Airway inflammation and the pathogenesis of asthma

Ai rway inflammation has been recogni1,ed for more than l 00 years lo be present in the a irways o f pat ients with severe asthma. Muc h more recently, airway intl ammation has been identified lo be central to the pathogenesis of al l asthma. The inllammation is of a characteristic type, with the presence o f activated eosinophils, mast ce lls and lymphocytes in bronchoalveolar lavage lluid and a irway biopsies from patients with even mild asthma. Stimuli that are known to worsen as thma, such as inhaled allergens, also increase the numbers of mast ce lls and cosinophils in asthmatic airways. In addition, treatment with inhaled cortico teroids the most effective treatment for asthma improves symptoms and reduces the numbers of eos inophil s, mast cells and lymphocytes in the airways. The precise funct ions of the cells in promoting in tlammation and causing asthma symptoms has not yet been fully elucidated. However, it is very likely th :tt cicosanoiJs, such as the cysteiny l leukotrienes, are produced by eo.,inophils and mast cells and are a major cause of bronchoconstriction in asthma. Also, these infl a mmatory cells can produce prointbmmatory cytokines, such as granulocytc-rnacrophage colony-stimulating factor. in terleukin (I L) 3 and IL-5, which may promote continuing intlammation in the airways. Lastly, the pers isting inflammatory cell infiltrate and products re leased from these cells arc ve ry lil-.d y the cause or the airway structural changes characteristic of asthma, such as epithelial damage, gohlet cell hyperplasia. smooth musck thi cken ing and deposition of collage n below the basement membrane. These changes have been suggested tn he the cause of airway hyperrcsponsiveness in asth ma. An improved understandi ng of the pre cise mechanisms by which airway intlammation is initiated, propagates and causes airway damage will hopefully a llow more precise treatment strategies to he developed for asthma than currently exist.

In 18()2, William Osler published the mcdic;il textbook The Pri11ci{Jol. 1 and Pructice of Medicine .In the chapter on asthma, Osler notes that "bronchial asthma ... in many cases is a special form of inflammation of the small er bronchiole s" (3 ).This reflected the then curre nt descriptions or the pathology of the airways of patients dying of severe, fatal asthma .A more comple te description of airway infla mmation in patients dying of acute asthma was provided hy Dun n ill and colleag ue s (4) in the I 960s.Thus , while airway inflammation and its structur,d consequences had been recog n ized for mo re than I()() years.it was believed that infl ammation was confined to the airways of asthmatic patients with very severe di sease.More recently.the importance of airway inflammation in the pathogenesis of asthma, ranging in severity from mild to severe.or in trans ie nt asthma after ex posure to an in fla mmatory stilllulus.has been recognized .T hi s review exarnines the ev idence that airway inflammation is important in Lhe pathogenesis or asth ma. and considers the possi ble conse4uc nccs of persi sting airway infl am mation on airway structure and function .

AIRWAY INFLAMMATION IN ASTHMA
Dorlands Medical Dictionary defi nes inflam mation as th e "condition into which tissues enter as a reaction to inj ury".A common manifes tation of inllammatiun is the prese nce , at some time in the process, of activated infl ammatory cells at the a ffected tissue site.The type nf inflammatory cell varies with the type of inllammation.The most important inflammatory ecll at sites or acute inflammation is the ncut ro phil.However, in other, nwre chronic inflammatory conditions, eosinophils , ly mphucy tes or mast cell s appear to be more promine nt. for the purposes of this article.airway inflammation is defined as the presence of activated infl a mma tory cells in the airways.W hile thi s definition is restric ti ve and excludes other import,1111 components or inflam mato ry events, such as edema and vasodilation.quantifying the numbers, and occasionally the state of activation. of infl ammatory cells has bee n the most commonly used index of airway intlammation in studies of asthma.
Transient asthma and airway inflammation: The identification of stimuli that can cau se tran sient ast hma has proven to he important in studi es of the pathogenes is of asthma.Thi s is because the evidence that the presence o f inflammatory cdls is causally related to the de ve lopment o f ai rway hy perresponsiveness and transient asthma in human subjects initi a lly dcpL•ndcd on studies that cx ,11nincd numbers of cell s   and cellular differenti als in bronc hoalveo lar lavage ll uid (BALF) be fo re and a fter inhalation of ozone (5). the occ upational sensiti1,ing agents tulucne di isocyanate (TO I) (6), and all e rgen (7), all of whi ch arc I-: now n to cau se airway hyperrespons iveness and transient asthma or to exacerbate persisting asthma.T hese studies all demo ns trated an acu te infla mmatory response in the ai rways associated with the dL•vclopme nt of varia ble airllow obstruct ion, airway hypem.:sponsivenessand as th ma .In additi o n, the studies suggested that the st imulus wh ich initiates the airway hy perresponsi vcness de term ines the type of cel lular response.For example. a substan tial increase in neutro phi ls and a smaller increase in eos inophils were described in BALF in su bj ec ts with airway hyperrespo nsiveness followin g TOI (6 ).In contrast, airway cha llenge with plic atic acid, rcspo nsihlc for western red cedar asthma.caused inc reases in eos ino phils, but not nc utrophils in BALF (7).Arter alle rgen challe nge, some studie, desc ribe increases in cos inophi!s (8).o r eosinophils and ncutrophils (9).or eosi noph ils .lymphocytes and baso ph il s ( 10).
Measu rements in these studies we re carried o ut at diffrrent time points and with differe nt challe nge techniques, whic h may e xplain, in part , the varied res ul ts.However. it appears likely that d ifferen t inhaled stim ul i, such as allergen and TD!. cause a different patte rn of cellular influx into BALF.
A more recent roe.us has been the e xamination of the state of activation of the in llammato ry cells arter allergen inhalation (II) .These studies identified that cosinophi!sarc activated , as indi cated by posi tive staining rnr the marker for cosinophil cationic protein (EG2 ), as early as 3 hand persi sted ror more than 24 h in BALF after allergen inh.11.ltion.and this change preceded the increase in to tal number or co,inophils after allergen inhalat ion .
More recently , a less invasive method than hronchoscopy has been developed , using sputum induced by the inhalation of hypcrtonic sali ne, to quan tiry and characte rize inrtamrnatmy cells in asth matic airways.Studies using this method demonstrated that eosinophi ls increase marked ly in sputu m samples of asthmatics undergoing a naturall y occurring exacerbation of their asth ma ( 12), as well as 24 h after allergen inhalat ion , and persist fo r th ree days ( 13).

Persisting asthma and airway inflammation:
A number of studies prov ided info rmation on cell populations in BALF in mild stable as thmatics with persistent airway hyperresponsiveness and asthma ( 14-16).A common findi ng in all of these studies, as well as in recen t examinations of bronchial mucosa!biopsies (17,18) is the presence of increased numbers of in fla mmatory cell s.suc h as eos inophi ls, lymphocytes and mast cell s, compared wi th normal control subjects with normal airway responsiveness.The eosinophils have shown signs of activation, as indicated by increased levels of granular proteins, maj or basic protei n ( 15) and eosinophi lic cationic protein ( 16).In the bronchial mucosa, the eosinophils have shown morphological feat ures of activation, as indicated by heterogeneity of the granular structure ( 17) or as eosinoph il granu les lying free in the mucosa!interstitium ( 17 ).Azzawi and colleagues ( 19) and Poston et al (20) have also demonstrated increased numbers o f EG2-pos itive cell s, as well as significant increases in acti vated T lymphocytes ( 19).Mast cells in the airway mucosa have exhibited various stages of degran ulati on ( 17) su ggesting that mediator release is an ongoing process in the airways of stable asthmatics with pers istent airway hyperrespo nsiveness.
Some studies have correlated nu mbers of inflammatory cells in BALF with the severity of methacholine airway hyperresponsi veness in stable asthmatics.Kirby et al (14) demonstrated clo e con•el ations between degree of airway hyperresponsiveness in subj ects with mild asthma and m11nber of mast cells and eosinophi ls in lavage fluid .Kelly and coworkers (21) showed correlation s between numbers of neu trophil s and airway hyperresponsiveness.They also fo und that the activity of both neutrophils and alveolar macrophages, as indicated by luminol-enhanced chemolum inesce nce, was increased in asthmati cs compared wit h normal control s, demonstrating that these cells were metabo-1 icall y active.
These studies can be summarized as demonstrating the presence o f activated in flamm atory cells.eosinophils, ncutrophils, lymphocytes and mast cdls in the ai rways of asthma tics, even at a time when they arc conside red stab le and asymptomatic.The nu mbers of cells increase follow ing a sti mu lus that causes airway hyperresponsiveness, as well as in natural exacerbations of asth ma.However, the precise role of these different cells in cau sing airway hyperresponsivcness has not yet been clari fied.

INFLAMMATORY MEDIATORS AND ASTHMA
Identifying a role for an in fla mmatory mediator in the pathogenes is of asthma has re lied on the collection of various types of ev idence.Generally.when the structure of the mediator is identified and synthesi zed, the mediator is gi ven (usually by inhalation) to asthmatics to identify whether it can mimic some component of the as th matic response.Subsequen tly , when assays for its measurement are available,

Pathogenesis of asthma
efforts arc made to measure the med iator or its metaboli te during as thmatic responses.When antagonists fo r the mediator or inhibitors of its synt hesis are avai lable, they are studied in cli nical models of asthma.The final , and most di fficu lt, hurdle is to determine whether the mediator antagonists or synthesase inh ibitors are useful in treat ing asth matic patients .A variety of med iators has been suggested to be important in the pathogenesis of asthma.T hese have included histamine, acetylcholinc, prostaglandi ns D2, and F2a, th romboxane , cysteiny l leukotrienes, platelet activati ng facto r, and.most recently, cytokines and growth fa ctors .
Histamine and acetylcholine: Histamine and acetylcholinc have biological effects relevant to asth ma; both are released in response to appropriate stimuli -acetylcholine from airway nerves and histam ine fro m airway mast cells ; both are potent bronc hoconstrictors , and histami ne has other effects such as vasodilation and increasing vascular permeability.In addit ion, HI receptor antagoni sts have been de monstrated to inhibit part ially some asthmatic responses, such as exerci seinduced bronchoconstriction (22).However, even very potent and long acti ng antihistam ines are not effective bronchodi lators, nor do they have a useful role in the management of asthma .Atropine and other anticholinergics, such as ipratropium bromide, are bronc hodi lators and have been used fo r many years to treat bronchoconstriction, and, more recently, to treat (together wit h inhaled ~2-agonists) acute severe asthma (23).This suggests that acetylcholi ne is released from airway nerves in asthmatics and causes bronchoconstriction .However, anticholinergics do not modi fy any other important component o f asthma, which indicates that acetylcholi ne release is not involved in the underlying pathogenesis of asthma.
Thromboxane: Thromboxane A2 is a potent constrictor or smooth muscle, and its analog, U466 l 9, constricts hu man airways (24).A recent stud y exam ining the effects of a th romboxane synthetase inhibitor on airway responses after allergen challenge demonstrated slight, but significant, inh ibition of the allergen-induced early as th matic responses by 20 to 25 %, but no inhibition of allerge n-induced late asthmatic responses and allergen-induced histam ine airway hyperresponsiveness 24 h post-allergen (25).This suggested that thromboxane may be released fo llowing allergen challenge and may be partly res pons ible for the earl y asthmatic respo nse, but is not import ant in causi ng othe r a ll e rgeninduced responses.The thromboxane A2 synthetase inhibitor, OKY 046, administered orally, has been shown to improve acetylcholine airway hyperrespons iveness in stable asthmatic subjects (26).However, these studies were uncontrolled and need to be repeated in a placebo controlled double-blind study before the res ults can be properl y interpreted.There are no published studies on the clinical efficacy of thromboxane receptor antagonists or synthetase inh ibitors in the management o f asthma.However. the weak activity in protecti ng agai nst allergen-induced asthmatic responses suggests that these compounds are un li kely to ha ve a major clinical effect.activation ol in fl a mma tory cell s, particularly cosino phils and ncutrophils (27).Inh a led PAF has a lso been desc ribed as causing hro rw hoconstrict io n and airway hyperrcsponsiVL'ncss in normal su bjec ts in some (2X) , but not other (29), studi es.PA F has a lso been demonstrated to be re leased into thL' plas ma fol lowing allergen inha lati o n in mild asthmatic subjects (30 ).For these reaso ns, PA F has been suggested to be an import ant mediator in the deve lopment of allerge nind uced late asthmatic respon ses and airway hypc rrcsponsi vcness.and was suggested to play an important role in the ongo ing a irway infl am mation o f as th ma .
A variety ol' very potent and sel ect ive PAF recepto r antagonists has heco rrn: available f'ur clin ica l st udy .Until rL' .cently, the 'gold standard' compound has been the lh ie no-triamludi azepinc , WEB 2086.This compound has been used lo characterize th e PAF rece pto r. and has been shown tu prcvrn t PAF-induced hrnnc hoconstricti on in human subjects ( 3 I ).RL•ccnt studies have de monst rated that W EB 2086, ur ano ther very potent PAF antagon ist, UK 74.505.was c ompletely ine ffec tiv e in inh ibi ting a llergenind uced early o r lat e as thmati c responses o r a llerge n-ind uced airway hy pcrrespons ive ncss (3 2,33 ).Thus, PAF docs nut appear tu be import;111t in the pathogenesis uf allergen responses in asthma.It is very unlikl'l y that oral PAF antago•nists will ha ve clinical e ffi cacy in asth ma, althoug h high concen trati ons de li vered topicall y by inhalat ion may be mo re effective.

Cysteinyl leukotrienes:
Cy steinyl leukot ri e ncs (LT) C.; and D.; arc the most potent bronchoconstricto rs yet studied in hu man subjects, a rc up to 10,000 times mo re po tent than rne th acholine in some normal subjects (34), and have a longe r duration of action Lhan inhaled hista mine (35).Increases in urinary levels of L TE.;, the metabolite of L TC.; and LTD4, have been demonstrated follow ing alle rgen-induced early responses (36.37) , in patients prese nt ing to hospita l with ac ute severe as th ma (37) and fo ll owi ng exe rciseinduced bronehoconstriction U 8). S tudies wi th potent and speci fic leukotriene antagoni~ts have supported an impo rtan t ro le for the leu kot rienes in cl inica l model s or asthma, such as exerc ise (39) (Figure 2).and alle rgen -(40) and ace tylsalicy li c ac id-induced as thma (4 1 ).Lc uko triene re lease is partl y responsible for spontaneous bronchoconstrict ion in asthma (42) .Recent studies ha ve suggested that the cysteinyl leukotrie nes c an also cause the release of inhi bitory prostag landins rn ast hmatic airway~.whic h pnibably is the cause o f exerci se refractori ness (red uced bronc hocons tri c ti o n with repeated exercise c hall enges) in asth matic subjects ( 43 ).Lastly , the initi al st ud ies of the le uko tricne antago ni sts and synthes is inhibiturs have demonstrated clinical e ffi cacy in aslhma (44).
These results taken together indicate that the cystciny l lcukotrienes arc important in the pathogenesi s o f asth ma .
Cytokines: More th a n 30 different protein mediators arc c lassed as cytoki ncs.Some o f' these have bee n impli cated in the pathogenes is o f ~,sthma, mainl y because o f the abil ity ot some o r these proteins to pro mote inllammatory ce ll growth and differentiat io n, or in ll ammatory cell mi g ration and ac tivation, o r they may cause changes in the structural cell s o r the airways.T he study o f the cy tokincs in ast hma is not YL'l as deve loped as for the lipid med iators, main ly because of th e lack o r specific ant ago nists L hat c an be stud ied in hu mans.Ho wever.the cytoki nes inte rl e uk in-3, interl eukin-5, and g ranulocytc macrophage-colo ny sti mulating factor (GM -CSF) may be important beca use of the ir abi li ty to promo te eosinophil diffe rentiati on, recruitme nt and activation into the airways.and prolo ng the survival or these cel ls o nce in the airways .A ll or these cytoki ncs.as well as interlcukin-4.which is necessary fo r immunoglohulin E producti on.arc produced by o ne type of hel per T ce ll , the TH 2 cell.wh ich is present in as thmatic a irways (45).In add itio n. oth er ai rway ce lls may be responsibl e for the productio n of these cy tokines.Increased amo unts of GM-CSF arc present in airway biopsies fro m mild asthmatics , and the levels increase after a llergen chal le nge (46,47 ).It appears very like ly that these, as we ll as o the r cytokines.arc respons ible fo r the presence o r persisti ng activated cosinophil s and mast ecl ls in a~thm atic airways; howeve r, further studies with drugs that block the ac tio n of specific cytokines will be nC'cdcd to cstahli sh preci sely their role in ast hma.

AIRWAY STRUCTURAL CHANGES IN ASTHMA
Severa l struct ural changes have been described in asthmatic airways.which appear to be c ha rac teristic or th e di sease and which may a lso be responsibl e fo r th e presence or pe rsis ting airway hype rrcspo nsi vcness in asthma.T hese c hanges inc lude : pa tc hy dcsq uamated cp ithcliurn; th icke ni ng o r the ret irnlar coll agen layer below the basement mcmhranc ; and airway smooth muscle hypertro phy.Both epithe lial damage and smoot h muscle hypert ruph y have bee n imp licated in th e pat hogenes is o f a irway hy pcrrcsponsivcness in as thma.
Airway epithelial damage: Epithelial darnage and desquamatio n are present in airways of patients w ith asthma Can Respir J Vol 1 No 3 Fall 1994 (48 ).OnL' .hyputhe.,is lo explain epithelial damage and desquamation in ast hmatic airways is that basic protein., re leased from activated eosinoph ils damage the epithelium.Levels of major basic protein have been shown to be increased in BALF from asthmatic patients ( 15).Major hasiL• protein has also been identi fied in tissue sections by mea ns ul immunofluorescence fro m airways of patients dy ing from se vere as thma (-+9).Even in sections where identifiable eosinophils could not be seen, evidence show•ed major basic protein to hL' .present in epithelium and submucosa giving signs of ti ssue da mage (49).
Severa l hypotheses have been proposed to ex plain how epithelial damage may res ult in airway hy pcrrcsponsiveness in asthma.These incl ude increased permeabi li ty of the airway epithet ium : loss of inhibitory medi ators generated by the airway epithdium: and luss of ne utral endope ptidase.Several :.tudies compared the permeability or the resp iratory ep ithelium in nor111al subjects.asthmatics and asymptomatic smokers .These studie., demonstrated that the clearancc o l inhaled r:idiol:1bcllcd aeroso ls from the lung into the blood was much fas te r in asy111ptomatic smokers with normal airway responsiveness than in ei ther norma l subjects with normal airw:1y respo nsiven ess or in asthmatic .,ubjects with airway hype rrcsponsivl'ncs s (50).These resu lts make it very unlikely that increases in ai rway epithelial permeability are tl1L' cause of airway hyperresponsi veness in asthma.
The epithelium has been reported to release a fal°lor that reduces the airway smooth muscle cont racti le responses to agoni sts such as histamine and acctylcholine (51 ). Thi s mediato r has been called epithclium-dcri vccl relaxing fac tor tEpDRF ).The hypothesis that loss of an inhibitory EpDRF may be responsible for airway hyperresponsi ve ness in asthma has not been poss ible to test in human subjects.Studies in dogs dcmunstratcd that the functi on of EpDRF in vitro, as cletn111i11cd by the ability to reduce the conlractill' responses uf the trachealis to acetylclmlinc.hi stamine ur seroto nin , was un.tltcred in dogs with airway hy pe1Tesponsivcness in vivo compared with control dogs (52).It is unlikely.therdore.that loss of Ep DRF is the cause of airway hyperresponsivcness, at least in this animal preparation .
Airway epithcli:tl cells arc k.nown to release prostaglandin E1 (SJ).which has potent inhibitory cfkcts in the ai rways .,uc h as presy napliL• modulation and inhibition ot .1cctylcholine release from muscarinic ncrws (54).In addi tion, prostaglandin E has been shown to reduce contractile responses to inhaled hist:1minc.,tcetylcholine and rnethacholine (55 ). as well as brondmcon strictor responses to a crcise in asthmatics (56).The airw,ty epithelium also L'Olltains enzy mes -neutral L '.ndupeptidases -capable of metaboli zing lachykinins such as substance P (57).which arL' bronchoconstric tor and prointlammatory med iato rs.Loss of these inhibitory mediators 111ay have importa nt results in asthmatic airways; howc wr.this hypothesis has not yet been tested in asthmatic subjects.Airway smooth muscle : Airway hype rresponsiveness in as thma is nonspec ific.This mea ns that asthmatic airway s arc more responsive to all hrunchoconstrictor med iators acti ng Can Respir J Vol 1 No 3 Fail 1994

Pathogenesis of asthma
on airw,ty smooth muscl e receptors.One explanation Im the lack ol spL'c ilicity is that the underlying abnor111ality in ast h•• matic airways resides in the smooth muscle.The responses of airway smooth muscl e from human subjects with airway hypcrrcsponsi vc ness in vivo have been studied by a nu mber of different investigators (58 ,59).No co nsis tent in reasc in smooth mu .,clcresponsiveness in vitro has been associated ,vith airw,ty s hypcrresponsiveness in vivo, Very kw stud ies have examined smooth muscle in vit ro from subjects with airway hy perresponsivcncss and asthma.A small num ber or studies of airway smooth muscle from asthmatic subjec ts suggest that the smooth mu scle i., hyperrcsponsiVL.lo agonists in vitro compared with ai rways lrurn nonas thmatic subjects ((10.61 ).Thus. an inherent defect may ex ist in ,tsthmatic airway s111001h muscle .which accounts for airw:1y hyperresponsi vcness.
Another poss ibility by wh ich the mcrease in ,tirway sm oo th muscl e vol ume in as thmatics ca uses airway hyperresponsiveness is by cau sing thickening ol asth matic ai rways.James ct al (62).using modelling studies.de monstrated that a small inc rease in the thickness of the airway wall.which i, not possible to demonstrate by changes in spirn111c1-riL' indexes.could result in airway hypc rrespons iwncss in asth111a.

CONCLUSIONS
Airway intfa,nmati on appears to be central to the pathogenesis of all of the clinical manifes ta ti ons of ast hma.Many .,tudies have now demonstrated the presence of activated cosinophil s and mas t cells in the airway lumen and airway wall or patients with asthma.even those with mild di sease.T he presence and survival of these in lla mmato ry cell s may be promoted by inc reased proinll ammatory eytok incs, such as GM -CSF, in ast hmatic airways.These ce lls have the capacity to release potent bronchoconstrictur mediators such as the cystcinyl leukotricnes.whi ch arc responsible, in part al IL'ast , for airway narrowing in asthma and for allergen .exercise and ace tyl sa[icylic acid-induced asthma.Othe r cells, suL•h as a su bset of T lymphocy tes (TH2) .may also be importan t in maintaini ng the infl ammatory cascade.Airway structu ra l changes cau sed by persisting inflammation.such ;1s airway epithe lial da mage, or altered smooth muscle !unction or volume.:trL' likely to he important in the pathogenes is of stab le lon g standi ng airway hypc rresr onsivc ncss.Mediators released from the in tlammatory cells may be respons ible fur these changes.Desp ite the great increase in knowledge :thoul airway inflammation in the pathogenes is or as thma.the sequence of eve nts leading to the presence or persisti ng airway infl ammatory cell s, airway structural changes and airway hyperresponsivcness in asthma remains to he clarified.