Up-regulation of nicotinic receptors by nicotine varies with receptor subtype

Recent evidence suggests that in addition to alpha4beta2 and alpha3-containing nicotinic receptors, alpha6-containing receptors are present in midbrain dopaminergic neurons and involved in the nicotine reward pathway. Using heterologous expression, we found that alpha6beta2, like alpha3beta2 and alpha4beta2 receptors, formed high affinity epibatidine binding complexes that are pentameric, trafficked to the cell surface, and produced acetylcholine-evoked currents. Chronic nicotine exposure up-regulated alpha6beta2 receptors with differences in up-regulation time course and concentration dependence compared with alpha4beta2 receptors, the predominant high affinity nicotine binding site in brain. The alpha6beta2 receptor up-regulation required higher nicotine concentrations than for alpha4beta2 but lower than for alpha3beta2 receptors. The alpha6beta2 up-regulation occurred 10-fold faster than for alpha4beta2 and slightly faster than for alpha3beta2. Our data suggest that nicotinic receptor up-regulation is subtype-specific such that alpha6-containing receptors up-regulate in response to transient, high nicotine exposures, whereas sustained, low nicotine exposures up-regulate alpha4beta2 receptors.     

Tropical soil microflora of spice-based cropping systems as potential antagonists of root-knot nematodes

Suppression of plant parasitic nematodes with nematode predators, parasites or antagonists is an eco-friendly approach than the toxic chemicals. In a study, soil borne fungi from the rhizosphere of major spice crops were collected from diverse cropping systems prevailing in three southern states of India. A series of in vitro studies were conducted using 73 freshly collected fungal isolates and 76 isolates obtained from other sources. Out of this 67 isolates were not parasitic on females of root-knot nematodes whereas 115 isolates, though colonized the egg masses, did not show any signs of parasitism on nematode eggs. Fifty-nine isolates showed 50-90% inhibition in egg hatch. Pochonia chlamydospora, Verticillium lecanii, Paecilomyces lilacinus, and few isolates of Trichoderma spp. showed >25% parasitism on root-knot nematode eggs. The most promising isolates in this study were one isolate each of Aspergillus (F.45), Fusarium (F.47), and Penicillium (F.59); three each isolates of Trichoderma (F.3, F.52, and F.60) and Pochonia (F.30 and Vc.3) Verticillium (Vl); and two isolates of fungi that could not be identified (F.28 and F.62). Parasitism by Aspergillus tamarii, Aspergillus ustus, Drechslera sp., Humicola sp., and Scopulariopsis sp. on root-knot nematode eggs or females, reported in the present study, are new reports.     

Supplementation of vitamin E and selenium prevents hyperoxaluria in experimental urolithic rats

Renal injury is considered as one of the prerequisites for calcium oxalate retention. In order to determine the role of lipid peroxidation related effects for hyperoxaluria, we evaluated the alterations in lipid peroxidation, antioxidants and oxalate synthesizing enzymes in lithogenic rats with response to vitamin E + selenium treatment. In kidney of lithogenic rats, the level of lipid peroxidation and the activities of oxalate synthesizing enzymes were found to be increased whereas the levels/activities of non-enzymatic and enzymatic antioxidants were found to be decreased. The urinary excretion of both oxalate and calcium were significantly elevated. Supplementation of lithogenic rats with vitamin E + selenium decreased the levels of lipid peroxides and the activities of oxalate synthesizing enzymes like glycolic acid oxidase (GAO), lactate dehydrogenase (LDH), xanthine oxidase (XO) with a concomitant increase in the activities of enzymatic antioxidants like superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and glucose-6-phosphate dehydrogenase (G6PDH) and increased levels of non-enzymatic antioxidants like ascorbic acid, alpha-tocopherol and reduced glutathione (GSH). The urinary excretion of oxalate and calcium were normalized. The antioxidants vitamin E + selenium thereby protected from hyperoxaluria.     

Development of a Quantitative Competitive Reverse Transcription Polymerase Chain Reaction (QC-RT–PCR) for Detection and Quantitation of Chikungunya Virus

Chikungunya is one of the most important emerging arboviral infections of public health significance. Due to lack of a licensed vaccine, rapid diagnosis plays an important role in early management of patients. In this study, a QC-RT–PCR assay was developed to quantify Chikungunya virus (CHIKV) RNA by targeting the conserved region of E1 gene. A competitor molecule containing an internal insertion was generated, which provided a stringent control of the quantification process. The introduction of 10-fold serially diluted competitor in each reaction was further used to determine sensitivity. The applicability of this assay for quantification of CHIKV RNA was evaluated with human clinical samples, and the results were compared with real-time quantitative RT–PCR. The sensitivity of this assay was estimated to be 100 RNA copies per reaction with a dynamic detection range of 10² to 10¹⁰ copies. Specificity was confirmed using closely related alpha and flaviviruses. The comparison of QC-RT–PCR result with real-time RT–PCR revealed 100% concordance for the detection of CHIKV in clinical samples. These findings demonstrated that the reported assay is convenient, sensitive and accurate method and has the potential usefulness for clinical diagnosis due to simultaneous detection and quantification of CHIKV in acute-phase serum samples.     

Synthesis and antibacterial activity of new N-linked 5-triazolylmethyl oxazolidinones

A new series of N-linked 5-triazolylmethyl oxazolidinones with varying substitution at the piperazine nitrogen 4-position were synthesized and tested against a panel of Gram-positive and Gram-negative bacteria including clinical isolates. Most of the compounds showed excellent antibacterial activity against susceptible and resistant Gram-positive organisms. One of the compounds showed enhanced antibacterial activity against Moraxella catarrhalis.     

Structure-Activity Relationship and Mode of Action of a Frog Secreted Antibacterial Peptide B1CTcu5 Using Synthetically and Modularly Modified or Deleted (SMMD) Peptides

All life forms are equipped with rapidly acting, evolutionally conserved components of an innate immune defense system that consists of a group of unique and diverse molecules known as host defense peptides (HDPs). A Systematic and Modular Modification and Deletion (SMMD) approach was followed to analyse the structural requirement of B1CTcu5, a brevinin antibacterial peptide amide identified from the skin secretion of frog Clinotarsus curtipes, India, to show antibacterial activity and to explore the active core region. Seventeen SMMD-B1CTcu5 analogs were designed and synthesised by C and N-terminal amino acid substitution or deletion. Enhancement in cationicity by N-terminal Lys/Arg substitution or hydrophobicity by Trp substitution produced no drastic change in bactericidal nature against selected bacterial strains except S. aureus. But the sequential removal of N-terminal amino acids had a negative effect on bactericidal potency. Analog B1CTcu5-LIAG obtained by the removal of four N-terminal amino acids displayed bactericidal effect comparable to, or in excess of, the parent peptide with reduced hemolytic character. Its higher activity was well correlated with the improved inner membrane permeabilisation capacity. This region may act as the active core of B1CTcu5. Presence of C-terminal disulphide bond was not a necessary condition to display antibacterial activity but helped to promote hemolytic nature. Removal of the C-terminal rana box region drastically reduced antibacterial and hemolytic activity of the peptide, showing that this region is important for membrane targeting. The bactericidal potency of the D-peptide (DB1CTcu5) helped to rule out the stereospecific interaction with the bacterial membrane. Our data suggests that both the C and N-terminal regions are necessary for bactericidal activity, even though the active core region is located near the N-terminal of B1CTcu5. A judicious modification at the N-terminal region may produce a short SMMD analog with enhanced bactericidal activity and low toxicity against eukaryotic cells.     

In vitro radioprotection studies of organoselenium compounds: differences between mono- and diselenides

Organoselenium compounds belonging to the class of monoselenides, such as selenomethionine (SeM) and methylselenocysteine (MSeCys) and diselenides including selenocystine (SeCys) and selenopropionic acid (SePA), were examined for their comparative radioprotective effects using in vitro models. Effects of these compounds on the inhibition of γ-radiation induced lipid peroxidation in liposomes, protein carbonylation in bovine serum albumin (BSA) and strand breaks in pBR322 plasmid DNA, assessed, respectively, by the formation of thiobarbituric acid reactive substances, formation of 2,2′-dinitrophenyl hydrazine (DNPH) carbonyl complex and horizontal gel electrophoresis, were used to compare their radioprotective ability. The IC₅₀ values for SeCys, SePA, SeM and MSeCys for lipid peroxidation were 27 ± 1, 33 ± 2, 200 ± 8 and 163 ± 4 μM, respectively, and the values for inhibition of protein carbonylation were >200, 300 ± 6, 464 ± 8 and 436 ± 3 μM, respectively. Inhibition of DNA strand break formation was tested at 200 μM for all the compounds and SePA and SeCys exhibited a protective effect on DNA, while SeM and MSeCys did not lead to any protection. The in vitro cytotoxicity studies in normal and tumor cells revealed that MSeCys and SeM were not cytotoxic to lymphocytes and EL4 tumor cells at the concentrations employed. In contrast, SeCys was toxic, with a higher effect on tumor cells than lymphocytes. Our studies suggest that the non-toxic diselenides like SePA should be explored as protective agents against γ-irradiation induced damage.     

Molecular docking analysis of compounds from Lycopersicon esculentum with the insulin receptor to combat type 2 diabetes

It is known that tomato (Lycopersicon esculentum) contains bioactive compounds to combat type-2 diabetes. Therefore, it is of interest to document data from the molecular docking analysis of compounds from Lycopersicon esculentum with the insulin receptors to combat type-2 diabetes. We report the binding features of cinnamic acid, chlorogenic acid, gallic acid & glucoside with insulin receptors for further consideration.     

Molecular probes for the A2A adenosine receptor based on a pyrazolo[4,3-e][1,2,4]triazolo[1,5-c]pyrimidin-5-amine scaffold

Pyrazolo[4,3-e][1,2,4]triazolo[1,5-c]pyrimidin-5-amine derivatives such as SCH 442416 display high affinity and selectivity as antagonists for the human A_2A adenosine receptor (AR). We extended ether-linked chain substituents at the p-position of the phenyl group using optimized O-alkylation. The conjugates included an ester, carboxylic acid and amines (for amide condensation), an alkyne (for click chemistry), a fluoropropyl group (for ¹⁸F incorporation), and fluorophore reporter groups (e.g., BODIPY conjugate 14, K_i 15 nM). The potent and A_2AAR-selective N-aminoethylacetamide 7 and N-[2-(2-aminoethyl)-aminoethyl]acetamide 8 congeners were coupled to polyamidoamine (PAMAM) G3.5 dendrimers, and the multivalent conjugates displayed high A_2AAR affinity. Theoretical docking of an AlexaFluor conjugate to the receptor X-ray structure highlighted the key interactions between the heterocyclic core and the binding pocket of the A_2AAR as well as the distal anchoring of the fluorophore. In conclusion, we have synthesized a family of high affinity functionalized congeners as pharmacological probes for studying the A_2AAR.     

Potential role of kinins in the effects of taurine in high-fructose-fed rats

The present work investigates the involvement of kinins in the effects of taurine in fructose-fed hypertensive rats. The effects of taurine on blood pressure, plasma glucose, insulin, and the insulin sensitivity index were determined. Angiotensin-converting enzyme (ACE) activity and nitrite content in plasma, plasma and tissue kallikrein activity, and taurine content were also investigated. The blood pressure changes in response to the coadministration of inhibitors of the synthesis of nitric oxide (NO), prostaglandins (PGs), or a kinin receptor blocker along with taurine was also evaluated. Fructose-fed rats had higher blood pressure and elevated plasma levels of glucose and insulin. Kallikrein activity, taurine, and nitrite contents were significantly lower in fructose-fed rats as compared with controls. The increases in systolic blood pressure, hyperglycemia, and hyperinsulinemia were controlled by taurine administration in fructose-fed rats. ACE activity was lower, while nitrite and taurine content and kallikrein activity were higher, in taurine-supplemented rats as compared with fructose-fed rats. A significant increase in blood pressure was observed in rats cotreated with the inhibitors Hoe 140 (a kinin receptor blocker), L-NAME (a NO synthase inhibitor), or indomethacin (a PG synthesis inhibitor) with taurine for 1 week as compared with taurine-treated fructose-fed rats. This suggests that the antihypertensive effect of taurine in fructose-fed rats was blocked by the inhibitors. Augmented kallikrein activity and, hence, increased kinin availability may be implicated in the effects of taurine in fructose-fed hypertensive rats.