Doxorubicin Induces Endotheliotoxicity and Mitochondrial Dysfunction via ROS/eNOS/NO Pathway.

Shuping Chen,Huan He,Dong Yin,Ming He,Yang Qiao,Qing Huang,Hongwei Li,Liang Wang,Qing Zhou

doi:10.3389/fphar.2019.01531

Abstract

Background: Doxorubicin (Dox) can induce endotheliotoxicity and damage the vascular endothelium (VE). The most principle mechanism might be excess reactive oxygen species (ROS) generation. Nevertheless, the characteristics of ROS generation, downstream mechanisms, and target organelles in Dox-induced endotheliotoxicity have yet to be elucidated. Methods and Results: In order to explore the related problems, the VE injury models were established in mice and human umbilical vein endothelial cells (HUVECs) by Dox-induced endotheliotoxicity. Results showed that the activities of lactate dehydrogenase (LDH) and creatine kinase of mice’s serum increased after injected Dox. The thoracic aortic strips’ endothelium-dependent dilation was significantly impaired, seen noticeable inflammatory changes, and brown TUNEL-positive staining in microscopy. After Dox-treated, HUVECs viability lowered, LDH and caspase-3 activities, and apoptotic cells increased. Both intracellular/mitochondrial ROS generation significantly increased, and intracellular ROS generation lagged behind mitochondria. HUVECs treated with Dox plus ciclosporin A (CsA) could basically terminate ROS burst, but plus edaravone (Eda) could only delay or inhibit, but could not completely cancel ROS burst. Meanwhile, the expression of endothelial nitric oxide synthase (eNOS) decreased, especially phosphorylation of eNOS significantly. Then nitric oxide content decreased, the mitochondrial function was impaired, mitochondrial membrane potential (MMP) impeded, mitochondrial swelled, mitochondrial permeability transition pore (mPTP) was opened, and cytochrome C was released from mitochondria into the cytosol. Conclusion: Dox produces excess ROS in the mitochondria, thereby weakens the MMP, opens mPTP, activates the ROS-induced ROS release mechanism, induces ROS burst, and leads to mitochondrial dysfunction, which in turn damages VE. Therefore, interrupting any step of the cycles, as mentioned above can end the related vicious cycle and prevent the occurrence and development of injury.

Highlights

Since its advent in the 1960s, Doxorubicin (Dox) has been widely used in the treatment of various malignant tumors because of its broad spectrum and high efficiency (Bonadonna et al, 1969)
The activities of serum Lactate Dehydrogenase (LDH) and Creatine Kinase (CK) in the Eda+Dox group and the pAD/endothelial nitric oxide synthase (eNOS) +Dox group were significantly improved (P < 0.01), suggesting that the treatment of Eda and pAD/eNOS could alleviate the abovementioned tissue and/or organ damage caused by Dox
Our studies show that excessive reactive oxygen species (ROS) generation by Dox toxicity trigger myocardial injury by mitochondria mediated (He et al, 2018; Chen et al, 2019), whether Human umbilical vein endothelial cells (HUVECs) toxicity by Dox will cause excessive ROS generation and cause damage? First, we found that HUVECs were treated by 1 mM Dox after added 100 mM Eda (Masuda et al, 2016), a free radical scavenger, and 1 mM ciclosporin A (CsA) (Teixeira et al, 2013), an mitochondrial permeability transition pore (mPTP) closing agent, coincubation, cell viability increased and LDH activity decreased (P < 0.01, Figures S2A, B of Supplementary Materials)

Summary

Introduction

Since its advent in the 1960s, Doxorubicin (Dox) has been widely used in the treatment of various malignant tumors because of its broad spectrum and high efficiency (Bonadonna et al, 1969). Many studies have found that there are various reasons for Dox’s cardiotoxicity or endotheliotoxicity (Wojcik et al, 2015; Cappetta et al, 2018; Renu et al, 2018), a common factor is that Dox itself may induce oxidative stress, resulting in excessive reactive oxygen species (ROS) generation (Wolf and Baynes, 2006; Octavia et al, 2012; Angsutararux et al, 2015; Cappetta et al, 2017). Edaravone (Eda), a free radical scavenger, is the only neuroprotective agent for acute ischemic stroke used in Japan (Matsumoto et al, 2018) It captures and reduces excessive ROS, preventing brain damage. The characteristics of ROS generation, downstream mechanisms, and target organelles in Dox-induced endotheliotoxicity have yet to be elucidated

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