Document ba1nzMaDBVmjnkQ5Yjqw7ZO0k

NtMNfl AL. 'Phwph Hasp *4 ICMf Im IMM-S.1,9*17 Ids NnmI Itythwii h*n Afe.w mitut* Jt, V7 Augusts. 1977 , r i COMPARATIVE TOXICITY OF PflfvrumtiMATcn puchvi AND POLYBROMINATED BIPHENYL IN THE RAT LIVER: LIGHT AND ELECTRON MICROSCOPIC ALTERATIONS AFTER SUBACUTE DIETARY EXPOSURE Louie Katza, Morris A. Weinberger Division of Pathology, Food and Drug Administration, Washing ton, D. C. David E. Hinton, Benjamin F. Trump, Chimanlal Patel Department of Pathology, University of Maryland, School of Medicine. Baltimore, Maryland Leonard f riedman, Larry H. Carthoff Division of Toxicology, Food and Drug Administration, Washing ton, D. C. The comparative toxicity of po/ychformated biphenyl fPCBj and polybrommated biphenyl(BBB) in liven was studied in male Hollzman rats, lout-weekold animals were ledat dietary levels of ft 5, JO, or 500 ppm tor 5 weeks and then sacrificed the mean hvei body weight ratios of the 50 and SCO ppm groups were increased. Wist pathohgk examination ot the livers revealed laity degenerative change associated with both compounds. This change was more marked al 500 than at 50 ppm. Various sized lamellar cytoplasmic inclusions were defected in livers ot animals led 500 ppm o/ either compound. However, the inclusions were more numerous in the 700-treated rats. Several animals led 50 ppm PBS had a lew inclusions. In rats that received 500 ppm PBB, hypertrophic degenerative hepafocyfes were present aiound the centra/ veins. On the periphery of this change there were occasionally muhtnucleated hepatocytes. Electron microscopic examination at a dose level of 5 ppm Jn both the 7C0 and 700 groups showed a slight proliferation ol smooth endoplasmk retkulum ($181 a moderate increase ol lipid droplets and some liposome*, and a marked proliferation ol Colgi condensing vesicles containing lipoprotein particles. A decreased number of mitochondria and lysosomes was also observed At 50 ppm, similar but more marked ultrattruciuta! alterations were seen, in ad dition, an increased number ol brant hed and cup-shaped profiles of mitot hondrta and a decreased number of Coigi condensing vesicles containing lipoprotein pani cles were observed Concentric membranous cytop/asmir whorls were encountered only in the 50 ppm POP-treated rats. At 500 ppm the number ol mitochondria tie creased in both groups. There was also a marked increase in the number ol S18 and liposome* concomitant with a decreased number ol Coigi condensing vesicles con taining lipoprotein granules. Membranous whorl* were also present m the 500 ppm Please send fequests lor reprints to Dr 1 oms Kas/a, Division oi Pathology (MM-1341.1 out and Drug Administration, 200 C Street. S W , Washington. D C 20204. U S A |wmwl oi environmental Pathology and Toxicology, 1:241-257 Copyright f)117l by Patholox Publishers, Inc. MOMS 001333 f *; m 4 242 RAS/Aff At. INTRODUCTION Based upon feeding experiments using commercial mixtures of poly chlorinated biphenyls (PCBs) (Kimbrough ef a/., 1972, Littcrst and Van Loon, 1972,1974; Litterst ef al., 1972; Turner and Green, 1974; Kasza ef a/., 1976) and chemically pure isomers of PCB with chlorine substitution at the 4 and 4' positions (Hansell and Ecobichon, 1974), a well-established series of events is seen in Ihe rat liver Proliferation of smooth endoplasmic reticulum (SER) (Kimbrough et af1972; Hantell and Ecobichon, 1974; Kasza et a/., 1976) occurs with a concomitant rise in activity of enzymes of the microsomal mixed-functionoxidalive system (Litterst and Van Loon, 1972,1974; Litterst et a/., 1972; Turner and Green, 1974) and an increase in microsomal protein (Litterst and Van Loon, 1972). These changes are frequently associated with hepatocytomegaly, increased liver weight, and increased liver-body weight ratios (Kimbrough eta/., 1972; Litterst and Van Loon, 1972; Kasza ef a/., 1976). Despite the fact that a markedly fatty liver develops (Kimbrough eta/., 1972; Litterst eta/., 1972; Kasza eta/., 1976). no necrosis occurs with a one-month exposure at dietary levels of up to 500 ppm (Kasza eta/., 1976). Another halogenated biphenyl compound that has received attention currently is polybrominated biphenyl (PBB) and, more specifically, the commercial mixture FireMaster* BP-6. Its importance has been emphasized by an accidental contamination of animal food, which led to a widespread poisoning of farm animals in Michigan Oackson and Halbert. 1974). Lee cf al. (1975a, 1975b) studied the morphologic alterations in rat liver brought about by oral intubation and short-term feeding exposure to octabromobiphenyl. The liver was enlarged, and the hepatocytes had hypertrophy, margination of basophilic cytoplasm, foamy cytoplasmic vacuolation, laminated cytoplasmic inclusions, and proliferation of SIR. The effect of TireMaster BP-6* on hepatic drug metabolism was also studied (Dent ef a/., 1976). Although the contamination of the environment (Gustafson, 1970; Kolbye, 1972) and the morphologic alterations associated with PCB toxicity have been the subject of several investigations (Kimbrough et al., 1972; Hansell and Ecobichon, 1975; Kasza ef al., 1976), limited morphologic study has been done on PBB (Lee ef a/.. 1975a, 1975b, Sleight and Sanger, 1976). The primary objective of the present study was to compare the relative hepatoloxicity of equivalent doses of PCB and PHB on liver morphology, using light and electron microscopy. The study described here is part of a larger study in which the biochemical and cytogenetic effects of feeding 5, 50. or 500 ppm of PCB or PBB for 2, 3, or 5 weeks were investigated (Carthof f ef al., 1977). MATERIALS AND METHODS M0NS 00 U3* Animals Four-wcek-old male Holtzman rats, divided into six groups consisting of seven rats each, were fed the following diets: 5, 50, or 500 ppm of either XASZAI1AI mn o( polyrst And Van Kaspa ef a/., iddionAt the iMshrd s Tips rndoplasmic chon, 1974, I enzymes of i Van | (Kin, n increase in changes are eight, and mt and Van hr (aHy liver *, 1976), no of up to 500 ad attention KkaNy, the has been nfckh Ird to wd Halbert, adorn in rat exposure to locylrs had rytopiasmic lion at SIR. m we* also rfsow. 1970; with PC rough rt il. (*), limited Tlib; Sleight ady was to :B and PBB The study emkal and t (or 2, 3, or Is romisling |m ol either PCS AND FBR. PATHOIOCV AND ELECTRON MICROSCOPY PCB (Aroclor 1254) or PBB(FireMaster BP-6) A seventh group of nine rats served as a common control. The PCB test material was obtained from lot number 370-71-698 and the PBB from lot number 1050-74-55, which arc held by the Food and Drug Administration to serve as reference materials The chemicals were ground into the commercial rat chow diet, using a large mortar and pestle. These concentrates were serially diluted to prepare the test diets; see Garthoff ef at (1977) for additional details. The control group was kept in a different room to avoid any possible cross-contamination. The animals were housed individually in stainless steel cages with wiremesh bottoms The daily lighting schedule was 12 hr of light followed by 12 hr of darkness. The rooms were kept at 72F and 50% humidity Feed and water were available ad libitum. The animals were fed with their dietary levels for 5 wk; control animals were fed the same food without test materials. After 5 wk, rats were sacrificed and, at the time of necropsy, whole body and liver weights for each animal were recorded to determine mean liver weights, mean body weights, and liver- body weight ratios All materials and excreta were incinerated to prevent environmental contamination. Histopathology Livers were collected from test animals and controls and were fixed in 10% buffered formalin. After fixation, the tissues were trimmed, dehydrated in a graded series of ethyl alcohol, infiltrated with xylol, and embedded inparaplast. Embedded tissues were cut-in 5-6pm sections and stained with Mayer's hematoxylin and eosin (H & E) for histopathologic examination. Histochemistry Special stains of selected liver sections were made. Trichrome stain was used to detect fibrosis. Clycogen in hepatocytes was evaluated by periodic acid Schiff reaction. To demonstrate lipid, oil red O-stained frozen sections of the liver were prepared Electron Microscopy Electron microscopic examination of livers was conducted on control animals and on animals exposed to 5, 50, or 500 ppm of PCB or PBB. Livers for electron microscopic examination were minced (1 mm1) in 3 percent glutaraldehyde in 0.1 M phosphate buffer (pH 7.4) and fixed for 2 hr. Following a single buffer rinse, tissues were postfixed in 2 percent buffered osmium tetroxide for 1 hr. dehydrated in graded ethyl alcohol solutions, embedded in Epon (Luft, 1961), and sectioned on a Sorvall MT-1 ultramicrotome. For correlation between light and electron microscopic observations, semithin sections (0 5-1 pm) of Epon-embedded material were stained with toluidine blue (Trump et at, 1961) and viewed under a light microscope Thin sections were stained with uranyl acetate (Watson, 1958) and lead citrate (Venable and Coggcshall, 1965) and viewed under a IEOLCO 2B electron microscope. ' HONS 001335 i 144 MS24II Al. Six electron micrographs were recorded from each section at a primary magnification of 14.000X. Random sampling of fields was ensured by positioning the fluorescent viewing screen of the microscope into one specified corner of six consecutive squares of the supporting copper grid as previously described (Weibel etal., 1969) RESULTS Weights The mean body and liver weights, mean liver-body weight ratios (expressed as percentages), and statistically significant differences between groups are given in Table 1. The mean liver-body weight ratios of the 50 and 500 ppm groups were increased over the control with both PCB and PBR. Histopathology In PCB-treated rats the most prominent liver alteration was fatty de* generation, which was most marked in the 500 ppm group. This change manifested itself by intracytoplasmic vacuolation which occurred either as a large, rounded empty space that occupied almost the entire cytoplasm of the hepatocyte or as fine intracytoplasmic vacuoles that gave the cells a foamy appearance (Fig. 1) Mononuclear infiltration was present in a few foci. The most common location of these foci was adjacent to the fatty degenerative areas (Fig. 2). The location* of intracytoplasmic vacuolation in the hepatic lobules were in the midzonal regions (Fig. 3). Most of the involved cells were hypertrophic, and cellular pleomorphism could be observed. Several of the nuclei were pushed to a marginal position within the cells. In those cells which had cytoplasmic vacuoles, oil red-O stain demonstrated fat droplets In livers of rats fed 50 ppm PCB, qualitatively similar changes were present, but the changes were less severe. In the PBB-treated rats, fatty degeneration occurred in a similar location to (hat seen in the livers of rats treated with PCB. Most of the fatcontaining vacuoles occurred as small droplets, giving a foamy appearance to the cytoplasm; there were large vacuoles too, but in smaller numbers than seen after PCB. In addition to the fatty changes in the PBB*treated rats, the most striking changes occurred in the pericentral zone. Here the cells became markedly hypertrophic with well-marked borders. The cytoplasm became rather homogeneous and stained lightly with eosin, giving a "ground-glass" appearance. The nuclei lost their central orientation, and several showed pyknosis In a few cells there were no nuclei These degenerating cells surrounded the central veins and. at the periphery of the lesions, there were multinucleated hepatocyte* (Fig 4). Bile duct hyperplasia was also observed At the 50 ppm dose level, all these changes were present but to a lesser degree. There were no changes at 5 ppm and in the controls. Lightly stained, eosinophilic, lamellar cytoplasmic inclusions were detected in both the PCB- and PBB-treated animals at the 500 ppm dose HONS 001336 i ftr$ 3 2 C? " f> V Hi; -3 ,, C 2-5 isLsi <^5< ? O O TABLE 1- Effects of PCS and PBB on Mean Liver and Body Weights and liver-Bodv Weight Ratios in Rats1 Dietary Level (ppa) Weighta PCI O' 5 50 500 5 PBB 50 500 Liver, g 11.49+0.50 13.2040.69* 13.82+1.7ft1 U. 88+1.95* 12.72+0.72* 19.44+1.42* 23.72+3.43* Body, g 352.0+28.2 380.0+14.1 384.5+34.4 306.5+26.9 364.0+23.3 388.0+16.8 286.2+50.6 Liver-body, X 3.28+0.18 3.47+0.11 4.17+0.25* 'Valuesare means SDforn 9 (controls) or 7 (treated groupt) JUsed as control value for ail treated groups 'Significantly different from control. p<0 05 6.44+0.18* 3.50+0.10 5.02+0.52* 6.36+0.95* >. JL, i riCUIIC 1. Fatty vacuolation o4 thp liver of a rat treated with MX) ppm PCD fm 5 wfc. Several ot the m tracyloplasmir va urrlej have a "foamy" appearance H ft I stain, 250X. r .--.l* 1 t ' k* j *> v ..`h i . t* * " ?vV"o-. 1 /, > ".* . rf?f * i.' -r. i . us. <** % '1 (ICUNI 2. Mononuclear infiltration present aritaccnt lo tie fatly dcgem-ta'ivr area m the liver of a ral treated with SOO |>pm PCD for S wit H ft t slain, 2SOX r- :v:a>y ^0[00'0; :4~; i - v: *i MCUftt 3. Intia< ytoplasmit varuoiatiun with a predominantly midronrl arrangement m the Irvrr pi a rat treated with SOO ppm PCD lor 5 wit lift l slam, 100X HONS 001338 KAUAIT Ai. Weird a rat PCS AND PH, PATHOLOGY AND ELECTRON MICROSCOPY 247 " *. s-T "* .**rf> r * w/, *. fU* 's p t> ! ; t Y- f e a- . . / /* t, ty i* `y,r '2 f :"-eV? tv. " *V e. *i ^ ^ l Vv'% ^J. * At s< c i> o ' ' i t *>' ' *<- ' r . --?.' -i-rtf. <m -*.*-Sr*. . IICURI 4. Markedly hypertrophic and degenerating hepatocytes with a "ground-glass" appearance, tumpndmi the central vein in a rat treated with 500 ppm POB for 5 wk. There are multmuclreted grant cells (arrows) MAI stain, 2S0X. level (Fig. 5). The number of inclusions was greater in the livers of PBB* treated rats. At the 50 ppm dose level, inclusions were present only in the PBB-treated group. Inclusions in the affected areas did not show specific zonal distribution. Hypertrophied cells were seen to contain several inclusions. Generally, the inclusions were spherical, varying in size from a few microns to such large dimensions that they occupied almost the entire cytoplasmic space. Several inclusions were surrounded by a narrow halo, and the ring-like inclusions surrounded empty spaces in the centers. High magnification of some inclusions revealed a laminated structure Inclusions were seen in both the slightly and markedly affected cells. Several inclusions were detected in cells where fatty changes were not seen by light microscopy. riGURI 5. Several intracyloplasmir inclusion* arc (nescnt (ariows) in a rat treated with 500 ppm PBH (or S wk A lew nuclei show pyknosis. HAL Siam, 400X MQNS 001339 24a KAS7A IT Al. Histochemistry The special stains to detect fibrous proliferation and an increase of glycogen did not react specifically The oil red-O stain demonstrated m increase of lipids in the cytoplasm of hepatocytes in both the PCN- and PBB-treated rats at the 50 and 500 ppm levels, with the greater increase at the 500 ppm level. At 5 ppm and in controls there was no lipid increase Electron Microscopy Toluidine blue-stained semithin sections of control and treated rat liver confirmed the findings previously noted In paraffin sections In (he treated groups markedly hypertrophied hepatocytes were seen, which contained large areas of cytoplasm free of dark-staining granules (mitochondria and lysosnies). By electron microscopy, these areas were sites of SER proliferation and decreased numbers of mitochondria ami lysosoroes. A survey micrograph of a typical control hepatocyte is shown in t ig 6 Control hepatocytes contained abundant stacks of rough endoplasmic reticulum (RliR) in parallel arrays. At the periphery of RER stacks, variable amounts of SER were encountered. Mitochondria were numerous, were randomly positioned within cytoplasm, and revealed typical crisMe ami matrix granules. The Colgi apparatus was encountered in most riCUR! 4. Portion of two hr*|Mtot ytrs o( control Ml Golgi hihi.h.iUis ((.A) show* riittpndttl < ish trw Nolr* o4 Rl R 16.000X MOMS 001340 HAWAII Al inc rease of xtMrated an w fCB- and increase at nrreasc. treated rat ions. In the een# which f granules areas were ontfria and tm in Fig. 6. ndoplasmic Iks, varblr *04*, were trfetae and f in most res ANO PBB. PATHOLOGY ANU IUCTRON MICROSCOPY 349 micrographs and often contained granules (250-000 A in diameter) of lipoprotein (Jones ef a/., 1967; Glaumann ef a/., 1975); these granules were particularly obvious in condensing vesicles (Fig 7). All aspects of the ultrastructure of control hepatocytes conformed to previous descriptions of normal liver (Bruni and Porter. 1965). The major ultrastructuraf findings in control and treated livers are given in Table 2 Both normal-appearing mitochondria and branching, cup-shaped forms (Fig 8) were seen in animals fed 5 ppm PCB or PBB. The RER was arranged in parallel stacks as seen in controls. The SFR of (lie cytoplasm of this group showed a slight increase over that of controls. SER was found both at the periphery of the cells and near the nuclei. A moderately increased number of intracytoplasmic lipid droplets was seen after 5 ppm exposure to PCB or PBB in comparison with controls. Cytoplasmic lipid usually appeared as round droplets of low electron density. These often appearecLio be impinging upon adjacent membranes, producing a "pseudomembranous" arrangement. This was described previously in livers of PCB treated rats (Kasza et a/., 1976). Other lipids appeared within membranes of the endoplasmic reticulum (Fig. 9). referred to as liposomes (Lombardy and Oler, 1967). Still other lipids were seen as lipoprotein particles within Golgi condensing vesicles. The number of Golgi condensing vesicles containing these particles was increased after exposure to 5 ppm PBC or PBB Exposure to 50 ppm PCB or PBB was associated with a marked proliferation of SER. RER appeared as meandering tracks. In addition, the number of mitochondrial profiles appeared to be decreased in both groups. The number of cytoplasmic lipid droplets was MNliHlrMMr nCURt 7. Portion n< Iwpalocvtr hum control al showing Golgi apparatus with adtactnr Golgi condensing vrsic Its (( V) lonlaming Upopiottin partk Its. %,00C)X HONS 001341 1A9U l Mint iMfiMtvttutii (mdinit in Control and PCB- and PBB-TreatedNat thaw1 -Gsoup Mitochondria Endoplasmic reticulum Smooth Bough No. of No. of lipid lipo droplets somes No. of Golgi condensing No. of vesicles containing membrane lipoprotein particles wherls Control Orthodox configuration PCD, 5 ppm Oecsalonal branched and cup-shsped profiles PBS, 9 ppm Occasional branched and cup-shaped profiles PCS, 50 ppm Increased number of branched and cup-shaped profiles nw, 50 PfMi Increased number of branched and cup-shaped profiles PCB, 500 PP* Decreased number of profiles m, 500 PI*"' Decreased number of profiles >1/3 field In 2 of 96 fields >1/3 field In 5 of 96 fields In parallel stacks In parallel stacks >1/3 field In 7 In parallel of 96 fields stacks >1/3 field In 62 Meandering of 96 fields tracks >1/3 field in 48 Meandering of 96 fields tracks >1/3 field In 39 Meandering of 96 fields tracks >1/3 field In 64 Meandering of 96 fields tracks 20 31 45 48 196 177 126 2 36 73 51 50 957 523 146 236 244 101 80 73 105 0 0 0 0 11 5 11 'ifmdiega af bated upon examined o< 6 micrographs (14,000 x original magnification) per block. 4 block* per animal, and 4 animals per group Only hepalm Via cytoplasm was studied loUl micrograph* per group 96. HONS 0013*2 res rta,and faimolocy and iuciron microscopy 2SI IlCUtl 4. Portion of hepatorytr rytoplatm from rat exposed to S |pm PBR for 5 wk Notr imtentnl surface of rmtochondrion forming "cup-shaped" profile with lipid droplet in concavity J5.000X increased over the 5 ppm groups. PBB-treated animals showed more lipid droplets than the PCB-treated animals The number of liposomes in PCBtreated animals was greater after 50 ppm than after 5 ppm exposure; Ihe reverse was true for the PBB-treated animals However, the number of liposomes produced was the same for both 5 ppm PCB and PBB Membrane whorls were encountered in the 50 ppm PBB-exposed animals but not in the 50 ppm PCB-exposed group: a total of 11 were seen in 1 different animals. These membrane whorls (Fig. 10) appeared to arise from smooth-surfaced endoplasmic reticulum and were always seen in areas of marked SER proliferation. The number of Golgi condensing vesicles containing lipoprotein particles was markedly increased in both groups at the 5 ppm exposure level. However, after 50 ppm, the number of these structures was less than that of controls. Animals exposed to 500 ppm of PCB or PBB revealed a reduction in Ihe number of mitochondria per field, marked SER proliferation (Fig 11). disarray of RER. and lipid accumulation. The number of cytoplasmic lipid droplets in 500 ppm PCB-treated rats was higher than that seen after 50 ppm. Fewer lipid droplets were seen after 500 ppm PBB than at 50 ppm, but the number was greater than in the controls and lower dose exposure groups. The number of liposomes concomitant with a decrease in the number of Golgi condensing vesicles containing lipoprotein granules was markedly increased in both groups In addition..exposed animals frequently contained Golgi condensing vesicles with flattened cisternac (Fig. 10). Membrane whorls were encountered in livers of rats exposed to both PCB and PBB at the 500 ppm concentration. DISCUSSION AND CONCLUSIONS The effects of PCB and PBB on the livers of 9-week-old male Holtcman rats were compared. The primary lesion produced by PCB was fatty HONS 001343 252 KASZA IT At r. > a' V i*.: ,,( < j* ' IJ 'K*. i \ * .* ** -*>*:,. * / " tvi `V.:... V' \;<C -:-,k ''M 4i .hNc'&m. ``Tilfni-ft IICURE 9. HriMtocyte cytoplasm from rat exposed to 50 ppm PCI* for 5 <*k Not*- kp*d of <Saractprdtk low efectroodanstty within membranes of smootHsoifaced endoplasmic reticulum 44.000X degeneration, multilobular and occurring in the mid- and pericentral zones of the lobules. A limited number of cytoplasmic inclusions was detected, but only at the 500 ppm dose levels. Cellular hypertrophy accompanied the fatty degeneration in the involved areas. The fatty degeneration was less intensive than we saw in Osborne-Mendel rats fed PCB at the same dose levels (Kasza et a/., 1976). Also, lamellar inclusions were not seen in that strain of rats, which might indicate strain differences in reaction to PCB. In the PBB-treated rats the fatty degeneration also occurred in the mid- and pericentral zones of the liver lobules. Lamellar cytoplasmic inclusions were numerous at the 500 ppm dose level, and a few were detected at the 50 ppm dose level. The intracytoptasmic inclusion bodies detected by light microscopy corresponded to the concentric laminated membranes of SER found by electron microscopy (Figs. 5 and 10). The most striking difference between the PCB- and PDBtreated rats was the presence of hypertrophic cells around the central veins in the PBB-exposed rats. The enlarged hepatocytes had ground-glass appearing cytoplasms. These cells surrounded the central veins at the depth of one to three cell thicknesses. Most of the inclusions occurred adjacent to these areas on the peripheries HONS 001344 RMIAil Al. PCS AND PM, PATHOLOGY AND CUCTRON MICROSCOPY . ;vw-y*r.. yr?rt:*aM9 2S3 erkentral lions was sertrophy The fatly t rats fed nrfusions defences Sion also lamellar el and a optamk I to the croxopy rod PBB- i central nd-glass is at the orcurred 'J V'J b i 'v .5 . mo; MCURI 10. Poition of two hrpatocytci from ral npmed to SO ppm PBB for 5 wk Smooth-uattsi t*d ndnpIlMnii reticulum Iwt mofifrittrd wid f ills cytoplasm. Col|t*pp*rtut.(GA) ttows fbttnwd tistniur 71.000X. Electron micrographs from control and PCB- and PBB-treated animals revealed qualitative similarities in ultrastructural alteration. Both compounds caused proliferation of SER, which was particularly pronounced in the 50 and 500 ppm groups. In addition, liver weights and liver-body weight ratios increased upon exposure to either compound at all concentrations studied (Table 1). Increased lipid within hcpatocytes followed exposure to both compounds; furthermore, the sites of lipid accumulation (i.c., as free fat droplets in the cytoplasm and as membranebounded liposomes) were identical. PCB and PBB caused an increase in cell size, with affected hepatocytes showing rounded margins with extensive areas of cytoplasm free of mitochondria and lysosomes. These findings confirm previous morphologic reports on PCB (Nishizumi, 1970; Kimbrough et al, 1972; Norback ct al., 1972; Koller and Zinkl, 1973; Kasza el al, 1976) and PBB (lee et al, 1975a, 1975b; Corbett et al, 1975; Sleight and Sanger, 1976). Although enzyme activity in the hepatic mixed-function oxidative system was not measured in the present study, it has been shown that proliferation of SER by halogenated biphenyls is associated with a rise in enzyme activity (Litlerst ct al, 1972; Dent et al, 1976; Sleight and Sanger. 1976). HONS 001345 KASZAtT At IICURC 11. Portion of hepatotylc from ral pojd to MX) ppm PBIt for S wk Nurievs (N) * at lop of field. Cytoplasm contains parallel membranes in wltorl arrangement. 21.Q00X. Quantitative differences were seen in relation to liver weight. Values for PBB-treated animals were higher at each exposure level (Table 1). In addition, membrane whorls or laminated cytoplasmic inclusions occurred after exposure to 50 ppm PBB but not 50 ppm PCR. At 500 ppm, membrane whorls were seen in the hepatocyte cytoplasm of both PCBand PBB-treated animals, but the number of whorls was higher with the latter compound. Although the significance of the membrane whorls has not been established, they arise from endoplasmic reticulum (Smuckler and Arcasoy, 1969) after exposure to various compounds, including phenobarbital (Burger and Herdson, 1966), toxic fat (Norback and Allen. 1969), DDT (Ortega, 1966), ethionine (Steiner ef a/., 1964), and inhibitors of protein synthesis (Hwang ef a/., 1974). S ince membrane whorls form during normal and regenerating processes, including embryonic formation of egg membrane during vitellogenesis (Sweeney ef a/., 1970) and regenerating liver (Becker and Lane, 1965). those whorl-like membrane structures may be indicative of a normal stage in membrane formation and/or repair (Hwang ef a/., 1974). Hutterer eta/. (1969) described the hypertrophic hypoactive SER as the transition between adaptation and degeneration or injury. Since membrane whorls represent hypertrophic SER. the question of the functional ability--of these membranes-following polyhalogenated HONS 001346 MUAItAl. I PI. Values tWrl) In loccuncd mw ppm, Wth PCB ' with the 'both has SmucfcJrr toelading ftd Alien, Mors of i* during on of egg iterating Mies may or repair kic hyj>oration or question Dgmatcd PCS AND PBB, PATHOLOGY ANO ELECTRON MICROSCOPY 25S biphenyl exposure is important. In this regard. Sleight and Sanger (1976) demonstrated enhanced activity of hepatic mixed function oxidative enzymes concomitant with membrane whorl formation. Based upon our observation of 96 randomly selected fields of identically magnified hepatocyte cytoplasm, (he number of Colgi condensing vesicles containing lipoprotein particles appeared to increase in both PCB- and PBE-treated animals after exposure to 5 ppm of the compounds (Table 2). A decrease occurred after exposure to 50 and 500 ppm, and, in these animals, the Golgi cisternae appeared to be flattened (Fig. 10). Concomitantly, in the 500 ppm animals, the number of liposomes increased 20-fold in the PCB-troatcd rats and 10-fold in the PBB-treated rats. Litterst et a/. (1972) found increases in hepatic triglyceride in PCBexposed rats. In the rat liver, the primary pathway for utilization of hepatic triglyceride is secretion into the circulation in the form of lipoproteins (Jones ef a!., 1967; Stein and Stein, 1967; Glaumann cf a/., 1975). This process appears to involve synthesis of protein, triglyceride, and other lipid moieties, conjugation of lipid and protein, intracellular transport via the endoplasmic reticulum and Golgi apparatus, intracellular migration under the influence of microtubules (Orci cf a/, 1973), and secretion into the extracellular space. Our data indicate that a blockade in the transport of lipid occurs upon exposure to PCB and/or PBB (Kasza et a/., 1976 and this paper). Furthermore, the blockade occurs at the Golgi apparatus or between the endoplasmic reticulum and the Colgi apparatus. In addition, Mehlman and associates (1974) found that exposure to PCB decreased the availability of free fatty acids for hepatic mitochondrial oxidation. Such decreased utilization would be expected to lead to fatty liver (Cederbaum et a/., 1975). 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