Postprandial Hypertriglyceridemia Predicts Development of Insulin Resistance Glucose Intolerance and Type 2 Diabetes

Insulin resistance (IR) and type 2 diabetes mellitus (T2DM) have been found to be associated with postprandial hypertriglyceridemia (PPHTg). However, whether PPHTg can cause IR and diabetes is not clear. We therefore investigated the role of PPHTg in development of T2DM in rat model of T2DM. 96 male Wistar rats were randomized into four groups (24 rats each). Control Group A, high sucrose diet (HSD) Group B, HSD+Pioglitazone (10mg/kg/day) Group C and HSD+Atorvastatin (20mg/kg/day) Group D. Fat and glucose tolerance tests were done at regular intervals in all groups besides insulin and body weight measurement. At 26 weeks, low dose streptozotocin (15mg/kg,i.p.) was given to half of the rats. All rats were followed up till 48 weeks. PPHTg developed as early as week 2 in Group B and stabilized by week 14. Group B displayed highest PPHTg compared to other groups. Atorvastatin treatment (Group D) abolished PPHTg which became comparable to controls, pioglitazone treatment partially blunted PPHTg resulting in intermediate PPHTg. Group B with highest PPHTg showed highest subsequent IR, glucose intolerance (GI) and highest incidence of prediabetes at week 26 and diabetes at week 34 and 46 compared to other groups. Group D rats displayed lower IR, GI, low incidence of prediabetes and diabetes at these time points compared to Groups B and C. ROC analysis showed that triglyceride area under the curve of each time point significantly predicts the risk of diabetes. Present study provides the evidence that PPHTg predicts the development of IR, GI and T2DM in rat model of diet induced T2DM.     

In Silico Derived Peptides for Inhibiting the Toxin–Antitoxin Systems of Mycobacterium tuberculosis: Basis for Developing Peptide-Based Therapeutics

Toxin–antitoxin (TA) systems of Mycobacterium tuberculosis (Mtb) is a prerequisite for the bacterium to survive in extreme conditions. Antimicrobial peptides inhibiting the formation of these complexes provide a novel strategy for TB drug discovery process. Absence of TA genes in human, makes these systems as an attractive target for drug development. In this study using Peptiderive server, we have derived a number of potential inhibitory peptides for nine TA complexes—VapBC3, VapBC5, VapBC11, VapBC15, VapBC26, VapBC30, RelBE2, RelJK, MazEF4 of Mtb. We have studied about the common interacting toxin residues with the antitoxin and with the derived peptide. Further, using Cluspro server, we compared the binding efficacy of the in silico derived peptides with the published potential peptides for the toxins VapC26, VapC30 and MazF. Thus, these in silico derived peptides would serve as basis for developing peptide based therapeutics for TA complexes of Mtb.

     

Completion of the Vimentin Rod Domain Structure Using Experimental Restraints: A New Tool for Exploring Intermediate Filament Assembly and Mutations

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Pharmacokinetic stability of macrocyclic peptide triazole HIV‐1 inactivators alone and in liposomes

Previously, we reported the discovery of macrocyclic peptide triazoles (cPTs) that bind to HIV‐1 Env gp120, inhibit virus cell infection with nanomolar potencies, and cause irreversible virion inactivation. Given the appealing virus‐killing activity of cPTs and resistance to protease cleavage observed in vitro, we here investigated in vivo pharmacokinetics of the cPT AAR029b. AAR029b was investigated both alone and encapsulated in a PEGylated liposome formulation that was designed to slowly release inhibitor. Pharmacokinetic analysis in rats showed that the half‐life of FITC‐AAR029b was substantial both alone and liposome‐encapsulated, 2.92 and 8.87 hours, respectively. Importantly, liposome‐encapsulated FITC‐AAR029b exhibited a 15‐fold reduced clearance rate from serum compared with the free FITC‐cPT. This work thus demonstrated both the in vivo stability of cPT alone and the extent of pharmacokinetic enhancement via liposome encapsulation. The results obtained open the way to further develop cPTs as long‐acting HIV‐1 inactivators against HIV‐1 infection.     

Differential modulation of voltage-dependent K+ currents in colonic smooth muscle by oxidants

The effect of oxidants on voltage-dependent K⁺ currents was examined in mouse colonic smooth muscle cells. Exposure to either chloramine-T (Ch-T), an agent known to oxidize both cysteine and methionine residues, or the colon-specific oxidant monochloramine (NH₂Cl) completely suppressed the transient outward K⁺ current (I_to) while simultaneously enhancing the sustained delayed rectifier K⁺ current (I_dr). In contrast, the cysteine-specific oxidants hydrogen peroxide (H₂O₂) and 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB) exhibited partial and slow suppression of I_to by inducing a shift in channel availability of -18 mV without affecting I_dr. After enhancement by NH₂Cl or Ch-T, I_dr was sensitive to 10 mM tetraethylammonium but not to other K⁺ channel blockers, suggesting that it represented activation of the resting I_dr and not a separate K⁺ conductance. Extracellular dithiothreitol (DTT) partially reversed the effect of H₂O₂ and DTNB on I_to but not the actions of NH₂Cl and Ch-T on either I_dr or I_to. Dialysis of myocytes with GSH (5 mM) or DTT (5 mM) prevented suppression of I_to by H₂O₂ and DTNB but did not alter the effects of NH₂Cl or Ch-T on either I_dr or I_to. Ch-T and NH₂Cl completely blocked I_to generated by murine K_v4.1, 4.2, and 4.3 in Xenopus oocytes, an effect not reversible by intracellular DTT. In contrast, intracellular DTT reversed the effect of H₂O₂ and DTNB on the cloned channels. These results suggest that I_to is suppressed via modification of both methionine and cysteine residues, whereas enhancement of I_dr likely results from methionine oxidation alone. colon; colitis; redox; ion channel     

Heterologous complementation of the exopolysaccharide synthesis and carbon utilization phenotypes of Sinorhizobium meliloti Rm1021 polyhydroxyalkanoate synthesis mutants

A reduced exopolysaccharide phenotype is associated with inability to synthesize polyhydroxyalkanaote (PHA) stores in Sinorhizobium meliloti strain Rm1021. Loss of function mutations in phbB and phbC result in non-mucoid colony morphology on Yeast Mannitol Agar, compared to the mucoid phenotype exhibited by the parental strain. This phenotype is attributed to reduction in succinoglycan synthesis. We have used complementation of this phenotype and the previously described D-3-hydroxybutyrate/acetoacetate utilization phenotype to isolate a heterologous clone containing a Bradyrhizobium japonicum phbC gene. Sequence analysis confirmed that this clone contains one of the five predicted phbC genes in the B. japonicum genome. The described phenotypic complementation strategy should be useful for isolation of novel PHA synthesis genes of diverse origin.     

Inactivation of Cyanobacterial Nitrogenase After Exposure to Ultraviolet-B Radiation

Exposure of the N₂-fixing cyanobacterium Anabaena BT2 to ultraviolet-B radiation (2.5 W m⁻²) for 30 min resulted in complete loss of nitrogenase activity but 100% cell killing occurred only after a 90-min exposure. Inactivation of nitrogenase activity was not specific to Anabaena BT2; other species also showed a similar effect. The time required for 100% killing and inactivation of nitrogenase activity differed in various species, and this difference may be ascribed to the presence of different levels of UV-B protection mechanisms in individual species. Inhibition of nitrogenase activity was immediate, since exposure of cultures to UV-B for as little as 5 min elicited some inhibition of activity. The activity of UV-B-inhibited nitrogenase did not recover upon transfer of exposed cells to fluorescent light, suggesting that the inhibition may be due to specific inactivation of the enzyme. By employment of inhibitors of protein synthesis and PS-II activity, it was demonstrated that restoration of nitrogenase activity in a UV-B-treated culture occurred by fresh synthesis of nitrogenase polypeptide. Our findings suggest that estimation of nitrogenase activity in diazotrophic species may be used as a marker enzyme for assessing the impact of UV-B radiation. Received: 13 June 2002 / Accepted: 22 July 2002     

Effect of ovariectomy and estrogen supplementation on brain acetylcholinesterase activity and passive-avoidance learning in rats

The effect of ovariectomy and estrogen treatment on the brain acetylcholinesterase activity and cognition in rats was investigated in this study. Ovariectomized and nonovariectomized rats were treated subcutaneously with estradiol dipropionate for 8 d. In the single-trial, passive-avoidance test all the groups showed significant learning and retention of memory as evident by the increase in transfer latency time in trial 2 as compared with trial 1. No-transfer response was significantly increased in the estradiol-dipropionate-treated ovariectomized (80%) and nonovariectomized (60%) group as compared with the ovariectomized (30%) group. Specific activity of acetylcholinesterase was assayed spectrophotometrically in salt-soluble and detergent-soluble fractions of various brain areas: frontal cortex, cerebral cortex, striatum, hippocampus and hypothalamus, thalamus, pons, medulla, and cerebellum. The effect of ovariectomy and estradiol dipropionate was varied in both fractions of these brain areas. Estradiol dipropionate treatment could restore the acetylcholinesterase activity to the control level only in the detergent-soluble fraction of hypothalamus and salt-soluble fraction of hypothalamus, thalamus, and medulla in ovariectomized rats. The results indicate that ovariectomy alters acetylcholinesterase activity in the brain areas but not in a uniform manner and affects only qualitative aspects of cognitive function, which could be improved by estrogen supplementation.     

Role of p38 mitogen-activated protein kinase phosphorylation and Fas-Fas ligand interaction in morphine-induced macrophage apoptosis

In this study, we evaluated the molecular mechanisms involved in morphine-induced macrophage apoptosis. Both morphine and TGF-beta promoted P38 mitogen-activated protein kinase (MAPK) phosphorylation, and this phosphorylation was inhibited by SB 202190 as well as by SB 203580. Anti-TGF-beta Ab as well as naltrexone (an opiate receptor antagonist) inhibited morphine-induced macrophage P38 MAPK phosphorylation. Anti-TGF-beta Ab also attenuated morphine-induced p53 as well as inducible NO synthase expression; in contrast, N(G)-nitro-L-arginine methyl ester, an inhibitor of NO synthase, inhibited morphine-induced P38 MAPK phosphorylation and Bax expression. Morphine also enhanced the expression of both Fas and Fas ligand (FasL), whereas anti-FasL Ab prevented morphine-induced macrophage apoptosis. Moreover, naltrexone inhibited morphine-induced FasL expression. In addition, macrophages either deficient in FasL or lacking p53 showed resistance to the effect of morphine. Inhibitors of both caspase-8 and caspase-9 partially prevented the apoptotic effect of morphine on macrophages. In addition, caspase-3 inhibitor prevented morphine-induced macrophage apoptosis. These findings suggest that morphine-induced macrophage apoptosis proceeds through opiate receptors via P38 MAPK phosphorylation. Both TGF-beta and inducible NO synthase play an important role in morphine-induced downstream signaling, which seems to activate proteins involved in both extrinsic (Fas and FasL) and intrinsic (p53 and Bax) cell death pathways.