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Cigarette smoke regulates the expression of TLR4 and IL-8 production by human macrophages.

Sarir H, Mortaz E, Karimi K, Kraneveld AD, Rahman I, Caldenhoven E, Nijkamp FP, Folkerts G - J Inflamm (Lond) (2009)

Bottom Line: In this study, we evaluated the effects of cigarette smoke medium (CSM) on TLR4 expression and interleukin (IL)-8 production by human macrophages investigating the involvement of ROS.CSM stimulation increased intracellular ROS-production and decreased glutathione (GSH) levels.TLR4 may be involved in the pathogenesis of lung emphysema and oxidative stress and seems to be a crucial contributor in lung inflammation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Division of Pharmacology and Pathophysiology, Departement of Pharmaceutical Sciences, Faculty of Sciences, Utrecht University, the Netherlands. e.mortaz@uu.nl.

ABSTRACT

Background: Toll-like receptors (TLRs) are present on monocytes and alveolar macrophages that form the first line of defense against inhaled particles. The importance of those cells in the pathophysiology of chronic obstructive pulmonary disease (COPD) has well been documented. Cigarette smoke contains high concentration of oxidants which can stimulate immune cells to produce reactive oxygen species, cytokines and chemokines.

Methods: In this study, we evaluated the effects of cigarette smoke medium (CSM) on TLR4 expression and interleukin (IL)-8 production by human macrophages investigating the involvement of ROS.

Results and discussion: TLR4 surface expression was downregulated on short term exposure (1 h) of CSM. The downregulation could be explained by internalization of the TLR4 and the upregulation by an increase in TLR4 mRNA. IL-8 mRNA and protein were also increased by CSM. CSM stimulation increased intracellular ROS-production and decreased glutathione (GSH) levels. The modulation of TLR4 mRNA and surface receptors expression, IRAK activation, IkappaB-alpha degradation, IL-8 mRNA and protein, GSH depletion and ROS production were all prevented by antioxidants such as N-acetyl-L-cysteine (NAC).

Conclusion: TLR4 may be involved in the pathogenesis of lung emphysema and oxidative stress and seems to be a crucial contributor in lung inflammation.

No MeSH data available.


Related in: MedlinePlus

IL-8 expression is ROS dependent after CSM exposure. MDMs (5 × 106 cells/ml) were pretreated with NAC (10 mM) for 30 min and then stimulated by CSM (0.03, 0.06 and 0.12 OD) for 4 h. RNA was extracted and mRNA levels of IL-8 were quantified by real-time PCR (A). Results are expressed as copies of IL-8 vs. copies of GAPDH mRNA. (B) MDMs (1 × 106 cells/ml) were pretreated with NAC (10 mM) for 30 min and then stimulated by CSM (0.06 OD) for 16 h Supernatants were collected after 16 h incubation and IL-8 production was quantified using ELISA methods. *P < 0.05 vs baseline # P < 0.05 vs CSM stimulated (n = 3).
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Figure 4: IL-8 expression is ROS dependent after CSM exposure. MDMs (5 × 106 cells/ml) were pretreated with NAC (10 mM) for 30 min and then stimulated by CSM (0.03, 0.06 and 0.12 OD) for 4 h. RNA was extracted and mRNA levels of IL-8 were quantified by real-time PCR (A). Results are expressed as copies of IL-8 vs. copies of GAPDH mRNA. (B) MDMs (1 × 106 cells/ml) were pretreated with NAC (10 mM) for 30 min and then stimulated by CSM (0.06 OD) for 16 h Supernatants were collected after 16 h incubation and IL-8 production was quantified using ELISA methods. *P < 0.05 vs baseline # P < 0.05 vs CSM stimulated (n = 3).

Mentions: Next, in order to investigate the involvement of ROS by CSM, MDMs were pre-treated with the antioxidant NAC (10 mM) for 30 min and then incubated with CSM (0.03, 0.06 and 0.12 OD) for 4 h. NAC suppressed the upregulation of TLR4 mRNA-induced by CSM compared to control (Fig. 3A). Moreover, NAC suppressed the expression of IL-8 at mRNA and protein levels (Fig. 4A and 4B).


Cigarette smoke regulates the expression of TLR4 and IL-8 production by human macrophages.

Sarir H, Mortaz E, Karimi K, Kraneveld AD, Rahman I, Caldenhoven E, Nijkamp FP, Folkerts G - J Inflamm (Lond) (2009)

IL-8 expression is ROS dependent after CSM exposure. MDMs (5 × 106 cells/ml) were pretreated with NAC (10 mM) for 30 min and then stimulated by CSM (0.03, 0.06 and 0.12 OD) for 4 h. RNA was extracted and mRNA levels of IL-8 were quantified by real-time PCR (A). Results are expressed as copies of IL-8 vs. copies of GAPDH mRNA. (B) MDMs (1 × 106 cells/ml) were pretreated with NAC (10 mM) for 30 min and then stimulated by CSM (0.06 OD) for 16 h Supernatants were collected after 16 h incubation and IL-8 production was quantified using ELISA methods. *P < 0.05 vs baseline # P < 0.05 vs CSM stimulated (n = 3).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2683828&req=5

Figure 4: IL-8 expression is ROS dependent after CSM exposure. MDMs (5 × 106 cells/ml) were pretreated with NAC (10 mM) for 30 min and then stimulated by CSM (0.03, 0.06 and 0.12 OD) for 4 h. RNA was extracted and mRNA levels of IL-8 were quantified by real-time PCR (A). Results are expressed as copies of IL-8 vs. copies of GAPDH mRNA. (B) MDMs (1 × 106 cells/ml) were pretreated with NAC (10 mM) for 30 min and then stimulated by CSM (0.06 OD) for 16 h Supernatants were collected after 16 h incubation and IL-8 production was quantified using ELISA methods. *P < 0.05 vs baseline # P < 0.05 vs CSM stimulated (n = 3).
Mentions: Next, in order to investigate the involvement of ROS by CSM, MDMs were pre-treated with the antioxidant NAC (10 mM) for 30 min and then incubated with CSM (0.03, 0.06 and 0.12 OD) for 4 h. NAC suppressed the upregulation of TLR4 mRNA-induced by CSM compared to control (Fig. 3A). Moreover, NAC suppressed the expression of IL-8 at mRNA and protein levels (Fig. 4A and 4B).

Bottom Line: In this study, we evaluated the effects of cigarette smoke medium (CSM) on TLR4 expression and interleukin (IL)-8 production by human macrophages investigating the involvement of ROS.CSM stimulation increased intracellular ROS-production and decreased glutathione (GSH) levels.TLR4 may be involved in the pathogenesis of lung emphysema and oxidative stress and seems to be a crucial contributor in lung inflammation.

View Article: PubMed Central - HTML - PubMed

Affiliation: Division of Pharmacology and Pathophysiology, Departement of Pharmaceutical Sciences, Faculty of Sciences, Utrecht University, the Netherlands. e.mortaz@uu.nl.

ABSTRACT

Background: Toll-like receptors (TLRs) are present on monocytes and alveolar macrophages that form the first line of defense against inhaled particles. The importance of those cells in the pathophysiology of chronic obstructive pulmonary disease (COPD) has well been documented. Cigarette smoke contains high concentration of oxidants which can stimulate immune cells to produce reactive oxygen species, cytokines and chemokines.

Methods: In this study, we evaluated the effects of cigarette smoke medium (CSM) on TLR4 expression and interleukin (IL)-8 production by human macrophages investigating the involvement of ROS.

Results and discussion: TLR4 surface expression was downregulated on short term exposure (1 h) of CSM. The downregulation could be explained by internalization of the TLR4 and the upregulation by an increase in TLR4 mRNA. IL-8 mRNA and protein were also increased by CSM. CSM stimulation increased intracellular ROS-production and decreased glutathione (GSH) levels. The modulation of TLR4 mRNA and surface receptors expression, IRAK activation, IkappaB-alpha degradation, IL-8 mRNA and protein, GSH depletion and ROS production were all prevented by antioxidants such as N-acetyl-L-cysteine (NAC).

Conclusion: TLR4 may be involved in the pathogenesis of lung emphysema and oxidative stress and seems to be a crucial contributor in lung inflammation.

No MeSH data available.


Related in: MedlinePlus