Regulatory Role of Nitric Oxide in the Reduced Survival of Erythrocytes in Visceral Leishmaniasis

Background: Nitric oxide (NO) plays a vital role in maintaining the survivability of circulating erythrocytes. Here we have investigated whether NO depletion associated with visceral leishmaniasis (VL) is responsible for the reduced survival of erythrocytes observed during the disease. Methods: Infected hamsters were treated with standard anti-leishmanial sodium stibogluconate (SAG) and NO donor isosorbide dinitrate (ISD). Erythrophagocytosis by macrophages was determined by labelling the cells with FITC followed by flow cytometry. Aggregation of band3 was estimated from band3 associated EMA fluorescence. Caspase 3 activity was measured using immunosorbent assay kit. Phosphatidylserine (PS) externalization and cell shrinkage were determined using annexin V. Aminophspholipid translocase and scramblase activities were measured following NBD-PS and NBD-PC internalization, respectively. Results: Impairment of both synthesis and uptake of NO resulted in decreased bioavailability of this signaling molecule in erythrocytes in VL. NO level was replenished after simultaneous treatment with ISD and SAG. Combination treatment decreased red cell apoptosis in infected animals by deactivating caspase 3 through s-nitrosylation. Drug treatment prevented infection-mediated ATP depletion and altered calcium homeostasis in erythrocytes. Improved metabolic environment effectively amended dysregulation of aminophospholipid translocase and scramblase, which in turn reduced cell shrinkage, and exposure of phosphatidylserine on the cell surface under the diseased condition. Conclusion and general significance: In this study, we have identified NO depletion to be an important factor in promoting premature hemolysis with the progress of leishmanial infection. The study implicates NO to be a possible target for future drug development towards the promotion of erythrocyte survival in VL

Combination Therapy with Andrographolide and D-Penicillamine Enhanced Therapeutic Advantage over Monotherapy with D-Penicillamine in Attenuating Fibrogenic Response and Cell death in the Periportal zone of Liver in Rats during Copper Toxicosis

Long treatment regime with D-penicillamine is needed before it can exert clinically meaningful benefits in the treatment of copper toxicosis. The consequence of long-term D-penicillamine treatment is associated with numerous side effects. The limitations of D-penicillamine monotherapy prompted us to search for more effective treatment strategies that could decrease the duration of D-penicillamine therapy. The present study was designed to evaluate the therapeutic potential of D-penicillamine in combination with another hepatoprotective drug, andrographolide in treatment of copper toxicosis in rats. D-penicillamine treatment led to the excretion of copper through urine. Addition of andrographolide to D-penicillamine regime appeared to increase protection of liver by increasing the biliary excretion of copper and reduction in cholestatic injury. The early removal of the causative agent copper during combination treatment was the most effective therapeutic intervention that contributed to the early rectification of fibrosis in liver. Combination treatment reduced Kupffer cells accumulation and TNFα production in liver of copper exposed rats. In particular, andrographolide mediated the anti-inflammatory effect by inhibiting the cytokine production. However, another possible mechanism of cytoprotection of andrographolide was decreasing mitochondrial production of superoxide anions that resulted in better restoration of mitochondrial dysfunction during combination therapy than monotherapy. Furthermore, ROS inhibition by combination regimen resulted in significant decline in activation of caspase cascade. Inhibition of caspases attenuated apoptosis of hepatocytes, induced by chronic copper exposure. In summary, this study suggested that added benefit of combination treatment over use of either agent alone in alleviating the hepatotoxicity and fibrosis associated with copper toxicosis

Product Inhibition of Biological Hydrogen Production in Batch Reactors

In this paper, the inhibitory effects of added hydrogen in reactor headspace on fermentative hydrogen production from acidogenesis of glucose by a bacterium, Clostridium acetobutylicum, was investigated experimentally in a batch reactor. It was observed that hydrogen itself became an acute inhibitor of hydrogen production if it accumulated excessively in the reactor headspace. A mathematical model to simulate and predict biological hydrogen production process was developed. The Monod model, which is a simple growth model, was modified to take inhibition kinetics on microbial growth into account. The modified model was then used to investigate the effect of hydrogen concentration on microbial growth and production rate of hydrogen. The inhibition was moderate as hydrogen concentration increased from 10% to 30% (v/v). However, a strong inhibition in microbial growth and hydrogen production rate was observed as the addition of H2 increased from 30% to 40% (v/v). Practically, an extended lag in microbial growth and considerably low hydrogen production rate were detected when 50% (v/v) of the reactor headspace was filled with hydrogen. The maximum specific growth rate (µmax), substrate saturation constant (ks), a critical hydrogen concentration at which microbial growth ceased (H2*) and degree of inhibition were found to be 0.976 h−1, 0.63 ± 0.01 gL, 24.74 mM, and 0.4786, respectivel

Diazirine-based photo-crosslinkers for defect free fabrication of solution processed organic light-emitting diodes

The fabrication of solution deposited OLEDs is fraught with difficulties, largely due to the interlayer mixing and surface erosion during sequential deposition of the layers. We demonstrate that these problems can be circumvented by using photopolymerizable diazirine-based cross-linker capable of converting soluble organic materials into highly cross-linked insoluble networks. 3-Trifluoromethyl(aryl)diazirines photolyze readily upon the 10–15 min exposure of 365 nm UV irradiation to generate carbenes, which react with polymers or small molecules via C–H bond insertion producing highly cross-linked materials. This photo-generated cross-linking does not require any catalyst, initiator or short-wavelength UV light and is performed at room temperature, releasing molecular nitrogen as the only byproduct. To show the cross-linked polymer layers do not display inter-layer mixing, we deposited red-emitting regioregular poly(3-hexylthiophene-2,5-diyl) (P3HT) over cross-linked (10% cross-linker) blue emitting dioctyl polyfluorene (PFO) layer. The overlaid layers showed clear and well-defined boundary with no interlayer mixing. The surface morphology of the solution deposited layers was investigated by AFM to show that the cross-linked layers exhibited significant decrease in surface roughness. This is also shown on the example of the hole transporting material 4,4′-bis[N-(1-naphthyl)-N-phenylamino]-biphenyl (NPB) which displayed roughness average to decrease from 6.4 nm to 1.0 nm. The effect of decreased surface roughness on the performance of phosphorescent OLEDs was investigated by fabricating devices with configuration of ITO/PEDOT:PSS/NPB:(0%/5%/10%) cross-linker/MCP:6% Ir(mppy)3/TPBI/CsF/Al. Following the diazirine-mediated cross-linking, the OLEDs displayed a decrease in turn-on voltage from 3.8 V to 3.0 V along with a six-fold enhancement of external quantum efficiency (EQEmax) from 1.1% to 6.8% and maximum luminous efficiency increase from 3.8 cd A−1 to 22.9 cd A−1. These results demonstrate that the simple diazirine mediated photo-cross-linking using mild conditions compatible with organic layers is a promising strategy for improving the performance of the solution-processed OLEDs.</p

S-allyl Cysteine in Combination with Clotrimazole Downregulates Fas Induced Apoptotic Events in Erythrocytes of Mice exposed to Lead

Background: Chronic lead (Pb2+) exposure leads to the reduced lifespan of erythrocytes. Oxidative stress and K+ loss accelerate Fas translocation into lipid raft microdomains inducing Fas mediated death signaling in these erythrocytes. Pathophysiological-based therapeutic strategies to combat against erythrocyte death were evaluated using garlic-derived organosulfur compounds like diallyl disulfide (DADS), S allyl cysteine (SAC) and imidazole based Gardos channel inhibitor clotrimazole (CLT). Methods: Morphological alterations in erythrocytes were evaluated using scanning electron microscopy. Events associated with erythrocyte death were evaluated using radio labeled probes, flow cytometry and activity gel assay. Mass spectrometry was used for detection of GSH–4-hydroxy-trans-2-nonenal (HNE) adducts. Fas redistribution into the lipid rafts was studied using immunoblotting technique and confocal microscopy. Results: Combination of SAC and CLT was better than DADS and CLT combination and monotherapy with these agents in prolonging the survival of erythrocytes during chronic Pb2+ exposure. Combination therapy with SAC and CLT prevented redistribution of Fas into the lipid rafts of the plasma membrane and downregulated Fas-dependent death events in erythrocytes of mice exposed to Pb2+. Conclusion and general significance: Ceramide generation was a critical component of Fas receptor-induced apoptosis, since inhibition of acid sphingomyelinase (aSMase) interfered with Fas-induced apoptosis during Pb2+ exposure. Combination therapy with SAC and CLT downregulated apoptotic events in erythrocytes by antagonizing oxidative stress and Gardos channel that led to suppression of ceramide-initiated Fas aggregation in lipid rafts. Hence, combination therapy with SAC and CLT may be a potential therapeutic option for enhancing the lifespan of erythrocytes during Pb2+ toxicity

Synovial IL-9 facilitates neutrophil survival, function and differentiation of Th17 cells in rheumatoid arthritis

Abstract Background Role of Th9 cells and interleukin-9 (IL-9) in human autoimmune diseases such as psoriasis and ulcerative colitis has been explored only very recently. However, their involvement in human rheumatoid arthritis (RA) is not conclusive. Pathogenesis of RA is complex and involves various T cell subsets and neutrophils. Here, we aimed at understanding the impact of IL-9 on infiltrating immune cells and their eventual role in synovial inflammation in RA. Methods In vitro stimulation of T cells was performed by engagement of anti-CD3 and anti-CD28 monoclonal antibodies. Flow cytometry was employed for measuring intracellular cytokine, RORγt in T cells, evaluating apoptosis of neutrophils. ELISA was used for measuring soluble cytokine, Western blot analysis and confocal microscopy were used for STAT3 phosphorylation and nuclear translocation. Results We demonstrated synovial enrichment of Th9 cells and their positive correlation with disease activity (DAS28-ESR) in RA. Synovial IL-9 prolonged the survival of neutrophils, increased their matrix metalloprotienase-9 production and facilitated Th17 cell differentiation evidenced by induction of transcription factor RORγt and STAT3 phosphorylation. IL-9 also augmented the function of IFN-γ + and TNF-α + synovial T cells. Conclusions We provide evidences for critical role of IL-9 in disease pathogenesis and propose that targeting IL-9 may be an effective strategy to ameliorate synovial inflammation in RA. Inhibiting IL-9 may have wider impact on the production of pathogenic cytokines involved in autoimmune diseases including RA and may offer better control over the disease

Additional file 1: Table S1. of Synovial IL-9 facilitates neutrophil survival, function and differentiation of Th17 cells in rheumatoid arthritis

Demographic and clinical characteristics of RA patients (n = 28). (DOCX 12 kb

Oxidation of hemoglobin and redistribution of band 3 promote erythrophagocytosis in visceral leishmaniasis

In visceral leishmaniasis (VL), oxidative assault on erythrocytes perturbs their cellular environment and makes them vulnerable to premature hemolysis. In this study, we assessed the contribution of oxidation-induced modifications of hemoglobin and membrane protein band 3 in the reduced survival of red cells in VL. Oxidative transformation of oxyhemoglobin to hemichrome enhanced its interaction with erythrocyte membrane in the infected animals. Association between denatured globin and band 3 contributed to the formation of insoluble copolymer of macromolecular dimension. Disulfide bonding appeared to be necessary in the making of high molecular weight aggregates during copolymerization. Hemichrome induced clustering of band 3 promoted generation of epitopes on erythrocyte cell surface. This provided a signal favoring immunologic recognition of redistributed band 3 by autologous IgG followed by erythrophagocytosis. An eventual outcome of the sequence of events pointed to early removal of affected red cells from circulation during the disease