halogenated analogs of thymidine

Vladimir I. Stobetsky sto at aha.ru
Sat Mar 14 01:54:19 EST 1998




Key words: halogenated analogs of thymidine, human lymphocytes, allocyclic
chromosomes

Cytogenetic  abnormalities in cultured human lymphocytes under treatment
with halogenated analogs of thymidine

After prolonged co-treatment of heteroploid Chinese hamster cells with 
colcemid, that induced cells with micronuclei, and halogenated analogs of 
thymidine (HAT) such as 5-iododeoxyuridine (5-IdU),  5-bromodeoxyuridine 
(5-BrdU)  and 5-chlorodeoxyuridine (5-ChldU) were discovered  dicentric 
chromosomes formed by "end-to-end fusions" (Stobetsky,1976). Registered 
dicentrics were a result of delayed disruption  of telomeric links existed 
in interphase nucleus
(Stobetsky,1978).

In  this study we investigated the cytogenetic abnormalities in cultured 
human lymphocytes under treatment with low doses HAT.  
 In experiments we used peripheral blood  of healthy donors - man at the 
age 44 years an woman at the age 35 years. We  added to 0,5 ml heparinized 
blood 0,2 ml PHA , 6,0 ml Eagle medium and 1,5 ml calf serum. The cells 
cultured at 37o C. HAT  (20 microg/ml)  administrated at 48 h of
cultivation 
for 20 h. The cells treated with colcemid (0,1 microg/ml) during  last 1,5
h. 
Chromosome reparations made with standard method. In each case we scored 
100 metaphases.

Results

Most frequently we observed  the delay of spiralization in  the region of 
constitutive heterochromatin  in chromosome No9. 
Table No1.Frequency of chromosome abnormalities in human lymphocytes under 
  treatment with HAT.	
Type of abnormality                                                        
                                               Intact  culture      5-IdU  
 5-BrdU  5-ChldU	
                                                        Man lymphocytes	
1.Metaphases with delay spiralization or break in heterochromatin region of
chr.9.	       0	        27	         5	        1	
2.Metaphases with onechromatid break or gap (except chromosomes 1,9 and 16)
       3	         4	         5 	        9	
                                                         Woman lymphocytes	
1.	                                                                        
                                                                      0	   
     43	         12	        7	
2.	                                                                        
                                                                      2	   
       1           4 	        10	

Table 1  shows that first abnormality  have a maximum under treatment with 
5-IdU and minimum under action  of 5-ChldU for man and 
woman.Heterochromatic region of chromosome 9   look like as  prophasic  or 
as "pulverization".  Sometimes this  region appeared  as  a break  i.e.  
was  achromatic. This  table shows also that frequency of onechromatid  
breaks  increased in he  row  5-IdU  -  5-BrdU  -  5-ChldU. Locus  of delay

spiralization of the chromosome 9  we consider as allocyclic  chromosome  
as it  is or in   early prophase or  in S-phase  i.e. such chromosomes  are

allocyclic  ( The behavior of allocyclic chromosomes in Bloom's syndrome. 
Otto P.G. Otto P.A. and Eeva Therman.  Cromosoma,1981,84:337-344). So HAT  

(particularly 5-IdU)  induces  the shift some phases of mitotic cycle.  In 

favour of this  hypothesis we have found some cases of  "pulverization" of 
whole  chromosome. Intensity of this process  depends  directly  from 
molecular weight  of  HAT. We  consider that the delay disruption of 
telomeric links in cells with micronuclei  in heteroploid Chinese cell 
lines  that  induced  with  colcemid and HAT which  appeared as dicentrics 

are also allocyclic chromosomes so  the disruption of telomeric links  
occur  at the early stages of mitosis  as telomeric fusion exists  at the 
interphase. Thus  we conclude   from our experiments  that cell response 
depends  on type of   HAT  , specific  properties of cells and  mode of 
treatment.

References
1.Stobetsky  V.I. (1976). Chromosome fusion by telomeres in cells treated 
with colcemid and  5-bromodeoxyuridine. (Russ). Bull.Exptl.Biol.Med. No9. 
1142-1144.
2.Stobetsky  V.I. (1978). Mammalian polykaryocytes  in vitro.II.The 
phenomenon of delayed disruption of telomeric links between  chromosomes. 
Biologisches Zentralblatt. 97:595-604.
3.Stobetsky  V.I.  (1995). Cytogenetic   abnormalities in cultured  human 
lymphocytes undertreatment  with  halogenated analogs of thymidine.(Russ). 
Bull.Exptl.Biol.Med.  No9. 326-328.

*******************************************************
Vladimir I. Stobetsky
Senior Scientist
Research Institute of Pediatric Hematology
Laboratory of Cytogenetics and Molecular Biology
App. 116, 6 Novoorlovskaya St. Moscow, 119633, Russia
tel: +7 095 731 8932   
e-mail address: sto at aha.ru  






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