Ozone exposure increased neutrophils in bronchoalveolar lavage (D closed bar).

<p>No other inflammatory cell type number was affected by either ozone or the dual p38/JNK MAPK inhibitors. *p<0.05 Significantly different from air exposed controls. Data are mean ± SEM. n = 3–6.</p

Blocking p38 and JNK MAPK completely prevented ozone-induced airway hyperreactivity mediated by the vagus nerves.

<p>In anesthetized and vagotomized guinea pigs, stimulation of the vagus nerves (10V, 0.2 ms pulse width, 1–25 Hz, 5 sec duration at 1 minute intervals) caused frequency dependent bronchoconstriction (A open circles; measured as an increase in inflation pressure in mmH₂O) that is significantly potentiated one day post-ozone exposure (A closed circles). Pretreatment with dual MAPK inhibitors V-05-013 (A closed squares), V-05-014 (B closed triangles), or V-05-015 (C closed inverted triangles) completely prevented ozone-induced airway hyperreactivity. All three dual MAPK inhibitors suppressed parasympathetic nerve activity (A open squares, B open triangles, C open inverted triangles). Ozone and air exposed control data are the same in A-C. *p<0.05, **p<0.01 Significantly different from air exposed controls. Data are mean ± SEM. n = 4–7.</p

Ozone potentiated vagally mediated falls in heart rate (measured as beats/minute; A–C closed circles) compared to air exposed animals (A–C open circles).

<p>Separate pretreatment with all three dual MAPK inhibitors (V-05-013: A closed squares; V-05-014: B closed squares; V-05-015: C closed squares) prevented the ozone-induced potentiation of frequency induced falls in heart rate. Fall in heart rate following intravenous acetylcholine administration was not changed by either ozone, or MAPK inhibitors (D–E). Ozone and air exposed controls are the same for A–C, and are the same for D–F. **p<0.01 Significantly different from air exposed controls. Data are mean ± SEM. n = 3–7.</p

Neither ozone nor the dual p38/JNK MAPK inhibitors affected inflammatory cell numbers in peripheral blood.

<p>Data are mean ± SEM. n = 3–6.</p

Bronchoconstriction (measured as an increase in inflation pressure in mmH₂O) in response to intravenous acetylcholine was significantly potentiated one day post-ozone (closed circles) compared to air exposed controls (open circles), and was not blocked by V-05-013 (A closed squares), or V-05-014 (B closed triangles).

<p>V-05-015 (C closed inverted triangles) attenuated ozone-induced smooth muscle hyperreactivity while air-exposed animals pretreated with V-05-015 were hyperreactive to intravenous acetylcholine (C open inverted triangles). Ozone and air exposed controls are the same in A-C. *p<0.05, **p<0.01 Significantly different from air exposed controls. Data are mean ± SEM. n = 3–7.</p

Baseline cardiovascular and pulmonary parameters.

<p>Values are means ± SEM. Baseline pulmonary inflation pressure significantly increased after ozone exposure. Treatment with dual p38/JNK MAPK inhibitor V-05-013 significantly reduced the ozone-induced increase in baseline pulmonary inflation pressure. *p<0.05 Significantly different from air exposure. ^‡p<0.05 Significantly different from ozone exposure.</p

In control (air exposed) guinea pigs electrical stimulation of the vagus nerves (3–30V, 0.2 ms pulse width, 15 Hz, 5 sec duration at 1 minute intervals) resulted in vagally induced bronchoconstriction (measured as an increase in pulmonary inflation pressure; 16±1 mmH₂O).

<p>An M₂ receptor antagonist, gallamine, potentiated vagally induced bronchoconstriction up to 6-fold in air exposed animals (open circles) demonstrating that functional M₂ receptors were limiting acetylcholine release. The potentiation by gallamine was decreased in ozone-exposed animals, demonstrating M₂ receptors were dysfunctional after ozone exposure (closed circles). V-05-013 partially prevented M₂ receptor dysfunction (C closed squares), while V-05-014 (B closed triangles) and V-05-015 (C closed inverted triangles) completely protected M₂ receptor function. Vagally induced bronchoconstriction in the absence of gallamine was not different from control among all groups. Ozone and air exposed controls are the same in A–C. *p<0.05, **p<0.01 Significantly different from air exposed controls. Data are mean ± SEM. n = 4–7.</p

Ki values for dual p38 and JNK MAPK inhibitors.

<p>All compounds have a Ki greater than 1 µM for all other kinases tested.</p>*<p>This one value is an IC50, not a Ki.</p

Chemical structures of dual p38 and JNK MAPK inhibitors.

<p>Chemical structures of dual p38 and JNK MAPK inhibitors.</p