AUTHOR=Rajgarhia Ayan , Ayasolla Kameshwar R. , Zaghloul Nahla , Lopez Da Re Jorge M. , Miller Edmund J. , Ahmed Mohamed 

TITLE=Extracellular Superoxide Dismutase (EC-SOD) Regulates Gene Methylation and Cardiac Fibrosis During Chronic Hypoxic Stress

JOURNAL=Frontiers in Cardiovascular Medicine

VOLUME=Volume 8 - 2021

YEAR=2021

URL=https://www.frontiersin.org/journals/cardiovascular-medicine/articles/10.3389/fcvm.2021.669975

DOI=10.3389/fcvm.2021.669975

ISSN=2297-055X

ABSTRACT=Chronic hypoxic stress induces epigenetic modifications mainly DNA methylation in cardiac fibroblasts, inactivating tumor suppressor genes (RASSF1A), and activating kinases (ERK1/2) leading to fibroblast proliferation and cardiac fibrosis. The Ras/ERK signaling pathway is an intracellular signal transduction critically involved in fibroblast proliferation. RASSF1A functions through its effect on downstream ERK1/2. The antioxidant enzyme, extracellular superoxide dismutase (EC-SOD), decreases oxidative stress from chronic hypoxia but it’s effects on these epigenetic changes has not been fully explored. To test our hypothesis we used in-vitro model; Wild type C57B6 male mice (WT) and transgenic males with an extra copy of human hEC-SOD (TG). Studied animals were housed in hypoxia (10% O2) for 21 days. Right ventricular tissue was studied for cardiac fibrosis markers using RT-PCR and Western Blot analyses. Primary C57BL6 mouse cardiac fibroblast tissue culture was used to study in-vitro model, the downstream effects of RASSF-1 expression and methylation and its relation to ERK1/2. Our findings showed a significant increases in cardiac fibrosis markers: Collagen 1, Alpha Smooth Muscle Actin (ASMA) and SNAIL, in the WT hypoxic animals  as compared to the TG hypoxic group (p< 0.05). Expression of DNA methylation enzymes (DNMT 1&3b)  was significantly increased  in the WT hypoxic mice as compared to the hypoxic TG mice (p<0.001). RASSF1A expression was significantly lower and ERK1/2 was significantly higher in hypoxia WT compared to the hypoxic TG group (p<0.05).  Use of SiRNA to block RASSF1A gene expression in murine cardiac fibroblast tissue culture led to increased fibroblast proliferation (p<0.05). Methylation of RASSF1A promoter region was significantly reduced in the TG hypoxic group compared to the WT hypoxic group (0.59 vs 0.75 respectively). Based on our findings, we can speculates that EC-SOD significantly attenuates RASSF1A gene methylation and can alleviate cardiac fibrosis induced by hypoxia.