Flavones derived from nature attenuate the immediate and late-phase asthmatic responses to aerosolized-ovalbumin exposure in conscious guinea pigs

Objective

Asthma is an inflammatory disease of the lung that is characterized by airway hyperresponsiveness and the increase of inflammatory cell infiltration into the airways. Naturally occurring flavones have potent anti-inflammatory effects, but their effects on asthmatic responses are still relatively unknown.

Methods

We evaluated the inhibitory effects of flavone derivatives having the chromone moiety on the immediate-phase asthmatic response (IAR) and the late-phase asthmatic response (LAR) to aerosolized-ovalbumin (OA) exposure in conscious OA-sensitized guinea pigs.

Results

Luteolin and apigenin (30 mg/kg, p.o.) significantly (P < 0.05) decreased not only the specific airway resistance (sRaw) in IAR and LAR, but also the recruitment of leukocytes and the release of histamine and activities of phospholipase A₂ (PLA₂) and eosinophil peroxide (EPO) in bronchoalveolar lavage fluid (BALF), compared to control. However, their anti-asthmatic activities were less than those of cromolyn sodium and dexamethasone.

Conclusions

These results indicate that flavones containing more hydroxyl radicals have a greater anti-asthmatic effect. The potencies of flavone anti-asthmatic activities are, in order: luteolin ≥ apigenin > baicalein > chrysin > flavone.     

Antigen-induced late-phase airway obstruction in the guinea pig?

Hartley or Dunkin-Hartley guinea pigs (n=136) were actively sensitized to ovalbumin orAscaris suum protein by five different regimes. Specific airway resistance (sR _AW) was measured in conscious animals by a plethysmographic procedure before, immediately following and at various intervals (up to 96 h) after aerosolized antigen or vehicle challenge. Each sensitization and challenge regime produced an immediate allergic response with positive responses (defined as a 2-fold increase in sR _AW) in 78–100% of animals. Recovery from the immediate response followed by late-phase responses was observed in only two out of a group of four animals. The results failed to substantiate literature reports of a high incidence of late responses in the guinea pig at 4–8, 17–24 or 72 h after allergen challenge.     

Density profile of bronchoalveolar lavage eosinophils in the guinea pig model of allergen-induced late-phase allergic responses.

Inhalation of aerosolized ovalbumin by guinea pigs both during sensitization and upon challenge induces a pulmonary eosinophilia as assessed by cells recovered in bronchoalveolar lavage fluid (BALF). In comparison with BALF eosinophil numbers in naive animals of 0.82 +/- 0.2 x 10(6) cells, those in sensitized animals before challenge and 17 and 72 h after challenge were 1.48 +/- 0.2 x 10(6), 2.60 +/- 0.6 x 10(6), and 4.2 +/- 0.7 x 10(6) cells, respectively. BALF eosinophils from all these groups were notable for their heterogeneity with respect to density, size, and appearance under the electron microscope. In comparison with peritoneal eosinophils, which had a single mean density peak of 1.088 +/- 0.001 g/ml, BALF cells comprised hypodense (less than 1.080 g/ml), normodense (1.080 to 1.096 g/ml), and hyperdense (greater than 1.096 g/ml) eosinophils. The percentage of hypodense eosinophils rose from 25% in naive animals to 63% in sensitized animals (P less than 0.001) and fell after challenge. In contrast, challenge induced the appearance of hyperdense eosinophils, which rose from 6% in sensitized animals to 42% 72 h after challenge (P less than 0.001). Blood eosinophils in naive animals showed a similar profile to those in the lung, but after sensitization and challenge no gross changes in the proportion of either hypodense or hyperdense eosinophils were observed. Flow cytometric analysis of BALF eosinophils indicated that hypodense eosinophils, with a mean diameter of 15.8 microns, were larger than both normodense and hyperdense eosinophils, which had mean diameters of 14.3 and 11.6 microns, respectively. Although the numbers and size of granules were not reduced in hypodense BAL eosinophils, electron microscopy morphology indicated a reduced granular content.(ABSTRACT TRUNCATED AT 250 WORDS)     

The effect of sympathomimetic drugs on immediate skin reactions and metabolic responses in asthmatic patients

The varied effects of intradermal injection of isoproterenol and propranolol on the immediate skin reaction were studied in relation to the metabolic responses to the intravenous injection of epinephrine. In asthmatic patients whose skin reaction was not suppressed with 10(-7) M isoproterenol, the hyperglycemic response after the injection of epinephrine was significantly reduced.     

Inhalation exposure to respiratory sensitising chemicals down-regulates guinea pig IgE and pulmonary responses

Inhalation exposure of the guinea pig to the respiratory sensitising chemicals trimellitic anhydride (TMA) or diphenylmethane di-isocyanate down-regulated the IgE antibody response to subsequent systemic challenge with alum-precipitated hapten-protein conjugate. The suppression was isotype and hapten specific, challenge with unrelated hapten-protein conjugate resulting in high-titre IgE and IgGl antibody responses. Preliminary results indicate that the down-regulation of the IgE response to TMA markedly reduced the capacity of the animals to undergo a bronchoconstriction response following challenge with an aerosol of TMA, suggesting that the initial route of exposure may have profound effects upon the development of occupational respiratory allergy.     

Prostaglandin D2 and histamine during the immediate and the late-phase components of allergic cutaneous responses

With a skin blister technique in which the mediators generated by the trauma of forming the blister are allowed to subside, we have collected human interstitial skin fluid during the course of allergic reactions to ragweed, and measured levels of histamine and prostaglandin D2 (PGD2). Of 18 ragweed-allergic individuals tested, 11 developed both an immediate and a late-phase reaction (LPR) with fivefold-elevated levels of histamine (40 ng/ml) at 30 minutes and a peak level of PGD2 (6.5 ng/ml) later at 2 1/2 hours after ragweed challenge. The other seven allergic individuals had immediate reactions without an LPR lesion and demonstrated somewhat smaller elevations of histamine (25 ng/ml) but much lower levels of PGD2 (1.6 ng/ml; p less than 0.05). The time course of appearance of these mediators was identical in both groups of patients. The fluids from unchallenged blisters of allergic and nonallergic patients and the fluids of nonallergic patients challenged with ragweed had similar levels of histamine, at the lower limit of detection, and undetectable PGD2 levels. The peak levels of PGD2 in allergic individuals correlated with the size of the LPR lesion (p less than 0.05). These data suggest that the LPR involves the secondary elaboration of mediators different from mediators responsible for the immediate manifestations of the allergic skin reaction.     

Human recombinant lymphokines and cytokines induce pulmonary eosinophilia in the guinea pig which is inhibited by ketotifen and AH 21-132

Subcutaneous or intraperitoneal injection of recombinant human granulocyte-macrophage colony-stimulating factor, interleukin 3, or mouse tumour necrosis factor alpha, but not recombinant human interferon gamma, platelet-derived growth factor, or transforming growth factor beta caused selective eosinophilia of the pulmonary airways in the guinea pig. Unlike responses to platelet-activating factor, there was no attendant detectable airway hyperreactivity, but in common with responses to platelet-activating factor, eosinophilia of the airways was prevented by pretreatment with ketotifen or AH21-132. Cytokines or lymphokines may contribute to pulmonary eosinophilia in diseases such as asthma.     

OFF responses in the auditory thalamus of the guinea pig

ON and OFF auditory responses were examined in the medial geniculate body (MGB) of the guinea pig. Single- and multiunit recordings were carried out on 12 anesthetized animals, and noise-burst or pure-tone stimuli were applied to the ear contralateral to the recording hemisphere. One hundred and thirty-five OFF or ON-OFF neurons and 160 ON neurons were studied, and the tuning curves of 21 ON-OFF or OFF neurons were examined from various nuclei of the MGB. The mean minimum threshold of the OFF responses (40.8 +/- 20.0 dB SPL, mean +/- SD; range: 0-80 dB SPL) was significantly higher than that of the ON responses (28.5 +/- 17.6 dB SPL, range: 0-60 dB SPL; n = 17, P < 0.001). Of 10 ON-OFF neurons that showed identifiable tuning frequencies for both ON and OFF responses, 7 showed a higher OFF than ON best frequency (BF), 2 showed the same BF for both ON and OFF, and only 1 showed a slightly lower OFF than ON BF. Most OFF responses sampled from the borders of the ventral (MGv) and the rostromedial (MGrm) nuclei of the MGB showed single-peaked tuning curves, similar to those of the ON responses in the MGv. The neurons located in the shell (MGs) and dorsal (MGd) nuclei of the MGB showed complicated-either multi-peaked or broad-tuning curves. All OFF responses showed long-duration-selectivity for acoustic stimuli: the mean half-maximum duration was 116.5 +/- 114.8 ms (n = 19, range: 27-411 ms). The latencies of 135 OFF responses were studied in various divisions of the MGB. The ventral border region of MGv showed the shortest latency, followed by the dorsal border region of the MGv, the MGrm, and the caudomedial nucleus (MGcm) of the MGB. The posterior nucleus of the thalamus (Po), the MGd, and the MGs showed much longer mean latencies of >30 ms (P < 0.05 compared with the border regions of the MGv, ANOVA), with Po showing the greatest mean latency of 60.3 ms and the greatest deviation of 25.5 ms). The latency of the OFF response (29.0 +/- 14.0 ms, n = 135) was significantly greater than that of the ON response (15.6 +/- 9.6 ms, n = 160, P < 0.001). The present results provide valuable information about the threshold, frequency tuning characteristics, minimal response latency, and duration selectivity of OFF neurons in the auditory thalamus.     

Aujeszky's disease in the guinea pig: Cellular and humoral responses following immunization

Summary The fatal disease caused by virulent ADV in guinea pigs was found to be identical to that seen in sheep and cattle. Intramuscular (i.m.) injection of an avirulent strain of ADV (Bartha) yielded better immunity to challenge after 3 weeks than did intranasal (i.n.) immunization, and this was reflected in differences in histopathological changes in the brain.Serum antibodies active in antibody-dependent cell-mediated cytotoxicity (ADCC) were titrated using polymorphonuclear leukocytes as effector cells. ADCC correlated fairly well with virus neutralization and was a far more sensitive technique. There was good, but not complete, correlation between ADCC and protection. Lymphocyte responsiveness to virus antigensin vitro was assessed by³H-thymidine uptake and lymphokine tests. Lymphocyte stimulation and mitogenic factor responses were low grade but blood lymphocyte stimulation was more pronounced in the better-protected animals. Macrophage migration inhibition correlated neither with serum ADCC nor with protection, being equally demonstrable in the two immunized groups.This work was carried out while this establishment under its former title Microbiological Research Establishment was under the management of the Ministry of Defence.     

Inhibitory effect of TMK-688 on late asthmatic responses as well as T-cell and eosinophilic infiltration in guinea pigs with asthmatic reactions

The effects of an oral anti-allergic agent, TMK-688, which inhibits 5-lipoxygenase, at doses of 3.2 and 10 mg/kg were studied in guinea pigs with dual-phase asthmatic response. We previously observed that pretreatment with TMK-688 inhibited the late asthmatic response (LAR) induced by ovalbumin inhalation exposure. The present study focused on the effect of TMK-688 on infiltration by T-cells and eosinophils. TMK-688 inhibited both T-cell and eosinophilic infiltration. These findings suggest that TMK-688 is effective in inhibiting infiltration of T-cells and eosinophilic chemotaxis, and thereby suppresses LAR.     

Specific adrenergic responses of smooth muscles in the vascular wall of guinea pig pulmonary arteries during ovalbumin sensitization

The adrenergic contractile responses of smooth muscles in the vascular wall of guinea pig pulmonary arteries were studied during ovalbumin sensitization. Sensitization was followed by inhibition of contractile responses to an α-adrenoceptor agonist mesatone, prevented endothelium-derived relaxation, and potentiated the contractile response to isoproterenol. Administration of a β-adrenoceptor agonist isoproterenol potentiated the increase in mechanical strain of smooth muscles in the pulmonary artery precontracted with high-potassium Krebs solution. Removal of the endothelium had no effect on the contractile response of smooth muscle segments from the pulmonary artery of intact and sensitized guinea pigs to β-adrenergic influences. The contractile responses of smooth muscles of the pulmonary artery are associated with activity of the cAMP-dependent signal system and play a role in the pathogenesis of ventilation-perfusion disturbances during atopic inflammation. __________ Translated from Byulleten' Eksperimental'noi Biologii i Meditsiny, Vol. 145, No. 6, pp. 617–620, June, 2008     

Lack of late-phase airway responses in conscious guinea pigs after a variety of antigen challenges

Guinea pigs were actively sensitized by parenteral injections of ovalbumin (OA), house dust extract (HD) orAscaris suum extract (As) in a variety of multidose regimens. At least 3 weeks after the initial sensitization injection, aerosols of the appropriate antigen were administered to conscious guinea pigs in a double-chamber body plethysmograph. OA elicited the most consistent and intense bronchoconstriction (BC) as measured by decreases in specific airway conductance (sGAW). The airway responses to As were clearly separable into responders and nonresponders. HD produced essentially no BC. However, intense lacrimation and rhinorrhea occurred in all HD-sensitized, but not unsensitized, animals. No late-phase changes in sGAW or increased reactivity to other spasmogens were seen up to 8h after any antigen challenge. Eosinophil influx of magnitude similar to that measured by 24h post-antigen bronchoalveolar lavage (BAL) occurred with all the three antigens. Animals which did not bronchoconstrict to As experienced an equal or greater pulmonary eosinophilia as airway responders. The present data with HD and As suggest that acute BC in response to antigen provocation is not a prerequisite for the eventual pulmonary eosinophilia. The lack of late-phase airway reactions in these models raises a doubt in the direct extrapolation to airway responses in allergic human asthma. The acute lacrimation and rhinorrhea to HD may suggest utility as a model of allergic rhinitis.     

Effect of San-Ao-Tang on immediate and late airway response and leukocyte infiltration in asthmatic guinea pigs

San-Ao-Tang (SAT), a traditional Chinese medicines, has been used to treat patients with the bronchial asthma for several centuries. However, the therapeutic mechanisms of this Chinese medicine are still far from clear. To understand the mechanism of antiasthmatic property of SAT, a guinea pig model of allergic asthma was used to investigate the effects of SAT on Dermatophagoides pteronyssinus-induced. immediate and late asthmatic responses and airway inflammation. Our results showed that administration of SAT (10g/kg) extracts singificantly inhibited the antigen induced immediate asthmatic responses (IAR) in actively sensitized guinea pig. Examination of bronchoalveolar lavage fluid (BALF) revealed that SAT significantly inhibited the increase in neutrophil in the airway at 1, 2, 4, 6, 8 hr after antigen challenge. Histopathologic examination showed SAT suppressed the neutrophil infiltration into lung tissue. These results suggest that the antiasthmatic effect of SAT be mainly due to its bronchodilator effect and its ability to inhibit the neutrophil into the airway. The precise mechanism of action of SAT in asthma remains to be elucidated.     

卵蛋白诱发哮喘发作时豚鼠大脑和肺内Fos蛋白的表达

为了解支气管哮喘后豚鼠大脑和肺组织c-fos基因表达的变化，探讨c-fos表达在豚鼠哮喘发病中的可能意义。我们复制卵蛋白致敏哮喘豚鼠模型，采用免疫组织化学ABC方法，对Fos蛋白在大脑和肺脏内的分布情况进行观察。结果发现：哮喘组豚鼠大脑和肺内c-fos表达较对照组明显增加，其Fos阳性产物在大脑主要分布于额顶皮质、边缘前脑（扣带皮质、梨状皮质和中央杏仁核等）、丘脑室旁核、下丘脑室旁核、视上核、下丘脑外侧区、下丘脑室周核、孤束核、最后区和延髓腹外侧区内，小脑内无明显Fos分布密集区。原癌基因c-fos的表达增强在豚鼠支气管哮喘发病过程中可能起一定作用。     

Vestibular primary afferent responses to sound and vibration in the guinea pig

This study tested whether air-conducted sound and bone-conducted vibration activated primary vestibular afferent neurons and whether, at low levels, such stimuli are specific to particular vestibular sense organs. In response to 500 Hz bone-conducted vibration or 500 Hz air-conducted sound, primary vestibular afferent neurons in the guinea pig fall into one of two categories--some neurons show no measurable change in firing up to 2 g peak-to-peak or 140 dB SPL. These are semicircular canal neurons (regular or irregular) and regular otolith neurons. In sharp contrast, otolith irregular neurons show high sensitivity: a steep increase in firing as stimulus intensity is increased. These sensitive neurons typically, but not invariably, were activated by both bone-conducted vibration and air-conducted sound, they originate from both the utricular and saccular maculae, and their sensitivity underpins new clinical tests of otolith function.     

Cardiovascular and metabolic responses during burn shock in the guinea pig.

The hemodynamic and metabolic responses of fatally burned, nonfatally burned, and unburned control guinea pigs were compared. The burns were induced in temporarily anesthetized animals by immersion to either the xyphoid process (70% fatal) or the midabdomen (100%survival) in boiling water for 3 s. Although cardiac output was reduced in all animals postburn, the survivors (MAG) has higher cardiac outputs at lower arterial pressures than the nonsurvivors (XPN). The postburn lactate levels in the XPN were higher than in the MAG, and the postburn values for pH, oxygen consumption, and core temperature were lower in the XPN. In each group, hyperglycemia was evident for 8 h postburn and terminal plasma glucose concentrations were usually elevated or similar to the prevalue. It was concluded that fatal and nonfatal burn shock were distinguished primarily by differences in tissue perfusion.     

A23187-induced pulmonary gas trapping and inflammation in the guinea pig

A brief A23187 aerosol exposure produced prolonged airway obstruction with granulocyte accumulation in conscious guinea pigs. Aminophylline, atropine, pyrilamine, salbutamol, SC-41930 (a leukotriene B₄ antagonist) and WEB 2086 (a platelet activating factor antogonist) were administered intravenously (i.v.) to evaluate their ability to prevent these changes. Inhaled salbutamol was also assessed. Aminophylline, atropine, and salbutamol (i.v. and aerosol) inhibited A23187-induced gas trapping (p<0.01). However, pyrilamine, SC-41930 and WEB 2086 did not influence this airway obstructive effect. Only atropine, inhaled salbutamol and SC-41930 inhibited the cell influx (p<0.01), while pyrilamine potentiated the inflammation (p<0.05). We conclude that A23187 produces a sustained bronchospasm and an intense granulocyte accumulation. The treatment agents tested differ considerably in their ability to alter A23187-induced airway obstruction and inflammation.