Expression of diabetes-related genes in rodent β-cell line (RIN-m) treated with monocarbonyl curcumin analogues (C66 and B2BrBC) in presence or absence of streptozotocin – PCR array data
Description
Curcumin monocarbonyl analogues C66* and B2BrBC** were synthesized at the Institute of Chemistry, Ss. Cyril and Methodius University in Skopje, Macedonia by substituting benzaldehyde with cyclohexanone in a base catalyzed aldol reaction. RIN-m rat β-cells (ATCC, Cat. # CRL-2057) were grown in RPMI-1640 medium (ATCC, Cat. # 30-2001) supplemented with 10% FBS (VWR, Cat. # 89510-186) and antibiotic/antimycotic mixture (MP Biomedicals, Cat. # 1674049) in incubator with standard conditions (5% CO2 and temperature of 37 °C). For experiments, 0.3 x 10(6) cells were seeded in 6-well plates with tissue culture growth media. The following day the media was replaced with serum-free media supplemented with 1.5 mM of streptozotocin (Sigma-Aldrich Cat. # 85882), 50 µM of C66, or B2BrBC dissolved in DMSO (VWR, Cat. # N182), and combination of streptozotocin with C66 or B2BrBC, for 72 hours. Total RNA was extracted using TRIzol reagent (Ambion, Cat. # 15596018). The quantification of RNA was made using NanoDrop One spectrophotometer (Thermo Fisher Scientific) after which the samples were normalized to 1 µg total RNA and converted to cDNA using qScript cDNA SuperMix (Quantabio, Cat. # 95048). PCR array was performed using QuantStudio 3 Real-Time PCR System (Thermo Fisher Scientific) and RT2 Profiler PCR Array – Rat Diabetes (Qiagen, Cat. # 330231, GeneGlobe ID PARN-023ZA). The Ct values were obtained using QuantStudio Design & Analysis Software. * (2E,6E)-2,6-bis[(2-trifluoromethyl)benzylidene]cyclohexanone, C22H16F6O ** (2E,6E)-2,6-bis(2-bromobenzylidene)cyclohexanone), C20H16Br2O