For overexpression of IRAK-M, IRAK-M cDNA clone and control vector (pCMV6-XL-4) were purchased from Origene. Raw macrophages exposed to alcohol for 7 days, were transiently transfected with IRAK-M expression vector and control vector using Lipofectamine and plus reagents (Invitrogen) according to the manufacturer’s instructions. The transfected cells were either stimulated with LPS for 24 h to check the effect of overexpression or for 6 h to determine the production of TNF-α in IRAK-M overexpressed cells.
