Isolation of Venous Endothelial Cells

Venous endothelial cells were freshly isolated from non-smokers, non-menthol cigarette smokers, and menthol cigarette smokers without cardiovascular disease, as previously described^{26 (link)–28 (link)}. A 0.018 inch J-wire (Arrow International, Reading, PA) was inserted through a 20 or 22 gauge catheter in a vein of the forearm and used to gently rub the inside of the vessel. Following removal, the J-wire was rinsed several times with red blood cell lysis and dissociation buffer. The sample was then centrifuged and cells applied to poly-L-lysine coated slides (Sigma, St. Louis, MO). Nitric oxide production and/or bioavailability was then assessed immediately following isolation as outlined below.