Document EvqeMEO697zEJBVodbbKBZwkn

Seminar in Occupational Medicine Mutagenicity of Vinyl Chloride External Chromosome Studies on Persons With and Without VC Illness, and on VC Exposed Animals I. Fleig, Dr. rer. nat., and A. M. Thiess, Dr. Med. Review of the Literature According to our knowledge, nine studies3 '0 13 relating to vinyl chloride (VC) problems have been undertaken. However, no conformity could be reached from these results. As Table 1 shows, five studies had a higher rate of chromosome aberration in the VC exposed group than in the control group. On the other hand, four further studies produced negative results. All those investigations made on animal and sub-mammalian species' "''4 are shown in Fig 1. Personal Investigations Investigations on persons concerned with the manufacture of VC/PVC with estimated exposure have been undertaken in our From the Department for Occupational Medicine and Health Protection, BASF Aktiengesellschaft, 6700 ludwigshafen W. Germany ^Corporate Medical Director Professor Dr Med, A M Thiess) Paper presented at the Second international Conference on Environmental Mutagens, July 11-15, 1977, in Edinburgh medical department.5 In this group we cannot make any definite statements regarding the rate of exposure, but it is most probable that the assumed values taken from abroad are also applicable to the Federal Republic of Germany.15 These values are: 1945-1950 -- 1,000 ppm: 1950-1960 -- 400-500 ppm; 1960-1970 -- 200400 ppm: and 1971-1973 -- below 150 ppm. In addition, investigations were undertaken on persons in the VC/PVC processing plants who have undergone monitored VC exposure (Fig 2).5 Both groups were combined as "Exposed per sons showing no symptoms of VC illness" (Table 3 and Fig 3'. Before commencing the study, anamnestic data were collected regarding sex, age. occupational exposure(sl, medical x-ray treat ment either for diagnostic or therapeutic purposes, recent viral diseases and vaccinations, consumption of drugs, and smoking habits, m order to discard those subjects who have multiple ex posures. We matched these subjects with healthy controls of the same sex (in this case male) and approximately the same age who Authon Fleig. Thiess (1974) Ducatman Nirschhofn Selikoff (1975) Funes-CraviotO et al (1975) Hansteen Hillestad Thns Evenson (1975) Kilian, Picciano (1975) Lange. Schwinger Veltman 11975) Purchase Richardson, Anderson <19751 fleig (1977) Leonard et al (1977) Table 1. -- A Survey of Results from VC Exposed Persons. No. of Protends 10 workers 11 workers 10 controls 7 workers 3 controls 39 workers 16 controls 203 workers ]08 controls 12 workers Ltngth of Exposure (tun) 426 428 9-29 -- Up to 30 32521 56 workers 24 controls 20 workers 20 controls -- 4 30 18 workers 10 controls 2-17 % Chromosome Aberration* VC Exposed Controls (Average) 75 55 Mutagenic Effect Negative 9 82 9 52 64 194 Positive Positive Pathological Results (VC Syndrome) No pathological results -- 1 thrombocytopenia 341 39 -- 8 13 112 1 9 resp 2S 1 79 55 -- 4 IS 55 26 Positive Negative -- __ Negative Thrombocytopenia sc^enomegalv acroosteolysis Raynaud s syndrome scleroderma Positive Positive More severe chromosome anomalies -- due to radiograph1 -- Thrombocytopenia sotenomegaly sclerodermia Raynajd s syndrome liver hbrosis esopragus and stomach varices of the fundus acroosteolysis -- Journal of Occupational Medicine/Vol. 20. No. 8/August 1978 ucc 110882 557 Author* Malawi* Montaaano 0975) flannuQ Johtnnoon Wachtmwrtw 0974) LA0VWOO M Ai (1978) M*Qmi*nn tarn* 09751 PklKMM Wchamaon Andanon (19751 Hwg 0977) T*t - Method t! AME5-Tst TA 1530, TA 1535, 0 46 b| AMS`T*t TA 1536 TA1S37. TA 1539 a) AMES-T*t TA 1535 b) AM6S-T**t TA 1535 TA 1537. TA 1635 1 AME$-Tt Schizostccharofflyces pom0 b> AMES'Tost Saccharomyca* cerv<*tM o host-mechated-assay (Moo*4) MUUR-5-M4thod (Drovophita) DOMINANT-LETHAL-T4*1 (M0U44) BONE MARROW (Hamit*) EipOMFI .000, 20.000 ond 200 000 ppm h nd 48 h 200,000 ppm 30. 60. 90 Mm* 4) 3.000 pom b) 3,000 ppm cj 700 mg/fcQ oral not known 3.000.10.000 and 30,000 ppm 2,500. 5,000 ppm Mutapantc EttPCt t) potitm b) n*o*ttv* a) positm b) napatn* | POSH** b> powtiva O po*mv* pO*rtrv4 n*QtTV* P0*ftiv4 AbtTM^n Typt Pott* mutMiOn* Potnt mutations Powtt mutation* Racvhm latnaj mutations No dotwiant tathal mutation* Chranoaoma ibonotwnt were not exposed to VC or to any other known chromosome damaging agent but worked in the same factory in different departments (e.g., clerical staff from the sales or technical department). The same questionnaire used for obtaining data on experimental subjects was also used for controls, and the same criteria for exclu sion from the study was applied. Lymphocyte cultures were prepared from both groups and finally processed using the so-called "micro-method " One hundred metaphases per person were in vestigated, making a total of 3,000, which were analyzed for structural chromosome anomalies. The frequency of aberrant metaphases totaled 3.1% excluding gaps and 7,5% including gaps. The correspond Fig 1. -- A survey of the results using VC in various tests. ing values in the control group were 2.1% and 5.5%, respectively. The individual datas Recording Point Ruid Mixer Ruid Mixer Exit Mixing-mill Mixing-mill directly above the nip Colander feeding point Colander take-off VC content in ppm <i <i <i 12 <1 <1 Remarks At headlevel in the position occupied by the operator At headlevel in the position occupied by the operator (after the mixing process.) At headlevel in the position occupied by the operator Position not occupied by personnel At headlevel in the position occupied by the operator At headlevel in the position occupied by the operator are depicted in Tables 2 and 3. There was no significant difference in the rate of chromosome aberrations compared with the control group (Fig 3). Investigations were also made on workers not employed in the BASF but with VC symptoms after an unknown rate of ex posure. Within the framework of a re search program from the insurance associa tion of the chemical industry and the Verband Kunststofferzeugende Industrie (VKE), 20 workers all showing symptoms of VC ill ness (Fig 4) were studied. Many of the workers examined showed signs of more Extruder jet Extruder feed-hopper < 1-4 < 1-28 Only temporarily occupied by the operator Only temporarily occupied by the operator than one symptom of VC illness. The statis tical evaluation revealed a significant differ ence between the rate of chromosome Fig 2. -- Recordings of VC concentrations in the atmosphere In the region of the KAE Technical Department. aberrations in these persons and those in the control group. The frequency of aberrant metaphases in the exposed group was 5.2% excluding Table 2. -- Chromosome Aberration Rate of Controls. gaps and 11.2% including gaps, compared with 2.1% and 5.5%, respectively, in the control group (Fig 5). The control group was Analyzed % Abttrtnt Matapha*#* No. Aft Uatapha** Iftcl. Gaps E*d. Gap* 18 42 100 7 3 22 50 100 3 26 33 100 -- 29 39 100 6 2 132 37 100 -- 149 56 100 7 2 153 46 100 -- 206 54 100 3 1 207 44 100 1 208 64 100 9 7 209 37 100 3 2)0 62 1DO 9 4 211 38 100 2 212 35 100 5 l 213 57 100 2 1 214 42 100 9 4 216 60 100 2 217 33 100 4 2 218 52 100 3 t 219 50 100 2 the same one mentioned earlier. The individual datas are depicted in Tables 2 and 4. External investigations were done on a patient with an angiosar coma due to VC exposure. According to information received Table 3. -- Chromotome Aberration Rate of Exposed Person! Showing No Symptoms of VC fllneia. Duration of Expoaura Anatyzad % Abamnt Mataphssat No. H* (Y**r*l Mataphatai loci. Gaps Exel. Gaps 5 51 11 100 6 39 7 100 8 14 57 20 100 10 76 52 25 100 9 82 43 15 100 83 52 84 54 20 29 100 100 8 6 86 46 4 100 8 87 55 26 100 8 101 34 6 100 4 1 2 2 3 3 5 4 4 5 2 ucc558 110883 Mutagenicity of Vinyl Chloride/Fleig and Thiess 10 - Control mci. gaps Control excl gaps Exposed tncl. gaps Exposed excl. gaps Fig 3. -- Chromosome aberration rate of exposed workers showing no symptoms of VC illness. from the Berufsgenossenschaft, 11 cases from a total of 57 cases of hemangioendothelial sarcoma, attributable to VC/PVC exposure, were discovered in the Federal Republic of Germany (Table 5), One of these 11 persons who is still alive, aged 43. was occupied in the PVC manufacturing area for 13 years. The rate of VC expo sure to which he personally was exposed during the last years was between 35 to 150 ppm..He was occupied for 20 to 30 hours per month manually cleaning the autoclaves. In addition, he some times took samples from the decontamination plant to the labora tories for testing. An analysis of 200 metaphases produced a rate of aberrant cells of 7.3% excluding gaps and isogaps, a significant increase com pared with a control group which had a rate of 2.1%. The rate of aberrant metaphases including gaps and isogaps was 16.6% com pared to the control group, with a rate of 5.5%. 10 - ma. > -co CO mmma 5 jafzfl 11.2 Control inci. gaps Control excl. gaps Exposed incl. gaps Exposed excl. gaps .g0<o>0 & Fig 5. -- Chromosome aberration rate of exposed workers showing symptoms of VC illness. Journal of Occupational Medicine/Vol 20. No, 8/August 1978 Ref Age Exposure Symptoms Nr in years % aberrant metaph Exchanges ind. gaps excl gaps 90 56 6 Uver fibrosis. oesophagus vane**, thrombocytopenia, splenomegaly 10 6 2 91 54 11 Liver fibrosis, portal hyper tension. thrombocytopenia, splenomegaly, oesophagus vances 11 6 1 119 33 7 Uver parenchyma damage, thrombocytopenia, splenomegaly 7 2- 1S8 37 5 Sclerodermia. liver fibrosis, Ray 7 4 1 naud* syndrome, acroosteolysi*. splenomegaly, thrombocytopenia 180 32 161 42 6 Uver parenchyma damage, thrombocytopenia 15 Thrombocytopenia, liver parenchyma damage 13 7 1 TO 5 f 162 60 30 Uver parenchyma damage. Raynauds syndrome, thrombo cytopenia, oesophagus vances 11 5 184 53 19 Uver fibrosis, thrombocytopenia 9 4 1 185 64 22 Uver fibrosis, thrombocytopenia, splenomegaly 11 42 167 52 26 Uver parenchyma damage, thrombocytopenia 20 9 1 170 34 4 Uver parenchyma damage, 931 thrombocytopenia, splenomegaly 171 44 9 Uver fibrosis. Raynaud* syndrome, acroosteolysis 11 5 1 172 52 5 Uver parenchyma damage, Raynauds syndrome, thrombocytopenia 8 31 173 41 5 Uver parenchyma damage, splenomegaly 8 31 174 62 10 Uver parenchyma damage, Raynauds syndrome 22 10 2 176 41 16 Liver fibrosis, thrombocytopenia, 14 6 1 oesophagus varices 177 36 11 Thrombocytopenia, hepatomegaly 18 8 2 183 49 10 Uver parenchyma damage, splenomegaly 7 5" 199 52 12 Uver parenchyma damage 8 51 203 42 8 Uver fibrosis, splenomegaly 11 52 Fig 4. -- VC symptoms and rate of chromosome aberration in the exposed probands. Apart from our investigations on human beings, tests were also performed using bone marrow of Chinese hamsters. Two types of tests were undertaken: (1) inhalation tests with dosages of 2,500 ppm and 5.000 ppm, respectively, for four hours a day, 24 hours apart over a five-day period, and (2) intraperitoneal injections of 300 mg/kg and 600 mg/kg, respectively, given in five injections, 24 hours apart, over a five-day period. Table 6 shows a survey of all important data concerning the arrangement of our animal in vestigations. Both tests (Figs 6 and 7) showed that the frequency of structural anomalies, inclusive and exclusive of gaps in the bone marrow cells, was significantly higher compared to the control group.4 Discussion The mutagenic effect of vinyl chloride mentioned in various sources of literature, bacterially tested, investigated on mammals, sub-mammals and human beings, has been confirmed by our in vestigations on mammals (2,500 ppm and 300 mg/kg) and man (5000 ppm and 600 mg/kg). The most important point to mention is that an increased rate of chromosome aberration could be found only in those persons showing symptoms of VC illness, as well as in an angiosarcoma case. One very important fact must, however, be taken into ac count. Nine months before blood samples were taken (February 1, 1977). the patient with the angiosarcoma was given polychemotherapy which was comprised of applications of 150 mg Adriblastin, 120 mg Vincristin, 3,000 mg Endoxan, and 6,000 ucc 110884 559 Table 4. -- Chromosome Aberration Rate of Exposed Persons Showing Symptoms of VC Illness. Duration of Exposure Anilynd % Aberrant Metaphases No. Age (Ymis) Metaphases (net. Gaps Exel. Gap* 90 56 6 100 10 91 54 u 100 11 119 33 7 100 7 158 37 5 100 7 160 32 6 100 13 161 42 15 too 10 162 60 30 100 11 164 53 19 100 9 165 64 22 100 11 167 52 26 100 20 170 34 4 100 9 171 44 9 100 U 172 52 5 100 8 173 41 5 100 8 174 62 21 100 22 176 41 16 100 14 177 38 11 100 18 183 49 10 100 7 199 52 12 100 8 203 42 8 100 11 6 6 2 4 7 5 5 4 4 9 3 5 3 3 10 6 8 5 5 5 mg DTIC (Dacarbazin). It is, therefore, not possible to say whether or not the results we have obtained are solely due to the patient's exposure to VC or whether his chemotherapeutical treatment, together with the exposure rate of VC, affected our results.2 Summary Chromosome analysis was undertaken on lymphocyte cultures taken from (1) six workers with estimated exposure to VC and four workers with monitored exposure to VC (employed in the BASF); (2) 20 workers showing symptoms of VC illness with unknown exposure and one angiosarcoma case, due to VC ex posure (not employed in the BASF); and, (3) on bone marrow cells of Chinese hamsters after inhalation of 2,500 ppm or 5,000 ppm, Tabta 6. -- T*t Proc*dure. Animal species Animal's age Animal's weight Substance Tt 1 Type of administration Interval of administration Dosage Time until sacrificed after last administration Test 2 Type of administration Solvent Interval of administration Quantity administered Dosage Time until sacrificed after last administration Chinese hamsters 10-14 weeks 26-30 g Vinyl chloride Inhalation 24 hours 2.500 ppm/5 days/4 hears 5,000 ppm/5 days/4 hours 6 hours i p injection Olive oil 24 hours 5 x 05 ml 300 mg/Kg body weight 600 mg/kg body weight 6 hours or after intraperitoneal injections of 300 mg/kg or 600 mg/kg. The proband groups showing symptoms of VC illness, the only living angiosarcoma case, and the animal test show a significant increase in the rate of aberrations in comparison to the control group. References 1. Bartsch H, Malaveille, and Montesano R: Human, rat and mouse liver mediated mutagenicity of vinyl chloride in S. typhimunum strains. Int I Cancer 15:429-437, 1975. 2. Bauchinger M and Schmid E: Cytogenetische Veranderungen in weiben Blutzellen nach Cyclophosphamidtherapie. Z Krebsforsch 72:77-87. 1969. 3. Ducatman A, Hirschhorn K, and Selikoff I]: Vinyl chloride exposure and human chromosome aberrations. Mutat Res 31:163-168, 1975. 4. Fleig I: Zur Mutagent&'t von VC -- Untersuchungen an Sauger und Mensch. (Thesis). Heidelburg University, Federal Republic of Germany, 1977. 5. Fleig I and Thiess AM: Chromosomen-Untersuchungen bei Vinylcblorid-Exposition. ASP 9(12):280-283. 1974. 560 Rtf. No. 1 2 3 4 5 6 7 8 9 10 11 T*bl 5. -- Survty of Angiourcomit of th Livor Duo to VC Exposira in th* Fodtnl Rtpublle of Gtnrany. Plant 4 4 12 8 11 7 8 4 4 2 2 Occupation PVC production autociaver PVC production autociaver VC application. Spray filler PVC production chemical worker PVC research dept lab assistant PVC production shift foreman Technical appl autoclave cleaner PVC production PVC production/ processing PVC production polymerization VC processing Date of Birth Q6/04/3Q 07/26/31 07/16/30 09/04/30 05/09/36 01/01/32 09/29/26 01/19/17 08/31/07 12/13/34 12/29/36 Otend 01/25/69 12/14/71 10/10/73 11/25/74 12/16/74 01/10/75 11/13/75 12/25/75 10/20/76 Alive 03/77 A* M DtafWtlt 38 39 43 44 38 43 49 58 -- 41 40 Rwtod of Etpuun (Ymk) 12 UtMicy Ptriod (Yun) 1/2 11 2 15 1/2 17 -- 59 12 1/2 2M/2 -- 201/3 9-2/3 M/2 5-1/3 14-1/4 10 until 1971 Alive 51/2 Mutagenicity of Vinyl Chloride/Fleig and Thiess UCC - 110885 10 - a <o E -f4Uc> Control incl gaps Control excl gaps 10 - Fig 7. -- Chromotome aberration rate of Chlnete hamtteri after being intraperitoneally injected with 300 mgAg and 600 mgAg VC. 60 Fig 6. -- Chromosome aberration rata of Chinese hamsters after Inhalation of 2500 and 5000 ppm VC, respectively. Control incl gaps Control excl gaps 10 3 Exposed incl gaps Exposed excl gaps 6. Funes-Cravioto F, Lambert B, Lindsten J, et al: Chromosome aberrations in workers exposed to vinyl chloride. Lancet 1:459. 1975. 7. Hansteen IL, Hillestad L. and Thtis-Evensen E: Chromosome studies in workers exposed to vinyl chloride. Fifth Annual Meeting, EEMS. Florence. 1975. 8. Kilian D| and Picciano Dl: Cytogenetic monitoring of vinyl chloride workmen. Fourteenth Annual Som. Cell Genetics Conference Houston, Texas, 1975, 9. Lange CE. Schwinger E. and Veltman G: Zur Frage der Chromosomenveranderungen bei Patienten mit Vinvlt hlond Krankheit Dtsch. Ges. f Arbeitsmed., Munchen 1975. 10. Leonard A, Decat C. Leonard ED, et al: Cytogenetic investigations on lymphocytes from workers exposed to vinyl chloride. I Toxicol Environ Health 2:1135-1141, 1977 11. Loprieno N, Abbondandolo A, Barale R, et al: Evaluation of the genetic effects by vinyl chloride monomer (VCM) under mammmalian metabolic activation: Studies in vitro and in vivo, \futaf Res 40 85-96, 1976. 12. Magnusson | and Ramel C: Mutagenic effects of vinyl chloride in drosophila melanogaster. Mufar Res 38 115, 1976. 13. Purchase IFH, Richardson CR, and Anderson D: Chromosomal and dominant lethal effects of vinyl chloride. Lancet 2:410-411. 1975 14. Rannug U. lohannson A, Ramel C. and Wachtmeister CA: The mutagenicity of vinyl chloride after metabolic activation Amhio 3 194-197, 1974. 15. Thiess AM and Frentzel-Beyme R: Retrospective survey of the alleged dseases associated with vinyl chloride in the Federal Republic of Germany I Occup Med 17:430-432. 1975. Journal of Occupational Medicine/Vol. 20, No. 8/August 1978 UCC - 110886 561