Doxorubicin induced sustained transcriptional reprogramming and mesenchymal changes in cardiac microvascular endothelial cells, with increased transgelin expression observed specifically in female mice.
Does doxorubicin induce sustained mesenchymal reprogramming in cardiac microvascular endothelium?
Doxorubicin induces sustained mesenchymal reprogramming in cardiac endothelial cells, providing mechanistic insight into anthracycline-induced cardiotoxicity.
Chemotherapeutic anthracycline drug, doxorubicin (Dox), is known to cause cardiomyopathy in treated cancer patients. Dox is clinically administered intravenously delivering high concentrations of the drug in cardiac endothelial cells which may contribute to enhanced endothelial injury by Dox. Although immediate exposure and accumulation of Dox occur during Dox treatment, delayed changes in expression and morphology develop after completion of treatment and systemic elimination of Dox. These delayed changes are indicative of cardiovascular endothelial sustained cell reprogramming by Dox. In this study, we examined the immediate and persisting effects of Dox treatment on the heart, focusing on endothelial cell involvement. We hypothesized that Dox targets cardiac endothelial cells, inducing lasting changes in transcription processes and protein expression leading to mesenchymal reprogramming in cardiac microvascular endothelium. We first measured Dox accumulation in vitro and in vivo over time comparing accumulation in endothelial versus non-endothelial cell nuclei. Endothelial cell nuclei developed greater and longer lasting accumulation of Dox implying a major role of this cell type in Dox induced damage. We have previously shown that Dox treated cells have increased TGF-β2 production and enhanced Smad3 phosphorylation. We used ChIP-seq to assess transcriptional reprogramming of Dox-treated human umbilical vein endothelial cells (HUVEC) in the presence of TGF-β2 after Dox washout compared to the control. Smad3 binding sites were identified by Partek Flow v 12.8.3 (Illumina) and enriched pathways were identified by Ingenuity Pathway Analysis (Qiagen) and Gene Sets Enrichment Analysis ( https://www.gsea-msigdb.org/gsea/index.jsp ). Mesenchymal and fibrotic pathways were significantly activated in the Dox and TGF-β2 treated cells showing persisting changes in transcription factor binding post Dox treatment. SB431542 (SB) is a specific ALK4/5/7 inhibitor that we used to block Smad2/3 phosphorylation and further study the TGF-β pathway involvement in Dox injury. HUVEC were treated with SB during Dox-treatment or during washout only with and without TGF-β2 and compared to control HUVEC. SB blocked Smad2/3 phosphorylation during Dox washout in response to TGF-β2. We evaluated mesenchymal changes in vivo by measuring expression of fibronectin and transgelin, key mediators in endothelial to mesenchymal transition, using immunohistochemistry in hearts of Dox treated mice. Dox treatment increased transgelin expression in cardiac tissues of Dox treated female mice while male mice showed no change in expression, indicating possible sex dimorphism in response to Dox. This study provides insight into the immediate effects and longer-term mesenchymal reprogramming affecting endothelial cells in Dox induced cardiovascular injury and provides a better understanding of the mechanisms underlying cardiotoxicity of Dox. These studies were supported by the National Heart, Lung, and Blood Institute of the National Institutes of Health under Award Numbers R15HL133873 and R15HL170243. This abstract was presented at the American Physiology Summit 2026 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.
Cobb et al. (2026) studied Doxorubicin-induced cardiomyopathy. Doxorubicin vs. Control was evaluated on Transcriptional reprogramming and mesenchymal changes in cardiac microvascular endothelium. Doxorubicin induced sustained transcriptional reprogramming and mesenchymal changes in cardiac microvascular endothelial cells, with increased transgelin expression observed specifically in female mice.