Project Summary The prevalence of heart attack, also known as myocardial infarction (MI), in pregnancy has increased over the past decades due in part to rising maternal age and lifestyle changes. MI during late pregnancy carries a worse prognosis than in non-pregnant (NP) women for unclear reasons. The late-pregnant (LP) rodent is more prone to myocardial ischemia-reperfusion injury (IRI) compared to the NP rodent. Furthermore, reactive oxygen species (ROS) production in the heart is higher in LP mice subjected to IRI. However, the underlying molecular mechanisms involved in the higher susceptibility of late pregnancy to IRI are not clear. MicroRNAs (miRs) have emerged as a powerful novel regulator of gene expression. In the heart, several miRs have been involved in IRI, suggesting that regulation of their function could be used as a novel cardioprotective strategy. MicroRNA microarray analysis showed that the expression of microRNA-98-5p (miR98) was highly increased in the LP heart compared to the NP heart at baseline. Strikingly, miR98 was increased further in the LP heart compared to the NP heart when subjected to IRI. These miR98 expression dynamics may underlie the higher vulnerability and predisposition of the LP heart to IRI. miR98 is also significantly increased in the plasma of pregnant women at high risk of cardiovascular disease compared to healthy pregnant patients and healthy non-pregnant patients in the reproductive age. In vitro data shows that the expression of miR98 is increased in female cardiomyocytes subjected to hypoxia/reoxygenation, which mimics IRI. Knockdown of miR98 reduces apoptosis, ROS production, and inflammatory markers in female cardiomyocytes subjected to H/R. To investigate the mechanism of action of miR98, TargetScan software was used to predict miR98 targets. STAT3 and PGC-1α were strongly predicted as miR98 targets. STAT3 and PGC-1α are known negative regulators of reactive oxygen species production and their deletion in cardiomyocytes in vivo has been shown to predispose mice to peripartum cardiomyopathy. Expression of STAT3 and PGC-1α are lower in the LP rat hearts subjected to I/R injury compared to the NP rat hearts subjected to I/R injury. Furthermore, knockdown of miR98 in vitro increases STAT3 and PGC-1α protein expression, whereas miR98 overexpression reduces their expression, suggesting they could be targets of miR98. The hypotheses of this proposal are, i) induction of miR98 in LP leads to higher cardiac vulnerability to IRI via targeting STAT3 and PGC-1α, leading to increased cardiomyocyte death, ROS production, and inflammation, and ii) miR98 has the potential to be served as a therapeutic modality and as a biomarker for early prediction of cardiovascular disease in pregnancy.