Epigenetics of Notch1 regulation in pulmonary microvascular rarefaction following extrauterine growth restriction.

BACKGROUND: Extrauterine growth restriction (EUGR) plays an important role in the developmental origin of adult cardiovascular diseases. In an EUGR rat model, we reported an elevated pulmonary arterial pressure in adults and genome-wide epigenetic modifications in pulmonary vascular endothelial cells (PVECs). However, the underlying mechanism of the early nutritional insult that results in pulmonary vascular consequences later in life remains unclear. METHODS: A rat model was used to investigate the physiological and structural effect of EUGR on early pulmonary vasculature by evaluating right ventricular systolic pressure and pulmonary vascular density in male rats. Epigenetic modifications of the Notch1 gene in PVECs were evaluated. RESULTS: EUGR decreased pulmonary vascular density with no significant impact on right ventricular systolic pressure at 3 weeks. Decreased transcription of Notch1 was observed both at 3 and 9 weeks, in association with decreased downstream target gene, Hes-1. Chromatin immunoprecipitation and bisulfite sequencing were performed to analyze the epigenetic modifications of the Notch1 gene promoter in PVECs. EUGR caused a significantly increased H3K27me3 in the proximal Notch1 gene promoter, and increased methylation of single CpG sites in the distal Notch1 gene promoter, both at 3 and 9 weeks. CONCLUSIONS: We conclude that EUGR results in decreased pulmonary vascular growth in association with decreased Notch1 in PVECs. This may be mediated by increased CpG methylation and H3K27me3 in the Notch1 gene promoter region.

Epigenetics of hyper-responsiveness to allergen challenge following intrauterine growth retardation rat.

BACKGROUND: Epidemiological studies have revealed that intrauterine growth retardation (IUGR) or low birth weight is linked to the later development of asthma. Epigenetic regulatory mechanisms play an important role in the fetal origins of adult disease. However, little is known regarding the correlation between epigenetic regulation and the development of asthma following IUGR. METHODS: An IUGR and ovalbumin (OVA)-sensitization/challenge rat model was used to study whether epigenetic mechanisms play a role in the development of asthma following IUGR. RESULTS: Maternal nutrient restriction increased histone acetylation levels of the endothelin-1 (ET-1) gene promoter in lung tissue of offspring, but did not cause significant alterations of DNA methylation. The effect was maintained until 10 weeks after birth. Furthermore, these epigenetic changes may have induced IUGR individuals to be highly sensitive to OVA challenge later in life, resulting in more significant changes related to asthma. CONCLUSIONS: These findings suggest that epigenetic mechanisms might be closely associated with the development of asthma following IUGR, providing further insight for improved prevention of asthma induced by environmental factors.

Decreased Kv1.5 expression in intrauterine growth retardation rats with exaggerated pulmonary hypertension.

Chronic hypoxia pulmonary hypertension (CH-PHT) in adulthood is likely to be of fetal origin following intrauterine growth retardation (IUGR). Oxygen (O2)-sensitive voltage-gated potassium channels (Kv channels) in resistance pulmonary artery smooth muscle cells (PASMCs) play an important role in scaling pulmonary artery (PA) pressure. Expression and functional changes of Kv channels are determined, in part, by embryonic development. We hypothesized that O2-sensitive Kv channels play an important role in exaggerated CH-PHT following IUGR. We established a rat model of IUGR by restricting maternal food during the entire pregnancy and exposed IUGR rats and their age-matched controls aged 12 wk to hypoxia for 2 wk. We found that hypoxia exposure significantly induced increased PA pressure and thicker smooth muscle layer in the IUGR group relative to controls. We compared the constriction of the resistance PA to inhibitors of K⁺ channels, 4-aminopyridine (4-AP), tetraethylammonium, and BaCl2. Despite the thickness of the smooth muscle layer, the constriction to 4-AP was significantly reduced in the IUGR group exposed to hypoxia. Consistent with these changes in pulmonary vascular reactivity, 2 wk of hypoxia induced weaker 4-AP-sensitive Kv currents in a single IUGR PASMC. Moreover, after 2 wk of hypoxia, Kv1.5 expression in resistance PAs decreased significantly in the IUGR group. Overexpression of Kv1.5 in cultured PASMCs could offset hypoxia-induced cell proliferation and hypoxia-inhibited Kv currents in the IUGR group. These results suggest that the inhibited expression of Kv1.5 in PASMCs contribute to the development of exaggerated CH-PHT in IUGR rats during adulthood.

Increased Expression of MicroRNA-206 Inhibits Potassium Voltage-Gated Channel Subfamily A Member 5 in Pulmonary Arterial Smooth Muscle Cells and Is Related to Exaggerated Pulmonary Artery Hypertension Following Intrauterine Growth Retardation in Rats.

Background Intrauterine growth retardation ( IUGR ) is related to pulmonary artery hypertension in adults, and mi croRNA -206 (miR-206) is proposed to affect the proliferation and apoptosis of pulmonary artery smooth muscle cells ( PASMC s) via post-transcriptional regulation. Methods and Results In an IUGR rat model, we found that the expression and function of potassium voltage-gated channel subfamily A member 5 (Kv1.5) in PASMC s was inhibited, and pulmonary artery hypertension was exaggerated after chronic hypoxia ( CH ) treatment as adults. micro RNA expression was investigated in PASMC s from 12-week-old male IUGR rats with CH by microarray, polymerase chain reaction, and in situ hybridization. The expression levels of Kv1.5 in primary cultured PASMC s and pulmonary artery smooth muscle from IUGR or control rats were evaluated with and without application of an miR-206 inhibitor. Right ventricular systolic pressure, cell proliferation, luciferase reporter assay, and IKv were also calculated. We found increased expression of miR-206 in resistance pulmonary arteries of IUGR rats at 12 weeks compared with newborns. Application of an miR-206 inhibitor in vivo or in vitro increased expression of Kv1.5 α-protein and KCNA 5. Also, decreased right ventricular systolic pressure and cell proliferation were observed in PASMC s from 12-week-old control and IUGR rats after CH , while inhibitor did not significantly affect control and IUGR rats. Conclusions These results suggest that expression of Kv1.5 and 4-aminopyridine (Kv channel special inhibitor)-sensitive Kv current were correlated with the inhibition of miR-206 in PA rings of IUGR - CH rats and cultured IUGR PASMC s exposed to hypoxia. Thus, miR-206 may be a trigger for induction of exaggerated CH-pulmonary artery hypertension of IUGR via Kv1.5.

Epigenetic Regulation and Its Therapeutic Potential in Pulmonary Hypertension.

Recent advances in epigenetics have made a tremendous impact on our knowledge of biological phenomena and the environmental stressors on complex diseases. Understanding the mechanism of epigenetic reprogramming during the occurrence of pulmonary hypertension (PH) is important for advanced studies and clinical therapy. In this article, we review the discovery of novel epigenetic mechanisms associated with PH including DNA methylation, histone modification, and noncoding RNA interference. In addition, we highlight the role of epigenetic mechanisms in adult PAH resulting from undesirable perinatal environments-Extrauterine growth restriction (EUGR) and Intrauterine growth retardation (IUGR). Lastly, we give a comprehensive summary for the remaining challenges and discuss future methods of epigenetic targeted therapy for pulmonary hypertension.

Extrauterine growth restriction on pulmonary vascular endothelial dysfunction in adult male rats: the role of epigenetic mechanisms.

OBJECTIVE: Early postnatal life is considered as a critical time window for the determination of long-term metabolic states and organ functions. Extrauterine growth restriction (EUGR) causes the development of adult-onset chronic diseases, including pulmonary hypertension. However, the effects of nutritional disadvantages during the early postnatal period on pulmonary vascular consequences in later life are not fully understood. Our study was designed to test whether epigenetics dysregulation mediates the cellular memory of this early postnatal event. METHODS AND RESULTS: To test this hypothesis, we isolated pulmonary vascular endothelial cells by magnetic-activated cell sorting from EUGR and control rats. A postnatal insult, nutritional restriction-induced EUGR caused development of an increased pulmonary artery pressure at 9 weeks of age in male Sprague-Dawley rats. Methyl-DNA immune precipitation chip, genome-scale mapping studies to search for differentially methylated loci between control and EUGR rats, revealed significant difference in cytosine methylation between EUGR and control rats. EUGR changes the cytosine methylation at approximately 500 loci in male rats at 9 weeks of age, preceding the development of pulmonary hypertension and these represent the candidate loci for mediating the pathogenesis of pulmonary vascular disease that occurs later in life. Gene ontology analysis on differentially methylated genes showed that hypermethylated genes in EUGR are vascular development-associated genes and hypomethylated genes in EUGR are late-differentiation-associated and signal transduction genes. We validated candidate dysregulated loci with the quantitative assays of cytosine methylation and gene expressions. CONCLUSION: These results demonstrate that epigenetics dysregulation is a strong mechanism for propagating the cellular memory of early postnatal events, causing changes in the expression of genes and long-term susceptibility to pulmonary hypertension, and further providing a new insight into the prevention and treatment of EUGR-related pulmonary hypertension.