The Effects Of Different Fatty Acids On The Expression And Secretion Of AgRP And Their Signaling Pathway In N38 Cell Of The Hypothalamus

Food intake is an important factor to affect the production performance of animal. It is now believed that the signal integration of appetite axis in hypothalamus plays an important role on “appetite regulation network”. In the hypothalamic ARC, the concentration change of nutrients such as glucose, fatty acid and amino acid sensed by orexigenic neurons, thus stimulats the expression and secretion of AgRP and takes part in controlling food intake and energy balance. To date, the regulation of food intake and its mechanism of different fatty acid are still not clear. In order to further investigate the effects of the regulation of food intake by different fatty acid, at the cellular level, we use the N38 cells of mice’s hypothalamus as a model. Using western blot and quantitative PCR technique respectively detect the expression and secretion of AgRP and the m RNA expression level of AgRP, CD36 and metabolism-related genes of different fatty acid.Meanwhile, the changes of inflammation and insulin / leptin signaling pathways were also studied by the methods of RNA interference and agonist treatment compared with fatty acids. This research carried out the following results:(1) Different fatty acids have different effects on the expression and secretion of AgRP in N38 cells. 100 μM of butyric, oleic and palmitic acid have no significangtly effect on the expression and secretion of AgRP. However, 100 μM of lauric and octanoic acid significantly increase such an effect of AgRP, as well as stearic and myristic acid(P < 0.05).In contrast, 100 μM of linoleic acid obviously decrease the expression and secretion of AgRP(P < 0.05).(2) Linoleic acid in 10 and 50 μM can significantly inhibit the secretion of AgRP(P <0.05).When the concentration of linoleic acid increases to 100 or 300 μM, it can also downregulate the expression of AgRP; But stearic and lauric acid in 10, 50 and 100 μM can significantly promote the expression and secretion of AgRP(P < 0.05).(3) 100 μM of linoleic acid incubating the N38 cells with LPS, the TLR4 activator, it demonstrate that LPS remarkably eliminated the inhibitory effects of linoleic acid on the protein expression of TLR4 and TLR4-dependent JNK and IKKa/β signaling pathway(P <0.05). After knock down TLR4, the activated expression of TLR4 and TLR4-dependent JNK are reversed by stearic acid(P < 0.05). Meanwhile, the promoted effect of TLR4 and IKKα/β is reversed by Lauric acid after the interference of TLR4 receptor in N38 cells(P <0.05). It is indicated that TLR4 is involved in regulating the expression and secretion of AgRP in N38 cells by the above fatty acids.(4) 100 μM of linoleic acid improves the sensitivity of leptin and insulin. Moreover,linoleic acid-induced enhancement of leptin and insulin signaling pathways are effectively blocked by LPS and reverse the inhibited effect of AgRP(P < 0.05). While stearic acid remarkably suppresse leptin signaling pathway. Furthermore, after knock down TLR4, the inhibition of IRS/Akt/Fox O1 or JAK2/STAT3 is eliminated and reversed(P < 0.05).However, 100 μM of lauric acid has no sign on the expression of insulin/leptin pathway.(5) 100 μM of linoleic acid can significantly reduce the m RNA expression of metabolism related genes FAS and CPT1(P < 0.05), the m RNA expression of CD36 is markedly increased(P < 0.05). While, stearic acid can significantly increase the expression of CPT1 and FAS and CD36(P < 0.05).100 μM of lauric acid in N38 cells, threre are no significant influence in the expression of ACCα/FAS/CPT1 and UCP2.In conclusion, linoleic acid can inhibit the inflammatory pathways by TLR4-dependent,then increasing the sensitivity of insulin/leptin pathway, and ultimately increase the expression and secretion of AgRP. Instead, stearic and lauric acid can increase inflammatory signaling pathway through TLR4. Stearic acid can also decrease the sensitivity of insulin/leptin pathway and upregulate the expression and secretion of AgRP.Meanwhile, our results involve in activating the AgRP neurons by CD36 and metabolic pathways in linoleic acid and stearic acid, it remains to be further research.