In addition to the circulatory renin-angiotensin-aldosterone system (RAAS), there is now a significant body of evidence supporting the concept of a “local tissue or cellular RAAS” that has important roles in the pathology of cardiovascular diseases [
Aldosterone activates its mineralocorticoid receptor (MR) in the nondiabetic heart and can cause structural and electrical remodelling, fibrosis, oxidative stress, inflammation, and arrhythmias [
It is well established that signaling network alterations in diabetes are such that potential therapies will need to be tailored for this pathological state [
ACEI and/or ARBs do not completely block end-organ damage in diabetes and/or hypertension, and clinical trials of ACEI and ARBs in combination have generally shown that they do not offer added benefits but rather lead to greater adverse effects such as electrolyte imbalance and renal complications [
17-week-old male Wistar rats were divided into 2 groups (
Diabetes was induced by a single intraperitoneal injection of 55 mg/kg body weight STZ. Basal glucose levels were determined prior to STZ injection and 48 h after STZ injection. Rats with a blood glucose concentration above 250 mg/dL were declared diabetic and any not meeting this criterion were excluded from the study. The animals’ diabetic state was re-assessed after 4 weeks just before sacrificing the animals.
Rats were anesthetized with Intraval Sodium (40 mg/kg body weight), and hearts were rapidly removed after intravenous heparinization (1000 U/kg body weight). The excised hearts were immediately mounted on the Langendorff perfusion assembly (Hugo Sachs Electronics, Freiburg, Germany) and were perfused initially with a constant pressure perfusion of 50 mmHg with the oxygenated (95% O2 + 5% CO2) Krebs-Henselit buffer (37°C) of the following composition (in mM): NaCl 117; KCl 4.39; CaCl2 2.5; NaHCO3 20.0; KH2PO4 1.21; MgCl2·6H2O 1.2; glucose 12.0; osmolarity 300 mOsm/L, pH 7.35. A water-filled balloon was introduced into the left ventricle and connected to a Statham pressure transducer (P23Db) and balloon volume was adjusted to give the baseline end-diastolic pressure of 5 mmHg. Left ventricular developed pressure (
Hearts removed from animals were perfused for 30 min and then subjected to 30 min of global ischemia (I) followed by a period of 30 min of reperfusion (
Data are presented as mean ± SEM of “
Diabetes induction by STZ injection led to a significant increase in blood glucose concentration. Hyperglycaemia persisted throughout the study period and was
The effects of various acute drug treatments (RU, Capt, Los, Capt + Los, Capt + RU, and Los + RU, Capt + Los + RU) administered during perfusion (before ischemia) or during reperfusion (after ischemia) on recovery of each cardiac function parameter following I/R in normal and diabetic hearts were recorded. The percent (%) recovery in
The effect of drugs given before ischemia on % recovery in cardiac function versus reperfusion time for (a)
The effect of drugs given after ischemia on % recovery in cardiac function versus reperfusion time for (a)
The effect of drugs given before ischemia on % recovery in cardiac function versus reperfusion time for (a)
The effect of drugs given after ischemia on % recovery in cardiac function versus reperfusion time for (a)
In untreated controls, % recovery (%
A comparison of the % change in left ventricular developed pressure (
Control (P)
Control (R)
Diabetes (P)
Diabetes (R)
Recovery in
%
A comparison of the % change in left ventricular end-diastolic pressure (LVEDP) in control and diabetic hearts following acute treatment with various drug regimens. Drugs were given to control hearts during perfusion (a) or reperfusion (b) and to diabetic hearts during perfusion (c) or reperfusion (d). The percent change in parameter is calculated relative to the % recovery seen in the respective nondiabetic or diabetic controls. RU: RU28318; Capt: Captopril; Los: Losartan; Triple: RU28318 + Captopril + Losartan.
Control (P)
Control (R)
Diabetes (P)
Diabetes (R)
In diabetes, LVEDP was significantly (
In untreated controls, % recovery of
A comparison of the % change in positive derivative of pressure (
Control (P)
Control (R)
Diabetes (P)
Diabetes (R)
Diabetes (8 ± 2%) led to about a 6-fold reduction in %
In untreated controls, % recovery (%
A comparison of the % change in negative derivative of pressure (
Control (P)
Control (R)
Diabetes (P)
Diabetes (R)
Diabetes (11 ± 3%) led to a about a 3.4-fold reduction in %
In untreated controls, % recovery (%
A comparison of the % change in coronary flow (CF) in control and diabetic hearts following acute treatment with various drug regimens. Drugs were given to control hearts during perfusion (a) or reperfusion (b) and to diabetic hearts during perfusion (c) or reperfusion (d). The percent change in parameter is calculated relative to the % recovery seen in the respective nondiabetic or diabetic controls. RU: RU28318; Capt: Captopril; Los: Losartan; Triple: RU28318 + Captopril + Losartan.
Control (P)
Control (R)
Diabetes (P)
Diabetes (R)
Diabetes (12 ± 3%) led to about a 3.8-fold reduction in %
A major goal of this study was to identify the optimal treatment strategy of RAAS blockade to prevent or treat ischemia-reperfusion injury in normal and diabetic hearts [
This study showed that treatment with RU was generally better in preventing or reversing ischemia-induced cardiac dysfunction in normal hearts compared to treatment with an ACEI (Capt) or ARB (Los) alone. In the case of diabetic hearts, RU was generally similarly effective as Capt or Los treatment. Also, dual therapies involving RU were similarly effective as Capt + Los therapy, whereas Triple combination was generally equal to or the most effective strategy in preventing or reversing ischemia-induced cardiac dysfunction in normal or diabetic hearts. For example, irrespective of whether drugs were administered before or after ischemia, combination therapies appeared more effective, whereby triple > double > single therapy in most of the study scenarios (i.e., 18 out of the 20 experimental scenarios of looking at 5 different cardiac parameters studied for both normal and diabetic hearts for when drugs were given either before or after ischemia). In contrast to nondiabetic hearts, Capt + Los dual therapy was the most effective therapy (alongside Triple therapy) in diabetic hearts which is consistent with the known diabetes-induced overactivity of the ACE/Ang II/AT1 signaling cascade [
In this study, looking at an MR antagonist in diabetic hearts, we show that indeed the diabetic pathology leads to altered cardiac response to treatments with RU as well as ARBs and ACEI. In general, with the exception of
Although RU-mediated improvement in cardiac function generally appears to be as good as ARBs and ACEIs, differences exist between RU and other drugs as to effectiveness when given before or after ischemia. In nondiabetic hearts, RU was the best or one of the best single agents when given before or after ischemia, whereas in diabetes, it generally provided minimal or least benefit when given after ischemia with the exception of
Our study also showed that irrespective of whether drugs were administered before or after ischemia, their effectiveness in recovering cardiac function appeared to be time dependent; typically, drugs rapidly improved cardiac function within the first 10 min of reperfusion followed by a period of steady improvement thereafter up to the 30 min study period (Figures
The possible mechanisms by which RU may be acting beneficially in normal and diabetic hearts subjected to I/R are not known but these may include blockade of aldosterone-induced oxidative stress, endothelial dysfunction, and inflammation in the heart and in the coronary vasculature that generally contribute to abnormal calcium homeostasis and cardiac dysfunction. Thus, RU-mediated blockade of these important processes likely also improves calcium handling and overload in hearts subjected to I/R. This assertion is supported by previous findings that aldosterone/MR activation induces cardiomyocyte ionic remodelling by modulating potassium and L-type calcium channel activity [
Aldosterone is also known to induce increased Na+/H+ exchanger-1 (NHE-1) activity via transactivation of the epidermal growth factor receptor (EGFR) and subsequent reactive oxygen species (ROS) formation which is thought to be an important signaling cascade in the genesis of many cardiac pathologies [
Whether the beneficial effects of RU may be via antagonism of aldosterone-mediated effects in cardiac muscle and/or vasculature is not clear but recent studies suggest the involvement of both. It appears that cardiomyocyte aldosterone/MR participates in the crosstalk between cardiomyocytes and coronary blood vessels, such as increased aldosterone synthesis by the cardiomyocytes resulting in coronary dysfunction [
In diabetic patients, in addition to cardiac dysfunction, there is also significant risk of renal damage where Ang II blockade is contraindicated as it will lead to attenuation of Ang II-driven glomerular filtration rate (GFR) and renal shutdown particularly in patients with renal artery stenosis. However, RU which does not affect GFR may be particularly useful for such patients. Further, addition of RU as a combination strategy with ACEI and ARBs may allow for a reduction in the dose of the latter agents such that they have minimal disruption on GFR and electrolyte balance whilst retaining their ability to reduce diabetes-induced proteinuria and their beneficial effects on cardiac function as highlighted in the present study.
Importantly, MR antagonists when combined with ACEI but not ARBs have shown significant reduction in total mortality in patients with CHF [
In diabetic patients, therapies involving aldosterone/MR antagonists may have other additional beneficial effects beyond the cardiovascular system. They may oppose aldosterone-mediated detrimental effects on structural and functional integrity of the pancreatic [beta]-cell resulting from islet cell inflammation and oxidative stress as well as aldosterone-induced insulin resistance [
If our findings reported here are reproduced in clinical studies, our study may have important clinical implications in the way these drugs should be administered in cardiac dysfunction. Firstly, our data implies that in normal patients, RU alone could be an effective therapy for prevention of cardiac dysfunction because as a single agent, it yields the best improvements in cardiac function when given before ischemia. Furthermore, for postischemic injury, although RU appears to be an effective therapy, ACEIs and/or ARBs appear to be the drugs of choice for diabetics as they yielded the best improvements in cardiac function when administered after ischemia. We also suggest that MR antagonists, since they act through a non-ACE/Ang II/AT1R pathway, may represent a novel class of RAAS inhibitor that potentially could overcome the limitations observed with the ACEI and ARB combinations of RAAS inhibitors.
Our study by selecting to administer drugs acutely in isolated hearts is advantageous in that it examines the effects of these therapeutic agents directly on the heart and avoids noncardiac contributions of these agents. Furthermore, this study highlights that in addition to the benefits observed by MR antagonists when administered systemically, these agents can also be beneficial when administered locally. Thus, our data implies that in the clinic RU treatment locally might be considered as an effective therapy or preventative measure in cardiac I/R injury for susceptible patients and possibly also preoperatively for patients undergoing aortic cross-clamping or other cardiac surgeries such as cardiopulmonary bypass or coronary artery bypass grafting. Our study also suggests that optimal usage of drug(s) alone or in combination may require their selection based on several criteria including their relative benefit in the normal versus pathological state, whether being considered for prevention or treatment strategy, on the specific cardiac parameter that might need to be improved (e.g., diastolic function in diabetes) and on the optimal rate of cardiac function recovery required for a given condition.
The authors declare that there is no conflict of interests.
This study was supported by Grant no. MR 04/09 from the Kuwait University.