Document GK83bq45JZVVm6dGJVoD6nMDx

1 ' ; l\ A i .i k vi ;44 -vi Adenofibrosis in the Rat Liver With Persistence of Polychlorinated Biphenyls in Adipose Tissue Rcnatc I). Kimbrough, MD; Ralph E< Linder; Virlyn IV Burse; Ralph IK Jennings, Chamblce, Cla Fifty mole Sherman strain rats were fed 500 ppm of a polychlorinated biphenyl (PCD) (Aroclor 1254) for six months. Five each were Killed zero, one, two, three, four, six, eight, and ten months after exposure to Aroclor had ceased. The livers of these rats were examined by light and electron mi croscopy. Liver lesions persisted although exposure to PCBs ceased. Ten months after exposure ceased, 1,192 ppm PCBs were still present in the rats' adipose tissue and 22.05 ppm in the rat livers. Aroclor patterns found in the tissues by electron capture gas chromatography dif fered from patterns of dietary Aroclors. Mass spectral analysis of livor end adipose tissue revealed three major Aroclor compo nents with masses of 324, 358, and 392. These contained isotopic clusters indica tive of tho presence of Cl,, Cl,, and C1(, respectively. In a previously reported study1 Sherman strain rats were fed poly chlorinated biphenyls (PCBs) (Aroclor 1254 and Aroclor 1260). The rats developed characteristic mor phological changes in the liver. These changes consisted of hypertrophy of the liver cells, a brown pigment, in Kupffer cells, lipid accumulation in the cytoplasm of hepatocytes and, at the higher dietary levels, adenofi brosis. Because of the wide distribu tion of PCBs in the environment- and Submitted for publication March 8* 1973; ac cepted April 19. From the Environmental Protection Agency, Bio-Effects Branch, Chamblce, Cn. Reprint request* to 1B5 Avery Dr NE, Atlanta 30309 (Dr, Kimbrough). their persistence, the hepatic lesion was studied further. The present study was undertaken to see whether, once cx|x>sure to PCBs was stopped, the morphological changes produced in the liver might disappear. ' Materials and Methods A Colnl of 50 mate wemiting, specifiepathogen-fiee, random-bird Sherman strain rats were tagged individually and group-caged, ten rats per cage. They were given Aroclor 1254 in their diet for six months. The dose of 500 ppm, which is equivalent to an average daily intake of 36.4 mg/kg body weight,1 was chosen be cause in the previous study at) ten c.xjiosed male rots developed adenofibrosis within a period of eight months. The Aroclor was mixed into ground laboratory chow ns pre viously described.1 After six months of Fig l.-Area ot adenofibrosis in liver section consisting primarily of duels surrounded by flattened epithelium. Ducts are surrounded by little fibrosis (homatoxylm-eosin, original magnification X 150). Fig 2.- Rat liver section showing cluster of smalt columnar cells resem bling pancreatic tissue adjacent to blood vessel. Hepatocytes surround ing lesion are vacuolated (hemaloxylin-eosin, original magnification x 175). I t 1 1It r i 4' I-',:;.-99954 T:'-9 -'9,*, v _* ` -: 4'is CV -| ''* . / ~ * V"-, a- ' ; *./'l ` W ' 'A* l; 1 39D Arch Cnvircm Heatth/Vot 27, Dec 1973 Exhibit 5,7 ' Acienofibrosis/Kimbrough et at DSW 029083 STLCOPCB4013045 Use of the present method produces nn average slope around I. For nick el, the slope changes from 0.222 in acid, to 0.C06 in this organic base. Since copper and zinc concentra tions arc determined after dilution, and therefore subject to a certain amount of dilution error, a National Bureau of Standards sample (No. 1577) of bovine liver v. as subjected to tbe same procedure and analyzed. The copper recovery was 113% i 5.0%, and zinc, 112% 6% in three sepa rate analyses. The precision of the method in re peated nnnlyses of control and ex posed lung tissue is presented in Ta ble 2. All lungs were divided into approximately five equal portions and each portion analyzed for cad mium, nickel, and zinc. Tbe standard deviations for each lurig varied from a low of 0.02 ppm to 1.56 ppm, depend ing somewhat on the exposure to the inhaled metal. Within measured ranges of cadmium and nickel con centrations, 200 to 300 mg of wet tis sue from an exposed animal are nec essary for precision. If the animal is unexposed, 300 to 400 mg are re quired for the cadmium analysis. Although one would not expect tiny difference in the nickel concentration before or after exposure to cadmium, tbe nickel concentration was below the level of detection in the control animals. Sample weights can be rc' dueed by ns much as 90% to 95%- for the analysis of zinc and copper.1 Using the data given in Table 3, the totnl amount of cadmium re covered from the lung was calculated and compared with the total amount of cadmium present in the inhalation chamber during that, period, as well ns with the estimated amount of cadmium inhaled by the rats. It can be seen tliut the percent retained (&% to 20%) of Hint "inhaled," is larger by a factor of 50 than the amount re tained from the totul exposure. These figures are somewhat comparable to those reported in the literature,11 which for short acute exposures var ied from 6% to 13% for rats, and un der chronic conditions, lasting eight months, are estimated at 2% for rab bits. Both of these studies were per formed at higher exposure levels than . those in the present experiments. Under the conditions of the present study, all animals are exposed to a relatively small particle size, (89%, are less than 0.53pm in diameter)1" which for cadmium oxide would be relatively soluble. The unimnls were killed 16 hours after rcmovul from the chamber, and the total exposures were not large. Experiments per formed in these laboratories after five- to six-week exposure periods, to be reported in the future, do not show much increase in lung retention. Under the conditions of these experi ments then, it would appear that of the codmium oxide aerosol absorbed, a substantial portion remains in the lung. In summary, data have been pre sented to illustrate a fast and rela tively precise method for the analysis of cadmium and nickel in animal tis sues.13 The technique lias been used to measure cadmium concentrations in the rat lung after exposure to cad mium oxide aerosols. This investigation was supported by the Na- tionn! Institute of Occupational Safety nnd Health grant 5-K01-OII 00347 02. ` Emil Pfitzer, PhD, assisted in the construction of tabulation experiments. William Barkley ran the inhalation chambers. Tony Horstntan, PhD, performed the particle Fir ing. 1. Friliorp I., I'iscnlor M, h'onttierg C: Prob lems of unnlysis, in Cadmium in the Environ mail. Cleveland, Chemical Rubber Co, 1971, p IS. 2. Chrislinn 0, Kelihnmi F: Atomic Atoorp iitin Sjicctivscopy- Now York, Wilcv-Intersciunce Publishers, 1970. 3. Analytical Methods for Atomic Absorption Spectrophotometry. Nonvnlk, Conn, Pci kin* Elmer Co. 1971. 4. Pulido P, Fuwa K, Vnllce 11: Determination of cadmium in biotoyical materials by atomic absorption spcrtrophotnnictrv. Anal Biochcm 14: 333 101, inns. ' 5. I'achard Technical Bulletin, publication 6051B. Downers drove. 111. Packard Instrument Co, 1970. References C. MucOcc J: Characterization of mammalian tissues and mtcworgnnisins by gus-liquid ebromnlbgrnphv. J Gas Chromatogr 6:1072-1073, 1970. ` 7. Jackson AJ, Michael LM, Schumacher HJ: Improved tissue solubilisation for atomic absorp tion. AnalChcm 4<1:106-1 -1065, 1972 8. Mustnfn MCI, Cross DO: Pulmonary alveo lar macrophage Biochemistry 10:4176-4185, 1971. 9. Bingham K, ct at: Alveolar macrophages: Reduced number in rats after prolonged inhala tion of lend seaquioxide. Science 162:1297-1299,. 19G8. 10. llorptmauS.ctal. Aerosols oflead, nickel, and cadmium: A method of generating soluble and Insoluble compounds. Arch Environ Health 2G.75-77,1973. 1). Sundcrmnn FW: Measurements of nickel in biological materials by atomic absorption spectrometry. Am J Clin Pathol 44,181-18$, 1965. 12. Fribcrg L, PtRcutor M, Nordbcrg G: Me tabolism, in Cadmium in the Encinmmcut. Cleveland, Chemical Rubber Co, 1971, pp 27-66. 13. Nlurlhy L. el at: Atomic absorption deter mination of zinc, copper, yadmium and lend in tissues solubilized by aqueous tclramclhylummoniuni hydroxide. Anal Diochctn, to be pub lished. Atcli Tnviron Henith/Vol 27, Dec 1973 Direct Dcterminalion/Kaplaii et a) 389 OSW 029084 STLCOPCB4013046 f Ig 3.-Section of hepatic tissue in latter part of recovery phase illustrating numerous small lipid vacuoles (LV) (lead cilrateuranyl acetate, original magnification x 51,3001. Fig 4.- Poition of hcpntocyle with tubules of endoplasmic reticulum (ER) arranged in disorderly fashion. N represents nucleus; M, mitochondria (lead citrale uranyl acetate, original magnificaticm x 20,235). Arch F.nviion Health/Vol 27, Dec 1973 . ' . Adenofibrosis/Kinihrouch et al 39 J DSW 029085 STLCOPCB4013047 ij JT *, - fV-J Fig 5. - Border of area of adenof ibrosis in liver. Note round clusters of dark granules intermixed with web-like clusters of vacuoles. These formations Illustrate appearance of brown pigment (BP). C represents collagen; LC, liver cell (lead citrate-urany! acetale. original magnification x 20,2351. receiving the experimental diet, the rnts residual level ten months after dietary in amounted to 3.34ft of the average j received no more Aroclor end were fed plain laboratory chow. At this time five take hud censed, adipose and hepatic com posites were analyzed by gas chromntog- body weight. The fraction of the body weight for the liver in control adult l rots wero killed. Additional killings of five randomly selected rats were made one, raphy-mnss spectrometry. The conditions for mass s|>octrnl analysis were as follows; male rats ranges from 2.44ft to 2.59ft.1 two, three, four, six,eight, and ten months after ex|>osurc to PCB had been discontin the mass spectrometer was interfaced with a gas chromatograph und equipped with Forty livers were studied micros * ued. At autopsy the livers were removed, - a mass marker accurate within 0.3 mass copically. The hepatocyles wen; en weighed, and the tissue fixed in phos units. The gas chromatographic column larged in 39 livers and all 40 livers phate-buffered 5ft glutnrnldehydc for temperature was 220 C; flash heater, 235 showed either vacuolated or foamy three hours for electron microscopic exam C; glass-coiled column, 3.05 mxO.GI cm; cytoplasm because of increased lipid ination, post-fixed in Jft osmium tetrox- 1.5ft OV-17/1.95ft QF-1 on 60/80 mesh accumulation. Cytoplasmic inclu ide, embedded in rtinraglnss, sectioned chromosorb "W" (high performance, acid sions similar to those described with a glass knife, and stained with lend citrate and urnnyl acetate. Tissue for the light microscope was fixed in 4ft buffered formaldehyde solution and stained with hematoxylin and eosin. Frozen tissue of selected formalin-fixed hepatic tissue was cut on the cryostnt and stained with oil red 0 to demount rale lipids. At the time of the lust sacrifice, ten months after exposure to washed, dimethyldichlorosilane treated),' helium prcssuie; 0.84 kg/sq cm; flow rate (rotameter setting 3), 45 m!/min; separa tor temperature, 225 C; source energy, 70 ev; decelerating voltage 3.5 kv; trap cur rent GOpamp; box current 50/rnmp, and leak current 8/tamp. . Results previously"1 were observed in the liv ens of 24 rats. Adenofibrosis, which in many instances was very extensive, was present in 36 of the 40 livers, and a brown pigment was noted in the KuplTcr cells and other macrophages in 25 livers. Of the four rats that did not develop adenofibrosis, one each PCH-contnining diets had been discontin During the course of the experi- was killed two and six months nfler ued, ndi|we tissue and liver from seven mctit three rnts died. The livers of the the poisoned.diets were discontinued. i previously exposed rats and the adipose tissue and liver from two rnts of the same rnts tlutl were killed at the lime expo sure to Aroclor 1254 was discontin The other two tats without adenofibrosis were killed ten months after age that had been fed only plain chow were analyzed lor the presence of I'Clls by election capture iKCt gas clp omalography according to methods described by Curley ct nip ued were enlarged, with average weights of 6.32ft of the body weight. When the experimental diets had been discontinued for four months, onset of exposure. Tlu> adenofibrosis ranged in size from grayish-while le sions measuring 0.5 to 1 ctn to metis of 5 cm with a pitted sm lace. Quite In oixler to determine which Aroclor tiro liver made up only 3.6ft of often the lesion was mull icent t ic. The const iluonts were ol'sueh chemirnl stabili the body weight, mid after eight tklcnofibrosis' is illustrated in Fig 1 ty that they remained at n measurable months recovery the average weight nnd2. In this study the appearance of 392 Arch Environ Henlth/Vol 27, Dec 1973 Adenofibrosis/Kinibrougli et al t DSW QZ9Q86 STLCOPCB4013048 Fig 6. - Gas cluomalograpliic traces of Aroclor 1254 standard (A), hepatic tissue extract (B), and extract of adipose tissue (C). Response expressed as percentages, with 100%-= 1.0inv. the lesion, particularly when it tens extensive, varied a p-onl deni. Small lesions consisted predominantly of glandular pale-staining epithelial cells that formed ducts and were sur rounded by very little fibrosis. The larger lesions had extensive fibrosis and contained collagen, ami often the ducts which were formed bv the epi thelial pale-staiuing cells were markedly dilated. The dilated ducts contained mucus or necrotic debris, and nl times formed small cysts mea suring up to 0.5 cm in diameter. Small dilated duets were lined by co lumnar epithelium, and larger ones by flattened epithelium (Fig 1). In areas of extensive fibrosis, and also adjacent to blood vessels hi areas of normal hepatic parenchyma (Fig 21, clusters of small glandular cells were seen in 15 of the livers which resembled the epithelium observed normally in the pancreas. These small clusters of pancreatic-like tissue have not previously been described in the livers of rats or of other species. Special stuins for esterase, esterase with sodium taurocholale, and tryp tophan showed a positive reaction indicative of salivary gland tissue. The significance of these cells is pres ently not understood. Normally tis sue of a pancreatic typo is not ob served in the livers of our rats. Clus ters of small columnar epithelial cells resembling pancreatic tissue also occurred in portal triads without l>cing surrounded by typical adenofibrosis. In the present study, one to two rats at each killing showed these small columnar cells with highly aci dophilic cytoplasm and a small, round nucleus. In the older animals, the fibrosis was quite extensive in some instances and the glandular proliferation was present mainly at the periphery of the lesion. Signs of regression of the lesion were not noted, except for the disappearance of epithelial cells from the center of the lesion. Furthermore the lipid accumulation and the hyper trophy of the hepatic cells remained constant throughout the study. Ex tensive areas of necrosis were ob served in three of the 40 livers. In addition to the change described previously,' election microscopic examinations of sections of liver tis sue showed numerous small lipid vacuoles in the cytoplasm of many hepatocyles, particularly in the latter part of the recovery phase (Fig 3). The smooth and rough endoplasmic retic ulum was arranged in a disorderly fashion (Fig 4). Many of the mito chondria were atypical and surround ed by double membranes. The electron microscopic appear ance of the brown pigment present in the macrophages and Kupfier cells is illustrated in Fig 5. I,nrgc clusters of dark granular material, intermixed with lipid vacuoles, were observed, as well ns round clusters of granular material about the size of mitochon dria. In our pivvious studies it was found that at least some of this pig- Arch Environ Health/Vo! 27, Dec 1973 Adenotibrosis/KImbrougli et at 393 DSW 02908? STLCOPCB4013049 I A 'i i / i (i .1 i ! 3 3 : Ai -J i meat contained iron1 but in addition high concent rat ions of urojiorphyrin (75ft 8-carboxyprophyrins and 25'/r 7-carboxyprophyrins) were found in JOO- the livers,31 which could account for some of the pigment. In addition to these findings, oil red 0 stain demon strated granules of red-staining lipid material in the area of the brown pigment. Since the morphological changes in the liver did not disappear, the re maining seven rats were killed after, a ten-month recovery phase and their adipose tissue and livers analyzed for Aroclor. In adipose tissue, the concen trations of PCBs ranged from 924 ppm to 1,688 ppm, with an arithmetic mean of 1,192 ppm. The concentra tions in the liver ranged from 17.30 ppm to 26.24 ppm, with an arithmetic mean of 22.65 ppm. The PCB levels in control adipose and liver tissues were <1.0 ppm. The gas chromatogram obtained from the EC analysis of adipose tissue did not differ from that of liver, but was significantly different from standard Aroclor 1254. An average or seven peaks was observed as a rest- . due in the tissues. Under, these gas chromatographic conditions", at least .18 peaks have been observed in the standard material (Fig 6). The differ ence is gas chromatographic patterns between stored and ingested Aroclor has been observed before in this labo ratory and by other researchers in the 0 ii 0 17 i i i r i ' ] i r "i i i i 3 A 5 6 7 8 9 10 11 12 13 14 field.,i-T The total ion current. (TIC) chro TIME (Minutes) matogram for the liver tissue com posite is shown in Fig 7. Mass spec Fig 7.-Tolal ion current chromatogram of sample of composite liver. Response expressed as percentages, with 100%^ 10.0 mv. . tral analysis revealed the presence of only three major Aroclor components chlorinuled moieties, while other" epithelial proliferation occurs in the with masses of 324, 35S, and 392. peaks observed in the liver TIC con periphery, while the central portion These components contained isotopic tained the following molecular ions: of the lesions contains primarily clusters indicative of the presence or peak 4 (324); peaks 5 to 7 (392 and fibrous tissue. The present study docs ClM Cl,;, and Cl., respectively. Adi 358); and peaks 8 to 9 (392). not completely resolve the question of pose tissue analysis by mass spec reversibility of this lesion since the trometry revealed the same patterns. . Comment Allbough only three different mo storage levels of PCBs in adipose tis sue and liver may have been suffi lecular components were present, The ndenofibrosis of the liver pro ciently high to stimulate continued examination of the EC and TIC duced in this study and the one pre growth of the altered hepatic tissue. chromatograms coupled with mea sured intensities from multiple scan ning or eluting peaks indicates that stereoisomers of these three mole cules tiro obviously present. Peaks 1 to 3 (Fig 7) did not contain any viously reported' is the result of Aro clor consumption. Whether the Aro clor itself, or a contaminant such as a chlorinated dibetr/.ofui an, is responsi ble still needs to be resolved. A study of the lesion shows that The levels in the liver suggest that a constant low-level turnover of PCBs takes place in the rats. In a previous study, Curley et t\P found levels of TO.OOO ppm Aroclor 1251 in adipose tissue of rats that had been fed 500 394 Arch Environ Heallli/Vol 27, Dec 1973 Adenofibiosis/Kimbrough ct al DSW 029088 STLCOPCB4013050 ppm of the mixlurc- in their diet for tight inoiilh'j. This finding imliailus Ihnt n t;re;il deal of the Aroelors had been mobilized from the adifKi.se tis sue and excreted in the present study. The storage levels found ten months after onset of recovery are in the same range of those of rats fed 100 ppm PCBs in their diet for eight months." The dietary level of 100 ppm produces: a much lower incidence of adenofibrosis,1 and it is possible that edenofibrosis represents a permanent lesion, particularly since the percent of body weight of the livers had de creased. The Aroclors found in the tissues differed from those fed in the diet. Metabolism of lower chlorinated bi phenyls is considered os a possible explanation for the differences ob served analytically between con sumed and stored PCB. Earlier work of Block and Cornish-1 has shown that biphenyls chlorobiphenyls can be hydroxylalcd and excreted in the urine of rabbits. A more recent study by Hutzingcr et aP indicates that tbe rat is capable of hydroxylnling mono-, di-( and tetrochlorohiphenyls, but not hexnchlorobiphenyls. Since Aroclor 1254 averages five chlorines per mol ecule, it is conceivable that the heav ier chlorinated moieties could remain ns a residue. P-dimethylaminoazobenz.ene also produces adenofibrosis. Edwards and White10 and Bennett et al" described the lesion in rats when a mixture of chlorinated naphthalenes and chlori nated diphenyls were fed. Unfortu nately, the finding was not well de scribed in Bennett et al's report, but Edwards nnd White mentioned Ben nett el al's finding as another exam ple of the induction of ndenofibrosis by chemicals in nn excellent descrip tion of the lesion. A critical review of the hislopnthogencsis or this lesion was made by Stewart and Snell1-. The presence of the brown pigment in Knpfler cells and macrophages is of interest nnd was to a lesser extent also observed in the earlier study.' Electron microscopic examination of the pigment shows that particularly the large clusters of it resemble very closely illustrations of ceroid pig ment.13 Experimentally ceroid pig ment can he produced by hemorrhage into fatty tissues rich in unsaturaled fatly acids, particularly in the ab sence of an antioxidant (vitamin E deficiency). Ceroid pigment is com monly referred to as lipofuscin. These lipid pigments are at times associated with hemosiderin and may contain iron-positive granules as in the pres ent case.1 Ceroid granules were also described by Edwards and White in the livers of rats fed butter yellow. Bennett et al also observed a brown pigment in rats fed a mixture of chlorinated naphthalenes and chlori nated biphenyls. These authors claimed that it failed to stain in a manner characteristic of either hemo siderin or hemofuscin. The porphy rins demonstrated in the livers of our rats fed Aroclor may also contribute to the overall pigment deposits. Con oid pigment increases in people with age and has therefore been called "Abnulzungspigmcnt" by the Ger mans. Attempts have been made to relate it to senility. It has been associ ated with neuronal ceroid-lipofusci nosis (Batten disease) in man. In minks, a disease called yellow fat disease" is associated with the accu mulation of excessive amounts of cer oid in adipose tissue. The accumula tion of ceroid in mink can be prevent ed by addition of vitamin E to the diet. It occurs when the food rations contain high percentages of fish scrap. However, Gorham et al suggest that oilier factors may also be in volved, since high dietary percent ages of fish scrap did not promote growth ns well ns the stock ration, even when vitamin E was added to the diet. These observations raise the question whether high doses of vita min E, nnd possibly the addition of choline to the diet, could prevent or minimize the observed effects of PCBs on the liver orouvrals. Richard k Moore essisii-d with lh- animal test*. kind**! W. Anderson os'dsted with preparation and interpretation of the electrotnicrugrnphs. Annie R. Alford performed tho tissue prepara tions. II. L Stewart, MD. and the tissue inlmr.itorv f the National Cancer Institute, Bcthesdu. Md, performed evaluot ions of liver slides. References I. Kimbrough UD, Linder RE. Gaines TB: Morphological change* in liver or raU fed pole- chk>rinntcd biphenyls. Arch Environ Health 25 354-304,1972. 2L Riscbrough R\V, et nl: Polychlorinuted bi phenyl* in the global ecosystem. Nature 220. 1078 1102. 1968. ` 3. Curley A, ct al; Polychlorinated biphenyls. Distribution and storage in body fluid.* end tis sues of shermnn rats, Em-iron Res 4:481 >495, 1971. 4. Kimbrough ItD,GnincsTB, Linder RL. Tho ulUastructurc of livers of rutn fed DOT and dieldrin. Arch Environ Health 22:460-467,1971. 6. Goldstein JA, Hickman P, Juo D: Hepatic porphytia induced by polychlorinated biphenyls (PCB'k): Relationship between porphyrin, 6- aminolevulinic acid synthetase ond cytochrome P-450, abstracted. Fed Proc 32:702,1973. fi. Grunt DL, Phillip* Wl-J, Villeinmec DC: Metabolism of,, a polychlorinated biphmiyl lAroclor 1254) mixture in the rat. Bull F.nnion Contain Toxicol 6:102*112,197). 7. Vos JO, Koemnn J1I; Comparative tosicol- rgic study with polychlorinnted biphenyls in chickens with special reference to |x>rphyria. odemn formation, liver necrosis nnd tissue resi due?. Toxicol Appl Pharmacol 17:656-658,1970. 8. Block WD, Cornish HU: Metabolism of bi phenyl nnd 4-chlorobiphcni I in the rabbit, J Biol Cheat 234:3301-3302, 1959. 9. HntringerO,et nl: Polychlruinatedbiphen yls: mvtabolic behavior of pure isomers in pi geons, rats, and brook trout. Science 17S.312- 313,1972. 10. Edward* JE, White J:Putho!ugic changes with special reference to pigmentation nnd e)ns- fifMMlipu of hepatic tumors in rats fed p-riimrih- ylmmnoazobenzene (butter yellow). J Xatf Cancer lust 2:157-163,0 9 ! 1. II. Bennett OA, Drinker CK. Warren Ml': Morphological changes in tho liver*; of rats re sulting from exposure to certain chlorinated by- 'drocarboits.t/ lad //.VA'Tuvirof 20:97*133. I9.1S. 12. Slowort |H.,Knell KC: The hialopotholngy f experimental tumors of the liver uf the rat; A critical review ot the histopotbogenesis. Arfn iUnh hit Contra Conemm 13:770-803, 1957. 13. Ifurlroft WS: Observation and interpreta tion oflipid pigments i LipofiNcinsi in the pathel- K'gy of lalnnatory animals. Crit Ret Tnxiail 3:379-41 J, 1972. 14. Gorham JK, Baker GA. Boe N: Observa tions on the etiolo^v or yellow fat disea.M* in imink: JYcliminarv report. Yet Med 46:100-103. 3951. DSW 029089 Arcti Environ Health/Vol 27, Dec 1973 Adonofibrosis/Kimbrough ct nl 395 STLCOPCB4013051