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ST00I !6 I 0 638 A ^t?Of'n:eo jre Verves a En^ironmenia, 'ifiwrn ~orn *96d volume tS ZoDyognt i<)F,Fj Amer.can Medcai Association Experimental Asbestosis \ Studies on the Progressiveness of the Pulmonary Fibrosis Caused by Chrysotile Dust Paul Groww. MD. and Robert T. P. de Treuillt, Pittwburth i\-V T HE unreliability of symptoms as a crite termined. This ia particularly true of asbea- rion for the progression of pulmonary fibro tosis. sis was underlined in a medical and psychi It is the purpose of this paper to define atric study of coal miners with respiratory some of the anatomic stjymata.-of wpgrea- complaints. Here, a very significant associa oTnot asbestb^^caused^y^uya^^^wffi' tion was observed between a recognizable psychoneurotic factor and an apparent progressive. worsening of the clinical condition: of 40 patients studied, only 12.5% had disability Methods and Materials on "physical" grounds alone: 35% had dis ability on psychiatric grounds alone; and the remainder on both.1 Furthermore, discerning clinicians recog nize that opinions concerning the progres Lung burdens of chrysotile asbestos dust were imposed upon rats, hamster*, and guinea pigs by exposure.ia inhalation cframhera as well as by intratn^he^Linjectfaflfc. Most of the ani mals exposed to dial .in an inhalationcfaembcx sion of a pulmonary fibrosis based upon the and reported in this study were port of a larger symptomatology (eg, shortness of breath) and a roentgenologic evaluation, even when reenforced by Lung function studies may be quite erroneous. In the presence of a superadded coincidental pulmonary illness, such signs and symptoms may be reversible in whole or in part between such episodes, which include covert and overt acute pneumonitides, pulmonary edema, and allergic or chronic bronchitis--with or without em physema, or emphysema without bronchitis. Such diagnostic difficulties were the subject of a recent symposium on emphysema in ;ndustry. As uncertain as are the clinical criteria of progression of pneumoconiosis, very little help has come from postmortem studies of human lungs or those of experimental ani mals in defining the criteria by which progression of pneumoconiosis may be de- investigation that will be reported separately. The inhalation chamber in Which D. animals (except four guinea pigs) were exposed to dust, measured 8 X 8 X feet. The animals - were housed in wine cage* that were suspended hi racks. Periodically, these cages were rotated *o that inequalities to dust exposure caused by po sition were largely obviated. The exposures were for six boon per day. five days per week. The total exposure varied; as listed hi the table. The chrymtila rebestaa was baUmifiad and then fed into a buanwnOl (modified from a design by Holt and Young*). This was prtnidtri with inlet and outlet tiddag so arranged that the comminuted aabestoa wan fed back eosrtuniatly into the hammermiH. At the same time, the ultrafines were allowed to waft upward Into the inhalation chamber. Two wf these hamrocnnilli provided the chamber wfffc sufflriaat-dnst to av erage 86 mg/cu m with a range at 42 mg/co m to 146 mg/cu m. The asbestos dust cloud was evaluated by sampling sfft a twe-stage rise se SubnritUd to* publication Aug 9, 1966: accepted Au* 25. From the Industrial Hygiene Foundation, Mellon Institute, Pittsburgh. Read before the Hatch Symposium* Graduate School of Public Health* University of Pittsburgh, Julv 18-19, 1966. Reprint requests to the Industrial Health Founda tion. Mellon Iortitute, 4400 Fifth Ave, Pittsburgh 15218 fDr. Gnm). lective device, similar la design to that pro posed by Wright* and operated at 20 ti tera/miuute. The first stage of the tostnaaaat waa a horisemtai elutriatar with selector char acteristics recommended by the Johanuedbmg Pneumoeoarioais Conferssra at 1.969, ft,----lv acceptance at all psrtidw havtoe Isrnfasl sattllng velocities greater **" frw c* m Tip sphere 6f ttohtity I gn>M> cm. S0% ecceptanoe Arch Environ Health--Vol 15, Nov 1567 PLAINTIFF'S EXHIBIT ST0011610 of particles, the terminal settling velocity of Animalt Exposed !o, or Infactad Intrat/achaalty which 19 equivalent tr> 5m spherical particles ..( With, Chrytotlla Aabasie* Dual 1 gm/cu cm density, and dropping to zero ac ceptance of particles having terminal settling velocities equal to a 1m sphere of l gm/cu cm density. All particles penetrating the first stage elutriator were considered "respirable" and were collected on a permeable-membrane filter which acted as the instrument's second stage Inhalation Exposure (Months) at Oust intra T.me, Concentration tracheal End o! Species (average) Injection Exposure No. [86 mg/ (20 mg/ No. 3.5-mg to Death Animals cu m) cu m) Inactions (Months) The filter was dried and weighed prior tn and after collection of the sample. In this man ner. it was found that 63% to 79% of the sus pended asbestos dust in the exposure chamber was respirable, ie, its terminal settling velocity was less than that of a 7.1m sphere of 1 gm/ou cm density. The exhaust air from the inhalation chamber was drawn by suction through a pair of elec trostatic precipitators. The chamber air was Rats 2 0.25 6 0.5 61 52 44 48 5l 52 44 10 6 U 13 1 3 24 5 II. 25-24 11.2523 0 S 21 3.5 17 21 21 21 21 consequently always under negative pressure as long as the hammermills were operating. 52 Hamsters When the hammermills were shut down, fresh air was pulled through the chamber before the 9 5 1 13.5-21 2 12.5-16.5 door was opened. 14- Periodically, dust from the electrostatic pre Quiftaa Rigs cipitators was collected and weighed. Since a record of the air flow through the chamber was 6 0.5-5.5 43 0 04 kept, it was possible to calculate the average 10 dust concentration in the chamber air by divid ing thB weight of the dust collected within a lion of the lung dust burden as shown in the time period by the volume of air flowing table. through the chamber within that time. All animals were autopaied, the lungs were As seen in the table, four guinea pigs were removed and expanded with 4% formaldehyde exposed to a concentration of chrysotile dust solution under a pressure of 12 cm water. averaging about 20 mg/cu m. For this exposure, Paraffin sections were cut at 6m- Cleared un a smaller chamber waa used and the dust was stained sections were examined under dark- prepared by atomizing a suspension of ballmilled chrysotile asbestos at a pressure of 100 lb/sq in and impinging the emergent jet a nearsonic velocity against a tool-steel baffle. The av erage fiber length of this dust waa 0.92m as de termined by measurements from electron pho tomicrographs. Inasmuch as other data on the size distribution of the dust are not available, it is not possible to state what percentage of the dust suspended in the chamber air was respira ble. These animals were exposed for three months to this dust. intratracheal injections, the dust collected from the electrostatic precipitators waa .sus pended'in water so that 1 ml contained 3.5 mg field conditions'* and appropriate fields were photographed on 35 mm film. These- sections were then stained with hematoxylin and eosin and the same fields rephotographed. After de colorizetion and silver impregnation (Gordon and Sweets), the fields were photographed for the third time. A fourth photograph of the same fields was made after the sections had been subjected to microincineration at 600 C for an hour and treated with concentrated hy drochloric acid after cooling. Representative lesions in the lungs of control animals were also photographed for purposes of comparison with the experimental basons. There were lfl rats. 29 hamsters, aad 13 guinea pigs that had been part of the shipments of the dust- The rats and hamsters were injected in- animals later put on test, but had been set Tratrachealiy, some repeatedly, as listed in the aside away from the dust exposure as laborato table. The injections were made under light ry controls. ether anesthesia with the aid of an illuminated The results of this investigation are based speculum that allowed visualization of the vo largely upon a comparative study of about 700 cal chorda. fields selected from the sections of 76 animals. Some animals died at various times following the imposition of the dust borders. Others were Results killed approximately two years after the imposi- Rats (Intratracheal Injection).--Immedi- Arch Environ Health--Vol 15, Nov 1967 CO --1 CD CD cn ST0011611 640 EXPERIMENTAL ASBESTOSIS--GROSS & DE TREVILLE 2 9 iIO O iSi Fig 1.--Ash pattern of scid*insolubt material In lung of rat Immediately following intratracheal injec tion of chrysotlle dust. Alveolar ducts are outlined by fairty thick, continuous coating of dust. Some dust, mostly In tha form of discontinuous deposits, is also found In tome distal alveoli. Section was subjected to microincineration followed by treatment with concen trated hydrochloric acid (X 150)- ately after the intratracheal injection of chrysotile, the dust was seen applied as a uniform and continuous, dense coating upon the respiratory surfaces of the alveolar ducts and their evaginating alveoli (Fig 1). The lumens were everywhere widely patent. Sur face coating by the dust of some of the distal alveoli was also observed, but not regularly. After 72 hours, the proximal portion of the racemus5 was a nearly solid, cellular structure in which the lumens of the respira tory bronchiole and alveolar ducts were oblit erated by a polypoid mass of ovoid and plump spindle-shaped cells which also oblit erated the lumens of the evaginating al veoli. A highly significant change in the distribution of the asbestos dust was now Fig 2.--Aah pattern of actd-iraolut>l* material, Hire* day* attar Intretrachasl ln|ootkM of chiytoMa dost In rat. Dust, now In tha form of flotation* massas of speculated aggregates, saams to term casts of alveolar duct* and fill* their lumen*. Vary UtUa dust Is noted in the peripheral atveoh (X 150). observed. The dust permeated the occlusive inflammatory tissue and thereby formed a cast of what was once the lumen (Fig 2). It was, however, no longer compactly disposed; rather, it was distributed in a fiocculent manner. Nevertheless, the total amount of dust in the lumen* of the ahreolar ducts ap peared to be considerably greater three days after the intratracheal injection than that present immediately after tbs bijection. At the same time, leas asbestos dust was ob served in the more peripherally situated al veoli It is probable that the increase in the amount of dust in the alveolar ducts was de rived from the more peripheral ahreolL Al though some of the finer dust may have been intracellular, the greater bulk, by far, was extracellular, being imprisoned in the inter- Arch Environ Health--Vol 15. Nov 1967 RT001161? EXPEMMEXT \L . S ..(.UOSS & DE TREYILLE 641 STOO!1613 Fig 3 --Minimal, moderately healed asbestotic lesion in rat exposed to chrysotile dust for one month, then pastured and killed 24- months later. Lesion consists of a partially thickened alveolar duct. Mural thickening >s caused by hypocellular connective tissue that has ob literated alveoli near bottom of held (hematoxylin and eosm. (x 150). stices of the precollagenous stroma of the inflammatory tissue. When examined one year or longer after the intratracheal dust injection, asbestotic fibrosis was found in all rats so injected. The characteristic lesion was sharply limited to the alveolar duct as well as to the short re spiratory bronchiole and consisted of moder ate to severe collagenous thickening of the wall, often also associated with striking hy perplasia of smooth muscle tin more than one third of the injected animals h The col lagenous mural thickening involved much of the wall and resulted in the obliteration of many of the evaginating alveoli. Generally speaking, the involvement of the alveolar ducts, even in animals injected with but a Fig 4.--Field same as in Fig 3 after silver impreg nation shows that scar is composed on nonbranching, largely parallel, thick collagen fibers that are fairly densely arranged (Gordon and Sweats. (X 150). single do6e of 3.5 mg of chrysotile dust, was greater than the minimal lesion observed in rats that had inhaled high concentrations of the dust for one month. As judged from the examination of a single section of both lungs of the rats, greater dust dosage was often, but not always, associated with an increased incidence of thickened alveolar ducts. The occurrence of larger collagenous scars con taining small remnants of air spaces seemed also to be related to the multiplicity of the asbestos dust injections, as was a metaplasia to the columnar variety of the epithelium Lining the surviving evaginating alveoli. The sharply delimited mural thickening of the alveolar ducts fFig 3) and respiratory bronchioles, with the collagenized, nonarborescent, thick stromal fibers fFig 4), and the associated reduced cellularity fFig 3), .trc<h Environ Health--Vol IS, Sou 1967 ST0011613 ,;42 .'.'.W'/'/.'/..,'.' ' ' \7 > i' -1 ',YJv s v JiE 7 Jit `! 1 LE oo Oh Fig 5.--Ash pattern of acld-lnsoluble material of same field as in Fig 3 after microincineration and treatment with concentrated hydrochloric acid, showing the presence of tangled masses of asbestos fibers in the regions of the scar tissue (X 1503. represented a healed or healing inflamma tion. Within this sear tissue, apparently sequestered compact masses of asbestos fibers were often strikingly prominent after microincineration ' Fig 5. ) On the other hand, msav scars were seen that contained little or to- demonstrable riu.=t. The sharp delimita i >n of the- scars was accentuated by the ad jacent normal delicate alveolar walls. These were characterized hv single, nonbranching argyrophilic fibers. Nevertheless, in some of the rats, there were also foci of a more active inflammation. Such foci were marked by cellular thickening of alveolar walls and by considerable desquamation. Here, the stroma was arborescent, the side branches tending to give structural support to the increased num ber of alveolar cells. Since similar inflammalorv foci of cellular alveolar mural thicken- Fig 6.--Hyperplasia of smooth muscle In rat injected twice with 3.5 mg chrysotile dust and killed 21 months later. Well-defined bundles of smooth muscle are found in relation to alveolar ducts where the only normally present muscle is situated anund mouths of evagmating alveoli (hematoxylin and eosm X 150). ing were also found in the lungs of control rats not exposed to. or injected with, dust, it is believed that they were caused by un related spontaneous disease. No nshestos bodies were found in rats ex posed to or injected w ith chrysotile asbestos dust. Rats (Inhalation t.--The characteristic minimal lesion, as found a year or more aft er the pulmonary deposition of chrysotile dust, consisted of focal mllagenous thicken ing affecting patchc- ,f alveolar duct wall between some of : . evaginating alveoli 'Fig 3 and 4). With more extensive deposi tion of the chrysotile fibers, tlie mural fibro sis tended first to narrow the mouths of the evaginating alveoli and then to close them Arrh Environ Health--Vo! Jo. ,\'ov 1967 ST0011614 1. \ I- JS.S AT HE TJlEYil.I.K f>4d wO CD O CT> cn Fig 7.--One cf smaller cellular loci in lungs of hamster injected twice intratracheaily with 3.5 mg chr/sotile dust. Animal died one year later. The alveo lar structure of cellular tissue is obliterated hy cellula' proliferations. There is a suggestion ol o polypoid mass slightly to the right and above the center of the fie'd (hematoxylin and eosm. y 150). Fig 8.--Silver impregnation of same field as in Fig 7 shows a plethora of branching and interconnecting argyrophilic fibers forming thick alveolar walls and projecting into air spaces as polypoid masses of van ous sizes. Largest polypoid mass is one refemed to In Fig 7. It fills the lumen of alveolar duct (Cordon anti Sweets. X 150) uft entirely. The fibrosis also dipped into some of the evaginating alveoli. partially or completely obliterating them. In this man ner. the respiratory' bronchiole and adjoin ing alveolar ducts mav become converted into relatively smooth-walled tubes with few or no evaginations. Most of the wall thick ness of these tubes was attributable to the fibroticallv obliterated alveoli that original ly evaginated from the former. The cellularity of the thickened structures varied considerably depending upon the time interval between the deitosition of the dust upon the parenchymal surfaces and the death of the animal. In the florid stage of the inflammation, as seen in a rat exposed to the chrysotile dust for four months and killed two weeks later, the cells in the affect ed regions were large and numerous while the stroma consisted of an arborescent net work of argyrophilic fibers in which the cellwere enmeshed. In time, the nrgyrophilii fibers became thickened, condensed, and lost their arborescence as well as their argyre philia: the latter coincident with their col lagenous conversion (Fig 4). During this process of stromal maturation, the cellulari tv of the tissue diminished and the cells elongated to resemble fibroevtes. In some of the rats examined a year or longer after the dust exposure, there were, in addition to the collagenous foci, cellular foci of more active inflammation that had no ap parent relation to the proximal portion of the An/i Km tt on Health--Vo! 13, .Voc 1067 ST0011615 . in-: . ! co --I CD CD cn Ch Fig 9.--Ash pattern cf acid-insoluble material shows mat region of polypoid mass in alveolar duct is densely permeated by tangled masses of asbestos fibers. Small er masses and aggregates of dust are found m periph eral alveoli and other air spaces (y, 150). racemus. Some of thcsr f>'ci ni.iv also onlain variable numbers ui poivmorphonucicar leukocytes. Similar cellular loci oi active inflammation were also found in some of the unexposed laboratory control rats of compar able ace. Lung sections of all lb ru.s exposed une month or longer to heavy concentrations of chrysotile dust exhibited minimal fibrotic pulmonary lesions one year or longer after the exposure. In contrast, in but one of dx rats exposed to the same dust for nnlv two weeks were similar dunces lound The oth er five exposed animals had lures that did not differ irom those of tinexposcd rats. In neither of two rats exposed to the same dust for only one week, was there any t-udemv of asbestotic fibrosis 11 and 1'1 months respec tively. after exposure. Fig 10.--Numerous luminescent asbestos i>c-d*?s in region of asbestotic inflammation, from the of a hamster that received intratracheal injection <rf 3.5 mg chrysotile dust and died !33/i months later (unstained section: 12.5/0.3 leitz 'directive and 0 9 Leltz darh field condenser x 150>. ( ompared wi ;>e amount oi acid-insolutile ash s., .. in :*! lime ovtions shortly aft er the riu-: .vis,sure, it apoeared that only a small frai i.un of ihis amount of dust could be seen in the lungs of rats one nr more years after the exposur. N sertheiess, in view of the ultramicroscopic size of much of the chrysotile dust, the failure nf this method to demonstrate mineral du.sl in the tissue must not be (v ineri to indic.de necessari ly that no dus" .- present. It is also of in terest til.ai tb-. i rkfield examination of the unstained m-. 1 -:> wi-rh dry objectives was negative for hiimmsccnt material, even in regions later shown to upturn abundant icid-irisnluble mineral a-.h In the lungs of a numi- r of rats exposed 11j chrysotile dust two mn.-uhs or more, there Art/' /-t'U'tti 111 tj! t }i--I'-af IA- .Vac-- lljr7 ST0011616 STOOI I 6 I 7 EX PERI M ESTA L A a BESTOS IS --GROSS Jt DE TREV1LLE 646 were bundles of smooth muscle in.it often tins'itutcd masses oij/j tu 500,.. :n I hick ness Fig hi. These muscle bundles wen' -hnrply delimited and related lo the wall of thii-kencd alveolar ducts. They had no con nection with blood or lymph rebels nor could a relationship to the muscuiaris of a terminal bronchiole be demonstrated. In the newly formed connective tissue about some of the alveolar ducts and respiratory bron chioles, there were scattered gland-like structures. These pseudo-glands represent surviving units of the evaginating alveoli that have become lined by tall columnar epi thelium, occasionally ciliated. No acid-insol uble mineral ash was demonstrable in the area occupied by the proliferated muscle fibers. The regions, pervaded by the more cellular and active inflammation, some con taining leukocytes, were also devoid of de monstrable mineral ash. Hamsters (Intratracheal Injection).--Of 19 hamsters, 16 died as a result of the asbestotic inflammation that followed the intra tracheal injection of chrysotile dust. The three survivors had received only one injec tion and were killed 21 months after the im position of the lung dust burden. The aver age survival of the hamsters that had one intratracheal injection was 18% months, with a range of 13% to 20% months. The animals that had received two intratracheal injections survived, on the average, 14l/4 months from the time of the first injection, with a range of 121. -2 to t6'/2 months. The three survivors that were killed after 21 months were emaciated and also had exten sive asbestotic disease. Many of the lung sections showed exten sive consolidation with relatively few patent air spaces. In other lung sections, the consol idation was patchy', seemingly concentrated about larger air spaces with relatively little aerated tissue between the consolidated por tions. The consolidated portions were com posed of masses of cells with vesicular round or oval nuclei--apparently alveolar cells i Fig 7). Leukocytes were not seen. Al veoli or larger air spaces usually could not lie identified in these regions except after silver impregnation Fig 8i. The argyrophilic mural stroma of all air spaces in the solid ified regions was greatly thickened by ar borescent fibers that formed a loose network encroaching upon the lumens. Occasionally, the argyrophilic stroma extended into t! lumen of an alveolar duct or a bronchiole :i a polypoid manner, occluding it. The cr...densation of thin argyrophilic fibers to for a parallel thick fibers devoid of arboresceme and the transformation into collagen, chmacteristic of the lesions in rats, was not st--n at ail or to a very minor extent in there hamsters. For the meet part, the occlusion ( the air spaces was caused mainly by mas***! alveolar cells, but the contribution of the <rgyrophilic stroma should not be dismishrd. It should also be emphasized that in con trast to the situation in rat lungs where me involvement was sharply limited to the dveolar duct and its evaginating alveoli, there was no such delimitation in the hamster lung. Here, the cellular as well as the stro mal proliferation extended without abate ment to the peripheral alveoli, thus account ing for the diffuser>e3s of the consolidation Following incineration, abundant aodinsoluble mineral dust was observed in the involved racemi. Many individual fibers could be identified; some straight or wavy, others coiled and forming tangled masse*. Compared with the amount of dust found in rat lungs, the chrysotile dust in the hamster lungs was much more abundant and nv.-ee diffusely distributed (Fig 91. Asbestos t> . ies were found in abundance throughout affected regions. They were considers smaller than these seen in human lungs : were readily recognized They were seen under dark-field illummation with objectives in unstained sections (Fig ' However, there was no parallelism bwthe number of luminescent asbestos ooO'~ seen in the unstained sections m-i itamount of dust found in the same field ifter microincineration. The amount of dus' ores ent was much greater than might sus pected from the number of asbestos hndie.observed. Most of the du9t and the asbestos bodies were situated in the air spares. When observed within tissue, the asbesr.vt Ixidies and the dust were found trapped w ithin the network of inflammatory argyrophilic fibers and associated cells that tended to occlude the air spaces. Guinea Pigs (Inhalation).--Barely r-cog nizable, minimal mural thickening of an oc casional alveolar duct and respiratory bron- Arch Er.tu un Health--Vol 13. Sot' 1967 ST0011617 r rtTSiS--<-Ht-sv A- DE TP.EY1LLE / I'T Vui c\|>osf<! : -r 'hire :: .'me.-ph.-re contain.ng .. 1 in n-v.-ituo <!-:.-5l. .'j.'tuLu [u !!.! i[ij.1 -1' nee o| :.-h l-.-aous in :i.r;i-t<-r-_ To mural Tivkin . i:'Usjslftl of ; rxii::v-in-d ul'.viu.u evils :-.:i if supporting .-trum;; of intern um et vyoiphniv idler-. ! he !i:ir; -Xiaunx i pigs i;it hud innaied high omci n: 'o mrua in; uf r'nr>-ntilr. ilur-t mr week.- i >r iou.er. manifested similar r.n- mural ashestotic thickening in the .vimni portion of die r.ici-muo. Ilimiiif, number uf uniLs r.ict-mi; involved was i.Mich greater than that in the animats ex posed to the lower dust concentration for nree months. No evidence of healing tsig- ;ic.mt reduction in ct'ilularitv of the lesion ..nd coilageni.uiLiun of the1 stroma 1 was seen in animals examined up to 'even months aft er the i t eaming of the exposure. Following microinciner.ition and treatment with acid, the amount of ash seen in the guinea pig lung sections was comparable to that noted m rats that had inhaled chrysotile dust. As bestos todies found in the lung sections were few and very small. disappearance <( chrysotile dust from the sipii. i it her hv c;.v*>lutior> cm- by transport, ii<.t a sine qua non of healing; but that heating, at least in the rat. does take place-- even in the presence of this dust. One would like to think of the limitation of the asiiestos tody as a protective mechanism by which the asbestos filler becomes sequestrated and the tissue? safeguarded from further irritant action by the li!x:r. To what extent this mechanism may apply is not known: but it does seem that in ruts, in which asbestos Ixxlies are not demonstrable (with the opti cal micro.scofie >,! healing occurs in the pres ence of apparently naked fibers. The diminution, in time, of the amount of dust demonstrable in the tissue and its apparent disappearance in some of the scars, [x).s<s an interesting question in regard to the mechanism bv which the asbestos fibers disappear. It is commonly believed that chrysotile fibers have a relatively high solu bility in tissue fluid. This would seem to ac count for the inability to demonstrate asbes tos fibers in some of the asbestos bodies found in human a.-Lcstotic lungs. However, dissolution of the fibers, particularly of the coarser ones, would require a long time--so Comment A chronic pulmonary inflammation may 1 <* tinned progressive if it extends from its original site into adjoining, previously nor mal air spaces, and if it remains active, re taining its argyrophiiic precollagenous stro ma and liich celhiluriiy. Such a pulmom rv inflammation may lx: considered healed if its argvrophilic stroma has teen completely < xliagcnizcd while its rcllttl.u itv has liocome considerably reduced. In the rat that has inhaled high concen tration:-. of chryviliie .j'bostos fibers for only a few months or has been injected intra Ir.acheally with this du.'t. the asbe-stotiv in flammation remains sharply limited to the proximal [lortion of the racemus and heals by becoming transformed into a hvpocellular collagenous scar. Thus. it would appear proper to classify adiestosis i eased by chrys otile dust as nonprogressive in the rat. It is of basic interest that appreciable amounts of aslvstos fibers are demonstrable within the scars of healed a- healing that it would lie difficult to explain the fail ure of jvripheral alveoli to become involved by inflammation. Furthermore, high solubil ity would no; !x> consistent with the 6harp inflammatory mmlir.atinn of the asbestotic lesion in the prudm.ii portion of the racemus in rats. It is iou. more reasonable to explain this sharp I.- ili/.uiun of the in flammation on a fairlv prompt removal of (lie inhalrd or injected irritant dust from the {XTiphera'i alveoli and the subsequent stag nation of the dust in the proximal portions of the racemus." The train-port of the chrys otile dust from the (wiripheral alveoli is ef fected by the alveolar clearance mechanism consisting of a proximally moving film of alveolar fluid." This transport is dramatical ly illustrated by the increase in, and concen tration of dust in the lumen of the alveolar duct within 72 hours after the intratracheal injection of chrvsniile dust fcf Fig 1 and 2). It may be of interest at this point to note that the localization of the early asbes totic lesion to the proximal portion of the racemus was first described by Vorwald et inflmrunation. This would suggest that the ol, and recently confirmed by Holt et al.* ,\r< h Ei.' 'run Ihnlth-- l`ol 15. iVoe 1561 <n --i CD CD cr> CD ST0011618 6 19 1 001Si EXEEIHMEXTaL .sBES'IeXiS--vnOSS & DE TREVILLE 647 Tile reaction of the houi-te: lung to chrys- >ri!i* dust is the antithesis of th.it ohseried 01 rots. Tiie lesion extends from Ine proxirmi [xirtion of tlie rau mus to the peripheral ilvE-oJi. I hereby producing extensive uin-niidutions in the hamster June's. Tlie consolida tions are composed of obliterated air spaces, llie lumens of which are filled with masses of alveolar cells, mostly desquamated 'Fig tv. Although the inflammation involves the en tire racemus in this animal, the proximal portion shows more severe involvement than the peripheral portion. It is this ex tensiveness of asbestotic involvement with consequent pulmonary inadequacy that ac counts for the high mortality of the ham sters. With the previously given deSnition of progression in mind, it is seen that the diffuse involvement of the rucemus in the hamster would fit the first requirement for progressiveness of the disease, if it can be shown that the inflammation originally was confined to the proximal portion of the racenius and then extended into the peripheral air spaces. This information, unfortunately, is not available from the present investiga tion. However, the finding of a more severe involvement of the proximal portion of the racemus favors the probability that such an extension took place. The second require ment of progression, ie, that the inflamma tion remain active and nonhealing, is a con spicuous feature of the asbestotic hamster lungs. The failure of the argyrophilic fibers (o lose their arborescence and to collagenize and the persistance of the extreme cellularily of the involved tissues nearly two years after the Lntrapulmonarv dust deposition is incontrovertible evidence of nonhealing. Two possible explanations come to mind for the difference in the disease produced in the rat and the hamster by the same chrysolile dust. One explanation is that the reactiv ity of hamster tissue to the dust is greater than that of rat tissue. The other is that the hamster's alveolar clearanoe mechanism is not as capable as that of rats in transporting the dust from the peripheral air spaoes proximally. As will be seen below, both explana tions apply. There is some evidence that the hamster's pulmonary clearance mechanism is not as effective as that of the rat. This is seen in tiie considerably larger amount of acid-insol uble mineral ash demonstrable in the ham ster lung sections than in those of rats. Fur ther evidence is seen in the diffuseness of the dust distribution in the hamster li t? 9) in contrast to its restriction to the proximal portion of the racemus as in the rat Fig 5). The. reactivity of hamster tissue to chrysotile,fibers is significantly different from that of rat tissue, the former being much more florid. This is seen not only in the plentiful production of asbestos bodies in hamster lungs but also in the failure of argyrophilic precollagenous fibers to mature into colla gen as well as in the undiminishing cellularity of the inflammatory tissue even after a lapse of nearly two years. This difference in the reactivity of lung tissue in the rats on the one hand and in hamsters on the other is not unique to chrysotile dust inasmuch as simi lar and quite comparable differences in these two species have been noted in their reactivity to quartz dust.10 It is highly probable that interspecies differences in the efficiency of the pulmo nary clearance mechanism play an impor tant role in the susceptibility of the different species to certain pulmonary diseases. It is recognized, for example, that horses and mules working during their lifetime in mines exposed to quartz dust will have accumulat ed a negligible amount of silica in the lungs and will demonstrate no silicosis;1' whereas miners in the same workings will have vary ing degrees of silicosis as well as a sig nificantly elevated pulmonary silica content. We have recently exposed rats and mice limultaneouslv to quartz dust in the same in halation chamber for the same length of time. All animals were killed immediately after the end of the dust exposure W. i defined silicotic nodules were present in the rat lungs; whereas, the mouse lungs were aT solutely normal, and no acid-insoluble <htjt could be demonstrated in the mouse-lung sections after microincineration (unpub lished data). The above observations are relevant to the present investigation insofar as such in terspecies differences tend to make extrap*, lation from animal data to human disease unreliable. The questions to be resolved ho whether the efficiency of the cleanri**mechanism of man approximates that of tlv Arch Environ Health--Vol 15, Sov W67 ST0011619 km'F.iir.\i \ 7(, 111 >ss & !>h. TRf VII !.K 649 The chunictcristic c:irlv .i.-i>e.-,tuiic lcion is situated in. and sharply localized to. (he proximal portion of the pulmonary rarernus which. in (lie rat. oinsists or a very 'hurt respiratory bronchiole and adjoining; aheolar ducts. Astx'stosis in rats, cwii.-ed hv chry>otile dust I- nonprogressive. ! The evidenre for the nonprogressiveness of chrvsotile asbestosis in rats consists of the observations that the minimal asbestotic lesion is limited to the wall of the respiratory bronchiole and of the adjoinin': alveolar ducts and does not extend into ad joining normal alveoli and that the minimal nsbestotir lesions heal by hecornin'; colly geni/.ed and hyj)oci*lliil.tr. t. The localization of the minimal asbestotic le-tons is attributed to the proximal transport of dust from peripheral air spaces by the alveolar clearance mechanism and sulvscquont stagnation of the transported dust in the proximal portion of the racemus. a. In rats chrysotile asbestotic lesions ho d in the absence of demonstrable \ optical miem-cupe asbestos bodies and in the pres ence a' chrysotile fibers, the latter becoming trappr-d in the scar tissue. u. There is a considerable reduction in the amount of chrysotile dust found in the I tine sections of rats one year or more after the im[>osition of the lung dust burden as at11'.pared with the amount of dust present shortly after the dust burden was imposed. T. The read ion of hamster lungs to chrysotile dust is the antithesis of that observed in rats The lesion is progressive, extending into [ieripher.il air spaces so as to involve the entire racemus and thereby producing 'xtensivv consolidations leading to the death of the animals. The lesions do not heal, since the precollagenous stroma and the high cellularitv persists through the twoyear [xriod of observation. At the same time, the amount of asbestos dust demon strable in the sections remains large and HilTa-cly distributed. T The progressiveness of the chrysotile ashcstotic lesion in hamsters is ascribed to a less effective pulmonary clearance mecha nism and tu a greater reactivity of the pul monary' tissue than exist in ruU I). The early chrysotile nsix-stotic lesion in guinea pigs resembles that ut t us. hm it has not been adequately rtudica 'o permit further characterization. !"his >tudv wis supported .n jm.' !>' ! ' >;it IR.dtl- vy*rvi e ^iant OH (XU32. John--M.ur. <*oir.pan\ through ihe rourtrsv of Kfini'i'i ^ "'ndh. MV -.upping} u\o rhrvsoMlo References 1. Ross, W.D.. ct a): Emotiofwd Asp** u of Rcspi raforv Disordci** A/non*; F.d JAM* 156:484 (Oct 2) 1954. 2. Holt, P.E., and Vt-unp. OK 5 Dust-Fw'i- Mechanism Suitable for Fibrous Du-t Aar Chv:,/- Hyg I960 3. Wriffht, R.M : A Size-Selecting Sampler `nr Air-Borne Dust. Br:t J Imh.dr Med 11:284 19M 4. Gross, P.. and Tolkvr. Eli Ou3t P.:--: :ti/ m Lung Sections: Some Notes >n Method.- o - r Visualization, Arrh /vntirr-n Health 12:21- 5. von Hayok. H : The Human Lung. V !. K-. ftrans), New York Hnfm*r !'ublishing . Inc., p 172. 6. Vorwald, A.-I : Dttrk.m. T M.: and Pratt. ! Experimental Studies of Asbestosis, ArrA lnd>. Hyg 3:1. 1951. 7. Gross. P.; Pfitzer, E.A.; and Hatch, T.F.: A veolar Clearance: Tts Relation to Lesions of the R. .*>piratory Bronchiole. Amrr lu r Rtsp Ois, 94`A* 196*1. fl. Hatch, T.F . *ruJ Gnws, P Pulmonarv Dt iio-. lion and Retention of Inhaled Aerosols. New Yu ^ Academic Press, Inr. 1964. p 69. 9. Holt. P.F.: Mills. J ; and Young, D.K.: Ear'*, Effects of Chrysotile Asbestos Dust on Rat Lung. ' Path Bad 87:15, 1964. 10. Gross, P.: dcVillirrs, A.J.: and HeTreMp- R.T.P.: Experimental Sdicosis. Arch Path 84:8" i 1W. 11. Einbrodt, H -I ; Klosterkotter. W : and V- * H.: Vergleichendp Untprsuchumron uber <tie K (jr'isson retinierter St.'ulv- m don Luneen - >n M und Ti**r. Beilr Sihfiosefnrsch 6:491. 12 Gross. P. and Smith, K.W.: 'I'he T.'-poirr*: ! )isl, dmtion of Mineral Ou^N in Sonu- I'm- . coniotic Lungs. Dis CFir*t 35:140 1959 13. Gross. P.: "Patholngv /if the I'neum*' / nioscs." in Ijin/ta. A.J.L '<'/} > 7 he I'acum/ennn New York: (Irune and Snaunn In/ . pr 14. Knox, J.F.. and Beattie J.: Mineral For-, of the I,unEp$ After Ex|>OMUir* io Asbestos I Hist. : Industr ff\g 10:23. 1950. 15 Nagels^hmidL G.: Some Observations c4 Dust Content and Comi>*sitifju in f.unip* With hestoais, Ann NY Acad Set 132:64-78. Ia-'V> 16. Rayi' SC.; Kinif, E J . and Harrison <' V.: i Action of Variable Amounts of Quartz on the 1 .. - of Rats, Brif J Induntr \frd 8:62, 19'd 17. SU-bert, FT. and Kislur. E.R.: Brunch-- r Emphysema. /If 'f I l\:!h 33:1137, 1357 \rrii f.nitron ffenlth--l'/d />. .Yot* 1967 oo --! O o or ro CD STOO11620