Gut Dysbiosis and Barrier Disruption Are Associated with Diastolic Dysfunction in a Novel Mouse Model of Heart Failure with Preserved Ejection Fraction

Abstract

Gut dysbiosis has recently been associated with the severity of the complex syndrome of heart failure (HF) in patients. The mechanisms of action involving gut microbes and immune responses are starting to be characterized in experimental models of HF. More than 50% of HF patients present with HF with preserved ejection fraction (HFpEF), characterized by impaired relaxation (diastolic function) and preserved contractility (systolic function), which do not respond to any of the available treatments for HF. Obesity and hypertension are major risk factors for HFpEF, and have been independently associated with gut dysbiosis, and modulation of the microbiota in experimental models of these risk factors has revealed mechanistic insights. However, whether and how the microbiota contributes to HFpEF is unknown. Recently, a novel mouse model of HFpEF induced by a combination of high-fat diet (HFD) and L-NAME, to mimic obesity and hypertension, respectively. We hypothesize that specific microbiota and derived metabolites, are altered in HFpEF and contribute to systemic inflammation and diastolic dysfunction. C57/BL6 WT mice were fed HFD and L-NAME (H/L) to induce HFpEF. We used invasive hemodynamics to determine diastolic and systolic function, and 16S rRNA sequencing in fecal samples to characterize gut bacteria and perform metabolic pathway analysis. The heart, the intestine and plasma were collected and evaluated for indicators of cardiac remodeling and gut permeability using flow cytometry and qPCR. As expected, H/L triggered diastolic dysfunction at 5 weeks. At this time, H/L intestinal epithelial cells (iECs) showed downregulation in gene expression of the junctional markers cadherin-1, ZO-1 and occludin, suggesting the induction of leaky gut. We also found increased concentration of proinflammatory molecules in serum (e.g. IL-6, IL-17 and CXCL1). Strikingly, H/L treatment resulted in gut dysbiosis in H/L mice compared to those that received standard diet (STD). The main phyla analysis showed a higher Firmicutes/Bacteroidetes ratio in H/L, which has been associated with obesity, compared to STD. At the family level, Ruminococcaceae, also linked to obesity in humans, showed elevated relative abundance in H/L group compared to STD, and Rikenellaceae and Christensenellaceae significantly decreased their numbers. Interestingly, gut bacteria from H/L-treated mice revealed an enrichment of metabolic pathways related to synthesis and degradation of fatty acids. We studied the gene expression of the main short-chain fatty acid (SCFA) receptors, associated with improvement in cardiovascular health. FFAR2 (GPR43) expression did not change in the heart neither the gut, but strikingly, FFAR3 (GPR41) expression was significantly upregulated in the heart in the H/L group compared to STD, but not in iECs. In conclusion, we demonstrated that a novel mouse model of HFpEF induced gut dysbiosis and alterations in gut permeability, and identified a dysregulation of fatty acid synthesis and degradation metabolic pathways. Mechanistic studies are needed to see the role of fatty acids and its receptors in cardiac inflammation during diastolic dysfunction.