Cxcl 1

Total RNA was extracted by Trizol reagent (Thermo), according to the manufacturer’s instructions after 24 h post-treatment. The total RNA concentration was determined in RNAase-free water using Nanodrop, while the concentration was determined using Qubit Fluorometer 3.0 (Thermo). 1 μg of total RNA was reverse transcribed using a 5X All-In-One RT MasterMix (Applied Biological Materials, Richmond, BC, Canada) into cDNA using thermo-block (Eppendorf). Finally, the real-time PCR was carried out on ABI 7300HT sequence detection system (ABI), containing 2X TaqMan Gene Expression Master Mix (Invitrogen, USA), DEPC water and 5 μL of cDNA and 1 μL the following primers: Prime Time qPCR Assay: GLUT4, CXCR2, IL-10, CXCL-1, TNFA, GLUT1 and GLUT4 qRnoCIP0027857 were purchased from Biorad, Des Plaines, IL, USA. Triplicate samples were run for each gene. The reference gene GAPDH qRnoCIP0050838 was applied as an internal control to normalize the expression of target genes. Relative expression levels were determined for each sample after normalization against the reference gene, using the ΔΔCt method to compare relative fold expression differences.

Bio-Plex pro reagent kit III and Bio-Plex Pro Human IL-6 set (171BK29MR2), IL-10 set (171BK32MR2), TNF-α (171BK55MR2), CXCL1 (171BK22MR2), and CXCL5 (171BK14MR2) were purchased from Bio-rad. The cytokines and chemokines were determined according to the manufacturer’s protocol. SF samples were measured on the Bio-Plex 200 system (Bio-Rad), and analysis was done using Bio-plex Manager 6.2 Software (Bio-Rad).