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I NoTiCf 11ds ssrteeiat ra" k Tobacco smoke "sensitivityrr-ls there an °i ~ immunologic basis? Sarnuai B. Lahr.r, Ph.D., Farouk Barbandi. M.D., Jsffsry P. Taylor, M.P.H., and John E. Safvaggio, M.D..Vew ©rlearts. La. This srtrdY was undertaken to determine ;f there is an immunologic basis for reporrsd tobacco-smoke hypersertsirlvirv in man. Nfrten•-rhree individuals who were recruited on the basis of their sntokin j hr`srory andlor claimed sensiriviry to tobacco smoke were skin prick tested with tobacco smoke and leaf essracrs and their sera a+salyzed for reagtnic and preeipirariR; anribodies ro these antigens. Results demonstrated that a sitnifraar mmber of the nsdividt.als who were tested had positive shirt test and RAST responses to tobacco leaf antifeass. whereas anly a small number responded to smoke antigens. RASf or skin rest responses of stt{dy subjects to leaf or smoke antigens did nor correlate with symptoms of tobacco-smoke "settsitivir;n" or smoking historv 6ur did correlate with atopic status. Prectpirins were detected only to tobacco leaf C in 46 of the 93 irtdividyais who were tested but did nor correlate with s+trotite= hisrory or smoke "senscrivtti." These results suggest that subjective rolracco•smatce seissirivir}• is not caused by kypersensitivny ro tobacco leaf or smoke attnaerts. !J At1tRGt' Cutv /xMt/trot 73:,40.245. /984.) t~) rsG. The claim that allergic or allergic-like symptoms develop in some atopic individuals on exposure to tobacco smoke has led to the suspicion that tobacco- smoke allergy may exist.'` However, the existence of tobacco-smoke allergy in man has as yet not been proved. and available data are controversial.'-" Re- ports of immediate skin reactivity of smoke-sensitive individuals to tobacco leaf antigens suggest that an immunologic basis for clinical "sensitivity" to to- bacco smoke may exist.' This proposition hai been supported by the studies of Becker et al.4 who have isolated a glycoprotein from tobacco smoke and leaf that they claim is allergenic in man (based on pro- duction of a wheaE-and-flate reaction in 12 of 31 subjects whose atopic status was not defined). How- ever. no attempt was made to correlate this response with clinical "iensitiviry`• to smoke, and tttest is contrQversy concerning the purity of tobacco gIyco- • protein isoiates' From the C7insaf tatAweoiop Seetion. Ttttane Uai+Kaitry School of Medicine. New Orieau. La. Supponad by gang from ttr Costneil for Tobacco itssearcls. USA Sp.ct.t Project 804. and National te:oua of Nnttlt granrc Al 13401. Reeesved for yubticatum Nov. 22. 1992. Aeeepnd for pubjicatton Au;ust 12.1983. Repnnt ztquests: Sanuei B. Lehrer. Pls.D.. 1700 letditto St.. New Orieutt. i.A 70112. 240 , The recent observation that tobacco smoke contains immunogens that can stimulate an immune response to tobacco leaf in expetimental animals has given new importance to earlier findings that humans react to tobacco leaf antigens.t4'"' Thus our ctttient study was undertaken to determine if immediate skin reactivity and antibodies (tzaginic and precipitating) to tobacco Ieaflstnoke antigens exist in man and whether or not these responses are related to subjective tobacco- smoke sensitivity. MATERIAL AND MEfHQDS Study subj.rss Study subjects weze selected on the basis of NinKal "sensitiviry.' to tobacco smoke. Tlssae included atoptc and nonatopic individuals as well as smokers. exstnokers. or ttonstnokers. Smoke "senstcive" individuals wett defined as those who claimed conjurtcttvai,, nasal. and/or bronchial tobacco-smoke induced symptoms i i.e.. intense lacnma- tion. rhittitis. srteezin=. wheezing. or cough) on passive ezpostm to tobacco smoke. Nonsmoking "sensttive" con- trot subjects (both atopic and tronatopicf consisted of an age and sex-esatttted gsoup of volunteers. Subjects of both groups were interviewed with a standard quesriotsnaire'to obtain infoernation that coecanied syrttp< toms of tespiratory atopic disease. smoking history. and type of symptoms that wets eneauntetad when passively exposed to tobacco smoke. In order to assess smoke sen- sttivity of the studysttblect parucipants. they were uked L_

5 VOtUW T! Tobacco smoke "stns+tivity" 241 NUflatilt 2 i4bbreviations used RAST: Radioallerfosorbent test PBS: Phosphate-buffered saline SE: Smoke extract NHS: Natural hutttatt 'setvm PR1ST: Paper radioiusmunosorbtnt test the following question: "Do you encounter any difficulty when exposed to tobacco smoke?-' Those answering ..Yes'n were classifted as smoke sensitive. whereas those answering "No" wae classified as ttonsensitive. Those answering "Yes" were asked to define their sensitivity with respect to clinical synsptotns as sumtttatised in Table 1. Atopic indi- viduals were defined as those with a personal andlor fattaly history of allergy plus wheal-and-Rate skin reactivity to two or more of a battery of 16 local inhalant allergens. Stnokers were defined as individuals who were smoking at least one cigarette per day in the last 6 tno preceding the interview. exsmokers were defined as individuals who had discon- tinued smoking for at least the last 6 tno preceding the interview. and nonsmokers were defined as individuals who had never smoked more than one cigarette per day for more than a 6 me period. Tobacco i.af and smoke antigens Leaf A extract was prepared from flue-cured tobacco leaf (NC 2323 vanety. donated by Wallace 3. Dickens. Border- belt Tobacco Research Station. Whitesville. N. C.1: Leaf B was prepared from Maryland leaf (donated by M K. Ayecock. Jr.. Department of Agronomy. University of Maryland. College Park. Md.)• Leaf C was prepared frotn air-cured burley tobacco leaf (donated by D. L. Davies. Department of Agronomy. University of Kentucky. Lex- ingtcnt. Ky.). Leaf extracts were prepared by homogenizing 100 pn of tobacco leaf in i L phosphate (0.0IM) buffered (pH. 7.2) saline (0.15M NaCI)-PBS that cottuizied 0.1* NaN3 tas described elsewhereSe• "). After overnight extraaiott on a shaker (24• C). residual leaf in the mixture was removed by passage through gauze. attd the alttste was centrifuged (40.000 x f). The lipid-like material at the surface was tetnoved by aspiration. and the remaining sttperttatattt was dialyzed (Spectraphor4. Speeam Medical is+dastries. Inc.) against PBS. After dialysis, aeetmottismt sulfate (100%) was added dropwise at +t' C to a dnal eoaeeactscron of 30%. and the solut#on was stitred for one bms and was cezttrifaged at 10,000 x g. Sttpernatants were discsrded, and pellets were tedissolwd in and dialyzad against PBS. SEs were prepared by passing stnohe from a total of I300 IR2F cigarettes (Tobacco and Health Instittae. Lexington. Ky.) that was produced with a 30-pott Bae=waldt stnokin: machine (donated by the R. l. Reynolds Company) through a standard gas bubbler containing 50 ml PBS tdesigitated SE) or through a 30 ml solution of 156 pooled NHS. ob- tained from hepatitis B negative nonsmokers. in PBS (des- igtuted S€-NHS).". " Extracts were cottcentrated on an Amicon UMOS and dry weights were determined. All anti- gen soluttons for skin testing were filtered (0.45) for bacte- tiologic stettlity before use. Sterility was checked by plating 0.1 ml of extract onto a plate of tryptic soy agar followed by 48 hr incubation at 37' C. None of the extracts demonstrated microbial growth. Skin testing Skin prick tests were performed on all study subjects with 16 common inhalant allergens (house dust. cat epithelia. dog epithelia. elta, oak. pecan. Johnson grass. Bermuda grass. giant ragweed. English plantain. marsh eider. Alter rtaria. Xormodettdttu+t. Xelnttnthoiporirat. ASpergilluS. and FtrsariYm (Grees Laboruories: Lenoir. N. C.) at I:20 (wtv), tobacco leaf extract (10 tnghnl). and smoke extracts (5 mgJml). To exclude nonspecific t+eactiviry, all tobacco leaf and SEs were demonstrated to be skin-test negative in normal nonatopic laboratory personnel. A wheal texpressed as a mean of the largest wheal diatiu- ter and the one perpendicular to it) of 2 mm or tnott: was classified as positive. Positive and negative controls were 1 mgintt histamine and the vehicle solution PBS. respec- tively. NHS was used as a control for SE-NHS, and indi- viduals positive to SE-NHS were negative or had smaller mean wheal diameters to NHS. Immunological assays Immunodiffusion was performed in 0.4% agarose in PBS containing 0.1% sodium azide. Wells. which were sepa- rated by approximately 3 mm. contained 150 jLl of serum and 25 µI of antigen at 20 tagrmt. All plates were incubated at room temperature for tS hr and then washed three times with O.I7M sodium cittate in PBS to remove nonspecific precipitation. RAST was performed as described by Ceska and Lttndkvist." Cyattogen bromide activated disks were coated with leaf A, leaf B, kaf C. SE. or SE-NHS. In addition disks were coated with SE or S€•NHS by passing smoke from cigarettes through 50 tN 0.1 M borate solution that contained 100 cyanopn btotttide-acstvated disks or through 30 tnI of borate solution that contained 100 disks that were coupled with NHS. These disks wete then treated in the same manner as those coated with proteins.'• RAST results with smoke-coated disks were the saate regardless of the coupling method. All NHS was obtained from a pool of sera from nanatopic voiutttesss.lh RAST assay wat:perfototed as follows: the antigen-coated disks were incubated in du- plicate with the test sttbjeet's serutn. washed. incubated with -s"I-label.d anti-httmatt Ig€ (Pharatacia). and washed again. The total number of egnt for each disk was deter- mined. means wess calcttiated. and t+esults were expressed as the ratio of this number of epm to the number of cpm -observed with control disks that wett coated with NHS. Ratios a 2 were arbitrarily considered to be positive. Ii€ determinations were done in duplicate using the Phadebas PRIST kit according to the directions of Phar- tnacia. IgE values tunits per milliliter) were computed from a standard curve.

'Z42 t.lhrir at i1. TABLE I. Characteristics of study population by smoke sensitivity stnoke s.nsiti+retY i4han apf Mafa suts*.els J. .ii.L€RGY :LfN MMUNOL FIlAUNtv IlM fetsal* subjaets Vlthib subi.ets Nonwhit. subjsets Sensitive (n - 60) 31.8 i9 (31.796) 4i (68.3%) 87 (7a.396) 13 (21.7%) Vonsensitive (n = 33) 30.4 IS (45.5961 18 (s4.s%) 2S (7S.a%) 8 (24.2%) All (n - 93) 31.3 34 (36.696) S9 (63.45Fr) 72 t77.49U) 21 (22.65tr) TABLE IL Symptoms reported by smoke-s.nsitive subjeces on passive smoke exposure Atoptr tRStus Snntto+ns Ata'pis /1=K t94I Nonaopis As26 1%) Total nst{o tx1 Wheezing or dyspnea 50.0 26.9 40.0 Rhinorttjea. sneezing. or nasal stuffiness 70.6 53.8 63.3 Eczetsta or skin pruritis 3.0 0.0 1.7 Urticans-angioedena 3.0 0.0 1.7 Other (conjunctival irrita- tion. headache. nausea) 41.0 69.2 53.3 Statistical analysis Logar:thmic transformation was applied before starting the ana!}sis. Logarithms of the tmmunogiobuiin concentrn- tiona have a more normal diatnbution. Comporssons of mean serum-IgE levels with smoking hrstones were done by nnalvsis of variance. The Student t test was used to teat for differences in serum-IgE tneans with atopu status. The chi-square test was used to test for association between skin tests or RAST ratios and smoking history or smoke "sen- stttvity. ** RESULTS Study population The study population (93 volunteers) was predomi- t>ahtly female subjects (63%) and white subjects (77%) as shown in Table i. The mean age of the population was 31.3 yr with a range of 19 to 60 yr of the totat study population. approximately a third of the subjects were in each of the tfuee smoking categories (stnokas. exsmokers. and nonsmokers). and 60 subjects (fS+i6) claimed clinical "sensitivity." to smoke. There wet+e no signiFatu (p > 0.05) dif- ferences in age, sex, or race among subjects that were categorized by smoking history or by smoke "sen- sitivity." Sytttptoms experienced by the 60 atopic and nonatopic "smoke settsitive- individuals are shown in Table II. The most commonly reported symptoms were respiratory in nature: 40% of the subpcts re- ported lower respiratory tract symptottts. and 63% of the subjects rtpotted upper respiratory ttact symptoms. The atopic individuals had a higher proportion of respiratory tract symptoms (primary'rhinias. wheez- ing, or dysptxa), whereas in the nonatopic groups, symptoms were usually those of conjunctival irrita- tion. iyeadaciu, andlor nausea. Symptom distri- butions were similar for smokers and nonsmokers. whereas exsmokers displayed a somewhat different pattern with fewer respiratory symptoms and a higher proportion of itritattt-iike or nonspecific symptoms such as conjunctival irt7tatiort. Ifeadaclfe. and nausea. The differences in symptom distribution were not significant (p > 0.05). Skin test reactivity As shown in Table IiI. almost 28% of the total study population had a positive immediate wheal. and-flare skin test to at least one of the three tobacco leaf extracts tested. whereas only about 12% of the study population had a positive skin test to one of the SEs. Twenty-two individuals (24%) had positive skin tests to two or mott leaf exttuts. primarily to Leaf B and Leaf C. and 10 individuals (I i qe ) had positive skin tests to all tiuee leaf extracts. These proportions differ, however, with atopic stuus; 56.8% of atopic subjects had at least one positive skin test to any to- bacco leaf extract compared to 2% among nonatopic subjects. A similar relationship was seen for SEs with 20% of atopic and 4% of nonatopic subjects testing positive to at least one SE. No significant differences were observed among ,.sensitive" and ..nonsensi- tive" subjects when the subjects were being con- trolled for atopy. RAST to tobacco Naf and smoke exuaets Twenty-tiiree individuals (24%) had positive RASTs (ratio x 2) to any one ieaf extrart: the most predominant response was to exttuts of leaves B and C. Twenty-one individuals (22.5%) had positive RAST ratios to two or mote leaf extracts with five individuals (S.3%) that had positive ItAST ratios to all three leaf extracts. Only three individuals 0.2%) had IgE antibodies to SE-vHS: none had tgE anti- bodies to SE. The prevalence of specific-serum IgE F__

vOLun+f » . MU.ntM 2 Tobeeeo sn,oke "s.nsnivity" 243 ` TABLE IIL Percentage of atopic and nonatopic smoke sensitive and nonsensitive subjects with positive skin tests to tobacco extracts St:+ft sttbi.e.t tnr A' s C Any s€ SE-NNS Atopic (s+t) 31.8 54.5 37.7 56.8 20.4 13.6 20.4 Sensitive (34) 32.3 58.8 50.0 58.8 20.5 14.7 20.6 Nonsensitive (10) 30.0 40.0 40.0 50.0 20.0 10.0 20.0 Nonatopic (49) 2.0 2.4 2.0 2.0 4.0 0.0 4.0 Sensitive (26) 0 0 0 0 7.6 0 7.7 Nortsettsitive (23) 4.3 4.3 4.3 4.3 9.0 0 9.0 TABLE IV. Percentage oi atopic and nonatopic smoke sensitive and nonsensitive subjects with positive RAST to tobacco antigens t,.et 1%) Smoke t%) lttsdy srsbj.et (n) A a C Any S€ SE-MMS Atopic (id) 11.4 43.2 43.2 45.4 0.0 4.5 4.5 Sensitive (34) 11.7 4+t. t 47.0 47.0 0.0 5.9 5.9 Notsssrtsitive ( 10) 10.0 40.0 30.0 40.0 0.0 0.0 0.0 Nonuopic (49) 4.1 4.1 6.1 6.1 0.0 2.0 2.0 Sensitive (26) - 7.7 3.E 7.7 7.7 0.0 3.9 3.8 Nonsensitive (23) 0.0 4.3 4.3 4.3 0.0 0.0 0.0 antibodies among atopic and nonatopic individuals is shown in Table IV. Immunologic responses to leaf and SEs that were measured by RAST were related to atopic status but not to smoking history. No sig- nificant difference was observed in specific-serum FgE antibodies to any tobacco leaf extract in smokers. exsmokets, or nonsmokers. Approximately the same proportion of smoke-sensitive and nonsettsitive sub- jects had positive levels of specific-serum IgE that was measured by RAST to any tobacco Ieaf extract. To assess any relationship between the magnitude of the RAST response aad subjective smoke sensitiv- ity. RAST values for each leaf or SE wers plotted as shown in Fig. 1. Ttte greatest ISE-antibocly responses were to tobacco leaves B and C: ho+vever. these did not significantly differ in ••sestsitive versus nottsensi- tive" subjects. There was atiaimai or no IjE-andbody response to Ieaf A or to SEs. Strutn 1g€ IevNs The mean sertun-IgE level for the study population was 360 Ulml. As expected. a significant differ P,,, P was also seen between atopic and nonatopic subjects with atopic sttbjects having higfter ISE levels (616 UJttt3) compared to nonatopics (112 U/mI). Differ- ences by smoking history or smoke sensitivity when subjects were controlled for uopic status and race were not statistically significant, and no consistent trends were observed. I treai i%) smotte t%1 Prseipit#n response to tobacco leaf and smoke extrsets The only serum precipitins that were detected were to tobacco leaf C and were present in 40% of "non- sensitive" to 61.8% of the -sensitive" study popula- tion. No relationship to smoking history or smoke sensitivity was found at p > 0.05. DISCUSSION Although individuals who reported symptoms after passive exposure to tobacco smoke generally are convinced tttst tbese symptoms are "allergic" in na- ttne. analysis of this problem has yielded conflicting results. Our results indicate that individuals who cotnptained of smoke "sensitivity" report a variety of predominantly upper and lower tespiratory, conjettx- t1val. and nonspecific symptoms. Cttr ttsuttS further indicate that a significant propordon of atopic indi- viduals have positive skin or RAST reactivity to kaf and to a lesser degree to smoke antigens. In spite of the positive t+eactivity, ttere was no correlation be- tween ISE antfbody or serum precipitins and clinical •'sensidvity' to smoke in atopic or nonatopic stsb• jeets. Ottr results thus do not support an immunologic ateahanissa for tobacco smoke "sensitivity." Our findings that almost 28% of the study poptt3a- tion have positive skin prick test to at least one to- bacco leaf extract do not differ significantly from the 33% of the study population rrpotud in a recent :3 1:5PP 0101,q,9k-

J. Ai,LERGY i.N MM(;NQL ~ 2" UAnr it al. sasnuMV 'ss. 1ar taMHi L!! • t soom iaw s, TMKq 4Nt c . i4F ~ f 2 ~ t ~ bwa H . • • : • 10 l.r.. ta+nn NrMn s.r.-sO. • Pho" OEM N 3 N 4Wrwe o.r+arc S N S N a -tt.r++.t . - sanxrn.t FiG. Z. iqf antibodies to tobacco antiqens in '•amotca sensitive and nonseruitivE" individuals. tpE antibodies are expressed as RAS7 ratios. The soGd circtss indicate results that were obtained with atopic individuals and the open circles indicate results ttut were obtained with nonatopic individuals. S indicates individuaia raspondinq "yes" rvMn asked at4out tobacco smoke san- sativity and N indicates individuals respondinQ "no" to this ouastion. study." However, when skin reactivity is analyzed to tobacco antigens with respect to reported smoke sen- sitivity, it is obvious that there is no difference be- tween the two study groups. This is a conclusion that differs from that of Becker at al." This may be due. in part. to the fact that the past study by Becker at al: ` used a very small population and did not attempt to comlate atopic status or smoke "sensitivity" of study subjects with positive tobacco skin tests: `Other investigations4 used iarger study populations but did not adequately control for atopy and used only to- bacco leaf "antigens." 2'herefoce the positive skin reactivity to tobacco leaf antigens that were observed in these studies4 was not necessarily related to smoke "sensitivity" but rather to atopic status. This is based on our results that demonstrate atopic individuals had a ltigh'incideace of positive skin and RAST reactivity to tobacco leaf antigetts wluther or not they were smoke "sensitive." Why do a significant number of study subjects have positive skin and RAST results to tobacco kaf ex- tracts? Since tobacco-smoke sensitivity may represent a heterogeneity of clinical responses. it is possible tttat some of these responses may be IgE•mediated reactions to tobacco-smoke antigens. although our data suggest that now of these responses are related to tobacco-smoke sensitivity. Alternatively it is poisible that these tosponses have nothing to do with tobacco- smoke exposure or smoke "sensitivity.' and merely represent immune responses of uopics to ctossreact- ing antigens. which are stimulated perhaps by other members of the plant family Solansceae to which to- bacco belongs. Indeed the recent report of Becker et al. that indicated tobacco glycoprotein is present in many different vegetable products supports this pos- sibility.'s This hypothesis is also in keeping with the fact that atopic subjects are known to develop IgE antibodies against a wide range of common environ- mental inhalant atlergens," An utxxpected finding in this study was that some of the smoke-sensitive individuals were in fact current smokers themselves. This observation does appear tl- logicai since the question arises, how can they tolet:te tobacco smoke generated by themselves? However. generally these individtsals do not repott adverse reactions when they we smoking their own cigarettes but only in the pttsenee of smoke generated by others. This may rellect differences in inhaled tmain- stleam) smoke as compared to sidesueam smoke. which has been well documented ctianicaliy." Al- though tltis appears to be an untrsual obse:vadon. it was demonstrated in a number of individuals and ap- pesrs to be a significant reaction. If reported clinical "setuitiviry" to tobacco smoke does not have an IgE immunologic basis. then what is the cause of this reaction? Other potential mecha- nisms to be coasidered are the irritant effects of to- bacco smoke that are reported for other biologic sys- tems'»- '• or activation of other biologic pathways such as nonspecific histamine release. alternate cotn- ~ ~A S T E ~-~~P 0

i VOLUi1RE 73 NNWER 2 plement pathway activation. activaxion of the Hage- man Factor (factor XII) of the coagulation cascade (in view of the repotud effeas of tobacco glycopto- tein 2°) or I:gG homocytatrapic:ntibody 4. Finally we must consider the psychologic aspects of tobacco- smoke hypersensitivity in that some reactions to ta huco smoke may be due to the suggestibility of the subjects rather than to a pharmacotogic or an allergic reaction since psychologic factors ast known to pro- duce bronchospasm in asthmuic sabjects.a' ,These are irnportant aspects of this very complex problem, all of which should be adequately investi- gated before any deifnitive statement about the occur- rence of immunologically specific-tobscco smoke hypersensitivity. AEf'El1EriCKS 1. Sp.Q F: Tobacco and die nonsmoker. Arch Eavuon #dealth i6:"3. 19bE 2. Savel H: Clinica! 6ypersatsiciviey to ci=estte smoke. Arch Emiran Iiealth 21: I jb. I 970 3. Taylor G: Tobacco smoke atlesgy-Daes it tsist9la; Rylsadsr K. 'editor: Environmental to0aeea saaoke eflaas on the nonsmoker. 1974. Univerury of Gmva. pp S0-SS 4. 7susman BM: Tobacco seasiovity, ia tla allezsic popuisnon. I Astlsma Res I I:I39. 197t S. Harkavy I: Tobacco allergy in ca:diovaseular disease: A te- vte+r. Ann Allergy 25:u7. I96b fi. Secker CG. Dublin T. Weidmann HP: Hvpesseasnivity to ta- Oacco antigen. Proc Nasl Aesd Set 73:I712. 1976 7. McDougall JC. Gie,ch CJ. Tobacco alkrty-Fact or fantasy. I Acl.cac:• CuH Iwwsvkal. $7:237. 197b !. Keiler NF. Doyle RI: A saeehaaism for tabaeeo smoke in ndw.ad aikr=y. I AiuscY CuN Iwrurrat J7:271. 1976 TobacGQ fradkl -NtlstfW' 246 9. aick It1.. Saftsa RL. Ktoeidc R.. Ffilhaan E. Fat..d J: Srodies related to tobacco =lyeaproain: A clanaed acsivaeor of coagulation /tbeortysu. iuaie and a claimed atlerSea. Thomb Haemost s6:?31. I9U 10. tshner SE. tVlis on MR. Salvaggto IE It>:mtnsageme pop.rs t,es of ioh.aco smoke. I Atcs.wY Ccct+ Isuccte,at b2:36t. I97S i i. Becker CG. Lsri R. ?sv.a J: Iadoerioa of IfE anoboQias to aauiSen ffolat.rt feom tobaeeo irsva aad from cipmes smoke eoedeasns. Am J Fatioi 96:249. 1979 212. Gkicts GT. Weisb !w: Immmocimmical aad physioodsaniai propenrc of soboeco e:am. Am Rev Respn Dis 220:995. 1919 13. Lsh::r SIi. Wilson MR. Saivaggio IE Iaueisaopaiesty of to- baeeo sanolca eonspoerau in rabbita aed mia. ImAreh Alkrty Appi Immwrol b2:I6. 1960 I14. Cpiu M. Lavidlcvist V: A ner and simple tadoiamsuaoassay a>sthod for ehe anamioaaoa of ISE. Imnunodnnuay, 9:1020. 2972 15. S.eter t3s. VanEian" N. Wagar W: Tobaoea. eooaa. eof= fae and ta=w.Q eronnaaint allergens tlss asavaae factor XII dependent pethYays. siood 3t:t61. 1961 16. Salvaggio 1. CavawuSis IA. Lowell FC. Lskowia S: A eom- Iqeisoo of the imsnweoiagic responses of normal and acoptc individeals so imiaasWy admiaistar.d aatigon. 1 Ara.tatiY 33:42. 19be 17. Blue JA: Gprane astfmta and te6aceo aliwV. Asn AlkrV 29:110. 1970 19. Hatns MC. Shaae N: Sensieivuy ehese diseues. Philadelphia. 1964. F. A. Davis Company. p. 93 19, Reeicer CO. Dubin T: Activation ot faetor ?QI by tobacco glyeoproteen. i Exp Med l+b•07. t977 20. Shephard R. Collins R. Silverman R: ••Passive" exposure of asthmuu snblects to cifasstu smoke. Environ Res 20:392. 1979 21. McFadden ER Jr. Ltrpuello T. Lyon HA. gl.a3rsr E The n+.ehaassm of assias of NIsesaon ie ttr iadsaenon of awts aselrisa. FsyMosoen Mad 31:134. 1969 . HA j C- `~~