Doxorubicin treatment modulates chemoresistance and affects the cell cycle in two canine mammary tumour cell lines
Abstract Background Doxorubicin (DOX) is widely used in both human and veterinary oncology although the onset of multidrug resistance (MDR) in neoplastic cells often leads to chemotherapy failure. Better understanding of the cellular mechanisms that circumvent chemotherapy efficacy is paramount. The...
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doaj-9e19a9b5e529431598bfe941ca04addb2021-01-24T12:15:30ZengBMCBMC Veterinary Research1746-61482021-01-0117111510.1186/s12917-020-02709-5Doxorubicin treatment modulates chemoresistance and affects the cell cycle in two canine mammary tumour cell linesMichela Levi0Roberta Salaroli1Federico Parenti2Raffaella De Maria3Augusta Zannoni4Chiara Bernardini5Cecilia Gola6Antonio Brocco7Asia Marangio8Cinzia Benazzi9Luisa Vera Muscatello10Barbara Brunetti11Monica Forni12Giuseppe Sarli13Department of Veterinary Medical Sciences, University of BolognaDepartment of Veterinary Medical Sciences, University of BolognaDepartment of Veterinary Medical Sciences, University of BolognaDepartment of Veterinary Sciences, University of TurinDepartment of Veterinary Medical Sciences, University of BolognaDepartment of Veterinary Medical Sciences, University of BolognaDepartment of Veterinary Sciences, University of TurinDepartment of Veterinary Medical Sciences, University of BolognaDepartment of Veterinary Medical Sciences, University of BolognaDepartment of Veterinary Medical Sciences, University of BolognaDepartment of Veterinary Medical Sciences, University of BolognaDepartment of Veterinary Medical Sciences, University of BolognaDepartment of Veterinary Medical Sciences, University of BolognaDepartment of Veterinary Medical Sciences, University of BolognaAbstract Background Doxorubicin (DOX) is widely used in both human and veterinary oncology although the onset of multidrug resistance (MDR) in neoplastic cells often leads to chemotherapy failure. Better understanding of the cellular mechanisms that circumvent chemotherapy efficacy is paramount. The aim of this study was to investigate the response of two canine mammary tumour cell lines, CIPp from a primary tumour and CIPm, from its lymph node metastasis, to exposure to EC50(20h) DOX at 12, 24 and 48 h of treatment. We assessed the uptake and subcellular distribution of DOX, the expression and function of P-glycoprotein (P-gp) and Breast Cancer Resistance Protein (BCRP), two important MDR mediators. To better understand this phenomenon the effects of DOX on the cell cycle and Ki67 cell proliferation index and the expression of p53 and telomerase reverse transcriptase (TERT) were also evaluated by immunocytochemistry (ICC). Results Both cell lines were able to uptake DOX within the nucleus at 3 h treatment while at 48 h DOX was absent from the intracellular compartment (assessed by fluorescence microscope) in all the surviving cells. CIPm, originated from the metastatic tumour, were more efficient in extruding P-gp substrates. By ICC and qRT-PCR an overall increase in both P-gp and BCRP were observed at 48 h of EC50(20h) DOX treatment in both cell lines and were associated with a striking increase in the percentage of p53 and TERT expressing cells by ICC. The cell proliferation fraction was decreased at 48 h in both cell lines and cell cycle analysis showed a DOX-induced arrest in the S phase for CIPp, while CIPm had an increase in cellular death without arrest. Both cells lines were therefore composed by a fraction of cells sensible to DOX that underwent apoptosis/necrosis. Conclusions DOX administration results in interlinked modifications in the cellular population including a substantial effect on the cell cycle, in particular arrest in the S phase for CIPp and the selection of a subpopulation of neoplastic cells bearing MDR phenotype characterized by P-gp and BCRP expression, TERT activation, p53 accumulation and decrease in the proliferating fraction. Important information is given for understanding the dynamic and mechanisms of the onset of drug resistance in a neoplastic cell population.https://doi.org/10.1186/s12917-020-02709-5Canine mammary tumourCell lineChemoresistanceDoxorubicinP-glycoproteinBCRP |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Michela Levi Roberta Salaroli Federico Parenti Raffaella De Maria Augusta Zannoni Chiara Bernardini Cecilia Gola Antonio Brocco Asia Marangio Cinzia Benazzi Luisa Vera Muscatello Barbara Brunetti Monica Forni Giuseppe Sarli |
spellingShingle |
Michela Levi Roberta Salaroli Federico Parenti Raffaella De Maria Augusta Zannoni Chiara Bernardini Cecilia Gola Antonio Brocco Asia Marangio Cinzia Benazzi Luisa Vera Muscatello Barbara Brunetti Monica Forni Giuseppe Sarli Doxorubicin treatment modulates chemoresistance and affects the cell cycle in two canine mammary tumour cell lines BMC Veterinary Research Canine mammary tumour Cell line Chemoresistance Doxorubicin P-glycoprotein BCRP |
author_facet |
Michela Levi Roberta Salaroli Federico Parenti Raffaella De Maria Augusta Zannoni Chiara Bernardini Cecilia Gola Antonio Brocco Asia Marangio Cinzia Benazzi Luisa Vera Muscatello Barbara Brunetti Monica Forni Giuseppe Sarli |
author_sort |
Michela Levi |
title |
Doxorubicin treatment modulates chemoresistance and affects the cell cycle in two canine mammary tumour cell lines |
title_short |
Doxorubicin treatment modulates chemoresistance and affects the cell cycle in two canine mammary tumour cell lines |
title_full |
Doxorubicin treatment modulates chemoresistance and affects the cell cycle in two canine mammary tumour cell lines |
title_fullStr |
Doxorubicin treatment modulates chemoresistance and affects the cell cycle in two canine mammary tumour cell lines |
title_full_unstemmed |
Doxorubicin treatment modulates chemoresistance and affects the cell cycle in two canine mammary tumour cell lines |
title_sort |
doxorubicin treatment modulates chemoresistance and affects the cell cycle in two canine mammary tumour cell lines |
publisher |
BMC |
series |
BMC Veterinary Research |
issn |
1746-6148 |
publishDate |
2021-01-01 |
description |
Abstract Background Doxorubicin (DOX) is widely used in both human and veterinary oncology although the onset of multidrug resistance (MDR) in neoplastic cells often leads to chemotherapy failure. Better understanding of the cellular mechanisms that circumvent chemotherapy efficacy is paramount. The aim of this study was to investigate the response of two canine mammary tumour cell lines, CIPp from a primary tumour and CIPm, from its lymph node metastasis, to exposure to EC50(20h) DOX at 12, 24 and 48 h of treatment. We assessed the uptake and subcellular distribution of DOX, the expression and function of P-glycoprotein (P-gp) and Breast Cancer Resistance Protein (BCRP), two important MDR mediators. To better understand this phenomenon the effects of DOX on the cell cycle and Ki67 cell proliferation index and the expression of p53 and telomerase reverse transcriptase (TERT) were also evaluated by immunocytochemistry (ICC). Results Both cell lines were able to uptake DOX within the nucleus at 3 h treatment while at 48 h DOX was absent from the intracellular compartment (assessed by fluorescence microscope) in all the surviving cells. CIPm, originated from the metastatic tumour, were more efficient in extruding P-gp substrates. By ICC and qRT-PCR an overall increase in both P-gp and BCRP were observed at 48 h of EC50(20h) DOX treatment in both cell lines and were associated with a striking increase in the percentage of p53 and TERT expressing cells by ICC. The cell proliferation fraction was decreased at 48 h in both cell lines and cell cycle analysis showed a DOX-induced arrest in the S phase for CIPp, while CIPm had an increase in cellular death without arrest. Both cells lines were therefore composed by a fraction of cells sensible to DOX that underwent apoptosis/necrosis. Conclusions DOX administration results in interlinked modifications in the cellular population including a substantial effect on the cell cycle, in particular arrest in the S phase for CIPp and the selection of a subpopulation of neoplastic cells bearing MDR phenotype characterized by P-gp and BCRP expression, TERT activation, p53 accumulation and decrease in the proliferating fraction. Important information is given for understanding the dynamic and mechanisms of the onset of drug resistance in a neoplastic cell population. |
topic |
Canine mammary tumour Cell line Chemoresistance Doxorubicin P-glycoprotein BCRP |
url |
https://doi.org/10.1186/s12917-020-02709-5 |
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