Sulfamethizole attenuates poloxamer 407-induced atherosclerotic neointima formation via inhibition of mTOR in C57BL/6 mice

Mammalian target of Rapamycin C1 (mTORC1) inhibition limits plaque progression in atherosclerosis. The present study evaluated the protective effect of sulfamethizole on poloxamer 407-induced atherosclerotic neointima formation in C57BL/6 mice via mTOR inhibition. Poloxamer 407 (P-407) (0.5 g/kg body weight) was administered intraperitoneally to male C57BL/6 mice every third day for 148 days to induce chronic hyperlipidemia. From Day 121 to 148, animals were additionally administered Sulfamethizole (5, 10, and 50 mg/kg, p.o.), Rapamycin (0.5 mg/kg, positive control), or vehicle (1 ml/kg). Plasma lipid levels were measured on Days 120 and 148. Upon sacrifice, histological studies were performed, and aortic tissue interleukin (IL)-6, tumor necrosis factor-α (TNF-α), and mTOR levels were evaluated. A molecular docking study was carried out to mimic the interaction of sulfamethizole with mTOR protein. Chronic P-407 administration significantly (p < 0.001) elevated plasma lipid levels, compared with those of the normal control group. Chronic hyperlipidemia resulted in increased tunica intima thickness, collagen deposition, and IL-6, TNF-α, and mTOR levels. Treatment with Sulfamethizole attenuated these parameters significantly in a dose-dependent manner. Molecular docking studies showed a significant interaction of Sulfamethizole with mTOR. In conclusion, this study suggests that sulfamethizole significantly limits poloxamer 407-induced atherosclerotic neointima formation in C57BL/6 mice via mTOR inhibition.     

(2')3',5'-Bisphosphate nucleotidase

(2')3',5'-Bisphosphate nucleotidase has been prepared in electrophoretically homogeneous form from guinea pig liver. The enzyme catalyzes the hydrolysis of the 2'- or 3'-phosphate from the appropriate nucleoside 2',5'- and 3',5'-bisphosphates and is active with 3'-phosphoadenosine 5'-phosphosulfate and with coenzyme A but not with ATP. The 40,000-dalton protein is a monomer that requires Mg2+ for activity.     

Binding and precipitating activities ofErythrina lectins with complex type carbohydrates and synthetic cluster glycosides. A comparative study of the lectins fromE. corallodendron,E. cristagalli,E. flabelliformis,andE. indica

Erythrina lectins possess similar structural and carbohydrate binding properties. Recently, tri- and tetra-antennary complex type carbohydrates with non-reducing terminal galactose residues have been shown to be precipitated as tri- and tetravalent ligands, respectively, with certainErythrina lectins [Bhattacharyya L, Haraldsson M, Brewer CF (1988) Biochemistry 271034-41]. The present work describes a comparative study of the binding and precipitating activities of fourErythrina lectins,viz. E. corallodendron, E. cristagalli, E. flabelliformis, andE. indica, with multi-antennary complex type carbohydrates and synthetic cluster glycosides. The results show that though their binding affinities are very similar, theErythrina lectins show large differences in their precipitating activities with the carbohydrates. The results also indicate significant dependence of the precipitating activities of the lectins on the core structure of the carbohydrates. These findings provide a new dimension to the structure-activity relationship of the lectins and their interactions with asparagine-linked carbohydrates.Abbreviations EAL, ECorL, ECL, EFL, and EIL represent the lectins from the seeds ofErythrina arborescens, - E. corallodendron, E. cristagalli, E. flabelliformis, andE. indica respectively - AFOS thetri-antennary complex type oligosaccharide from asialofetuin - AFGP the tri-antennary glycopeptide from asialofetuin - MeGal methyl -d-galactopyranoside Unless stated otherwise all sugars are in thed-configuration.     

Chromium(III)-insulin derivatives and their implication in glucose metabolism

Insulin has been reacted with five chromium(III) complexes that are capable of relatively facile substitution of aquo ligands. The new Cr(III) insulin derivatives have been characterized by means of electronic and infrared spectra, and evidence for major changes in the protein structure, including the state of aggregation, has been presented. Supporting evidence for the arguments favoring the beneficiary role of chromium(III) in glucose metabolism has been obtained using in vivo studies, and it has been shown that insulin derived with Cr(salen) (H2O)2+ is capable of reversing the blood sugar, serum cholesterol, and phospholipids levels to those of normal rats. The results emphasize the dependence of biopotency on the structure of Cr(III) complexes used for derivation of insulin and discount the postulates that Cr(III) serves to assemble insulin and receptor units through metal-sulphur bonding. The influence of Cr(III) on the structural stability and state of aggregation of insulin and their possible role in glucose metabolism is discussed.     

Chymotrypsin-catalyzed hydrolysis of chromium (III) derivatives of insulin: evidence for stabilization of the protein through interactions with metal ions

We investigated the chymotrypsin-promoted hydrolysis of a series of chromium(III)-insulin complexes containing chelating or macrocyclic ligands. It has been shown that Cr(III) stabilizes insulin against the chymotrypsin-promoted hydrolysis of the protein. The molecular weights of Cr(III) containing peptides have been estimated to be of the order of 2,700-3,700 daltons. The Cr(III) containing peptides are richer in glutamic acid than the intact insulin and are devoid of any isoleucine. High molecular weights and the observed glutamic acid/histidine ratios in Cr(III) containing peptides have been rationalized in terms of Cr(III) being associated with insulin aggregates rather than the monomer of the protein. The chymotrypsin hydrolysis of Cr(III) insulin derivatives is influenced markedly by the nature, charge, and type of Cr(III) complex with which the protein has been reacted. Arguments have been advanced that chymotrypsin-promoted hydrolysis of insulin Cr(III) derivatives does not lead to cleavages at or near every tyrosine residue.     

Characteristics of tripeptide transport in human jejunal brush-border membrane vesicles

These studies are aimed at characterizing the transport of the tripeptide, glycylglycyl-L-proline (GlyGlyPro) across human jejunal brush-border membrane vesicles. GlyGlyPro (0.65 mM) was hydrolyzed by brush-border membrane vesicles with the extent of hydrolysis per mg protein being 23% at 0.5 min, 57% at 1 min and complete hydrolysis at 60 min. Treatment of the membrane vesicles with gel-complexed papain (to remove membrane peptidases) resulted in minimal hydrolysis of GlyGlyPro up to 10 min of incubation. Measurement of GlyGlyPro influx with papain-treated vesicles in the presence of increasing medium osmolarity showed that uptake occurred into an osmotically reactive intravesicular space. Transport of GlyGlyPro with normal and papain-treated membrane vesicles was similar in the presence of an inward Na+ or K+ gradient. No overshoot phenomenon was observed in the presence of an inward proton gradient (extravesicular pH 5.5; intravesicular pH 7.5). An interior negative membrane potential induced by a K+ diffusion potential in the presence of valinomycin stimulated the uptake of the peptide. The effect of increasing concentrations on initial rates of GlyGlyPro uptake revealed the presence of a saturable component as well as a diffusional component. Preloading the membrane vesicles with 20 mM glycylsarcosylsarcosine stimulated uptake by 4-fold. Uptake of GlyGlyPro was inhibited greater than 50% by dipeptides and tripeptides and less than 15% by free amino acids. These results indicate that GlyGlyPro uptake in jejunal brush-border membrane vesicles is not energized by a Na+ or proton gradient and that transport occurs by carrier-mediated and diffusional processes.     

Effect of cytokines on polymorphonuclear neutrophil infiltration in the mouse. Prostaglandin- and leukotriene-independent induction of infiltration by IL-1 and tumor necrosis factor

The i.p. injection of mice with highly purified recombinant human rIL-1 alpha or beta resulted in the rapid influx of a large number of polymorphonuclear neutrophils (PMN) into the peritoneal cavity. Significant increases in the number of PMN were induced by doses of IL-1 which ranged from 0.005 to 5 ng/injection. Interestingly the dose response for PMN influx was bell-shaped because 50 ng of IL-1 did not result in a significant increase in peritoneal PMN. IL-1 induced PMN infiltration was detectable by 1 h with peak levels of PMN obtained by about 2 h, followed by a subsequent decline by 24 h. Other cytokines, IL-2, IFN-gamma, IFN alpha beta, granulocyte-CSF, granulocyte-macrophage-CSF, IL-3, TNF-alpha, and TNF-beta were compared to IL-1 for their ability to induce a PMN influx into the peritoneum. Only TNF-alpha or TNF-beta (lymphotoxin) were able to induce a significant influx of PMN within 2 h. However, based on total protein administered, about 100 times more TNF than IL-1 was required to produce a comparable PMN infiltration. Intraperitoneal injection of inhibitors of the cyclooxygenase or lipoxygenase pathways did not inhibit the IL-1-induced influx of PMN. Also, neither IL-1 nor TNF triggered an increase in PG or leukotriene release from peritoneal cells in vitro. Furthermore, direct peritoneal injection of leukotriene B4, a potent PMN chemoattractant in vitro, did not induce any significant increase in PMN in the peritoneal cavity indicating that chemotactic activity alone is insufficient for inducing peritoneal infiltration. These results suggest that the local production of very low levels of IL-1 in vivo would be sufficient to initiate a sequence of events that results in a rapid accumulation of PMN. Because IL-1 was not chemotactic for PMN in vitro, our data suggest that IL-1 induces production of factors that are chemotactic for PMN. Alternatively, IL-1 may act on other stages of the complex sequence of events that regulates the emigration of PMN into tissue sites in vivo. The synergy apparent in PMN influx when suboptimal concentrations of IL-1 and TNF were injected suggests that the local production of very low concentrations of these cytokines in situ could play a critical role in the emigration of PMN during infection.     

Prevalence of Muellerius capillaris in free-ranging spotted deer (Cervus axis) in India and its experimental cross-transmission to goats

A survey to assess the prevalence of parasitic infections among axis deer (Cervus axis) in three National Parks in India revealed infections with the lungworm Muellerius capillaris. Clinical signs were not evident in infected animals. Therefore, it is suggested that C. axis is probably a carrier of the infection. Under laboratory conditions, terrestrial molluscs (Macrochalamys sp.) were infected with first stage larvae of M. capillaris collected from fecal pellets of C. axis. Feeding of third stage larvae collected from these snails on day 14 post exposure produced patent infections in goats. On day 31 post infection, adult M. capillaris could be collected from the lungs of infected goats. This study establishes the possibility of cross-transmission of M. capillaris between wild and domestic animals in India.     

Intracellular calcium does not appear to be essential for arachidonic acid release from stimulated macrophages as shown by studies with Quin-2

The present investigation was undertaken to study the potential role of intracellular calcium on the release of arachidonic acid from mouse peritoneal macrophages activated by inflammatory stimuli. The intracellular calcium concentration, as measured using fluorescent probe Quin-2, was 112 +/- 8.4 nM. The chelation of intracellular calcium with Quin-2 did not affect the release of arachidonic acid from macrophages upon stimulation with phorbol myristate acetate, opsonized zymosan or calcium ionophore A23187. However, the removal of calcium from the extracellular medium resulted in a 30-50% decrease in arachidonic acid release from phorbol myristate acetate- and zymosan-stimulated macrophages and also the stimulation of arachidonic acid release from calcium ionophore-stimulated cells were nullified. These studies indicated the existence of calcium-dependent and independent mechanisms modulating the release of arachidonic acid from macrophages subjected to inflammatory stimuli.