Doxorubicin‑induced oxidative and nitrosative stress: Mitochondrial connexin 43 is at the crossroads

Abstract

Oxidative stress is widely accepted as a key factor of doxorubicin (doxo)-induced cardiotoxicity. There is evidence to indicate that nitrosative stress is involved in this process, and that doxo interacts by amplifying cell damage. Mitochondrial connexin 43 (mitocx43) can confer cardioprotective effects through the reduction of mitochondrial reactive oxygen species production during doxo-induced cardiotoxicity. The present study aimed to evaluate the involvement of mitocx43 in doxo-induced nitrosative stress. Rat H9c2 cardiomyoblasts were treated with doxo in the absence or presence of radicicol, an inhibitor of Hsp90, the molecular chaperone involved in cx43 translocation to the mitochondria that underlies its role in cardioprotection. FAcS analysis and RT-qPcR revealed that doxo increased superoxide dismutase, and catalase gene and protein expression. As shown by hypodiploid nuclei and confirmed by western blot analysis, Doxo increased caspase 9 expression and reduced procaspase 3 levels, which induced cell death. Moreover, a significant increase in the activation of the NF-κB signaling pathway was observed. It is well known that the increased expression of inducible nitric oxide synthase results in nitric oxide overproduction, which then rapidly reacts with hydrogen peroxide or superoxide generated by the mitochondria, to form highly reactive and harmful peroxynitrite, which ultimately induces nitrotyrosine formation. Herein, these interactions were confirmed and increased effects were observed in the presence of radicicol. On the whole, the data of the present study indicate that an interplay between oxidative and nitrosative stress is involved in doxo-induced cardiotoxicity, and that both aspects are responsible for the induction of apoptosis. Furthermore, it is demonstrated that the mechanisms that further increase mitochondrial superoxide generation (e.g., the inhibition of cx43 translocation into the mitochondria) significantly accelerate the occurrence of cell death.

Connexins in the Heart: Regulation, Function and Involvement in Cardiac Disease

“…In this regard, it has been shown that pharmacological inhibition of Hsp90 with radicicol, a treatment reducing mitochondrial Cx43 expression in H9c2 cardiomyoblasts, increased cardiotoxicity to doxorubicin, as determined by an enhancement in doxorubicin-induced ROS production, in mitochondrial calcium overload and in cytochrome C release [ 535 ]. Additionally, an increase in nitrosative stress was also observed after radicicol treatment [ 558 ]. Similar findings have been obtained with trastuzumab, the gold standard in the treatment of HER2+ breast cancer [ 559 ].…”

Diazoxide Needs Mitochondrial Connexin43 to Exert Its Cytoprotective Effect in a Cellular Model of CoCl2-Induced Hypoxia

“…Transmembrane Cx43 contributes to cell–cell communication and electrical coupling by the formation of gap junction channels, while on the mitochondrial membrane Cx43 exists as a hemichannel and is involved in mitochondrial volume regulation and respiration [ 6 ]. During stress conditions, Cx43 translocates to the mitochondria with a mechanism that involves the Hsp90/Tom20 machinery system [ 4 , 7 ], and many studies conducted using the Hsp90 inhibitor radicicol supported this theory [ 8 , 9 , 10 ]. Mitochondrial Cx43 (mCx43) acts as an important regulator of apoptosis, allowing the passage of molecules that induce apoptosis such as Ca 2+ , IP 3 and cAMP ions [ 11 ] and influencing mitochondrial respiration, matrix ion fluxes and ROS production [ 9 ].…”

“…During stress conditions, Cx43 translocates to the mitochondria with a mechanism that involves the Hsp90/Tom20 machinery system [ 4 , 7 ], and many studies conducted using the Hsp90 inhibitor radicicol supported this theory [ 8 , 9 , 10 ]. Mitochondrial Cx43 (mCx43) acts as an important regulator of apoptosis, allowing the passage of molecules that induce apoptosis such as Ca 2+ , IP 3 and cAMP ions [ 11 ] and influencing mitochondrial respiration, matrix ion fluxes and ROS production [ 9 ]. Indeed, mitochondria are at the same time the most powerful intracellular source and the primary target for damaging effects of ROS.…”

“…Furthermore, mitochondrial ROS overproduction facilitates the calcium dependent mitochondrial permeability transition, which plays a key role in caspase-mediated apoptosis [ 12 ]. The inhibition of Cx43 translocation on mitochondria induces a drastic increase in ROS production, interferes with calcium homeostasis and accelerates mitochondrial membrane depolarization, thus inducing apoptosis [ 7 , 8 , 9 , 13 ].…”

Chek2 Signaling Is the Key Regulator of Oocyte Survival After Chemotherapy

“…In contrast, we did not detect a significant increase in DSB markers in primordial oocytes after DOX or ETO treatment, suggesting a different mode of toxicity for these agents. It is possible that DOX induces late DNA damage in oocytes indirectly by increasing oxidative stress and reactive oxygen species (ROS) (80)(81)(82)(83). Indeed, co-administration of drugs reducing oxidative stress have been shown to decrease its ovotoxicity (84-86) and γH2AX-positive oocytes were reported after in vivo treatment with DOX (57,87).…”