Heterologous desensitization of response mediated by selective PKC-dependent phosphorylation of G(i-1) and G(i-2)

This study examined the ability of protein kinase C (PKC) toinduce heterologous desensitization by targeting specific G proteinsand limiting their ability to transduce signals in smooth muscle.Activation of PKC by pretreatment of intestinal smooth muscle cellswith phorbol 12-myristate 13-acetate, cholecystokinin octapeptide, orthe phosphatase 1 and phosphatase 2A inhibitor, calyculin A,selectively phosphorylated G_i-1 and G_i-2,but not G_i-3 or G_o, and blockedinhibition of adenylyl cyclase mediated by somatostatin receptorscoupled to G_i-1 and opioid receptors coupled toG_i-2, but not by muscarinic M₂ and adenosineA₁ receptors coupled to G_i-3. Phosphorylationof G_i-1 and G_i-2 and blockade of cyclaseinhibition were reversed by calphostin C and bisindolylmaleimide, andadditively by selective inhibitors of PKC and PKC. Blockade ofinhibition was prevented by downregulation of PKC. Phosphorylation ofG-subunits by PKC also affected responses mediated by-subunits. Pretreatment of muscle cells withcANP-(4-23), a selective agonist of the natriureticpeptide clearance receptor, NPR-C, which activates phospholipase C(PLC)-3 via the -subunits of G_i-1 andG_i-2, inhibited the PLC- response to somatostatin and[D-Pen^2,5]enkephalin. The inhibition waspartly reversed by calphostin C. Short-term activation of PKC had noeffect on receptor binding or effector enzyme (adenylyl cyclase orPLC-) activity. We conclude that selective phosphorylation ofG_i-1 and G_i-2 by PKC partly accounts forheterologous desensitization of responses mediated by the - and-subunits of both G proteins. The desensitization reflects adecrease in reassociation and thus availability of heterotrimeric G proteins.

     

Vaccine potential of poly-1-6 beta-D-N-succinylglucosamine, an immunoprotective surface polysaccharide of Staphylococcus aureus and Staphylococcus epidermidis

Staphylococcus aureus and S. epidermidis are among the most common causes of nosocomial infection, and S. aureus is also of major concern to human health due to its occurrence in community-acquired infections. These staphylococcal species are also major pathogens for domesticated animals. We have previously identified poly-N-succinyl beta-1-6 glucosamine (PNSG) as the chemical form of the S. epidermidis capsular polysaccharide/adhesin (PS/A) which mediates adherence of coagulase-negative staphylococci (CoNS) to biomaterials, serves as the capsule for strains of CoNS that express PS/A, and is a target for protective antibodies. We have recently found that PNSG is made by S. aureus as well, where it is an environmentally regulated, in vivo-expressed surface polysaccharide and similarly serves as a target for protective immunity. Only a minority of fresh human clinical isolates of S. aureus elaborate PNSG in vitro but most could be induced to do so under specific in vitro growth conditions. However, by immunofluorescence microscopy, S. aureus cells in infected human sputa and lung elaborated PNSG. The ica genes, previously shown to encode proteins in CoNS that synthesize PNSG, were found by PCR in all S. aureus strains examined, and immunogenic and protective PNSG could be isolated from S. aureus. Active and passive immunization of mice with PNSG protected them against metastatic kidney infections after intravenous inoculation with eight phenotypically PNSG-negative S. aureus. Isolates recovered from kidneys expressed PNSG, but expression was lost with in vitro culture. Strong antibody responses to PNSG were elicited in S. aureus infected mice, and a PNSG-capsule was observed by electron microscopy on isolates directly plated from infected kidneys. PNSG represents a previously unidentified surface polysaccharide of S. aureus that is elaborated during human and animal infection and is a prominent target for protective antibodies.     

Dietary input of microbes and host genetic variation shape among-population differences in stickleback gut microbiota

To explain differences in gut microbial communities we must determine how processes regulating microbial community assembly (colonization, persistence) differ among hosts and affect microbiota composition. We surveyed the gut microbiota of threespine stickleback (Gasterosteus aculeatus) from 10 geographically clustered populations and sequenced environmental samples to track potential colonizing microbes and quantify the effects of host environment and genotype. Gut microbiota composition and diversity varied among populations. These among-population differences were associated with multiple covarying ecological variables: habitat type (lake, stream, estuary), lake geomorphology and food- (but not water-) associated microbiota. Fish genotype also covaried with gut microbiota composition; more genetically divergent populations exhibited more divergent gut microbiota. Our results suggest that population level differences in stickleback gut microbiota may depend more on internal sorting processes (host genotype) than on colonization processes (transient environmental effects).     

Development of Modified-Release Tablets of Zolpidem Tartrate by Biphasic Quick/Slow Delivery System

Zolpidem tartrate is a non-benzodiazepine analogue of imidazopyridine of sedative and hypnotic category. It has a short half-life with usual dosage regimen being 5 mg, two times a day, or 10 mg, once daily. The duration of action is considered too short in certain circumstances. Thus, it is desirable to lengthen the duration of action. The formulation design was implemented by preparing extended-release tablets of zolpidem tartrate using the biphasic delivery system technology, where sodium starch glycolate acts as a superdisintegrant in immediate-release part and hydroxypropyl methyl cellulose as a release retarding agent in extended-release core. Tablets were prepared by direct compression. Both the core and the coat contained the drug. The pre-compression blends were evaluated for angle of repose, bulk density, and compressibility index. The tablets were evaluated for thickness, hardness, weight variation test, friability, and in vitro release studies. No interaction was observed between zolpidem tartrate and excipients from the Fourier transform infrared spectroscopy and differential scanning calorimetry analysis. The results of all the formulations prepared were compared with reference product Stilnoct®. Optimized formulations showed release patterns that match the United States Pharmacopeia (USP) guidelines for zolpidem tartrate extended-release tablets. The mechanism of drug release was studied using different mathematical models, and the optimized formulation has shown Fickian diffusion. Accelerated stability studies were performed on the optimized formulation.KEY WORDS: biphasic delivery system technology, hydroxypropyl methyl cellulose, modified release, sodium starch glycolate, zolpidem tartrate     

Birth Prevalence of Neural Tube Defects and Orofacial Clefts in India: A Systematic Review and Meta-Analysis

Background

In the last two decades, India has witnessed a substantial decrease in infant mortality attributed to infectious disease and malnutrition. However, the mortality attributed to birth defects remains constant. Studies on the prevalence of birth defects such as neural tube defects and orofacial clefts in India have reported inconsistent results. Therefore, we conducted a systematic review of observational studies to document the birth prevalence of neural tube defects and orofacial clefts.

Methods

A comprehensive literature search for observational studies was conducted in MEDLINE and EMBASE databases using key MeSH terms (neural tube defects OR cleft lip OR cleft palate AND Prevalence AND India). Two reviewers independently reviewed the retrieved studies, and studies satisfying the eligibility were included. The quality of included studies was assessed using selected criteria from STROBE statement.

Results

The overall pooled birth prevalence (random effect) of neural tube defects in India is 4.5 per 1000 total births (95% CI 4.2 to 4.9). The overall pooled birth prevalence (random effect) of orofacial clefts is 1.3 per 1000 total births (95% CI 1.1 to 1.5). Subgroup analyses were performed by region, time period, consanguinity, and gender of newborn.

Conclusion

The overall prevalence of neural tube defects from India is high compared to other regions of the world, while that of orofacial clefts is similar to other countries. The majority of studies included in the review were hospital based. The quality of these studies ranged from low to moderate. Further well-designed, high quality community-based observational studies are needed to accurately estimate the burden of neural tube defects and orofacial clefts in India.     

Global Spectrum of Copy Number Variations Reveals Genome Organizational Plasticity and Proposes New Migration Routes

Global spectrum of CNVs is required to catalog variations to provide a high-resolution on the dynamics of genome-organization and human migration. In this study, we performed genome-wide genotyping using high-resolution arrays and identified 44,109 CNVs from 1,715 genomes across 12 populations. The study unraveled the force of independent evolutionary dynamics on genome-organizational plasticity across populations. We demonstrated the use of CNV tool to study human migration and identified a second major settlement establishing new migration routes in addition to existing ones.     

Bioactivity-guided isolation of mosquitocidal constituents from the rhizomes of Plumbago capensis Thunb

A bioassay-guided fractionation and chemical examination of chloroform extract of Plumbago capensis roots resulted in isolation and characterization of two new napthaquinone derivatives (4, 8) along with six known compounds (1–3, 5–7). Their structures were determined on the basis of extensive spectroscopic (IR, MS, 1D and 2D NMR) data analysis and by comparison with the literature data. All the compounds were tested for their mosquito larvicidal activity against fourth instar larvae of Aedes aegypti, and compared with that of rotenone. Among the tested compounds, isoshinanolone (3) and plumbagin (1) showed excellent toxicity with LC₅₀ values of 1.26 and 5.43 μg/mL. New compound (8) displayed moderate toxicity against the tested mosquito species.     

Laccase production from oil palm industry solid waste: Statistical optimization of selected process parameters

Oil palm frond parenchyma tissue was used as a solid substrate for the production of laccase via solid‐state fermentation using the white rot fungus Pycnoporus sanguineus. With a rectangular aluminium tray as solid‐state fermentation bioreactor, process parameters such as bed height, moisture and supplemented nitrogen (as urea solution) levels were studied and optimized using a statistical design of experiment. The moisture level exerted a significant effect on the process. The interaction effect observed between bed height and supplemented nitrogen level suggested that uniform distribution of supplemented nitrogen into the substrate bed was important. The proposed regression model sufficiently predicted the process response over the experimental range tested. The optimum parameter combination for laccase production was a 3‐cm bed height, 72% w/w moisture and 0.21% w/v supplemented nitrogen. Laccase productivity remained constant when the tray size was increased from 1.4 to 3.4‐fold.     

Somatic embryogenesis and plant regeneration from cell suspension culture of niger (Guizotia abyssinica Cass.)

Somatic embryogenesis was achieved from cell suspension cultures of niger (Guizotia abyssinica Cass.). Initially, friable embryogenic calluses were induced from cotyledonary leaves of niger on Murashige and Skoog (MS) agar medium containing 5 μM 2,4-Dichlorophenoxyacetic acid (2,4-D) and 0.5 μM kinetin (KIN). Cell suspension cultures were established by using embryogenic calluses in MS liquid medium containing 5 μM 2,4-D and 0.5 μM KIN. Initiation of somatic embryogenesis and development up to globular stage from embryogenic cell clumps occurred in the liquid medium itself. Thereafter embryogenic cell aggregates were transferred to MS agar medium supplemented with 3 μM KIN for embryo differentiation, whereas maturation of somatic embryos occurred in MS agar medium containing 10 μM abscisic acid.     

A role for ATP-citrate lyase, malic enzyme, and pyruvate/citrate cycling in glucose-induced insulin secretion

In pancreatic beta-cells, metabolic coupling factors generated during glucose metabolism and pyruvate cycling through anaplerosis/cataplerosis processes contribute to the regulation of insulin secretion. Pyruvate/citrate cycling across the mitochondrial membrane leads to the production of malonyl-CoA and NADPH, two candidate coupling factors. To examine the implication of pyruvate/citrate cycling in glucose-induced insulin secretion (GIIS), different steps of the cycle were inhibited in INS 832/13 cells by pharmacological inhibitors and/or RNA interference (RNAi) technology: mitochondrial citrate export, ATP-citrate lyase (ACL), and cytosolic malic enzyme (ME1). The inhibitors of the di- and tri-carboxylate carriers, n-butylmalonate and 1,2,3-benzenetricarboxylate, respectively, reduced GIIS, indicating the importance of transmitochondrial transport of tri- and dicarboxylates in the action of glucose. To directly test the role of ACL and ME1 in GIIS, small hairpin RNA (shRNA) were used to selectively decrease ACL or ME1 expression in transfected INS 832/13 cells. shRNA-ACL reduced ACL protein levels by 67%, and this was accompanied by a reduction in GIIS. The amplification/K(ATP)-independent pathway of GIIS was affected by RNAi knockdown of ACL. The ACL inhibitor radicicol also curtailed GIIS. shRNA-ME1 reduced ME1 activity by 62% and decreased GIIS. RNAi suppression of either ACL or ME1 did not affect glucose oxidation. However, because ACL is required for malonyl-CoA formation, inhibition of ACL expression by shRNA-ACL decreased glucose incorporation into palmitate and increased fatty acid oxidation in INS 832/13 cells. Taken together, the results underscore the importance of pyruvate/citrate cycling in pancreatic beta-cell metabolic signaling and the regulation of GIIS.