Mitochondrial Mediated Thimerosal-Induced Apoptosis in a Human Neuroblastoma Cell Line (SK-N-SH)
Department of Pharmacology, Joan C. Edwards School of Medicine, Marshall University, Huntington, WV 25704-9388, USA. NeuroToxicology
(Impact Factor: 3.38).
07/2005; 26(3):407-16. DOI: 10.1016/j.neuro.2005.03.008
Environmental exposure to mercurials continues to be a public health issue due to their deleterious effects on immune, renal and neurological function. Recently the safety of thimerosal, an ethyl mercury-containing preservative used in vaccines, has been questioned due to exposure of infants during immunization. Mercurials have been reported to cause apoptosis in cultured neurons; however, the signaling pathways resulting in cell death have not been well characterized. Therefore, the objective of this study was to identify the mode of cell death in an in vitro model of thimerosal-induced neurotoxicity, and more specifically, to elucidate signaling pathways which might serve as pharmacological targets. Within 2 h of thimerosal exposure (5 microM) to the human neuroblastoma cell line, SK-N-SH, morphological changes, including membrane alterations and cell shrinkage, were observed. Cell viability, assessed by measurement of lactate dehydrogenase (LDH) activity in the medium, as well as the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) assay, showed a time- and concentration-dependent decrease in cell survival upon thimerosal exposure. In cells treated for 24 h with thimerosal, fluorescence microscopy indicated cells undergoing both apoptosis and oncosis/necrosis. To identify the apoptotic pathway associated with thimerosal-mediated cell death, we first evaluated the mitochondrial cascade, as both inorganic and organic mercurials have been reported to accumulate in the organelle. Cytochrome c was shown to leak from the mitochondria, followed by caspase 9 cleavage within 8 h of treatment. In addition, poly(ADP-ribose) polymerase (PARP) was cleaved to form a 85 kDa fragment following maximal caspase 3 activation at 24 h. Taken together these findings suggest deleterious effects on the cytoarchitecture by thimerosal and initiation of mitochondrial-mediated apoptosis.
Available from: Maria Fernanda Hornos Carneiro
- "This indicates doserelated mitochondrial potential damage and concurrent apoptosis , as was seen in the previous section. The property of etHg contained in thimerosal disturbing mitochondrial function has been examined mostly in neuronal and immune cell lines (Makani et al., 2002; Humphrey et al., 2005; Yel et al., 2005; Migdal et al., 2010; Zieminska et al., 2010). We believe ours is the first report on the effects of thimerosal on mitochondrial permeability transition in the kidney. "
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ABSTRACT: Thimerosal is an ethyl mercury-containing compound used mainly in vaccines as a bactericide. Although the kidney is a key target for mercury toxicity, thimerosal nephrotoxicity has not received the same attention as other mercury species. The aim of this study was to determine the potential cytotoxic mechanisms of thimerosal on human kidney cells. Human kidney proximal tubular epithelial (HK2) cells were exposed for 24 h to thimerosal (0-2 µM), and assessed for cell viability, apoptosis, and cell proliferation; expression of proteins Bax, nuclear factor-κB subunits, and transforming growth factor-β1 (TGFβ1); mitochondrial health (JC-1, MitoTracker Red CMXRos); and fibronectin levels (enzyme-linked immunosorbent assay). Thimerosal diminished HK2 cell viability and mitosis, promoted apoptosis, impaired the mitochondrial permeability transition, enhanced Bax and TGFβ1 expression, and augmented fibronectin secretion. This is the first report about kidney cell death and pro-fibrotic mechanisms promoted by thimerosal. Collectively, these in vitro results demonstrate that (1) thimerosal induces kidney epithelial cell apoptosis via upregulating Bax and the mitochondrial apoptotic pathway, and (2) thimerosal is a potential pro-fibrotic agent in human kidney cells. We suggest that new evidence on toxicity as well as continuous surveillance in terms of fibrogenesis is required concerning thimerosal use. © 2014 Wiley Periodicals, Inc. Environ Toxicol, 2014.
Available from: Ramsés García
- "The mitochondrion is a primary target of mercury-induced injury and the mitochondrial electron transport chain is the most likely site where excess ROS are generated to induce oxidative stress in mercury toxicity (Yee and Choi, 1996). Mitochondrial effects of mercury in vivo and in vitro, include mitochondrial dysfunction (Chávez and Holguín, 1988; Chávez et al., 1989; Hare and Atchison, 1992; Dreiem et al., 2005; Hernández-Esquivel et al., 2011), membrane permeability transition pore opening (Limke and Atchison, 2002; Polunas et al., 2011) and apoptosis (Shenker et al., 1999; Kim and Sharma, 2004; Humphrey et al., 2005). "
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ABSTRACT: Occupational or environmental exposures to heavy metals produce several adverse health effects. The common mechanism determining their toxicity and carcinogenicity is the generation of oxidative stress that leads to hepatic damage. In addition, oxidative stress induced by metal exposure leads to the activation of the nuclear factor (erythroid-derived 2)-like 2/Kelch-like ECH-associated protein 1/antioxidant response elements (Nrf2/Keap1/ARE) pathway. Since antioxidant and chelating agents are generally used for the treatment of heavy metals poisoning, this review is focused on the protective role of curcumin against heavy metals liver injury. Curcumin has shown, in clinical and preclinical studies, numerous biological activities including therapeutic efficacy against various human diseases and anti-hepatotoxic effects against environmental or occupational toxins. Curcumin reduces the hepatotoxicity induced by arsenic, cadmium, chromium, copper, lead and mercury, prevents histological injury, lipid peroxidation and glutathione (GSH) depletion, maintains the liver antioxidant enzyme status and protects against mitochondrial dysfunction. The preventive effect of curcumin on the noxious effects induced by heavy metals has been attributed to its scavenging and chelating properties, and/or to the ability to induce the Nrf2/Keap1/ARE pathway. However, additional research is needed in order to propose curcumin as a potential protective agent against liver damage induced by heavy metals.
Available from: Vered Abitbol
- "Cell viability, assessed by the MTT assay, as well as the 7-AAD flow cytometry assay, showed a time- and concentration-dependent decrease in cell survival upon thimerosal exposure. Mitochondria play a crucial role in regulating apoptosis 
, and it was found involved in thimerosal-induced cell death of several tumor cell types 
. To identify the apoptotic pathway associated with thimerosal-mediated cell death of primary lymphocytes, we used the probe DiOC6(3), which allows to study variations in the mitochondrial transmembrane potential ΔYm during intrinsic cell death . "
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ABSTRACT: Thimerosal is a preservative used widely in vaccine formulations to prevent bacterial and fungal contamination in multidose vials of vaccine. Thimerosal was included in the multidose non-adjuvanted pandemic 2009 H1N1 vaccine Panenza. In the context of the analysis of the ex-vivo T cell responses directed against influenza vaccine, we discovered the in vitro toxicity Panenza, due to its content in thimerosal. Because thimerosal may skew the immune response to vaccines, we investigated in detail the ex-vivo effects of thimerosal on the fate and functions of T cells in response to TCR ligation. We report that ex-vivo exposure of quiescent or TCR-activated primary human T cells to thimerosal induced a dose-dependent apoptotic cell death associated with depolarization of mitochondrial membrane, generation of reactive oxygen species, cytochrome c release from the mitochondria and caspase-3 activation. Moreover, exposure to non-toxic concentrations of thimerosal induced cell cycle arrest in G0/G1 phase of TCR-activated T cells, and inhibition of the release of proinflammatory cytokines such as IFN gamma, IL-1 beta, TNF alpha, IL-2, as well as the chemokine MCP1. No shift towards Th2 or Th17 cells was detected. Overall these results underline the proapoptotic effect of thimerosal on primary human lymphocytes at concentrations 100 times less to those contained in the multidose vaccine, and they reveal the inhibitory effect of this preservative on T-cell proliferation and functions at nanomolar concentrations.
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