Cadmium-induced Mitochondrial Damage And Intervention

Cadmium (Cd) is an industrial and environmental pollutant. Once Cd was uptake by human, it could accumulate in the body and affect multiple systems and organs in human. Both liver and kidney are the main target organs for cadmium accumulation and cadmium toxicity. Mitochondria is an organelle which is very sensitive to stimulus and xenobiotics. Since mitochondria plays important role in oxidative phosphorylation and mediating cell apoptosis, it becomes a life-and-death switch in cells.In this study, effects of Cd to liver and kidney mitochondria and relative mechanism were studied through in vivo and in vitro experiments. Ultrastructura of mitochondria was observed by using TEM. Evaluation of mitochondrial function includes mitochondrial respiratory test (with Clark's electrode); mitochondrial swelling test (spectrophotometry); mitochondrial membrane potential (ΔΨm) (with fluorescent probe of rhodamine 123); mitochondrial viability test (MTT test); mitochondrial superoxides level (NBT test). Drugs such as ruthenium red (RR), the blocker of mitochondrial calcium uniporter (MCU); cycroporin A (CsA), specific inhibitor of mitochondrial permeable transition pore (MPTP); dithiothreitol (DTT), a reducing agent and SH-groups protector; EGTA, a chelator of calcium, were used to study the mechanisms and intervention of Cd damage to mitochondria.In 5 days subcutaneous Cd administration study, morphological changes of rat kidney mitochondria in 1.2 mg Cd/kg bw and 1.8 mg Cd/kg bw groups were found, including breakdown of cristae, matrix vacuolation, and mitochondria swelling of mitochondria through TEM. Matrix darkening, proximal tubular cell vacuolation, and intracellular lysosome increase in 1.8 mg Cd/kd bw group indicated Cd damage in subcellular level. Repsiration control ratio (RCR) of rat kidney mitochondrial respiratory function decreased with the increase of administration dose of Cd, and significiently lower in 1.8 mg Cd/kg bw group compared with the control group. With increase of Cd administration dose, swelling effect of kidney mitochondria were aggravated,ΔΨm level were decreased, and mitochondrial superoxide level were significantly increased in 1.2 mg Cd/kg bw and 1.8 mg Cd/kg bw groups. In 12 weeks subchronic Cd administration study, morphological changes of rat kidney mitochondria were found in 4 weeks administration. Swelling and vacuoation of kidney mitochondria were found in 1.0 mg Cd/kg bw and 2.0 mg Cd/kg bw groups which indicated severe toxicity of Cd. With increase of Cd administration dose and time, RCR of rat kidney mitochondrial respiratory function showed trend of decrease, which suggested the dose-dependent and time-dependent manner of Cd toxicity. Superoxide level of rat kidney mitochondria increased with increase of Cd administration dose in 4 weeks. Afterwards, superoxide level decreased with decrease of mitochondrial viability.In vitro study showed that Cd2+ could inhibit RCR of isolated rat liver and kidney mitochondria in a dose-dependent manner. As to respiratory function, liver mitochondria was more sensitive to Cd toxicity than kidney mitochondria. Cd2+ could dose-dependently induce swelling of isolated rat liver and kidney mitochondria. The EC50 of liver and kidney mitochondria were 8.27μM and 16.27μM, respectively. The maxim swelling effect of liver and kidney mitochondria were 40% and 30% as swelling%. Comparing effects of Cd2+ and Ca2+ to liver mitochondrial swelling, we found that both of Cd2+ and Ca2+ could dose-dependently induce swelling of mitochondria, the EC50 were 8.27μM and 47.43μM; and maxim swelling effect were 40% and 25% as swelling%. Cd2+ could cause dissipation ofΔΨm of liver and kidney mitochondria. The EC50 of liver and kidney mitochondria were 10.55μM and 26.83μM, respectively. Cd2+ could also dose-dependently decrease mitochondrial viability of rat liver and kidney mitochondria, the EC50 were 8.27μM and 7.73μM, respectively.RR (5μM) could obviously inhibit Ca2+ and low dose (10μM) Cd2+ induced swelling of mitochondria and dissipation ofΔΨm With increase of Cd2+ concentration, effect of RR weakened. CsA could effectively inhibit swelling effect induced by Ca2+. However, CsA was only partly effectively to Cd2+ on liver mitochondria and almost void on kidney mitochondria.Since Cd shows high affinity to thiol-dependent domain and similarity with Ca property, DTT and EGTA were used for intervention of Cd damage. Both DTT and EGTA could dose-dependently prevent liver mitochondria from Cd2+ damage, including swelling of mitochondria, dissipation ofΔΨm, and reduction of mitochondrial viability. Protective effect of DTT is better than EGTA to Cd on liver mitochondria.In sum, through in vivo study, kidney mitochondria was confirmed as target of subcellular Cd toxicity. Cd could induce morphological change as well as function damage including respiratory function disturbance and mitochondrial viability damage in rat kidney mitochondria. In vitro study showed that Cd could dose-dependently affect respiratory function, induce swelling, lose ofΔΨm and viability damage of isolated rat liver and kidney mitochondria. Different affect could be found in mitochondria isolated from different organs. Liver mitochondria is more sensitive to Cd toxicity compared with kidney mitochondria. Cd can enter mitochondria through MCU and induce opening of MPTP. RR could inhibit damage of Cd to some degree, and CsA could partly inhibit Cd toxicity. Since Cd could affect protein structure and function through combination with SH group, DTT was used to counteract Cd toxicity. EGTA were also used to counteract Cd toxicity since it is specific Ca chelator.