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~ IMPIRIM ~vo . .. . •` 'L'f,. ~ tW a . , ~.. . _..~.... 1976 REI'ORT o/ TIIE COUNCIL EOR TOBACCO RESEARCII-U.S.A., lae. TNE COUNCIL FOR TOBACf.O Rt:St:ARfal-U.!{.A., Inc. 110 Ea.l S9t1.91ree1, New Yorh, N.Y. 10022 44

I SCIENTIFIC ADVISORY IIOARI) to The Council for Tobacco Rcsearch-U.S A.. Inc. as of December 31. 1976 SHELDON C. SOMMERS. M.D.. Chairman Director of Laborator(es. Lenoi 11ill Hospital Clinical Pro/rssor of Pathology College of Physicians B Surgeons of Columbia University New York, New York RICHARD M. BING, M.D. Director of Cardiology and Intramural Medkinr Hunlington Memorial Hospital, Pasadena, California Professor o/ Medicine University of Southern California School of Medicine Loa Angeles. California JOSEPH D. FELDMAN. M D. Head. De artment of Immunopathology Scripps CPnic and Research Foundation La lolla, California WILLIAM U. GARDNER. PH.D. Scientific Diractor, The Council for Tobacco Research-U.S.A., Inc. E. K. Hunt Pro/essor of Anatonry (emeritus) Yak University School of Medicine New Naven, Connecticut ROBERT 1. IIUEBNER. M.D. Chief. Laboratory of RNA Tumor Viruses National Canoer inslitute Bethesda. Marrlaod I I I LFON O. JACOBSON. M.D. Director. The Franklin McLean Memorial Research Institute Regenstein Pro/essor of Biological Sciences University of Chka" Chkato, Illinois u. AVERILL A. LIEBOW. M.D. 1 u+ ~ Pro/rssor of Pathology (emeritus) University of California School of Medicine Saa Diego. C.lifornia , t HENRY T. LYNCN, M.D. i Pro/risor and Chairman Department of Preventive Medicine and Public ltealth Creighton University School of Medicine Omaho, Nebraska HANS MEIER. D.V.M.. Dr. Med. Vct., M.R.S.11. senior Stae Scientist The Jackson Laboratory Bar Harbor, Maine I.EF W. WATTENBERG, M.D. Professor of Pathology IhCartment of Laboratory Medicine and Patnolo6Y University of Minnesota Medical School Minneapolis, Minnesota JOHN P. WYATT, M.D. Director Tobacco and Health Research Institute University of Kentucky Lexington. Kentucky 9dd.tlfe St.ff .f The Council WILLIAM U. GARDNER. Prt.D. Sckatl/ic Director ROBERT C. HOCKETT, PH.D. Reaerrb Director JOHN 11. KREISHER. PN.D. VINCENT F. LISANTI. D.M.D. Associate Research Director Associate Research Director DAVID STONE. PH.D. Astorlare Research Director

0 CONTENTS Studies Related to ('ardiovascular !)iccascs and Function .... 5 Abstracts o( Reports . . , . ... - . , , , , . . . . 14 Cancer-Rclated Studies . . . . . . . . , . , , 14 The Respiratory System . . . . . . . . . , , , 29 Heart and Circulation . . . . . . . . . . . . . 41 Ncuropharmacolo6y and Physiology . . . . . . . . 54 Immunology and Adaptive Mechanisms - . . . . . - 61 Epidcmiolo6y . . . . . . . . . . . . . . . 64 Active Projects . . . . . . . . . . . . . . . . . 67 Compkted Pro jects . . . . . . . . . . . . . . . . 75 IndeR of Principal Investigators . . . . . . . . . . . . 85 Ldex of Senior Authors . . . . . . . . . . . . . . 86 t Studies Related to Cardiovascular Diseases and Function Previous annual reports have presented the general plan and rationak of Council-sponsored sludies of earcirapnesis and the etiology of chronic pul- rrN.nary disnrders. Their purpose was to provide a framework in which Ihe rekvance of individual eontribulions would he more easily apparent. In the present issue, we describe similarly some of our appruaches to study of cardiovascular diseases and function. Cardiovascular Dl.e..es Among Ihe many disorders of the cardiovascular system, those deriving from progressive alherosckrosis rank On1 as causes of disability and death in the United Srates. These include heart attacks (myocardiat in/arclion), angina, slroke, arlerial blockages in the Iirnbs, and some cases of congestive hear/ failure. Hypertension is believed no1 only to accelerate alherosclerosis, but alut to precipitate acute events In damaged circulatory syslems. Atherosclerosis is, lherefore, a principal focus of the present discussion Epidemiology o/ At6ero.clero.ii-Related Di.eo.e. Numerous epidemiological studies during the lasl 20 years have southt possible "eauses" (primary or contributory) of the alherosckrosis-relaled dis- eases. These studies often surnmariud their findings in terms of "risk /aclors " The "risk faclor" is essenliaMy a statistical concept based upon mathematical relationships without necessarily any known or established mechanism by which it might contribute 1o etiolM. Among the many twch "risk faelora" frequently reported are hypertensinn, elevated serum choleslerol, diabetes or a prediabelic dialhesis, cillareue smoking. personality type, Inadequate physical aelivity. and emotional stress. (]enetic predisposition (e.g., hyperlipoproleinemia, hcxmaysteinemia) is another such /aclor, both In its own tight and as a conbibut.x to most ur pi>sibly aN of the olhen. Biochemisls, physiologists, pharmaaologists, psychuk.gisls, and wtKo scienlists have thus been challenged to /11) the gaps and learn whether. how. to what eatent and in what kinds of persons each factor might aclually he operative. The stimulus to eoniec/ure, speculation and hypothesis has Accn strong, and many scientists have apparently seited the occasion lo lest Ihcu personal hunctxs in the hope of a hxky strikc. This was entirely Icgilimatr, but has tended Io produce a weller of (ra`menlary, confusing and inconclusive observations subjected to speculative projections. Clarifkaiion may come as cotxrent, integrative I"ia r:metile that can accommodate all the findinp that prove vdid. S

I Cigarette Smoking as a "Risk Factor" The identification of a"risk factor" can be no more reliahle than the stalislics on which il is based. Smr+ken selccl themselves from among the Seneral population on a basis or bases that are little understood Is q legitimate to c.onspare them with nonsmoker controls as animal esperimcnlers compare their test animals with genetically identical htter-malcs? Tobacco use has many variations. both qualitative and quantitative. To what ealent does voluntary adoption of any particular behavicral pattern select out a sub-population with innNe characteristics or life-styles that are linked to cardiovascular disease swceptihilily? There are many evidences (rons studies of man that genetic predisposition plays aa important part in delermining which individuals are poter tial victims of aUetosckrosis-relaled diseases. In several types ot rather e.lreree suscepli- bilily, the patterns of inheritance have been well established. Review of the "tisk factors" listed above indicates that most of them are already knowa to be inlhrenced, it not determined, by heredity. Studia of sponuneous alcohol cornumption by inbred mice have showa subslanli.l straia differences in appNite, in behavioral responses and in mela- bolism. Analogous mane Mudies of spontaneous nicotine and tobacco amoke Intake have now been inaugurated by a Council grant to learn whether similar genetic variations occar. l'aiw Studies /deetkal Iwins provide the human coumerpart of animals of inbred strain. S/udies of all like-seaed Iwins in Sweden and Finland are being assisted by Tle Couneil in an eRorl to determine broadly the relative inffuences of heredity versus envirronmenl upon the incidence of atherosckrosis-bsed cardiovaseular diseases. A key slratesy Is to compare the incidence of such disorders in smoking and non.moking iden/ical Iwins, including symplomalology during life and, eventuafly, age al death. the primary cause of decease and posl•mortem evaha- lioo of vascular pathology. Non-idenlical, like-sesed twin siblings serve as owMro4. Whik the incidence of discordance in smoking practices is low among IdeNical twiru, an observation that in itself evidences a slrong role of heredi- tvy influences (even among Iwins reared aparl) in determining Initiation and maintenance of snaking, the numbers of discordanta are Sreat enough to accrve b iacrnsingly significant levels In substantial twin populations followed over a Mrllcienl petiod of tLne. These studies have now been eaended beyond Identical twins to other rdalives of various defined degrees of consanguinitr. such as h.lf-aibs and Ihe oRap.ing of twins. 'I1se.w lirneecwssuming iwvesliplinns will require a eonsiderahk induclion perlod before comprehensive new reports emerele. They may evenlually provide more direct evidence whether cigarette smoking per re is a truly si`niAeanl "ri.k factor" in these and perhaps son.e other diseases. They eannN, however, be erpected to add greatly to elucidation of palhotenie processes or to provide 6 9 methods useful in reducing risks due to biochemical aberratinn% imparted by the genes from ancestors. For Ihis, other types of rcaearch are rcquired Cholesterol Metabolism Another prevakM "risk factor" in epidemiological studies has been "ele vated serum choleUerd." Contradictory findings over the years suggest that this concept was an oversirnplificalion, especially since many victims of heart attacks had no hislory of high ehokNerol levels. That cholesterol is implicalcd In a/herosekrosis has Seen assurned because mature arterial plaques contain the steroid both free vtd in combinations. Neverlhekss, long lerm adminis- Iration of drugs that lavet serum ehoksterol has not been as generally elfeclive In arresting or reversinZ the process in man as had been hoped. Accumulating infetmalion about the states of combination of cholesterol in blood is directing aueetion 1o the several c-hole.terol<ontaining combina- lions, classified as ehy:onskrons, very low density lipoproteins (VL1)1.), low density lipoproleins (LOI.) and high density lipoproteins (11D1.). The t Uls are the major carriers of hlood chokslerd and there are a mrmher of eaperi- menlal as well as epiderniologieal /Mrdings that an elevated serum level of VLDI. is correlated with progression of alherosckrosis, whereas ekvaled Hl)1. is a favorable indicdion. Thus certain "lipoprotein proliks° are now considered by many to he better indicators of susceptibility /o atherosclerosis than elevated total chrd- esterol. Certain of these prollks, associated with very high susceptibility to disease, have been showw to be lIeneticaM)f dNermined. tfarring some unforeseeable empirieal discovery (which does occur in medical research and is a perennial hope). the rational route toward effective control is through systematic study of the palboWneais of alherosckrosis, at biochemical and physiob&al kveb. This obviously must include a heltcr understanding of the regulalion of synthesis, lransprxl, tunctiun. and elimin ation of cholesterol and of the aberrations in disease in the hope of altering these by targeted treNmenl. The Council is presently reviewing the mosl promising u.ncepls and kads that now esisl in this eompka field with the intent of increasing its suppoft of basic study of athererosckrosis. The inlent is not only 1o assist these im- porlanl devek+prnenls, but also 10 seek more directly relevant assay systerns for assessing the possible effects of cigarette smoke inhalation. The task Is additionally diflleuH beeause the atherosckrotic disnrders appear to be peculiarly human ones with few eounlerparts among animals under natural eorditions. liighly contrived manipulations to produce animal "oounlerpart:' tend 1o diminish probability of relevance to hrrman esperience Iluman studies have been impeded by the lack of non-interventiwns tech- wiques for assessing iniliNiow and prolresswn of the atherogencsis processes in the vasculalure of man. New eaperimental techniques tor visualizing the condition of the arterial walls, iecluding scintillation photography, appear ar ahow promise for the future. T

In Vitro Studies o/ Arteriei and Vein. Meanwhile, in vitro techniques provide one method of sluJying synthesis .rd uptake of lipids and cholesterol by human arteries and veins, both "normal" and atherosckrolic, in direct comparison with those of .nimais. Though in virro conditions cannot be made to duplicate the in vivo situation perfeclly. they may provide useful guides to in vivo research. Such studies have already been asade with CTR support as described in the Council's Annual Reports for 1972-1973. 11 has been reported (and is reiterated in a current review) that human .Iherosckrotic and nornNl coronary arteries as well as sapheswus veins, perfused under puls.lik .rler.l pressures M(h human plasma containing labeled cholesterol and acetale, do not synthe- site cholesterol from aeetale and produce only small .mounts of cholesterol eqers. Choksterd is laken up in Identical amoums by norm.l and diseased vessels .nd Ihis uptake is increaaed at higher pulse pressures, s.hkh seems eonsiMant with the reputed effects of hypertension. Labeled acetnle is Incor- porated In1o various lipids. both normal and diseased coronary rlerks and saphenous veins synthesize free /atly acids, triglycerides and plwspholipids. Addition of nicotine to the perfusion fluid did not influena' cholesterol uplake. Analogous esperinxnls with dog arteries appeared to shown .n In- fluence of nicotine upon choiesterol uptake. If this is confirmed. iI wttlests caution M estrapolations from this species to man (1972 reporl ). Carbon rna+oside In the perfused plasma was reported to euhanee ehol- ealerd uptake by afl arteries in this in vitro system. Several new studies report a dramatic inhibition of cholesterol uptake by both human and animal arlcrks, under these in vitro eoeditions, by 7-keto- ehoiesterof. Living rabbits also showed an inhibition of cholesterol uptake. hul to a much snsalkr eslent. TAe low solubilily of the 7-keto compound imposed teehnk.l problems. Improved techniques may increase the efficiency of the tRect. Meanwhik. the disparity between the in vitro and whole animal te- sponan is another resninder that eatrapolations must be cautious. Other current studies deal with elucidation of possible mechanisms of the Inhibition and with the metabolic /Ne of the injected 7-keto compound ht rabbits. Role o/ Smooth Mu.c/e Cells in Athero.clerotie Plaque. TAe heretofo.e prevaknl theory of alherosckrotic plaque formation postu- lated that the inflhrNion of (atly substances from the bloodstream inlo the arterid wall gives rise to cholesterol deposits that act as an Irritant, causing In/lammation and proliferation of cells by processes akin to ordinary healing. This "itnudalion" concept was eondstenl with the associations of atherosckr- oda with elevated blood chokslerol, high blood pressure and high lat diets. Animals fed large amoun/s of .aturaed f.ts snd cholesterol. sometimes sup- plrmenled with Iw.nmun.l rw other 1reNmenlt, devek.ped (imirms superfki.lly re.emMrns th...r .+.vr.rd in man •1 aa.trqav 7 he.e lesions often regressed In snrm.l• %tr.• rn. drro oo,e.e atr.-/ •n ~ 1.urvarun that stimulated human dretarr wul.r. rn..a..n~ re.u,r.e.rr ..f atr-IrNer,d snd saturated fat eonsump- r Iion. Complicated by difficulties of conlrol, the results in man have been equivocal. Recent electron microscopic studies of human arterial plaques have re- veakd that the major component of authentic playucs is proliferating smtwnh muscle cells like thrne normally composing the center 1 medial ) layer nf arter- ial walls rather than 6brohiasl cells such as pradiferate to heal a wound. l:arly human arterial lesions (streaks) contain Ii111e lipid. sugResling that insudalion is not the prime factor in typical atherosclerosis. Choleslerol .kptnits and cellular debris appear later. Present debate centers on what incites normal smooth muscle cells to migrate from the medial layer and what causes them to proliferate. Presum.bly, damage to the inner surface cell layer of the vessels is implicated and a number of theories as to how this endrHhelial layer may become damaged have been advanced. Many of the plaques generated In animal arteries by n..r-physiological manipulations are reported to be radically differenl in composition Irom those of genuine human atherosclerosis .nd their relevance to the human diseases is therefore questionable. Recognition of the difference has, however, led to reporta that certain animah can develop human-type atherosclerosis under other more appropriate conditions. Snaolh musck eeYs from primate arteries are now being maintained successfully in culture media. Low-density lipoproteins added to the media stimulate them to multiply. FtMher, il has been reported that the ever-present blood plalelels, normally involved in Ihe clotlin6 process in response to injury and in other functions, ahio secrett a substance that strongly stimulates pro- liferalion of Ihew smooth muscle ee11s. Damage to etsdcNhelial and inlimal cell layers of an artery, which ordinarily separate the bhxxl from contact wilh the smooth muscle cells of the medial layer, can bring the.e cells into contact with platelets and presumably Ineite multiplication in the artery wall. Another concept holds that the snsoMh muscle cells mulliply as the consequence of a mut./ion akin to that which transforms other normal cells into malignant ones. This `nsonocbeaP' Iheory, which is supported by suikin6 evidence but is neverthekss in sonse dispute, would suggest quite diRerent mechanisms of action by e.lernd agents than those describcd heretofore Endothelial C.lti and Blood Plateleta A single layer of endotheliat cells eomposes the thin memhrane lining the inner surfaces of arleries, performing an important function in relainrnt the red napuscks while allowing waer and s.ame other suh.taoces to pass throuth. As mentioned, damage to the endaMhchum is heGeved t+r crrntnhua to all three of the snechanisms of atherosclerosis described. Many p.nubtc causes of endothelial damage have been auRRestcd and are being sturlied It is possible that several may be involved. '1 he damaeed enduthelium can repair itself, rather slowly. but if damage is sustained ar repeated uw often. repair may mN he achieved. (.'lots (Ihrombi) forming in an artery are thuuRht tu in/urc Ihe endo thelium by pressure and by impeding the access a/ oxyRen "1 he hl.wxl plNelets have long been- known to play a role in the ctNnplet events that produce 9 0

tbrrxnbosis ie both arteries and veins, and may thus contribute indirectly to endothelial injury. At Ihe aame lime, it has been thought likely that the plate- kts may contribute In some way to maintaining integrity of the endothelium. Recently developed Iechniques for growing human endothelial cells in culture media, a(ler harvest from umbilical cords, have made it possible to study tacion that influence their replication. A Council-sponsored investigator has reported that platelets added to the medium, as well as some individual subslasces secreted by the platekts, will slimulate their growth. It is, therefore, suggested that !n v/.o, platelets may espedite endolhelial repair. !le has also Irolated from normal blood substances that inhibit platelet adhesion and thus itnpede the processes of thrombus formation. The implication is that a delicate balance among opposing (unctions maintains inlegrity under normal conditions. Estensbn of such studies should help delineate the mechanisms of throm- boaM and atherosckrosis. Another inve.titator has used /w vitro techniques to cuilurr, nt heart wNrele atsd epithelioid cetb /o determine the effects of chokslerd, choksterol- e+less and 6-lipoprwein in producing cellular lipid inclusions and in labilizing }yaowera or mitochoe8ria. Lreldhins Cholesterol Acy/ Tr.ns/er.se (LCAT) Leeilhin: cholesterol acyl transferase is an enzyme thought to be respon- dbie for the esteriAcalinn of lipoprsxein cholesterol in plasma. It i! postulated by seret.l inveMitalon to be important in the mechanisms that renave chol- ~MOrol frorn the arterial wall. A number of previous papen, abslracted in these Annual Reports, have reported dinkal studies of 1.('AT activity in the plaswra of animals and maw sder diRerent circumstances to elucidate its clinical significance. Instability of the estrytwe oomplkaled Ihese studies. E.perimentation to delineate the Mtyswe's wsodes of action required its isolation. purification and stabilization so dmt en:ya+e oooeenlrations could be controlled and substrate compositions betw defined. A reoent publication reports that a concentrated effort has achieved the Iadatiow of LCAT in substantial quuuily in a virtually pure state with greatly Irnproved stability. Development of a radioimmunoassay Is reported to be under way. Two publications on applied studies of LCAT have also appeared reoealti. Availability of the purified enzyme and of i1s radioirnmunoassay should sUmulata and facilitate studies of choksterol metabolism and the roks of the se.esd lipoprolcins and contribute to understanding of atherosckro.is. Massbolic Actioities o j Puln.otaary Endot/te/iel Ceils Pive years of research assisled by The Council have produced nunserous reporta on this subietl. including several published during (976 It has been ,howw that the vasnacsivt potypepUdes bradyhinin and angirAensin I are mtla- bolkdly alsned durrnt a single passage IhrrMith the vast pulrm+nary, vascular bed Bradykimw. a s+.b.taece that tends to lower blood pressure, is completely inactivated while ansiotenein 1 is converted to an6iotemin 11. which is a potent hypertensive agent. Bradykinin, under suitable conditions, can inhibit this antiotensin conversion. These relationships suggest a role of the pulmonary circulation in blood pressure regulation. These metabolic changes have been traced to peplide hydrolase enzymes of the lun`, which are not present in the bioodstream but are attached to Ihe luminal surfaces of endothelial eells, especially those of capillaries and venuks The same enzyme inaclivates bradylinin and converts ansiolensin 1. Steps in the hydrolytic process have been described in considerable detail; antiirxiia lo the enzyme were prepared and labeled to provide lools (or research. New functions of bradytinin are now being discovered, including an effect on prostatlandin synlhetase which remains to be esplored in depth It also remains for future studies to Inquire how dysfunclions of these systems may be related to vascular or puhno..ry, diseases. Chronic Smoke Inad.tion by Doss A technique for chronic eaposure of beagle dogs to cigarette smole via tracheostomy was employed by a CourKtil-supported scientist to took for possi- bk effects on clotling snechawisrns and on cardiovascular function. ikspite recognized IimitNiom ol this technique as a nwQel of human practice, rather e.tensive eaperieece wN,h i4 use for other purposes suggested that e.tension of observations to the cardiovascular system might provide some preliminary data and guidelines (or twure .ludies. No dinically evident dise.se was towrd at/er 1s months' eaposure under different dietary regirnes. Some Indications of possible enhancement of coatu• 1alion mechanisms and of relatively minor alterations in function were re• porled. The author described these minutely wilh cautions against transfer to man in view of eaperisuealal limilalioru, pecies dilferenees, and Ihe treater tbrnpkaNy of human e.viroament..' Studies Innotrins Nleotiwe A number of prW Couneil-sponsored studies, mainly using animals and with a variety ot objectives, have invo(ved the administration of nkotine. Eaperience through the yean wuests that comments on sorne of Ihe problems involved In selection of dosages are pertinent lor evaluati.ns of past results and for designing new and better esperimenls. SmoMers receive nicotine by way of the oral cavity and Ihe luns, in irnolf successive doses over a period of several minutes and repealed at variahle intervals. The entire dosage range they etperience is very low in conlras/ to the levels often used by pharmacologisb in e.ploratory studies of mcoNne's pharmacological potentials. The view has been e.pressed that ordinary smut- ing rarely. U ever, produces high enough levels to act upon the sympathetic ganglia, but slimulates only the several special sensory receptors to incite raprd but brief systemic responses of re(ka origin 7hese responses may be quite contradictory and paradoaical in nature, depending on the conditions that ptedomioate at any moment. 11 10

Yet, within the low range of dosages that can he achieved from smokint, there is still very considerable variation Rate of absorption via oral cavity or lung is highly dependent upon the acid-hase balance in Ihe smoke. Maintenance of blood level is strongly aRected by the hydrogen-ion concentration of the bladder contents. The detree of acidity influences nicotine rewrption and is altered by diet and nervous state. Nicotine metabolism is rapid and melabolic rates not only vary markedly among normal humans but are probably in- fiuetsced by duration of smoking esperience, a recognized but little understood phersoexnon. In addition are the variations due to choice of tobacco product used .nd Individual diRerences in puff volume, puff /rerauency, depth of inhalation. duration of snsoke retenlion, etc. Design of animal esperi.xnts intended to mimic human smoking e>t- perience, long or short, must include consideration of .11 the factors enurner- ated and species diflereeces as well. EaperimeMal control or Mandardiralion of all the variables mentioned in any large-acak or /ong-continued human study appears to be virtually im- posibk. However, an eapan.iors of information on the actual ran" or dura- tions of plasma nicoline levels attained by human smokers (and users of other forms of tobacco) under actual conditions of life should be attainable. Recent studi.s by laborious snethods have provided such data for small sumbers of human subjecls. These are a most valuable interim guide to the daign of animal esperimenls However, mass data on large representative populations are needed to de/lne, on a sound atatistical basis, the ranges, peaks and medians of plasma nicotine on a time scak, if important refinements are to be achieved in epidemiological studies of smoking. R.dioinswsunological A..ay o/ Nicotine Sensilive, specific and rapid assays for plasma nicotine and its major metabolites have long been needed. They should be able to he carried out with very small samples of blood. with simple. inespensive and mobile epuipmenl, and by competent technicians without etaraoadinary special Iraining, have long been needed for such purposes. Rad'aimmunoass.ys would appear to have promise of meeting these re- quiremenls and The Council is continuing to support studies in this area. Antibodies sensitive to nicotine have heen obtained and report.d t1973)• Continuing ellorts are now directed toward improving sensitiviq, testing for specificity and simplifying procedures for broad applicalion. Nicotine Bffecfa on Coronary Circulation in (:on.cioua noR• To avoid the eRects of anesthelics on the pharmacobRical r.•sponses to nieoline, a merhrxf was perfected lor imracar.rlid adminidratiun of the alka- loid to conscir.us do`s in doses de.cribed by the investigator as "realistic." A striking increa.e rn cor(Mary hhM.d flow was oh.erved which was (raeed to Iwo separate /actars Tbe marx component was found to he due indirectly to + an increase in the depth of respiration produced by nicotinc. Ihe minor one to a chemorefiea via a different pathway. F.PIDF,AIIOI.OGIC STUDY OF SiU)KIN(: Ct:SSATION A general difflcully in making and interpreting studies of smoking cessa• tion is that those who discontinue smoking are not cMnen at random hul by their own decision, so that the influence of acleclion on comparafahly of the groups has been unknown. Nor has it generally been possible to as.css the influences of concomitant or "eompensalory" changes in life-stylc, such as alleralions in dier, eaereise, use of akohesi or drusa. etc. A Council-spunsored eomparison of continuing snwskers, with smokers who have disconlinued, and with smokers who have stopped /rK a periwd and then resumed, sometimes repeatedly, is under way. In the population under sludy, a large body of data had been collected by uniform methods when all were smokers, before any had Mopped. Hence, some bases may be found lor assessing selection biases that rn.y have occurred in the aubsequent groupings into the calegories mentioecd. 12

j Abstracts of Reports Following are ahsuacls, approved by the aulhon, of reports on new re- search acknowkdgin` support (rom The Councit that have sppeareJ in scientifk }ournals since publication of the 1975 Report. 1 he name of the recipient is in ilalics. ~ The abstracts are grouped under these headingi: 1. Cancer-Rclated Studies. (1. The Respiratory System. 111. Heart and Circolation, IV. Neuropharn-ucolosy and Physiology. V. ImmunolM and Adaptive Mechanisms. VI. Epidemiology. 1. Cancer-Related Studies HYDROCARBON-NITROSAMINE SYNE:R(31SM AS A POSSIBLE AMPLIFYINO FACTOR IN LUNO TUMORIGENESIS BY TOBACCO SMOKE A number of studies have shown that inducers and repressors of microsomal miaed-funetion o.idases can powerfully inlluence the eflecls of chemical ear- einopens. A significant incidence of pulnwmary tumon has been reported in both rats and mice as a result of the simultaneora administration of )-metfiyl- eholanthrene (MC) and dimelhylnilrotamine (1)MN) at levels at which neither compound h carcinogenic in the lung Nittosamines, including DMN, as well as 7,1-bentopyrene and ),1-bentoAuoranthene whoae carcinogenic properties are very similar to thosc of MC, are present in tobacco snwke. Therefore, it is pro- posed that a substantial proportion of the lung tumor incidence of sn.okers is due to synergism between the carcinogenic hydrocarbons and nitrosamines in the smoke, rather than to hydrocarbons alone. Two alternative tnechanisms which may account for this synerlism are considered. Preliminary results, how- ever, support the concept that DMN increases the hydrocarbon epoaide pool by lowering the activity of microsomal eposide hydrases In the lung while substan- tially Increasing the arylhydrocarbon hydrosylase activity. The demonstration of a DMN-hydrocarbon synergism in human lung earcinobenesis could tn- oours{e uneapbred approaches for the development of partial means of pro- Ieclion for smokers, such as the use of DMN-dcmelhylase repressors and in- hibibrs to block the DMN-hydroearbon synergism. It is also suRgr.tcd that dietary nitrosamines or prenitrosamine components, such as nNrites anJ secon- dary amines, may be found to have sorne role in the etiology of lung cancer. Argus, M. F. and Arros. 1. C. Iorrnd of 7Aro.eN,d d/oloffy 36:491-1911, 1976. Other support: National Cancer Institute. From the Seamen's Memorial Research I ahnratory, I1. S. Public Ileallh Service 11n•pital, and the Uepartment of Medicine. lulane University Medical Center, New Orleans 1 SIRU('IURAI. I.IMIIS OF SPF.CH'1('ITY OF MEIIIYI.('ll()LANIHRENE-REPRES.SIBI.E NIfROSAMINI N-VEALKYLAStS. INHIBITION BY ANALO(: SUBSIRAII-s In order to gain sn insight into the structural specificily of durNthyln,tru,.. mine (DMN)-demethylase, a systematic investigation was carrrcJ wn ,ar lhe cnMnparative dealkylation of DMN, higher diatkylnilrosamines and DMN .n., k+ts, as well as on the inhihilion of DMN-demethylatiun by the DMN ,nawgs_ Other e.periments in this framework eapbred the eQec1 of pretreatnknt by 1. mcthyl choianthrene (MC). a potent repressor of DMN-demcthylase, on th..e different dealkylases. Resuhs showed that the dealkylation of dimethyl-. drcthy6 and dipropylnilrosamine by hepatic microsomes of Sprapre-Dawky rats was tepreaseJ by pretreatment of the anirnah with MC. 7his repression prolus. sively decreased with the Increase d.Ikyl chain length. In conlrast trr iti effect on Ihe demethylation of DIKN. In viro phenoharhital induced rathet than re- pressed the deethylation of dielhylnitrosamine (DEN). lhe rates of dcmcthyla lion of the DMN analog subslrales, although low as compared to 1)MN, in. creased with the acyl chain length. These analogs were potent in virro inhibaon of DMN demethylation when tned in combination with DMN as substrates, and the inhibition decreased with the length of the acyt chain. Although the rate of demethylation of inelhylphenylniuosamine was not influenced by M('- pretrestmenl, tine compound was, however, a potenl inhibitor of demelhylalion when used as substrate in combination wilh DMN. Moreover, beyond the ap- parent distinctness of DMNdemelhylase and DI:N-deethylasc there is now in dication that more Ihan one en:yme, having the same substrate specificity but different kinetic and regulatory eharaeleristics, underlie DMN-demethylase ac- tivity. Arcot, /. C. et e/. Zeirscibi/t Jiir KrebiJo.xArnP nnd It`lin/iche OnAotoRie e6:171-at), 1976. Otller ur'prtr National Cancer lmtilwe. From the Seamen's Memorial Research l.aboralory, U. S. Public lieal,h Service Hos pilal, and the Department of Medkine, Tulane University Medical ('enler. New Orleans. DIMETHYt.N11ROSAMINE-DEMETl/Y1.ASli: ABSEN( E 01; INCRI?ASEI) FN7.YME ('ATABO1 ISM AND MI)1 1/1'1 1('I I Y(1F EFFE('fOR SIIES IN REPRI:SSIUN. Hh.MUI•ROII-IN INVUI VI•MI NI Evidence is presented here that Ihe rate ot decay of Jimcthylnirru.anminc (DMN)-dernethylase following pretreatment with )-merhykhol.rnrhrene (Mt ) is no greater than that so be eapetled (rom rnuoul enryme cat.Mrlism '1 hrse results also show that the observed decrease of 1)MN dcmethyl..e V,,,, Iolluw inb MC administration is rat due to increascJ rate of breakdown MN to dc crea.ed Aa, novo synthesis. Other eapcrimenb in this study inJiaare that dr/fcr eet receptor sites are involved in the repressinn of I)MNdemethyla.e by hydru carbons and by phcnobarbilsl (PB), and that a P-ISI) type nuarosnmal cyto chrome is involved in the demethylalinn of UMN. lhe totality of these e. petimental observaliona presents an apparent patadoa which may tic aummar-