Hyperglycemia of Maternal Diabetes Induces Cardiac Isl1 Positive Progenitor Dysfunction Leading to Heart Defects

妊娠期糖尿病是一种与先天性心脏缺陷（CHDs）增加5倍相关的非遗传因素。以Isl1标记的第二心场（SHF）祖细胞在环形态发生和心室形成过程中驱动心管延伸。糖尿病诱导冠心病的潜在机制尚不清楚，但其中一种机制可能与母亲糖尿病抑制Isl1+ SHF祖细胞驱动的心脏发生有关。在过去的十年中，我们专注于冠心病患者心脏祖细胞的生物学和再生能力。为了最大限度地发挥其在冠心病患者中的再生和保护潜力，确定体内糖尿病和体外高糖对胚胎发生和出生后Isl1+祖细胞的生物学效应至关重要。因此，我们的总体假设是，在心脏发育的关键时期，母亲糖尿病的高血糖通过增加氧化应激，主要UPR传感器IRE1α及其下游转录因子XBP1的激活，诱导Isl1+ SHF祖细胞功能障碍，这是负责DNA高甲基化和SHF基因沉默，导致RNA甲基转移酶METTL14和m6A RNA甲基化的抑制。抑制细胞应激或调节DNA/RNA甲基化可改善SHF祖细胞缺陷、冠心病形成和这些祖细胞的潜在再生能力。目的1将确定母亲糖尿病的高血糖是否通过氧化应激诱导心脏发育过程中Isl1+ SHF祖细胞功能障碍。我们假设糖尿病通过诱导氧化应激导致线粒体功能障碍和心脏形态发生，而通过超氧化物歧化酶1 （SOD1）减轻氧化应激可减轻糖尿病妊娠期间冠心病的形成。目的2将确定主要的UPR传感器IRE1α及其下游效应物XBP1在糖尿病妊娠中导致冠心病的Isl1+ SHF祖细胞中的作用。我们将验证氧化应激对Isl1+ SHF祖细胞中的ER应激和UPR负责的假设，以及通过使主要的UPR传感器IRE1α或其下游转录因子XBP1失活性来抑制ER应激-UPR通路可减少糖尿病诱导的冠心病。目的3将确定Isl1+ SHF祖细胞中DNA甲基转移酶抑制的RNA甲基化是否有助于糖尿病诱导的冠心病，以及这些祖细胞的治疗意义。我们预计，增加的DNA甲基化抑制了Isl1+祖细胞功能所必需的RNA甲基转移酶样14 （METTL14）和RNA N(6)-甲基腺苷（m6A），减少DNA甲基化或恢复Isl1+祖细胞特异性的RNA甲基化可减少冠心病并增加这些细胞的治疗价值。

Pregestational maternal diabetes is a noninherited factor associated with a fivefold increase in congenital heart defects (CHDs). The second heart field (SHF) progenitors, marked by Isl1, drive the heart tube extension during looping morphogenesis and cardiac chamber formation. The underlying mechanism of diabetes-induced CHDs is unknown but one mechanism may involve the inhibition of Isl1+ SHF progenitor-driven cardiogenesis by maternal diabetes. During the last decade, we have focused on the biology and regenerative capability of cardiac progenitors in CHD patients. It is critical to determine the biological effects of diabetes in vivo and high glucose in vitro on Isl1+ progenitors during embryogenesis and postnatally in order to maximize their regenerative and protective potentials in CHD patients. Therefore, our overarching hypothesis that hyperglycemia of maternal diabetes induces Isl1+ SHF progenitor dysfunction during the critical period of cardiac development through heightened oxidative stress, activation of the major UPR sensor IRE1α and its downstream transcription factor XBP1, which is responsible for DNA hypermethylation and SHF gene silencing leading to repression of RNA methyltransferase METTL14 and m6A RNA methylation. Suppressing cellular stress or modulating DNA/RNA methylation ameliorates defects in SHF progenitors, CHD formation and potential regenerative capacity of these progenitors. Aim 1 will determine whether hyperglycemia of maternal diabetes induces Isl1+ SHF progenitor dysfunction during heart development through oxidative stress. We hypothesize that diabetes causes mitochondrial dysfunction and during cardiac morphogenesis through the induction of oxidative stress and that mitigation of oxidative stress by superoxide dismutase 1 (SOD1) alleviates CHD formation in diabetic pregnancy. Aim 2 will determine the role of the major UPR sensor IRE1α and its downstream effector XBP1 in Isl1+ SHF progenitors leading to CHDs in diabetic pregnancy. We will test the hypothesis that oxidative stress is responsible for ER stress and UPR in Isl1+ SHF progenitors and that suppressing the ER stress-UPR pathway by inactivating either the major UPR sensor IRE1α or its downstream transcription factor XBP1 reduces diabetes-induced CHDs. Aim 3 will determine whether DNA methyltransferases-suppressed RNA methylation in Isl1+ SHF progenitors contributes to diabetes- induced CHDs and the therapeutic implications of these progenitors. We expect that that increased DNA methylation represses RNA methyltransferase-like 14 (METTL14) and RNA N(6)-methyladenosine (m6A) essential for Isl1+ progenitor function and that reducing DNA methylation or restoring RNA methylation specifically in Isl1+ progenitors reduces CHDs and increases the therapeutic values of these cells.