. There was substantial variation in CL measurements, both by TVS and by TAS, from 36 to 40 wk gestation, although the two sets of measurements correlated closely. Mean CL changed significantly over the last three weeks before delivery. However, only one-third of the women showed CL change of >5 mm per week in the last three weeks. There was poor correlation between gestational age at delivery and the last measured CL, either by TVS or TAS. Length >3.1 mm, measured by TVS at 38 wk gestation, predicted post-dated pregnancy to a limited extent., Interpretation & Conclusions: Inter-individual variations in CL and in CL changes were large. Thus, it was not practical to predict spontaneous onset of labour by sonographic CL measurement near term. Post-dated pregnancy may be predicted with limited success. Further studies should explore other parameters, in addition to CL., Competing Interests: None

A correction to this article has been published and is linked from the HTML version of this paper. The error has been fixed in the paper.

Encephalitozoon cuniculi is a microsporidian species which can induce subclinical to serious disease in mammals including rabbits, a definitive natural host. The pathophysiology of infection has not been comprehensively elucidated. In this exploratory study, we utilized two mass spectrometry approaches: first, the analysis of the humoral response by profiling the microsporidian antigens as revealed by Western blot screening, and second, implementing the iTRAQ®-labeling protocol to focus on the changes within the host proteome during infection. Seven E. cuniculi proteins were identified at one-dimensional gel regions where specific seropositive reaction was observed by Western blot, including polar tube protein 3, polar tube protein 2, and for the first time reported: heat shock related 70kDa protein, polysaccharide deacetylase domain-containing protein, zinc finger protein, spore wall and anchoring disk complex protein EnP1, and translation elongation factor 1 alpha. In addition, there was a significant increase of nine host proteins in blood samples from E. cuniculi-diseased rabbits in comparison with non-diseased control subjects undergoing various inflammatory processes. This included serum paraoxonase, alpha-1-antiproteinase F precursor and alpha-1-antiproteinase S-1 which have presumptive catalytic activity likely related to infection control, and cystatin fetuin-B-type, an enzyme regulator that has been poorly studied to date. Notably, 11 proteins were found to be statistically increased in rabbits with neurological versus renal clinical presentation of E. cuniculi infection. Overall, this novel analysis based on mass spectrometry has provided new insights on the inflammatory and humoral responses during E. cuniculi infection in rabbits.

AbstractSeveral epidemiological studies have indicated the presence of asymptomatic infections with Leishmania donovani in the Indian subcontinent, where parasite transmission is considered anthroponotic. In India, such asymptomatic Leishmania cases have been identified in the state of Bihar. We explored here, the presence of asymptomatic Leishmania infection among healthy individuals living in two districts in the state of West Bengal, India, using serological and molecular tests. Blood samples of 246 healthy individuals were collected from nine villages of Malda and Murshidabad districts in West Bengal, considered endemic for visceral leishmaniasis (VL). Real-time quantitative polymerase chain reaction (qPCR) was performed for the quantification of parasite load in the blood. In addition, two serological tests were carried out to demonstrate anti- Leishmania antibodies: rK39 strip test and anti-total soluble Leishmania antigen IgG using enzyme-linked immunosorbent assay method. Nearly one-fifth (53/246) of the screened population was positive in qPCR as against 10.97% (27/246) positive in rK39 strip test. A range of parasite load was observed in the blood of identified asymptomatic Leishmania cases with a median value of 7.7 parasites/mL (range = 1-65). There was poor agreement between qPCR and serological tests (κ = 0.089, P = 0.13), and 29.62% and 20.54% of the population were qPCR positive in seropositive and seronegative groups, respectively. Combined molecular and serological tests enhanced the capacity to detect asymptomatic Leishmania infection in healthy individuals residing in the endemic areas of VL. A significant proportion of asymptomatic Leishmania individuals was detected in the examined endemic regions of West Bengal that might play a role in promoting VL transmission.

The main and only modifiable risk factor in glaucoma, the group of usually late onset progressive and irreversible blinding optic neuropathies, is elevated intraocular pressure (IOP). The increase in IOP is due to impeded aqueous humor (AH) outflow through the conventional pathway. The aberrant increased resistance at the trabecular meshwork (TM), the filter-like region in the anterior eye chamber is the major contributory factor in causing the impeded outflow. In normal as well as in glaucoma eyes the regions of the TM are divided into areas of high and low flow. The collector channels and distal outflow regions are now increasingly being recognized as potential players in contributing to impede AH outflow. Structural and molecular make-up contributing to the segmental blockage to outflow is likely to provide greater insight. Establishing segmental blockage to outflow in model systems of glaucoma such as the mouse in parallel to human eyes will expand our repertoire of tools for investigation. Further study into this area of interest has the potential to ultimately lead to the development of new therapeutics focused on lowering IOP by targeting the various components of segmental blockage of outflow in the TM and in the distal outflow region., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

AbstractVisceral leishmaniasis (VL), popularly known as kala-azar, is essentially a disease of poverty. Kala-azar is caused by a parasite, Leishmania donovani . Recent review indicates that worldwide 98 countries are endemic for kala-azar. Approximately 0.2-0.4 million new VL cases occur each year worldwide. More than 90% of global VL cases occur in Bangladesh, Brazil, Ethiopia, India, South Sudan, and Sudan. This trend is slowly changing due to the progress in kala-azar elimination in southeast Asia, where Bangladesh has reported an average of some 600 new cases in 2014-2015. With the advancement in our knowledge about the disease and development of tools to diagnose and treat VL, it was considered that elimination of kala-azar was possible from India, Nepal, and Bangladesh. The three countries signed a memorandum of understanding in 2005 for collaboration. Miltefosine is the first ever oral drug developed to treat VL, which was later replaced by lipid amphotericin B. The main components of the strategy are early diagnosis using rK39 strip test and complete treatment utilizing miltefosine for 28 days. Dichlorodiphenyltrichloroethane or pyrethroids were deployed for vector control. There was much to be desired for better performance of the vector control activity. Pharmacovigilance and monitoring of drug resistance were the weakest part of the program. In the post-elimination phase, surveillance reinforced by active case finding will of a crucial factor for sustainability of the elimination. A strong political will is required to ensure elimination of kala-azar from the Indian subcontinent and its sustainability in the post-elimination phase.

In the eye, intraocular pressure (IOP) is tightly regulated and its persistent increase leads to ocular hypertension and glaucoma. We have previously shown that trabecular meshwork (TM) cells might detect aqueous humor fluid shear stress via interaction of the extracellular matrix (ECM) protein cochlin with the cell surface bound and stretch-activated channel TREK-1. We provide evidence here that interaction between both proteins are involved in IOP regulation. Silencing of TREK-1 in mice prevents the previously demonstrated cochlin-overexpression mediated increase in IOP. Biochemical and electrophysiological experiments demonstrate that high shear stress-induced multimeric cochlin produces a qualitatively different interaction with TREK-1 compared to monomeric cochlin. Physiological concentrations of multimeric but not monomeric cochlin reduce TREK-1 current. Results presented here indicate that the interaction of TREK-1 and cochlin play an important role for maintaining IOP homeostasis. [Corrected].

In glaucoma, lowered intraocular pressure (IOP) confers neuroprotection. Elevated IOP characterizes glaucoma and arises from impaired aqueous humor (AH) outflow. Increased resistance in the trabecular meshwork (TM), a filter-like structure essential to regulate AH outflow, may result in the impaired outflow. Flow through the 360° circumference of TM structures may be non-uniform, divided into high and low flow regions, termed as segmental. After flowing through the TM, AH enters Schlemm's canal (SC), which expresses both blood and lymphatic markers; AH then passes into collector channel entrances (CCE) along the SC external well. From the CCE, AH enters a deep scleral plexus (DSP) of vessels that typically run parallel to SC. From the DSP, intrascleral collector vessels run radially to the scleral surface to connect with AH containing vessels called aqueous veins to discharge AH to blood-containing episcleral veins. However, the molecular mechanisms that maintain homeostatic properties of endothelial cells along the pathways are not well understood. How these molecular events change during aging and in glaucoma pathology remain unresolved. In this review, we propose mechanistic possibilities to explain the continuum of AH outflow control, which originates at the TM and extends through collector channels to the episcleral veins., (Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Imaging mass spectrometry (IMS) is a novel quantitative technique used to investigative diverse biomolecules in tissue sections. Specifically, IMS uses analytical separation of mass spectrometry to determine the spatial distribution of certain lipids and/or proteins located directly on biological sections from a single tissue sample. Typically, IMS is combined with histological analysis to reveal additional distribution details of characterized biomolecules including cell type and/or subcellular localization. In this chapter, we describe the use of Matrix-Assisted Laser Desorption/Ionization (MALDI) Time-Of-Flight/Time-Of-Flight (TOF/TOF) to analyze various cholesterol and phosphatidylcholine species in atherosclerotic plaque of murine heart aortic valves. In particular, we detail animals used, tissue collection, preparation, matrix application, spectra acquisition for generating a color-coded image based on IMS spectral characteristics.

Electrospray ionization mass spectrometry has revolutionized the way lipids are studied. In this work, we present a tutorial for analyzing class-specific lipid spectra obtained from a triple quadrupole mass spectrometer. The open-source software MZmine 2.21 is used, coupled with LIPID MAPS databases. Here, we describe the steps for lipid identification, ratiometric quantification, and briefly address the differences to the analyses when using direct infusion versus tandem liquid chromatography-mass spectrometry (LC-MS). We also provide a tutorial and equations for quantification of lipid amounts using synthetic lipid standards and normalization to a protein amount.

Introduction: Irrational use of Topical Corticosteroid (TC) is quite common in India due to unrestricted availability and use of TC not only by general public but also by physicians and chemists due to quick relief of symptoms in different dermatological conditions., Aim: The present study was conducted to evaluate and analyse the prevalence of misuse of TC and the causes behind misuse of TC among patients in a dermatology outpatient department in New Delhi., Materials and Methods: This was a cross-sectional observational questionnaire based study conducted over a period of 5 months (1 st June 2015 to 30 th November 2015). Patients were questioned and assessed for misuse of TCs in terms of indication, dose, frequency, duration and source of recommendation of TC., Results: During the study period, 256 (11.77%) patients presented with inappropriate use of TC out of 2174 patients attending OPD of dermatology unit of a government hospital. A total of 250 patients presented with adverse effects of TC resulting from the misuse of these drugs. There were 60%female patients and 20-29 years (38%) was the most common age group affected. We found fungal infection (38%) was the most common reason of abuse followed by facial acne (29%) and lightening of skin colour (8.4%). Friends and family (33.2%) were found to be the most influencing factors for misuse of TC. Betamethasone (72.8%) was the most commonly used TC preparation and tinea incognito (26.4%) followed by facial acne (25.6%) were the most common side effect experienced., Conclusion: Outcome of the misuse of TC could be dreadful and the cause for same lies at all levels- patients, family, physicians and the regulatory authorities.

Background: The inflammatory response system has been implicated in the pathophysiology of major depression. The pro-inflammatory cytokines like interferon-γ induce the enzyme indoleamine-2,3-dioxygenase (IDO) of the kynurenine pathway of tryptophan metabolism. The induction of IDO reduces the availability of tryptophan for serotonin synthesis. Furthermore, the metabolites of kynurenine pathway have neurotoxic property, which along with decreased serotonin may account for depression-like illness., Methods: The aim of this study was to compare the effects of treatment with fluoxetine and 1-methyl-L-tryptophan (1-MT) on Bacillus Calmette-Guerin (BCG)-induced inflammatory model of depression in mice. Behavioral tests included locomotor activity, forced swim test (FST) and tail suspension test (TST). Oxidative stress was assessed by examining the levels of thiobarbituric acid reactive species (TBARS) and non-protein thiols (NP-SH) in homogenized whole brain samples. Comet assays were performed to assess neurotoxicity., Results: The results of this study demonstrate that BCG treatment resulted in an increase in duration of immobility in FST and TST as compared to the saline group. Further, it produced a significant increase in the brain TBARS levels and decrease in the brain NP-SH levels. The hippocampal tissue from BCG group had significantly more comet cells than the saline group. 1-MT and fluoxetine were able to reverse the BCG-induced depression-like behavior and the derangement in oxidative stress parameters. Fluoxetine and 1-MT also reversed the BCG-induced neurotoxicity in such mice., Conclusions: 1-Methyl-L-tryptophan exhibits antidepressant-like effect comparable to that of fluoxetine in treating BCG-induced depression-like behavior in mice.

Background: Soluble Klotho functions as an endocrine factor that plays important roles in a variety of pathophysiological processes. Soluble Klotho contains 130 KDa and 65 KDa isoforms. However, their distinct individual functional heterogeneity remains uncertain. Herein, we investigated the regulatory role of two soluble Klothos on cardiac fibrogenic responses., Methods and Results: The effect of soluble Klothos on myofibroblast differentiation, proliferation, and collagen synthesis/degradation were examined in cultured mouse cardiac myofibroblasts. The role of 130 KDa Klotho on fibrosis in hypertensive heart disease were examined in wild type (WT) and Klotho transgenic (Tg/+) mice receiving chronic angiotensin (Ang)II infusion. Our in vitro studies revealed that addition of 130 KDa soluble Klotho isoform increased collagen synthesis in a dose dependent manner. Furthermore, 130 KDa Klotho significantly stimulated myofibroblast differentiation, proliferation, and ERK phosphorylation, which were abolished by fibroblast growth factor (FGF) receptor antagonist (SU5402). In contrast, 65 KDa soluble Klotho treatment significantly suppressed myofibroblast proliferation and collagen synthesis. In vivo study further demonstrated that chronic AngII infusion lead to cardiac fibrosis in both WT and Tg/+ mice. However, cardiac collagen, TGF-β1, TIMP-2, and α-smooth muscle actin (SMA) levels were markedly upregulated in Tg/+ mice compared to WT cohort., Conclusion: Taken together, these findings implicate that 130 KDa soluble Klotho plays a stimulatory role in cardiac myofibroblast growth and activity through FGF pathway, whereas 65 KDa soluble Klotho exerts an anti-fibrotic effect in cardiac myofibroblasts. Thus, two distinct isoforms of soluble Klotho appear to play the counter-regulatory roles in cardiac fibrogenic responses., (© American Journal of Hypertension, Ltd 2016. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Introduction: Vaccinating a buffer of individuals around a case (ring vaccination) has the potential to target those who are at highest risk of infection, reducing the number of doses needed to control a disease. We explored the potential vaccine effectiveness (VE) of oral cholera vaccines (OCVs) for such a strategy., Methods and Findings: This analysis uses existing data from a cluster-randomized clinical trial in which OCV or placebo was given to 71,900 participants in Kolkata, India, from 27 July to 10 September 2006. Cholera surveillance was then conducted on 144,106 individuals living in the study area, including trial participants, for 5 y following vaccination. First, we explored the risk of cholera among contacts of cholera patients, and, second, we measured VE among individuals living within 25 m of cholera cases between 8 and 28 d after onset of the index case. For the first analysis, individuals living around each index case identified during the 5-y period were assembled using a ring to define cohorts of individuals exposed to cholera index cases. An index control without cholera was randomly selected for each index case from the same population, matched by age group, and individuals living around each index control were assembled using a ring to define cohorts not exposed to cholera cases. Cholera attack rates among the exposed and non-exposed cohorts were compared using different distances from the index case/control to define the rings and different time frames to define the period at risk. For the VE analysis, the exposed cohorts were further stratified according to the level of vaccine coverage into high and low coverage strata. Overall VE was assessed by comparing the attack rates between high and low vaccine coverage strata irrespective of individuals' vaccination status, and indirect VE was assessed by comparing the attack rates among unvaccinated members between high and low vaccine coverage strata. Cholera risk among the cohort exposed to cholera cases was 5-11 times higher than that among the cohort not exposed to cholera cases. The risk gradually diminished with an increase in distance and time. The overall and indirect VE measured between 8 and 28 d after exposure to a cholera index case during the first 2 y was 91% (95% CI 62%-98%) and 93% (95% CI 44%-99%), respectively. VE persisted for 5 y after vaccination and was similar whether the index case was a young child (<5 y) or was older. Of note, this study was a reanalysis of a cholera vaccine trial that used two doses; thus, a limitation of the study relates to the assumption that a single dose, if administered quickly, will induce a similar level of total and indirect protection over the short term as did two doses., Conclusions: These findings suggest that high-level protection can be achieved if individuals living close to cholera cases are living in a high coverage ring. Since this was an observational study including participants who had received two doses of vaccine (or placebo) in the clinical trial, further studies are needed to determine whether a ring vaccination strategy, in which vaccine is given quickly to those living close to a case, is feasible and effective., Trial Registration: ClinicalTrials.gov NCT00289224., Competing Interests: I have read the journal's policy and the authors of this manuscript have the following competing interests: FJL received grants from the Bill and Melinda Gates Foundation. All other authors have no competing interests.

Adenosine monophosphate-activated protein kinase (AMPK) is a protein kinase involved in maintaining energy homeostasis within cells. On the basis of human genetic association data, AMPK activators were pursued for the treatment of diabetic nephropathy. Identification of an indazole amide high throughput screening (HTS) hit followed by truncation to its minimal pharmacophore provided an indazole acid lead compound. Optimization of the core and aryl appendage improved oral absorption and culminated in the identification of indole acid, PF-06409577 (7). Compound 7 was advanced to first-in-human trials for the treatment of diabetic nephropathy.

Post-translational protein modifications such as citrullination have been linked to the breach of immune tolerance and clinical autoimmunity. Previous studies from our laboratory support this concept, demonstrating that autoantibodies targeting citrullinated isoforms of heat shock protein 90 (HSP90) are associated with rheumatoid arthritis complicated by interstitial lung disease. To further explore the relationship between citrullination and structural determinants of HSP90 immunogenicity, we employed a combination of ELISA-based epitope profiling, computational modeling, and mass-spectrometric sequencing of peptidylarginine deiminase (PAD)-modified protein. Remarkably, ELISAs involving selected citrullinated HSP90β/α peptides identified a key epitope corresponding to an internal Arg residue (R502 [HSP90β]/R510 [HSP90α]) that is normally buried within the crystal structure of native/unmodified HSP90. In vitro time/dose-response experiments reveal an ordered pattern of PAD-mediated deimination events culminating in citrullination of R502/R510. Conventional as well as scaled molecular dynamics simulations further demonstrate that citrullination of selected Arg residues leads to progressive disruption of HSP90 tertiary structure, promoting exposure of R502/R510 to PAD modification and subsequent autoantibody binding. Consistent with this process, ELISAs incorporating variably deiminated HSP90 as substrate Ag indicate a direct relationship between the degree of citrullination and the level of ex vivo Ab recognition. Overall, these data support a novel structural paradigm whereby citrullination-induced shifts in protein structure generate cryptic epitopes capable of bypassing B cell tolerance in the appropriate genetic context., (Copyright © 2016 by The American Association of Immunologists, Inc.)

Background: Water-related diseases are of great concern in developing countries like Nepal. Every year, there are countless morbidity and mortality due to the consumption of unsafe drinking water. Recently, there have been increased uses of bottled drinking water in an assumption that the bottled water is safer than the tap water and its use will help to protect from water-related diseases. So, the main objective of this study was to analyze the bacteriological quality of bottled drinking water and that of municipal tap water., Methods: A total of 100 samples (76 tap water and 24 bottled water) were analyzed for bacteriological quality and pH. The methods used were spread plate method for total plate count (TPC) and membrane filter method for total coliform count (TCC), fecal coliform count (FCC), and fecal streptococcal count (FSC). pH meter was used for measuring pH., Results: One hundred percent of the tap water samples and 87.5 % of the bottled water samples were found to be contaminated with heterotrophic bacteria. Of the tap water samples, 55.3 % were positive for total coliforms, compared with 25 % of the bottled water. No bottled water samples were positive for fecal coliforms and fecal streptococci, in contrast to 21.1 % and 14.5 % of the tap water samples being contaminated with fecal coliforms and fecal streptococci, respectively. One hundred percent of the tap water samples and 54.2 % of the bottled water samples had pH in the acceptable range., Conclusions: All of the municipal tap water samples and most of the bottled drinking water samples distributed in Dharan municipality were found to be contaminated with one or more than one type of indicator organisms. On the basis of our findings, we may conclude that comparatively, the bottled drinking water may have been safer (than tap water) to drink.

In the present study, we investigated whether chromium (Cr) administered to the dams (F0) during lactation period could affect memory and oxidative stress in F1 generation mice in their adulthood and whether quercetin could modulate these effects. Morris water maze (MWM) was used to test for spatial memory. Passive avoidance task and elevated plus maze were used to test for acquisition and retention memory. Oxidative stress was evaluated by measuring glutathione-S-transferase (GST), catalase activity and malonaldehyde (MDA) levels in the brain tissue. The results of MWM showed that the animals in the Cr-treated group compared to control have better spatial memory that was further enhanced when Cr was administered along with quercetin (50 mg/kg). The elevated plus maze test also showed the Cr-treated group to improve acquisition as well as retention memory compared to control. Co-treatment with quercetin (all doses) also exhibited enhanced acquisition and retention memory compared to control. The passive avoidance task demonstrated no significant improvement in memory in the Cr-treated mice but co-treatment with quercetin (100 mg/kg) showed improved acquisition memory compared to control which was significantly better than the animals treated with chromium alone. GST activity was significantly increased in the Cr-treated animals, and this was further increased in groups treated with Cr and quercetin (all doses). Chromium when administered alone and in combination with quercetin (all doses) significantly reduced MDA levels. However, Cr treatment did not show significant change in catalase activity. Nevertheless, co-treatment with quercetin (25 and 50 mg/kg) resulted in significant decrease in catalase activity. Thus, our study demonstrates that Cr exposure during lactation could be beneficial for pups with respect to augmentation of cognitive function and reduction of oxidative stress. Quercetin could probably enhance this effect to some extent.

Background and Purpose: Ghrelin increases growth hormone secretion, gastric acid secretion, gastric motility and hunger but decreases glucose-dependent insulin secretion and insulin sensitivity in humans. Antagonizing the ghrelin receptor has potential as a therapeutic approach in the treatment of obesity and type 2 diabetes. Therefore, the aim was to pharmacologically characterize the novel small-molecule antagonist PF-05190457 and assess translational pharmacology ex vivo., Experimental Approach: Radioligand binding in filter and scintillation proximity assay formats were used to evaluate affinity, and europium-labelled GTP to assess functional activity. Rat vagal afferent firing and calcium imaging in dispersed islets were used as native tissues underlying food intake and insulin secretion respectively., Key Results: PF-05190457 was a potent and selective inverse agonist on constitutively active ghrelin receptors and acted as a competitive antagonist of ghrelin action, with a human Kd of 3 nM requiring 4 h to achieve equilibrium. Potency of PF-05190457 was similar across different species. PF-05190457 increased intracellular calcium within dispersed islets and increased vagal afferent firing in a concentration-dependent manner with similar potency but was threefold less potent as compared with the in vitro Ki in recombinant overexpressing cells. The effect of PF-05190457 on rodent islets was comparable with glibenclamide, but glucose-dependent and additive with the insulin secretagogue glucagon-like peptide-1., Conclusions and Implications: Together, these data provide the pharmacological in vitro and ex vivo characterization of the first ghrelin receptor inverse agonist, which has advanced into clinical trials to evaluate the therapeutic potential of blocking ghrelin receptors in obesity and type 2 diabetes., (© 2016 The British Pharmacological Society.)

Blockers of the renin-angiotensin-aldosterone system (RAAS), such as angiotensin-converting enzyme inhibitors and angiotensin receptor blockers are routinely used in patients with chronic kidney disease because of their cardiovascular (CV) and renoprotective effects. However, there are no uniform recommendations about RAAS blockers for CV protection in the end-stage renal disease (ESRD) population other than the preferred drug class for blood pressure control. This uncertainty stems from the fact that patients with ESRD were generally excluded from randomized controlled trials evaluating the cardioprotective benefits of RAAS blockers. It is important to weigh the potential harms associated with the use of RAAS blockers, such as electrolyte disturbances and worsening anemia, with their role in protection of residual kidney function, alleviation of thirst and potential CV benefits. The objective of this review is to summarize the current knowledge about the use of RAAS blockers in patients with ESRD., (Copyright © 2016 Southern Society for Clinical Investigation. Published by Elsevier Inc. All rights reserved.)

Oxygen and carbon isotope ratios in planktonic foraminifera Globigerina bulloides collected from tow samples along a transect from the equatorial Indian ocean to the Southern Ocean (45°E and 80°E and 10°N to 53°S) were analysed and compared with the equilibrium δ(18)O and δ(13)C values of calcite calculated using the temperature and isotopic composition of the water column. The results agree within ~0.25‰ for the region between 10°N and 40°S and 75-200 m water depth which is considered to be the habitat of Globigerina bulloides. Further south (from 40°S to 55°S), however, the measured δ(18)O and δ(13)C values are higher than the expected values by ~2‰ and ~1‰ respectively. These enrichments can be attributed to either a 'vital effect' or a higher calcification rate. An interesting pattern of increase in the δ(13)C(DIC) value of the surface water with latitude is observed between 35°S and~ 60°S, with a peak at~ 42°S. This can be caused by increased organic matter production and associated removal. A simple model accounting for the increase in the δ(13)C(DIC) values is proposed which fits well with the observed chlorophyll abundance as a function of latitude.

Vascular endothelial growth factor (VEGF)-D is a crucial mediator of angiogenesis. Following myocardial infarction (MI), cardiac VEGF-D and VEGF receptor (VEGFR)-3 are significantly upregulated. In addition to endothelial cells, myofibroblasts at the site of MI highly express VEGFR-3, implicating the involvement of VEGF-D in cardiac fibrogenesis that promotes repair and remodeling. The aim of the current study was to further explore the critical role of VEGF-D in fibrogenic response in myofibroblasts. Myofibroblast proliferation, migration, collagen synthesis, and degradation were investigated in cultured cardiac myofibroblasts subjected to VEGF-D with/without VEGFR antagonist or ERK inhibitor. Vehicle-treated cells served as controls. Myofibroblast proliferation and migration were detected by BrdU assay and Boyden Chamber method, respectively. Expression of type I collagen, metalloproteinase (MMP)-2/-9, tissue inhibitor of MMP (TIMP)-1/-2, and ERK phosphorylation were evaluated by Western blot analyses. Our results revealed that compared to controls, (1) VEGF-D significantly increased myofibroblast proliferation and migration; (2) VEGF-D significantly upregulated type I collagen synthesis in a dose- and time-dependent manner; (3) VEGFR antagonist abolished VEGF-D-induced myofibroblast proliferation and type I collagen release; (4) VEGF-D stimulated MMP-2/-9 and TIMP-1/-2 synthesis; (5) VEGF-D activated ERK phosphorylation; and (6) ERK inhibitor abolished VEGF-D-induced myofibroblast proliferation and type I collagen synthesis. Our in vitro studies have demonstrated that VEGF-D serves as a crucial profibrogenic mediator by stimulating myofibroblast growth, migration and collagen synthesis. Further studies are underway to determine the role of VEGF-D in fibrous tissue formation during cardiac repair following MI.

Objectives: To evaluate the role of oral ondansetron in facilitating successful rehydration of under-5-year-old children suffering from acute diarrhea with vomiting and some dehydration., Study Design: Children (n = 170) aged 3 months to 5 years with acute diarrhea with vomiting and some dehydration were enrolled in this double blind, randomized, placebo-controlled trial. The participants were randomized to receive either single dose of oral ondansetron (n = 85) or placebo (n = 85) in addition to standard management of dehydration according to World Health Organization guidelines. Failure of oral rehydration therapy (ORT), administration of unscheduled intravenous fluids, and amount of oral rehydration solution intake in 4 hours were the primary outcomes. Secondary outcome measures included duration of dehydration correction, number of vomiting episodes, adverse effects, and caregiver satisfaction., Results: Failure of ORT was significantly less in children receiving ondansetron compared with those receiving placebo (31% vs 62%; P < .001; relative risk 0.50, 95% CI 0.35-0.72). Almost one-half of the children in the ondansetron group received intravenous fluids compared with those in the placebo group, but it was not statistically significant (P = .074; relative risk 0.56, 95% CI 0.30-1.07). The oral rehydration solution consumption was significantly more in the ondansetron group (645 mL vs 554 mL; mean difference 91 mL; 95% CI: 35-148 mL). Patients in the ondansetron group also showed faster rehydration, lesser number of vomiting episodes, and better caregiver satisfaction., Conclusion: A single oral dose of ondansetron, given before starting ORT to children <5 years of age with acute diarrhea and vomiting results in better oral rehydration., Trial Registration: Clinical Trial Registry of India: CTRI-2011/07/001916., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Background: The test-negative design (TND) has emerged as a simple method for evaluating vaccine effectiveness (VE). Its utility for evaluating oral cholera vaccine (OCV) effectiveness is unknown. We examined this method's validity in assessing OCV effectiveness by comparing the results of TND analyses with those of conventional cohort analyses., Methods: Randomized controlled trials of OCV were conducted in Matlab (Bangladesh) and Kolkata (India), and an observational cohort design was used in Zanzibar (Tanzania). For all three studies, VE using the TND was estimated from the odds ratio (OR) relating vaccination status to fecal test status (Vibrio cholerae O1 positive or negative) among diarrheal patients enrolled during surveillance (VE= (1-OR)×100%). In cohort analyses of these studies, we employed the Cox proportional hazard model for estimating VE (=1-hazard ratio)×100%)., Results: OCV effectiveness estimates obtained using the TND (Matlab: 51%, 95% CI:37-62%; Kolkata: 67%, 95% CI:57-75%) were similar to the cohort analyses of these RCTs (Matlab: 52%, 95% CI:43-60% and Kolkata: 66%, 95% CI:55-74%). The TND VE estimate for the Zanzibar data was 94% (95% CI:84-98%) compared with 82% (95% CI:58-93%) in the cohort analysis. After adjusting for residual confounding in the cohort analysis of the Zanzibar study, using a bias indicator condition, we observed almost no difference in the two estimates., Conclusion: Our findings suggest that the TND is a valid approach for evaluating OCV effectiveness in routine vaccination programs., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Rationale: CO2 and O2 can exchange their oxygen isotopes rapidly in the presence of hot (~670 °C) platinum and this has led to a method for determining the δ(17)O value of a CO2 sample. We have improved the method to achieve a precision of 0.008 ‰ (1-σ standard deviation) in the determination of δ(17)O values. Such high precision is essential to identify the stratospheric component in tropospheric CO2 and use it for global carbon flux studies. The crucial issue in the accurate determination of the δ(17)O value is estimation of a correction factor, which depends on the amount ratio CO2/O2. An attempt was also made to investigate the mechanism of exchange with their controlling parameters., Methods: The oxygen isotopes of a CO2 sample gas are exchanged with those of an appropriate amount of tank O2 in the presence of hot platinum. The pre-exchange CO2 and O2 gas samples as well as the post-exchange O2 sample are analyzed by isotope ratio mass spectrometry. A mixing model was developed involving the δ(18)O value of the CO2 and δ(17)O and δ(18)O values of pre- and post-exchange O2 to obtain the δ(17)O value of the CO2 sample. A correction to the measured value was determined to obtain the actual value with high accuracy and precision., Results: To obtain a precision better than 0.01 ‰ requires the amount ratio CO2/O2 to be controlled to better than ~15 %. We also find that the oxygen isotopes are nearly homogeneously distributed between the O2 and the CO2 molecules. In addition, determination of the (16) O(13)C(18)O/(16)O(12)C(16)O isotopologue ratio in the CO2 shows that the abundance of (16)O(13)C(18)O is close to that expected for random partitioning of the isotopes among the CO2 isotopologues., Conclusions: The isotopic scrambling between O2 and CO2 that occurs on hot platinum allows one to accurately determine the δ(17)O values of CO2 through isotopic analysis of O2., (Copyright © 2015 John Wiley & Sons, Ltd.)

Cadmium (Cd) is a known pollutant present in the environment at low levels and is reported to affect reproduction in many ways. The present study was undertaken to explore the effect of Cd in F1 generation mice on cognitive parameters, and to further investigate whether quercetin could modulate these effects. In this study, female lactating mice were exposed to cadmium for seven days just after delivery. The new born pups in their adulthood were tested for learning and memory parameters by passive avoidance task and Morris water maze (MWM) test. It was observed that pups exposed to Cd showed significant impairment of memory in step down latency test, which was reversed by quercetin (100 mg/kg). In MWM test for spatial memory, animals exposed to Cd exhibited increased escape latency, which was reversed by quercetin (50 mg/kg) significantly. Quercetin alone (50 and 100 mg/kg) also demonstrated improved spatial memory, and showed improved retention memory in the passive avoidance paradigm at dose 50 mg/kg. On testing oxidative stress parameters, we observed significantly increased malondialdehyde (MDA) levels in brain tissue of Cd-treated mice. Moreover, co-treatment with quercetin (50 mg/kg) and Cd significantly reduced these MDA levels. The other doses (25 and 100 mg/kg) also showed reduction in MDA levels as compared to the group exposed to Cd alone, though the difference was not statistically significant. Hence, this study highlights the possibility of cognitive impairment in adulthood if there is Cd exposure during lactation and oxidative stress could possibly attribute to this effect.

Signalling pathways that control endothelial cell (EC) permeability, leukocyte adhesion and inflammation are pivotal for atherosclerosis initiation and progression. Here we demonstrate that the Sterile-20-like mitogen-activated protein kinase kinase kinase kinase 4 (MAP4K4), which has been implicated in inflammation, is abundantly expressed in ECs and in atherosclerotic plaques from mice and humans. On the basis of endothelial-specific MAP4K4 gene silencing and gene ablation experiments in Apoe(-/-) mice, we show that MAP4K4 in ECs markedly promotes Western diet-induced aortic macrophage accumulation and atherosclerotic plaque development. Treatment of Apoe(-/-) and Ldlr(-/-) mice with a selective small-molecule MAP4K4 inhibitor also markedly reduces atherosclerotic lesion area. MAP4K4 silencing in cultured ECs attenuates cell surface adhesion molecule expression while reducing nuclear localization and activity of NFκB, which is critical for promoting EC activation and atherosclerosis. Taken together, these results reveal that MAP4K4 is a key signalling node that promotes immune cell recruitment in atherosclerosis.

Mitogen-activated protein kinase 4 (MAP4K4) regulates the MEK kinase cascade and is implicated in cytoskeletal rearrangement and migration; however, identifying MAP4K4 substrates has remained a challenge. To ascertain MAP4K4-dependent phosphorylation events, we combined phosphoproteomic studies of MAP4K4 inhibition with in vitro assessment of its kinase specificity. We identified 235 phosphosites affected by MAP4K4 inhibition in cells and found that pTP and pSP motifs were predominant among them. In contrast, in vitro assessment of kinase specificity showed that MAP4K4 favors a pTL motif. We showed that MAP4K4 directly phosphorylates and coimmunoprecipitates with FERM, RhoGEF, and pleckstrin domain-containing protein 1 (FARP1). MAP4K4 inhibition in SH-SY5Y cells increases neurite outgrowth, a process known to involve FARP1. As FARP1 and MAP4K4 both contribute to cytoskeletal rearrangement, the results suggest that MAP4K4 exerts some of its effects on the cytoskeleton via phosphorylation of FARP1.

Background: Case-control studies have not been examined for their utility in assessing population-level vaccine protection in individually randomized trials., Methods: We used the data of a randomized, placebo-controlled trial of a cholera vaccine to compare the results of case-control analyses with those of cohort analyses. Cases of cholera were selected from the trial population followed for three years following dosing. For each case, we selected 4 age-matched controls who had not developed cholera. For each case and control, GIS was used to calculate vaccine coverage of individuals in a surrounding "virtual" cluster. Specific selection strategies were used to evaluate the vaccine protective effects., Results: 66,900 out of 108,389 individuals received two doses of the assigned regimen. For direct protection among subjects in low vaccine coverage clusters, we observed 78% (95% CI: 47-91%) protection in a cohort analysis and 84% (95% CI: 60-94%) in case-control analysis after adjusting for confounding factors. Using our GIS-based approach, estimated indirect protection was 52% (95% CI: 10-74%) in cohort and 76% (95% CI: 47-89%) in case control analysis. Estimates of total and overall effectiveness were similar for cohort and case-control analyses., Conclusion: The findings show that case-control analyses of individually randomized vaccine trials may be used to evaluate direct as well as population-level vaccine protection., (Copyright © 2015. Published by Elsevier Ltd.)

Recent studies in adipose tissue, pancreas, muscle, and macrophages suggest that MAP4K4, a serine/threonine protein kinase may be a viable target for antidiabetic drugs. As part of the evaluation of MAP4K4 as a novel antidiabetic target, a tool compound, 16 (PF-6260933) and a lead 17 possessing excellent kinome selectivity and suitable properties were delivered to establish proof of concept in vivo. The medicinal chemistry effort that led to the discovery of these lead compounds is described herein together with in vivo pharmacokinetic properties and activity in a model of insulin resistance.