Background:

Diabetes mellitus type II (T2DM) and metabolic syndrome are associated with an elevated risk of heart failure. Treatment options in particular for heart failure with preserved ejection fraction (HFpEF) are insufficient. Nitro-fatty acids (NO₂-FAs) like nitro-oleic acid (NO₂-OA) are electrophilic molecules with anti-inflammatory and anti-oxidative effects, for which beneficial actions in several mouse models of cardiovascular diseases have been shown. Given that NO₂-OA reduced blood glucose in a genetic mouse model of the metabolic syndrome in a previous study, we sought to investigate whether this effect could improve the left ventricular (LV) function in a diabetic mouse model of HFpEF induced by high fat diet (HFD) combined with the established endothelial nitric oxide synthase (eNOS) inhibitor N_ω-Nitro-D-arginine methyl ester hydrochloride (L-NAME). 

Methods and Results:

Four weeks old C57bl/6N mice were fed with HFD (60% fat, 20% carbohydrates, 20% protein) and L-NAME provided via the drinking water (0.5 g/L) for 11 weeks. Thereafter, mice received either NO₂-OA, oleic acid (OA) or vehicle via mini-osmotic pumps for 4 weeks.

LV morphology and function were determined by echocardiography. To assess direct effects of NO₂-OA on cardiomyocytes, cardiomyocytes derived from human induced pluripotent stem cell line NP0040-8 (UKKi011-A) (iPSC-CM) were treated with 10 mM glucose, 10 nM endothelin-1 and 1 µM hydrocortisone (diabetic condition) and NO₂-OA.

After 4 weeks of treatment with NO₂-OA, glucose tolerance in HFD+L-NAME treated animals was significantly improved compared to vehicle (p=0.05, N=5). LV diastolic function reflected by E/E´ and E´/A´ were significantly impaired after 11 weeks of HFD+L-NAME administration (mean ±SEM, normal chow vs. HFD+L-NAME, E´/A´: 1.12 ± 0.03, N=9 vs. 0.75 ± 0.05, N=27, p<0.0001; E/E´: 28.54 ± 2.17, N=9 vs. 47.86 ± 5.24, N=25, p=0.04) whereas systolic LV function remained unchanged. Diastolic function was significantly improved after 2 and 4 weeks of treatment with NO₂-OA compared to vehicle (mean ± SEM, HFD+L-NAME+NO₂-OA vs. HFD+L-NAME+vehicle, E/E´ 2 weeks: 25.50 ± 5.18, N=5 vs. 53.65 ± 9.62, N=6,  p= 0.05; E/E´ 4 weeks: 35.69 ± 3.31, N=7 vs. 88.99 ± 10.64, N=3, p=0.002; E´/A´ 2 weeks: 1.00 ± 0.12, N=5 vs. 0.57 ± 0.08, N=6, p=0.02). Treatment with OA had no effect on glucose tolerance and cardiac function.

In iPSC-CM hypertrophy, as assessed by measurement of cell area in troponin T-stained cells, was significantly reduced upon NO₂-OA treatment after 48h (diabetic condition vs. diabetic condition+NO₂-OA: 3700 µm²± 205.2 vs. 2967 µm² ± 134.5, p=0.01, N=12), which was accompanied by a less pronounced increase in mRNA and protein expression of natriuretic peptides (diabetic condition vs. diabetic condition+NO₂-OA, rel. ANP mRNA expression: 214.8 ± 21.6 vs. 109.5 ± 31.7, p=0.01, N=10; rel. BNP mRNA expression: 513.2 ± 93.3 vs. 159.5 ± 57,5, p=0.005, N=10; rel. ANP protein expression: 2.45 ± 0.03, N=7 vs. 1.51 ± 0.16, N=8, p=0.04).

Conclusion:

Diastolic LV dysfunction induced by cardiometabolic disease was alleviated by NO₂-OA application. Further experiments are ongoing to expand these findings and to elucidate underlying mechanisms.