EVALUATION OF THE CHANGES RESULTING FROM TAMOXIFEN ADMINISTRATION. A COMBINED DNA FLOWCYTOMETRIC AND HISTOPATHOLOGICAL STUDY
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1 EVALUATION OF THE CHANGES RESULTING FROM TAMOXIFEN ADMINISTRATION. A COMBINED DNA FLOWCYTOMETRIC AND HISTOPATHOLOGICAL STUDY
2 Tamoxifen Tamoxifen is a non steroidal tiphenylethylene y first synthesized in The drug was initially developed as an oral contraceptive, but instead of blocking ovarian function, tamoxifen was found to induce ovulation.
3 Tamoxifen Tamoxifen is widely used as an anti estrogin adjuvant therapy for breast cancer patients ts and as a chemopreventive agent for healthy women at high risk for breast cancer.
4 Tamoxifen Since 1971 it has been used successfully to treat many millions of women. Treatment results showed an increaseindiseasefreesurvivaland decrease inrecurrencerateof breast cancer.
5 Tamoxifen On October 29,1998, the Food and Drug Administration (FDA) approved Nolvadex (tamoxifen citrate) for reducing the incidence of breast cancer in women at high risk for developing the disease.
6 Tamoxifen With the widespread therapeutic and emerging prophylactic use of tamoxifen, there has been much discussion about side-effects of the drug, particularly its carcinogenicity
7 Tamoxifen Numerous studies have established an increased incidence of endometrial cancer among women taking tamoxifen. Recent results confirm not only an increased incidence of endometrial cancer but also increased mortality from the disease
8 Tamoxifen A number of studies are compatible with genotoxic activity. Tamoxifen induces micronuclei in metabolically competent human cells causes aneuploidy and chromosomal aberrations
9 Tamoxifen In rats, tamoxifen is a potent t hepatocarcinogen in both males and females.
10 Tamoxifen It has been characterized as a human carcinogen by the International Agency for Research on Cancer.
11 Aim The aim of the present study is to evaluate the effect of tamoxifen administration on uterus, kidney, spleen and liver tissues in mice by detecting the histopathologic changes as well as the DNA ploidy and cell cycle phase analysis.
12 Material and methods A total of 40 female Swiss Albino mice were involved in the study. The animals were divided at random into 2 groups: Mice of the first group were served as control group, were orally administered with normal saline 3 times /week for 6 weeks. Mice of the second group were orally administered with 20mg/kg tamoxifen 3 times /week for 6 weeks.
13 Material and methods Animals were sacrificed and the uterus spleen liver and kidneys were removed and each organ was divided into 2 pieces. one piece was fixed in 10% formaldehyde solution dehydrated, embedded in wax, sectioned, deparafinised d and stained with H&E for histopathological examination.
14 Material and methods The other parts of the previously mentioned organs were directly transferred to the flow cytometry laboratory where single cell suspensions were prepared and stained with propidium p iodide (PI) and subjected to DNA flow cytometric analysis.
15 RESULTS Uterine tissue: The results of the histopathological examination indicated that there is a pathological change in the form of squamous metaplasia and dysplasia of the glandular epithelial cells.
16 Dysplastic Squamous cells, lining endometrum (following drug application) are large in size, with vesicular nuclei (1) and prominent nuclei (2), kerato hyaline granules (3) are abundant in upper layers.
17 The effect of TAM treated on DNA ploidy in the uterine tissue. 100 ploid c ells % of dipoi id and aneu Diploid Aneuploid Control TAM Percent of diploid and aneuploid cells in the uterus of the normal control and 6 weeks treated animals
18 Percent of cell cycle phases of diploid and aneuploid cells in the uterus Percen nt of cell l cycle ph hases 100% 80% 60% 40% 20% 0% S G2-M G0-G1 control TAM control TAM Diploid cells Aneuploid cells
19 Results Histopathological examination of Kidney tissues showed a minimal change in the form of focal renal lesion in the renal tubular epithelium.
20 renal tubular epithelial cells with higher nuclear cytoplasmic ratio, hyper chromatic nuclei (»), density esinophilic cytoplasm
21 The effect of TAM treatment on DNA ploidy in the kidney tissues. % of di ipoid and an neuploid ce ells Diploid Aneuploid Control TAM Percent of diploid and aneuploid cells in the kidney of the normal control and 6 weeks treated animals.
22 Percent of cell cycle phases of diploid and aneuploid cells in The kidney Perce ent of cel ll cycle phases 100% 80% 60% 40% 20% 0% S G2-M G0-G1 G1 control TAM control TAM Diploid cells Aneuploid cells
23 Results No histopathologicl changes was No histopathologicl changes was detected in the spleen.
24 spleen of mice treated with tamoxifen for 6 weeks revealing no pathological changes
25 The effect of TAM treatment on DNA ploidy in the spleen tissues lls % of dip poid and an neuploid ce Diploid Aneuploid Control TAM P ercent of diploid and aneuploid treated anim als. cells in the spleen of the normal control and TAM
26 Percent of cell cycle phases of diploid and aneuploid cells in The spleen Perc cent of cell cycle phases s 100% 80% 60% 40% 20% 0% S G2-M G0-G1 control TAM control TAM Diploid cells Aneuploid cells
27 Results No histopathological changes were detected in the liver tissues of tamoxifen treated animals.
28 section from liver in tamoxifen treated mice, showing no pathological changes
29 The effect of TAM treated on DNA ploidy in the liver tissues neuploid ce ells % of dipoid and a Diploid Aneuploid 0 0 Control TAM Percent of diploid and aneuploid cells in the liver of the normal control and TAM treated animals
30 Percent of cell cycle phases of diploid and aneuploid cells in The liver. 100% S 80% G2-M G0-G1 60% 40% 20% 0% Control TAM Control TAM Diploid cells Aneuploid cells
31 Conclusion From the study we can conclude that tamoxifen may induce ploidy changes and disturbance in the cell cycle in the uterus, kidney and spleen tissues of female mice. Tamoxifen may induce cell proliferation in the cells oftheuterusandspleenandthisisclearfromthe increased percentage of cells in the S phase in these two tissues. Beside the histopathological investigation we may need to use the DNA flow cytometric analysis to detect the early changes resulting from tamoxifen administration that may results into malignant transformantion.
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