Free radical scavenging and anti-inflammatory activities of the extracts of Astraeus hygrometricus (Pers.) Morg.

The present study was aimed to evaluate free radical scavenging and anti-inflammatory potential of extracts of Astraeus hygrometricus - a tropical wild edible mushroom. Free radical scavenging potential of crude, boiled and ethanolic extracts was studied using different in vitro antioxidant models. The anti-inflammatory activity of the potential extract was evaluated in carrageenan and dextran induced acute and formalin induced chronic inflammatory model in mice. Among all the extracts, ethanolic extract possesses significant in vitro superoxide anion, hydroxyl radical scavenging and lipid peroxidation inhibition activities. The IC50 values of ethanolic extracts of A. hygrometricus represented 357.95, 81.2 and 87.96 μg/ml respectively. Furthermore, the ethanolic extract showed remarkable anti-inflammatory activity in all models comparable to the standard reference drug diclofenac. The results suggest that anti-inflammatory activity of the ethanolic extract of A. hygrometricus is possibly attributed to its free radical scavenging properties.Colegio de Farmacéuticos de la Provincia de Buenos Aire

Pesticide-induced oxidative stress in laboratory and field populations of native honey bees along intensive agricultural landscapes in two Eastern Indian states

International audienceAbstractPesticides have been cited as one of the major drivers of pollinator loss. However, little is known about pesticide impacts on natural populations of native honey bee species. This study looked into the effect of pesticides with respect to oxidative stress in the laboratory and in field populations of two native Indian honey bee species (Apis dorsata and A. cerana) by examining a combination of biomarkers, e.g., superoxide dismutase, catalase and xanthine oxidase. A significant upregulation of all three biomarkers was observed in both treated individuals in laboratory experiments and field populations sampled from a pesticide use gradient. This study reports, for the first time, an increase in expression of xanthine oxidase in an invertebrate system (honey bees) exposed to pesticides

Differential and conditional activation of PKC-isoforms dictates cardiac adaptation during physiological to pathological hypertrophy.

A cardiac hypertrophy is defined as an increase in heart mass which may either be beneficial (physiological hypertrophy) or detrimental (pathological hypertrophy). This study was undertaken to establish the role of different protein kinase-C (PKC) isoforms in the regulation of cardiac adaptation during two types of cardiac hypertrophy. Phosphorylation of specific PKC-isoforms and expression of their downstream proteins were studied during physiological and pathological hypertrophy in 24 week male Balb/c mice (Mus musculus) models, by reverse transcriptase-PCR, western blot analysis and M-mode echocardiography for cardiac function analysis. PKC-δ was significantly induced during pathological hypertrophy while PKC-α was exclusively activated during physiological hypertrophy in our study. PKC-δ activation during pathological hypertrophy resulted in cardiomyocyte apoptosis leading to compromised cardiac function and on the other hand, activation of PKC-α during physiological hypertrophy promoted cardiomyocyte growth but down regulated cellular apoptotic load resulting in improved cardiac function. Reversal in PKC-isoform with induced activation of PKC-δ and simultaneous inhibition of phospho-PKC-α resulted in an efficient myocardium to deteriorate considerably resulting in compromised cardiac function during physiological hypertrophy via augmentation of apoptotic and fibrotic load. This is the first report where PKC-α and -δ have been shown to play crucial role in cardiac adaptation during physiological and pathological hypertrophy respectively thereby rendering compromised cardiac function to an otherwise efficient heart by conditional reversal of their activation

Cardiomyocyte-specific regression of nitrosative stress-mediated S-Nitrosylation of IKKγ alleviates pathological cardiac hypertrophy

IKKγ prototypically promotes NFκBp65 activity by regulating the assembly of the IKK holocomplex. In hypertrophied cardiomyocytes, the p65-p300 complex-induced regenerative efforts are neutralized by the p53-p300 complex-mediated apoptotic load resulting in compromised cardiac function. The present study reports that nitrosative stress leads to S-Nitrosylation of IKKγ in hypertrophied cardiomyocytes in a pre-clinical model. Using a cardiomyocyte-targeted nanoconjugate, IKKγ S-Nitrosylation-resistant mutant plasmids were delivered to the pathologically hypertrophied heart that resulted in improved cardiac function by amelioration of cardiomyocyte apoptosis and simultaneous induction of their cell cycle re-entry machinery. Mechanistically, in IKKγ S-Nitrosyl mutant-transfected hypertrophied cells, increased IKKγ-p300 binding downregulated the binding of p53 and p65 with p300. This shifted the binding preference of p65 from p300 to HDAC1 resulting in upregulated expression of cyclin D1 and CDK2 via the p27/pRb pathway. This approach has therapeutic advantage over mainstream anti-hypertrophic remedies which concomitantly reduce the regenerative prowess of resident cardiomyocytes during hypertrophy upon downregulation of myocyte apoptosis. Therefore, cardiomyocyte-targeted delivery of IKKγ S-Nitrosyl mutants during hypertrophy can be exploited as a novel strategy to re-muscularize the diseased heart

Differential expression profile of Protein Kinase-C isoforms.

<p>(<b>A</b>) Western blot analyses showing change in expression of PKC α and PKC-δ (phospho and total) in groups C, H, E, H<i>^X</i> and E<i>^R</i>. PKC-δ cleavage product (41 kD) is exclusive in group H, H<i>^X</i> and E<i>^R</i>. RPL32 was used as loading control. (<b>B</b>) Western blot analyses showing phosphorylation of PKC-δ and PKC-α in adult cardiomyocytes isolated from different experimental groups. RPL32 was used as loading control. (<b>C</b>) Western blot analyses showing changes in expression of phosphorylated and total PKC-δ and PKC-α in exercise withdrawn animals (E<i>^R</i>) rested for different time periods. RPL32 was used as loading control. (<b>D</b>) Immunofluorescence study showing expression of phospho-PKC-δ and -α in different experimental group. Tissue sections showing phospho-PKC-δ expression in panels b, f, j, n and r and phospho-PKC-α in panels b’, f’, j’, n’ and r’ (green fluorescence). Sections were counter stained with alpha sarcomeric actinin antibody (panels c, g, k, o and s for PKC-δ and panels c’, g’, k’, o’ and s’ for PKC -α; red fluorescence). Nuclei were stained with DAPI (panels a, e, i, m and q for PKC-δ and panels a’, e’, i’, m’ and q’ for PKC-α; blue fluorescence) and merged images are shown in panels d, h, l, p and t for PKC-δ and panels d’, h’, l’, p’ and t’ for PKC-α. Increased expression of phospho-PKC-δ was observed in groups H and E<i>^R</i> whereas, phospho-PKC-α expression was induced in group E and H<i>^X</i> (Scale bar = 50 µm, Magnification = 40X).</p

Assessment of hypertrophy and estimation of collagen in all the experimental groups.

<p>(<b>A</b>) Graph showing HW/BW ratio in all five experimental models: pathological hypertrophy (H), physiological hypertrophy (E), exercise-trained pathological hypertrophy (H<i>^X</i>), mice kept at rest after 4 weeks of exercise training (E<i>^R</i>) and representative control (C) (*p<0.05 for H versus E or C; #p<0.05 for H versus H<i>^X</i>; ¶p<0.05 for E versus E<i>^R</i>). (<b>B</b>) Graph showing cardiomyocyte cross-sectional area (in µm²) in groups C, H, E, H<i>^X</i>, and E<i>^R</i> (***p<0.001 for H versus E or C; ##p<0.01 for H versus H<i>^X</i>; ¶p<0.05 for E versus E<i>^R</i>). (<b>C</b>) Expression profile of pathological hypertrophy markers (<i>ANF</i> and <i>β-MHC</i>) and physiological hypertrophy marker (<i>IGF-1</i>) estimated by RT-PCR. <i>GAPDH</i> was used as loading control. (<b>D</b>) Graph showing ventricular collagen concentration in groups C, H, E, H<i>^X</i>, and E<i>^R</i> estimated by hydroxyproline assay.</p

PKC-δ associated downstream target proteins.

<p>(<b>A</b>) Western blot analysis with the nuclear protein revealed significantly increased translocation of cleaved PKC-δ (41 kD) to nucleus in group H, H<i>^X</i> and E<i>^R</i> compared to either C or E. Significantly increased phospho-p53 (at Ser 46 and Ser 15) and total p53 was observed in group H and E<i>^R</i> compared to E or C whereas significantly reduced phospho-p53 (at Ser 46 and Ser 15) and total p53 was observed in groups E and H<i>^X</i> compared to H or E. RPL32 and Lamin B were used as loading control for cytosolic proteins and nuclear proteins respectively. (<b>B</b>) Subcellular fractionation followed by western blot analyses with mitochondrial protein showing significantly increased translocation of cleaved PKC-δ (41 kD) to mitochondria in group H, H<i>^X</i> and E<i>^R</i> compared to either C or E along with increased expression of PLS3, t-Bid, Bax, cytochrome-c proteins. RPL32 and COX IV were used as loading control for cytosolic proteins and mitochondrial proteins respectively. (<b>C</b>) Western blot analysis showingcleavage of caspase-3 and PARP in group H and E<i>^R</i> compared to C or E. RPL32 was used as loading control. Caspase-3 activity assay showing similar changes in all the experimental groups. No significant difference in caspase-3 activity was detected between groups E and C (*p<0.05 for H versus C; ###p<0.001 for H versus H<i>^X</i>; ¶¶p<0.01 for E versus E<i>^R</i>). (<b>D</b>) Subcellular fractionation followed by western blot analyses showed significantly increased translocation of cleaved PKC-δ (41 kD) to mitochondria along with caspase-3 cleavage in adult cardiomyocytes isolated from group H and E<i>^R</i> compared to either C or E or H<i>^X</i>. RPL32 was used as loading control. (<b>E</b>) Western blot analysis showing phosphorylation status of STAT3 and P38 MAPK in groups C, E, E<i>^R</i> and H.</p