Document 7Oxoz61G09Qrm7pKw99gqYj7B

WASHINGTON DISTRICT OF COLUMBIA TM\DAVIT ) ) ss. ) BEFORE ME, tb undersigned Notary Public, personally cane and appeared MICHAEL RHODE, a person of the full age of majority, being of sound mind and body, who, after being first duly sworn by me, did depose and say: I, MICHAEL RHODE, am an employee of the Armed Forces Institute of Pathology, Washington, D.C. I am custodian of the original archives of Dr. Arthur J. Vorvald. Pages 1 through X2. attached hereto are true, correct and authentic copies of documents from Dr. Arthur Vorvald's archives. The pages attached are____ ASBSSTOSIS - Experimental Studies by .The Sar.a.nflc_LaboratQrvJ Report to the Johns-Manville Corporation, dated September 30. 1948 Dr. Vorvald was the Director of the Saranac Laboratory of the Trudeau Institute. Upon his death, Dr. Vorvald's archives were donated to the Armed Forces Institute of pathology by his vidov. Dr. Vorvald's archives are in such condition as to raise no suspicion concerning their authenticity; they were received by the Armed Forces Institute of Pathology from Dr. Vorvald's widow after ?J?ODE A/FOAVTT - 1 * * .V Dr. Vorvald's de*th: thsy *re presently cn file <'S authorized by law in tha National Museua of Health * Medicine, Armed Forces Institute of Pathology, Building 54, Walter Reed Army Medical Center, Washington, D.C. 20306-6000; and the archives have been in existence for 20 (twenty) or aore years. A__ 6uFIANT SWORN TO AND SUBSCRIBED before ms, the undersigned Notary Public, on this the day of April, 1992. NOTARY PUBLIC Printe:eddNaamaee: LtiCr RHODE ATTEDAVTT - P*o 1 at t f\ 4 ASB3SX0SIS Bsporlmatal Studies by THE SAJUXAC UL30HAT0ET SAEAEAC LAKE, HE* TOES Haport to tha JOHVS-UOTZLLE COBPORAXZOI JOT TORE, REV TOOT Soptoabor 30, 1948 . AC./j2t t 2. Introduction 3. Asbestos Mistrals *. A33T2ACT 'c* 17. ZSTSHSmL 1SHE5TCS25 attiods 6. Spades Susceptibility' 7. Peculiar Giaraeterirtici cf Asbestos 3. ?-ate of Tissue Fraction to labostos Fibers ?. isbestosis Bodies i. nRAunoa espsaisans n. Xing's Floats asbestos Dust 12. Dusting Material 13. Dust Composition 1L. Dust Concentration 15. Reaction is 16. Guinea Pigs If( Bats and IVpe of Reaction IS. Progression ^ 19. Infection Coincident dth Dost Inhalation 20. Infection after Dost Inhalation 21. Aebestosis Bodies 22. Babbits 23. a*ts 2li. Sursiery and Interpretation 6 7 -' 7 8 0n e 8 9 9 n 1C H ^2 12 12 13 t c i; 9 YA CM Short--fiber Asbestos Dust 26. Dusting Uaterial 27- Dost Composition 28. Dust Concentration 29. Size-frequency of Dust 30. Enaction in Aniaals 31. Ouinea Pigs 32. Hate and Type of Reaction 33. Progression 3k. Asbestosis Bodies 35. Rats 36. Rate and Type of Reaction 37. Cats 38. Rate and Type of Reaction 39. X-ray Chances ko. Asbestosis Bodies 111. Rabbits Ii2. Rate and Type of Reaction Ii3. Asbestoeis Bodies Ui. Stannary and 2hterpretation us. 100 Par Cant Ball-silled Asbestos Dost Ii6. . Dusting Material L7. Dust Cosposlticn Ii8. Dost Concentration k9. Sice-frequency of Dust - i ;i - Page Ill 111 15 15 15 15 15 15 17 17 13 18 19 19 19 20 20 2G 20 21 21 21 22 22 23 ^ A '* W 41 MU 4 50. Reaction Is 51. Gainsa rigs 52 Rata and Type or Reaction 53. Progression Si. lymph Node Involvement 55. Asbestosis Bodies 56. Bats and !2Lee 57. Sunnary and Interpretation 50. Long-fiber Asbestos Dust 55. Dosting :Material 60. Dost Composition 61. Dost Concentration .62 Size-frs<iuenc7 of Lust 63. Reaction in Animale 61u Guinea Pigs .65. Rate and Type of Reaction 66 Progression 67. Lymph Node Involvement .68 Asbestosis Bodies 69. Cats 70. Rate end Type of Reaction 71. X-ray Changes 72. ?-ats 73. Rate and Type of Reaction 23 23 23 2L 2k 2k 2k 25 26 26 26 26 27 27 27 27 28 29 29 30 30 30 30 31 ilicc 'iats ani ^: .'zzurf fci;-. r..icr-v^ta<v.. tocth. isjacnoa .o:-i=.r. -ss ?C, Intratracheal 'irrrrlrastc 7.7. Coapariscr. of - r.brous ''cr.-.'.Mrsc.* . trirf 7'C. Comparison of Tariots Lonr-fibsr Dust? 21. Ccsparison of locc-flbcr and 5>.rrt-fi*:\_- Trots 32. Intravenous Superusats 33. Intraperitnasal Ifeperinsnts Ul il> ICTC7. ocra s.C'Zizr-:i2 5. Protective Action of Ali^isnn Ccrsiiacs.: 6. Fcraation of Asbcstosia Todies IJCCTII. THSCP-7 -? L.R.ITA'JT ACTIO" I* UT7III. COMPLICATIONS 9. Infection ?C. Susceptibility to Tusareulous Infection 71. Susceptibility to ncn-fcutjercuiras Infcotton P2. rteoplass ;:cin a<*(.**+* **r- T W*VJ IWWV .W xcrv 'Mr >32 jo 3t JtT 35 35 36 37 37 3S 33 -jc i;C m - ~r 72 /3ST3ACT Asbestoals is a pulmonary disease caused by the Inhalation of asbestos dust. In animals it is characterised by a peribronchiolar fibrosis which seems to be the result of mechanical, rather than chemical, irritation of the tissue by asbestos fibers. Only the long fibers produce a typical reaction; short fibers are relatively inert. The fllaaented structure of the fibers is an essential factor in the mechanism of irritation. A character istic tissue response eat be produced by noa-elliceous as well as siliceous fibrous minerals. Inhalation of asbestos dost apparently does not alter significantly the course of experimental tuberculosis la gulasa pigs. The asbestosis body, which is a specific coacoaltant of asbestosls and forms soon after the entrance of the asbestos fiber into the lung, is believed to prevent further damage to the tissue by the fiber and thus to limit pro gression of the reaction when exposure ceases, ilunrinum does not exert a protective action against the tissue irritation of asbestos fibers as it does against that of quarts particles. 1. XPra23HTAL 3TITIT5 AS3Z2TCSIS 2. Introduction lsb3tc3is is a Terr c- pnetraoconiosis resulting iroa prolonged inhala tion o- ssbestes civst. Tuc nans asbestos, literally "unburaable, is not that ex* a particular mineral but is a term applied to a nuaber of different minerals Those characteristic feature is a structure composed of long, rar- n^l I -- allcl, flamiblg fibers. This structure is unique because the fibers are capable of repeated longitudinal subdivision to units of molecular proportions. In length the fibers ''ary free a few microns to six or more Inches. 3sirs varieties are stiffer than others bat many are sufficiently flexible to be spun into yarn and ro7en on modified textile machinery. 3. Asbestos Minerals The asbestos ninerals are silicates of variable composition and belong tc the serpentine and the anphibele groups. Listed held* are the more coamon varieties. Acahibole Group Anfchophyllite (Kg, ?e) silicate i'oosita (Eg, ?> H) silicate Amphibole (Ca, Vgt ?e, Al, Na, ) silicate Treaclito (Ca, Kg) silicate Actir.elite (Ca, Ug, ?o) silicate Crocicoliie (Kaf Te) silicate ?e silicate Serpentine Groca fhcvrcsile ' ' - iig silicate (hydrous) -2- The bulk of the esbestos of ccmercs is cnry;* ;ii, J0.2Si02.2H2 vtn-h ie ninaa iri Lie Thetford regisa of the Tronr: cf T---CC. Crociaoiite and anosite ore also used coccnrciallj" bat in -.-kcr. saiHer assents. Chrysotile occurs as veins in serpentina, a ninarai siriia* in chemical com position to chrysotile but which exists in massive Core ani is cade up of microscopic fibers without the parallel orientation cherac isristic of chrysetile. The massive blue black serpentine, which is xaooth nsd soapy to the touch, is -traversed by veins of fibrous cbrysotlle varying in vidth from a barely perceptible line to six or more inches. The fibers ra across the vein and not lengthwise with the formation. Attention is directed to the mineral brucite, agO.K^, -which is often found in the same formations with serpentine aad chrysotils and may be fibrous in structure* It has no commercial value at present because its fibers are not sufficiently flexible to be used in textiles but they are capable of repeated longitudinal subdivision. Ohlike other asoestlfora zdnerals, brudte is not a silicate aad for this reason it has been a valuable tool in an experimental evaluation of the action of fibrous ninerals upon lung tissue. it. npssasmi asbzstosis . For many years studies have been carried on by the Saranac Laboratory in an investigation of the cause, nature and development of asbesteals. The present re port is devoted to Experimental Asbestosis. In it are described the animal ex periments with various kinds of asbestos dust. Another report, to be prepared aid issued later, will be concerned with Human Asbestoais and will cover the health aspects of workers who have been exposed to asbestos dust in an industrial environment. Although asbestosls is & is & chronic disease shich requires touts to vitfV U PwwilMe reproduce in one or aore species of i char-- istle tissue changes ohieh are similar to the lesions of human asbestosIs. Since the life-span of the experimental animal is relatively short, it is not possible to develop the characteristic lesions in i under the industrial con ditions* Consequently, to obtain a complete evaluation of the Mff"a response to inhaled particulate and fibrous material, it is necessary to accelerate the reaction by employing higher concentrations of dust than would ordinarily be en countered in industry. TThila conditions of exposure ere thus different, the information yielded by sxparlaenta eith animals is Invaluable in furnishing a better understanding of the reaction of the human organism to Inhaled asbestos dust. 5. Experimental Methods For investigating the biological reaction of the experimental animal to the verious asbestos minerals, tiro types .of technique have been emplorsd, namely, the inhalation method and the injection method. 2h inhalation experiments, groups of **t* - up to 100 or more guinea pigs ad sometimes smaller umbers of rsbbits, cats, dogs, rets or min - are kept for eight hours a day in a cubical dost roam, eight fat in dimension, in shich e cloud of asbestos dost is maintained At intervals during the experiment, a few ere sacrificed and the tissue examined to determine the nature and extent of the dust reaction. Some animals are oeposed for periods 19 to three years. The injection experiments, in ahich the dust, either dry or suspended in fluid, is introduced into the animal by the intravenous, lutraperitohaal or intratracheal procedure, are used to determine ehether or not a particular dust has a potential capacity to prodnee tissue reaction. _ Long-torn inhalation experiments i\mish information upon which great reli ance is placed when erwmacijG *;* usgrt* ^o rrJr- a dv-t aright.be hazardous to industrial workers. 'bather cr not atmospheric dust, even though potentially dangerous, can be inhaled, pass the natural defense barrier* and reach the pul monary tissue in quantities sufficient to cause damage can be determined only by inhalation procedur-s. Injection experiments arc useful because in then contact between the dust particles end tissues is assured and the potential capacity or the dust to produce reaction can be estimated accurately. When dealing with fi brous minerals like asbestos, the intratracheal sethed is valuable since it per mits observing the effect of the fibers on pulmonary tissue. 6. Species Susceptibility Unlike free silica, asbestos does not exert its specific effect in all organs of all species of animal (Table 1). Injection of fine quarts into various organs of the guinea pig, rabbit, rat, cat, dog, chicken and even tadpole will produce silicotic nodules. However, similar injections of long or short fiber asbestos have resulted in a fibrous reaction in the lung and, to a lesser extent, in the peritoneum but not In other organs. 7. Pec^Ha*. Characteristics of Asbestos Experience has demonstrated that most of the particulate matter Inhaled into the lungs of man and is ID microns or lass In aarlima diameter. Larger particles apparently are excluded by the protective mechanism of the upper respir atory tract. In tbs case of fibrous materials, however, this restriction does not apply and fibers 100 and even 200 microns in length have been found in the termin al air spaces of human lungs. In snail laboratory animals exposed to asbestos dust the maximum length cf fiber found in the lung rarely exceeds 60 microns.. Hot every w-ind of fibrous caterial is inhaled with equal readiness; for example. the synthetic fibers if glass rmzl apparently are too inflexible to pass easily through t>o rr ae, pharynx, trachea and bronchi and seibom rsaca tlx* c.--.. --r-* bronchioles and alveoli. Inhaled particulita natter coses to rest throughout the terminal air spaces (alveolar duet, atria, alveoli) in all parts of the Imps; inhaled asbestos fibers are first retained in the respiratory bronchioles. These vary mall tubes are immediately distal to bronchioles Lined by ciliated epithelium. Their own essen tial lining is a lore ccboidai type of epithelium but, as their name implies, they actually function in respiration through lateral alveoli given off as pouches along their vails. Either these pouches, or the abrupt change in the character of the lining epithelium, or the decrease in diameter of the tube, or perhaps the combination of all three factors is responsible for local retention of the inhaled fiber. Only after asbestoeis is veil established are appreciable numbers of fi bers carried Into the more peripheral air spaces. 3. Rate of Tissue Reaction to Asbestos Fibers _ The rate of tissue reaction to asbestos is much acre rapid than to an active dust like quarts. Evidences of tissue response appear as soon as fibers have localised In sufficient concentration in specific areas. Xa rats receiving asbes tos fibers by intratracheal Injection this evidence is visible us early as tvo eeekm after injection; far quarts dust the latent period sight be tso months or The behavior of the tissue reaction to Inhaled dust after the termination of exposure is not the same in silicosis as in asbestosis. In silicosis, the young nodules become larger; in asbestosis, young scar tissue,that msf have farmed,con tracts and becomes more dense but the area of involvement 'decreases In site. If exposure to asbestos dust is tertd rated after a brief period, the recently-inhaled fibers in the lung aay cause the fibrous tissue response to continue for a -r.cn until the fiber* hare been coated. "Tiis progression is oi oru.y a slight degree and of little significance. ?. Asbestosis oodles - The peculiar structure knoro as the asbestosis body or curious bode* is z specific concomitant of asbestosis. The typical body is a golden-yellcw, beadaa or fcanstrxted rod vhich may be either straight or cursed. Often one or both ends are bulbous like a dumb-bell. The bodies tat7 considerably1 in length, and d- mansions up to 2$0 microns hare been recorded. It is believed that asbestosis bodies are due to a deposit of protein az^ iron pigment upon the surface of fibers. Is guinea pigs they fora after about 60 days of contact sith the tissue. They ars abundant in sun and the guinea pig (see Table 1) but are such larger is the former, probably because the larger- sized air tubes admit fibers of greater dissnsion. In eats, rabbits and alee thsre is an atypical coating of a few of the fibers after much longer residence is the lungs. In rats end dogs no bodies could be discovered. Although the evldeice is incomplete, it appears that the formation of tha asbestosis body prevents dasage to the tissue by the fiber. . I. DHAUTXCK ETPnUMENTS Four comprehensive Inhalation experiments have been conducted at the Saranac Laboratory eith various fossa of asbestos dust. - In each of these investigations aore than l0 animals sere used snd the experiments sere carried on for periods ranging from 2 to aore than $ years. The four kinds of asbestos dust employee are identified as King's floats, short fiber, 100 per sent ball-milled, and long-fiber asbestos dust. 11. Inhalation Experimant with c'Ling 's Floats" Asbestos Dost The first icn*ia;Lou ^pennant ;t *h* e`~r.-r laboratory with asbestos dust was begun in 1923. Animals inKatrf the dust for periods up to nearly three years and seme guinea pigs lived for about four years after their first oeposure to dust. A prnliginary report giving observations after 29 months of exposure appeared In the r ebruary, 1931 issue of TBE J01RK1L OP INDU STRIAL HTGIENE.* At that tine observations covered a period of only 2-1A years and the conclusions as to the ultimate offsets of Inhaled asbestos dust sere provisional. Results of the completed study show that most of the conclusions drawn in the preliminary report were substantiated. A complete review of this experiment follars. 12. The dusting material was a csaasrcial variety of asbestos dust known as King's floats and was composed of short fibers and particles of variable size. It was obtained from tbs Thetford, Quebec plant of the Asbestos Corporation of Aserica. 13. The dust composition (Table 2} reveals that the amount of fibrous chrysotils was only 1U per cent, a rather low value. However, there was suf ficient fibrous material to produce a characteristic fibrosis. 111. The duat concentration at first was quite low and for impinger samples taken soon after the experiment was started, the average light field count: by the standard technique was only 6.0 million particles per cubic foot of air. An appreciable number of large particles (or fibers) also were present, as S72DI2S OK EKPTR-KPTtAL PHEUIS0C0NIXI5. VI. Inhalation of Asbestoo Dust., Gardner, L.T.j and Cummings, D.S. J. Xnd. Hyg., 13: 65-81, 97-llii, 1931. -S- sV an Tn coart of 0.5 Billion far oarticles ereit:,. i.1^ ?0 micro?. Iftor tho inhalation experiment had boon under 007 for about too 70ars, tho speed of tho rotating paddle in tho dusting nachino tu increased and fcr tho remain-- lng 9 or 10 sooths of tho sspcriant considerably or* dust m dispersed into tho atmosphere. Average dust counts for ispingor samples eolloetod after ehange ware 53.7 adHian fhr the usual light-field method. end 1JS Billion fbr psrtlelss larger than 10 nicrons. . 15. Reaction in ininala to Inhaled "King*8 Floats8 Asbestos Dust. Basalts of the investigation, briefly susBarised in Table 3j shoe that inhala tion of King's floats asbestos dust produced a typical peribronchiolar fibrosis in pigs but not in rabbits or ruts. 15. Guinea Pigs. Soron groups of goiaea pigs -wore used. In throe groups the effect of a continuous and of an interrupted dust exposure was studied; in too other groups the relationship between infection and dnst exposure was investigated. The raaaining two groups were infection controls 17. Rate and Type of Sanction. Guinea pigs inhaling this dust for periods up to 33 aoaths developed a characteristic fibrosis occurring in conical pstebss about the reepiratojy bronchioles. During tvi4 exposure the peripheral alveoli were not involved. The particulate elancets in the dust were transported to the lymphatic system where they caused no signi ficant reaction; the fibrous elements remained fixed at the site of original localization and were seldom detected In tbs lymphoid tissue. Flsurisy and fi brosis in the septa were observed only when infection complicated the process. after exposure of approximately a year, a vrall amount of cannier reaction had been produced about mazy respiratory bronehxslss. is acre dust was inhaled, ih continued to aceusulate in the see location and later stages of the disease consisted of extensions or the original lesions. Own areas were not involved. Apparently, the innaled nears ear* caught In the pockst-lika elTeoli that are given off fkom the lateral walls of the respiratory bronchioles. There they rare phagocytixcd and easy oT then were carried into the wall by olgratcry cells. i'cssiBicleer leucocytes attracted to the area cessed as appreciable thickening of the bronchial wall. After 16 aonths a delicate fibrosis aide ft* appearance. The process evolved so gradually that aitotic division of fibroblasts could rarely be discovered; nevertheless, the masher of fins intercellular collagenous fibers (fibrosis) steadily increased* As this fibrosis contracted, it partially closed the alveoli, and with this atelectasis the n*nf cpitheliua assaned its embryonic coboidal fora. The result was the adencaa-lika appearance that Wills described In guinea pigs Inhaling silicon carbids. The longer exposures resulted only in more thickening of the walls of ths air spaces, largely due to an in crease in the amount of fibrosis. The fibrous tissue always remained cellular and never showed the hyalinixation characteristic of silicosis. IS. Progression. The reaction produced in exposed guinea pigs did sot pro gross significantly during a subsequent period of 37 months when the animals lived in a normal atmosphere. Batsmen 8 and 11 aonths after exposure ceased, the cellular reaction bad bean oosqilstely replaced br thin strands of fibrous tissue. Observed still longer, the scar tissus decreased la amount but in tbe last sacrificed, 37 aonths after discontinuing dust exposure, some fibrosis was still visible. 19. Infection Coincident with 3ost Inhalation. Of the group of 1x0 guinea pigs infected with atten uated tubercle bacilli (R^ strain; 31 died or mare sacrificed before two years of dust closure and were reported in the paper by Gardner and Coamings mentioned above. Seventeen of these dies from interearrant pneumonia. Briefly, the re sults sere as follows * 10 revealed sous eriosnce of spread of the tuberculous process; in 6 of these it was confined to the longs and in the other U the ahrioginal viscera also were involved. Usually a slight local extension of the tuberculous infection had occurred but subsequent healing had resulted in fi brosis of both the poisonary lesions and the nai niiiTu j - -- a other organs. .w The healed pa1nonary lesions showed sore fibrosis than is characteristic of either tuberculosis or asbestosia alone. The 9 animals which were still alive after two years of dust exposure were sacrificed at intervale during tho following year. In I* of then the primary foci of infection had healed with fibrosis and even calcification and thera was no evidence of progression. In the other $ the tuberculous fed showed evidence of having previously spread locally: in L of them it had healed, by the tine of autopsy, with excessive fibrosis; in the other animal there was a generalised chronic tuberculous pneumonia in one loba and isolated primary tubercles, which were still active but had not spread in the other lobea. Evidence of extension of the infection was first seen after 7 months of dust Inhalation; during the next 20 months more than half of the animals thawed an actively spreading tuberculosis and in 3 of them small cavities had developed. During the last 8 months no animals exhibited any evidence of active infection although la half of them tho healed fibrous scars of previous extensions were ob vious. Sixty per cant of the guinea pigs with spreading pulmonary tuberculosis showed tuberculosis of the spleen and liver. 20. Infection 3uperimposed Open an Established lebeetoeis. Twelve guinea pigs, after in haling asbestos dust for nearly 26 months, were infected with tubercle bacilli and then removed to normal air. The aubplsural tubercles in the dusted animals were no nore numerous than in non--ducted controls, but a considerably pjr,-'v- ^er fcund in the depths of the lung about foci of aabeslusis. The reaction to infac tion showed only slight local extension about the orlglrsl sites in the lungs and tracheobronchial lymph nodes. The a>vnnr^fif; riscera r.arc involved in only one animal. Caseation was found in tubercles 1-1/4 souths eld but by 5-1/2 aonths it had cosplstely disappeared, leaving only sear tissue. The latter still persisted in the last animal, which was killed lit months after infection. 21. AsbestosIs Bodies. Ibdarate numbers of asbestosis bodies occurred in - the lungs of the guinea pigs, becoming sore nuaerous and nore distinctly segseated in later aonths. 22. Rabbits. Rabbits exposed to the asbestos dust for periods up to 19 souths developed a low-grade foreign-body type of reaction but no fibrosis. Although their lungs contained particulate elcaents of the dust, fibers were not present, indicating that the upper respiratory sechanisa of the rabbit is adequate to exclude fibrous foreign bodies. Two rabbits, after inhali-g dust for 6 end 16 aonths, lived in normal air for sort than tso /ears, it autopsy neither anissl showed any evidence of cellular reaction or fibrosis In the term inal bronchioles nor ware there aqy asbestoela bodies. 23. Rats. All tbs whits rats had acquired an infection, resulting in the formation of pulaonary abscesses, before they cane to autopsy. Apparently, so such heavy sucus obstructed their bronchi that very few fibers could have entered their lungs. In a few of the rats, an occasional asbestosls body was discovered but there was no fibrosis. O 21a. Suuar7 and mtartrefcsxlor. of Inhalation Sanerimant with Ring's Floats Prut. I'iiis findings in the experiment vitn king's 1'ioai... cc.-. -y- . Lzed under three headings. 1. Effect of the inhaled dust cn normal g-i-at*. The ling's floats dost caused a charac teristic peribronchiolar fibrosis in guinea pigs but cot la rabbits or rats. The fibrosis did not progress after the dust exposure ms discontinued and the guinea pigs transferred to normal air. 3. Effect of the Inhaled dust on tiiberculoela in guinea nigs. In guinea pigs in fected with attenuated tubercle '.gtrivu and then placed in the dost room, the results were nore variable than is usual in an experiment of this type. A few showed no sign of progression; in most of then there was evidence of tem porary progression with subsequent healing; in one animal there was continuous progression to death. In contrast, when guinea pigs, after being infected, are exposed to quarts instead of asbestos dust, the infectious process continues to progress and eventually causes the death of the animals. On the other hand, ex posure of Infected to a harmless dust libs calotte or gypsum does not lead to any progression of the infection. Guinea pigs infected with attenuated tubercle following the termination of about two years1 exposure to asbestos dust did not develop progressive disease. The only modification of the infection was in its localization, a few th being retained in the fibrous terminal 'bronchioles forming tubercles there in addition to the usual foci beneath the pleura. ' In view of this variability, the unusual nature of the response and the high proportion of deaths from intarcurreou pneumonia it is felt that definite conclu sions as to the influence of this dust on the course of tuberculous infection are not justified.~ _ - - >-- C. Effect of tuberculous infection on the reactloc to T.nfo'**4--"r-?'*' **'' -i*** 1 fogr ojf the duet to produce fibrous tissue. This change -x:curr-jd whether or not the bacterial lesion ns in contact with the area of dust reaction. 25. Inhalation Experiment with Short-fiber Asbestos Duet. Since hazardous dusts like quarts are nost effective in producing fibroais when the particles are 3 microns and less in size, an inhalation experiment ms carried on to determine whether this condition is true also for asbestos dust. It was thought that h7 using a short-fiber asbestos dust consisting almost entirely of fibers and particles smaller than 3 mierens an accelerated tissue response might be Initiated and an advanced reaction obtained in a short time. The previous inhalation experiment with King's floats asbestos, which contained fibers frao 1 ssl. to 1 sicron or less in length as well as a great daal of particulate natter and which produced a typical peribronchiolar fibrosis in seposed guinea pigs, served as a basis of eosparisoa. 26. The destine material for this experInsertt was forwarded from the KanvUle plant of the Tiitins IfinrlTlii Corporation. It was the remains of fibers collected in dust bins after a carding operation and screened to pass 200 mesh. Since the material as received contained many long fibers, it wae ground in a steel ball sill to reduce practically all the partlelea to 3 aierons or less in size. Khan used alone In a standard dusting machine, this finely-ground asbestos tended to pack In the hopper and it became necessary to adz one volume of the un ground material with three volumes of the ground to generate a satisfactory dust cloud. The addition of the quantity of ungreund asbestos was unfortunate becanae it confused the interpretation of results. Probably the minor amount of reaction that developed was due to the Long fibers in the mixture although the data of this experiment do not prove the point 27. The coeaoaition of the short-fiber asbestos u rscaired is disclosed by the chemical and petrographic analyses given in Tibia h. Samples taken before and after grinding 71aided about the same mines on analysis, Indicating that there am co contnTi nation from the elll or lose of eater content. 28. The duet concentration varied somewhat during the experiment and light .. field counts for atmospheric samples collected inside the aniaal cages with the inpinger apparatus ranged from 83 to 182 Billion. The average of counts was 130 million for the first Tear of the experiment, 13U million for tiw second Tear and liiO million for the third year. 29. Slse-ffreauency measurements of air-floated dust from inside the cages at a magnification of 13Q0Z revealed a great preponderance of fine par ticles (Table 5). Searly 90 per cent of the pertieles seen were smaller than 3 microns. 30. Reaction in Animals to Inhaled Short--Fiber Asbestos Dust. Four species of animals - guinea pigs, white rata, cats and rabbits - were used in this experiment. The results of the dust exposure, which are eunsaarised in Table 6, mill be considered more in detail below. 31. Omnwa Pigs. Eighty- guinea pigs ware originally placed in the dust room but 21 of them were later eliminated from the experiment and killed because of enlarged lymph nodee. Of the other 59 animals, I16 remained in the dust room until they were sacrificed or died from natural ceases aid 13, after being exposed to dust far 20 months, were transferred to a normal atmosphere _ }2. Rate and Tree of Reaction. The type of tissue reaction to the inhaled short-fiber asbestos t*c essentially the sans as that already observed la the scperinen*. with Ki^'j floats asbestos. The rate oi reaction also was - aately the same but the extent of involvement with the short-fiber dust was very uch less aid after 16 to 2li sooths of expoeure only a very few foci of reaction, which generally required microscopic wraarf nation for detection, were produced in the guinea pigs. Until closures had continued for approximately one year, there was little tendency for dust-containing phagocytes to collect into clumps. By l aanths phagocytes had begun to collect about the walls of a few of the respiratory bron chioles with a little proliferation or infiltration of mononuclear cells in these walls* There were also some sultinccleated cells but they were always of the inert foreign-body type. It 20 to 2ii aoaths the cellular eltzsps were sometimes quite narked and sometimes changes In the epithelium resulted In the adenoma-like or "adenomatoid" appearance previously described in Section 17* In most of the subsequent members of the series, the reaction remained cellular in type. In a few, however, fibrous elegante dominated the picture* In the latter case, the collagen was pals in color and tenuous with no heavy swollen hyalinization. As in the rats described below, the alveolar walls might be made up of a band of collagen supporting a layer of epithelim, but with no contained capillaries. In the tracheobronchial lymph nodes the reaction was more pronounced in this experi ment than Is the previous one with King's floats asbestos, probably because of the transportation of an excess of fine particles to the nodes in animals inhaling short-fiber asbestos. The reaction was essentially an increase in reticulm, rather than a fibrosis, with preservation cf the original cells between the thickened reticular fibers. Diffuse chronic pleurisy without evidence of pulmonary infection was present in a few animals. -- 33. Progression. Ir. the 13-2A months following tha cessation of 20 months' axpc3urs v: drst, progression of disease was not definitely demonstrated but neither could it be absolutely disproved, owing to the varia bility of the response in different animals, it the end of the dust exposure of 20 months two pigs were rend as and arte as 2*. Among the 13 removed from dust the findings were variable: in 2 the reactisn wzs i; in li it was ; in 3 it was y+i in 3 it was U*; and in one animal it was 5+. It Is quits possible that those with the most marked changes had already developed more reaction than the remain der by the time exposure ceased. Since tbs more severe reactions occurred spora dically and bore no relationship to the length of tiae after cessation of expo sure the differences were attributed to variation in individual susceptibility. This view received support from thn chemical analyses (Table 7), which often re vealed comparable amounts of ash and silica in lungs with widely different amounts of tissue change. For example, the ash and silica values were quite similar for three animals in dust 20 months and then in normal air 13-3/lt months, yet the tis sue reaction for one animal was U+; far another, ; and for tha third, only . 3ii. Asbestoeis Bodies. The formation of asbestoois bodies was at first ex tremely limited. After $ months' exposure only a very rare short body could be found, usually inside of cells. Around tha finest Intracellular partleles there were yellow deposits haring tbs earns color as the asbestosis body. E^osure of one year bad permitted an lecucolation of many longer fibers about which the asbestosis-body coating dsveloped. Host of these were still short enough to be partially or entirely within phagocytic cells. By the 2Cth axrath and thereafter, they were comparatively numerous although still rare in caparison with the findings ir. the King's floats experiment. 35. Rats. Sevcrty--three white rat..* were expose tn ataospharic short-- fiber arrestor dust ibr yrrtc^f rr tc 32 norths. Sacrt- fiaiags during the first 10 aonths wo made Mmnthly and for the remainder of the experiment at less frequent intervals. 36. Hate and Type of Reaction. The dust cells until 8 aonths sere widely scattered and existed in foci only sporadi cally. Reaction w Halted to occasional slight thickenings of the septa about snail accumulations of dust calls. In a fear rata at 10 months, there was a sug-- fastion of early fibrosis but the change was so slight that it would probably be overlooked without the eluap of dost calls to attract attention to the area. Only 10 animals were exposed froa 12 to 32 aonths. In each of them the lungs showed minute patches of well-defined fibrosis distributed like that of asbestosia but without asbeetosia bodies. The lesions, visible only at a signification of 150 diameters or store, consisted of patches along alveolar ducts in which the walls of the air spaces were very thick, dm to seollen collages framework. Connective tissue and Foot-Sielacbowsld. silver preparations revealed complete loss of capil lary bed locally. Outside the collagen was a thin layer of epithelial cells. This did not resemble the 'adenomatoid" change characteristic of guinea pig asbsstosis. Bo pleurisy was present. Hoar the lesions the air spaces were filled with phagocytes containing gray to yellow particulate dust and a rare long naked asbestos fiber. Careful search failed to reveal even a suggestion of an asbestosis body... The tracheobronchial nodes showed compact focal collections of mono cytic cells at 12 aonths and, at 20 nonths, some diffuse thickening of the reticulum. In a few rate there was definite fibrosis along the margins of the node ard extending into the mediastinal areolar tissue. Coopered with the response to active dusts lilse quarts and chert the reaction to short-fiber asbestos was negligible. Results of chemical analyses made cn the stilt e rats ore given In Table 3 and ihe average vaXtas have bees tabulated is Taeie y for comparison -ill. a:.., values far rats Inhaling other dusts. The concentration of atmospheric particles to which the animal s were exposed was approximately the same for asbestos end quarts; for the gypsum-quarts mixture, it was about tales as high end for chert fire times as high. It will be noted that the percentages for asbestos are lover than those for quarts or chert but are similar to those for the g/paua-quarta mixture, in which atmospheric agglutination tended to reduce the aneunt of dust inhaled. It might be inferred that the total. quantity of asbestos dust inhaled w low or that it had been eliminated from or dissolved within the lungs. In the present stats of our knowledge evaluation of these hypotheses is not possible. 37. Cats. Twenty eats were used in this inhalation experiment with the short-fiber asbestos. Eighteen were kept in the dust rocs until death, the exposure period ranging from one month to nearly L-l/2 years, and two, after a dust exposure of 31*1/2 months, ware removed to normal air. One of these was sacrificed $ months, and the other 2li months, later. 38. Rate and Type of Reaction. The reaction was essentially that to an inert dust, even after more than ii years of expo sure. The tissue response in this species was confined to microscopic foci of fibrosis is the walls of groups of subplsural alveoli, rather than in tha peri bronchiolar areas* In one animal the change was extensive enough to be Tiraa.llzed cn gross inspection of the section. 39. I-5ar Changes. Only in the animal with the longest exposure did the - X-ray reveal definitely abnormal shadows. After 2?-3/c. mouths tie picture was negative; after 1x5 aentis a faint mottling could be deterged throughout both lumgs. At autopsy, 5 months later, there war only microscopic fibrosis in tha subplaurai tone plus heavy lynphocytic infiltration about snail bronchioles. IiC. Asbestosls S&ciac. Cn prolonged search a few yellow atypical asbestosis bodies, swath and without haustrations, were found in two arrival * ezsosed for sore than a rear. 111. Rabbits. Eight rabbits were ecpoecd to dust for periods extending from one to nora then fire yeare. The last was restored from the dust rooa and loft in normal air 6 months before being sacrificed. 112. Rate end Type of Roaction. There was never enough fibrosis to be de tected grossly and there was no chronic adhesive pleurisy. Microscopic evidence of alveolar wall thickening was first detected after about 3 ymars of exposure and was seen in all five animals examined thereafter. In one animal that died of paralysis after nearly four years of ex posure the reaction was extensive enough to be visible on gross inspection of tissue sections. The possibility of pulmonary infection in this animal could not be excluded. Hoearer, in another animal dying two years later the focal fibrosis was not nearly as obvious or as advanced, Areas of involvement, which were largely visualized because of phagocytic reaction within the air spaces, tended microscopically to become core fibrous with the passage of time but there waa never much encroachment upon the lumen of air spaces and the architecture of the lung was presred. 113. -Asbestosis Bodies. Asbestos bodies were not detected in rabbits that died early in the experiment but were seen in all an<-alt that had been chores to the cost for sore than three years. lii. Sua.iArr and Interprahation. The original purpose of the experiment nu to evaluate the chemical theory of the pathogenesis or as'oestosis. It mas felt that if the tissue reac tion to asbestos were chenical in origin as accelerated or accentuated response would result from exposure to finely-divided asbestos, as is the ease with quarts. This experiment, in which the reaction was slower and less extensive than with King's floats, indicates that the reaction probably is not primarily chenical in nature. _.. _ Of the four species exposed in this experiment only the guinea pig and rat reacted with characteristic peribronchiolar fibrosis. The cat reacted with atypical sub-pleural fibrosis and In the rabbit the fibrosis which occurred could not be positively attributed to the dust because of a strong possibility of pulmonary Infection. U5. Inhalation Zxperiaant with 100 Per Cent Ball-milad Asbestos Dust. In the inhalation experiment with short-fiber asbestos dust a snail quantity of usground asbestos was adzed with ground material in order to produce a suitable dust cloud. Hhen evidence of a dust reaction appeared in the guinea pigs during the experiment, it was not dear whether this was a tissue response to the small number of long fibers in the unground asbestos or was a delayed effect of the more abundant fine dust. Consequently, another inhalation experiment was started in which no unground material was used. -- U6. The dusting material cas the ground short-fiber asbestos used in the previous inhalation experiment but no inground material was mixed with it. To obtain a sufficient ascsvmr: of atmospheric dust, the design of the dusting apparatus was changed to an open type of hopper and fresh dust was added daily. Grrir^ to the tendency oi* the material te fom sphsrulan '!' ic'-i nrcrantar if the fibreus portion froa floating out of the hopper, the cicpci'oal of the dust -ran not entirely satisfactory and after 7 months of operation, the dart ing us.ciu.n3 ess rocccrcrtei to its original design. To present r7iilinr:i or o.v formation of spherules of asbestos, steel sire brushes ware attached tc the in side stirface of tha hopper and to the rotating candle. This arraegoEsst gave satisfactory results and was used for the remaining Z1 months of th/ ^spsriaent. i?* The cossssition of the ram material (sbort-fiber asbestos; and cf atia'spheric dust liberated froa the ball-milled product in the dusting ma chine .is given in Table 10. These values are based upon petrograpai: stud-- and X-ray diffraction analysis. The atmospheric saa^le vas collected rath an elec trostatic precipitator after sire brushes had been installed in the dusting machine. Previous to this, the ehrysotile content of the air-suspended material was undoubtedly Lees than the 15 per cent value given in Table 1C. In. an inter in report, it was stated that tha air-borne dust contained about 5 per cent of ehrysotile before the wire brushes were used and up to 8 per cent aftersards, but these values ware probably loir. Quantitative estimates on ball-silled' asbestos dust may be somewhat inaccurate because it is difficult to determine how much of a dust sa&ple is fibrous ehrysotile and henr much is nos-fibrous serpentine. UB. Bie dust concentration for the first 7 months of the experiment was about IX million particles per cubic foot of air. After the wire brushes had been installed, the dust counts were a little higher and the overall average for the first year, was 106 million. The average of courts for the second year was l3 million and for the third year lL5 aillion. Is. 7:.' sizs-ft'cc-ieucr z:. the crrccpsutz of atnoepheric dust collected c,hs omia-'. rutr. the electrostatic apparatus is reported in Telle 11- -"o espies rrcre tslv.u, cnce before the wire brushes mere installed and one after. It "ill hr nc-tc:.' r.hct cf tor the sirs breehes were in use a greater proportion of very* fine particles and also of longer fibers was released into the air. 50. Reaction in inlr-ils to Inhaled ICO ?er Cent Ball-Killed Asbestos Oust. Guinea piss, rats end .v.ra rvre used in the inhalation experiment with the ICO per sent ball-mil led asbestos dust. The results are sismrized in Table 12. 51. C-uisea Pigs. Tae experiment uas started with 100 guinea pigs. Is the dust exposure proceeded} there rare 39 accidental deaths j 32 of pneumonia in an epidemic. Iftar 23 sonthe of dusting the 16 surviving guinea pigs rere transferra-c to normal air. " 52. Rate and Type of laactioa. For the first year of exposure practically the only reaction to the dust was the pre sence of scattered phagocytes and an occasional minute aabestosis bedy. At 16 and 20 cocth3 no gross response vas visible on the tissue section but sicroseopieally peribronchiolar fod of inflonsatory cells could be seen. At 2b months there was still no change large enougu to be seen with a hand lens although microscopic examination revealed cellular accumulations about terminal bronchioles and nany* more asbe3tosi3 bodies, chiefly within cells. ' Chemical analTtes cf the lungs (Table 13) reveal that in spits of the United ti33ue roactisn ccrridsrcbla dust had been retained in the lung. 53. 7rogrc3gj.on. The lungs of animals arucscd for she full dusting period (28 norrtas; anc then living in ncm&l air for 2 months revealed the changes described above and also very slight peribronchiolar fibro sis. After 0 conths in norral air the findings were sAr~'t:-y but at 12 oonths 3 of Ia animals showed grcs3ly-visihls characteristic peribronchiolar fibrosis with adenomatoid change. Sli. Lyrroh gsde Involvement. The tracheobronchial nodes were essentially - nagativo until exposure had been continued for more than a year and a half. Anl tj.1s sacrificed at 12 months and 16 months revealed a fee minute collection* of phagocytes containing particles but prac tically no fibers large enough to be recognized as such. After 23 months of exposure many monocytes filled with yellow grannies were present, it 3C months there had been a slight increase in reticulum but no fibrosis. No further changes occurred in the nodes. Asbestosis bodies were not seen in the nodes of any of the guinea pigs. $$. isbestoais Bodies. UiTrrfx* asbestosis bodies were observed as early as 3 months aftsr .jhpoems began, but they did not be come numerous until 16 mouths had elapsed. The bodies were abort and practically an were intracellular, although at 20 months some wre long enough to project beyond the cell borders. It is important to cote that in the later months of exposure there was a dis tinct increase in the amber of long fibers Qip to 70 microns in length) in the lungs and that after exposure ceased characteristic long asbestosis bodies were seen. 56. frhlte Hats and l-!lce. In this wperiasnt \i0 rata were exposed for periods up to 20 ccmths and 21. mice for periods up to 12 -caths. "either srecirj irvclop-ed <ttsi a cuygcsticn of anbcsteai.s s~.d reacvior. -3 Limited to uhagocvtosis or inhaled particles '07 wiceiy-scatterad dust ceils Tziich retained free In. air spaces or were transported to the tracheobronchial lynch nodes, ib asbeswocis bodies cuts found in the rats but in the nice thsrc were a 7ery few stall non-haustrated foras within phagocytes. In 21 souse lung3 sectioned there were 3 instances of palaonsry adenoma 37. Stgnsarr and Interpretation. The tissue reactions observed in this enporiment ware orjch loss sresneire and slcwer in development than in the previous investigation rith short-fibsr asbestos. Since presumably there were fsvar fibers longer than 3 microns in the notarial used in this erparia-nt, the results tend to confirm the intei-pretation made in Section U2i of the short-fiber ezperisent that the reaction probably is not primarily cheslcal in nature. The finding of long asbestosis bodies in aniiwi* inhnl.lng the bsll-aiilsd material is an example of the difficulty of completely eliminating long fibers from an asbestos preparation. In regard to progression of reaction after renoval from dust, -riich-was ob served in this experiment but sot in the others, the following interpretation 1s offered: 'Chen the reaction is well-developed at the termination of exposure.tbe contraction of the fibrous tissue would obscure any possible progression. In this experiment, however, since only the earliest stage of reaction was present at the time of removal from dost, its subsequent progress was apparent. It should be noted that the degree of progression was so slight that it can have little, if any, practical significance. 58. Inhalmtion Exoorimant with Long-fiber isbastos Dost. After ar-Lzzls inhaling short-fiber asbestos dust fen ..-<= tcum a 7ir tJ^d failed to develop significant reactien, the hypothesis that aabestosls is pro duced by the nschaaical irritation of Ions fibers was given added support. Since the ILir.g's floats asbestos urea in the first inhalation experiment haw a rather low content of fibrous ciuysctile and contained considerable serpentine and other impurities, it was decided to conduct a new inhalation experiment with a purer fora of chrysotila which would be richer in long fibers. - 5?. The dusting arterial employed in this investigation was obtained from the Uanvllle plant of the Johns-Danville Corporation. Samples of se veral varieties of asbestos dust were first submitted to the Saranac laboratory for examination and one kind, identified as lot D, which was low in magnetite and chromite and had a fibrous content estimated to be about 75 per cent, was selected as most suitable. Steel wire brushes were fastened to the inside sur face of the hopper and to the rotating paddle in order to open up the bundles of asbestos and liberate more fibers into the atmosphere. 6C. The composition of the long-fiber asbestos used in this experiment, is indicated by the chemical and petrographic analyses given in Table lit. It appears that this material was a such purer fora of asbestos than the shortfiber dust used in other expert.cants. This is borne out by- comparing the approx imate analyses of the long-fiber and short-fiber ldust in Table 15. 61. The dust concentration as revealed by inpiager'samples taken inside the animal cages was ouch lower than the concentration for the experiments with short-fiber-or ball-milled cost. For the first year of the experiment with long-fiber asbestos the averse of the light field counts waa 32 million; for the second Tear, uo rillior for tho third Tear, 3? million; and for the fourth year, U3 mUULion. ixaminaticn of swplea with dark field illualn- ation disclosed that many fine particles less then one rdcrcn in size accompanied the larger particles and dark fiald counts */sre, on tiia arerage, about 5 or 6 tines larger than the light field counts. 62. The sino-frequency of atmospheric sangles cf the long-fiber asbestos duet and of the ball-silled dust is shewn in Table 16. Both saeples sere collected with the electrostatic precipitator. It will be noted that there was far more fibrous material in the long-fiber cunt. 63. Reaction in Animals to Inhaled long-Tiber isoestos Dust. Guinea pigs, eats, rats and sice ware employed in the initiation experiment with ~ long-fiber asbestos. Results of the experiment, summarized In Table 17, are described in greater detail below. 6ii. Guinea Pigs. The experiment was started with ICO guinea pigs, ifter exposure had been carried on for a Tear, a severe epidemic of pneumonia arose in the dust room and about one-third of the animals died or were killed. To replace then, 38 more guinea pigs were added to the surviving group in the dust roan. 65. Bate and Type of Reaction. Histological examination revealed grossly visible lesions in the lungs after 6 months of exposure to dust, consisting of cellular infiltration about the terminal bronchioles. It 12 months, there were adenomatoid changes in the air spaces and by ti* 16th month & definite fibrosis was present in these areas in half the-- animals. The fibrous lesion could be seen macroscopically at 20 months. Tram this ti^ on the reaction inrressed in extent ar.d is tho amount of collagen.and 'sj tils 3iith month, it had fanned out into the paranchTtsc.. Tda lesions were rather sharply localized and the extensions dboa different bronchioles shooed no tendency to fuso, even in animals exposed Tor the nnxinsa period (3 years), although the intra-oulioonary reaction sometimes reactrd the pleura, there ms no involvement of that mcrbrane. J-io emphysema was vis.'.b.'.s at any point. Some thickening of tha larger bronchi ritii a chronic infU-naatory infiltration was re vealed, but it probably was no core than traulci be produced by t similar exposure to any dust. For the first 3 conths the phagocytes corsicteti of ?.or.:c.rtss or very small giant cells; Later, giant cell formation res mere preninsut. After 16 sooths the giant calls were large, fUlod with yeMowish-hrean pigment and sometimes vacuolated. An occasional aninal showed an Admixture of polymorpho nuclear leukocytes and, in guinea pigs exposed fer r. eou.'.'.d.srabla period, eosinophiles. The reaction was at first entirely cellular but by 15 souths fi brous tissue formation was definite. However, it never attained a stags of hyalinization suggestive of silicosis. 1 aocerate individual variation occurred among the exposed z. lulls, both in the rate of developing lesions and in the stage of development atta.ced at the end of exposure. Analyses of the longs (Table IS) disclosed that although the tissu. response wae amch greater in these guinea pigs than in those exposed to either she: t--fiber or bail-milled asbestos, the amount of mineral matter in the lung ash tas Lee. 66. Progression. In guinea pigs exposed to the dust for 20 months and tinu removed to neraal air, there was a narted tendency for cellular inflarusatory reaction to clear. This effect, accompanied by contraction of the fibrous tissue, resulted in a diminishing sirs of the focal lesions. ':z-s of thase animals, tills! ot- various periods up to iii months after exposure. . lc-~sr^. so lar^s so thoss in tho poup sacrificed at the end of the 20-aonth exposure period or those in animals which remained in the dust room ' - 2" *:.' --..... ---. in four of the six remaining guinea pigs were so small that taoy- were visible only with a hard lens. Reaction in the group eicpoeed for 27 months and then transferred to a normal atmosphere was quite similar to the response in the 20-rcnth exposure animals cmtiered above. However, small foci were always visible on gross inspection of sections of all guinea pigrs of the 27-<acnth series bat in no instance was there evidence of srtension of the reaction. 67. Lynch Node Involvement. Reaction in the tracheobronchial lymph nodes was first visible at the third month of ex posure. it the 6th month patches of cellular connective tissue began to appear in the medulla and by the llxth month most of the node had been replaced by cellular connective tissue. This picture, which resembled that in early silicosis, persisted to the end of the experiment. Some animals, as a variant, showed heavy sheets of diffose monocytes and large active giant cells but there was never any necrosis or hyaline formation. The spindle-shaped new cells were yellowish in color from fine pigment granules that stained for iron. So fibers or asbestosis bodies were seen. 68. Asbestosis Bodies. Although asbestosis bodies were seen as early as one month after exposure began, they were rare and hard to find. At ? aonths aore were visible, -chiefly coiled inside giant cells, and at 3 months many bodies were free in connective tissue. They became fairly abur-iant as sepssure progressed although in some later animals the asbestosis bodies were only moderately tmeniis.  - 6?. Cats. Four cate inhaled the lor.q-fiiac* asbestos dust for periods of Hu, J-' aEu tu; cc.ithj, respectively, and were immediately sacrificed. Two ether cats, after being exposed to duet for 18 Booths, lived in a normal atmosphere for an additional Cii months. 70. Rate and Type of Reaction. Exposure for Hi months vas sufficient to produce cellular accumulations of phagocytes around terminal bronchioles and peripheral arterioles together with compact col lections of similar cells in the tracheobronchial lymph nodes. At that tism there were no typical as'oestosis bodies, but smooth pointed yellow fibers were seen very rarely* ffith continued exposure, up to L2 months, reaction in the locations noted progressed to the formation of cellular connective tissue which made well-defined 3heaths about the respiratory bronchioles and arterioles, marked lymphoid hyper plasia and lymphoid infiltration of brouchiolar walls. The bronchiolar epith elium was low and flattened, giving the tubes a smooth contour. Typical asbestosis bodies were not forasd although there was an occasional yellow, smooth, pointed fiber. No pleurisy was jresent. The reaction was similar in location to that in the guinea pigs, but fibrosis was much slower in development end had not reached the same degree of maturity. (X. X-Kay Changes. Roentgenograms of three eats were made after exposure periods of 25, 33 and L2 months, but tissue changes were not dense enough to be seen on an X-rgy file. 72. Rats. Although 20 rats were placed in the dust room, many died from pneu monia and were not suitable for study. Five animals, of which-cue was exposed for 1? months and four for 2$ months, ware frae fcoa pulmonary infec tion and offer a basis for conclusions. 73. Rata and Tyne of Reaction. All four sacrificed at 2$ norths showeu a /.eu pe.ibrouuhioiar fibrosis In the 15-month animal, reaction was Just beginning - Asbestosis bodies were practically absent at both 15 and 2 5 months although two small smooth bodies tt.s--e found in the 19-month animal after a long search. Thus, these exhibited fibrosis without asbestosis bodies. Ta. Hice. Out cf 20 white nice used in this experiment, 11 lived a year er acre in dust and died or were killed without showing an appre ciable degree of pulmonary Infection. 75. Rate and Type of Reaction. Reaction was limited to phagocytosis by mononuclear cells. Usually these were wide ly scattered through the air spaces; a lisd.ted number were grouped about the ter minal bronchioles producing some thickening of their walls. There was no sug gestion of fibrosis. The striking feature of the experiment was that 9 out of the 11 mice (92 per cent) exposed to dust for a year or more showed pulmonary tumors, usually adenomatous in type*. These lesions did not contain dust or asbestosis bodies. _ Numerous asbestosis bodies wore observed in killed late in the experi ment. Thus, these animals exhibited asbestosis bodies without fibrosis. 76. Sunnary and Interpretation. ~ The purpose of this experiment was to evaluate the importance of long fibers in the tissue response to inhaled asbestos. The results Indicate strongly that long fibers are chiefly responsible for the reaction. Thus, in guinea pigs reaction developed earlier ar.d b*cane-more extensive than in previous expert cents in spite of a smaller concsr.tr at ion of atmospheres dust and a lower mineral con tent in the lungs. Furthertcrs. z. typical paribrsnuhiolar ti'urcsis ves trtr::--; in cats although in a previous experiment with short-fiber dust it did not `t. thi* species. The cause of the cellrtar fibrosis in the lymph nodes of the guinea pigs is ncu clear. It ill cc-.-cr 1.: ccher inhalation :perinsnts vitr. ascestcs. Lcnru. iNJECTiow stpsrihb<ts ' In order to determine to what extent the various fibrous minerals possess the capacity to produce tissue damage, numerous injection experiments were per formed. In these experiments guinea pigs and rabbits were used and the mineral dust was injected by the Intratracheal, intraperitoneal and intravenous tech niques. For the purpose of simplification the findings in each series of testa have been condensed and reported in tables, to which reference will be* made later. ^ 70. Experiments Using Intratracheal Technique. -- Since the asbestos alnarals do not cause a typical advanced fibrosis in extra-pi1 nonary tissue, the Intratracheal technique is the perferred way of in troducing fibrous dust into the experimental animal. In this method, the dust suspension is injected by means of a special needle or catheter deep into the trachea, Cram which it flows into the lungs. 79. Comparison of Fibrous and Son-Fibrous Dusts. To demonstrate that the ability of asbestos to produce fibrosis resides in its fibrous character, the series of injection experi aents reported in Table 19 were ->*r formed. The tests were made with unheated long-fiber chrysotila and 'rith that had been ignited to deszrzr/ its flexible structure or call-Tilled t r-:rj?c ''~i Icr-^-'r. of fib?r : r ' si crons end 'c.- -. same tin.-*. crr.trcl tos'i.r. ,r.\r * 2 rii'i rarreutine, widen nas -he same chemical composition as chry?otile "vi is rxr.-Jibreu::- A review of the _7.niii.tro reverie c:..y t1..-. Fibers subjected to igniricn or shortens-:r. by hail-rilling iuid lest their capacity to cause serious tissue darace. Ignition or ;e teed intortaut chs-5-3 he chry- sotila fibers, among them 'ceing loss ?f ---vt-ir. an riteracion frees a flexible to a brittle structure and possibly ether chansji:. 30. Comparison of 7arious Lopg-Fibsr Ovists. 5one rery interesting findings * ere disclosed 'ey the results of the experiments included in Table 2C. First, all the long-fibei- asbestos minerals tested, with the exception of anthephyHite, produced a typical fibrosis. It is not entirely clear why anthophyllite beiiavad differently from tha other asbestos minerals. Unfortunately, 5 of 3 animals died of pneumonia within the first two weeks of the experiment and the remaining animals were sacrificed at 1, 3 and 12 months; thus observations were not mads at the optima periods of 2 and It months. Second, with the mineral brucite, which is not a silicate but is a fibrous form of magnesium hydroxide, a characteristic fibrosis lila that of the asbestos minerals was obtained. Since ths brucite used contained only C.?0 per cent silica (as an impurity), it is obrious that a siliceous component is not an essential factor In the development of asbestoeis. Third, no fibrosis resulted froa ths injection of glass wool fibers, even ~' e though glass wool reseeibles asbestos in some ways. There are fundamental differ ences, however. A glass wool fiber 3 microns in diameter is a solid rod and, in short lengths, is fairly rigid, while an asbestos fiber of the same diameter is a bundle of extremely fine filaments wnich impart to the fiber a high degree cf flex' ibility. It would seem -hit thi :e arc the assneoateo i leans ^ - r- are important factors governing the capacity of a mineral to oroduce peribronchiolar fibrosis. 81. Comparison of Long-Fiber and Short-Fiber Posts. With quartz dust it has been demonstrated that the smaller the particles, the more intense is the tissue reaction, and that there is little reaction to particles larger than 3 microns In diameter. In the case of asbestos, however, the reverse is true and apparently only long fibers have aiy specific effect. This is confirmed by the data of Table 21, in which a series of tests with fibrous minerals is reported. When the Injected dust consisted of fibers 20 to 50 microns long, all the minerals tested (except anthophyllite, as noted in Section 80) produced a fibrosis j whan the material was prepared by first grinding the fibrous dust until tbs length of fibers was reduced to 20 microns and lees (or, in some eases, 3 microns and last), none of the Injected mineral dusts canned fibrosis. 82. Sroeriments Being Intravenous Tect"**q*- The experiments, described is Table 22, in which the intravenous method of injection was employed, shoe that the asbestos minerals are far different from quarts in thalr action on tissue. It has bees repeatedly demonstrated that intra venous injection of quarts particles 3 aderons and lees la diameter will cause a typical tissue reaction with the development of fibrosis in extrapulmonary sites, such as the liver and spleen. Asbestos minerals, however, on intravenous Injection generally produce ocly an Inert type of reaction, as is revealed by the results given in the table. The reason for the early deaths in the experiment with ehryio- lile particles is not clear; it say hare been caused by silicic add liberated by the finely-ground mineral. ' 33. Experiments Pslag Intraperltoneal Technique. r& results of Injection experiments with the iatrapentoneal technique are givsa In Table 23. It will be noted that the long-fiber dust* produced a fibrous reaction while dusts composed of particles 3 microns and lass in siae caused only an inert type of response. These experiments indicate also that the fibrosis Initiated by the irritation of asbestos flbsrs is not rsatrietad to tha lungs, as was formerly assumed, but can be produced in the peritoneum as well. LXX237. OTHER EtPEXIIgRTS WITH ASBESTOS HISEFii-s 1 number of additional experiments were conducted to throw nore light os specific phases of the asbestosis problaa. 3?. Protective Action of Alnalnna Co"tTM,t*4*- ___ Intratracheal injection of a suspension of long-fiber ehrysotile to which colloidal alnnrtnnw hydrmids had been added revealed that the addition of the allied mm wwipcMaai did not prevent the tissue irritation produced by ctsysotils. If anything, the acute Inflen--tory response to the Injected fibrous mineral was accelarated. Otoe month after the last injection of tha dust suspension the bronchiolitis was hemming fibrous. 86. Formation of Asbestosis Bodies. The iron la the costing of the asbestosis body appears to be derived from blood or tissue elements sad not, as has been suggested, from the mineral fiber. Following subcutaneous injection of two kinds of chrysotile into the groin of guinea pigs - one kind containing 2 per cent and the other 0.2 per cent ?e20ithe asbestosis bodies were equally numerous at both sites of injection. An attempt to produce asbestoeis bodies in guinea^pigs by implantation of three silk bags oonteuung nonras cnrysotile ess pnsuccassful. One bag planted e subcutaneously in the abdominal nil disappeared; the other tee bags, placed in the peritoneal cavity, produced a little foreign body reaction but no asbeatoala bodies in a year. Intratracheal injection Into guinea pigs of asbestosia bodies recovered from human lung tissue failed to prodace the typical tissue reaction to asbestos fibers. The injected material mas obtained by digesting with sodium hypochlorite solution lung tissue removed at autopsy from an asbestos mortar. The asbestosia bodies could be seen in the guinea pigs for at least a year after injection. This experiment shoes that the asbestosia body has a rather resistant coating * ehlch la not destroyed by moderate hypochlorite treatment and may be maintained In riro far a year or longer. LXXXTXX. THEORY CDF* IRRITANT ACTION OP ASBESTOS UIHZRAI3 Two hypotheses have been proposed to explain the tissue irritation and reac tion cammed by asbestos fibers* the chemical and the merhenlnl. In the chemical theory, which Is based upon experience with quarts, it is assumed that tha asbeetoe rrfnarnij dissolve la the bodbr fluids and that in this process their bases are assy to leave silica is a fora eapable of lrtitating tissues. Aceordisq to this hypothesis, aabeetosls would be migely an Indirect dUeMii. Several facta mata the chemical theory untenable t (1} intratracheal injections of brueite fibers, which had a silica content of only 0.90 per cent, caused a typical fibrosis like that produced by the asbestos minerals; (2) ftree silica particles Increase in potency as tbs partirle sire becomes less, but asbestos fibers shorter than about 10 to 20 .uiermc 3."- rslativel hydroxide neutralises the irritatin/; effect of quarts but not of asbestos; (2a) serpen--Ljx bcc tr.c ;^z9 ccnp^siticr c? Iczg -fiber chrysotila bat it does not produce tha sane kind of tissue reaction; (5) there Is a vide range in the chemical composition of the nioerals uhicc. do cause asbestosis (see Table 2k). In rimr of this evidence it seems acre likoly that asbestoeia Is caused by an unusual mechanical irritation from Lone asbestos fibers. Probably this irritation is related to the peculiar filisested structure of the fiber end fati the associated flexibility, which are poasessed by no other foreign body2 Ter * example, ignition of chrysotile fibers changed their structure and made them inert while the same fibers, before being heated, would produce fibrosis (sea Table 19) Further support for the theoz7 of mechanical irritation is that asbestosis occurs in an organ of high mobility - the lung - and that a fibrous reaction esa be produced by injection of asbestos fibers into the peritoneum, where there is also a degree of mobility, bat not in other estrapalmonaiy organs. mimi. COEPLICATIOSS The experimental investigation with asbestoa minerals was concerned primarily with the effect of the dust on normal tissue but some attention was given to other phases, such as susceptibility to infection and occurrence of malignancy. 89* Infection. The only experiment in which the effect of inhaler! asbestos dust os a pul monary infection was studied was tha first inhalation experiment, carried on vith Sing's floats* dust. It is, perhaps, unfortunate that infection studies were not made in the other inhalation experiments also. SO. Susceptibility* to Tuberculous Infection, Tb- of a uer- culcnxa process Initiated at the beginning of exposure to asbestos dust, and also of an infection superimposed upon an established asbestosls, was described in Sections 1? sad 20 of **** re port. It will bs noted that asbestos, when classified according to the effect of a dust on tuberculous infection, would be placed below sa active dost quarts but above inert dusts, such as caldte and gypsuuu In infected with attenuated tubercle bacilli, quarts will cause the infections process to progress until the animal dies of tuberculosis. Inert dnsta will have no affect on the infection and the lesions will usually heal and the **<--<. disappear. Asbestos dost is in a different category. *ben the fibrous dost vas being in haled during the evolution of the infection, there was a spreading of the tubercolons process for a tlaa but usually tbs stimulus far caatimzsd proliferation of the tubercle bacilli was not sustained, the progression was arrested and followed. At guinea pigs infected with attenuated tubercle fallowing the completion of nearly three yean of exposure to asbestos dust, progressive disease did not develop. The only modification of the infection vas one of localisation, a for being retained la the fibrous terminal bronchioles and farming tubercles there in addition to the usual fod beneath tba pleura. Such tubercles healed in a few months and there was nothing to suggest ary lnflnancs on tbs course of the disease. 91. Susceptibility to Bon-Tuberculous Infection. There was no pointed x- perlasat concerning tbs effect of inhaled asbestos dust on non-tuberculous infection. Intercut'rent pnen- :nia among exposed to asbestos dust was rather common, the frequency in guinea pigs exposed in the four inhalation experiments ranging from 16 to jp per cent. This incidents* evidence suggests the possibility" of an effect of duso on non--uaercoicua infection. Nevertheless, since such epidemics are not uneassaon in inhalation experiments -nith other dusts and eves In the colcry of normal animals, it is felt that the inhalation of asbestos dust does net exert a significant effect on the susceptibility to noo-tuberculous pulmonary infection. 92. Beoclaaau Vb specific experiment -was conducted to detersine whe'.her the inhalation of asbestos favors the development of neoplastic disease but certain observations on this subject sere recorded in the outline of the propoeed eomgraph on asbeatosis submitted by the late Dr. L. 0. Gardner in February 19it3. In it he called atten tion to the high Incidence of lung cancer among alee inhaling lc5g-flber aabestos. In his experimental notes, however, he referred to theee lesions us adenomas. There is an important distinction between adenoma and cancer rhich should be made dear. A cancer is a tumor, or neoplasm, capable of local iivvision and de struction of tissue, which can distribute cells through the lymphatics or blood stream to produce Isolated foci, Craa which new tenors develop. Thii phenomenon of dissemination is known as metastasis and so? tumor ifclch exhibits it is a malignant growth, of #iich cancer is one type. An tdennre, on the o .her bend, is a so-called benign or aon-oallgnaat tumor (neoplasm) which may.or mrr not be capable of local invasion but which does sot aatastaaiae. In order to clarify the exact nature of these lesions the path 'logical mater ial is being carefully examined. Since it is felt desirable to her, the benefit of Sector 7omald*s judgswit, a review of the data on this cbjer' post poned until after his return from Europe. Hather than delay the eat'.*9 report, further discuseion will be reserved for a euuplssent to be issued lat 1. Ccing to the tas* crnunt cf data include.! in ibis report it seess aost convenient to state the ccnciusions derived dree the investigation and. whan necessary, folic? each one -sith a briaf rasuxt of the evidence. A. farises form .of asbestos fibers produce & peribronchiolar fibrosis cf the lungs of guiiea. pigr, rats, cats and rabbits bet not of mica and dogs. -- Both inhalation and injection axperiaents provide ample support for this conclusion, figures 5 and 6 she* the reaction to ten different IdLncs of asbestos aineral. 3. The node of action appears to bo primarily mechanical rather than chemical in nature. The evidence is given Is section LZTZVITt Figures 1, 2, 3, li and 7 illustrate the important points. The fibrous filasented structure of asbestos appears to play an essential part in the irritating action, since the solid fibers of glass wool do not > produce fibrosis (see Figure 6). C. Short asbestos fibers do cot produce fibrosis. The conclusion la implied is the evidence cautioned in para graph B above. Ssperisents which further support this fiadlnt are reported in Tables 21 and 23. 0. Typical fibrosis eon be produced by an ataospheric suspension of asbestos dust containing only an ertreoely snail proportion of long fibers. ... In the IrlcsleticR experiment with ICO per cent bail-oilled asbestos dust a typical, though delayed, fibrosis was --uO-- .**.:** <3-.*v'`'lvN'tT,,r.-. Vui-:z 1.' ' i-.-.'.o rrv. a-. drsal'Lng f.ctica <;.->iT-dasJ sx,7 i rili -i- -ul azscrA. of it vr -rrri furthar i-troato rrr-i-r ci this r'-iasc of is.^rtocis -.5 cir.si"icrrr'. IL:c feruitioa of car-estosis todies snc.r to rrurr-^err; a c zz.tL-.~ the iicars er.d -T.s-'iT.ts Lz Anars or tij* nbilitr to rro.isco i'icroni ,T:itrat.i'ac*x?l ir^rctisn o? ssiiaatoaia halier r-JLI :c- 'x- (/ prcd'ica t.lie tjpical tissua rsection (saa section. . TI:? cassation or prosressxra i-sactica to lrhalad azbosbos dvai serrn after rscrosiaro toraisatce osr *ic du to tbs roiTiti.oo of asbestcoio bedims. -42-