CItnicul ami E.xperimentul Allergy. 1992. Volume 22. pages 75-81
Protection of nedocromil sodium on bronchoconstriction induced by inhaled neurokinin A (NKA) in asthmatic patients N. CRIMI, F. PALERMO, R. OLIVERI, B. PALERMO. R. POLOSA and A. MISTRETTA Institute of Respiratory Diseases. University of Catania. Italy Summary
Neurokinin A (NKA) has been shown to exert a potent contractile action on bronchial smoolh muscles both in vitro and in vivo. Although this etTeci seems to be due either to a direct action of this peptide on specific muscular receptors or to an indirect eflect on mast cells and/or nerves, its mechanism of action in bronchial asthma is still unknown. In the present study we have investigated ihc airway response to inhaled NKA in 10 asthmatic subjects and Ihe activity of the novel pyranoquinoline dicarboxylicacid drug, nedocromil sodium, on ihis response. Ten asthmatic patients with stable aslhma took part in the study consisting of four separate visits. On the Hrst two occasions we derived histamine and NKA PDii values in absence ofany drug treatment. On the following two visits the inhalation challenge with NKA was performed aflcr administration oleither nedocromil sodium or matched placebo administered as pressurized aerosols via metered dose inhalers in a randomized double-blind order. Inhaled NKA produced a dose-related fall in FEV| in all the subjects studied. Inhaled nedocromil sodium had a significant effect on the FEVi response to NKA inhalation, the geometric mean PD15 value increasing from 16-6 to 32 2 x 10"'' mol. We conclude that nedocromil soditim attenuates subsequent responsiveness to inhaled NKA in asthmatic subjects. However, further studies arc necessary lo investigate the exact role and tbe mode of action of tacbykinins in human airways. Clinical and E-xperimental Allergy, Vol. 22, pp. 75 81. Received 11 February 1991; revised 24 July 1991: accepted 29 July 1991. Introduction
The importance of neuropeptides in bronchial tone regulation and in the pathogenesis of bronchial asthma has been stressed in several reports [1-5]. Immunochemical studies [6.7]. have demonstrated the presence of neuropeptides in pulmonary tissue, giving the possibility that its presence could be related to the regulation of bronchomotor lone. The presence of substance P (SP) and other related peptides (i.e. neurokinin A, CGRP) has been demonstrated in the small unniyelinaled fibres both in central and in peripheral sensory nerves endings [6.8.9]. In addition, immunohistochemical studies [6,8.10] demonstrated thai submucosal glands. Correspondence: Dr N. Crimi. Instituie of Respiratory Diseases. L'niversity tif Catania. Via Passo Gravin;i. 187, 95125 Catania, llaly.
and pulmonary arteries and veins, are both innervated by peplidergic nerves. Neurokinin A (NKA) exerts a potent contractile action on guinea pig bronchial smooth muscles both in vitro and //( vivo [1.11.12]. In addition NKA appears to be particularly active in human isolated bronchi [13]. possibly via stimulation of a NK-2 receptor type [14.15]. In guinea-pig airways and isolated human airways the conslriclor effect of tachykinins was initially reported lo be direct [16,17]. However, sodium cromoglycate and nedocromil sodium significantly reduced the bronchoconslricLor response to NKA in rals [IS]. Similar lindings were obtained in mild asthmatic subjects, thus suggesting that the mechanism of action of NKA-induced bronchoconstriclion in aslhma is largely due to an indirect pathway and ihis could be mediated by an effect on inllammatory cells and/or nerves [19.20]. 75
76
A'. Crinti et al.
Tabie I. Patients characteristics Patient no. 1 2 3 4
5 6 7
8 9 10 Mean ±s.d.
Age Height Weight "A. FFVI NKA PD,^ predicted (X 10 •'mol) Sex (years) (cm) (kg) M F F F F M F M F M
25 43 27 18 27 16
41 18 23 39 27-7 99
175 154 164 165 164 169 165 174 161 167
69 70 65 65 60 65 67 67 52 66
82 82 98 79 97
165 6
64 5
89 7
15 9 293 123 56 17'2 36 5 10 3 119 6-0 5-0
9!
89 99 97 83
Skin-test
Histamine PD,5 (mg)
G-W DW W-G D G-W W G W D W-G
004 006 003 0-03 006 0-04 006 006 004 004
12-2* 7-5-19-8**
0'046* 004-006**
G, grass; W, wall pellitory; D. dermalophagoides. * Geometric mean. ** 95'^^ confidence limits.
Nedocromil sodium, the disodium salt o f t h e pyranoquinoline dicarboxylic acid, is a drug which has been proposed as adjuvant treatment of bronchial asthma [21]. Its administration prevents early and late allergic reactions in asthmatic allergic patients [22]. In addition to attenuating mast cell activation, nedocromil sodium also possesses the ability to inhibit non-cholinergic neural reflexes [21]. Thus if it is true that tachykinins arc able to induce a contractile effect via indirect pathways, then nedocromil sodium should attenuate the airway response induced by inhaled N K A . This study was conducted in order to observe the airway response to inhaled N K A in asthmatic subjects and to evaluate the protective effect of nedocromil sodium, on NKA-induced bronchoconstriction. Materials and methods Subjects Ten asthmatic patients (four males and six females; age ranging from 16 to 43 years) took part in the study. All the patients had a history of dyspnoea with wheezing or tightness after exposure to airborne allergens. All were non-smokers and had positive skin prick tests to one of more common aeroallergens (Table 1). At the beginning of the study all patients were asymptomatic with values for the forced expiratory volume in one second (FEV|) not less than 80*^ of the predicted normal values (Table 1).
None o^ the patients had recently acute respiratory infections or other respiratory diseases. Inhaled beta^agonists. and theophylline were withheld for at least 48 hr before the beginning of the trial. None of them were receiving sodium cromoglycate. oral or inhaled steroids and antihistamines within the previous month. Neurokinin A challenge Neurokinin A (Peninsula Laboratories Belmont, California. U.S.A.). was diluted in lO'V;. albumin in order to produce doubling doses ranging from 5 75 to 92 x 10"^ mol. It was administered as an aerosol generated by a nebulizer (DeViibiss 646 De Vilbiss. Somerset. Pennsylvania. U.S.A.) giving 2 ml of solution at an airflow of 121/ min. The output for each breath was 16 6 6 / 1 and the particle size mass diameter was between 1 5 and 3 5 / m . Each dose was administered through a mouthpiece, while wearing a nose-clip, by 15 tidal breaths from a compressed air nebulizer controlled by a dosimeter (Mefar— Brescia Mefar, Brescia, Italy). The nebulized solution was generated only during inspiration time by a microcomputer which was linked to a solenoid valve. The inhalation (0 8 sec) and exhalation (1-6 sec) times had been previously standardized, Airway response was measured as FEV| using a Fleish pncumotachograph (Pulmonary System 47I20A Hewlett Packard instrument Fort Collins. Colorado. U.S.A.). F E V I was initially measured at 1. 3.5. 10 and 15 min after
Nedocromil sodium on hronchoconstriction induced bv inhaled NKA
the administration of each dose, and thereafter every 5 min until a value higher than the previous one was found. We slopped the inhalations when FEV| had decreased by more than 15'^ of the value obtained after control solution inhalation (10% albumin Sigma A-8763, St Louis. Missouri. U.S.A.). The dose of NKA which produced a 15% change in FEV| (PD15) was calculated using the individual semilogarithmic dose-response curve. Airway response was also evaluated by the measurement of the area under the dose-response curve which was calculated by the integration of the least-squares fitted, parabolic dose-response curve to NKA inhalation; provocative dose at 15% (PD15) is the right-hand limit (xmax) of the integral [23].
Table 2. FEVi mean ( + s.d.) values (I) on each study day
Pre-drug
A%
Table 3. NKA PD,.. values ( x 10 " mol) to NKA after placebo and nedocromil treatment
Patient no. 1
Subjects attended the laboratory on four separate visits at the same time each morning. On the first visit, in the absence of any drug treatment, ihe patients attended to undertake a dose response study with inhaled histamine (doubling doses from 003 to 4 mg); the provocative dose of histamine producing a 15% fall in FEV| from the post-saline baseline value (PD|s) was derived. On the second study day, NKA was inhaled in order to determine baseline PD|s value. On the following two visits (not less than 72 hr aparl) patients undertook dose-response studies with inhaled NKA 30 min after treatment with either nedocromil sodium (2 mg x 2 actuations) or placebo in a randomized double blind manner. On each occasion three baseline measurements of FEVi were recorded at intervals of 3 min. followed by the administration of nedocromil sodium or matched placebo. Nedocromil sodium and placebo were delivered by a pressurized aerosol via MDI and the inhalation of pressurized aerosols was conducted by one of the conducting investigators (RO). The MDI was held 2 cm away from the mouth and pressed during a slow, deep inspiration with a breath holding pause of 10 sec; the second administration followed 30 sec after the first pufl". The protocol was approved by the Ethics Review Committee of the Department of Respiratory Diseases {University of Catania) and the consent was obtained from each subject after the nature and reason for the study had been explained in detail.
Post-drug
Baseline 3-02(±0'54) Placebo 3-02 ( ± 0 4 5 ) 3-04(±058) 0-21 ( ± 3 3) Nedocromi! sodium 302 ( ± 0 5 6 ) 3-05 ( ± 0 57) 0-82 ( ± 1 4 )
Baseline
Placebo
8
15 9 293 I2'3 56 172 36 5 10'3 119
9 10
6-0 5-0
19-9 21 5 5-7 5-9
12-2 7'5-198
16-7 7'7-36-3
2
Study design
77
3 4 5 6 7
GM 95"., confidence limit
110
392 7-9 5-0 769
>920
Nedocromil >92 0 524 >92-0 74 >92 0 60 5 47-5 38 5 62 50 32-2* 14-2 73-4
*P-00019). In four cases, nedocromil sodium pretreatment doubled the values obtained after placebo administration (Table 4). Discussion In a previous study [24] we have demonstrated the protective elfect exerted by nedocromil sodium on SPinduced bronchoconstriction in asthma. The results ofthe present investigation confirm previous data showing the bronehoconstrictor activity of inhaled NKA [5.25] and the significant protection afiTorded by nedocromil sodium against the airway response to this neuropeptlde [19], Although NKA was effective in inducing bronchospasm. bronchial response to this peptide was not related to the level of non-specific bronchial hyper-reactivity to histamine in these patients in agreement with previous reports [19]. The lack of correlation cannot account for differences in hislamine baseline lung function on the study days and it could suggest a diiferent mechanism of action of these agents involving different pathways of bronchial response. The mechanism by which NKA, or other tachykinins, induce bronchospasm is still unclear. A possible involvement of mast cell activation in this response has been suggested by the large inhibitory effect of methysergide. a selective serotonin antagonist, on the bronchial response to NKA in the rat [18]. In addition in the same animal model both sodium cromoglycate and nedocromil sodium attenuated the bronchospastic response to NKA [18]. Both drugs seem to inhibit mediator release from a range of cell types including mast cells [21]. However, in dogs, they also possess the ability to inhibit non-cholinergic neural reflexes from stimulation of sensory nerves [26.27]. in human dispersed skin mast cells. NKA caused significant concentration-related histamine release although considerably less potent than substance P [28]. In support of this. Fuller et al. [29] have shown that in man the Hphistamine antagonist terfenadine (60 mg) significantly inhibited the weal and flare response to intradermally injected NKA. A different response to the same peplidergic stimuli may be elicited in mast cells harvested from different tissues. Indeed SP stimulation causes histamine release from cutaneous mast cells but not from pulmonary mast cells in vitro [30]. In contrast, recent studies have demonstrated histamine release in
80
A'. Cr/m/etal.
bronchoaiveolar lavage fluid recovered after adenosine and NKA infusion in a rat model [31] thus supporting the view that airways mast cell activation may follow tachykinin exposure and hypothesize an indirect mechanism of neuropeptide-induced bronchospasm. However, our recent observations (N. Crimi, unpublished data) have shown that pharmacological pretreatment with terfenadine (180 mg) failed to protect the airways of 10 asthmatic subjects against the constrictor effect of NKA. These data question the role of endogenous histamine as mediator of NKA-induced bronchoconstriction in human asthma. We have also demonstrated that premedication with the potent anti-allergic compound astemizole did not protect against substance P-induced bronchoconstriction [32]. In the same study the bronchospastic response induced by inhaled substance P was weakly inhibited by the anticholinergic drug ipratropium bromide. These findings suggest that activation o\' a cholinergic component rather than release of mast cell mediators in SPinduced bronchospasm. Stimulation of post-ganglionic nerve endings on rodents' airways demonstrated the release of acetylcholine and suggested that the bronchoconstrictor activity of tachykinins may be due to a cholinergic activation [8,18]. Furthermore eapsaicin, the pungent agent coming from hot peppers, is capable of inducing smooth muscle contraction and subepithelial oedema, possibly via liberation of SP and other neuropeptides from some primary atlerent neurons [2]. In human airways in vitro capsaicin exposure induces a slight and transient contraction which is inhibited by anticholinergic drugs [17]. In this study, nedocromi! sodium administration gave a significant protection when NKA was inhaled. This drug, introduced as a prophylactic treatment for inhaled use in asthma, has no intrinsic bronchodilator activity and against a range of acute challenges with several nonspecific bronchoconstrictor stimuli such as SOi [33]. nebulized distilled water (fog) [34], cold dry air [35], adenosine [36] and bradykinin [37] exerts a potent inhibitory effect probably by an action exerted on the afTerent reflexes arising from stimulation of lung irritant receptors. Protective action given by nedocromil sodium treatment against NKA could be determined by its neural activity on vagai fibres. Although anatomical, pharmacological and functional studies, conducted both in rivo and in vitro, attribute to neuropeptides an important role in the pathophysiology of bronchial asthma, the constrictor response to inhaled NKA in asthmatic airways does not necessarily imply that ihis peptide may have a specific role in the pathogenesis of bronchial asthma. The mechanism of action of various tachykinins, including NKA and SP, could be due to a cholinergic reflex activation, or to a
direct aetion exerted on specific receptors of bronchial smooth muscle rather than to mast cell mediator release. Further studies are necessary to investigate the exact role and the mode of action of tachykinins in human airways. References 1 Uchida Y, Nomura A, Ohlsuka H, Hasegawa S. Goto K, Kimura S, Sugita Y. Uchiyiama Y. Neurokinin A as a potent bronchoconslrictor. Am Rev Respir Dis 1987; 136:718-21. 2 Lundberg JM. Saria A. A bronchia! smooth muscle contraction induced by .stimulation of capsaicin sensitive sensory neurons. Acta Physiol Scand 1982: 116:473-6. 3 Barnes PJ. Asthma as an axon reflex. Lancet 1986: i:242-5. 4 Tanaka DT, Grunstein MM. Mechanisms of substance Pinduced contraction ofrabbit airway smooth muscle. J Appl Physiol 1984:57:1551 7 5 Joos G. Pauwels R. Van Der Straeten M. The effect of inhaled substance P and neurokinin A on the airway of normal and asthmatic subjects. Thorax 1987; 42:779-83. 6 Lundberg JM. Hokfelt T. Martling CR, Saria A, Cuello C. Substance P immunoreaetive sensory nerves in the lower respiratory iraci of various mammals including man. Cell Tissue Res 1984; 235:251-61 7 Saria A.Theodorsson-Norheim E. Gamse R, Lundberg JM. Release ofsubstance P and substance K-Iikc immunoreaclivities from the isolated perfused guinea-pig lung. Eur J Pharmacol 1985; 106:207 8. 8 Terenghi G. McGregor GP. Ghultacharji S. Wharton J, Bloom SR. Polak JM. Vagal origin ofsubstance P-containing nerves in lhe guinea pig lung. Neurosci Letl 1983: 36:229-36. 9 Lundberg JM. Franco-Cereceda A, Hua X. Hokfelt T. Fischer J A. Co-existence ofsubstance P and ealeitonin generelated peptide-like immunnreactivities in sensory nerves in relation to cardiovascular and bronchoconstrictor effects of capsaicin. Eur J Pharmacol 1985; 108:315-9. 10 Nilsson G., Dahlberg K. Brodin E. Sundler F. Strandberg K. Distribution and constrictor effect of substance P in guinea-pig tracheal bronchial tissue. In: Von Euler US. Pernow B, eds. Substance P. New York: Raven Press. 1977:75-81. 11 Theodorsson-Norheim E. Hua X, Brodin E. Lundberg JM Capsaiein treatment decreases tissue levels ol neurokinin Alike immunoreactivity in ihc guinea-pig. Acta Physiol Scand 1985: 124:129-31. 12 Karlsson JA, Einney MJB, Persson CGA, Post C. Substance P antagonists and the roleoflachykinins in non-cholinergic bronchoconstriction. Life Sciences 1984; 35:2681-91. 13 Martling CR. Theodorsson-Norheim E, Lundberg JM. Occurrence and effects of multiple taehykinins: substance P, neurokinin A and neuropeptide K in human lower airways. Life Sciences 1987.40:1633 43. 14 Advenier C. Naline E. Drapeau G. Regoii D. Relative potencies of neurokinins in guinea pig trachea and human bronchus. Eur J Pharmacol 1987; 139:233-7.
Nedocromil sodium on bronchoconstriction induced bv inhaled NKA
15 Naline E, Devillier P, Drapeau G, Toly L. Bakdach H. Regoli D, Advenier C. Charaeterization of ncurokinin effeets and receptor selectivity in human isolated bronchi. Am Rev Respir Dis 1989; 140:679-86. 16 Niisson G, Dahlberg K. Brodin F.. Sundler F. Strandberg K. Distribution and constrictor effect of Substance P in guinea pig tracheobronchial tissue. In: von Euler US, Pernow B, eds. Substance P. New York: Rav^^n Press, 1977:75-81. 17 Lundberg JM. Martling CR. Saria A. Substance P and capsaicin-induced contraction of human bronchi. Acta Physiol Scand 1983; 119:49 53. 18 Joos GF. Pauwels RA. Van Der Stractcn ME, The mechanism of tachykinins induced bronchoconstriction in the rat. Am Rev Respir Dis 1988; 137:1038-44. 19 Joos GF. Pauwels RA. Van Der Straeten ME. The elTecl of nedocromil sodium on the bronchoconstrictor effect of neurokinin A in subjeets with asthma. J All Clin Immunol 1989; 83:663-8. 20 Joos GF\ Pauwels RA- Mechanism involved in neurokininindueed bronchoconstriction. Arch Inl Pharmacodyn 1990; 303:132-46. 21 Church MK, Polosa R, Rimmer JS. Cromolyn sodium and nedocromil sodium: mast cells stabilizers, neuromodulators or anti-inflammatory drugs? In: Kaliner MA, Barnes PJ, Persson CGA, eds. Asthma: its pathology and treatment. New York: Marcel Dekker, 1991:541 69. 22 Dahl R, Pcderscn B. Influence of nedocromil sodium on the dual asthmatic reaction after allergen challenge: a double blind, placebo controlled study. Eur J Respir Dis 1986; 69 (suppl. l47):263-5. 23 Townley RG, Bewtra AK. Nair NM. Brodkey FD, Watt GD, Burke KM. Methacholine inhalation challenge studies. J All Clin Immunol 1979; 64:569 74. 24 Crimi N. Palermo F. Olivcri R. Palermo B. Vyncheri C, Polosa R. Mistretta A. Effect of nedocromil on bronchospasm induced by inhalation of substance P on asthmatic subjects. Clin Allergy 1988; 18:375. 25 Evans TW, Dixon CMS. Clarke B. Conradson TB, Barnes PJ. Comparison of neurokinin A and substance P on cardiovascular and airway function in man. Br J Pharmac 1988; 25:273 5. 26 Jackson DM. Richards IM. The effect of sodium cromoglyeate on histamine aerosol induced reflex bronchocon-
27
28
29
30
31
32
33
34
35
36
37
81
striction in the anaesthetized dog. Br J Pharmacol 1977; 61:257-67. Kauffman MP, Coleridge HM, Coleridge JCG, Baker DG. Bradykinin stimulates afferent vagal C-fibrcs in intrapulmonary airways of dogs. J Appl Physiol 1980; 48:51 i 7. Lowman MA. Bcnyon RC, Church MK. Characterization of neuropeptide-induced histamine release from human dispersed skin mast cells. BrJ Pharmacol 1988; 95:121-30. Fuller RW, Conradson TB, Dixon CMS, Crossman DC, Barnes PJ- Sensory neuropeptide effects in human skin. BrJ Pharmacol 1987; 92:781 8. Ali H, Leung KBP. Pearee FL. Hayes NA, Foreman JC. Comparison ofthe histamine releasing action of substance P on mast cells and basophils from different species and tissues. Int Arch Allergy Appl Immunol 1986; 79:413-8. Pauwels R. Joos G. Kips J. Van Der Straeten M. Adenosine and Ncurokinin A cause histamine release in the airways. Am Rev Respir Dis 1988; 137:A2I2. Crimi N. Palermo F. Oliveri R. Palermo B. Vancheri C. Polosa R. Mistretta A. Influence of antihistaminc (Astemizole) and anticholinergic drugs (Ipratropium bromide) on bronchoconslriction induced by substance P. Annals of Allergy 1990; 65:115-20. Fuller RW. Dixon CMS. Barnes PJ. Nedocromil sodium inhibits sulfur dioxidc-indticcd bronchoconstriction. Am Rev Rcspir Dis 1986; 55:133Robuschi M. Vaghi A. Simone P. Bianco S. Prevention of fog-induced bronehospasm by nedocromil sodium. Clin Allergy 1987; 17:69. Juniper EF. Kline PA. Morris MM. Hargreave FE. Airway constriction by isocapnic hyperventilation of cold dry air: comparison of magnitude and duration of protection by nedocromil sodium and sodium cromoglycate. Clin Allergy 1987; 17:523Crimi N. Palermo F, Oliveri R. Vanchcri C. Polosa R, Palermo B. Maccarrone C. Mislretta A. Comparative study of the effects of nedocromil sodium (4 mg) and sodium cromoglycate (10 mg) on adenosinc-indiiced bronchoconstriction in asthmatic subjects. Clin Allergy 1988; 18:367-74. Dixon CMS. Barnes PJ. Bradykinin induced bronchoconstriction: inhibition by nedocromil sodium and sodium cromoglycate. Br J Clin Pharmacol 1989; 27:831-6.
Protection of nedocromil sodium on bronchoconstriction induced by inhaled neurokinin A (NKA) in asthmatic patients N. CRIMI, F. PALERMO, R. OLIVERI, B. PALERMO. R. POLOSA and A. MISTRETTA Institute of Respiratory Diseases. University of Catania. Italy Summary
Neurokinin A (NKA) has been shown to exert a potent contractile action on bronchial smoolh muscles both in vitro and in vivo. Although this etTeci seems to be due either to a direct action of this peptide on specific muscular receptors or to an indirect eflect on mast cells and/or nerves, its mechanism of action in bronchial asthma is still unknown. In the present study we have investigated ihc airway response to inhaled NKA in 10 asthmatic subjects and Ihe activity of the novel pyranoquinoline dicarboxylicacid drug, nedocromil sodium, on ihis response. Ten asthmatic patients with stable aslhma took part in the study consisting of four separate visits. On the Hrst two occasions we derived histamine and NKA PDii values in absence ofany drug treatment. On the following two visits the inhalation challenge with NKA was performed aflcr administration oleither nedocromil sodium or matched placebo administered as pressurized aerosols via metered dose inhalers in a randomized double-blind order. Inhaled NKA produced a dose-related fall in FEV| in all the subjects studied. Inhaled nedocromil sodium had a significant effect on the FEVi response to NKA inhalation, the geometric mean PD15 value increasing from 16-6 to 32 2 x 10"'' mol. We conclude that nedocromil soditim attenuates subsequent responsiveness to inhaled NKA in asthmatic subjects. However, further studies arc necessary lo investigate the exact role and tbe mode of action of tacbykinins in human airways. Clinical and E-xperimental Allergy, Vol. 22, pp. 75 81. Received 11 February 1991; revised 24 July 1991: accepted 29 July 1991. Introduction
The importance of neuropeptides in bronchial tone regulation and in the pathogenesis of bronchial asthma has been stressed in several reports [1-5]. Immunochemical studies [6.7]. have demonstrated the presence of neuropeptides in pulmonary tissue, giving the possibility that its presence could be related to the regulation of bronchomotor lone. The presence of substance P (SP) and other related peptides (i.e. neurokinin A, CGRP) has been demonstrated in the small unniyelinaled fibres both in central and in peripheral sensory nerves endings [6.8.9]. In addition, immunohistochemical studies [6,8.10] demonstrated thai submucosal glands. Correspondence: Dr N. Crimi. Instituie of Respiratory Diseases. L'niversity tif Catania. Via Passo Gravin;i. 187, 95125 Catania, llaly.
and pulmonary arteries and veins, are both innervated by peplidergic nerves. Neurokinin A (NKA) exerts a potent contractile action on guinea pig bronchial smooth muscles both in vitro and //( vivo [1.11.12]. In addition NKA appears to be particularly active in human isolated bronchi [13]. possibly via stimulation of a NK-2 receptor type [14.15]. In guinea-pig airways and isolated human airways the conslriclor effect of tachykinins was initially reported lo be direct [16,17]. However, sodium cromoglycate and nedocromil sodium significantly reduced the bronchoconslricLor response to NKA in rals [IS]. Similar lindings were obtained in mild asthmatic subjects, thus suggesting that the mechanism of action of NKA-induced bronchoconstriclion in aslhma is largely due to an indirect pathway and ihis could be mediated by an effect on inllammatory cells and/or nerves [19.20]. 75
76
A'. Crinti et al.
Tabie I. Patients characteristics Patient no. 1 2 3 4
5 6 7
8 9 10 Mean ±s.d.
Age Height Weight "A. FFVI NKA PD,^ predicted (X 10 •'mol) Sex (years) (cm) (kg) M F F F F M F M F M
25 43 27 18 27 16
41 18 23 39 27-7 99
175 154 164 165 164 169 165 174 161 167
69 70 65 65 60 65 67 67 52 66
82 82 98 79 97
165 6
64 5
89 7
15 9 293 123 56 17'2 36 5 10 3 119 6-0 5-0
9!
89 99 97 83
Skin-test
Histamine PD,5 (mg)
G-W DW W-G D G-W W G W D W-G
004 006 003 0-03 006 0-04 006 006 004 004
12-2* 7-5-19-8**
0'046* 004-006**
G, grass; W, wall pellitory; D. dermalophagoides. * Geometric mean. ** 95'^^ confidence limits.
Nedocromil sodium, the disodium salt o f t h e pyranoquinoline dicarboxylic acid, is a drug which has been proposed as adjuvant treatment of bronchial asthma [21]. Its administration prevents early and late allergic reactions in asthmatic allergic patients [22]. In addition to attenuating mast cell activation, nedocromil sodium also possesses the ability to inhibit non-cholinergic neural reflexes [21]. Thus if it is true that tachykinins arc able to induce a contractile effect via indirect pathways, then nedocromil sodium should attenuate the airway response induced by inhaled N K A . This study was conducted in order to observe the airway response to inhaled N K A in asthmatic subjects and to evaluate the protective effect of nedocromil sodium, on NKA-induced bronchoconstriction. Materials and methods Subjects Ten asthmatic patients (four males and six females; age ranging from 16 to 43 years) took part in the study. All the patients had a history of dyspnoea with wheezing or tightness after exposure to airborne allergens. All were non-smokers and had positive skin prick tests to one of more common aeroallergens (Table 1). At the beginning of the study all patients were asymptomatic with values for the forced expiratory volume in one second (FEV|) not less than 80*^ of the predicted normal values (Table 1).
None o^ the patients had recently acute respiratory infections or other respiratory diseases. Inhaled beta^agonists. and theophylline were withheld for at least 48 hr before the beginning of the trial. None of them were receiving sodium cromoglycate. oral or inhaled steroids and antihistamines within the previous month. Neurokinin A challenge Neurokinin A (Peninsula Laboratories Belmont, California. U.S.A.). was diluted in lO'V;. albumin in order to produce doubling doses ranging from 5 75 to 92 x 10"^ mol. It was administered as an aerosol generated by a nebulizer (DeViibiss 646 De Vilbiss. Somerset. Pennsylvania. U.S.A.) giving 2 ml of solution at an airflow of 121/ min. The output for each breath was 16 6 6 / 1 and the particle size mass diameter was between 1 5 and 3 5 / m . Each dose was administered through a mouthpiece, while wearing a nose-clip, by 15 tidal breaths from a compressed air nebulizer controlled by a dosimeter (Mefar— Brescia Mefar, Brescia, Italy). The nebulized solution was generated only during inspiration time by a microcomputer which was linked to a solenoid valve. The inhalation (0 8 sec) and exhalation (1-6 sec) times had been previously standardized, Airway response was measured as FEV| using a Fleish pncumotachograph (Pulmonary System 47I20A Hewlett Packard instrument Fort Collins. Colorado. U.S.A.). F E V I was initially measured at 1. 3.5. 10 and 15 min after
Nedocromil sodium on hronchoconstriction induced bv inhaled NKA
the administration of each dose, and thereafter every 5 min until a value higher than the previous one was found. We slopped the inhalations when FEV| had decreased by more than 15'^ of the value obtained after control solution inhalation (10% albumin Sigma A-8763, St Louis. Missouri. U.S.A.). The dose of NKA which produced a 15% change in FEV| (PD15) was calculated using the individual semilogarithmic dose-response curve. Airway response was also evaluated by the measurement of the area under the dose-response curve which was calculated by the integration of the least-squares fitted, parabolic dose-response curve to NKA inhalation; provocative dose at 15% (PD15) is the right-hand limit (xmax) of the integral [23].
Table 2. FEVi mean ( + s.d.) values (I) on each study day
Pre-drug
A%
Table 3. NKA PD,.. values ( x 10 " mol) to NKA after placebo and nedocromil treatment
Patient no. 1
Subjects attended the laboratory on four separate visits at the same time each morning. On the first visit, in the absence of any drug treatment, ihe patients attended to undertake a dose response study with inhaled histamine (doubling doses from 003 to 4 mg); the provocative dose of histamine producing a 15% fall in FEV| from the post-saline baseline value (PD|s) was derived. On the second study day, NKA was inhaled in order to determine baseline PD|s value. On the following two visits (not less than 72 hr aparl) patients undertook dose-response studies with inhaled NKA 30 min after treatment with either nedocromil sodium (2 mg x 2 actuations) or placebo in a randomized double blind manner. On each occasion three baseline measurements of FEVi were recorded at intervals of 3 min. followed by the administration of nedocromil sodium or matched placebo. Nedocromil sodium and placebo were delivered by a pressurized aerosol via MDI and the inhalation of pressurized aerosols was conducted by one of the conducting investigators (RO). The MDI was held 2 cm away from the mouth and pressed during a slow, deep inspiration with a breath holding pause of 10 sec; the second administration followed 30 sec after the first pufl". The protocol was approved by the Ethics Review Committee of the Department of Respiratory Diseases {University of Catania) and the consent was obtained from each subject after the nature and reason for the study had been explained in detail.
Post-drug
Baseline 3-02(±0'54) Placebo 3-02 ( ± 0 4 5 ) 3-04(±058) 0-21 ( ± 3 3) Nedocromi! sodium 302 ( ± 0 5 6 ) 3-05 ( ± 0 57) 0-82 ( ± 1 4 )
Baseline
Placebo
8
15 9 293 I2'3 56 172 36 5 10'3 119
9 10
6-0 5-0
19-9 21 5 5-7 5-9
12-2 7'5-198
16-7 7'7-36-3
2
Study design
77
3 4 5 6 7
GM 95"., confidence limit
110
392 7-9 5-0 769
>920
Nedocromil >92 0 524 >92-0 74 >92 0 60 5 47-5 38 5 62 50 32-2* 14-2 73-4
*P-00019). In four cases, nedocromil sodium pretreatment doubled the values obtained after placebo administration (Table 4). Discussion In a previous study [24] we have demonstrated the protective elfect exerted by nedocromil sodium on SPinduced bronchoconstriction in asthma. The results ofthe present investigation confirm previous data showing the bronehoconstrictor activity of inhaled NKA [5.25] and the significant protection afiTorded by nedocromil sodium against the airway response to this neuropeptlde [19], Although NKA was effective in inducing bronchospasm. bronchial response to this peptide was not related to the level of non-specific bronchial hyper-reactivity to histamine in these patients in agreement with previous reports [19]. The lack of correlation cannot account for differences in hislamine baseline lung function on the study days and it could suggest a diiferent mechanism of action of these agents involving different pathways of bronchial response. The mechanism by which NKA, or other tachykinins, induce bronchospasm is still unclear. A possible involvement of mast cell activation in this response has been suggested by the large inhibitory effect of methysergide. a selective serotonin antagonist, on the bronchial response to NKA in the rat [18]. In addition in the same animal model both sodium cromoglycate and nedocromil sodium attenuated the bronchospastic response to NKA [18]. Both drugs seem to inhibit mediator release from a range of cell types including mast cells [21]. However, in dogs, they also possess the ability to inhibit non-cholinergic neural reflexes from stimulation of sensory nerves [26.27]. in human dispersed skin mast cells. NKA caused significant concentration-related histamine release although considerably less potent than substance P [28]. In support of this. Fuller et al. [29] have shown that in man the Hphistamine antagonist terfenadine (60 mg) significantly inhibited the weal and flare response to intradermally injected NKA. A different response to the same peplidergic stimuli may be elicited in mast cells harvested from different tissues. Indeed SP stimulation causes histamine release from cutaneous mast cells but not from pulmonary mast cells in vitro [30]. In contrast, recent studies have demonstrated histamine release in
80
A'. Cr/m/etal.
bronchoaiveolar lavage fluid recovered after adenosine and NKA infusion in a rat model [31] thus supporting the view that airways mast cell activation may follow tachykinin exposure and hypothesize an indirect mechanism of neuropeptide-induced bronchospasm. However, our recent observations (N. Crimi, unpublished data) have shown that pharmacological pretreatment with terfenadine (180 mg) failed to protect the airways of 10 asthmatic subjects against the constrictor effect of NKA. These data question the role of endogenous histamine as mediator of NKA-induced bronchoconstriction in human asthma. We have also demonstrated that premedication with the potent anti-allergic compound astemizole did not protect against substance P-induced bronchoconstriction [32]. In the same study the bronchospastic response induced by inhaled substance P was weakly inhibited by the anticholinergic drug ipratropium bromide. These findings suggest that activation o\' a cholinergic component rather than release of mast cell mediators in SPinduced bronchospasm. Stimulation of post-ganglionic nerve endings on rodents' airways demonstrated the release of acetylcholine and suggested that the bronchoconstrictor activity of tachykinins may be due to a cholinergic activation [8,18]. Furthermore eapsaicin, the pungent agent coming from hot peppers, is capable of inducing smooth muscle contraction and subepithelial oedema, possibly via liberation of SP and other neuropeptides from some primary atlerent neurons [2]. In human airways in vitro capsaicin exposure induces a slight and transient contraction which is inhibited by anticholinergic drugs [17]. In this study, nedocromi! sodium administration gave a significant protection when NKA was inhaled. This drug, introduced as a prophylactic treatment for inhaled use in asthma, has no intrinsic bronchodilator activity and against a range of acute challenges with several nonspecific bronchoconstrictor stimuli such as SOi [33]. nebulized distilled water (fog) [34], cold dry air [35], adenosine [36] and bradykinin [37] exerts a potent inhibitory effect probably by an action exerted on the afTerent reflexes arising from stimulation of lung irritant receptors. Protective action given by nedocromil sodium treatment against NKA could be determined by its neural activity on vagai fibres. Although anatomical, pharmacological and functional studies, conducted both in rivo and in vitro, attribute to neuropeptides an important role in the pathophysiology of bronchial asthma, the constrictor response to inhaled NKA in asthmatic airways does not necessarily imply that ihis peptide may have a specific role in the pathogenesis of bronchial asthma. The mechanism of action of various tachykinins, including NKA and SP, could be due to a cholinergic reflex activation, or to a
direct aetion exerted on specific receptors of bronchial smooth muscle rather than to mast cell mediator release. Further studies are necessary to investigate the exact role and the mode of action of tachykinins in human airways. References 1 Uchida Y, Nomura A, Ohlsuka H, Hasegawa S. Goto K, Kimura S, Sugita Y. Uchiyiama Y. Neurokinin A as a potent bronchoconslrictor. Am Rev Respir Dis 1987; 136:718-21. 2 Lundberg JM. Saria A. A bronchia! smooth muscle contraction induced by .stimulation of capsaicin sensitive sensory neurons. Acta Physiol Scand 1982: 116:473-6. 3 Barnes PJ. Asthma as an axon reflex. Lancet 1986: i:242-5. 4 Tanaka DT, Grunstein MM. Mechanisms of substance Pinduced contraction ofrabbit airway smooth muscle. J Appl Physiol 1984:57:1551 7 5 Joos G. Pauwels R. Van Der Straeten M. The effect of inhaled substance P and neurokinin A on the airway of normal and asthmatic subjects. Thorax 1987; 42:779-83. 6 Lundberg JM. Hokfelt T. Martling CR, Saria A, Cuello C. Substance P immunoreaetive sensory nerves in the lower respiratory iraci of various mammals including man. Cell Tissue Res 1984; 235:251-61 7 Saria A.Theodorsson-Norheim E. Gamse R, Lundberg JM. Release ofsubstance P and substance K-Iikc immunoreaclivities from the isolated perfused guinea-pig lung. Eur J Pharmacol 1985; 106:207 8. 8 Terenghi G. McGregor GP. Ghultacharji S. Wharton J, Bloom SR. Polak JM. Vagal origin ofsubstance P-containing nerves in lhe guinea pig lung. Neurosci Letl 1983: 36:229-36. 9 Lundberg JM. Franco-Cereceda A, Hua X. Hokfelt T. Fischer J A. Co-existence ofsubstance P and ealeitonin generelated peptide-like immunnreactivities in sensory nerves in relation to cardiovascular and bronchoconstrictor effects of capsaicin. Eur J Pharmacol 1985; 108:315-9. 10 Nilsson G., Dahlberg K. Brodin E. Sundler F. Strandberg K. Distribution and constrictor effect of substance P in guinea-pig tracheal bronchial tissue. In: Von Euler US. Pernow B, eds. Substance P. New York: Raven Press. 1977:75-81. 11 Theodorsson-Norheim E. Hua X, Brodin E. Lundberg JM Capsaiein treatment decreases tissue levels ol neurokinin Alike immunoreactivity in ihc guinea-pig. Acta Physiol Scand 1985: 124:129-31. 12 Karlsson JA, Einney MJB, Persson CGA, Post C. Substance P antagonists and the roleoflachykinins in non-cholinergic bronchoconstriction. Life Sciences 1984; 35:2681-91. 13 Martling CR. Theodorsson-Norheim E, Lundberg JM. Occurrence and effects of multiple taehykinins: substance P, neurokinin A and neuropeptide K in human lower airways. Life Sciences 1987.40:1633 43. 14 Advenier C. Naline E. Drapeau G. Regoii D. Relative potencies of neurokinins in guinea pig trachea and human bronchus. Eur J Pharmacol 1987; 139:233-7.
Nedocromil sodium on bronchoconstriction induced bv inhaled NKA
15 Naline E, Devillier P, Drapeau G, Toly L. Bakdach H. Regoli D, Advenier C. Charaeterization of ncurokinin effeets and receptor selectivity in human isolated bronchi. Am Rev Respir Dis 1989; 140:679-86. 16 Niisson G, Dahlberg K. Brodin F.. Sundler F. Strandberg K. Distribution and constrictor effect of Substance P in guinea pig tracheobronchial tissue. In: von Euler US, Pernow B, eds. Substance P. New York: Rav^^n Press, 1977:75-81. 17 Lundberg JM. Martling CR. Saria A. Substance P and capsaicin-induced contraction of human bronchi. Acta Physiol Scand 1983; 119:49 53. 18 Joos GF. Pauwels RA. Van Der Stractcn ME, The mechanism of tachykinins induced bronchoconstriction in the rat. Am Rev Respir Dis 1988; 137:1038-44. 19 Joos GF. Pauwels RA. Van Der Straeten ME. The elTecl of nedocromil sodium on the bronchoconstrictor effect of neurokinin A in subjeets with asthma. J All Clin Immunol 1989; 83:663-8. 20 Joos GF\ Pauwels RA- Mechanism involved in neurokininindueed bronchoconstriction. Arch Inl Pharmacodyn 1990; 303:132-46. 21 Church MK, Polosa R, Rimmer JS. Cromolyn sodium and nedocromil sodium: mast cells stabilizers, neuromodulators or anti-inflammatory drugs? In: Kaliner MA, Barnes PJ, Persson CGA, eds. Asthma: its pathology and treatment. New York: Marcel Dekker, 1991:541 69. 22 Dahl R, Pcderscn B. Influence of nedocromil sodium on the dual asthmatic reaction after allergen challenge: a double blind, placebo controlled study. Eur J Respir Dis 1986; 69 (suppl. l47):263-5. 23 Townley RG, Bewtra AK. Nair NM. Brodkey FD, Watt GD, Burke KM. Methacholine inhalation challenge studies. J All Clin Immunol 1979; 64:569 74. 24 Crimi N. Palermo F. Olivcri R. Palermo B. Vyncheri C, Polosa R. Mistretta A. Effect of nedocromil on bronchospasm induced by inhalation of substance P on asthmatic subjects. Clin Allergy 1988; 18:375. 25 Evans TW, Dixon CMS. Clarke B. Conradson TB, Barnes PJ. Comparison of neurokinin A and substance P on cardiovascular and airway function in man. Br J Pharmac 1988; 25:273 5. 26 Jackson DM. Richards IM. The effect of sodium cromoglyeate on histamine aerosol induced reflex bronchocon-
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striction in the anaesthetized dog. Br J Pharmacol 1977; 61:257-67. Kauffman MP, Coleridge HM, Coleridge JCG, Baker DG. Bradykinin stimulates afferent vagal C-fibrcs in intrapulmonary airways of dogs. J Appl Physiol 1980; 48:51 i 7. Lowman MA. Bcnyon RC, Church MK. Characterization of neuropeptide-induced histamine release from human dispersed skin mast cells. BrJ Pharmacol 1988; 95:121-30. Fuller RW, Conradson TB, Dixon CMS, Crossman DC, Barnes PJ- Sensory neuropeptide effects in human skin. BrJ Pharmacol 1987; 92:781 8. Ali H, Leung KBP. Pearee FL. Hayes NA, Foreman JC. Comparison ofthe histamine releasing action of substance P on mast cells and basophils from different species and tissues. Int Arch Allergy Appl Immunol 1986; 79:413-8. Pauwels R. Joos G. Kips J. Van Der Straeten M. Adenosine and Ncurokinin A cause histamine release in the airways. Am Rev Respir Dis 1988; 137:A2I2. Crimi N. Palermo F. Oliveri R. Palermo B. Vancheri C. Polosa R. Mistretta A. Influence of antihistaminc (Astemizole) and anticholinergic drugs (Ipratropium bromide) on bronchoconslriction induced by substance P. Annals of Allergy 1990; 65:115-20. Fuller RW. Dixon CMS. Barnes PJ. Nedocromil sodium inhibits sulfur dioxidc-indticcd bronchoconstriction. Am Rev Rcspir Dis 1986; 55:133Robuschi M. Vaghi A. Simone P. Bianco S. Prevention of fog-induced bronehospasm by nedocromil sodium. Clin Allergy 1987; 17:69. Juniper EF. Kline PA. Morris MM. Hargreave FE. Airway constriction by isocapnic hyperventilation of cold dry air: comparison of magnitude and duration of protection by nedocromil sodium and sodium cromoglycate. Clin Allergy 1987; 17:523Crimi N. Palermo F, Oliveri R. Vanchcri C. Polosa R, Palermo B. Maccarrone C. Mislretta A. Comparative study of the effects of nedocromil sodium (4 mg) and sodium cromoglycate (10 mg) on adenosinc-indiiced bronchoconstriction in asthmatic subjects. Clin Allergy 1988; 18:367-74. Dixon CMS. Barnes PJ. Bradykinin induced bronchoconstriction: inhibition by nedocromil sodium and sodium cromoglycate. Br J Clin Pharmacol 1989; 27:831-6.
