Guinea Pig Pulmonary Mechanics: Altered Sensitivity to Carbachol by Cadmium and/or Selenium

Male Hartley guinea pigs (480–610 g) were treated intratracheally as follows: saline, cadmium (Cd, 0.3 mg), selenium (Se, 0.3 or 0.06 mg), or Se (0.06 mg) and Cd (0.3 mg) simultaneously. Selenium and Cd were administered as sodium selenite and cadmium chloride, respectively. Twenty-four h later, dynamic lung compliance (C_dyn) and pulmonary resistance (R_p) were measured before (baseline C_dyn and R_p) and after carbachol administration (0.0001, 0.001, 0.01, and 0.1 μmol/kg, intravenously). Results indicated a significant decrease in baseline C_dyn caused by 0.3 mg of Cd, 0.3 mg or 0.06 mg of Se, and 0.3 mg of Cd with 0.06 mg of Se (p < 0.05). A significant increase in baseline R_p due to 0.3 mg of Se was observed (p < 0.05). Carbachol decreased C_dyn significantly below baseline, evident after lower doses of carbachol, in guinea pigs pretreated with 0.3 mg of Se, whereas a significant improvement in C_dyn was seen after 0.0001 μmol/kg carbachol in the group pretreated with Se and Cd simultaneously (p < 0.05) compared with the respective baseline values of the saline-treated group. Similarly, a significant increase in R_p was observed after carbachol in groups pretreated with 0.3 mg of Cd or Se (p < 0.05). Results also indicated a significant increase in large airway constriction caused by Cd and/or Se (p < 0.05). A leftward shift in the carbachol dose-response curve indicated increased sensitivity to carbachol in Cd- and/or Se-pretreated guinea pigs. Accepted for publication: 14 March 1997     

Bronchial Hyperresponsiveness in Farmers: Relation to Respiratory Symptoms, Lung Function, and Atopy

There is only limited information on the factors associated with nonspecific bronchial hyperresponsiveness (BHR) in farmers. Our purpose was to examine the relationship between BHR and respiratory symptoms, atopy, and abnormalities of lung function in a sample of French farmers. Farmers scheduled for a preventive medicine check-up in northeastern France were examined. Occupational exposure, respiratory symptoms, and work-related symptoms were assessed by questionnaire, sensitization to 34 common and agricultural allergens by skin prick tests, and BHR by the single-dose (1,200 μg) acetylcholine (ACh) challenge test. Data were obtained from 741 farmers (95% of those invited). Seventy-seven subjects (10.3%) had BHR defined as a fall in forced expiratory volume in 1 s (FEV₁) ≥ 10% after the inhalation of ACh or, for those with a poor lung function, an increase in FEV₁ > 10% and > 200 ml after the inhalation of 200 μg of salbutamol. The proportion of asthmalike symptoms, especially wheeze during work, positive skin tests to acarian (storage mites) and cereal dust allergens, and low levels of lung function was significantly greater among reactors than among nonreactors. Stepwise logistic regression analysis showed a significant and independent association between BHR and wheezing during work (OR = 4.99; 95% CI = 2.29–10.89; p= 0.0001) and baseline FEV₁ (OR = 1.49; 95% CI = 1.05–2.20; p= 0.026). In conclusion, hyperreactive farmers had significantly more asthmalike symptoms, positive skin tests, and abnormal lung function than normoreactive farmers. Work-related wheeze and low baseline FEV₁ were significantly and independently associated with BHR. Accepted for publication: 26 January 1999     

A Murine Model of Latex Allergy-Induced Airway Hyperreactivity

Sensitization to latex proteins can cause immediate IgE mast cell-mediated reactions. Health care workers have been found to be particularly at risk because of high exposure. Latex allergy can be produced in mice as demonstrated by IgE and eosinophil responses. Thus the mouse is a potential animal model for studying this disease, but the airway response to latex sensitization in mice has not been evaluated previously. In the present study, we immunized BALB/c mice intranasally with nonammoniated latex proteins. Animals were anesthetized, and lung mechanics were evaluated plethysmographically. Changes in pulmonary conductance (G_L) and compliance (C_dyn) were measured in response to a nonspecific challenge with methacholine or to a direct challenge with intravenous latex antigen. Latex sensitization resulted in elevated levels of IgE and latex-specific IgG₁ as well as interstitial infiltrates consistent with an allergic response. The methacholine dose-response ED₅₀ for G_L was 116.4 μg for the control mice and fell significantly to 20.9 μg for latex-sensitized mice. The ED₅₀ calculated for C_dyn was also significantly lower after latex sensitization. The G_L in latex-sensitized mice challenged with latex antigen fell significantly from a prechallenge value of 1.87 ± 0.41 (S.E.) to 0.198 ± 0.03 ml · s⁻¹· cmH₂O after latex antigen challenge. The results indicate that latex-sensitized mice did exhibit increased airway reactivity in the methacholine challenge test. The latex allergic response in mice is unique in that direct challenge with latex antigen itself also resulted in a significant airway response. Accepted for publication: 10 September 1998     

Cough Receptor Sensitivity to Capsaicin and Tartaric Acid in Patients with Mycoplasma Pneumonia

There has been no detailed study of cough sensitivity during acute lower respiratory infection. The aim of this study was to clarify cough sensitivity in Mycoplasma pneumonia, which is a well known acute lower respiratory infection with persistent nonproductive cough. We examined cough sensitivity to inhaled capsaicin and tartaric acid in both the acute and the convalescent phases of Mycoplasma pneumonia, cell differentials in bronchoalveolar lavage fluid, and pathologic findings of transbronchoscopic bronchial biopsy specimens. Although dry cough was observed in all patients during Mycoplasma pneumonia, cough sensitivity in the acute phase [capsaicin: 19.8 (GSEM, 0.214) μM, tartaric acid: 0.26 (GSEM, 0.356) M] were not enhanced compared with those in both control subjects [capsaicin: 27.9 (GSEM, 1.24) μM, tartaric acid: 0.316 (GSEM, 0.079) M] and patients in the convalescent phase [capsaicin: 15.7 (GSEM, 0.219) μM, tartaric acid: 0.50 (GSEM, 0.326) M] when all symptoms including cough had disappeared. The percentage of lymphocytes and neutrophils in bronchoalveolar lavage fluid BALF was significantly greater than in the control subjects, and lymphocyte-dominant bronchitis was observed in biopsied specimens. We conclude that cough threshold to inhaled capsaicin or tartaric acid was not enhanced during acute Mycoplasma pneumonia with lymphocyte-predominant bronchitis. This is the first report examining cough sensitivity in patients with acute lower respiratory infection with pneumonia. Accepted for publication: 4 December 1997     

Induction of Manganese Superoxide Dismutase Gene Expression in Bronchoepithelial Cells after Rockwool Exposure

Superoxide dismutases play an important protective role in the lung defense against the pro-oxidative effect of fibrous dusts (e.g. crocidolite fibers). Particularly crocidolite, but also other asbestos fibers, are known to induce cellular antioxidant defense. Although rockwool, a man-made fiber made from rock, is used widely for insulation purposes, its effects on the superoxide dismutases in bronchoepithelial cells have not been investigated. Thus, the purpose of this study was to determine whether human bronchoepithelial cells (BEAS 2B) respond to rockwool fibers (115-4 experimental rockwool fiber) by induction of MnSOD mRNA and an increase of MnSOD activity levels. The results were compared with BEAS 2B cells exposed to silica (α-quartz: DQ12; SiO₂) and UICC (Union Internationale Contre le Cancer) crocidolite (concentrations of all dusts: 0, 2, 5, 10, 25, 50 μg/cm²= 0, 2.4, 6, 12, 30, 60 μg/ml; 24-h exposure) as control fibers. Scanning electron microscopy confirmed close dust cell contact under all experimental settings. Very low MnSOD mRNA baseline levels rose significantly (p < 0.001) in BEAS 2B cells exposed to all three dusts at 2 μg/cm². However, at >25 μg/cm² MnSOD mRNA levels in silica- and crocidolite- but not in rockwool-exposed cells decreased. Slight (no significance) increases of MnSOD activity were observed which decreased at higher dust (>5 μg/cm²) concentrations. These results suggest that: (1) like crocidolite and silica, rockwool accelerates MnSOD gene expression in bronchoepithelial cells; (2) an increase of MnSOD mRNA levels is not accompanied by MnSOD activity elevation; (3) in contrast to rockwool, high concentrations (≥25 μg/cm²) of crocidolite and silica reduced MnSOD activity and MnSOD mRNA levels. Because oxidants (H₂O₂) and crocidolite fibers were shown to reduce SOD activity, lack of active MnSOD protein may be caused by inactivation on a post-translational level. Furthermore, the decline of MnSOD mRNA and MnSOD activity levels coincides with increasing cytotoxicity. In conclusion, rockwool was demonstrated to induce MnSOD gene expression, perhaps because of its pro-oxidative effect in bronchoepithelial cells. In contrast to crocidolite and silica, rockwool fibers are not cytotoxic in this experimental setting. Accepted for publication: 21 August 1997     

Methacholine Dose-Response Slopes from Maximal Bronchial Challenge Tests in Asthmatic Children: Methodological Aspects

To determine whether the slope of a maximal bronchial challenge test (in which FEV₁ falls by over 50%) could be extrapolated from a standard bronchial challenge test (in which FEV₁ falls up to 20%), 14 asthmatic children performed a single maximal bronchial challenge test with methacholine (dose range: 0.097–30.08 μmol) by the dosimeter method. Maximal dose-response curves were included according to the following criteria: (1) at least one more dose beyond a ΔFEV₁≥ 20%; and (2) a MFEV₁≥ 50%. PD₂₀ FEV₁ was calculated, and the slopes of the early part of the dose-response curve (standard dose-response slopes) and of the entire curve (maximal dose-response slopes) were calculated by two methods: the two-point slope (DRR) and the least squares method (LSS) in % ΔFEV₁×μmol⁻¹. Maximal dose-response slopes were compared with the corresponding standard dose-response slopes by a paired Student's t test after logarithmic transformation of the data; the goodness of fit of the LSS was also determined. Maximal dose-response slopes were significantly different (p < 0.0001) from those calculated on the early part of the curve: DRR_20% (91.2 ± 2.7 ΔFEV₁% ·μmol⁻¹) was 2.88 times higher than DRR_50% (31.6 ± 3.4 ΔFEV₁% ·μmol⁻¹), and the LSS_20% (89.1 ± 2.8% ΔFEV₁·μmol⁻¹) was 3.10 times higher than LSS_50% (28.8 ± 1.5% ΔFEV₁·μmol⁻¹). The goodness of fit of LSS_50% was significant in all cases, whereas LSS_20% failed to be significant in one. These results suggest that maximal dose-response slopes cannot be predicted from the data of standard bronchial challenge tests. Accepted for publication: 12 December 1996     

The Importance of Polymorphonuclear Leukocytes in Lipopolysaccharide-Induced Superoxide Anion Production and Lung Injury: Ex Vivo Observation in Rat Lungs

The purpose of this study is to determine if the polymorphonuclear leukocyte (PMN) is a major causative agent for lipopolysaccharide (LPS)-induced lung injury and responsible for the excess production of superoxide anion in the lung. We measured superoxide anion production from the lung and pulmonary capillary permeability in rats with and without PMN depletion. The superoxide anion production from the lung was measured using a purpose-built ex vivo chemiluminescence apparatus. Pulmonary capillary permeability was evaluated by the Evans blue dye extravasation method. PMN sequestration was determined by counting PMNs in histologic tissue specimens using microscopy. All rats received 3 mg/kg LPS intravenously. Examinations were undertaken at 2, 6, and 12 h after the LPS injection. The PMN-depleted group received cyclophosphamide 4 days before the LPS injection, which resulted in a PMN count of less than 200 cells/μl. In rats without PMN depletion, Evans blue dye extravasation increased significantly at 12 h after the LPS injection; PMN sequestration increased at 2, 6, and 12 h after the LPS injection; and superoxide anion production increased at 6 h and remained elevated at 12 h after the LPS injection. The increased permeability, PMN sequestration, and superoxide anion production were not seen in the PMN-depleted group. The contribution of the xanthine/xanthine oxidase system and alveolar macrophages to the observed superoxide anion production was negligible. We conclude that, in rats, the PMN is a major causative agent in LPS-induced lung injury and is responsible for the excess production of superoxide anion in the lung. Accepted for publication: 3 March 1997     

Airway Eosinophilic Inflammation and Bronchial Hyperresponsiveness after Allergen Inhalation Challenge in Asthma

Allergen exposure in atopic asthmatic patients is associated with recruitment and activation of eosinophils in the airways. Once activated, eosinophils release toxic products, including the eosinophil cationic protein (ECP), able to damage bronchial structures and to increase bronchial hyperresponsiveness. With this background, the present study was designed to evaluate whether ECP levels in bronchoalveolar lavage (BAL) fluid could reflect, better than BAL eosinophil counts, the cellular activation that follows allergen exposure in atopic asthmatics. Twenty-two atopic patients attended the laboratory on two separate days. On the 1st day, they underwent methacholine (MCh) inhalation challenge to detect the degree of nonspecific bronchial hyperresponsiveness. On the 2nd day, they underwent fiberoptic bronchoscopy and BAL, at baseline or 4–6 h after allergen inhalation challenge. In this latter patient group, MCh challenge was repeated 3–5 h after allergen challenge, 1 h before fiberoptic bronchoscopy. The analysis of the mean baseline FEV₁ values and the degree of bronchial reactivity to MCh (MCh Pd₂₀) on the 1st study day did not demonstrate differences between the two patient groups (p > 0.1, each comparison). In addition, in the allergen-challenged group, MCh Pd₂₀ was decreased significantly after allergen challenge (151.4 μg/ml and 67.6 μg/ml, respectively, before and after challenge; p < 0.05). Evaluation of the different BAL cell types demonstrated that the proportions of eosinophils and epithelial cells were increased significantly in the allergen-challenged group compared with the group evaluated at baseline (p < 0.01 and p < 0.05, respectively). Moreover, ECP levels, corrected by the correspondent albumin levels (ECP/Alb), were higher in the allergen-challenged group compared with the group evaluated at baseline (p < 0.05). In addition, although a positive correlation was demonstrated between BAL eosinophil percentages and ECP/Alb values (r= 0.72, p < 0.05) in the group evaluated at baseline, no links were found between these parameters in the allergen-challenged group (p > 0.1). However, in this latter group, a weak positive correlation was demonstrated between eosinophil percentages and ΔMch, i.e., the increased nonspecific bronchial reactivity, which is observed after allergen challenge (r= 0.55; p < 0.05). Thus, in stable asthmatic patients an ongoing activation of eosinophils parallels their migration, but this eosinophilic inflammation is not strictly related to bronchial reactivity to Mch. By contrast, after allergen inhalation challenge, eosinophil recruitment and activation seem to follow different temporal kinetics, and eosinophilic inflammation may be partially associated with the degree of airway hyperresponsiveness. Accepted for publication: 15 September 1997     

Byssinosis: Role of Polymer Length on the Effect of Tannin on the Airway β-Adrenergic Receptor

Tannin, isolated from cotton bracts and implicated in the pathogenesis of byssinosis, inhibits isoproterenol and forskolin-stimulated cAMP release from airway cells in part by decreasing cell surface β-adrenergic receptor number and uncoupling the β-adrenergic receptor from its stimulatory G-protein (G_s) and in part by inhibiting adenylyl cyclase activity. We have hypothesized that cotton tannin, because of its long polymer length, interacts with the hydrophobic binding pocket of the β-adrenergic receptor and alters β-adrenergic receptor binding and G_s coupling. In these studies, tannins of three different polymer lengths and molecular masses were isolated from cotton bracts using sequential Amicon ultrafiltration [molecular mass > 10,000 (YM10 retentate), 1,000–10,000 (YM10 filtrate), and 1,000–5,000 Da (YM2 retentate)]. The YM10 retentate (25 μg/ml) decreased chloride secretion (Jnet = 1.11 ± 0.28 (control) to 0.59 ± 0.18 μEq/cm²·h, p < 0.05, n= 6), decreased cell surface β-adrenergic receptor number (18.0 ± 1.8 (control) to 10.6 ± 0.9 fmol/mg protein, p < 0.02, n= 4), and inhibited forskolin-stimulated cAMP release (5,254 ± 1,290 (control) to 2,968 ± 620 pmol/mg protein, p < 0.01, n= 8). In contrast, neither the YM10 filtrate nor the YM2 retentate had any effect on net chloride secretion, β-adrenergic cell surface receptor number, or forskolin-stimulated cAMP release. We conclude that polymer length is essential for the effect of tannin on the β-adrenergic receptor and on adenylyl cyclase. Accepted for publication: 28 June 1998     

Effects of vagal stimulation on slowly adapting pulmonary stretch receptors and lung mechanics in anesthetized rabbits

An in vivo preparation was designed to investigate the effect of vagus nerve stimulation-induced bronchoconstriction on the relationship of slowly adapting pulmonary stretch receptor (SAR) activity and lung mechanics. SAR activities were recorded from the left vagus nerve. The responses of SARs, total lung resistance (R_L), and dynamic lung compliance (C_dyn) to electrical stimulation of the peripheral end of the cut right vagus nerve (10–15 V, 5–30 Hz, 0.2 ms) were examined before atropine and 5 and 10 min after atropine (2 mg/kg) in anesthetized, artificially ventilated, bilaterally vagotomized rabbits. In the time course profile during vagal stimulation, an increase in R_L and a decrease in C_dyn occurred simultaneously, and these opposite changes were frequency dependent. The average responses of SAR activity, RL, and Cdyn to vagal stimulation became more pronounced as the frequencies of the stimulation were increased. The responses obtained during vagal stimulation (5–30 Hz) were blocked or diminished greatly by the administration of atropine. Repeated vagus nerve stimulation in the presence of atropine did not show any significant change in SAR activity and lung mechanics. These results suggest that changes of SAR activity, R_L, and C_dyn induced by vagal stimulation occur as a result of smooth muscle contraction in the airways, which is mediated mainly by muscarinic receptor activation and which is not involved in the release of neurotransmitters to relax airway smooth muscle. Offprint requests to: S. Matsumoto     

Attenuation of Colonic Inflammation by PPARγ in Intestinal Epithelial Cells: Effect on Toll-like Receptor Pathway

The peroxisome proliferator-activated receptor gamma (PPARγ) is a nuclear receptor highly expressed in the colon and playing an anti-inflammatory role through inhibition of the NF-κB pathway. Toll-like receptor 4 (TLR4) has been known to mediate LPS-induced cellular signaling through activation of NF-κB pathway in intestinal epithelial cells. The aims of this study were to evaluate attenuation of inflammation by PPARγ in intestinal epithelial cells and to study the possible relation between PPARγ and TLR4. HT-29 human epithelial cells were stimulated with LPS (20 μg/ml) and PPARγ ligand, 15d-PGJ₂ (10 μM), or with LPS (20 μg/ml) alone for 24 hr. COX-2, IL-8, TLR4, and PPARγ mRNA expression was assessed by RT-PCR. IL-8 protein levels and TLR4 protein expression were analyzed by ELISA and Western blot, respectively. To evaluate the action mechanisms of PPARγ ligand, Western blot analysis for IκBα degradation was performed. Costimulation with LPS and PPARγ ligand in comparison to LPS stimulation alone (1) decreased COX-2, IL-8 mRNA expression and IL-8 protein secretion, (2) decreased TLR4 mRNA and protein expression, and (3) decreased PPARγ mRNA expression. PPARγ ligand delayed LPS-induced IκBα degradation. These findings suggest that PPAR-γ ligands suppress inflammation in intestinal epithelial cells. PPARγ and TLR, these two antagonistic signaling pathways in intestinal epithelial cells may be partially cross-linked.     

Budesonide Down-Regulates Eosinophil Locomotion But Has No Effects on ECP Release or on H2O2 Production

Treatment of allergic asthma with inhaled corticosteroids results in local down-regulation of proinflammatory cytokine synthesis and in marked decrease in tissue eosinophilia. Blood concentrations of inhaled corticosteroids, although significantly lower than those measured in the lung, may still have antiinflammatory effects on circulating eosinophils, reducing their ability to migrate. The aim of our study was to evaluate in vitro the activity of budesonide on blood eosinophils by measuring their chemotactic response, eosinophil cationic protein (ECP) release, and hydrogen peroxide (H₂O₂) production in the presence of different drug concentrations similar to those obtained at airway level (10⁻⁸ and 10⁻⁷ M) and at blood level (10⁻¹⁰ and 10⁻⁹ M). Partially purified blood eosinophils, isolated from 23 asthmatic subjects, were used to evaluate the activity of budesonide on: (1) chemotaxis toward the activated fifth component of complement (C5a, 0.1 μg/ml) or recombinant human (rh) interleukin (IL)-5 (200 pg/ml), (2) ECP release by cells stimulated with tetradecanoylphorbol acetate (TPA) and (3) H₂O₂ production by TPA-activated cells. The chemotactic response to C5a was down-regulated significantly by budesonide only by the highest concentrations tested (10⁻⁸ and 10⁻⁷ M); differently, budesonide was effective in inhibiting eosinophil migration toward rhIL-5, at all concentrations tested (p < 0.01, each comparison). By contrast, no drug-induced modifications were observed in ECP release or in H₂O₂ production (p > 0.05, each comparison). We conclude that concentrations of budesonide similar to those obtained in vivo are effective in inhibiting eosinophil locomotion but not in down-regulating the release of reactive oxygen species and granule-associated proteins. Accepted for publication: 11 February 1999     

Central AT1 receptors are involved in the enhanced cardiac sympathetic afferent reflex in rats with chronic heart failure

The purpose of this study was to determine if the cardiac sympathetic afferent reflex (CSAR) was augmented in rats with coronary ligation-induced chronic heart failure (CHF), and if central angiotensin II (ANG II) was involved in this enhancement. Under α-chloralose and urethane anesthesia, mean arterial pressure (MAP), heart rate (HR) and renal sympathetic nerve activity (RSNA) were recorded in sino-aortic denervated and cervical vagotomized rats. An intracerebroventricular cannula was implanted. The CSAR was examined by epicardial application of bradykinin (BK, 0.04 and 0.4 μg in 2.0 μl) or capsaicin (0.04 and 0.4 μg in 2.0 μl) to the anterior and posterior wall of the left ventricle. The CSAR evoked by BK or capsaicin was augmented in rats with CHF. In sham rats, there was no significant difference of the CSAR induced by BK or capsaicin between anterior and posterior epicardial application. However, in rats with CHF, the CSAR induced by BK (0.04 μg) to anterior epicardial application was blunted compared with posterior application. Intracerebroventricular injection of losartan (500 nmol) normalized the enhanced CSAR in rats with CHF, but had no significant effects on the CSAR in sham rats. However, intravenous application of the same dose of losartan only decreased the baseline MAP, but did not alter baseline RSNA or the CSAR in sham or CHF rats. Pre-treatment with epicardial application of lidocaine to the anterior wall abolished the CSAR evoked by application of BK or capsaicin but had no effects on the CSAR evoked by epicardial application of BK or capsaicin to the posterior wall. These results suggest that the CSAR induced by epicardial application of BK and capsaicin is enhanced in the rats with CHF, and the enhanced CSAR is mediated by central AT₁ receptors. Received: 17 December 2001/Returned for revision: 7 January 2002/Revision received: 4 February 2002/Accepted: 26 February 2002     

Effects of calcium channel and H1-receptor blockers on the responses of slowly adapting pulmonary stretch receptors to histamine in vagotomized rabbits

We studied the effects of calcium channel antagonists (verapamil and nifedipine) and H₁-receptor blockers (mequitazine) on changes in the slowly adapting pulmonary stretch receptors (SARs) located below the carina in response to right atrial injections of histamine (60 and 80 μg/kg) in anesthetized artificially ventilated rabbits with bilateral vagotomy. After histamine was injected into the right atrium, the SARs became more active during expiration but decreased their activity during inspiration. These changes were more pronounced by increasing the dosage of histamine. However, administration of histamine had no significant effect on tracheal pressure (P_T). Verapamil treatment (1 mg/kg) did not alter the SAR response to histamine, whereas the responses of SARs to histamine at different dosages were significantly diminished by treatment with nifedipine (1 mg/kg). Mequitazine (1 mg/kg), a potent H₁-receptor blocker, blocked completely all the responses of SAR activity to histamine. These results suggest that the effect of histamine 60–80 μg/kg on SAR activity is mediated by the activation of H₁-receptors of the peripheral airway smooth muscle and that this activation, at least in part, involves the opening of calcium channels of the airway smooth muscle.     

Suppressive Effect of Prostaglandin E1 on Pulmonary Hypertension Induced by Monocrotaline in Rats

The effect of administering prostaglandin E₁ (PGE₁) on the extent of monocrotaline (MCT)-induced pulmonary hypertension and cytokine production [interleukins (IL) 1 and 6 and tumor necrosis factor (TNF)] by macrophages during MCT induction of pulmonary hypertension was studied. Right ventricle/left ventricle plus septum weight ratios (RV/LV + S) were used as an index of the development of pulmonary hypertension. Administering PGE₁ at a dose of 0.2 mg/kg/day for 4 weeks reduced significantly the RV/LV + S ratio from 0.428 ± 0.070 to 0.243 ± 0.059 (p < 0.01) and decreased the production of these cytokines: IL-1, from 4.675 ± 3.558 to 1.800 ± 0.722 units; IL-6, from 0.322 ± 0.121 to 0.060 ± 0.039 units; and TNF, from 0.578 ± 0.369 to 0.004 ± 0.004 units. In another series of experiments, a significant reduction of the RV/LV + S ratio was noted for only 1 week when we administered PGE₁ immediately after the injection of MCT. We confirmed that histopathologic improvements of lungs were noted by administering 0.2 mg/kg PGE₁ for 4 weeks. In another experiment, PGE₁ at a concentration of 2 μg/ml suppressed a rise in the cytosolic Ca²⁺ concentration of lipopolysaccharide-stimulated peritoneal macrophages of rats in vitro, suggesting that PGE₁ suppressed cytokine production by macrophages through the suppression of the Ca²⁺ influx. These results suggest that administering PGE₁ may be effective in the treatment of some forms of pulmonary hypertension in humans. Accepted for publication: 20 August 1998     

Effects of GABA-B Agonist Baclofen on Bronchial Hyperreactivity to Inhaled Histamine in Subjects with Cervical Spinal Cord Injury

Bronchial provocation studies performed in our research center have consistently demonstrated airway hyperresponsiveness to both inhaled methacholine and histamine in subjects with chronic cervical spinal cord injury (SCI). More recently, we reported that the airways of such subjects maintained on chronic baclofen (γ-aminobutyric acid) therapy were not hyperreactive to inhaled methacholine. In this study we determined whether baclofen also blocks the effects of the bronchoprovocative agent histamine in subjects with cervical SCI. Twenty-four male subjects with cervical SCI participated in this study; 14 were maintained on oral baclofen, and 10 served as age-matched controls. The subjects were challenged with increasing concentrations of aerosolized histamine until either a 20% fall in forced expiratory volume in 1 s (FEV₁) from baseline (defined as PC₂₀) was observed, or a maximum of 25 mg/ml histamine was administered. We found that 11 of the 14 baclofen subjects (78.5%) and 8 of the 10 control subjects (80%) responded (PC₂₀ < 8 mg/ml) to the histamine challenge. Mean PC₂₀ values among responders in the baclofen (PC₂₀= 2.91 ± 2.3) and control (PC₂₀= 2.18 ± 1.9) groups did not differ significantly. Because histamine acts directly on histamine receptors and indirectly on cholinergic pathways, our findings that baclofen blocks bronchoconstriction due to inhaled methacholine, but not that due to histamine, suggests that hyperresponsiveness in subjects with cervical SCI may be secondary to nonspecific airway hyperreactivity. Accepted for publication 21 January 1997     

Developmental Regulation of the Effects of Surfactant Protein A on Phospholipid Uptake by Fetal Rat Type II Pneumocytes

Surfactant protein A (SP-A) enhances the uptake of phospholipid by type II cells derived from adult and late gestation fetal rat lung. The present study was performed to examine more fully the developmental biology of the effects of SP-A on phosphatidylcholine (PC) uptake, to determine the effect of SP-A on the cellular location of bound and internalized phospholipid and on the metabolism of internalized phospholipid by morphologically undifferentiated (18-day) and morphologically differentiated (19-day) fetal type II cells. SP-A enhanced uptake almost twofold in a dose-dependent manner in 19-day fetal cells, but it had no effect on uptake by 18-day fetal cells at any concentration. Stimulation of uptake by 19-day fetal cells was saturable at concentrations above 1 μg/ml SP-A. Maximal uptake was 1.12 nmol of PC/mg of protein, and the effective concentration that yields 50% maximal response, K_Φ, was 58.9 ng/ml (84.1 pM). The effect of SP-A on uptake by 19-day fetal cells was detectable as early as 1 min of exposure. Uptake correlated significantly with time both in the absence (r= 0.98, p < 0.001) and presence of 5 μg/ml SP-A (r= 0.979, p < 0.001). The rate of uptake in the presence of SP-A (0.019 ± 0.002 nmol of PC/mg of protein/min) was twice the rate of uptake in controls (0.009 ± 0.001 nmol of PC/mg of protein/min). SP-A had no effect on binding to plasma membranes and uptake of phospholipid into lamellar bodies by 18-day fetal cells. On the other hand, SP-A significantly enhanced binding of dipalmitoyl phosphatidylcholine to plasma membranes (two- to threefold) and uptake into lamellar bodies (threefold) of 19-day fetal cells. SP-A caused a significant reduction in the degradation of internalized phospholipid by differentiated fetal type II cells. Based on the lack of effect of exogenous SP-A on 18-day fetal cells, we conclude that the response to SP-A is under developmental control. SP-A enhances the initial binding to the plasma membranes of fetal type II cells and subsequent internalization into the lamellar bodies. This effect is associated with a protection of internalized phospholipid from metabolic degradation. Both of these processes are developmentally regulated during the transition from the canalicular to the saccular phase of lung development. Accepted for publication: 15 May 1997     

The acute cardioprotective effect of glucocorticoid in myocardial ischemia–reperfusion injury occurring during cardiopulmonary bypass

The purpose of this study was to evaluate the acute cardioprotective effect of high-dose methylprednisolone (25 mg/kg) in the controlled in vivo model of myocardial ischemia–reperfusion injury occurring during cardiopulmonary bypass. Forty nondiabetic male patients with three-vessel disease undergoing first-time bypass surgery were enrolled for this double-blind prospective study. Patients were randomized to be given 25 mg/kg methylprednisolone (Group I) and saline (Group II) 1 h before cardiopulmonary bypass. The levels of cardiac troponin-I (cTnI) were used as a marker of myocardial tissue damage in myocardial ischemia–reperfusion injury. The cTnI levels were measured before surgery, at the second hour after cardiopulmonary bypass, at the 6th and 24th hours, and 5th day postoperatively. There was no significant difference between the two groups in respect to the duration of ischemia and reperfusion. The preoperative cTnI levels were 0.22 ± 0.29 ng/ml in Group I and 0.23 ± 0.28 ng/ml in Group II. cTnI levels increased to 2.40 ± 1.0 ng/ml in Group I and 3.19 ± 0.88 ng/ml in Group II at the 2nd hour after cardiopulmonary bypass. When the differences between T1 and T0 level that showed the amount of troponin release occurring due to ischemia–repefusion injury was calculated and then compared, there was a significant difference between Groups I and II (P = 0.024). The cTnI levels measured at 6 h after CPB were 1.98 ± 0.63 ng/ml in Group I and 2.75 ± 1.15 ng/ml in Group II (P = 0.049). cTnI levels decreased to 0.22 ± 0.10 ng/ml in Group I and 0.49 ± 0.25 ng/ml in Group II on the postoperative day 5 (P = 0.0001). Univalent regression analysis showed that preoperative high-dose corticosteroid usage decreased the troponin release in about 12% and this effect was statistically significant (R² = 0.12, P < 0.05). A single dose of intravenous methylpredisolone (25 mg/kg) given 1 h before ischemia reduced myocardial ischemia–reperfusion injury. These results demonstrated that the acute cardioprotective effect of corticosteroids has much potential in the future for reducing ischemia–reperfusion injury occurring during cardiopulmonary bypass when it is inevitable.     

Effects of Concanavalin A on Na+-Dependent and Na+-Independent Mechanisms for H+ Extrusion in Alveolar Macrophages

Alveolar macrophages (mφ) possess two parallel mechanisms for plasmalemmal H⁺ extrusion: a V-type H⁺ pump (V-ATPase) and a Na⁺/H⁺ exchanger (NHE). To investigate the coordinated functioning of these H⁺ extruders for mφ intracellular pH (pH_i) regulation, we investigated the effects of the plant lectin concanavalin A (ConA) on resident alveolar mφ from rabbits. ConA (1 μM, 30-min pretreatment) activated the mφ for phagocytosis of opsonized Escherichia coli. ConA activation did not affect the baseline pH_i of mφ or the initial rate of pH_i recovery (dpH_i/dt) from an intracellular acid load (acid-loaded pH_i nadir ≈ 6.9). However, the contributions of Na⁺-independent H⁺ transport (i.e. V-ATPase activity) and Na⁺-dependent H⁺ transport (i.e. NHE activity) to dpH_i/dt were altered significantly. The lectin stimulated Na⁺/H⁺ exchange and inhibited V-ATPase activity. In control mφ, V-ATPase-mediated H⁺ extrusion was responsible for >80% of dpH_i/dt. Conversely, in ConA-treated mφ, Na⁺/H⁺ exchange was responsible for ∼65% of dpH_i/dt, and V-ATPase activity was responsible for only 35% of dpH_i/dt. These results underscore the complex mechanisms and signaling pathways that coordinate the activities of cellular acid-base transporters in mφ pH_i regulation. Accepted for publication: 18 March 1997     

Iron status in Danes updated 1994. I: Prevalence of iron deficiency and iron overload in 1332 men aged 40–70 years. Influence of blood donation, alcohol intake, and iron supplementation

 Iron status, S-ferritin, and hemoglobin (Hb) were assessed in a population survey in 1994 (DAN-MONICA 10) comprising 1332 Caucasian Danish men equally distributed in age cohorts of 40, 50, 60 and 70 years. Blood donors (n=186) had lower S-ferritin, median 76 μg/l, than nondonors, median 169 μg/l (p<0.0001). S-ferritin in donors was inversely correlated with the number of phlebotomies (r _s=–0.57, p<0.0001). S-ferritin in nondonors (n=1146) was similar in men 40–60 years of age, median 176 μg/l, and subsequently decreased at 70 years of age to a median of 146 μg/l (p=0.01). In the entire series, the prevalence of small iron stores (S-ferritin 16–32 μg/l) was 2.7%, that of depleted iron stores (S-ferritin <16 μg/l) 0.45%, and that of iron deficiency anemia (S-ferritin <13 μg/l and Hb <129 g/l) 0.15%. Among nondonors, the prevalence of iron overload (S-ferritin >300 μg/l) was 20%. S-ferritin in nondonors correlated with body mass index (r _s=0.19, p=0.0001) and with alcohol intake (r _s=0.26, p=0.0001). In the entire series, 28% of the subjects took supplemental iron (median 14 mg ferrous iron daily). Iron supplements had no influence on iron status. Nondonors (n=170) treated with acetylsalicylic acid had lower S-ferritin, median 136 μg/l, than nontreated, median 169 μg/l (p<0.001) and those treated with H₂-receptor antagonists (n=30) had lower S-ferritin, median 142 μg/l, than nontreated, median 171 μg/l (p<0.04). Compared with the DAN-MONICA 1 iron status survey of Danish men in 1984, the prevalences of iron depletion and iron deficiency anemia are unchanged whereas the prevalence of iron overload has increased significantly. In Denmark, iron fortification of flour was abolished in 1987. This apparently had no negative effect on iron status in men. Received: November 19, 1998 / Accepted: April 25, 1999