siRNA-Like Double-Stranded RNAs Are Specifically Protected Against Degradation in Human Cell Extract

RNA interference (RNAi) is a set of intracellular pathways in eukaryotes that controls both exogenous and endogenous gene expression. The power of RNAi to knock down (silence) any gene of interest by the introduction of synthetic small-interfering (si)RNAs has afforded powerful insight into biological function through reverse genetic approaches and has borne a new field of gene therapeutics. A number of questions are outstanding concerning the potency of siRNAs, necessitating an understanding of how short double-stranded RNAs are processed by the cell. Recent work suggests unmodified siRNAs are protected in the intracellular environment, although the mechanism of protection still remains unclear. We have developed a set of doubly-fluorophore labeled RNAs (more precisely, RNA/DNA chimeras) to probe in real-time the stability of siRNAs and related molecules by fluorescence resonance energy transfer (FRET). We find that these RNA probes are substrates for relevant cellular degradative processes, including the RNase H1 mediated degradation of an DNA/RNA hybrid and Dicer-mediated cleavage of a 24-nucleotide (per strand) double-stranded RNA. In addition, we find that 21- and 24-nucleotide double-stranded RNAs are relatively protected in human cytosolic cell extract, but less so in blood serum, whereas an 18-nucleotide double-stranded RNA is less protected in both fluids. These results suggest that RNAi effector RNAs are specifically protected in the cellular environment and may provide an explanation for recent results showing that unmodified siRNAs in cells persist intact for extended periods of time

Para-tertiary butyl catechol induces eryptosis in vitro via oxidative stress and hemoglobin leakage in human erythrocytes

Exposure of human population to industrial chemicals is believed as a significant contributing factor to the outgrowth of occupational diseases especially in developing countries due to improper safety measures and sanitary conditions. Para-tertiary butylcatechol (PTBC) widely employed in petrochemical, thermofax and phototypesetting industries, induces melanocytotoxicity and contact dermatitis leading to occupational leukoderma/vitiligo. Few vitiligo patients were reported for oxidative stress-induced hemolytic anemia and thrombocytopenia, however its impact on blood components is still not clear. Erythrocytes are the major cell population in circulation and play a prominent role in various diseases. In this work, the effect of PTBC on human erythrocytes is evaluated in vitro. PTBC induces oxidative stress-mediated eryptosis (erythrocyte death) causing detrimental changes such as depleted antioxidant levels, altered surface morphology, hemoglobin denaturation and heinz body formation. These findings validate that PTBC could induce toxic effects on human erythrocytes. Exposure of humans to toxic chemicals constitutes an important issue in various industries; one such issue is the exposure of PTBC at work place resulting in a spectrum of dermal complications. Therefore, it is imperative to appraise the long-term toxicities in order to further delineate the mechanisms of resultant disorders associated with PTBC and to establish the therapeutic interventions

Para-tertiary butyl catechol induces eryptosis in vitro via oxidative stress and hemoglobin leakage in human erythrocytes

Exposure of human population to industrial chemicals is believed as a significant contributing factor to the outgrowth of occupational diseases especially in developing countries due to improper safety measures and sanitary conditions. Para-tertiary butylcatechol (PTBC) widely employed in petrochemical, thermofax and phototypesetting industries, induces melanocytotoxicity and contact dermatitis leading to occupational leukoderma/vitiligo. Few vitiligo patients were reported for oxidative stress-induced hemolytic anemia and thrombocytopenia, however its impact on blood components is still not clear. Erythrocytes are the major cell population in circulation and play a prominent role in various diseases. In this work, the effect of PTBC on human erythrocytes is evaluated in vitro. PTBC induces oxidative stress-mediated eryptosis (erythrocyte death) causing detrimental changes such as depleted antioxidant levels, altered surface morphology, hemoglobin denaturation and heinz body formation. These findings validate that PTBC could induce toxic effects on human erythrocytes. Exposure of humans to toxic chemicals constitutes an important issue in various industries; one such issue is the exposure of PTBC at work place resulting in a spectrum of dermal complications. Therefore, it is imperative to appraise the long-term toxicities in order to further delineate the mechanisms of resultant disorders associated with PTBC and to establish the therapeutic interventions

5-Methyl-3-phenylisoxazole-4-carboxylic acid

In the title compound, C11H9NO3, the phenyl and isoxazole rings form a dihedral angle of 56.64 (8)°. The carboxy group is almost in the same plane as the isoxazole ring with a C—C—C—O torsion angle of −3.3 (2)°. In the crystal, pairs of O—H...O hydrogen bonds link the molecules into head-to-head dimers. C—H...N hydrogen bonds and π–π stacking interactions between phenyl rings [centroid–centroid distance = 3.9614 (17)Å] link the dimers into a three-dimensional network

Lupeol derivative mitigates echis carinatus venom-induced tissue destruction by neutralizing venom toxins and protecting collagen and angiogenic receptors on inflammatory cells

Background: Echis carinatus bite is a serious threat in South-Asian countries including India, as it causes highest number of deaths and terrifying long-term tissue destruction at the bitten site. Although venom metalloproteinases and hyaluronidases are the suggested key players, studies on the effect of venom on polymorphonuclear cells, peripheral blood mononuclear cells and platelets, and their role in long-term tissue destruction are still in infancy. While, the effect of venom on collagen receptors, integrin alpha 2 beta 1/GP VI/DDR1 and CX3CR1 chemokine receptor present on these cells is an untouched area. Methods: Lupeol, lupeol acetate, its synthetic derivatives 2-8 were screened for inhibition of E. carinatus venom induced-hemorrhage in mouse model where compound 8 was found to be the most potent. Further, compounds efficiently neutralized venom induced hemorrhage, edema, dermonecrosis, myonecrosis, myotoxicity, procoagulant, oxidative stress, inflammatory cytokines and cleavage of collagen and CX3CR1 receptors on inflammatory cells in in vivo, in silico, ex vivo and in vitro studies. Conclusions: This study for the first time demonstrated the cleavage of collagen receptors and the receptor for angiogenesis and wound healing by the venom and its inhibition by compound 8, as these are important for firm adhesion of inflammatory cells at the damaged site to resolve inflammation and promote tissue repair. General significance: This study provides a lead in venom pharmacology, wherein, compound 8 could be a therapeutic agent for the better management of viper venom-induced long-term tissue destruction. (C) 2015 Elsevier B.V. All rights reserved

Protective Effect of Tamarind Seed Coat Ethanol Extract on Eryptosis Induced by Oxidative Stress

Suicidal erythrocyte death, or eryptosis, is the key event in eliciting anemia in numerous pathological conditions, including diabetes, chronic kidney disease, cancer, sepsis, etc. Oxidative stress is an important trigger in the acceleration of erythrocyte loss via eryptosis and an underlying mechanism of anemia emergence in the above pathologies. Therefore, there is an increasing demand for identification of antioxidants and anti-eryptotic agents forthe management of stress-related ailments. Here, we demonstrated the antioxidant and anti-eryptotic properties of the tamarind seed coat ethanol extract (TSCEE) against 2,2'-azobis(2-amidinopropane) dihydrochloride (AAPH)-induced oxidative stress and eryptosis. The presence of probable secondary metabolites in the TSCEE extract was investigated by RP-HPLC. Active groups present in the TSCEE were studied by the Fourier-transform infrared spectroscopy. Cyclic voltammetric studies confirmed the antioxidant potential of TSCEE. The protective effect of TSCEE on red blood cells was confirmed by assessing various eryptotic markers, such as reactive oxygen species generation, intracellular calcium levels, and phosphatidylserine exposure. TSCEE reduced lipid peroxidation and protein carbonyl content and restored the levels of glutathione, antioxidant enzymes, and enzymes involved in glutathione replenishment. In conclusion, TSCEE was found to exhibit multiple therapeutic properties, which makes it a. promising agent for treating oxidative stress-induced eryptosis and subsequent anemia in various pathologies