Vanadate induces necrotic death in neonatal rat cardiomyocytes through mitochondrial membrane depolarization
Sandra Sofia Soares1, Fernando Henao, Manuel Aureliano
1Comparative Cardiovascular Physiopathology Group (GFCC), Faculty of Environmental and Marine Sciences, Centre of Marine Sciences (CCMAR), University of Algarve, Campus de Gambelas, 8005-139 Faro, Portugal. sssoares@ualg.pt
Abstract:
Besides the well-known inotropic effects of vanadium in cardiac muscle, previous studies have shown that vanadate can stimulate cell growth or induce cell death. In this work, we studied the toxicity to neonatal rat ventricular myocytes (cardiomyocytes) of two vanadate solutions containing different oligovanadates distribution, decavanadate (containing decameric vanadate, V 10) and metavanadate (containing monomeric vanadate and also di-, tetra-, and pentavanadate). Incubation for 24 h with decavanadate or metavanadate induced necrotic cell death of cardiomyocytes, without significant caspase-3 activation. Only 10 microM total vanadium of either decavanadate (1 microM V 10) or metavanadate (10 microM total vanadium) was needed to produce 50% loss of cell viability after 24 h (assessed with MTT and propidium iodide assays). Atomic absorption spectroscopy showed that vanadium accumulation in cardiomyocytes after 24 h was the same when incubation was done with decavanadate or metavanadate. A decrease of 75% of the rate of mitochondrial superoxide anion generation, monitored with dihydroethidium, and a sustained rise of cytosolic calcium (monitored with Fura-2-loaded cardiomyocytes) was observed after 24 h of incubation of cardiomyocytes with decavanadate or metavanadate concentrations close to those inducing 50% loss of cell viability produced. In addition, mitochondrial membrane depolarization within cardiomyocytes, monitored with tetramethylrhodamine ethyl esther or with 3,3',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolcarbocyanine iodide, were observed after only 6 h of incubation with decavanadate or metavanadate. The concentration needed for 50% mitochondrial depolarization was 6.5 +/- 1 microM total vanadium for both decavanadate (0.65 microM V 10) and metavanadate. In conclusion, mitochondrial membrane depolarization was an early event in decavanadate- and monovanadate-induced necrotic cell death of cardiomyocytes.
Insights
Vanadate compounds, decavanadate and metavanadate, induce necrotic cell death in cardiomyocytes by causing early mitochondrial membrane depolarization. This occurs without significant caspase-3 activation, highlighting a novel toxicity pathway for vanadium in cardiac cells.
Area of Science:
- Cardiovascular Biology
- Toxicology
- Cellular Physiology
Background:
- Vanadium compounds, particularly vanadate, are known to affect cardiac muscle function.
- Previous research indicates vanadate can either promote cell growth or induce cell death.
- The specific mechanisms and forms of vanadate toxicity in cardiomyocytes require further elucidation.
Purpose of the Study:
- To investigate the toxicity of two distinct vanadate solutions, decavanadate and metavanadate, on neonatal rat ventricular myocytes.
- To determine the specific cellular events leading to vanadate-induced cell death in cardiomyocytes.
- To compare the toxicological effects of different vanadate oligomers on cardiac cells.
Main Methods:
- Neonatal rat ventricular myocytes were incubated with decavanadate and metavanadate solutions.
- Cell viability was assessed using MTT and propidium iodide assays.
- Vanadium accumulation was measured by atomic absorption spectroscopy.
- Mitochondrial superoxide anion generation, cytosolic calcium levels, and mitochondrial membrane potential were monitored using specific fluorescent probes (dihydroethidium, Fura-2, TMRM, JC-1).
Main Results:
- Both decavanadate and metavanadate induced necrotic cell death in cardiomyocytes after 24 hours, with 50% cell death occurring at approximately 10 microM total vanadium.
- Caspase-3 activation was not significantly observed, suggesting a non-apoptotic cell death pathway.
- Vanadium accumulation was similar for both vanadate forms.
- Early mitochondrial dysfunction, including depolarization and decreased superoxide generation, occurred within 6-24 hours, preceding significant cell death.
- Cytosolic calcium levels increased concurrently with mitochondrial dysfunction.
Conclusions:
- Mitochondrial membrane depolarization is an early and critical event in vanadate-induced necrotic cell death of cardiomyocytes.
- The toxicity of decavanadate and metavanadate is comparable at equivalent total vanadium concentrations.
- Vanadate toxicity in cardiomyocytes involves disruption of mitochondrial function and calcium homeostasis, leading to necrosis rather than apoptosis.
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