WDNM1 is Associated with Differentiation and Apoptosis of Mammary Epithelial Cells

In this study, we show that expression of the Westmead DMBA8 nonmetastatic cDNA 1 (WDNM1) gene was increased upon SFM and/or TNFα treatment, with a corresponding increase in apoptotic cells, and gradually decreased following re-stimulation with serum in HC11 mammary epithelial cells. TNFα induced WDNM1 expression showed the NFκB-dependent mechanism since it's expression was abrogated in IκBαM (super-repressor of NFκB)-transfected cells, but not those transfected with control vector. Furthermore, overexpression of WDNM1 suppressed growth and differentiation, and accelerated apoptosis of HC11 cells. Thus, our results demonstrate that WDNM1 gene expression, regulated by the TNFα-NFκB signal pathway, is associated with HC11 cell apoptosis.     

Coenzyme Q 10 Depletion is Comparatively Less Detrimental to Human Cultured Skin Fibroblasts than Respiratory Chain Complex Deficiencies

The oxidative stress possibly resulting from an inherited respiratory chain (RC) deficiency was investigated in a series of human cultured skin fibroblasts presenting either ubiquinone depletion or isolated defect of the various RC complexes. Taken as an index for superoxide overproduction, a significant induction of superoxide dismutase activity was observed in complex V-deficient fibroblasts harboring the NARP-mutation in the ATPase 6 gene. Superoxide dismutase induction was also noticed, albeit to a lesser extent, in complex II-deficient fibroblasts with a mutation in the nuclear gene encoding the flavoprotein subunit of the succinate dehydrogenase. No sign of oxidative stress could be found in ubiquinone-depleted fibroblasts. In all cases but complex IV-defect, increased oxidative stress was associated with increased cell death. In glucose-rich medium, apoptosis appeared as the main cell death process associated with all types of RC defect. However, similar to the great variations in oxidative stress associated with the various types of RC defect, we found that apoptotic features differed noticeably between defects. No indication of increased cell death was found in ubiquinone-depleted fibroblasts.     

The Cytotoxic and Proapoptotic Activities of Hypnophilin are Associated with Calcium Signaling in UACC‐62 Cells

Hypnophilin (HNP) is a sesquiterpene that is isolated from Lentinus cf. strigosus and has cytotoxic activities. Here, we studied the calcium signaling and cytotoxic effects of HNP in UACC‐62 cells, a human skin melanoma cell line. HNP was able to increase the intracellular calcium concentration in UACC‐62 cells, which was blocked in cells stimulated in Ca²⁺‐free media. HNP treatment with BAPTA‐AM, an intracellular Ca²⁺ chelator, caused an increase in calcium signals. HNP showed cytotoxicity against UACC‐62 cells in which it induced DNA fragmentation and morphological alterations, including changes in the nuclear chromatin profile and increased cytoplasmatic vacuolization, but it had no effect on the plasma membrane integrity. These data suggest that cytotoxicity in UACC‐62 cells, after treatment with HNP, is associated with Ca²⁺ influx. Together, these findings suggest that HNP is a relevant tool for the further investigation of new anticancer approaches. © 2013 Wiley Periodicals, Inc. J BiochemMol Toxicol 27:479‐485, 2013; View this article online at wileyonlinelibrary.com . DOI 10.1002/jbt.21507     

Hydrogen sulfide alleviates uranium-induced kidney cell apoptosis mediated by ER stress via 20S proteasome involving in Akt/GSK-3β/Fyn-Nrf2 signaling

Abstract

Hydrogen sulfide (H₂S) shows antioxidative, anti-inflammatory, antiapoptotic, and cytoprotective effects in kidneys. Recently, H₂S has been reported to alleviate uranium-induced rat nephrotoxicity through oxidative stress and inflammatory response via Nrf2-NF-κB pathways. Here, the protective effect and molecular mechanism of H₂S on uranium-induced apoptosis were examined in normal rat kidney proximal cells (NRK-52^E) in vitro. The results indicate that NaHS (an H₂S donor) administration in uranium-intoxicated kidney cells ameliorated uranium-induced reactive oxygen species generation, caspase-3-dependent apoptosis, and endoplasmic reticulum (ER) stress identified through several key markers including GRP78, C/EBP homologous protein (CHOP), and caspase-12. NaHS treatment in uranium-intoxicated kidney cells abolished the effects of uranium on Akt phosphorylation, GSK-3β activation, increased Fyn nuclear expression, and concomitantly decreased Nrf2 nuclear expression. NaHS administration in uranium-treated kidney cells resorted uranium-decreased the expression of two key subunit PSMA6 and PSMB7 in 20S proteasome. But, DRB (an Nrf2 inhibitor) administration abrogated the effects of NaHS on PSMA6 and PSMB7 expression in uranium-contaminated kidney cells. Bortezomib (a proteasome inhibitor) treatment in NaHS pulsing uranium cotreated kidney cells reversed the effects of NaHS on not only PSMA6 and PSMB7 but also GRP78 and CHOP. Taken together, all data suggest that H₂S can attenuate uranium-induced kidney cell apoptosis mediated by ER stress via 20S proteasome involving in Akt/GSK-3β/Fyn-Nrf2 signaling axis.     

Carotenoids Induce Apoptosis in the T-lymphoblast Cell Line Jurkat E6.1

Epidemiologically, a high-carotenoid intake via a fruit- and vegetable-rich diet is associated with a decreased risk of various forms of cancer. The mechanisms by which carotenoids exert this protective effect are controversial. In this study, we examined the potency of a range of carotenoids commonly found in human plasma to induce apoptosis in Jurkat E6.1 malignant T-lymphoblast cells. At a concentration of 20 &#119 M, the order of potency to induce apoptosis after 24 h was: &#103 -carotene > lycopene > lutein> &#103 -cryptoxanthin=zeaxanthin. Canthaxanthin failed to induce apoptosis under these conditions. &#103 -Carotene induced apoptosis in a time- and concentration-dependent manner with a lowest effective concentration of about 3 &#119 M. Pre-conditioning of &#103 -carotene for 72 h destroyed its pro-apoptotic activity almost completely, whereas degradation for 6 h or less did not, indicating that either &#103 -carotene itself and/or an early degradation product of &#103 -carotene are the death-inducing compounds. Apoptosis induced by &#103 -carotene was characterized by chromatin condensation and nuclear fragmentation, DNA degradation, PARP cleavage and caspase-3 activation. The antioxidant BO-653 inhibited the degradation of &#103 -carotene in vitro and significantly increased its cytotoxicity, indicating that a pro-oxidant effect of &#103 -carotene is unlikely to cause its pro-apoptotic activity. The induction of apoptosis in transformed cells by carotenoids may explain their protective effect against cancer formation in humans. Possible pathways for induction of apoptosis by carotenoids are discussed.     

Melatonin Prevents Oxidative Stress and Hepatocyte Cell Death Induced by Experimental Cholestasis

The induction of oxidative stress precedes liver injury during experimental obstructive jaundice (OJ). In this sense, different evidences suggest that melatonin (MEL), as antioxidant, may be useful in the protection against apoptosis and necrosis during experimental cholestasis. In addition, we will also assess if MEL-dependent protection is related to a recovery of antioxidant status disturbances induced by OJ. Cholestasis was achieved by double ligature and sectioning of the principal bile duct. MEL was injected intraperitoneally (500?μg/kg/day). Lipid peroxidation was evaluated by the measurement of malondialdehyde (MDA) content in liver. Different parameters related to antioxidant status, such as reduced glutathione (GSH), glutathione peroxidase (GPx), catalase and superoxide dismutase (SOD) were determined in liver. Liver injury was assessed by alanine aminotransferase (ALT) in serum, histological examination, DNA fragmentation and TUNEL assay. The activation of perisinusoidal stellate cells was evaluated by immunohistochemical measurement of α-smooth muscle actin in liver sections. The induction of OJ increased all the parameters related to apoptosis and necrosis in liver. The induction of liver injury was associated with stellate cell activation, as well as an increase in MDA (p<0.0001) and a reduction in GSH, GPx, catalase and SOD content (p<0.0001) in liver. MEL reduced hepatic apoptosis and necrosis (p<0.004) with a significant improvement in all oxidative stress markers. In conclusion, our results showed that MEL recovered the antioxidant status and reduced apoptosis and necrosis induced by experimental cholestasis.     

Changes of PI3K/AKT/BCL2 signaling proteins in congenital Giant Nevi: melanocytes contribute to their increased survival and integrity

Abstract

Congenital Giant Nevi (CGN) are rare melanocytic lesions with the potential to regress into malignant melanoma. Simultaneous up-regulation and cooperative interactions of signaling pathways are crucial events in the pathogenesis of melanocytes. Our study aimed to identify changes in the expression and activation of proteins controlling survival and/or apoptosis of the key signaling pathways PI3K/AKT/BCL2 and Wnt/β-catenin of CGN melanocytes. We applied a model of cultured melanocytes from paired CGN and normal appearing skin, and Western blot (WB) analyzed the expression and activation profile of survival and anti-apoptotic proteins of these signaling pathways, growth pattern, cell cycle and apoptosis. WB analysis demonstrated a significant higher expression level of activated AKT and of BCL2 proteins in the CGN melanocytes compared with paired melanocytes from normal appearing skin. A relative increase in the level of GSK3 and FOXO1 proteins, down stream targets of AKT, as well as of pβ-catenin was also detected in the CGN melanocytes compared with the controls. These changes were not affected by growth of the CGN melanocytes in reduced serum (starvation). Both cell populations shared a similar growth pattern, with no significant differences in the proportion of apoptotic cells and in cell cycle fractions. These data demonstrate for the first time, changes in signaling proteins of cultured CGN melanocytes. Further, suggesting that the changes in AKT/BCL2 signaling molecules might mediate growth and anti-apoptosis processes at least in part, thus increasing the survival potential of CGN melanocytes and maintaining their integrity.     

The role of insulin against hydrogen peroxide-induced oxidative damages in differentiated SH-SY5Y cells

Abstract

Exogenous hydrogen peroxide (H₂O₂) can easily penetrate into biological membranes and enhance the formation of other reactive oxygen species (ROS). In the present study, we have investigated the neuroprotective effects of insulin on H₂O₂-induced toxicity of retinoic acid (RA)-differentiated SH-SY5Y cells. To measure the changes in the cell viability of SH-SY5Y cells at different concentrations of H₂O₂ for 24?h, a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT)-based assay was used and a 100?µM H₂O₂ was selected to establish a model of H₂O₂-induced oxidative stress. Further assays showed that 24?h of 100?µM H₂O₂-induced significant changes in the levels of lactate dehydrogenase (LDH), nitric oxide (NO), ROS, and calcium ion (Ca²⁺) in neuronal cells, but insulin can effectively diminish the H₂O₂-induced oxidative damages to these cells. Moreover, cells treated with insulin increased H₂O₂-induced suppression of glutathione levels and exerted an apparent suppressive effect on oxidative products. The results of insulin treatment with SH-SY5Y cells increased the Bcl-2 levels and decreased the Akt levels. The treatment of insulin had played a protective effect on H₂O₂-induced oxidative stress related to the Akt/Bcl-2 pathways.