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Modulation of insulin response in rat soleus muscle by omega-3 polyunsaturated fatty acids: effects on protein phosphorylation, PPAR activity, eicosanoids production and role of caveolae

Grant number: 06/00345-4
Support Opportunities:Scholarships in Brazil - Doctorate (Direct)
Effective date (Start): July 01, 2006
Effective date (End): April 30, 2011
Field of knowledge:Biological Sciences - Physiology - Physiology of Organs and Systems
Principal Investigator:Rui Curi
Grantee:Jarlei Fiamoncini
Host Institution: Instituto de Ciências Biomédicas (ICB). Universidade de São Paulo (USP). São Paulo , SP, Brazil


Insulin resistance that occurs in obesity and type 2 diabetes mellitus is associated with hyperglycemia and increase in plasma free fatty acids (FA) level, which has been suggested to play a role in the establishment of this condition. In contrast to saturated FA, polyunsaturated FA (PUFA) omega-3 improves insulin response. Some mechanisms by which FA modulate insulin response have been described. Palmitic acid (saturated FA) induces insulin resistance by interfering in its signaling pathway. Another possible mechanism of insulin action is the modulation of expression of genes involved in the metabolism of FA and glucose. Opposite effects were found for saturated FA and PUFA on expression of enzymes involved in energetic metabolism. The discovery of transcription factors activated by FA and derivatives - the peroxissome proliferators activated receptors (PPAR) - evidenced the importance of modulation of gene expression by these metabolites. PUFA are intracellularly metabolized generating eicosanoids, which can modulate cell function; for instance, prostaglandin E1 (PGE1) increases the response to insulin. Another possible mechanism by which FA may modulate insulin response is the change in cell membrane phospholipid composition. There is evidence that caveolae - cell membrane microdomains, rich in cholesterol and sphingolipids - are involved in mechanisms of signal reception and transduction. Proteins involved in the insulin signaling cascade are concentrated in caveolae and interact with proteins found in these domains - caveolins. The modulation of FA composition of caveolae and related structural alterations have been suggested as one of the possible mechanisms of PUFA action on insulin signaling. The aim of this project is to investigate the effects of PUFA omega-3 on insulin response in rat soleus muscle, comparing the effects of the PUFA omega-3 with that of palmitic acid. Soleus muscles will be isolated and pre incubated for up to 6 h in Krebs-Ringer bicarbonate buffer containing glucose in the absence or presence of 100 µM of palmitic, a-linolenic, eicosapentaenoic and docosahexaenoic acids and a combination of palmitic acid with PUFA omega-3. After this period, muscles will be incubated in the same buffer added with 2-deoxi-[2,6-3H]-glucose and [U-14C]-glucose. Muscle uptake of 2-deoxi-[2,6-3H]-glucose and the incorporation of [U-14C]-glucose into glycogen with or without insulin stimulus will be measured. It will be investigated if the effects of PUFA omega-3 on insulin sensitivity lead to alterations in expression and phosphorylation of proteins involved in insulin signaling cascade through Western blotting analysis. The modulation of expression of genes involved in glucose and FA metabolism will be investigated through real time PCR analysis. The activity and expression of PPARs alpha e beta will be determined using real time PCR and electrophoretic mobility-shift assay. The role of caveolae in the effect of PUFA omega-3 on insulin sensitivity will be investigated by FA composition of the caveolae and the content of insulin signaling pathway proteins (IR, IRS-1/2 and Akt). Finally, the involvement of eicosanoids in the effect of PUFA omega-3 on insulin response will be determined through the quantification of PGE1 released from the muscle and evaluation of the effects of ciclooxygenase and lipooxygenase inhibitors and addition of PGE1 in the incubation medium. The results of this study will contribute to determine the relationship between PUFA omega-3 and insulin sensitivity. (AU)

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Academic Publications
(References retrieved automatically from State of São Paulo Research Institutions)
FIAMONCINI, Jarlei. Effects of diets with different omega 3 fatty acids content on energy metabolism - modulation of peroxisomes function.. 2011. Doctoral Thesis - Universidade de São Paulo (USP). Instituto de Ciências Biomédicas (ICB/SDI) São Paulo.

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