Bronchoalveolar lavage fluid from asthmatic subjects is mitogenic for human airway smooth muscle.

Airway smooth muscle proliferation may contribute to the airway wall remodeling seen in asthma. In this study we tested for the presence of airway smooth muscle mitogenic activity in bronchoalveolar lavage (BAL) fluid obtained from 12 atopic asthmatics before and serially after segmental allergen challenge, and from four normal subjects who did not undergo allergen challenge. Mitogenic effect was assessed by coincubating BAL fluid with human airway smooth muscle cells, and measuring its effect on (3)[H]thymidine incorporation and cell number. Induction of ERK phosphorylation and cyclin D(1) protein abundance were also assessed. Compared with serum-free medium alone, BAL fluid obtained from normal subjects increased thymidine incorporation, cell number, ERK phosphorylation, and cyclin D(1) abundance. BAL fluid from asthmatic subjects prior to allergen challenge induced even greater increases in all measures, except for cell number, which was similar to that observed with normal subjects' BAL fluid. Incubation with lavage fluid obtained 48 h after segmental allergen challenge in atopic asthmatics caused yet further increases in thymidine incorporation, cell number, and cyclin D(1) protein abundance. Molecular sieving of prechallenge BAL fluid from three asthmatic subjects demonstrated that mitogenic activity was present exclusively in the > 10 kD fraction. These results provide the first direct demonstration that fluid lining the airways of asthmatics contains excess mitogenic activity for human airway smooth muscle, and that this activity increases further after allergen challenge.

Relationship of cellular transmigration and airway response after allergen challenge.

We examined the relationship between eosinophil migration into the bronchoalveolar space and change in FEV(1) after endobronchial allergen challenge (EBAC) in atopic asthmatic (AA) and atopic nonasthmatic (ANA) subjects. The purpose of this study was to obtain continuous, intrasubject controlled assessment of the relationship between cell migration in control and allergen-challenged segments in the same individuals over 96 h. In AA subjects, the eosinophil (Eos) count in the bronchoalveolar lavage fluid (BALF) increased from a baseline of 7,896 +/- 3,865 to 416,476 +/- 231,012 Eos/ml by 72 h (p = 0.001) in the challenged segment post-EBAC. For ANA subjects, the postsegmental challenge count was 29,874 +/- 474 Eos/ml (p = 0.03 versus baseline and p < 0.05 AA peak versus ANA peak). In both groups, there was a comparable decrease in peripheral blood eosinophil count beginning 5 h after challenge, which resolved at 24 h. In AA subjects, 416,476 +/- 231,012 Eos/ml was obtained from the allergen-challenged segment and 23,522 +/- 8,298 Eos/ml was obtained from the sham-challenged segment (p < 0.001) at 72 h. In contrast, there was no difference in the Eos count obtained from the BALF between the antigen- and sham-challenged segments of ANA subjects. We also found that increased airway neutrophils were present in equal numbers in allergen-challenged and sham-challenged segments in both AA and ANA subjects. We conclude that augmented eosinophil migration after EBAC is a characteristic of atopic asthma and is not present in atopic subjects who do not have asthma. We find that BAL eosinophilia in ANA patients as well as neutrophilia in both ANA and AA subjects are nonspecific consequences of bronchoscopy. Finally, we find no relationship between specific airway eosinophil migration into the BALF and FEV(1) < 72 h after challenge; however, at 96 h, there is a substantial decrease in FEV(1) that accompanies BALF eosinophilia.

Near-fatal heat stroke during the 1995 heat wave in Chicago.

BACKGROUND: In July 1995, Chicago sustained a heat wave that resulted in more than 600 excess deaths, 3300 excess emergency department visits, and a substantial number of intensive care unit admissions for near-fatal heat stroke. OBJECTIVE: To describe the clinical features of patients admitted to an intensive care unit with near-fatal classic heat stroke. Patients were followed for 1 year to assess delayed functional outcome and mortality. DESIGN: Observational study. SETTING: Intensive care units in the Chicago area. PATIENTS: 58 patients admitted to the hospital from 12 July to 20 July 1995 who met the case definition of classic heat stroke. MEASUREMENTS: The data collection tool was designed to compile demographic and survival data and to permit analysis of organ system function by abstracting data on physical examination findings, electrocardiography and echocardiography results, fluid resuscitation, radiography results, and laboratory findings. Data on functional status at discharge and at 1 year were collected by using a modified Stanford Health Assessment Questionnaire. RESULTS: Patients experienced multiorgan dysfunction with neurologic impairment (100%), moderate to severe renal insufficiency (53%), disseminated intravascular coagulation (45%), and the acute respiratory distress syndrome (10%). Fifty-seven percent of patients had evidence of infection on admission. In-hospital mortality was 21%. Most survivors recovered near-normal renal, hematologic, and respiratory status, but disability persisted, resulting in moderate to severe functional impairment in 33% of patients at hospital discharge. At 1 year, no patient had improved functional status, and an additional 28% of patients had died. CONCLUSIONS: Near-fatal classic heat stroke is associated with multiorgan dysfunction. A high percentage of patients had infection at presentation. A high mortality rate was observed during acute hospitalization and at 1 year. In addition, substantial functional impairment at discharge persisted 1 year. The degree of functional disability correlated highly with survival at 1 year.

Differential effects of cyclosporine A after acute antigen challenge in sensitized cats in vivo and ex vivo.

1. We determined the effect of cyclosporine A (CsA) treatment on mast cell degranulation and lung resistance (R(L)) in vivo, and tracheal smooth muscle (TSM) contraction ex vivo after antigen challenge in sensitized cats. We also determined the direct effects of addition of CsA to the tissue bath on antigen-induced responses of TSM in vitro. 2. Cats (n=10) were sensitized by i.m. injection of Ascaris suum antigen (AA); 5 cats (CsA+) received CsA twice daily for 2 weeks before acute antigen challenge in doses sufficient to suppress interleukin-2 secretion from feline peripheral blood mononuclear cells ex vivo. 3. Lung resistance increased comparably within 10 min of exposure to AA (P<0.03). Histamine content in bronchoalveolar lavage fluid from both groups increased comparably within 30 min of antigen challenge, from undetectable levels to 542+/-74 pg ml(-1) post AA for CsA+ and from 74+/-19 pg ml(-1) at baseline, to 970+/-180 pg ml(-1) post AA CsA- (P<0.05; P=NS vs CsA+). 4. In excised TSM, active tension elicited by exposure to AA in vitro was 107+/-38% KCl in the CsA+ group vs 144+/-56% KCl in the CsA- group (P=NS). However, contraction of TSM (n=4) harvested from both groups was abolished or greatly diminished after AA challenge when tissues were pre-incubated with 1 microM CsA in vitro (8+/-8% KCl, P<0.05 vs CsA+ and CsA-). This was associated with inhibited release of 5-hydroxytryptamine into the organ bath fluid of tissues treated with CsA in vitro only. 5. We demonstrated that CsA treatment in vivo does not inhibit the early phase asthmatic response or mast cell degranulation following antigen challenge in sensitized cats. Additionally, the effects of CsA on mast cell function ex vivo do not reflect lack of effects of CsA on mast cell function in vivo in this animal model of atopic asthma.

EP1 receptor blockade attenuates both spontaneous tone and PGE2-elicited contraction in guinea pig trachealis.

We assessed the effect of prostaglandin (PG) E2 on tone of guinea pig tracheal smooth muscle (TSM) strips in vitro. In the presence of spontaneous tone [ST; i.e., no indomethacin (-Indo)], exogenous PGE2 caused a significant relaxation of ST at concentrations > 10(-6) M [to -127 +/- 40.8% electric field stimulation (EFS); P = 0.001 vs. baseline ST] and at concentrations < 10(-6) M caused a variable change in contractile force (51.6 +/- 29.6% EFS; P = NS vs. baseline ST). In the absence of ST (i.e., +Indo) 10(-10) to 10(-7) M PGE2 elicited contraction of TSM to 126.3 +/- 10.5% EFS (P = 0.001 vs. baseline) and no relaxation. Addition of prostanoid EP1 receptor antagonist (either AH-6809 or SC-19220) to Indo-treated TSM caused a substantial rightward shift and attenuation of contraction in response to PGE2 (55.9 +/- 16.8% EFS for 10(-5) MAH-6809; P = 0.007 vs. +Indo alone, and 80.5 +/- 12.7% EFS for 10(-5) M SC-19220, P = 0.03 vs. +Indo alone). We further assessed the effect of EP1 and EP4 receptor antagonism on the ST of guinea pig TSM strips. Concentration-response curves to the EP1 receptor-specific antagonists AH-6809 or SC-19220 and the EP4 receptor-specific antagonist AH-23848B were generated (10(-7) to 10(-5) M); AH-6809 caused relaxation of ST to 11.4 +/- 2.9% ST (P = 0.001 vs. initial ST) and SC-19220 caused relaxation to 31.0 +/- 12.7% ST (P = 0.02 vs. initial ST). However, AH-23848B did not significantly affect ST (to 60.9 +/- 7.7% ST; P = 0.07 vs. initial ST). Furthermore, AH-6809 specifically inhibited contraction elicited by the EP1 receptor agonist Iloprost but had no effect on contraction elicited by the EP3 receptor agonist Enprostil. We demonstrate that in the presence of ST (-Indo), exogenous PGE2 elicits a biphasic response in guinea pig TSM in which relaxation predominates. In the absence of ST (+Indo), exogenous PGE2 elicits contraction of guinea pig TSM strips that is inhibited by EP1 receptor-specific antagonism. Spontaneous tone of guinea pig TSM strips also is inhibited by EP1 receptor antagonism. Our data suggest that both PGE2-elicited contraction and ST of guinea pig TSM are mediated through activation of EP1 prostanoid receptors.

Muscarinic hyperresponsiveness of antigen-sensitized feline airway smooth muscle in vitro.

OBJECTIVE: To determine the effect of in vivo antigen sensitization (Ascaris suum) of cats on tracheal smooth muscle (TSM) and bronchial smooth muscle (BSM) muscarinic reactivity in vitro. ANIMALS: Healthy domestic shorthair cats of either sex. PROCEDURE: Cats were sensitized and were long-term antigen (or sham) challenge exposed for 6 weeks by aerosolization with soluble Ascaris suum. Tracheal and BSM preparations were obtained and stimulated in vitro by electrical field stimulation (EFS), acetylcholine (ACh, a muscarinic agonist), and physostigmine (an AChase inhibitor). Responses were compared with responses of comparable tissues from sham antigen challenge-exposed cats. RESULTS: Tracheal and BSM from sensitized, compared with sham-sensitized (control), cats had greater isometric contraction (expressed as percentage of the response observed for isotonic, 63 mM KCl-elicited contraction [% KCl]) in response to endogenous (EFS) and exogenous muscarinic receptor activation (ACh). Contractions in response to EFS by TSM from control cats were 74% KCl vs 97% KCl for antigen-sensitized TSM (P < 0.04). Muscarinic responses were augmented comparably by in vivo sensitization; TSM from control cats contracted to 190% KCl vs 230% KCl (P < 0.03) for TSM from immune-sensitized cats. Physostigmine augmented responses of all tissues to ACh so that TSM from control (290% KCl) and antigen-sensitized (257% KCl) cats were similar. Responses of BSM from antigen-sensitized cats had similar augmentation of contractile response to EFS and ACh. CONCLUSIONS: Long-term in vivo antigen sensitization increases numbers of muscarinic receptors on airway smooth muscle or decreases the availability or activity of AChase in cats. CLINICAL RELEVANCE: Modulation of muscarinic receptors may be useful for treatment of asthmatic cats with in vivo airway hyperreactivity.

Differential tachykinin receptor subtype activation in capsaicin and KCl contractions of guinea pig trachealis.

We assessed the role of endogenously secreted tachykinins in mediating contraction caused by potassium chloride (KCl) in guinea pig tracheal smooth muscle (TSM) strips in vitro. Maximal isometric contraction was elicited with approximately 45 mM KCl and was 196 +/- 8% of the response to electrical field stimulation (% EFS) in the same tissues. Muscarinic receptor blockade with atropine modestly attenuated this contraction caused by KCl to 175 +/- 9 %EFS (P < 0.05), and treatment with a selective neurokinin subtype 1 (NK1) receptor antagonist, LY-297911, caused even greater inhibition of KCl-elicited contraction to 124 +/- 8 %EFS (P < 0.001). By contrast, SR-48968, a selective NK2 antagonist, had no effect on contraction caused by KCl (183 +/- 9 %EFS; P = NS vs. KCl alone). However, given together at the same concentration, SR-48968 augmented the inhibition of contraction caused by LY-297911 to 93 +/- 15 %EFS (P < 0.05 vs. LY-297911 alone). In contrast to the effect on KCl-induced contraction, LY-297911 caused only moderate inhibition of the contraction caused by capsaicin to 81 +/- 13 %EFS (P < 0.05 vs. control, 114 +/- 15 %EFS), whereas SR-48968 caused substantial attenuation of contraction caused by capsaicin to 23 +/- 5 %EFS (P < 0.005 vs. LY-297911). We demonstrate that a significant portion of the contraction caused by KCl, in addition to capsaicin, is elicited in guinea pig TSM through neurokinin secretion. However, NK1 receptors predominantly mediate contraction caused by KCl, and NK2 receptors predominantly mediate contraction elicited by capsaicin in guinea pig airway smooth muscle.

Cyproheptadine-induced attenuation of type-I immediate-hypersensitivity reactions of airway smooth muscle from immune-sensitized cats.

We assessed the effect of serotonergic inhibition by cyproheptadine on the responsiveness of tracheal smooth muscle (TSM) strips and epithelium-intact third-generation bronchial rings from immune-sensitized (Ascaris suum) cats after exposure to antigen. Cats were sensitized by IM administration of antigen and adjuvant twice over a 4-week period. Sensitization was confirmed in vivo by skin testing with antigen and by physiologic airway responses after exposure to nebulized antigen. Tissues were tethered isometrically to force transducers and were actively equilibrated by exposures to KCl-substituted perfusate. Maximal response after exposure to antigen (expressed as percentage of maximal contraction of each tissue to 63 mM KCl (%KCl) was 169 +/- 18% KCl for sensitized TSM and 43 +/- 18% KCl for sensitized TSM pretreated with cyproheptadine (P < 0.001). Similarly, maximal response to antigen was 81 +/- 27% KCl for sensitized bronchial rings, compared with 16 +/- 14% KCl for sensitized bronchial rings pretreated with cyproheptadine (P = 0.05 vs control). Blockade of leukotriene synthesis by 10(-6) to 10(-4) M A-64077, a selective 5-lipoxygenase inhibitor, had no significant effect on peak response for either TSM (193 +/- 13% KCl vs 169 +/- 18% KCl for sensitized untreated TSM) or bronchial rings (79 +/- 20% KCl vs 81 +/- 27% KCl for sensitized untreated bronchial rings). Release of serotonin from airway tissues was confirmed by the presence of serotonin in the perfusate of 8 sensitized TSM preparations after, but not before, antigen challenge.(ABSTRACT TRUNCATED AT 250 WORDS)

Immune sensitization augments epithelium-dependent spontaneous tone in guinea pig trachealis.

We examined epithelial modulation of tracheal smooth muscle (TSM) responsiveness in vitro from guinea pigs receiving active immune sensitization in vivo. Initially, guinea pigs were either ovalbumin sensitized (by aerosol) or sham sensitized with normal saline; TSM responsiveness was assessed isometrically as active tension (AT) after equilibration by electrical field stimulation in vitro. For epithelium-intact (Epi+) tissues, sensitization caused an increase in baseline active spontaneous tone (1.89 +/- 0.20 g AT) vs. sham-sensitized tissues (1.18 +/- 0.28 g AT; P = 0.02). Spontaneous tone in sensitized TSM in which the epithelium was removed (Epi-) (1.01 +/- 0.14 g AT) was substantially less than from Epi+ tissues (P = 0.01) and did not differ from sham-sensitized epithelium-denuded tissues (0.82 +/- 0.24 g AT; P > 0.05). Indomethacin caused a reduction in spontaneous tone to comparable magnitude for all treatment paradigms. Immune sensitization caused physiological reduction in the ability to relax in response to isoproterenol; the concentration of isoproterenol eliciting 50% relaxation of spontaneous tone was 7.10 +/- 0.13 (-log M) for TSM from sensitized guinea pigs compared with 8.20 +/- 0.27 (-log M) for sham-sensitized tissues (P = 0.006). However, after precontraction with exogenous acetylcholine, relaxation caused by isoproterenol was not affected by either indomethacin or epithelial removal. Muscarinic responsiveness to acetylcholine was augmented by immune sensitization; however, the increase in response to acetylcholine was attenuated by epithelium removal or cyclooxygenase blockade.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of airway inflammation on smooth muscle shortening and contractility in guinea pig trachealis.

We studied the effect of either 1) immunogenic inflammation caused by aerosolized ovalbumin or 2) neurogenic inflammation caused by aerosolized capsaicin in vivo on guinea pig tracheal smooth muscle (TSM) contractility in vitro. Force-velocity relationships were determined for nine epithelium-intact TSM strips from ovalbumin-sensitized (OAS) vs. seven sham-sensitized controls and TSM strips for seven animals treated with capsaicin aerosol (Cap-Aer) vs. eight sham controls. Muscle strips were tethered to an electromagnetic lever system, which allowed isotonic shortening when load clamps [from 0 to maximal isometric force (Po)] were applied at specific times after onset of contraction. Contractions were elicited by supramaximal electrical field stimulation (60 Hz, 10-s duration, 18 V). Optimal length for each muscle was determined during equilibration. Maximal shortening velocity (Vmax) was increased in TSM from OAS (1.72 +/- 0.46 mm/s) compared with sham-sensitized animals (0.90 +/- 0.15 mm/s, P < 0.05); Vmax for TSM from Cap-Aer (0.88 +/- 0.11 mm/s) was not different from control TSM (1.13 +/- 0.08 mm/s, P = NS). Similarly, maximal shortening (delta max) was augmented in TSM from OAS (1.01 +/- 0.15 mm) compared with sham-sensitized animals (0.72 +/- 0.14 mm, P < 0.05); delta max for TSM from Cap-Aer animals (0.65 +/- 0.11 mm) was not different from saline aerosol controls (0.71 +/- 0.15 mm, P = NS). We demonstrate Vmax and delta max are augmented in TSM after ovalbumin sensitization; in contrast, neurogenic inflammation caused by capsaicin has no effect on isolated TSM contractility in vitro. These data suggest that airway hyperresponsiveness in vivo that occurs in association with immunogenic or neurogenic inflammation may result from different effects of these types of inflammation on airway smooth muscle.

Physiologic significance of epithelial removal on guinea pig tracheal smooth muscle response to acetylcholine and serotonin.

We studied the modulatory effect of airway epithelium on guinea pig tracheal smooth muscle (TSM) contraction. Isometric force was measured in vivo before and after removal of the tracheal epithelium. In parallel studies, TSM contraction was also measured isometrically in epithelium-intact and epithelium-denuded TSM strips in vitro. Epithelial removal in vivo did not alter the contractile response of TSM to acetylcholine (ACh) or serotonin. In nine guinea pigs, active tension (AT) caused by 3 x 10(-7) mol/kg of intravenous ACh was 0.74 +/- 0.14 g force per longitudinal length of the segment (g/cm) in the presence of epithelium versus 0.89 +/- 0.16 g/cm after removal of airway epithelium (confirmed histologically) (p NS). The threshold response to ACh was also unchanged (-8.0 +/- 0.3 log mol/kg control versus -8.3 +/- 0.3 log mol/kg after epithelial removal, p NS). In six guinea pigs, the AT caused by 3 x 10(-8) mol/kg of intravenous serotonin was 1.92 +/- 0.63 g/cm with an intact epithelium versus 2.15 +/- 0.70 g/cm after epithelial removal in vivo (p NS). Epithelial removal in vitro increased the sensitivity of TSM contraction to ACh when the data were expressed as the percentage maximal response to ACh. The concentration of ACh causing 50% of the maximal response (EC50) was -5.74 +/- 0.25 log M in eight epithelium-intact TSM strips versus -6.37 +/- 0.16 log M after epithelial removal in controls (n = 8) (p = 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)