Epigenetic regulation of miR-181c-5p in a cardiorenal mouse model with co-occurring thrombotic microangiopathy
Authors: Boen J R A , Kruger D , Dendooven A , Tubeeckx M , Bruyns T , Van Fraeyenhove J , Feyen E , Gevaert A B , Segers V F M , Van Craenenbroeck E M 
Published: October 2022

Abstract

Background
MiR-181c-5p is described to induce heart failure (HF), while its role in renal pathology and healthy mice is rather undetermined. Renal dysfunction is present in 40–60% of HF patients and associated with high morbidity and mortality rate.

Purpose
This study aims to investigate the role of miR-181c-5p in a new mouse model of metabolic cardiorenal disease (CRD). Our hypothesis states a protective effect of miR-181c-5p inhibition on HF development by regulation of Tgfbr1.

Methods
CRD was induced by feeding male C57BL/6J mice (n=20) a high-fat diet (HFD) and L-NAME in drinking water (5g/L) for 6 weeks, angiotensin-II was co-administered via osmotic minipumps (1000ng/kg/min) during the final 2 weeks. Healthy controls (n=16) were given normal chow and drinking water, and underwent sham-surgery. Mice were randomly assigned to weekly injections (40mg/kg) with miR-181c-5p antagomiR (INH) or scrambled control for the duration of the study. We assessed cardiac function (echocardiography, invasive hemodynamics), renal function (plasma creatinine), target expression (RT-qPCR), and histology.

Results
CRD animals showed mild systolic and diastolic cardiac dysfunction compared to healthy controls characterized by reduced dP/dt min (−4795±1164 vs −7728±1693 mmHg/s; p=0.01) and dP/dt max (6222±1069 vs 8706±1739 mmHg/s; p=0.038), and increased tau (9.88±3.09 vs 6.07±0.73 ms; p=0.02) with preserved ejection fraction (45±21 vs 51±8%; p=0.53). Histology shows cardiac fibrosis (2.5±0.3 vs 1.8±0.2% area; p=0.0004) and hypertrophy (0.11±0.03 vs 0.08±0.01g/cm; p=0.005). Renal dysfunction presents with kidney atrophy (0.07±0.006 vs 0.09±0.01g/cm; p=0.02), increased plasma creatinine (21±6 vs 10±5; p=0.01), renal fibrosis (0.26±0.22 vs 0.005±0.21% area; p=0.036) and glomerular abnormalities (glomerulosclerosis, hyperfiltration, mesangial matrix expansion, reduced podocyte number). CRD+INH animals had comparable cardiac phenotype to CRD (p&gt;0.05), except a significantly reduced cardiac output compared to healthy controls (6±3 vs 18±3 μl/s; p=0.035). Their renal phenotype was exacerbated with elevated glomerular damage (26±3 vs 18±9; p=0.04) and significantly increased mortality rate (50%) (Kaplan-Meier p=0.01) compared to healthy controls (0%) or CRD (20%), associated with increased occurrence of tubular atrophy, endothelial swelling and systemic thrombotic microangiopathy (TMA) that manifested in kidney and the heart. RT-qPCR analysis identified Vegf as potential target of miR-181c-5p in kidney and showed significantly reduced levels of Tgfbr1 in cardiac tissue of CRD+INH mice.

Conclusion
This study demonstrates a detrimental effect of miR-181c-5p inhibition on renal function in a CRD mouse model, driven by glomerular damage and TMA through Vegf signaling. Despite identification of Tgfbr1 as potential target of miR-181c-5p in the heart, cardiac function was rather unaffected.

Funding Acknowledgement
Type of funding sources: Public Institution(s). Main funding source(s): University of Antwerp