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Titre : | Traitement du cancer du sein pendant la grossesse :Détermination de leffet de la chimiothérapie sur ledéveloppement placentaire |
Auteurs : | Alexandre MORJAU, Auteur ; Christophe L. Depoix, Promoteur |
Type de document : | Travail de fin d'études |
Editeur : | Bruxelles : Institut Paul Lambin, 2020 |
Langues: | Français |
Index. décimale : | TFE - Biologie médicale |
Résumé : |
Background About 0,2% of pregnant women will be diagnosed with cancer. Even if this number is very low, the number of cases is rising with time. Breast cancer is the most common cancer diagnosed during pregnancy. Chemotherapy is frequently used to cure this condition, but not without side effects on the fetus. It can be started only after the first trimester, but it is known to cause fetal growth retardation. Since there is a correlation between the weight of the placenta and the baby, we assume that chemotherapy negatively affects the development of the placenta.
Methods Cytotrophoblasts were isolated and purified from placentas collected just after delivery. Cells were allowed to differentiate into syncytiotrophoblast for 48-hours before treatment with the drugs. Four drugs have been tested: epirucin, docetaxel, 5-fluorouracile (5-FU) and 4-hydroperoxy-cyclophosphamid. Firstly, because 5-FU and 4-hydroxyperoxy-cyclophosphamide were assayed for the first-time, their cytotoxicity has been evaluated using a wide range of concentrations. Cell mortality has been measured with CytoTox-Glo Cytotoxicity Assay after a 24-hour treatment with the drugs. Secondly, the effect of the four drugs on cell viability and apoptosis has been measured after 6 and 24 hours with ApoLive-Glo Multiplex Assay. Three different concentrations have been tested: Epirubicin [0.01, 0.1, and 1 μM), Docetaxel (0.01, 0.1, and 1 μM), 5-FU (5, 50, and 50 μM), and 4-hydroxyperoxy-cyclophosphamid (0.03, 0.3, and 3 μM). Thirdly, gene expression has been evaluated. Total RNA was extracted, and retrotranscribed into cDNA for subsequent gene amplification and quantification by real-time PCR. Finally, cell fusion was assessed by immunofluorescence. The cell membrane was marked with a fluorescent molecule-conjugated anti-desmoplakin antibody, and the nuclei were dyed with NucBlue. Cell fusion was then determined by counting the number of cells with more than three nuclei (syncytiotrophoblasts). Those with less than three nuclei were not considered as differentiated (cytotrophoblast). Results Only 4-hydroxyperoxy-cyclophosphamide presented cytotoxic effect with a rise in the number of dead cells at the concentration of 50 μM. After a 24-hour treatment with 1 μM of epirubicine, cell viability decreased to ~60%. The other drugs had no effect on cell viability. Concerning apoptosis, after a 6-hour treatment, caspase 3/7 activity increased by 2-fold with 1 μM of epirubicin compared to the DMSO-treated group (control). After a 24-hour treatment, there was an 8-fold increase in caspase activity over control. The treatments had no significant effect on gene expression and cell fusion. Conclusion Only epirubicine had negative effects on cell viability and apoptosis. This can be explained by the ability of epirubicin to cause oxidative stress. Nor cell fusion or gene expression seemed to be affected by the treatments. Those results are in accordance with the fact that chemotherapy is well supported by the fetus, even if some side effects have been reported. In the future, it will be interesting to use drugs combination as in classical chemotherapy and increase the number of placentas to lower the variations in the results. |
Accès : | Identifiez-vous avant d'accéder au document électronique |
Disponible en ligne : | Oui |
Lieu du stage : | Institut de recherche expérimentale et clinique Laboratoire dobstétrique |
Département : | Biologie médicale |
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TFE biologie médicale Adobe Acrobat PDF |