Effects of a single bout of strenuous exercise on platelet activation in female ApoE/LDLR-/- mice.

Strenuous physical exercise leads to platelet activation that is normally counterbalanced by the production of endothelium-derived anti-platelet mediators, including prostacyclin (PGI2) and nitric oxide (NO). However, in the case of endothelial dysfunction, e.g. in atherosclerosis, there exists an increased risk for intravascular thrombosis during exercise that might be due to an impairment in endothelial anti-platelet mechanisms. In the present work, we evaluated platelet activation at rest and following a single bout of strenuous treadmill exercise in female ApoE/LDLR-/- mice with early (3-month-old) and advanced (7-month-old) atherosclerosis compared to female age-matched WT mice. In sedentary and post-exercise groups of animals, we analyzed TXB2 generation and the expression of platelet activation markers in the whole blood ex vivo assay. We also measured pre- and post-exercise plasma concentration of 6-keto-PGF1α, nitrite/nitrate, lipid profile, and blood cell count. Sedentary 3- and 7-month-old ApoE/LDLR-/- mice displayed significantly higher activation of platelets compared to age-matched wild-type (WT) mice, as evidenced by increased TXB2 production, expression of P-selectin, and activation of GPIIb/IIIa receptors, as well as increased fibrinogen and von Willebrand factor (vWf) binding. Interestingly, in ApoE/LDLR-/- but not in WT mice, strenuous exercise partially inhibited TXB2 production, the expression of activated GPIIb/IIIa receptors, and fibrinogen binding, with no effect on the P-selectin expression and vWf binding. Post-exercise down-regulation of the activated GPIIb/IIIa receptor expression and fibrinogen binding was not significantly different between 3- and 7-month-old ApoE/LDLR-/- mice; however, only 7-month-old ApoE/LDLR-/- mice showed lower TXB2 production after exercise. In female 4-6-month-old ApoE/LDLR-/- but not in WT mice, an elevated pre- and post-exercise plasma concentration of 6-keto-PGF1α was observed. In turn, the pre- and post-exercise plasma concentrations of nitrite (NO2-) and nitrate (NO3-) were decreased in ApoE/LDLR-/- as compared to that in age-matched WT mice. In conclusion, we demonstrated overactivation of platelets in ApoE/LDLR-/- as compared to WT mice. However, platelet activation in ApoE/LDLR-/- mice was not further increased by strenuous exercise, but was instead attenuated, a phenomenon not observed in WT mice. This phenomenon could be linked to compensatory up-regulation of PGI2-dependent anti-platelet mechanisms in ApoE/LDLR-/- mice.

Haematological parameters, serum lipid profile, liver function and fatty acid profile of broiler chickens fed on diets supplemented with pomegranate seed oil and linseed oil.

The objective of the present study was to determine effect of pomegranate seed oil (PSO) and linseed oil (LO) on haematological parameters, serum lipid profile and liver enzymes as well as fatty acids profile of adipose tissue in broilers. Broilers (n = 400) were fed on diets containing graded PSO levels (0.0%, 0.5%, 1.0%, 1.5%) with or without 2% LO. After 6 weeks of feeding, 6 male broilers from each group were slaughtered and abdominal fat, liver and blood samples were collected. Mixtures of pomegranate seed oil (0.5%, 1%) with linseed oil increased white blood cell level in broilers. Total cholesterol was elevated after LO supplementation whereas administration of PSO (1.5%) significantly decreased this parameter. PSO administration caused c9,t11 conjugated linoleic acid (CLA) concentration-dependent deposition in adipose tissue. By LO addition α-linolenic acid (ALA) content was enhanced, decreasing the n-6/n-3 ratio. PSO and ALA also affected oleic acid proportion in adipose tissue. Neither pomegranate seed oil nor linseed oil had any effect on liver parameters. Pomegranate seed oil had no negative effects on broiler health status and can be considered as a functional poultry meat component.

Identification of a biochemical marker for endothelial dysfunction using Raman spectroscopy.

In the present work, we propose the spectroscopic approach to identify biochemical alterations in endothelial dysfunction. The method is based on the quantification of the ratio of phenylalanine (Phe) to tyrosine (Tyr) contents in the endothelium. The synthesis of Tyr from Phe requires the presence of tetrahydrobiopterin (BH4) as a cofactor of phenylalanine hydroxylase (PAH). Limitation of BH4 availability in the endothelium is a hallmark endothelial nitric oxide synthase (eNOS) dysfunction that may also lead to PAH dysfunction and a fall in Tyr contents. Using Raman spectra, the ratio of marker bands of Tyr to Phe was calculated and the pathological state of the endothelium was detected. We provide evidence that Phe/Tyr ratio analysis by Raman spectroscopy discriminate endothelial dysfunction in ApoE/LDLR(-/-) mice as compared to control mice.

Exercise capacity and cardiac hemodynamic response in female ApoE/LDLR(-/-) mice: a paradox of preserved V'O2max and exercise capacity despite coronary atherosclerosis.

We assessed exercise performance, coronary blood flow and cardiac reserve of female ApoE/LDLR(-/-) mice with advanced atherosclerosis compared with age-matched, wild-type C57BL6/J mice. Exercise capacity was assessed as whole body maximal oxygen consumption (V'O2max), maximum running velocity (vmax) and maximum distance (DISTmax) during treadmill exercise. Cardiac systolic and diastolic function in basal conditions and in response to dobutamine (mimicking exercise-induced cardiac stress) were assessed by Magnetic Resonance Imaging (MRI) in vivo. Function of coronary circulation was assessed in isolated perfused hearts. In female ApoE/LDLR(-/-) mice V'O2max, vmax and DISTmax were not impaired as compared with C57BL6/J mice. Cardiac function at rest and systolic and diastolic cardiac reserve were also preserved in female ApoE/LDLR(-/-) mice as evidenced by preserved fractional area change and similar fall in systolic and end diastolic area after dobutamine. Moreover, endothelium-dependent responses of coronary circulation induced by bradykinin (Bk) and acetylcholine (ACh) were preserved, while endothelium-independent responses induced by NO-donors were augmented in female ApoE/LDLR(-/-) mice. Basal COX-2-dependent production of 6-keto-PGF1α was increased. Concluding, we suggest that robust compensatory mechanisms in coronary circulation involving PGI2- and NO-pathways may efficiently counterbalance coronary atherosclerosis-induced impairment in V'O2max and exercise capacity.

Effects of Low Carbohydrate High Protein (LCHP) diet on atherosclerotic plaque phenotype in ApoE/LDLR-/- mice: FT-IR and Raman imaging.

Low Carbohydrate High Protein (LCHP) diet displays pro-atherogenic effects, however, the exact mechanisms involved are still unclear. Here, with the use of vibrational imaging, such as Fourier transform infrared (FT-IR) and Raman (RS) spectroscopies, we characterize biochemical content of plaques in Brachiocephalic Arteries (BCA) from ApoE/LDLR(-/-) mice fed LCHP diet as compared to control, recomended by American Institute of Nutrition, AIN diet. FT-IR images were taken from 6-10 sections of BCA from each mice and were complemented with RS measurements with higher spatial resolution of chosen areas of plaque sections. In aortic plaques from LCHP fed ApoE/LDLR(-/-) mice, the content of cholesterol and cholesterol esters was increased, while that of proteins was decreased as evidenced by global FT-IR analysis. High resolution imaging by RS identified necrotic core/foam cells, lipids (including cholesterol crystals), calcium mineralization and fibrous cap. The decreased relative thickness of the outer fibrous cap and the presence of buried caps were prominent features of the plaques in ApoE/LDLR(-/-) mice fed LCHP diet. In conclusion, FT-IR and Raman-based imaging provided a complementary insight into the biochemical composition of the plaque suggesting that LCHP diet increased plaque cholesterol and cholesterol esters contents of atherosclerotic plaque, supporting the cholesterol-driven pathogenesis of LCHP-induced atherogenesis.

Characterisation of Atherogenic Effects of Low Carbohydrate, High Protein Diet (LCHP) in ApoE/LDLR-/- Mice.

INTRODUCTION: Low Carbohydrate High Protein diet represents a popular strategy to achieve weight loss. OBJECTIVE: The aim of this study was to characterize effects of low carbohydrate, high protein diet (LCHP) on atherosclerotic plaque development in brachiocephalic artery (BCA) in apoE/LDLR-/- mice and to elucidate mechanisms of proatherogenic effects of LCHP diet. MATERIALS AND METHODS: Atherosclerosis plaques in brachiocephalic artery (BCA) as well as in aortic roots, lipoprotein profile, inflammation biomarkers, expression of SREBP-1 in the liver as well as mortality were analyzed in Control diet (AIN-93G) or LCHP (Low Carbohydrate High Protein) diet fed mice. RESULTS: Area of atherosclerotic plaques in aortic roots or BCA from LCHP diet fed mice was substantially increased as compared to mice fed control diet and was characterized by increased lipids and cholesterol contents (ORO staining, FT-IR analysis), increased macrophage infiltration (MOMA-2) and activity of MMPs (zymography). Pro-atherogenic phenotype of LCHP fed apoE/LDLR-/- mice was associated with increased plasma total cholesterol concentration, and in LDL and VLDL fractions, increased TG contents in VLDL, and a modest increase in plasma urea. LCHP diet increased SCD-1 index, activated SREBP-1 transcription factor in the liver and triggered acute phase response as evidence by an increased plasma concentration of haptoglobin, CRP or AGP. Finally, in long-term experiment survival of apoE/LDLR-/- mice fed LCHP diet was substantially reduced as compared to their counterparts fed control diet suggesting overall detrimental effects of LCHP diet on health. CONCLUSIONS: The pro-atherogenic effect of LCHP diet in apoE/LDLR-/- mice is associated with profound increase in LDL and VLDL cholesterol, VLDL triglicerides, liver SREBP-1 upregulation, and systemic inflammation.

Effects of margarine supplemented with t10c12 and C9T11 CLA on atherosclerosis and steatosis in apoE/LDLR -/- mice.

OBJECTIVES: The objective of this study was to evaluate functional effects of margarine supplemented with individual CLA isomers trans-10, cis-12 and cis-9, trans-11 in apoE/LDLR -/- mice. DESIGN: In LONG experiment (LONG), two-month old mice with no atherosclerosis were assigned to experimental groups and fed for the next 4 months. In SHORT experiment (SHORT), four-month old mice, with pre-established atherosclerosis, were assigned to experimental groups and fed for the next 2 months. The experimental diets were: AIN-93G (margarine), AIN-93G + 0.5% trans-10, cis-12 CLA (t10c12), and AIN-93G + 0.5% cis-9, trans-11 CLA (c9t11). RESULTS: In both experiments (LONG and SHORT), liver weight was significantly (P<0.05) increased in mice fed t10c12 CLA. Hepatic steatosis was found in animals fed t10c12 diet and no signs of the steatosis was observed in mice fed c9t11 CLA. Dietary treatments with t10c12 CLA significantly increased total plasma cholesterol and plasma triacylglycerols. There were no isomer-specific effects of CLA isomers on area of atherosclerotic plaque in aortic root. CONCLUSION: In conclusion, t10c12 CLA significantly increased liver weight in mice in LONG and SHORT experiments. Our results do not support the notion that CLA isomer supplementation to the margarine possess anti-atheroclerotic effect. Therefore, no isomer-specific effects of CLA on development of atherosclerosis were observed.

Hypercholesterolemia does not alter endothelial function in spontaneously hypertensive rats.

In humans, hypercholesterolemia and hypertension are associated with endothelial dysfunction. Here, we assess whether hypercholesterolemia induces endothelial dysfunction in rats with pre-existing hypertension. Spontaneously hypertensive rats (SHR) and normotensive controls (WKY) were fed with a high-cholesterol diet for 12 weeks, and endothelial function was assessed in isolated thoracic aortic rings. In SHR and WKY rats, the hypercholesterolemic diet resulted in the elevation of total cholesterol and low-density lipoprotein levels by approximately 2.5- and 4.5-fold, respectively. However, in aorta, the basal nitric oxide (NO) production--as assessed by the magnitude of L-NG-nitroarginine methyl ester-induced vasoconstriction as well as the NO-dependent relaxation induced by acetylcholine or histamine--were not diminished either in SHR or in WKY rats fed with the hypercholesterolemic diet. Interestingly, prostacyclin (PGI2) production in aortic rings from SHR rats was higher than in the aorta from WKY rats. However, the hypercholesterolemic diet had no further effects on PGI2 production in the aorta either of SHR or WKY rats. The monocyte chemoattractant protein 1 level in plasma was slightly elevated in SHR and WKY rats fed with the hypercholesterolemic diet compared with their normocholesterolemic counterparts. In summary, even in the presence of pre-existing hypertension, hypercholesterolemia fails to modify NO-dependent and PGI2-dependent endothelial function in SHR rats; it also does not induce a robust inflammatory response. Both are prerequisites for the development of atherosclerosis.