Document MJ887BbrJjRG3NvRvpZnyY39L

DOW CHEMICAL U.S.A. 1803 Building 26 Noveroher 1984 MIDLAND. MICHIGAN 48640 Brian Bennett, M.O. Imperial Chemical Industries, Ltd. Hi 11 house Site P.0. Box No. 4, Thornton-Cleveleys Blackpool FY5 4QD ENGLAND Dear Dr.Bennett: You may find the attached paper by Tamburro, Makk and Popper of interest (Hepatology 4(3):413-418 (1984)). I'm sure Maurey Johnson can fill you in on details. I'm proceeding on the assumption that Sir Richard Doll will be asked to review the epidemiology literature on vinyl chloride. I passed the information to Dr. Johnson since he is Chairman of the CMA and the SPL vinyl committees concerned with health and toxicology. Maurey indicated strong interest and support. I will do all I can to expedite the formalities, but I cannot imagine anything less than wholehearted support by U.S. industry. The timing is appropriate since we should have the Environmental Health Associates' update of the old Tabershaw-Cooper study fairly well along by then. The Doll review will supplement the EHA report and may even indicate areas that need special analysis. It is essential that the review be published. I will be glad to do all I can do to aid him in getting papers and reports. Thank you for the information on your conversations with Sir Richard. I hope your joint review of the angiosarcoma registry is proceeding well. Sincerely yours. Theo'dore R. Torkelson, ScD. Health and Environmental Sciences Enc. cc: M. N. Johnson, M.D. - B. F. Goodrich Co. Carol Stack, PhD. - CMA AN OPERATING UNIT OP THC DOW CHEMICAL COMPANY 1934 DOW RESEARCH 1M4` CMA 009795 - 0270-9139/84/0403-0413*02.00/0 H -PATOLOCY Copyn*ht O 1984 by Um Am*ric*n Anocistion for th* Study of Liver DUtun VoL 4. No. 3, pp. 413-418,1964 Pnnud in U.S.A, Early Hepatic Histologic Alterations Among Chemical (Vinyl Monomer) Workers Carlo H. Tamburro, Laszlo Mark and Hans Popper Departments of Medicine, Community Health, Pathology, and Liver Research Center, University of Louisville, and St. Anthony Hospital, Louisville, Kentucky, 40292, and the Stratton Laboratory for the Study of Liver Disease, Mount Sinai School of Medicine of the City University of New York, New York, New York 10029 Focal hepatocellular hyperplasia and focal mixed (hepatocytes and sinusoidal cells) hyperplasia are early histological alterations indicative of vinyl monomer exposure. To evaluate their uses in screening chemical workers, 93 liver biopsy specimens from 78 persons were examined in d ubleblind duplicative fashion. Forty-eight specimens were from exposed chemical workers, 35 f them having liver biopsy(ies) for hepatic test abnormalities and 13 for nonliver-related reasons. A comparison group consisted of 30 nonchemical workers who had undergone liver biopsy for nonliver related reasons. Twenty-three of the exposed workers (48%) had hepatic lesions consistent with exposure; 17 (35%) of these had focal hepatocytic hyperplasia, while 6 (13%) had focal mixed ' hyperplasia or more advanced lesions. Only five of the comparison group had like findings: four (13%) had focal hepatocytic hyperplasia; one had focal mixed hyperplasia and sinusoidal dilatati n. This individual had persistent hepatic test abnormalities with the focal mixed hyperplasia and a sinusoidal dilatation, and on subsequent biopsy, angiosarcoma (and a history of using hair spray containing vinyl chloride propellant). Ten individuals had 25 multiple biopsies also read doubleblindly; 10 had two or more readings of the same biopsy. Duplicate 21 of 23 (91%) and multiple 27 of 28 (96%) biopsy interpretations in the same individual were identical. Only 6% of either duplicate and/or multiple readings disagreed. Both focal hepatocellular and mixed hyperplasia were always associated with abnormalities in hepatic test results of which indocyanine green clearance was the most sensitive and 7-gIutamyl transpeptidase the least specific. Focal hepatocel lular hyperplasia is a more common finding and occurs earlier than the focal mixed hyperplasia among vinyl monomer exposed workers. Follow-up for 5 to 7 years yielded no biochemical r histological evidence of spontaneous progression of this lesion in a work environment where vinyl chloride levels were 10 ppm with time-weighted averages of 1 to 2 ppm. Focal mixed hyperplasia is, as noted, a later and less frequent finding among vinyl monomer-exposed workers. Som individuals with this lesion develop hepatic angiosarcoma independent of subsequent vinyl mon omer exposure. In screening for chemical liver injury, indocyanine green clearance supplementary to routine tests is recommended with confirmation by liver biopsy. Industrial exposure to vinyl chloride, other vinyl mon omers, and related chemicals is associated with various histological lesions in the liver, notably subcapsular, portal, and perisinusoids! fibrosis, and hyperplasia of hepatocytes and sinusoidal cells (1-4). Focal hepatocytic and focal mixed hyperplasia (hyperplasia of hepatocytes and sinusoidal cells) are early histological alterations (5, 6). To determine their applicability for screening exposed workers, liver biopsies from 78 individuals were studied Received Much 18, 1983; accepted November 2,1983. Portion* of this work were supported by NCI Contract NO-l-CN55212. Address reprint requests to: Carlo H. Tamburro. MD, Liver Recatch Center, University of Louisville, School of Medicine, Room 119A, Louisville, Kentucky 40292. in duplicate blind fashion to determine whether these lesions were: (i) associated with vinyl chloride exposure, and (ii) occurred in nonexposed populations without hepatic disease. MATERIALS AND METHODS Liver biopsy specimens from 48 vinyl monomer chem ical workers and 30 nonchemical workers with and with out hepatic biochemical abnormalities were studied. He patic tests were performed annually or semiannually on all 48 chemical workers; these included determination of activities of SGOT, SGPT, alkaline phosphatase (AP), and y-glutamyl transpeptidase (GGT), as well as total bilirubin, prothrombin time, total serum protein, albu min and indocyanine green clearance (ICG). In addition, a history and physical examination, radioisotopic liver- 413 CMA 009795 ' < 414 TAMBURRO ET AL. Hepatology ~ spleen scan and abdominal plain film were done an 1):546, Abstract] obtained after prolonged vinyl chloride nually. Thirteen of the chemical workers had liver biop exposure have identified lesions in the nontumorous sies performed for nonliver-related reasons, and 35 had parenchyma (i) Foci of enlarged hepatocytea with in biopsies done because of abnormalities in screening tests creased cytoplasm, and large hyperchiomatic nuclei were and/or the liver-spleen scan. seen; the hepatocytea varied in size and more often ha^^ A comparison population consisted of 29 individuals several nuclei. The foci were multiple, poorly circur^B who were not chemical -workers but who had undergone scribed and differed from the surrounding parenchynu^^ an abdominal operation and gave informed consent for where there were twin hepatic plates with nuclei adjacent a liver biopsy, done during the same 3-year period at the to the sinusoidal border (Figure 1). In these areas, sinu same hospital, by the same medical-surgical team. The soids often were slightly dilated and the reticulin frame same hepatic tests, except for GGT and ICG clearance, work was focally increased as shown by silver impreg were done on this group. None had a history of exposure nation (Figure 2, A and B). (ii) Sinusoidal cells were to halogenated hydrocarbons. During this time, one ad increased in number, particularly in areas of sinusoidal ditional individual underwent liver biopsy because of widening, and were associated with nuclear and cyto persistent hepatic test abnormalities. She had a history plasmic enlargement (Figure 3A). They consisted of: (a) of contact with vinyl chloride used as a propellant in ellipsoid endothelial cells which usually were flat or had hair spray. nuclei, diffusely periodic acid-Schiff-positive cytoplasm Pathological Examination and projected "pillow-like" into the often dilated sinu soids (Figure 3B); (b) Kupffer cells (macrophages) with Observations on humans (7-13, Tamburro, C. H. et periodic acid-Schiff-positive granules; (c) fat storing (Ito) al., Gastroenterology 1979; 77:A43, Abstract) and animals cells with fine vacuoles; (d) occasional fibroblasts with [14-20, Boorman, G. A. et al., Fed. Proc. 1980; 39(Part elongated, almost rectangular nuclei with loose vesicular chromatin patterns and associated with perisinusoidal fibrosis best recognized by silver and trichrome stain (Figures 3C and 4), and (e) few lymphocytes and occa sional plasma cells or segmented leukocytes. Pure hepatic changes were referred to as focal hepa- tocytic hyperplasia (FHH). Sinusoidal cell hyperplasia associated with the hepatocytic changes were referred to as focal mixed hyperplasia (FMH). The liver biopsies were coded and then interpreted blindly by two' of the authors (H. P. and L. M.). The cases with disease were subdivided into those characteristic of nonchemical in^ jury without steatosis, with steatosis alone or steato^B with fibrosis, and into those with evidence of chemic^^ injury. Chemical injury was identified by. (a) FHH with or without focal increased reticulin framework; (b)fFHH with increased reticulin deposition with or with ut sin usoidal dilatation; (c) portal fibrosis with sinusoidal cell Fig. 1. Focal hepatocytk hyperplasia (early stage)- 111-defined focal dysplasia, and (d) hepatic angiosarcoma. Focal hepatic area of enlarged hepatocytea with increased cytoplasm (curved arrows). hyperplasia, with or without focal increases in reticulin H & E, 100x. in the absence of other evidences of hepatocellular dis- FlG 2. Focal hepatocytk hyperplasia (later stage). (A) Focal hyperpU*ia of the hepatocyte* (curved arrows) with variation in cell *iaa well as cell nuclei which ate frequently double, polychromatophilic or can be vacuolated. H4E, lOOx. <B) Slight focal inciaaaei of reticulin framework (arrow*). Silver impregnation, lOOx. CMA 009797 '"Vol. 4, No, 3. 1984 m HEPATIC ALTERATIONS AMONG CHEMICAL WORKERS 415 Fig. 4. Perismutoidal fibrosi*. Perisinusoids! fibrosis in tptce of Dias* surrounding hepatocyte* (arrow*) with numerous fat-storing sinusoidal cells. Trichrome, oiL Histories of work with and exposure to 22 potentially hepatotoxic chemicals were available for all chemical workers. An estimate of total and average exposure to each of the 22 chemicals was made by using a rank order system of estimating exposure based on each worker's occupational history (21, 22). Exposure to each chemical was ranked on a scale of 1 (lowest) to 6 (highest). These rankings were done a priori for each job for each year of the plant's operation based on best estimates from avail able information and industrial expertise. Each worker's cumulative exposure was based on the various job clas sifications beld during employment Thus, separate, av erage, and cumulative exposure indices were developed for each employee. Fig. 3. Focal mi&d hyperplasia. (A) Hyperplati* of timieoidal cell* (curved arrow*) and hepatocytee (itraight arrow*) with sinuaoidal dilatation. H & E, 250x. (B) Sinusoidal cell dysplasia with conspicuous proliferation of polymorph sinusoidal cells (ttraight arrow*) tome of which show bizam nuclei, e*, helmet shaped (curved arrow*). H & E. 400x. (C) Same area showing focal increases in reticulin framework (arrow*) with early sinusoidal dilatation. Silver impregnation, 250x. ease or steatosis, was considered the minimum presump tive evidence of chemical exposure. FMH with increased reticulin and dilatation of sinusoids was considered a more advanced stage. Portal fibrosis with FMH and sinusoidal dilatation was taken as being even more ad vanced. Sinusoidal cell dysplasia and/or evidence of ma lignant transformation, was looked upon as terminal stages of chronic vinyl chloride injury. RESULTS Hepatic lesions consistent with chemical exposure were found in 37% of the exposed workers with hepatic test abnormalities. Among the exposed workers without test abnormalities, 23.5% (3/13) had hepatic lesions con sistent with exposure. In the nonchemical worker com parison group, none with normal test results (0/11) and only one (6%, 1/18) with abnormal liver screening tests had hepatic lesions consistent with chemical exposure. This latter individual had no history of chemical expo sure and/or alcohol consumption, but had undergone surgery for biliary stones. The person biopsied because of persistent abnormalities in hepatic tests had FMH and marked sinusoidal dilatation. This individual was later found to have hepatic angiosarcoma possibly caused by continued use of hair spray which contained vinyl chloride as a propellant. Table 1 lists the type and frequency of hepatic lesions in those individuals with and without liver test abnormalities. Reproducibility Reproducibility of the biopsy readings was evaluated in 17 individuals with 32 biopsies. Seven individuals had only one biopsy (2 needle, 5 wedge); 10 individuals had multiple biopsies (25 total). Biopsies were examined at least twice in a double-blind manner with 3 to 9 months between readings. Biopsy findings from each reading were recorded directly on a semiquantitative computer- 009799 V( s 416 TAMBURRO ET AL. Hepatology Table l. Correlation of Hepatic Biochemical and Histologic Abnormalities in Chemical Workers and Nonoccupationally Exposed Persons Hepetic test result*4 Biopsy diagnosis C heroically Nonworker eipossd (no.) companion (no.) workers group Normal (24) Normal Steatosis (5%) Sinu*oid*l dikuuon HcpatooUular variAiions Portal fibrosis Portal it'lamination FHH FMH (13) 1 1 0 0 0 0 0 (ID 0 0 3 2 3 0 0 Abnormal (54) Noncheznicftl liver injury (22) (17) Steatosis (fcSX) Hspautia Grtnulomu Alcoholic injury Other lesions FHH FMH 11 2 3 2 4 7 2 4 i 3 1 0 2 0 Chemical liver injury FocaI peliotift Angiosarcoma FHH FMH (13) l 0 13 0 * SCOT, SGPT, AP, uul bilirubin tnd prothrombin tun*. 1 1 2 1 (2) Table 2. Comparison or Biopsy Readings Identification no. Needle biopsies* Wedge biopsies* __(1_5_) ______ ________(1_7_) ___ 1st 2nd 3rd 1st 3 A, A 10 A A, A 14 aa 20 AAD 33 AD 26 A 35 A A 36 A A 24 A A 17 A 22 A. A A 24 A, A A 6 A A AA 23 A A A 27 A, A A AD 8 A, A 30 --A, A, A * A, agreement; D, disagreement. biopsy duplicate interpretations were identical. Ninetysix per cent (43/45) of all biopsy readings, whether duplicate or from multiple biopsies, yielded reproducible, consistent interpretations. Technical artifacts (excessive thickness, tom or frag^^ mented specimens) greatly impede correct identificatiom^r Similarly, fatty infiltration and/or chronic disease (hep atitis, granuloma) may obscure the characteristic lesions. We determined, however, the presence or absence of these lesions in six individuals who had a concomitant granulomatous process (four individuals with granuloma were repeatedly read as negative, and two individuals with granuloma were repeatedly read as positive for chemical liver injury). Correlation of Histological Lesions with Exposure From the histological observations, the biopsies of the chemical workers fell into three categories: (i) no histo logical evidence of liver disease (N); (ii) liver disorders inconsistent with chemical injury, (iii) chemical liver injury inferred from histological criteria specified. Indi vidual exposure ratings were not disclosed until after all final histological determinations and categorizations were completed. Comparisons of average and cumulative exposure rat ings of each worker foxvinyl chloride showed that chem ical workers with histological evidence of chemical liver injury had the most persons with highest exposure rat ings to vinyl chloride. Earlier studies showed that all chemical workers who developed hepatic angiosarcoma had an average vinyl chloride exposure rating of 3.5 greater (23). Of the chemical liver injury group, 55% an average vinyl chloride exposure rating of 3.5 greater. Only 22% of chemical workers with liver disease and 32% of exposed workers without evidence of liver disease had ratings of >3.5 (Figure 5). VINYL chloride EXPOSURE 0 * IM MWM AVESAat ized questionnaire which included the 24 histological features under study. The consistent reproducibility of the biopsy readings is shown in Table 2. The consistency in the readings between multiple biop sies in the same individual was very high. In only one case was one of the four biopsies read differently; the wedge was read differently from the needle biopsy. The reproducibility of biopsy readings from multiple needle versus multiple wedge biopsies from a single individual in all readings was consistent. Multiple biopsy readings at different times from the same biopsies showed that all needle-biopsy duplicate readings, and 12 of the 14 wedge- GROUP RATINGS FIG. 5. The high (Hi, *2.5). low (La, <3.5) and outrage C) vinyl chloride exposure ratings among the different histological groups. CLI, group with histological evidence of chemical liver injury; LD, group with histological evidence of liver disease (with or without steatosis) without evidence of chemical injury; SD, standard or "normal" grou| no histological evidence of liveT injury. CMA 009799 ' Vol. 4,No. 3, 1984 HEPATIC ALTERATIONS AMONG CHEMICAL WORKERS 417 Correlation of Biochemical Studies and Histological Findings Both FHH and FMH were never encountered when hepatic test results were normal. As in previous studies, ICG clearance, at 0.5 mg per kg dose and GGT were the most sensitive of the laboratory screening tests in the detection of all types of liver disease, followed by SGOT, SGPT, and AP (Figure 6). The laboratory tests most sensitive in detecting chemical liver injury and having the highest specificity were ICG and AP, while GGT and SGOT were more sensitive in detecting nonchemical liver disease (Figure 6A). Specificity was associated par* ticularly with abnormalities in AP activity and ICG clearance (Figure 6B). GGT was the least specific and most sensitive with a false-positive rate of 20%. The pattern is the same for the frequency with which the abnormalities occurred in these workers. In evaluating nonchemical injury in both exposed and nonexposed persons, significant steatosis as well as hepatitis and granulomas was found to be regularly associated with abnormalities in hepatic test results. DISCUSSION The histologic findings extend the original observa tions made in vinyl chloride-associated hepatic angiosar coma and hepatic injury (1, 5, Tamburro, C. H. et al.. Gastroenterology 1979; 77:A43, Abstract). Focal hepatocytic megalocytosis in the absence of other evidence of acute or chronic lesions seems the earliest detectable change after vinyl monomer exposure. Later, hyperplasia and hypertrophy of sinusoids accompanies the similar A. ALL NON-CHEMICAL CHEMICAL LIVER DISEASE LIVER DISEASE LIVER DISEASE a LIVER BIOPSIED WORKERS FlC. 6. The occurrence of biochemical screening test abnormalities in various histological groups with liver injury. (A) Sensitivity (percentage of biopsied worker* correctly identified a* having liver disease) of laboratory screening tests in all types of liver disease, chemical and nonchemical liver injury. (B) Specificity (percentage of biopsied work ers correctly identified a* not having liver disease) of laboratory screen ing testa in liver biopsied workers. AST, SGOT; ALT, SGPT focal hepatocytic process which now assumes a poorly defined shape and is associated with perisinusoidal fibro sis and proliferation of fibroblasts and/or their precursor (Ito) cells. The sinusoidal cell activation involves macrophagic and endothelial-lining cells as well as focal sinusoidal enlargement. Eventually the mixed hyperpla sia (hepatic and sinusoidal cells) is followed by sinusoidal cell dysplasia, palisading of sinusoidal-lining cells and, finally, by malignancy as seen in both man and experi mental animals (6, 7, 8,18). Our data, supplemented by individual exposure his tory, demonstrates then that FHH is the earliest consis tently identifiable histological lesion in workers exposed to vinyl monomer chemicals. It is relatively rare in nonchemical workers without liver disease but resembles focal regenerative hyperplasia (24) sometimes accompa nied by portal hypertension. By contrast FMH, being a more advanced lesion, was only found in chemically exposed workers and in a person with a history of expo sure (hair spray). This supports the concept that these lesions are related to prolonged or repeated chemical exposure. Whether the early lesions (FHH) may be precursors in the development of angiosarcoma or only evidence of cellular adaptation without progression requires follow up. However, the consistency of the results of the hepatic screening tests over a 7-year follow-up period and the absence of changesin the histological finding on serial biopsy of these workers suggest that FHH is not pro gressive when the worker is removed from exposure or exposure is very low (25). By contrast, FMH.'especially sinusoidal cell dysplasia, may be associated with devel opment of angiosarcoma as illustrated in one of our cases; quite likely, it is a nonreversible lesion. Diagnosis of these histological lesions in both needle and wedge liver biopsy is highly reproducible. Inconsis tency appeared more often among the duplicate readings of wedge biopsy specimens and between wedge and needle biopsy specimens from the same individual. This did not occur between duplicate readings of needle biopsies nor in reading of multiple needle or multiple wedge biopsies. Silver impregnation for assessment of the reticulin framework is useful in verifying FMH. The finding of early histologic lesions only in those individuals with hepatic test abnormalities gives objective support to the usefulness of hepatic tests to indicate liver injury. How ever, as in previous studies (26-28) GGT has proven to be excessively sensitive as a screening test for liver injury. Abnormalities of AP activity provide especially good correlation with the presence of subclinical chemical liver disease but lack the sensitivity needed for a primary screening test. ICG clearance provides both the sensitiv ity and specificity needed for primary screening and/or confirmatory study for those workers with elevated ac tivities of the commonly used hepatic enzymes. This, along with AP activity provide sufficient specificity for identifying those workers at high probability of chemical liver injury calling for confirmation by liver biopsy to reveal the specific lesion(s). The type of histological lesions may account for these biochemical findings in that the cellular hyperplasia and sinusoidal dilatation and perisinusoidal fibrosis interfere particularly with CMA 009800 418 hepatic microcirculation (29-31). This impairment is reflected in diminished ICG and less so by elevated serum activities of the intracellular enzymes, such as the ami notransferases whose release better mirrors hepatocel lular dysfunction. These correlations strongly suggest a need to change the presently required federal screening for chemical exposure workers. Acknowledgments: We wish to thank Drs. Seymour H. Hutner and Kathleen M. O'Connell for their manuscript review, and Vicky Strong for help in preparing the manuscript. REFERENCES 1. Thomas LB, Popper H. 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Environmental Perspectives 1981; 41:117-122. 28. Tamburro CH. Chemical hepatitis, pathogenesis, detection end management. Med Clin N Amer 1979; 63:545-566. 29. Lange CE, Juhe S, Steine G, et aL So-called vinyl chloride disease. Is it an occupational systemic sclerosis? Int Arch Arbeitsmed 1974; 32:1-32. 30. Kurokawa S, Inagaki T, Okyama S. Electron microscopic obser vation of the liver in portal hypertension following chronic expo sure to vinyl monomer. Gastroenterol Jpn 1977; 12:117-124. 31. Marsteller HJ, Lelbach WK. Muller R, et al. Chroniachtoxiscbe leberschaeden bei arbeitern in der PVC-production. Dtsch Med Wochenachr 1973; 98:2311-2314. CMA 00980