Gender differences in the expression and cellular localization of lipin 1 in the hearts of fructose-fed rats.

To give new insight to alterations of cardiac lipid metabolism accompanied by a fructose-rich diet (FRD), rats of both sexes were exposed to 10 % fructose in drinking water during 9 weeks. The protein level and subcellular localization of the main regulators of cardiac lipid metabolism, such as lipin 1, peroxisome proliferator-activated receptor α (PPARα), peroxisome proliferator-activated receptor γ coactivator-1 α (PGC-1α), carnitine palmitoyltransferase I (CPTI), and CD36 were studied. Caloric intake in fructose-fed rats (FFR) of both sexes was increased. Circulating triacylglyceroles (TAG) and non-esterified fatty acids were increased in male FFR, while females increased visceral adiposity and blood TAG. Total expression of lipin 1 in cardiac cell lysate and its cytosolic and microsomal level were increased in the hearts of male FFR. PPARα and PGC-1α content were decreased in the nuclear extract. In addition, cardiac deposition of TAG in male FFR was elevated, as well as inhibitory phosphorylation of insulin receptor substrate 1 (IRS-1). In contrast, in female FFR, lipin 1 level was increased in nuclear extract only, while overall CPTI expression and phosphorylation of IRS-1 at serine 307 were decreased. The results of our study suggest that fructose diet causes gender-dependent alterations in cardiac lipid metabolism. Potentially detrimental effects of FRD seem to be limited to male rats. Most of the observed changes might be a consequence of elevated expression and altered localization of lipin 1. Increased inhibitory phosphorylation of IRS-1 is possible link between cardiac lipid metabolism and insulin resistance in FFR.

Does oestradiol attenuate the damaging effects of a fructose-rich diet on cardiac Akt/endothelial nitric oxide synthase signalling?

Fructose-rich diets (FRD) cause cardiac insulin resistance manifested by impairment of Akt/endothelial NO synthase (eNOS) signalling. In contrast, oestradiol (E2) activates this signalling pathway in the heart. To study the ability of E2 to revert the detrimental effect of fructose on cardiac Akt/eNOS, female rats were subjected to a FRD and ovariectomy followed with or without E2 replacement. We also analysed the effects of the FRD and E2 on cardiac extracellular signal-regulated kinase (Erk 1/2) signalling related to their role in cardiac hypertrophy development. Expression of Akt, eNOS and Erk 1/2, as well as regulatory phosphorylations of these molecules were determined. The protein expression of cardiac Akt and eNOS was not affected by the diet or E2 treatment. However, the FRD was accompanied by a decrease in Akt phosphorylation at Ser(473) and Thr(308), and eNOS at Ser(1177), while the phosphorylation of eNOS at Thr(495) was increased. E2 replacement in ovariectomised fructose-fed rats caused a reversion of the diet effect on Akt and eNOS serine phosphorylation, but mostly had no effect on threonine phosphorylation of the molecules. The FRD and E2 treatment did not influence Erk 1/2 expression and phosphorylation and heart mass as well. The data show that E2 selectively suppress the negative effects of a FRD on Akt/eNOS signalling and probably point to the different effects of E2 on kinase/phosphatase pathways responsible for phosphorylation/dephosphorylation of Akt and eNOS. Furthermore, the results suggest that the heart of females in the reproductive period is partially protected against the damaging effects of increasedfructose intake.

Estradiol enhances effects of fructose rich diet on cardiac fatty acid transporter CD36 and triglycerides accumulation.

Fructose rich diet increases hepatic triglycerides production and has deleterious cardiac effects. Estrogens are involved in regulation of lipid metabolism as well, but their effects are cardio beneficial. In order to study effects of fructose rich diet on the main heart fatty acid transporter CD36 and the role of estrogens, we subjected ovariectomized female rats to the standard diet or fructose rich diet, with or without estradiol (E2) replacement. The following parameters were analyzed: feeding behavior, visceral adipose tissue mass, plasma lipids, cardiac CD36 expression, localization and insulin regulation, as well as the profile of cardiac lipids. Results show that fructose rich diet significantly increased plasma triglycerides and decreased plasma free fatty acid (FFA) concentration, while E2 additionally emphasized FFA decrease. The fructose diet increased cardiac plasma membrane content of CD36 in the basal and insulin-stimulated states, and decreased its low density microsomes content. The E2 in fructose-fed rats raised the total cardiac protein content of CD36, its presence in plasma membranes and low density microsomes, and cardiac deposition of triglycerides, as well. Although E2 counteracts fructose in some aspects of lipid metabolism, and separately they have opposite cardiac effects, in combination with fructose rich diet, E2 additionally enhances CD36 presence in plasma membranes of cardiac cells and triglycerides accumulation, which paradoxically might promote deleterious effects of fructose diet on cardiac lipid metabolism. Taken together, the results presented in this work are of high importance for clinical administration of estrogens in females with a history of type 2 diabetes.

Impairment of cardiac insulin signaling in fructose-fed ovariectomized female Wistar rats.

BACKGROUND: Fructose consumption produces deleterious metabolic effects in animal models. The sites of fructose-induced insulin resistance are documented to be the liver, skeletal muscle, and adipose tissue, but effects of fructose-rich diet on cardiac insulin signaling and action were not investigated. PURPOSE AND METHODS: In order to study the potential fructose effects on development of cardiac insulin resistance, we analyzed biochemical parameters relevant for insulin action and phosphorylation of insulin signaling molecules, plasma membrane glucose transporter type 4 (GLUT4) content, and phosphorylation of endothelial nitric oxide synthase (eNOS), in ovariectomized female rats on fructose-enriched diet, in basal and insulin-stimulated conditions. RESULTS: Fructose-fed rats (FFR) had increased content of visceral adipose tissue, but not body weight. Food intake was decreased, while fluid and caloric intake were increased in FFR. Additionally, fructose diet increased plasma insulin, blood triglycerides level, and HOMA index. Stimulation of protein kinase B (Akt) signaling pathway by insulin was reduced in rats on fructose-enriched diet, but effect of fructose on extracellular signal-regulated kinase (Erk 1/2) phosphorylation was not observed. Furthermore, insulin-induced GLUT4 presence in plasma membranes of cardiac cells was decreased by fructose diet, as well as insulin stimulation of eNOS phosphorylation at Ser(1177). CONCLUSION: In summary, these results strongly support our hypothesis that fructose diet-induced changes of plasma lipid profile and insulin sensitivity are accompanied with decrease in cardiac insulin action in ovariectomized female rats.

Interference between insulin and estradiol signaling pathways in the regulation of cardiac eNOS and Na(+)/K(+)-ATPase.

Insulin and estradiol share some of signaling pathways and regulate same target molecules exerting mostly beneficial cardiac effects. In order to study their cardiac interaction, ovariectomized female rats were treated with hormones, separately or simultaneously (20, 30 or 40min before analysis), and the phosphorylations of protein kinase B (Akt), extracellular signal-regulated kinases 1 and 2 (ERK 1/2), endothelial nitric oxide synthase (eNOS) were analyzed, as well as the plasma membrane content of α2 subunit of Na(+)/K(+)-ATPase. Insulin, particularly, and estradiol stimulate Ser(473) Akt phosphorylation. The combined treatment was stimulatory, but less than insulin alone was. The general increase of Thr(308) Akt phosphorylation by insulin was stronger than at Ser(473) and reduced in the presence of estradiol, which also stimulated this phosphorylation given alone. The estradiol induction of ERK 1/2 phosphorylation was inverted to the decrease by the combined treatment, while insulin had no effect. Only insulin increased the plasma membrane content of α2. Estradiol did increase the phosphorylation of eNOS, whereas the insulin effect was controversial. The effect of the combined treatment on target molecules was generally opposite to single hormone treatment. In summary, both hormones exerted an effect on Akt phosphorylation, but only estradiol stimulated ERK 1/2 phosphorylation. The α2 plasma membrane content was increased only by insulin, while estradiol increased eNOS phosphorylation more consistently. Finally, if these hormones were administered together, it seems that they disturb each other in having a full effect on cardiac Akt, ERK 1/2, and downstream effectors, eNOS and Na(+)/K(+)-ATPase.

Role of PI3K/AKT, cPLA2 and ERK1/2 signaling pathways in insulin regulation of vascular smooth muscle cells proliferation.

Vascular smooth muscle cells (VSMCs) respond to arterial wall injury by intimal proliferation and play a key role in atherogenesis by proliferating and migrating excessively in response to repeated injury, such as hypertension and atherosclerosis. In contrast, fully differentiated, quiescent VSMCs allow arterial vasodilatation and vasoconstriction. Exaggerated and uncontrolled VSMCs proliferation appears therefore to be a common feature of both atherosclerosis and hypertension. Phosphorylation/dephosphorylation reactions of enzymes belonging to the family of mitogen-activated protein kinases (MAPKs), phosphatidylinositol 3-kinase (PI3K) and protein kinase B (Akt) play an important role in the transduction of mitogenic signal. We have previously shown that among extracellular signal-regulated protein kinases (ERKs), the 42 and 44 kDa isoforms (ERK1/2) as well as Akt and cytosolic phospholipase 2 (cPLA2) participate in the cellular mitogenic machinery triggered by several VSMCs activators, including insulin (INS). The ability of INS to significantly increase VSMCs proliferation has been demonstrated in several systems, but understanding of the intracellular signal transduction pathways involved is incomplete. Signal transduction pathways involved in regulation of the VSMCs proliferation by INS remains poorly understood. Thus, this review examines recent findings in signaling mechanisms employed by INS in modulating the regulation of proliferation of VSMCs with particular emphasis on PI3K/Akt, cPLA2 and ERK1/2 signaling pathways that have been identified as important mediators of VSMCs hypertrophy and vascular diseases. These findings are critical for understanding the role of INS in vascular biology and hyperinsulinemia.

Impact of estradiol on insulin signaling in the rat heart.

It is well known that variation in the concentration of estrogens affects insulin action. In this study we examine the impact of estradiol (E2) on insulin signaling in the rat heart. Ovariectomized female rats were treated with E2 6 h prior to analysis of basal protein and mRNA content of insulin signaling molecules, and additionally with insulin 30 min before the experiment to delineate E2 effects on phosphorylations and molecular associations relevant for insulin signaling. The results show that E2 decreased insulin receptor (IR) tyrosine phosphorylation, while it did not alter IR protein and mRNA content. E2 administration did not change IR substrate 1 (IRS-1) protein content and tyrosine phosphorylation, while decreased mRNA content and increased its association with the p85 subunit of phosphatidylinositol 3-kinase (PI3K). E2 decreased protein and mRNA content of IR substrate 2 (IRS-2), while did not change IRS-2 tyrosine phosphorylation and IRS-2 association with p85. The increase of IRS-1/p85 is accompanied by increase of p85 protein and mRNA levels, and by stimulation of protein kinase B (Akt) Ser(473) phosphorylation. In contrast, Akt protein and mRNA content were not changed. In summary, although in some aspects cardiac insulin signaling is obviously improved by E2 treatment (increase of p85 mRNA and protein levels, enhancement of IRS-1/p85 association and Ser(473)Akt phosphorylation), the observed decrease of IR tyrosine phosphorylation, IRS-2 protein content, and IRSs mRNA contents, suggest very complex interplay of beneficial and suppressive effects of E2, both genomic and non-genomic, in regulation of heart insulin signaling.