Long-term 17alpha-ethinyl estradiol treatment decreases cyclin E and cdk2 expression, reduces cdk2 kinase activity and inhibits S phase entry in regenerating rat liver.
OBJECTIVE
The synthetic estrogen 17alpha-ethinyl estradiol (EE), a potent tumor promoter in rat liver, stimulates growth during short-term treatment but inhibits hepatocyte proliferation upon prolonged treatment. To identify the molecular targets of the mitoinhibitory effect of EE, the expression of proteins regulating G(1)- and S-progression were analyzed during the first cell cycle in EE-treated female Wistar rats.


METHODS
Long-term (60 days) EE treatment. Immunohistochemical staining for proliferation cell nuclear antigen (PCNA) to detect cells in S phase and quantification of mitosis. Western blot to monitor protein expression. Cdk2 kinase assay to examine histone H1 phosphorylation.


RESULTS
EE reduced the number of cells in S phase and mitosis by about 70%. Cyclin D1 and D3 were unaffected, while cdk4 was moderately decreased. Cyclin E and cdk2 were markedly decreased with concomitant marked reduction of cdk2 kinase activity. EE also decreased cyclin A and increased G1 levels of p53 and p21.


CONCLUSIONS
EE causes a cell cycle block before S-phase. The reduction of the cdk2 kinase activity, essential for G1/S-transition, might be involved in the cell cycle block. Also, EE treatment results in p53 activation and upregulation of the cdk inhibitor p21 that might contribute to the G1 arrest.
Koroxenidou
Lena
L
Department of Natural Science, Södertörns Högskola (University College), Box 4101, S-141 89 Huddinge, Sweden. koroxenidou@yahoo.se
Ohlson
Lena C E
LC
Porsch Hällström
Inger
I
D000818:Animals; D042846:CDC2-CDC28 Kinases / Q000235:genetics* / Q000378:metabolism; D019938:Cyclin D1 / Q000235:genetics; D056743:Cyclin D3; D019927:Cyclin E / Q000235:genetics*; D051357:Cyclin-Dependent Kinase 2; D016213:Cyclins / Q000235:genetics; D004261:DNA Replication; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects; D008099:Liver / Q000187:drug effects / Q000502:physiology*; D008115:Liver Regeneration / Q000187:drug effects / Q000502:physiology*; D008938:Mitosis / Q000187:drug effects; D009641:Norethynodrel / Q000031:analogs & derivatives* / Q000627:therapeutic use; D051381:Rats; D017208:Rats, Wistar; D016196:S Phase / Q000187:drug effects / Q000502:physiology*
C534698:Ccnd3 protein, rat; D056743:Cyclin D3; D019927:Cyclin E; D016213:Cyclins; D019938:Cyclin D1; C002029:17alpha-ethynylestr-5(10)-ene-3alpha,17beta-diol; D009641:Norethynodrel; D042846:CDC2-CDC28 Kinases; C495892:Cdk2 protein, rat; D051357:Cyclin-Dependent Kinase 2