Background-The mechanisms involved in the dysfunction of both endothelium-dependent vasodilatation (EDV) and NO biosynthesis related to smoking are unclear. In this study, EDV was assessed in healthy smokers and nonsmokers in vivo and, using serum from the same individuals, was related to the NO biosynthetic pathway in vitro. Methods and Results-Flow-mediated EDV of the brachial artery was measured in 23 male patients (8 nonsmokers and 15 smokers). Serum was collected, added to confluent (Ϸ85%) monolayers of human umbilical vein endothelial cells (HUVECs), and incubated for 12 hours. Basal and substance P-stimulated NO production was measured. The HUVECs used for measuring basal NO production were lysed, and both endothelial NO synthase (eNOS) protein expression and eNOS activity were determined. EDV was lower in smokers compared with nonsmokers (PϽ0.001). HUVECs treated with serum from smokers compared with nonsmokers showed significantly lower basal (PϽ0.0001) and stimulated (PϽ0.02) NO production, higher eNOS expression (PϽ0.0001), but lower eNOS activity (PϽ0.004). There was a significant positive correlation between in vivo EDV and in vitro substance P-stimulated NO production (rhoϭ0.57, PϽ0.01) and between basal NO production and eNOS activity (rϭ0.54, PϽ0.008) and a negative correlation between basal NO production and eNOS protein expression (rϭϪ0.60, PϽ0.003). Conclusions-This is the first study to combine an in vivo model with a near-physiological in vitro model to demonstrate an association between decreased NO production and reduced EDV. Cigarette smoking was associated with reduced EDV, NO generation, and eNOS activity in the presence of increased eNOS protein expression.

These results indicate light smoking may have similar detrimental effects on EDV and NO biosynthetic pathway as does heavy smoking. These data may have important implications concerning the amount of active cigarette exposure that imparts cardiovascular risk.

Background-Data about the effects of smoking on thrombo-hemostatic factors (tissue factor [TF] and tissue factor pathway inhibitor ) are limited and on fibrinolytic factors (tissue plasminogen activator [t-PA] and plasminogen activator inhibitor-1 [PAI-1]) are debatable. The present study investigated the smoking-related, endothelial cell (EC)-specific responses for these factors and their relation to nitric oxide (NO) production in vitro. Methods and Results-Serum from 8 nonsmokers and 15 smokers were incubated with confluent (Ϸ85%) human umbilical vein endothelial cells (HUVECs) in 24-well tissue-culture plates for 12 hours. After the incubation, basal NO, t-PA, PAI-1, TF, TFPI-1 production, and substance P (SP)-stimulated NO, t-PA, and PAI-1 production were determined. HUVECs treated with smokers' serum showed lower basal (PϽ0.02) and SP-stimulated (Pϭ0.059) t-PA production but similar basal and stimulated PAI-1 production (Pϭ0.9 and Pϭ0.6) compared with nonsmokers. Basal t-PA/PAI-1 molar ratio was significantly reduced in smokers (PϽ0.005). TFPI-1 level in the cell culture supernatant was also significantly lower in smokers compared with the nonsmoker group (PϽ0.05) with no difference in TF level between both groups (Pϭ0.5). As previously reported, both basal (PϽ0.001) and SP-stimulated (PϽ0.05) NO production were significantly reduced in smokers. Basal TFPI-1 in culture correlated positively with basal NO production (rϭ0.42, Pϭ0.04) and negatively with serum cotinine level (rϭϪ0.6, Pϭ0.01). Conclusions-These results indicate that cigarette smoking is associated with alterations in EC-derived fibrinolytic (t-PA)and antithrombotic (TFPI-1) factors. To our knowledge, this is the first demonstration that EC-derived TFPI is affected by smoking and endogenous NO or that the degree of smoke exposure may influence TFPI levels in an EC milieu.

The effects of acute and prior exposure to lipopolysaccharide (LPS) and staphylococcal enterotoxin B (SEB) on superoxide release by monocytes were examined in control subjects and in patients with sepsis and septic shock during the acute stage and recovery. High doses of LPS, PMA (phorbol 12-myristate 13-acetate), and SEB stimulated monocyte superoxide release in control subjects (P < 0.05). Pretreatment of normal monocytes with these doses of LPS, PMA, and SEB induced significant hyporesponsiveness to subsequent challenge (P < 0.01), and evidence of cross-tolerance was observed. Monocytes isolated from patients with sepsis and septic shock demonstrated high spontaneous superoxide release compared with those of control subjects (P < 0.05). Stimulation of patient monocytes with LPS or SEB resulted in less superoxide production than that spontaneously released by controls (P < 0.01). In patients recovering from their initial infection, spontaneous superoxide release was less than that released during acute stage. In addition, the superoxide release in response to the same stimuli was significantly increased when compared with release during the acute stage (P < 0.05). These data demonstrate that both LPS and SEB induce hyporesponsiveness to LPS- or SEB-stimulated superoxide release. A similar pattern of hyporesponsiveness was observed during sepsis that may represent a mechanism for modulating the inflammatory response during severe infections.