RNA isolation and RT-PCR
Total RNA isolated from 200 µL bupropion treated and untreated PBMCs
using Total RNA Mini Kit (Yektatajhiz, Iran) and cDNA synthesized
afterward using a cDNA synthesis kit (Yektatajhiz, Iran) according to
the manufacturer’s instruction. Relative mRNA expression of target genes
including tumor necrosis factor-a (TNF-a), Interleukine1(IL-1β), Janus
Kinase 2 (JAK2), and Signal transducer and activator of transcription 3
(STAT3) quantification conducted on Step One Plus real-time polymerase
chain reaction (PCR) system (Applied Biosystems, ABI, Foster City, CA,
USA) using SYBR Green qPCR Master Mix (Applied Biosystems, ABI, Foster
City, CA, USA). β-actin mRNA levels used for results’ normalization.
Primer sequences are mentioned in Table 1.