Embryonic stem cell-derived exosomes inhibit doxorubicin-induced TLR4-NLRP3-mediated cell death-pyroptosis

Abstract

Doxorubicin (Dox)-induced cardiac side effects are regulated through increased oxidative stress and apoptosis. However, it remains unknown whether Dox induces the specific inflammatory-mediated form of cell death called pyroptosis. The current study is undertaken to determine whether Dox induces pyroptosis in an in vitro model and to test the potential of exosomes derived from embryonic stem cells (ES-Exos) in inhibiting pyroptosis. H9c2 cells were exposed to Dox to generate pyroptosis and then subsequently treated with exosomes to investigate the protective effects of ES-Exos. Mouse embryonic fibroblast-exosomes (MEF-Exos) were used as a cell line control. We confirmed pyroptosis by analyzing the presence of Toll-like receptor 4 (TLR4)-pyrin domain containing-3 (NLRP3) inflammasome that initiates pyroptosis, which was further confirmed with pyroptotic markers caspase-1, IL-1β, caspase-11, and gasdermin-D. The presence of inflammation was confirmed for proinflammatory cytokines, TNF-α, and IL-6. Our data show that Dox exposure significantly ( P < 0.05) increases expression of TLR4, NLRP3, pyroptotic markers (caspase-1, IL-1β, caspase-11, and gasdermin-D), and proinflammatory cytokines (TNF-α and IL-6) in H9c2 cells. The increased expression of inflammasome, pyroptosis, and inflammation was significantly ( P < 0.05) inhibited by ES-Exos. Interestingly, our cell line control, MEF-Exos, did not show any protective effects. Furthermore, our cytokine array data suggest increased anti-inflammatory (IL-4, IL-9, and IL-13) and decreased proinflammatory cytokines (Fas ligand, IL-12, and TNF-α) in ES-Exos, suggesting that anti-inflammatory cytokines might be mediating the protective effects of ES-Exos. In conclusion, our data show that Dox induces pyroptotic cell death in the H9c2 cell culture model and is attenuated via treatment with ES-Exos. NEW & NOTEWORTHY Doxorubicin (Dox)-induced cardiotoxicity is mediated through increased oxidative stress, apoptosis, and necrosis. We report for the first time as per the best of our knowledge that Dox initiates Toll-like receptor 4 and pyrin domain containing-3 inflammasome formation and induces caspase-1-mediated inflammatory pyroptotic cell death in H9c2 cells. Moreover, we establish that inflammation and pyroptosis is inhibited by embryonic stem cell-derived exosomes that could be used as a future therapeutic option to treat Dox-induced cardiotoxicity.

Exosome Treatment Enhances Anti-Inflammatory M2 Macrophages and Reduces Inflammation-Induced Pyroptosis in Doxorubicin-Induced Cardiomyopathy

“…caesius , a family of anthracyclines. This powerful drug is an antineoplastic therapeutic agent used to treat a wide variety of solid organ tumors and hematologic malignancies, including leukemia, lymphoma, breast cancer, and lung cancer in adults and pediatric patients [1,2,3]. However, medicinal use of this drug is limited due to acute and chronic side effects such as alopecia, vomiting, nausea, cardiac dysfunction, and heart failure [4,5].…”

“…Apoptosis has also been reported in non-cardiac myocytes such as endothelial cells [9,10]. Recent studies suggest that DIC may involve infiltration of M1 macrophages that induces inflammation and pathogenesis [1,11]. However, this remains unknown whether presence of inflammation induces inflammation-mediated cell death—termed pyroptosis in DIC.…”

“…Therefore, recent studies have focused on embryonic stem cell-derived exosomes (ES-Exos) as a safer alternative approach to ESC therapy [23,24,25]. Exosomes (Exos) are cell-derived vesicles, which contain proteins, lipids, growth factors, miRNAs, and anti-inflammatory cytokines that are released through exocytosis [1,26,27]. Recent literature suggests that ES-Exos have the potential to repair infarcted heart [28].…”

PD-1 inhibitor inducing exosomal miR-34a-5p expression mediates the cross talk between cardiomyocyte and macrophage in immune checkpoint inhibitor–related cardiac dysfunction

“…Exosomes are cell-derived vesicles, which contain proteins, lipids, growth factors, microRNAs (miRNAs), and proinflammatory cytokines released through exocytosis. 14 The recent literature suggests that condition-modified exosomes induce cardiac injury. 15 Macrophage-derived exosomes in patients with diabetes showed proinflammation function, leading to cardiac fibrosis.…”

<p>Spinal TLR4/P2X7 Receptor-Dependent NLRP3 Inflammasome Activation Contributes to the Development of Tolerance to Morphine-Induced Antinociception</p>

“…18,19 In addition, systemic TLR4 antagonists attenuate morphine-induced tolerance. 16 Furthermore, in other inflammatory disease models, 20,21 TLR4 acts as the upstream of NLRP3 inflammasome. Therefore, we assessed whether the activation of the NLRP3 inflammasome is dependent on TLR4 during the development of morphine-induced antinociceptive tolerance.…”