Apolipoprotein A-I mimetic peptide L-4F prevents myocardial and coronary dysfunction in diabetic mice

Abstract

Diabetes is a major health problem associated with adverse cardiovascular outcomes. The apolipoprotein A-I mimetic peptide L-4F is a putative anti-diabetic drug, has antioxidant and anti-inflammatory proprieties and improves endothelial function. In obese mice L-4F increases adiponectin levels, improving insulin sensitivity, and reducing visceral adiposity. We hypothesized that the pleiotropic actions of L-4F can prevent heart and coronary dysfunction in a mouse model of genetically induced Type II diabetes. We treated db/db mice with either L-4F or vehicle for 8 weeks. Trans-thoracic echocardiography was performed; thereafter, isolated hearts were subjected to ischemia/reperfusion (IR). Glucose, insulin, adiponectin, and pro-inflammatory cytokines (IL-1β, TNF-α, MCP-1) were measured in plasma and HO-1, pAMPK, peNOS, iNOS, adiponectin, and superoxide in cardiac tissue. In db/db mice L-4F decreased accumulation of subcutaneous and total fat, and increased insulin sensitivity and adiponectin levels while lowering inflammatory cytokines (P < 0.05). L-4F normalized in vivo left ventricular (LV) function of db/db mice, increasing (P < 0.05) fractional shortening and decreasing (P < 0.05) LV dimensions. In I/R experiments, L-4F prevented coronary microvascular resistance from increasing and LV function from deteriorating in the db/db mice. These changes were associated with increased cardiac expression of HO-1, pAMPK, peNOS, and adiponectin and decreased levels of superoxide and iNOS (P < 0.01). In the present study we showed that L-4F prevented myocardial and coronary functional abnormalities in db/db mice. These effects were associated with stimulation of HO-1 resulting in increased levels of anti-inflammatory, anti-oxidative, and vasodilatatory action through a mechanism involving increased levels of adiponectin, pAMPK, and peNOS.

Fig. 1

Time course of body weight and glucose levels during the 8 weeks of L-4F or vehicle treatment in vehicle-treated wild-type mice, wild-type mice after L-4F treatment, db/db mice, and db/db mice after L-4F treatment mice.

Fig. 2

Upper: Effect of L-4F on blood levels of glucose and insulin in vehicle-treated wild-type mice (control), wild-type mice after L-4F treatment (control L-4F), db/db mice (db), and db/db mice after L-4F treatment (db L-4F) mice. Bottom: Correlation between HOMA index and body weight in the four groups of animals. The results are means ± SEM; *p<0.01 vs control and control L-4F, †p<0.05 vs db L-4F, ‡p<0.05 vs control and control L-4F.

Fig. 3

Effect of L-4F on blood levels of adiponectin and cytokines levels (TNFα, IL1β and MCP1) in vehicle-treated wild-type mice (control), wild-type mice after L-4F treatment (control L-4F), db/db mice (db), and db/db mice after L-4F treatment (db L-4F) mice. The results are means ± SEM; *p<0.01 vs control and control L-4F, †p<0.05 vs db L-4F, ‡p<0.05 vs control and control L-4F.

Fig. 4

Effect of L-4F on cardiac morpho-functional parameters. Panel A: representative M-mode echocardiograms. Panel B: the LV chamber diameters were measured at the end of diastole (LVEDD) and systole (LVESD) and averaged from 3–5 beats. Percent fractional shortening (FS%) was calculated as follows: FS% = (LVEDD − LVESD)/LVEDD × 100. The results are means ± SEM. *p<0.05 vs other groups

Fig. 5

Effect of L-4F on coronary resistance and LV function. Ex vivo hearts from vehicle-treated wild-type mice (control), wild-type mice after L-4F treatment (control L-4F), db/db mice (db), and db/db mice after L-4F treatment (db L-4F) were studied in Langendorff configuration with a protocol of ischemia/reperfusion. Coronary resistance (CR), left ventricular developed pressure (LVDevP), dP/dtmax and dP/dtmin in each stage of ischemia/reperfusion, i.e. baseline (bas), low pressure perfusion (Low P) and reperfusion (Rep) were monitored. The results are means ± SEM. * p<0.05 vs other groups, †p<0.001 vs db mice, ‡ p<0.001 vs other groups

Fig. 6

Cardiac levels of HO-1 and superoxide. Above: Representative immunoblots of HO-1 and HO-2 protein are shown. Bottom: Cardiac HO-1/actin ratio and superoxide levels in hearts from vehicle-treated wild-type mice (control), wild-type mice after L-4F treatment (control+L-4F), db/db mice (db), and db/db mice after L-4F treatment (db L-4F) mice. The results are means ± SEM. * p<0.05 vs other groups

Fig. 7

Cardiac tissue homogenates were subjected to Western blotting for AMPK, pAMPK, eNOS, p-eNOS, iNOS and adiponectin protein determination. The relative representative immunoblots (above) and the relative densitometry are shown in vehicle-treated wild-type mice (control), wild-type mice after L-4F treatment (control+L-4F), db/db mice (db), and db/db mice after L-4F treatment (db L-4F) mice. The results are means ± SEM. *p<0.05 vs control mice, †p<0.01 vs L-4F treated animals, ‡ p<0.05 vs other groups.