Document mq6wbxL093ZaeGX8KbNbo21vZ

k ;ii s: a:u':i ji. nui- htjj C::cc^;or.:c Lo^cnce of 2zt Skin, Lungs, r.d 3or.cs to V A4*Jr" * *wW ILLEGIBLE P. L. Viola, A. B-\;otti, and A. Caputo Rcfiiu cU'iu Institute for Cancer Research. Rome, fitly DOCUMENT SUMMARY ., Rats (Ar/JRE Wisiar strain) exposed for 12 months, to vapors of vinyl chloride developed tumors of the skin, lungs, and bones. Tie cutaneous tumors, which always appeared in the area ir. which submaxillary and parotid glands are located, have been histologically recognized as epidermoid carcinomas, papillomas, and mucoepidermoid carcinomas. The morphological characteristics of lung tumors, which occurred in a lower percentage, were mainly of the adenocarcinoma type, with the exception of a single epidermoid tumor originating from the epithelial covering cells. In a minor number of rats, a large proliferation of cartilaginous tissue diagnosed as osteochondroma developed in the metacarpal and metatarsal regions of the four limbs. INTRODUCTION The oncogenic properties of some chemical organic compounds used for the preparation of "plastics" haw been widely investigated, anu their limits and effectiveness have been well established. Detailed information on this subject may be found in the literature (5, 6, 8); however, all information and references are exclusively related to highly polymerized compounds of roughly the same size as used in various industries. Oncogenic polymers produce sarcomatous tumors, with the exception of polyuretluns which, as reported by Hucper (6), also induce adenocarcinomas. Recently, it has been demonstrated that sarcomatous tumors (1-4, 7) are transplantable, and it has been suggested that they may originate from the tissues of the capsule that gradually covers the plastic film. Thus, Brand et al (2-4) have observed cremaligr.ant areas made of poorly differentiated fibroblasts which were firmly attached to the plastic film up to the moment of malignant transformation. This investigation demonstrates that even the monomer vinyl chloride possesses oncogenic properties when used with an appropriate model different from the models previously reported by several authors (9-12). The carcinogenic response 1A prelinklury report of the remits reported here wa* civen at the Tenth International Cancer Congress, Houston, Texas, May 22-24, 11970. " Reodvey July 31,1970; accepted January 12,1971. to vinyl chloride follows a singular pattern for the different tissues and organs of the rat. MATERIALS AND METHODS The experiments were performed with vinyl chloride (CHjsCHCl, the monohalogenate derivative of'ethylene) of a commercial grade (99% purity) assumed to contain insignificant amounts of various noncarcinogenic contaminants. Three-month-old Wistar (Ar/IRE) male albino rats (about 1 SO g body weight) were exposed to vmyi chloride vapors for 4 hr a day, S days a week, for 12 months. Tire animals were kept in metal or plastic, air-tight cages in which a constant flow of air, containing 3% v/v (equal to 30,000 ppm) of vinyl chloride, was introduced. Twenty-five rats of the same strait, were the control group. At the end of the treatment, the surviving animals were killed at 20-day intervals, and the most important tissues and organs were examined histologically by standard methods. During the period of exposure, the animals were slightly soporific; however, the first few months of treatment were well tolerated and no changes in crowd) or behavior were noticed. After 10 months of treatment, some animals began to show a hard mass in ti e pataauocubr region which became progressively larger until t. `cached the size of a walnut or slightly larger, tn most cases, the swelling was unilateral; it was bilateral in only a few animals. After 1 or 2 months, the growing masses became ulceratcc and discharged necrotic debris, while a certain amount of tumorous tissue began to form on their surfaces. In addition we observed that file masses were an integral part of ,.u. "trau*.ricular region and could not be distinguished from r... .vui tissues. Caseous necrotic zones were found. Pleura and pericardium often showed diffuse inflammation of a fibrous nature and. in many cases, the lungs were covered with a number of white formations at large as grains of rice or even larger and horde: than the lungs themselves. In 2 eases, the lungs were hemorrhagic with milky, thick fluid in the pleural cavities. Yr.e liver was sometimes increased in size and very fragile. Tne animals were subjected to X-ray analyses at different interval! from the beginning of treatment in order to control the state of the skeletal bones. All the animals that inhaled vinyl chloride showed .. senes of parenchymal lesions. Among these, most prom'm&s were the disappearance of granular and Purltinje cells, degenerat-oa 516 CANCER RESEARCH VOL- 31 it iff to 00 1X3 -J -4 R&S 028278 Oncogenic Activity of Vin ci Chloride of the i-orv\v!l,];*.i. were chronic hepatitis, interstitial W.iny sulvniricul.ir growths occurred in some rat-.. Ti/C pneumonic.. nr.u :noJe:.::c swelling of tt;o kidney parenchyma, histological picture showed papilla.' epiiecl.a! prohtcratrto, often assuming the pattern of tubuloncphrosis. with progressive increase in the thickness of the epidermis 4^SULTS t T!:e rates of survive! a::J the main findings are summarized in Table I. Almost all the animals developed tumors of the s'.:r. a"J litres. Very few animals developed bone tumors; in :ite>e eases, the tumors were localized in the metacarpal and metatarsal bones of all A extremities: Skin tumors were by far the most frequent, amounting to 65 or 70%. Skin Tumors. Trie tumor that developed most frequently in the patuufieula: region was the epidermoid carcinoma, but wc aiso observed papillomas and rarely mucoepidermoid carcinomas. In ail cases, the neoplasms were of epithelial nature. The 3 patterns noticed cannot be compared to various ktstotypes bu: rather to a transition of one type into the other or, and which is more likely, to different stages of the same proliferative type. The papillary warts of exophytic type had a fibrous vascuio.* stroma and various degrees of infi.i:r.iuau.ry lymphocytic infiltration, marked liypcrkcratosit, parakeratosis, acanthomas, a:.J areas of individual dyskeratosis or pearl-like horr.y formations. The epithelial cells of the penetrating columns (fig. 2) were irregularly arranged and were frequently accompanied by an inflammatory infiltration of the dermis. The typical features of the Malpighian iaycr and of the stratum eorneum of the prickle cells and of germinal layers were easily recognized (Fig. 3). The horny layer was composed largely of cell nests, which appeared at certain points within the epithelial masses and assumed the well-known appearance of "homy pearls" (r:g. 3, arrows); they were made of flattened and compressed prickle cells without nuclei and were located around a central core of keratin. Table l Oncocnic effects ofinhaled vinyl chloride as a function of time The animals were exposed to vinyl chloride vapors for 4 hr a day, 5 days a week, foe a total of 12 months, in a constant flow of air containing 3% v/v vinyl chloride. Rat 1 X3 4 .s 6 7 8 14 Id. 17 21 22 23 24 25 2d Survival rate 300 280--3<X>\ 310 333 337 347 347 354 359 -- 380 380 ?5d 380 380 380 Tumors Skin Lunps Hones MuawpitWrmoul curfiiumiu Adcnoacanthoroa Epidermoid No tumor carcinoma, kera tinizing type Epidermoid Adenocarcinoma carcinoma, kera tinizing typo Papilloma, keratotic No tumor type Epidermoid No tumor carcinoma Epidermoid No tumor carcinoma Mucoepidermoid No tumor carcinoma Epidermoid No tumor carcinoma Epidermoid Adenocarcinoma carcinoma Epidermoid t No tumor carcinoma Epidermoid Adenocarcinoma carcinoma Epidermoid No tumor carcinoma Epidermoid Mucus-producing carcinoma adenocarcinoma (alveolar cell carcinoma?) Epidermoid No tumor carcinoma Epidermoid carcinoma Squamous cell carcinoma Osteochondroma No tumor No tumor Osteochondroma Osteochondroma No tumor Osteochondroma No tumor No tumor No tumor Osteochondroma No tumor No tumor No tumor No tumor MAY 1971 ILLEGIBiLE DOCUMENT 517 R8tS 028279 \v,,s r.r.'g.r.ai and often parakeratosis wax cartilaginous zones possessing regressive features, s.-.s.h ,,t obscrx cd areas of ri;e tumor in which tlte cells were fibrosis and hyalirtosis. The perichondrium appeared often ji a loosely ..ggtee.'.ied and unde,-going an individual rather than a compressed and structurally altered fiiir >us tn.s.ie. rolleetive ;.v of kcrat.ni/a: ton (Fig. *1). In dyskeratotic areas, In some cases, the tumor growth Was related to the stare o; few cp;;hc!:.:i tended to form peails. The tumor seldom the er.Joehondr.il ossification occurring hcluw rite "e;..;-.I;ystal showed an undifferentiated growth with cellular plate.*' O.vurication was irregular, so that ovwvtre tr.:hfce.;i.e plcomornitism and several mltoiie l\emes (Fig- 5). A tew varied greatly in thickness ami contours, hi other caves, foci uf tumors .showed little nests of isolated pale cells (Figs. 6 and 7) active cellular proliferation, cartilaginous areas, chor.drutd of 3 types: mucin-producing cells (criminating tram the duet developmental nests, and calcified bones occurred in she epithelium of sweat glands or salivary glands); squamous cells; deeper portions of the trabeculae (Fig. 18). and Intermediate cells with minor tendencies towards The new formation appeared to be an osteochondroma anti differentiation, . ' consisted essentially, of a bony protuberance capped by Respiratory Tract. These tumors, although occurring rarely, cartilage and a fibrous layer, which represented the were mainly* a Jcnocarctnomatous. Only in 1 rat did we observe perichondrium. art epidermoid tumor originating from the cpithclium-eovering The fibrous layer was continuous with the periosteum of cells. Sometimes, the tumors were seen in their early the adjacent cortical bone and extended inward to form septa development and consisted predominantly of cubic or separating and enclosing lobules of cartilage. columnar cells arranged as regular or irregular tubular and Control Animals. The control rats were kept under the same papillary elements (Fig. 8) and supported by poorly developed conditions as the experimental rats and at the appropriate time fibrous stroma (Fig. 9). were subjected to a constant flow of air'without vinyl In other eases, glandular structures were often imperfectly chloride. None of these animals developed tumors or the ty; formed and appeared as sheet-like proliferations cf of parenchymal lesions developed by the rats that iahal undifferentiated or pleomorphic character. Tire cells showed a vinyl chloride. In a very few of the control rats, there was tendency' to extend into the pulmonary parenchyma, tints swelling of the liver and kidneys. simulating the microscopic features of the so-called alveolar cell carcinoma (Fig. 10). Sometimes, this was the prevailing pattern. Tne pulmonary air sacs were limited by 1 or "more DISCUSSION layers of cubic, columnar, or polyhedral cells with abundant cytoplasm that was faintly eosinophilic (Fig. 11); frequently, As in many carcinogenesis studies in which multiple turnon adcnopapillary excrescences spreading into the alveolar spaces jrcere present. In some areas, loci of cellular polymorphism with arise in different tissues and organs, the data must be evaluated in terms of a more prompt positive response to be obtained with (lie ideal concentration of the carcinogenic compound. hyperchrontatie nuclei were noticed: mucin was produced in The cutaneous system is the most susceptible to the oncogen* varying amounts and sccretcJ into the lumen of the tubules effects of vinyl chloride. and acini. There were small pools of mucin in which signet ring Our experimental data do not explain why the cutaneous cells occurred, eithc: alone or in small clusters (Fig. 12). The tumors developed in the same site. ic.. the region including alveolar walls were often quite thick and, at times, showed an tiie area in which the submaxillary and parotid gland* are inflammatory infiltration. located. It is possible that the salivary glands may by involved As mentioned before, a single tumor showed squamous in the concentration or excretion of vinyl chloride or some of structures and appeared to have been formed mainly by its active decomposition products. This hypothesis is strongly spindle and oval undifferentiated cells (Fig. 12). supported by morphological findings showing the specific i Bones. In the metacarpal and metatarsal regions of the 4 tendency of the developed tumor to appear as a : limbs, a large proliferation of cartilaginous tissues arose mucoepidermoid carcinoma, which indicates die active -. outward from the periosteum and, from the appearance of its contribution of the mucus glandular cells to the tumorous cells, seemed to derive directly from the cortical bone (Fig. growth. Such kinds of histotypes are often related to some 14) . The periosteum also grew and In some areas spread as glandular aggregates, and, therefore, the histogenesis of human . finger-like prolongations into the newly formed cartilage (Fig. mucoepidermoid carcinoma has been restricted to the cells of . 15) . In these places, a gradual transition between fibrous intercalated duett. This hypothetical interpretion must be - cartilage, periosteum, and bone could be noticed. confirmed by future experiments in which the action of the j The newly formed cartilage appeared irregular with atypical vinyl chioride should be restricted to the salivary system. At I areas; the celis, which had nuclei larger titan the normal the concentrations used, saturation and wetting of the fur ! chondrocytes, lay in well-formed, capsulatcd lacunae. The cells might well be expected. Under these circumstance*, the occurred singly, in pairs, or in tetrads and, although of natural cleansing habit of the ru. '"g!.: add a significant different sice and shape, they usually contained a single, ingestion problem with subsequent concentration of vinyl darkly stained nucleus. The cartilaginous growth was not chioride in tlte salivary glands. Tlte local extreme homogeneous,- as shown by the extension of the fingcr-Uke concentration could be the result of the difficulty of a prolongations into the boundary of newly formed tissue, complete cleansing by the rat. esidcs the chondroblastic, chondrocytic, and angiomatous The neoplastic response of the lower respiratory tract. I reas which indicated a rapid growth, there were also although of lesser magnitude, is of relevant interest, since some CANCER RESEARCH VOL. 31 OCUMENT Oncogenic Ac/hity of Vinyl Chioriiic x tuwere ;r.oipho',oi:U'.i!ly similar ki those tloscriiicJ the .'N\n, Vi;.' hypothesis ;h,t: itiiietts-proihieir;;: colls iiuy x' c.sjuY.e of :e;.iinit'.$ vinyl chloride or its decomposition :o hr relevant once ir.*'rc. Obviously. a wii'.vtett; ;;;;e:p.e;,i;io:t is required tor the pathogenesis ol (he 00^,0 trots -i-J lor ;he;r si.mil.at.eous incidence ut the level o.fi!Ul::nbs. REFEliENCES j. ii.i.,e:*oe, y... j.-.J Dates, R. Prevalence tf llctcroploidy in Plastic lYii.'.-mduced Primary Sarcomas. Brit. J. Cancer, 20: -355-563, . :9oe>. . 2. Brand. G.. a::i Buocr., L. Polymer Tumorigencsis: Multiple Prer.coplasrie Clones ia Priority Order with Don'al inhibition. Proc. Soc. Uxptl. Hiol. Meil., I2S: 1153-115S. 190S. 5. Brand. G.. Baoen, L,, ar.d Brand, I. Malignant Transformation and Maturation in Non-dividing Ceils during Polymer Tumorigencsis. Proc, Soe. Exptl. Biol. MeJL. 675 -678. 1967. J. Brand. G., Baoen, L,, and Brand. I. Carcinogenesis from Polymer Implants: New Aspects from Chromosomal and Transplantation Studies during Premaiignancy. J, NatL Cancer Inst., 39: 663-679, 1970. 5. Drurkrey, It. I .Xjiciiiiiriiivllc ltei:r,e;:c mn.; V..-, n ,.-.is...,n C r C,ire:Mi)gi-tie Wi/k.my. Ar/neiiuitrel-l oiw:.,. /. V,> yt . I,;]. 6. llucpcr, W. C. (Nmvr Induction l>y l,iilyi..vtu.,;; .n.I |*,,l.".il,twr.c Plastic*. J. S'.,II. C.::.vr Ins!.. .U: IMH 11,27. I 7. Johiwin. k., Huu, a, L., itruit.i. I., a..d i ^ i1..., u.c: Tuuiitri.icnesis Clonal Detcrimnat.on uf lii.t ;.'no. C.ia. j, termin', dur.ug l.a.'iy .1i,i:,..ici1. s,.:; it,-!.,;,., ; . Karyotype, Mouse Strain, and See. j. X..II, C_me: -'V 7KS-793. 1970. 8. Kogan, A", Experimental Itlastomogcnetis (Nosed l-> Syr.ti-.et.c Polymers (Plasties). Mediums. Moscow, /; 215-237. 1665. 9. Mastromalteo, la-.' Fisher. A. M.. Christi, II.. and N-r./igcr. D. Acute Inhalation Toxicity of Vinyl Chloric to L-boratt-ry Animals. Am. Ind. iiyg. Avsoc.-J..21: 394-397. I960. 10. Torircrson, T. R,, Oyen; F., and Rowe. V. X. The Toxicity.of Vinyl Chloride as Determined by Repealed- Exposure of Laboratory Animals. Ind. iiyg. J,, 354-361, October 196' 11. Van Duren, B. L-. Sivaie, A.. Goldschmidt, li. M., Katz, C.. ar.d Mclehionnc, S. Carcinogenicity of Halo-Ethers- i. Natl. Cusccr Inst.. 4J.'481-486, 1969. 12. Viola, P. L. Pathology of Vinyl Chloride. 18th.' International Congress of Occupational Health. (Communication No. 38). ' Tokyo, 1969. ILLEGIBLE DOCUMENT Figures are -omitted because of failure to reproduce'adequately R&S 028280 Figs. 1 to 18. All seetions were stained with H A E. Fig. I, Squamous cell papilloma of the skin. X 25. Fig. 2. Transitional stage from papilloma to infiltrative type of cancer. X 80. Fig. 3. Finger-like trabeculae branch to form secondary processes showing homy pearl formation (errotvx). X 25. Fig. 4. Dyxkeratolie area. Few epithelial cells with a tendency to form whorls. Lillie capacity to develop into horny pearls. X 250. Fig. 5. Atypical cells partly lacking prickles and showing considerable variation in sin, shape and many mitotic figures. X 250. Fig. 6. Mucoepidermoid skin tumor. Occurrence of hydropic epidermoid ceils, basal cells, columnar cells, and oxyphilic cells in various proportions. X 100. Fig. 7. Mucoepidermoid skin tumor, showing differentiation of masses of squamous epithelium from columnar cells, lining and proliferating into distended tubular spaces. X 100. Fig. 8. Micronodular adenocarcinoma of the lung arising from a segmental bronchus. X 25, Fig. 9, Detail of Fig. 8 showing tubular aggregates made up by cells with byperchromatic nuclei Fig. 10. Nodular bronchiolar alveolar mucus-sccrcting adenocarcinoma. X 100. Fig, 11. Detail of Fig. 10, showing an alvcologcnic pattern. X 250. Fig. 12. Mucus-producing cells with byperchromatic and pleomorphic nuclei and occurrence of signet ring cells. X 250. Fig. 13. Squamous cell carcinoma of the lung. X 100. Fig. 14. Osteochondroma developing as a protuberance from the small bones of metacarpus and metatarsus. X 25. Fig, 15, Magnification of Fig. 14. The protuberance consists of bone trabeculae, capped by cartilage, and a fibrous layer functioning as perichondrium, x 100. Fig. 16. Osteochondroma. Cartilaginous growth characterized by finger-like proliferation. X 100. k Fig. 17. Osteochondroma. Area of esdochondrial ossification. X 100. r ' _ 18. Cellular area of an osteochondroma showing the transition from cartilage to osteoid. X 100. MAY 1971 519