Despite intense interest, the exact mechanisms by which SGLT2 inhibition provides cardiac protection remains unclear. Herein we demonstrate that despite RV hypertension, the SGLT2i dapagliflozin
1- reduced RV mass
2- reduced RV and LV water content
3- improved proteins involved in nutrient sensing, contractility, and cellular homeostasis in a model of stable, compensated RV hypertrophy induced by pulmonary artery banding.
These findings suggest that a possible primary mechanism of cardiac protection involves the osmotic diuresis and reduced intra/extracellular water content, with secondary beneficial effects upon important molecular remodeling parameters.
With multiple therapies now available to treat left ventricular remodeling, attention has focused upon the RV. Importantly, RVH is an independent predictor of outcomes in a range of diseases . Like the LV, in response to a pathologic stimulus the RV undergoes a robust remodeling response, characterized by alterations in matrix and cellular composition, resulting ultimately in RV fibrosis, RV hypertrophy, along with changes in metabolism and function. However, unlike the LV, therapies such as renin angiotensin system inhibition (RASi), which have been associated with the prevention of LVH and HF demonstrate different responses in the RV and RAS activation (and inhibition) is not as efficient in inducing or protecting against RVF. As a result, novel therapies are needed to address this limitation.
SGLT2i offer promise in this regard, being shown to improve renal and incident and prevalent HF outcomes, with a variety of mechanisms proposed. We have previously shown that the SGLT2i empagliflozin, when compared to “standard of care” reduced LV mass in persons with type 2 diabetes and established atherosclerotic disease, with the effect upon LV mass being independent upon changes in blood pressureFurthermore, we recently assessed extracellular volume (ECV) fraction in a sub study of the EMPA HEART study. We showed that empagliflozin, reduced ECV, with a trend towards reducing intracellular volume fraction or iICV. These findings suggest that part of the direct effect of SGLT2i in vivo upon LV mass regression is mediated by changes in extracellular, and possibly intracellular water content. In the current study, we showed that LV water content was increased in PAB + vehicle animals when compared to sham + vehicle, and that dapagliflozin reduced LV water content, thus supporting our human studies.
As it relates to the RV, experimental data using the monocrotaline model of pulmonary hypertension has demonstrated a direct impact of the SGLT2i empagliflozin upon right ventricular hypertrophy. However, the aforementioned study did not assess whether the reduction in RV mass was due to a change in myocyte size and mass, or whether it was related to changes in intra and extracellular volume fraction and water content.
Using the highly selective SGLT2i dapagliflozin, we observed two findings. The first was a reduction in RVH independent of changes in blood pressure, however the reduction in RV mass was primarily driven by a reduction in water content, as opposed to changes in myocyte hypertrophy assessed by cross sectional area. Secondly, these changes were associated with normalization of key proteins involved in nutrient sensing, contractility and cellular homeostasis, such as AMPKinase. Furthermore, in keeping with our previous studies in non-diabetic models assessing LV remodeling, we did not see a reduction in collagen type I in dapagliflozin treated PAB animals. The reduction in water content in the RV parallels the LV finding seen in the EMPA HEART LV ECV substudy. How this translates into the global prevention and treatment of RVF unfortunately remains unclear.
Postulated mechanisms for the change in RV and LV water content include the osmotic diuresis caused by SGLT2i, demonstrated by the increase in urinary sodium and glucose. We have previously demonstrated that SGLT2i results in an early natriuresis, but longer therapy likely results in compensatory distal tubule sodium reabsorption, ameliorating the natriuresis instead promoting osmotic diuresis. Other mechanisms to account for the cardioprotective properties of SGLT2i involve improved myocardial energetics because of either altered substrate delivery. Here we demonstrate that the protein expression of the energy sensor AMPK was reduced by PAB, and normalised by dapagliflozin. This finding extends the observations of Hawley et al which demonstrated that the SGLT2i canagliflozin, but not dapagliflozin or empagliflozin improved AMPK activity in HEK-293 cells, mouse embryonic fibroblasts as well as mice hepatocytes. The “thrifty substrate” hypothesis has been suggested to account in part for the protective effects of SGLT2i. Whilst we did not assess for metabolism markers in the current study, previous work by our group has failed to demonstrate significant changes in gene expression in a non-diabetic rodent model of HFpEF, despite SGLT2i improving cardiac function and normalizing LV mass, supporting the concept that metabolic changes may be a secondary effect of SGLT2i and not the primary mode of action of the drug to improve cardiac outcomes. Packer has recently suggested that glucose lowering drugs may impact autophagy secondary to changes in AMPK and sirtuin activity. Of note, our study identified a reduction in the LC3B I/II ratio, which was prevented by dapagliflozin therapy. The impact of improved energy biosensing and autophagic flux along with a reduction in oxidative stress is suggested to restore mitochondrial homeostasis and prevent myocardial dysfunction and the development of pathological RVH. Whilst the current study did not assess mitochondrial function, the improved energy biosensing and autophagic flux in a non-diabetic model with RVH, as opposed to the diabetic heart suggests this may be a common mechanism to explain the beneficial effects of SGLT2i regardless of diabetic status.
Abnormal calcium handling is a hallmark of ventricular remodeling, with downregulation of proteins involved in calcium handling, such as the Sarcoplasmic calcium uptake ATPASE, SERCA 2A. Here in we found that PAB reduced SERCA 2A protein content. Treatment with dapagliflozin improved SERCA 2A content. Improved SERCA levels is associated with improved cardiac function and remains a novel finding. The mechanisms leading to normal SERCA 2A expression remain unclear and require further investigation, but our data supports the concept that SGLT2i prevents the downregulation of SERCA, as opposed to directly modifying transcriptional activity and increasing protein expression of SERCA.
Our study was not without limitations. First, our PAB model recapitulated some, but not all aspects of RV remodeling and animals were in the compensated phase of remodeling, as evidence by preserved cardiac function and RV pressures being <2/3 systemic pressures. However, type 2 PH, the commonest cause of PH typically demonstrates only modest elevations of RVESP, thus making the finding of reduced RV mass with dapagliflozin potentially more clinically relevant. Secondly, dapagliflozin was initiated soon after PAB, but the effects of SGLT2 inhibitors on cardiac remodeling may differ depending on timing of administration. Additional studies where these drugs are initiated at different times during the remodeling process will help to better understand whether the cardiac features can be altered late after the onset of RVF. Thirdly, SGLT2i has been shown to increase hematocrit secondary to increased erythropoietin production, which may result in improved oxygen delivery. We did not observe a change in Hct or Hb levels, suggesting that this is not the mediator of the mechanisms behind RV mass regression in PAB. Whether the lack of change in Hct in response to PAB or dapagliflozin impacts the relevance of these findings in humans and other models is unknown.
Finally, the exact mechanisms buy which dapagliflozin reduced RVH remains unclear. We have previously shown that empagliflozin reduced ECV in persons with T2DM and ASCVD, which accounted for part of the reduction in LV mass observed in the EMPA HEART trial. Whether the same impact was observed in the RV was unable to be assessed in that study. Further studies using CMR focussing on the RV to assess ECV and cardiomyocyte compartments[33,34] is required to better understand the observed reduction in RVH, in persons both with, and without type 2 DM.
In conclusion, the SGLT2 inhibitor, dapagliflozin caused an osmotic diuresis and reduction in RV mass without significant changes in blood pressure, along with improvements in key nutrient, autophagic and contractility proteins within the RV, at an early compensated phase of RV remodeling. These finding suggest dapagliflozin may have a benefit for disease characterized by RVH induced by RV pressure loading.