Dental pulp stem cells (DPSCs) have previously demonstrated potential pericyte-like topography and function. However, the mechanisms regulating their pericyte function are still unknown. In this study, murine DPSC angiogenic and pericyte function were investigated. Tie2-GFP mouse DPSCs were negative for GFP, indicating the absence of endothelial cells in DPSC cultures. Endothelial cells co-cultured with DPSCs formed more mature in vitro tube-like structures as compared with those co-cultured with bone marrow stromal cells (BMSCs). Many DPSCs were located adjacent to vascular tubes, assuming a pericyte location. Subcutaneous DPSC transplants in mice with matrigel (MG) (DPSC-MG) induced more vessel formation than BMSC-MG. Soluble Flt (sFlt), an angiogenic inhibitor that binds VEGF-A, significantly decreased the amount of blood vessels in DPSC-MG, but not in BMSC-MG. sFlt inhibited VEGFR2 and downstream ERK signaling in DPSCs. Similar to sFlt inhibition, VEGFR2 knockdown in DPSCs resulted in down-regulation of Vegfa, Vegf receptors, and EphrinB2 and decreased angiogenic induction of DPSCs in vivo. Therefore, the capacity of DPSCs to induce angiogenesis is VEGFR2-dependent. DPSCs enhance angiogenesis by secreting VEGF ligands and associating with vessels resembling pericyte-like cells. This study provides first insights into the mechanism(s) of DPSC angiogenic induction and their function as pericytes, crucial aspects for DPSC use in tissue regeneration.

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Mutations in TAB2 (transforming growth factor β activated kinase 1 binding protein 2) have been implicated in the pathogenesis of dilated cardiomyopathy and/or congenital heart disease in humans, but the underlying mechanisms are currently unknown. Here we identified an indispensable role for TAB2 in regulating myocardial homeostasis and remodeling by suppressing RIPK1 (receptor-interacting protein kinase 1) activation and RIPK1-dependent apoptosis and necroptosis. Cardiomyocyte-specific deletion of Tab2 in mice triggered dilated cardiomyopathy with massive apoptotic and necroptotic cell death. Moreover, Tab2-deficient mice were also predisposed to myocardial injury and adverse remodeling following pathological stress. In cardiomyocytes, deletion of TAB2, but not its close homologue TAB3, promoted TNFa-induced apoptosis and necroptosis, which was rescued by forced activation of TAK1 or inhibition of RIPK1 kinase activity. Mechanistically, TAB2 critically mediates RIPK1 phosphorylation at Ser321 via a TAK1-dependent mechanism, which prevents RIPK1 kinase activation and the formation of RIPK1-FADD-caspase-8 apoptotic complex or RIPK1-RIPK3 necroptotic complex. Strikingly, genetic inactivation of RIPK1 with Ripk1-K45A knock-in effectively rescued cardiac remodeling and dysfunction in Tab2-deficient mice. Together, these data demonstrate that TAB2 is a key regulator of myocardial homeostasis and remodeling by suppressing RIPK1-dependent apoptosis and necroptosis. Our results also suggest that targeting RIPK1-mediated cell death signaling may represent a promising therapeutic strategy for TAB2 deficiency-induced dilated cardiomyopathy.

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Objective: Despite the well-studied pro-survival function of nuclear factor-κB (NFκB), recent studies suggest that NFκB may also play a pathogenic role in myocardial ischemia injury and adverse remodeling. This study aims to define a new pro-cell death role of NFκB in response to oxidative stress and the functional implications in ischemia reperfusion (I/R) injury. Methods and Results: We identified an unexpected pro-cell death role of NFκB in oxidative stress-induced necrosis, and provide new mechanistic evidence that NFκB, in cooperation with HDAC3, negatively regulates NRF2-ARE anti-oxidative signaling through transcriptional silencing. Genetic deletion of NFκB-p65 inhibits, whereas activation of NFκB promotes, oxidative stress-induced cell death and HMGB1 release, a biomarker of necrosis. Moreover, simulated ischemia reperfusion (sI/R) and doxorubicin (Dox) treatment both induce NFκB-luciferase activity in cardiomyocytes, and inhibition of NFκB diminishes sI/R- and Dox- induced necrosis. Importantly, NFκB negatively regulates NRF2-ARE activity and the expression of anti-oxidant proteins. Mechanistically, co-immunoprecipitation reveals that p65 is required for the association between NRF2 and HDAC3 and transcriptional silencing of NRF2-ARE activity. Further, the ability of HDAC3 to repress NRF2-ARE activity is lost in p65-/- cells. The HADC inhibitor TSA and NFκB inhibitor BMS-345541 both increase NRF2-ARE activity and promote cell survival following sI/R. In vivo , NFκB transcriptional activity in the heart is significantly elevated after I/R injury, which is abolished by cardiac-specific deletion of p65. Moreover, ablation of p65 using p65 fl/fl-Nkx-Cre mice reduces myocardial infarct size after acute I/R, and prevents chronic remodeling and contractile dysfunction after myocardial infarction. Conclusions: Our results identified NFκB as a key regulator of oxidative stress-induced necrosis by suppressing the NRF2-ARE anti-oxidant pathway through an HDAC3-dependent mechanism. Ablation of NFκB-p65 attenuates oxidative stress-induced necrosis and I/R injury, suggesting a new pathogenic role of the NFκB pathway, and thus a therapeutic target, in myocardial ischemic injury and remodeling.