Pregestational Diabetes Induced Congenital Heart Defects and Coronary Artery Malformations; Mechanisms and Preventative Therapies

Congenital heart defects (CHDs) arise from perturbations in complex molecular and cellular processes underlying normal embryonic heart development. CHDs are the most common congenital malformation, occurring in 1 to 5% of live births, and are the leading cause of pediatric mortality. Adverse genetic and environmental factors can impede normal cardiogenesis and increase the likelihood of CHDs. Pregestational maternal diabetes increases the risk of CHDs in children by more than four-fold. As the prevalence of diabetes rapidly rises among women of childbearing age, there is a need to investigate the mechanisms and potential preventative strategies for these defects. The aim of this thesis was to explore the pathogenesis of pregestational diabetes-induced CHDs and coronary artery malformations (CAMs), while testing the efficacy of two clinically relevant pharmacotherapies. To this end, using a mouse model of pregestational diabetes, I examined the impact of hyperglycemia-induced elevations in oxidative stress and miR-122 on heart development, concurrently determining the preventative capabilities of sapropterin or antimiR-122 treatment. I confirmed that pregestational diabetes results in spectrum of CHDs, CAMs and cardiac function deficits, and that their incidence is significantly lowered with either sapropterin or antimiR-122. Specifically, sapropterin treatment lowered the incidence of CHDs and CAMs from 59% and 50% to 27% and 21%, respectively. Similarly, antimiR-122 therapy reduced this incidence of CHDs from 57% to 23%. These morphological malformations range in severity, and include septal and outflow defects (OFT), myocardium deficiencies, and hypoplastic coronary arteries. Lineage tracing experiments revealed a diminished commitment of second heart field progenitors to the OFT, endocardial cushions and ventricular myocardium in embryonic hearts from diabetic dams. In addition, deficits in cardiogenic gene expression, enzyme activity, cell proliferation, and epicardial EMT, induced by pregestational diabetes, contribute to these defects, and were prevented by both treatments. Specifically, sapropterin treatment reestablished the functional eNOS dimer and restored its phosphorylation in embryonic hearts of diabetic dams, leading to normal cardiovascular development. Conversely, antimiR-122 attenuated the targeting and inhibition of key genes responsible for cardiogenesis by miR-122. These results suggest that sapropterin and antimiR-122 may have therapeutic potential in preventing CHDs in children of women with pregestational diabetes

Outcomes of Patients Referred for Arteriovenous Fistula Construction: A Systematic Review

Chronic Kidney Disease (CKD) affects 10-16% of the US population and its incidence is rising due to increasing prevalence of associated risk factors. Renal replacement therapy is required to treat late stage CKD and hemodialysis is the preferred modality for many patients. Vascular access is required for hemodialysis and arteriovenous fistulas (AVF) are currently the gold standard. This review intended to collate current knowledge on AVF outcomes regarding both the patient and fistula. Scopus and Medline were utilized to identify relevant literature. Inclusion and exclusion criteria were applied to narrow search results. Among CKD patients, 33.5-77.4% require a central venous catheter (CVC) before dialysis through a fistula. Many patients (33-51%) use a CVC regardless of AVF creation due to fistula immaturity or failure. There are large variations in AVF creation policies internationally; 16% of American hemodialysis patients use a fistula compared to 72% of German patients. Primary patency and primary AVFs' failure ranges from 60-70% and 20-26%, respectively. AVFs reduce morbidity and mortality in CKD. At present, too many patients are receiving hemodialysis through a CVC. Inadequate referral times for AVF creation can lead to fistula immaturity or failure in the intervention. Many countries are lagging behind recommended AVF creation rates published by the Kidney Disease Outcomes Quality Initiative. There is a paucity of literature concerning when a patient should be referred for AVF creation. It is paramount to have better predictive outcome measures and more clarity as to when patients will benefit from an AVF

The Role of Intraindividual Carotid Artery Variation in the Development of Atherosclerotic Carotid Artery Disease: A Literature Review

Carotid artery disease (CAD) is associated with numerous risk factors, including hypertension, hyperlipidemia, hypercholesterolemia, diabetes mellitus, and smoking. These systemic risk factors do not affect the carotid arteries equally in most patients, resulting in asymmetrical bilateral and unilateral CAD. It is unclear if anatomic variations in the carotid arteries predispose an individual to formation of atherosclerotic CAD. We wanted to assess (1) the inter-individual or intra-individual anatomical variations in the carotid arteries and (2) whether anatomical variations predispose the development of atherosclerotic CAD. PubMed and Medline were utilized to identify relevant literature for critical appraisal, summarization and documentation. Inclusion and exclusion criteria were applied to narrow results and articles were critically appraised and analyzed. Evidence suggests that a low outflow/inflow ratio, elevated bifurcation height, and bifurcation angle are associated with increased risk for CAD. Sex and age demonstrated positive correlation with the disease. Additionally, tortuosity and kinking of the carotid arteries may affect the formation of CAD but coiling of the arteries is a natural age-dependent process and does not affect CAD development. This review suggests there are anatomic variations in the carotid arteries that increase the risk of developing carotid artery disease. The most significant risk factors include a low outflow/inflow ratio, increased internal carotid artery tortuosity, elevated bifurcation height, and bifurcation angle

Say NO to ROS: Their roles in embryonic heart development and pathogenesis of congenital heart defects in maternal diabetes

Congenital heart defects (CHDs) are the most prevalent and serious birth defect, occurring in 1% of all live births. Pregestational maternal diabetes is a known risk factor for the development of CHDs, elevating the risk in the child by more than four-fold. As the prevalence of diabetes rapidly rises among women of childbearing age, there is a need to investigate the mechanisms and potential preventative strategies for these defects. In experimental animal models of pregestational diabetes induced-CHDs, upwards of 50% of offspring display congenital malformations of the heart, including septal, valvular, and outflow tract defects. Specifically, the imbalance of nitric oxide (NO) and reactive oxygen species (ROS) signaling is a major driver of the development of CHDs in offspring of mice with pregestational diabetes. NO from endothelial nitric oxide synthase (eNOS) is crucial to cardiogenesis, regulating various cellular and molecular processes. In fact, deficiency in eNOS results in CHDs and coronary artery malformation. Embryonic hearts from diabetic dams exhibit eNOS uncoupling and oxidative stress. Maternal treatment with sapropterin, a cofactor of eNOS, and antioxidants such as N-acetylcysteine, vitamin E, and glutathione as well as maternal exercise have been shown to improve eNOS function, reduce oxidative stress, and lower the incidence CHDs in the offspring of mice with pregestational diabetes. This review summarizes recent data on pregestational diabetes-induced CHDs, and offers insights into the important roles of NO and ROS in embryonic heart development and pathogenesis of CHDs in maternal diabetes

Myocardium-specific deletion of rac1 causes ventricular noncompaction and outflow tract defects

Background: Left ventricular noncompaction (LVNC) is a cardiomyopathy that can lead to arrhythmias, embolic events and heart failure. Despite our current knowledge of cardiac development, the mechanisms underlying noncompaction of the ventricular myocardium are still poorly understood. The small GTPase Rac1 acts as a crucial regulator of numerous developmental events. The present study aimed to investigate the cardiomyocyte specific role of Rac1 in embryonic heart development. Methods and Results: The Nkx2.5-Cre transgenic mice were crossed with Rac1f/f mice to generate mice with a cardiomyocyte specific deletion of Rac1 (Rac1Nkx2.5 ) during heart development. Embryonic Rac1Nkx2.5 hearts at E12.5–E18.5 were collected for histological analysis. Overall, Rac1Nkx2.5 hearts displayed a bifid apex, along with hypertrabeculation and a thin compact myocardium. Rac1Nkx2.5 hearts also exhibited ventricular septal defects (VSDs) and double outlet right ventricle (DORV) or overriding aorta. Cardiomyocytes had a rounded morphology and were highly disorganized, and the myocardial expression of Scrib, a planar cell polarity protein, was reduced in Rac1Nkx2.5 hearts. In addition, cell proliferation rate was significantly decreased in the Rac1Nkx2.5 ventricular myocardium at E9.5. Conclusions: Rac1 deficiency in the myocardium impairs cardiomyocyte elongation and organization, and proliferative growth of the heart. A spectrum of CHDs arises in Rac1Nkx2.5 hearts, implicating Rac1 signaling in the ventricular myocardium as a crucial regulator of OFT alignment, along with compact myocardium growth and development

Maternal nicotine exposure induces congenital heart defects in the offspring of mice

Maternal cigarette smoking is a risk factor for congenital heart defects (CHDs). Nicotine replacement therapies are often offered to pregnant women following failed attempts of smoking cessation. However, the impact of nicotine on embryonic heart development is not well understood. In the present study, the effects of maternal nicotine exposure (MNE) during pregnancy on foetal heart morphogenesis were studied. Adult female mice were treated with nicotine using subcutaneous osmotic pumps at 0.75 or 1.5 mg/kg/day and subsequently bred with male mice. Our results show that MNE dose-dependently increased CHDs in foetal mice. CHDs included atrial and ventricular septal defects, double outlet right ventricle, unguarded tricuspid orifice, hypoplastic left ventricle, thickened aortic and pulmonary valves, and ventricular hypertrophy. MNE also significantly reduced coronary artery size and vessel abundance in foetal hearts. Moreover, MNE resulted in higher levels of oxidative stress and altered the expression of key cardiogenic regulators in the developing heart. Nicotine exposure reduced epicardial-to-mesenchymal transition in foetal hearts. In conclusion, MNE induces CHDs and coronary artery malformation in mice. These findings provide insight into the adverse outcomes of foetuses by MNE during pregnancy

Maternal voluntary exercise mitigates oxidative stress and incidence of congenital heart defects in pre-gestational diabetes

Women with pre-gestational diabetes have a higher risk of producing children with congenital heart defects (CHDs), caused predominantly by hyperglycemia-induced oxidative stress. In this study, we evaluated if exercise during pregnancy could mitigate oxidative stress and reduce the incidence of CHDs in the offspring of diabetic mice. Female mice were treated with streptozotocin to induce pre-gestational diabetes, then mated with healthy males to produce offspring. They were also given access to running wheels 1 week before mating and allowed to exercise voluntarily until E18.5. Heart morphology, gene expression, and oxidative stress were assessed in foetal hearts. Maternal voluntary exercise results in a significantly lower incidence of CHDs from 59.5% to 25%. Additionally, diabetes-induced defects in coronary artery and capillary morphogenesis were also lower with exercise. Myocardial cell proliferation and epithelial-mesenchymal transition at E12.5 was significantly lower with pre-gestational diabetes which was mitigated with maternal exercise. Cardiac gene expression of Notch1, Snail1, Gata4 and Cyclin D1 was significantly higher in the embryos of diabetic mice that exercised compared to the non-exercised group. Furthermore, maternal exercise produced lower reactive oxygen species (ROS) and oxidative stress in the foetal heart. In conclusion, maternal exercise mitigates ROS and oxidative damage in the foetal heart, and results in a lower incidence of CHDs in the offspring of pre-gestational diabetes. Exercise may be an effective intervention to compliment clinical management and further minimize CHD risk in mothers with diabetes

Sapropterin Treatment Prevents Congenital Heart Defects Induced by Pregestational Diabetes Mellitus in Mice.

Background Tetrahydrobiopterin is a cofactor of endothelial NO synthase ( eNOS ), which is critical to embryonic heart development. We aimed to study the effects of sapropterin (Kuvan), an orally active synthetic form of tetrahydrobiopterin on eNOS uncoupling and congenital heart defects ( CHD s) induced by pregestational diabetes mellitus in mice. Methods and Results Adult female mice were induced to pregestational diabetes mellitus by streptozotocin and bred with normal male mice to produce offspring. Pregnant mice were treated with sapropterin or vehicle during gestation. CHD s were identified by histological analysis. Cell proliferation, eNOS dimerization, and reactive oxygen species production were assessed in the fetal heart. Pregestational diabetes mellitus results in a spectrum of CHD s in their offspring. Oral treatment with sapropterin in the diabetic dams significantly decreased the incidence of CHD s from 59% to 27%, and major abnormalities, such as atrioventricular septal defect and double-outlet right ventricle, were absent in the sapropterin-treated group. Lineage tracing reveals that pregestational diabetes mellitus results in decreased commitment of second heart field progenitors to the outflow tract, endocardial cushions, and ventricular myocardium of the fetal heart. Notably, decreased cell proliferation and cardiac transcription factor expression induced by maternal diabetes mellitus were normalized with sapropterin treatment. Furthermore, sapropterin administration in the diabetic dams increased eNOS dimerization and lowered reactive oxygen species levels in the fetal heart. Conclusions Sapropterin treatment in the diabetic mothers improves eNOS coupling, increases cell proliferation, and prevents the development of CHD s in the offspring. Thus, sapropterin may have therapeutic potential in preventing CHD s in pregestational diabetes mellitus

Myocardium-Specific Deletion of Rac1 Causes Ventricular Noncompaction and Outflow Tract Defects

Background: Left ventricular noncompaction (LVNC) is a cardiomyopathy that can lead to arrhythmias, embolic events and heart failure. Despite our current knowledge of cardiac development, the mechanisms underlying noncompaction of the ventricular myocardium are still poorly understood. The small GTPase Rac1 acts as a crucial regulator of numerous developmental events. The present study aimed to investigate the cardiomyocyte specific role of Rac1 in embryonic heart development. Methods and Results: The Nkx2.5-Cre transgenic mice were crossed with Rac1f/f mice to generate mice with a cardiomyocyte specific deletion of Rac1 (Rac1Nkx2.5) during heart development. Embryonic Rac1Nkx2.5 hearts at E12.5–E18.5 were collected for histological analysis. Overall, Rac1Nkx2.5 hearts displayed a bifid apex, along with hypertrabeculation and a thin compact myocardium. Rac1Nkx2.5 hearts also exhibited ventricular septal defects (VSDs) and double outlet right ventricle (DORV) or overriding aorta. Cardiomyocytes had a rounded morphology and were highly disorganized, and the myocardial expression of Scrib, a planar cell polarity protein, was reduced in Rac1Nkx2.5 hearts. In addition, cell proliferation rate was significantly decreased in the Rac1Nkx2.5 ventricular myocardium at E9.5. Conclusions: Rac1 deficiency in the myocardium impairs cardiomyocyte elongation and organization, and proliferative growth of the heart. A spectrum of CHDs arises in Rac1Nkx2.5 hearts, implicating Rac1 signaling in the ventricular myocardium as a crucial regulator of OFT alignment, along with compact myocardium growth and development

NOX2 Is Critical to Endocardial to Mesenchymal Transition and Heart Development

NADPH oxidases (NOX) are a major source of reactive oxygen species (ROS) production in the heart. ROS signaling regulates gene expression, cell proliferation, apoptosis, and migration. However, the role of NOX2 in embryonic heart development remains elusive. We hypothesized that deficiency of Nox2 disrupts endocardial to mesenchymal transition (EndMT) and results in congenital septal and valvular defects. Our data show that 34% of Nox2-/- neonatal mice had various congenital heart defects (CHDs) including atrial septal defects (ASD), ventricular septal defects (VSD), atrioventricular canal defects (AVCD), and malformation of atrioventricular and aortic valves. Notably, Nox2-/- embryonic hearts show abnormal development of the endocardial cushion as evidenced by decreased cell proliferation and an increased rate of apoptosis. Additionally, Nox2 deficiency disrupted EndMT of atrioventricular cushion explants ex vivo. Furthermore, treatment with N-acetylcysteine (NAC) to reduce ROS levels in the wild-type endocardial cushion explants decreased the number of cells undergoing EndMT. Importantly, deficiency of Nox2 was associated with reduced expression of Gata4, Tgfβ2, Bmp2, Bmp4, and Snail1, which are critical to endocardial cushion and valvoseptal development. We conclude that NOX2 is critical to EndMT, endocardial cushion cell proliferation, and normal embryonic heart development