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Sensitivity of Down's syndrome lymphocytes to mitomycin C and X-irradiation measured by sister chromatid exchange frequency
Sensitivity of Down's Syndrome Lymphocytes to Mitomycin C and XIrradiation Measured by Sister Chromatid Exchange Frequency Peter E. Crossen and William F. Morgan
ABSTRACT: The frequency of sister chromatid exchanges {SCE) was studied in lymphocytes from Down's syndrome patients and normal donors after in vitro exposure to mitomycin C(0.01 ~g/ml) and X-irradiation. There was no difference in the response of Dawn's syndrome cells and normals following treatment with mitomycin C. However, Dawn's syndrome patients shewed a significant increase in SCE following exposure ta both 50 and 100 reds, whereas normal donors showed an elevated SCE rate only after exposure to 100 rads.
INTRODUCTION Down's s y n d r o m e (trisomy 21) is a congenital disorder associated with an increased i n c i d e n c e of l e u k e m i a [1,2]. Chromosomes from Down's s y n d r o m e patients exhibit an increased frequency of aberrations following X-irradiation [3,41, viral infection [5], and treatment with chemical mutagens [6,7]. Increased c h r o m o s o m e fragility in response to X-rays and chemical mutagens as well as an increased i n c i d e n c e of malignancy is also found in ataxia telangiectasia, F a n c o n i ' s anemia, and Bloom's synd r o m e and has lead to the suggestion of a possible association between c h r o m o s o m e fragility and m a l i g n a n c y [8]. Although the precise m e c h a n i s m s of sister c h r o m a t i d exchange (SCE) are not understood, the technique offers a means of investigating the possible relationship between c h r o m o s o m e aberrations and DNA repair processes. Ataxia telangiectasia, Fanconi's anemia, and Down's s y n d r o m e all have normal SCE in cultured l y m p h o cytes [9 - 11], whereas Bloom's s y n d r o m e cells show a 12-fold increase in SCE [12]. To gain further information about the cellular defects in Down's syndrome, we have s t u d i e d the i n c i d e n c e of SCE in Down's s y n d r o m e l y m p h o c y t e s following challenge with m i t o m y c i n C (MMC) and X-irradiation. MATERIALS AND METHODS
Peripheral blood l y m p h o c y t e s were isolated from defibrinated blood by gelatin sedim e n t a t i o n and cultured in H a m ' s FIO m e d i u m containing p h y t o h e m a g g l u t i n i n (PHA) 12.5 ~ l / m l (Burroughs Wellcome). B r o m o d e o x y u r i d i n e (BrdUrd) was a d d e d at From the CytogeneticsUnit, Cancer Society of New Zealand, Christchurch Hospital, Christchurch, New Zealand. Address requests for reprints to Dr P.E. Crossen, Cytogenetics Unit, Christchurch Hospital. Christchurch, New Zealand. Received February 15, 1980; accepted May 8, 1980. 281
Copyright O Elsevier North Holland, Inc., 1980 52 Vanderbilt Ave., New York, NY 1 0 0 1 7
Cancer Genetics and Cytogenetics2, 281-285 (1980) 0165-4608/80/05028105502.25
282
P . E . Crossen and W. F. Morgan Table 1
SCE in Down's s y n d r o m e and normal l y m p h o c y t e s following treatment with MMC a
Down's Syndrome
Normal
Baseline
MMC
× increase
Baseline
MMC
× increase
10.74 (8)
22.88
2.16
10.78 (20)
23.86
2.35
~Each entry is the mean number of SCE from the number of donors (in parentheses} studied.
a final concentration of 10.0 ~g/ml after 24 hr and cultures kept in a lightproof box to prevent photolysis of the BrdUrd-substituted DNA [13]. Colcemid (Ciba Laboratories) was a d d e d 4 hr prior to harvest at a final concentration of 0.1 ~g/ml and c h r o m o s o m e preparations made by the flame drying technique of Scherz [14]. Slides were treated for sister chromatid differential staining by a slightly m o d i f i e d version of the fluorescence plus Giemsa (FPG) technique of Perry and Wolff [15]. All slides were c o d e d and the incidence of SCE scored in 20 well spread metaphases from each patient. MMC was a d d e d to cultures at the same time as the BrdUrd (24 hr} at a final concentration of 0.01 ~g/ml. Cultures for X-ray treatment were grown in Falcon T30 flasks and X-rays a d m i n i s t e r e d by a Phillips RT 100 X-ray m a c h i n e (100 kV; 8 m A using a 0.4 m m c o p p e r filter). Cultures were irradiated at 44 hr with doses of 50 and 100 rads. RESULTS There was no difference between baseline SCE frequency in Down's s y n d r o m e patients (10.74) and normals (10.57). Although slightly higher than w e have p r e v i o u s l y reported [16,17], a baseline SCE frequency of a r o u n d 10 is in good agreement with our later studies [18]. Exposure to MMC caused an a p p r o x i m a t e doubling of SCEs in both the Down's s y n d r o m e patients and normal controls (Table 1). Fifty fads X-irradiation caused a significant increase in SCE in cultures from Down's s y n d r o m e patients (t -- 3.2, 3 degrees of freedom, p < 0.05), but not in norreals (Table 2). Both normals and Down's s y n d r o m e patients showed a significant increase in SCE following 100 rads (Table 3}. DISCUSSION The hypersensitivity of Down's s y n d r o m e l y m p h o c y t e s to X-irradiation is well established in studies showing increased c h r o m o s o m e aberrations and m i c r o n u c l e i formation [3,4]. Although radiation is a c o m p a r a t i v e l y poor i n d u c e r of SCEs [19,20], we
Table 2
SCE in Down's s y n d r o m e l y m p h o c y t e s following 50 rads X-irradiation.
Normal donors (6) Baseline 50 rads x increase over baseline
Down's syndrome patients I II III IV 10.33 ~ t0.60 10.75 a 11.81 1.04 1.11
9.30 13.95 1.50
9.60 16.75 1.74
aMean SCE rate of the number of donors (in parentheses) studied.
12.30 15.35 1.24
Incidence of SCE in D o w n ' s S y n d r o m e Lymphocytes Table 3
283
SCE in Down's s y n d r o m e l y m p h o c y t e s following 100 rads X-irradiation Down's Syndrome Patients V VI VII VIII
Normal donors (8) Baseline 100 fads x increase over baseline
10.61 a 14.61 a 1.37
11.90 14.00 1.17
11.55 17.70 1.53
14.00 18.30 1.30
10.75 13.45 1.25
~Mean SCE rate of the number of donors (in parentheses) studied.
found that 50 reds of X-irradiation significantly increased the SCE rate in Down's s y n d r o m e cells, but not in normals. It is u n l i k e l y that the increase in SCE following 50 rads is due to differential BrdUrd incorporation b e t w e e n Down's s y n d r o m e and normal ceils. Not only are baseline SCE frequencies almost identical, but increasing the BrdUrd concentration does not result in an increase in X-ray i n d u c e d SCEs [21]. A l t h o u g h the precise DNA lesion leading to SCE formation is at present u n k n o w n , it appears that lesions resulting in SCE formation are i n d u c e d in Down's s y n d r o m e ceils at very m u c h lower doses of X-irradiation than in normal cells. These results are in contrast to the situation in ataxia telangiectasia cells, w h i c h are also more sensitive to X-irradiation, but do not show an increase in SCE at the X-irradiation doses s h o w n [22]. X-ray doses of 100 rads caused a significant increase in the n u m b e r of SCE both in normal and Down's s y n d r o m e ceils. However, Ku~erov~ and Polikov~i [23] found no increase in SCE in either normal or trisomic cells exposed to 100 rads. The reason for this difference is not clear, but m a y reflect the time of irradiation, as there is evidence that irradiation of G o l y m p h o c y t e s does not increase SCE [21,24]. W h i l e Down's s y n d r o m e cells show an abnormal response to X-irradiation, our studies and those of Ku~erov~ and Polikova [23] show that they have a normal response to challenge by MMC. MMC is a bifunctional aklylating agent that can also i n d u c e cross-links in DNA [25]. It w o u l d a p p e a r that MMC induces a similar n u m b e r of lesions in both Down's s y n d r o m e and normal cells, and that they are repaired efficiently. X-ray i n d u c e d c h r o m o s o m e breaks are repaired more r a p i d l y in Down's s y n d r o m e cells [4], while the uptake of 3H-thymidine following ultraviolet light exposure is r e d u c e d [26], suggesting an a b n o r m a l i t y in DNA repair m e c h a n i s m s in Down's syndrome. Although the relationship between SCE and DNA repair is unclear, Evans [27] has suggested that SCE m a y represent a general error correcting process of high fidelity, e n c o m p a s s i n g a w i d e variety of DNA damage. Our data indicate that SCE functions n o r m a l l y in Down's syndrome, correcting both X-ray- and M M C - i n d u c e d lesions. This work was supported by the Central Districts, the Canterbury and West]and Divisions of the Cancer Society of New Zealand, and the Medical Research Council of New Zealand. The authors wish to thank Dr. P.H. Fitzgerald for helpful suggestions and Mrs. J. Stewart for assistance with the statistical analysis.
REFERENCES 1. Holland WW, Doll R. Carter CO (1962): The mortality from leukaemia and other cancers among patients with Down's syndrome (Mongolism) and among their parents. Br J Cancer, 16, 177-186.
284
P.E. Crossen and W. F. Morgan 2. Jackson EW, Turner JH, Klauber MR, Norris FD (1968): Down's syndrome: variation of leukemia in institutionalized populations. J Chronic Dis 2 1 , 2 4 7 - 253. 3. Sasaki MS, Tonomura A (1969): Chromosomal radiosansitivity in Down's syndrome. Jap J Hum Genet 14, 8 1 - 9 2 . 4. Countryman PI, Heddle JA, Crawford E (1977): The repair of X-ray induced chromosomal damage in trisomy 21 and normal diploid lymphocytes. Cancer Res 37, 5 2 - 58. 5. Higurashi M, Tamura T, Nakatake T (1973): Cytogenetic observations in cultured lymphocytes from patients with Down's syndrome and measles. Pediatr Res 7,582 - 587. 6. O'Brien RL, Pooh P, Kline E, Parker JW (1971): Susceptibility of chromosomes from patients with Down's syndrome to 7,12-dimethylbenz(a) anthracene induced aberrations in vitro. Int J Cancer 8,202 - 210. 7. Kaina B, Waller H, Waller M, Rieger R (1977): The action of N-methyl-N-nitrosourea on nonestablished cell lines in vitro. I. Cell cycle inhibition and aberration induction in diploid and Down's fibroblasts. Murat Res 43,387 -400. 8. German J (1972): Genes which increase chromosome instability in somatic cells and predispose to cancer. Prng Med Genet 8, 61 - 101. 9. Galloway SM, Evans HJ (1975): Sister chromatid exchanges in h u m a n chromosomes from normal individuals and patients with ataxia telangiectasia. Cytogenet Cell Genet 15, 17 - 29. 10. Latt SA, Stetten G, Juergens LA, Buchanan GR, Gerald PS (1975): Induction by alkylating agents of sister chromatid exchanges and chromatid breaks in Fanconi's anemia. Proc Natl Acad Sci USA 72, 4066-4070. 11. Lezana EA, Bianchi NO, Bianchi MS, Sabala-Suarez JE (1977): Sister chromatid exchanges in Down syndromes and normal h u m a n beings. Mutat Res 45, 85 - 90. 12. Chaganti RSK, Schonberg S, German J (1974): A many fold increase in sister chromatid exchanges in Bloom's syndrome lymphocytes. Proc Natl Acad Sci USA 71, 4508-4512. Ikushima T, Wolff S (1974): Sister chromatid exchanges induced by light flashes to 5-bro13. modeoxyuridine and 5-iododeoxyuridine substituted Chinese hamster chromosomes. Exp Cell Res 87, 15 - 19. 14. Scherz RG (1962): Blaze drying by igniting the fixative for improved spreads of chromosomes in leucocytes. Stain Technol 37,386. 15. Perry P, Wolff S (1974): New Giemsa method for the differential staining of sister chromarids. Nature 251,156-158. 16. Crossen PE, Drets ME, Arrighi FE, Johnston DA (1977): Analysis of the frequency and distribution of sister chromatid exchanges in cultured h u m a n lymphocytes. Hum Genet 35, 345-352. 17. Morgan WF, Crossen PE (1977): The incidence of sister chromatid exchanges in cultured h u m a n lymphocytes. Mutat Res 42,305 - 312. 18. Crossen PE, Morgan WF (1980): Sister chromatid exchange in cigarette smokers. Hum Genet 53,425 -426. 19. Perry P, Evans HJ (1975): Cytological detection of mutagen-carcinogen exposure by sister chromatid exchange. Nature 258, 121 - 125. 20. Crossen PE, Morgan WF (1979): The effects of ~ radiation on sister chromatid exchanges in cultured h u m a n lymphocytes. Mutat Res 62,125 - 129. 21. Morgan WF, Crossen PE (in press): X-irradiation and sister chromatid exchange in cultured h u m a n lymphocytes. Envirnon Mutagen. 22. Galloway SM (1977): Ataxia telangiectasia: The effects of chemical mutagens and X-rays on sister chromatid exchanges in blood lymphocytes. Mutat Res 45,343 - 349. 23. Ku~erov~ M, Polikova Z (1978): In vitro comparison of normal and trisomic cell sensitivity to physical and chemical mutagens. In: Mutagen Induced Chromosome Damage in Man, HJ Evans, DC Lloyd, eds. Edinburgh University Press, Edinburgh, pp. 185 - 190. 24. Littlefield LG, Colyer SP, Joiner EE, Dufrain RF (1979): Sister chromatid exchanges in human lymphocytes exposed to ionizing radiation during G 0. Radiat Res 78, 514 - 521. 25. Kersten H (1975): Mechanism of action of mitomycins. In: Antineoplastic and ImmunoSuppressive Agents, Vol. 2, AC Sartorelli, DG Johns, eds. Springer-Verlag, New York, pp. 47 - 64.
I n c i d e n c e of SCE in D o w n ' s S y n d r o m e L y m p h o c y t e s
285
26. Lambert B, Hansson K, Bui TH, Funes-Cravioto F, Lindsten J, Holmberg M, Strausmanis R (1976): DNA repair and frequency of X-ray and u.v.-light induced chromosome aberrations from patients with Down's syndrome. Ann Hum Genet 39,293 - 303. 27. Evans HJ (1977): Some facts and fancies relating to chromosome structure in man. In: Advances in Human Genetics, Vol. 8, H Harris, K Hirschhorn eds. Plenum Press, New York, pp. 347-438.
ABSTRACT: The frequency of sister chromatid exchanges {SCE) was studied in lymphocytes from Down's syndrome patients and normal donors after in vitro exposure to mitomycin C(0.01 ~g/ml) and X-irradiation. There was no difference in the response of Dawn's syndrome cells and normals following treatment with mitomycin C. However, Dawn's syndrome patients shewed a significant increase in SCE following exposure ta both 50 and 100 reds, whereas normal donors showed an elevated SCE rate only after exposure to 100 rads.
INTRODUCTION Down's s y n d r o m e (trisomy 21) is a congenital disorder associated with an increased i n c i d e n c e of l e u k e m i a [1,2]. Chromosomes from Down's s y n d r o m e patients exhibit an increased frequency of aberrations following X-irradiation [3,41, viral infection [5], and treatment with chemical mutagens [6,7]. Increased c h r o m o s o m e fragility in response to X-rays and chemical mutagens as well as an increased i n c i d e n c e of malignancy is also found in ataxia telangiectasia, F a n c o n i ' s anemia, and Bloom's synd r o m e and has lead to the suggestion of a possible association between c h r o m o s o m e fragility and m a l i g n a n c y [8]. Although the precise m e c h a n i s m s of sister c h r o m a t i d exchange (SCE) are not understood, the technique offers a means of investigating the possible relationship between c h r o m o s o m e aberrations and DNA repair processes. Ataxia telangiectasia, Fanconi's anemia, and Down's s y n d r o m e all have normal SCE in cultured l y m p h o cytes [9 - 11], whereas Bloom's s y n d r o m e cells show a 12-fold increase in SCE [12]. To gain further information about the cellular defects in Down's syndrome, we have s t u d i e d the i n c i d e n c e of SCE in Down's s y n d r o m e l y m p h o c y t e s following challenge with m i t o m y c i n C (MMC) and X-irradiation. MATERIALS AND METHODS
Peripheral blood l y m p h o c y t e s were isolated from defibrinated blood by gelatin sedim e n t a t i o n and cultured in H a m ' s FIO m e d i u m containing p h y t o h e m a g g l u t i n i n (PHA) 12.5 ~ l / m l (Burroughs Wellcome). B r o m o d e o x y u r i d i n e (BrdUrd) was a d d e d at From the CytogeneticsUnit, Cancer Society of New Zealand, Christchurch Hospital, Christchurch, New Zealand. Address requests for reprints to Dr P.E. Crossen, Cytogenetics Unit, Christchurch Hospital. Christchurch, New Zealand. Received February 15, 1980; accepted May 8, 1980. 281
Copyright O Elsevier North Holland, Inc., 1980 52 Vanderbilt Ave., New York, NY 1 0 0 1 7
Cancer Genetics and Cytogenetics2, 281-285 (1980) 0165-4608/80/05028105502.25
282
P . E . Crossen and W. F. Morgan Table 1
SCE in Down's s y n d r o m e and normal l y m p h o c y t e s following treatment with MMC a
Down's Syndrome
Normal
Baseline
MMC
× increase
Baseline
MMC
× increase
10.74 (8)
22.88
2.16
10.78 (20)
23.86
2.35
~Each entry is the mean number of SCE from the number of donors (in parentheses} studied.
a final concentration of 10.0 ~g/ml after 24 hr and cultures kept in a lightproof box to prevent photolysis of the BrdUrd-substituted DNA [13]. Colcemid (Ciba Laboratories) was a d d e d 4 hr prior to harvest at a final concentration of 0.1 ~g/ml and c h r o m o s o m e preparations made by the flame drying technique of Scherz [14]. Slides were treated for sister chromatid differential staining by a slightly m o d i f i e d version of the fluorescence plus Giemsa (FPG) technique of Perry and Wolff [15]. All slides were c o d e d and the incidence of SCE scored in 20 well spread metaphases from each patient. MMC was a d d e d to cultures at the same time as the BrdUrd (24 hr} at a final concentration of 0.01 ~g/ml. Cultures for X-ray treatment were grown in Falcon T30 flasks and X-rays a d m i n i s t e r e d by a Phillips RT 100 X-ray m a c h i n e (100 kV; 8 m A using a 0.4 m m c o p p e r filter). Cultures were irradiated at 44 hr with doses of 50 and 100 rads. RESULTS There was no difference between baseline SCE frequency in Down's s y n d r o m e patients (10.74) and normals (10.57). Although slightly higher than w e have p r e v i o u s l y reported [16,17], a baseline SCE frequency of a r o u n d 10 is in good agreement with our later studies [18]. Exposure to MMC caused an a p p r o x i m a t e doubling of SCEs in both the Down's s y n d r o m e patients and normal controls (Table 1). Fifty fads X-irradiation caused a significant increase in SCE in cultures from Down's s y n d r o m e patients (t -- 3.2, 3 degrees of freedom, p < 0.05), but not in norreals (Table 2). Both normals and Down's s y n d r o m e patients showed a significant increase in SCE following 100 rads (Table 3}. DISCUSSION The hypersensitivity of Down's s y n d r o m e l y m p h o c y t e s to X-irradiation is well established in studies showing increased c h r o m o s o m e aberrations and m i c r o n u c l e i formation [3,4]. Although radiation is a c o m p a r a t i v e l y poor i n d u c e r of SCEs [19,20], we
Table 2
SCE in Down's s y n d r o m e l y m p h o c y t e s following 50 rads X-irradiation.
Normal donors (6) Baseline 50 rads x increase over baseline
Down's syndrome patients I II III IV 10.33 ~ t0.60 10.75 a 11.81 1.04 1.11
9.30 13.95 1.50
9.60 16.75 1.74
aMean SCE rate of the number of donors (in parentheses) studied.
12.30 15.35 1.24
Incidence of SCE in D o w n ' s S y n d r o m e Lymphocytes Table 3
283
SCE in Down's s y n d r o m e l y m p h o c y t e s following 100 rads X-irradiation Down's Syndrome Patients V VI VII VIII
Normal donors (8) Baseline 100 fads x increase over baseline
10.61 a 14.61 a 1.37
11.90 14.00 1.17
11.55 17.70 1.53
14.00 18.30 1.30
10.75 13.45 1.25
~Mean SCE rate of the number of donors (in parentheses) studied.
found that 50 reds of X-irradiation significantly increased the SCE rate in Down's s y n d r o m e cells, but not in normals. It is u n l i k e l y that the increase in SCE following 50 rads is due to differential BrdUrd incorporation b e t w e e n Down's s y n d r o m e and normal ceils. Not only are baseline SCE frequencies almost identical, but increasing the BrdUrd concentration does not result in an increase in X-ray i n d u c e d SCEs [21]. A l t h o u g h the precise DNA lesion leading to SCE formation is at present u n k n o w n , it appears that lesions resulting in SCE formation are i n d u c e d in Down's s y n d r o m e ceils at very m u c h lower doses of X-irradiation than in normal cells. These results are in contrast to the situation in ataxia telangiectasia cells, w h i c h are also more sensitive to X-irradiation, but do not show an increase in SCE at the X-irradiation doses s h o w n [22]. X-ray doses of 100 rads caused a significant increase in the n u m b e r of SCE both in normal and Down's s y n d r o m e ceils. However, Ku~erov~ and Polikov~i [23] found no increase in SCE in either normal or trisomic cells exposed to 100 rads. The reason for this difference is not clear, but m a y reflect the time of irradiation, as there is evidence that irradiation of G o l y m p h o c y t e s does not increase SCE [21,24]. W h i l e Down's s y n d r o m e cells show an abnormal response to X-irradiation, our studies and those of Ku~erov~ and Polikova [23] show that they have a normal response to challenge by MMC. MMC is a bifunctional aklylating agent that can also i n d u c e cross-links in DNA [25]. It w o u l d a p p e a r that MMC induces a similar n u m b e r of lesions in both Down's s y n d r o m e and normal cells, and that they are repaired efficiently. X-ray i n d u c e d c h r o m o s o m e breaks are repaired more r a p i d l y in Down's s y n d r o m e cells [4], while the uptake of 3H-thymidine following ultraviolet light exposure is r e d u c e d [26], suggesting an a b n o r m a l i t y in DNA repair m e c h a n i s m s in Down's syndrome. Although the relationship between SCE and DNA repair is unclear, Evans [27] has suggested that SCE m a y represent a general error correcting process of high fidelity, e n c o m p a s s i n g a w i d e variety of DNA damage. Our data indicate that SCE functions n o r m a l l y in Down's syndrome, correcting both X-ray- and M M C - i n d u c e d lesions. This work was supported by the Central Districts, the Canterbury and West]and Divisions of the Cancer Society of New Zealand, and the Medical Research Council of New Zealand. The authors wish to thank Dr. P.H. Fitzgerald for helpful suggestions and Mrs. J. Stewart for assistance with the statistical analysis.
REFERENCES 1. Holland WW, Doll R. Carter CO (1962): The mortality from leukaemia and other cancers among patients with Down's syndrome (Mongolism) and among their parents. Br J Cancer, 16, 177-186.
284
P.E. Crossen and W. F. Morgan 2. Jackson EW, Turner JH, Klauber MR, Norris FD (1968): Down's syndrome: variation of leukemia in institutionalized populations. J Chronic Dis 2 1 , 2 4 7 - 253. 3. Sasaki MS, Tonomura A (1969): Chromosomal radiosansitivity in Down's syndrome. Jap J Hum Genet 14, 8 1 - 9 2 . 4. Countryman PI, Heddle JA, Crawford E (1977): The repair of X-ray induced chromosomal damage in trisomy 21 and normal diploid lymphocytes. Cancer Res 37, 5 2 - 58. 5. Higurashi M, Tamura T, Nakatake T (1973): Cytogenetic observations in cultured lymphocytes from patients with Down's syndrome and measles. Pediatr Res 7,582 - 587. 6. O'Brien RL, Pooh P, Kline E, Parker JW (1971): Susceptibility of chromosomes from patients with Down's syndrome to 7,12-dimethylbenz(a) anthracene induced aberrations in vitro. Int J Cancer 8,202 - 210. 7. Kaina B, Waller H, Waller M, Rieger R (1977): The action of N-methyl-N-nitrosourea on nonestablished cell lines in vitro. I. Cell cycle inhibition and aberration induction in diploid and Down's fibroblasts. Murat Res 43,387 -400. 8. German J (1972): Genes which increase chromosome instability in somatic cells and predispose to cancer. Prng Med Genet 8, 61 - 101. 9. Galloway SM, Evans HJ (1975): Sister chromatid exchanges in h u m a n chromosomes from normal individuals and patients with ataxia telangiectasia. Cytogenet Cell Genet 15, 17 - 29. 10. Latt SA, Stetten G, Juergens LA, Buchanan GR, Gerald PS (1975): Induction by alkylating agents of sister chromatid exchanges and chromatid breaks in Fanconi's anemia. Proc Natl Acad Sci USA 72, 4066-4070. 11. Lezana EA, Bianchi NO, Bianchi MS, Sabala-Suarez JE (1977): Sister chromatid exchanges in Down syndromes and normal h u m a n beings. Mutat Res 45, 85 - 90. 12. Chaganti RSK, Schonberg S, German J (1974): A many fold increase in sister chromatid exchanges in Bloom's syndrome lymphocytes. Proc Natl Acad Sci USA 71, 4508-4512. Ikushima T, Wolff S (1974): Sister chromatid exchanges induced by light flashes to 5-bro13. modeoxyuridine and 5-iododeoxyuridine substituted Chinese hamster chromosomes. Exp Cell Res 87, 15 - 19. 14. Scherz RG (1962): Blaze drying by igniting the fixative for improved spreads of chromosomes in leucocytes. Stain Technol 37,386. 15. Perry P, Wolff S (1974): New Giemsa method for the differential staining of sister chromarids. Nature 251,156-158. 16. Crossen PE, Drets ME, Arrighi FE, Johnston DA (1977): Analysis of the frequency and distribution of sister chromatid exchanges in cultured h u m a n lymphocytes. Hum Genet 35, 345-352. 17. Morgan WF, Crossen PE (1977): The incidence of sister chromatid exchanges in cultured h u m a n lymphocytes. Mutat Res 42,305 - 312. 18. Crossen PE, Morgan WF (1980): Sister chromatid exchange in cigarette smokers. Hum Genet 53,425 -426. 19. Perry P, Evans HJ (1975): Cytological detection of mutagen-carcinogen exposure by sister chromatid exchange. Nature 258, 121 - 125. 20. Crossen PE, Morgan WF (1979): The effects of ~ radiation on sister chromatid exchanges in cultured h u m a n lymphocytes. Mutat Res 62,125 - 129. 21. Morgan WF, Crossen PE (in press): X-irradiation and sister chromatid exchange in cultured h u m a n lymphocytes. Envirnon Mutagen. 22. Galloway SM (1977): Ataxia telangiectasia: The effects of chemical mutagens and X-rays on sister chromatid exchanges in blood lymphocytes. Mutat Res 45,343 - 349. 23. Ku~erov~ M, Polikova Z (1978): In vitro comparison of normal and trisomic cell sensitivity to physical and chemical mutagens. In: Mutagen Induced Chromosome Damage in Man, HJ Evans, DC Lloyd, eds. Edinburgh University Press, Edinburgh, pp. 185 - 190. 24. Littlefield LG, Colyer SP, Joiner EE, Dufrain RF (1979): Sister chromatid exchanges in human lymphocytes exposed to ionizing radiation during G 0. Radiat Res 78, 514 - 521. 25. Kersten H (1975): Mechanism of action of mitomycins. In: Antineoplastic and ImmunoSuppressive Agents, Vol. 2, AC Sartorelli, DG Johns, eds. Springer-Verlag, New York, pp. 47 - 64.
I n c i d e n c e of SCE in D o w n ' s S y n d r o m e L y m p h o c y t e s
285
26. Lambert B, Hansson K, Bui TH, Funes-Cravioto F, Lindsten J, Holmberg M, Strausmanis R (1976): DNA repair and frequency of X-ray and u.v.-light induced chromosome aberrations from patients with Down's syndrome. Ann Hum Genet 39,293 - 303. 27. Evans HJ (1977): Some facts and fancies relating to chromosome structure in man. In: Advances in Human Genetics, Vol. 8, H Harris, K Hirschhorn eds. Plenum Press, New York, pp. 347-438.