The Effects Of Vaspin On PI3K/Akt And NF-kappa B Signaling Pathways In Islet Beta Cells Under High Lipid

Objective:1. To investigate the mechanisms of vaspin induced insulin resistance and inflammation in INS-1 islet beta cells.2. To determine whether vaspin modulates the function of INS-1 cells through PI3K/Akt and NF-kappa B signaling pathways. Methods:In order to study the effects of vaspin on PI3K/Akt and NF-kappa B signaling pathways and determine the appropriate concentration of vaspin, INS-1 cells were cultured and divided into five groups:(1)the control group: treated with culture medium only;(2) the PA group: treated with 0.5 mmol/l palmitic acid;(3) PA + vaspin(80 ng/ml) group: treated with 0.5 mmol/l palmitic acid and 80 ng/ml vaspin;(4) PA + vaspin(160 ng/ml) group: treated with 0.5 mmol/l palmitic acid and 160 ng/ml vaspin;(5) PA + vaspin(320 ng/ml) group: treated with 0.5 mmol/l palmitic acid and 320 ng/ml vaspin. The m RNA levels of IRS-1, IRS-2, Akt, NF-κB were determined by Real-Time PCR. The protein levels of IRS-2, p IRS-2, Akt, p Akt, NF-κB P65 were determined by western blot.In order to determine whether vaspin improve insulin secretion of INS-1 cells through PI3K/Akt and NF-κB signaling pathways, INS-1 cells were divided into three groups respectively:(1) the control group: treated with 0.5 mmol/l palmitic acid;(2) vaspin groups: treated with 0.5 mmol/l palmitic acid and 320 ng/ml vaspin;(3) vaspin + ly294002(PI3K inhibitor) or TPCK(NF-κB inhibitor): treated with 0.5 mmol/l palmitic acid, 320 ng/ml vaspin and 25 μmol/l PI3 K inhibitors ly294002 or 20 μmol/l NF-κB inhibitors TPCK. The m RNA levels of Akt and NF-κB were determined by Real-Time PCR. The protein levels of Akt, p Akt, NF-κB P65 were determined by western blot. The glucose stimulated insulin secretion(GSIS) were determined by ELISA. Results:1. The effects of vaspin on PI3K/Akt and NF-κB signaling pathwaysCompared with control group, PA significantly decreased the level of IRS-2 m RNA, total IRS-2 protein and Akt phosphorylation, while increased the NF-κB m RNA and NF-κB P65 protein levels and the IRS-2 phosphorylation levels(P<0.05). Pretreatment of cells with different concentration of vaspin reversed these PA treatment related effects(P<0.05).2. The effects of vaspin on PI3K/Akt signaling pathway induced by PAAkt phosphorylation and GSIS levels were significantly increased after treatment with vaspin compared to the control group(P<0.05). Pretreatment with PI3 K inhibitor ly294002 blocked this effect of vaspin(P<0.05).3. The effects of vaspin on NF-κB signaling pathway induced by PACompared with control group, vaspin and NF-кB inhibitor, TPCK significantly decreased the m RNA and protein levels of NF-κB and GSIS(P< 0.05). ConclusionVaspin protects the islet beta cells from palmitic acid induced insulin resistance and inflammation through PI3K/Akt and NF-κB signaling pathways.