Fatty acid incorporation in endothelial cells and effects on endothelial nitric oxide synthase

International audienceThe nature of fatty acids provided by the diet as well as plasma lipid metabolism can modify the composition and properties of plasma membrane and thus the activity of membrane proteins. In humans, as well as in experimental models, diabetes is associated with both an alteration in serum lipid profile and a documented endothelial dysfunction. This in vitro study investigated on an immortalized human endothelial cell line (EA.hy 926) the specific effects of several free fatty acids (FFAs) on the composition of cellular membranes and the regulation of endothelial nitric oxide synthase (eNOS). 0.1% of lipid deprived serum was added to the incubation medium with 25 mM glucose in order to study the effects of individual fatty acids: myristic acid, palmitic acid, stearic acid, oleic acid or linoleic acid at 100 microM bound with albumin. The effects of the FFAs on the endothelial nitric oxide synthase were investigated on mRNA level by quantitative PCR, on protein level and Ser1177 phosphorylation by Western blot and on enzymatic activity on living cells using radiolabelled arginine. Free linoleic acid increased the membrane content in n-6 fatty acids (mainly C18: n-6 and its metabolites) with a decrease in saturated and monounsaturated fatty acids. These conditions decreased the basal eNOS activity and reduced the phosphorylation of eNOS-Ser1177 due to activation by histamine. Free palmitic acid enriched the membranes with 16 : 0 with a slight decrease in monounsaturated fatty acids. These conditions increased eNOS activation without increasing Ser1177 phosphorylation upon histamine activation. The addition of the other FFAs also resulted in modifications of membrane composition, which did not to affect eNOS-Ser1177 phosphorylation. Among the fatty acids used, only modification of the membrane composition due to linoleic acid supply disturbed the basal enzymatic activity and Ser1177 phosphorylation of eNOS in a way that limited the role of histamine activation. Linoleic acid might involve the dysfunction of both eNOS basal activity and its phosphorylation status and may then contribute to an impaired vasodilatation in vivo

Depressed mitochondrial transcription factors and oxidative capacity in rat failing cardiac and skeletal muscles

Congestive heart failure (CHF) induces alterations in energy metabolism and mitochondrial function that span cardiac as well as skeletal muscles. Whether these defects originate from altered mitochondrial DNA copy number and/or mitochondrial gene transcription is not known at present, nor are the factors that control mitochondrial capacity in different muscle types completely understood. We used an experimental model of CHF induced by aortic banding in the rat and investigated mitochondrial respiration and enzyme activity of biochemical mitochondrial markers in cardiac, slow and fast skeletal muscles. We quantified mitochondrial DNA (mtDNA), expression of nuclear (COX IV) and mitochondrial (COX I) encoded cytochrome c oxidase subunits as well as nuclear factors involved in mitochondrial biogenesis and in the necessary coordinated interplay between nuclear and mitochondrial genomes in health and CHF. CHF induced a decrease in oxidative capacity and mitochondrial enzyme activities with a parallel decrease in the mRNA level of COX I and IV, but no change in mtDNA content. The expression of the peroxisome proliferator activated receptor gamma co-activator 1α (PGC-1α) gene was downregulated in CHF, as well as nuclear respiratory factor 2 and mitochondrial transcription factor A, which act downstream from PGC-1α. Most interestingly, only the level of PGC-1α expression was strongly correlated with muscle oxidative capacity in cardiac and skeletal muscles, both in healthy and CHF rats. Mitochondrial gene transcription is reduced in CHF, and PGC-1α appears as a potential modulator of muscle oxidative capacity under these experimental conditions

Consequences of changes in BDNF levels on serotonin neurotransmission, 5-HT transporter expression and function: studies in adult mice hippocampus.

In vivo intracerebral microdialysis is an important neurochemical technique that has been applied extensively in genetic and pharmacological studies aimed at investigating the relationship between neurotransmitters. Among the main interests of microdialysis application is the infusion of drugs through the microdialysis probe (reverse dialysis) in awake, freely moving animals. As an example of the relevance of intracerebral microdialysis, this review will focus on our recent neurochemical results showing the impact of Brain-Derived Neurotrophic Factor (BDNF) on serotonergic neurotransmission in basal and stimulated conditions. Indeed, although the elevation of 5-HT outflow induced by chronic administration of selective serotonin reuptake inhibitors (SSRIs) causes an increase in BDNF protein levels and expression (mRNA) in the hippocampus of rodents, the reciprocal interaction has not been demonstrated yet. Thus, the neurochemical sight of this question will be addressed here by examining the consequences of either a constitutive decrease or increase in brain BDNF protein levels on hippocampal extracellular levels of 5-HT in conscious mice

5-HT4 receptor agonists increase sAPPα levels in the cortex and hippocampus of male C57BL/6j mice

BACKGROUND AND PURPOSE: A strategy to treat Alzheimer's disease (AD) is to increase the soluble form of amyloid precursor protein (sAPPalpha), a promnesic protein, in the brain. Because strong evidence supports beneficial effects of 5-hydroxytryptamine 5-HT(4) receptor agonists in memory and learning, we investigated the role of 5-HT(4) receptors on APP processing in 8 weeks-old male C57BL/6j mice. EXPERIMENTAL APPROACH: Mice were given, subcutaneously, prucalopride or ML 10302 (s.c.), two highly selective 5-HT(4) receptor agonists and, up to 240 min later, the hippocampus and cortex were analysed by Western blot for sAPPalpha determination. KEY RESULTS: Prucalopride (5 or 10 mg kg(-1)) significantly increased sAPPalpha levels in the hippocampus and cortex, but did not modify the expression level of APP mRNA as detected by quantitative RT-PCR. A selective 5-HT(4) receptor antagonist, GR125487 (1 mg kg(-1), s.c.) inhibited prucalopride induced- increase in sAPPalpha levels. In addition, levels of sAPPalpha were increased by ML10302 only at 20 mg kg(-1) and was limited to the cortex. Also, prucalopride increased sAPPalpha levels in the cortex of a transgenic mouse model of AD, expressing the London mutation of APP. Furthermore, the combined injection of a selective acetylcholinesterase inhibitor, donepezil and prucalopride induced a synergic increase in sAPPalpha levels in the cortex and hippocampus. CONCLUSIONS AND IMPLICATIONS: Our results demonstrate that the 5-HT(4) receptor plays a key role in the non-amyloidogenic pathway of APP metabolism in vivo and give support to the beneficial use of 5-HT(4) agonists for AD treatment.status: publishe