Figure 1.
TNF-α is required for the increase in PAI-1 transcription induced by inhalational exposure to CAPs.
Wild-type (IL-6+/+) and IL-6−/− mice were exposed contemporaneously to CAPs (PM2.5) or filtered air (FA) for 8 hours daily on 3 consecutive days and BAL fluid was harvested at the end of the third day of exposure for measurement of Interleukin-6 (IL-6) (A), Monocyte Chemotactic Protein-1 (MCP-1) (B) and Tumor Necrosis Factor-α (TNF-α) (C). White adipose tissue was harvested from identically treated mice for measurement of the levels of PAI-1 mRNA (D). Wild type mice were treated with etanercept (10 mg/kg i.p.) or vehicle (saline) 3 days before and on the first day of exposure to CAPs or FA and white adipose tissue levels of PAI-1 mRNA were measured after the third day of exposure (E). (*p<0.05 CAPs vs. FA, n≥6/group).
Figure 2.
Interleukin-6 is required for activation of coagulation after inhalational exposure to concentrated ambient particles (CAPs) and the instillation of urban PM.
Wild-type (IL-6+/+) and IL-6−/− mice were exposed contemporaneously to CAPs (PM2.5) or filtered air (FA) for 8 hours daily on 3 consecutive days and lung tissue and BAL fluid were harvested at the end of the third day of exposure. Lung levels of mRNA encoding IL-6 and its transcriptional targets surfactant protein B (SFPB) and tissue factor (TF) (A–C) and plasma levels of thrombin antithrombin (TAT) complexes (D) were measured. Wild-type (IL-6+/+) and IL-6−/− mice were treated intratracheally with urban PM (200 µg/mouse) and 24 hours later BAL fluid was obtained for measurement of IL-6 levels (ELISA) (E) and plasma was collected for measurement of thrombin-antithrombin complexes (F). (*p<0.05 CAPs vs. FA, n≥6/group).
Figure 3.
Urban PM induces the IL-6-dependent expression of tissue factor and deposition of fibrin in the lung.
Wild-type (IL-6+/+) and IL-6−/− mice were treated with PM (200 µg/mouse) or PBS for measurement of tissue factor (TF) protein (A) (immunoblotting and densitometry analysis shown) (n = 4/treatment) and mRNA (B) (qRT-PCR) in whole lung homogenates (n = 4/treatment). In identically exposed mice, BAL fluid fibrin was measured as the difference in the levels of D-Dimer pre- and post-digestion with exogenously administered plasmin (C). (*p<0.05, PM vs. PBS, n = 6/treatment).
Figure 4.
PM-induced lung injury is similar in wild-type and IL-6−/− mice.
Wild-type (IL-6+/+) and IL-6−/− mice were treated with PM or PBS and harvested 24 hours later for histologic evaluation (A). Representative microphotographs of lung sections (hematoxylin and eosin stain, ×100 magnification, ×600 (inset) are shown. (B) Assessment of lung injury severity using a 5 point scoring system. (* p<0.05, PM vs. PBS, n≥4/group) Wild-type and IL-6−/− mice were treated with urban PM (200 µg/mouse) or PBS and BAL fluid was obtained 24 hours later for measurement of (C) total protein (D) cell count and (E) differential cell count. (*p<0.05, PM vs. PBS, n = 8/group).
Figure 5.
TNF-α but not IL-6 is required for urban PM-induced local and systemic release of PAI-1.
Wild-type (IL-6+/+) and IL-6−/− mice were treated with PM (200 µg/mouse) or PBS and lung and white adipose tissues, BAL fluid and plasma were harvested 24 hours later. (A) Whole lung PAI-1 mRNA, (B) BAL fluid PAI-1 antigen, (C) adipose tissue PAI-1 mRNA and (D) BAL fluid TNF-α levels were measured. Wild-type mice were treated with either etanercept, a TNF-α inhibitor (10 mg/kg i.p.) or vehicle (saline) 3 days and on the day of exposure to PM (200 µg/mouse) or PBS and BAL fluid, white adipose tissue and plasma were harvested or a bleeding time after tail vein cut performed 24 hours later. BAL fluid PAI-1 antigen (E), white adipose tissue PAI-1 mRNA (F), bleeding time after tail vein cut (G), plasma prothrombin time (PT) (H), partial thromboplastin time (PTT) (I) and plasma thrombin-antithrombin (TAT) complexes (J) were measured. (p<0.05, *PM vs. PBS, n≥4/group).