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Identifier

000378503

Title

The role of breast cancer resistance protein 1 (BCRP1) in the cardiac side population cells

Alternative Title

Μελέτη του ρόλου της πρωτείνης BCRP1στα κύτταρα του πλευρικού πληθυσμού της καρδιάς

Author

Sereti, Konstantina-Ioanna

Author

Σερέτη, Κωνσταντίνα -Ιωάννα

Thesis advisor

Roglih Liao

Reviewer

Ζαννής, Βασίλης
Παπαδάκη, Ε.
Ηλιόπουλος, Α.
Καραγωγέως, Δ.
Καρδάσης, Δ.
Γραβάνης, Α.
Μπούμπας, Δ.

Abstract

Cardiovascular diseases represent the leading cause of death in the industrialized world. Several scientific advances have achieved significant improvement in disease treatment and patient quality of life. However, with the exception of heart transplantation which represents the most efficient treatment, none of the current therapies focus on replacing the lost cardiac tissue. The identification of mitotic myocytes and more recently, of resident cardiac progenitor cells has abolished the long standing dogma that the heart is a terminally differentiated organ. Cardiac regeneration, through stem cell based therapies has become a promising area of research and has opened new horizons for the treatment of cardiovascular diseases. Cardiac side population (CSP) cells represent a resident cardiac progenitor cell population. CSP cells are identified based on the ability of ABC-cassette membrane transporters to efflux the DNA-binding dye Hoechst 33342. ABC-transporters Abcg2 and Mdr1 have been shown to efficiently export Hoechst. Upon proper stimulation, CSP cells are able to differentiate into all major cardiac cell types. Abcg2 and Mdr1 belong to the large family of ABC-transporters. ABC proteins are involved in the trafficking of a large variety of substrates across the cell membrane and intracellular organelles. Abcg2 and Mdr1 were initially identified in cancer drug resistant cells lines and have been associated with chemotherapy drug resistance observed in cancers. In particular, in addition to cancer resistance, Abcg2 has been suggested to play a protective role in crucial tissues such as the brain and fetus against xenobiotic transfer 9 through the blood-brain and placenta barriers. Abcg2 has also been shown to confer protection from cell death under hypoxic conditions. Most importantly, Abcg2 has been identified as the molecular determinant of the bone marrow SP phenotype. However, its contribution to the cardiac SP phenotype remains unclear. Abcg2 expression has been further linked to the proliferation of cancer cells. Additionally, in bone marrow SP cells, Abcg2 over-expression promoted cell proliferation while its inhibition resulted in abrogation of hematopoiesis. In the heart, Abcg2 expression has been demonstrated to be beneficial following injury by protecting microvascular endothelial cell function. The fundamental characteristic of stem/progenitor cells is the capacity to selfrenew and differentiate within one cell division. Stem/progenitor cell homeostasis is achieved through a balance between symmetric and asymmetric cell divisions. Regulation of the switch between these two types of division is of particular importance in normal and cancer stem cells. Deregulated divisions can lead to stem cell pool exhaustion or cancer cell over-proliferation. Cell fate decisions are made during the cell cycle and particularly during the G1- phase. The “cell cycle length” hypothesis suggests that lengthening of G1 is associated and required for stem/progenitor cell asymmetric division and differentiation. The goal of my study is to investigate the role of Abcg2 in the CSP phenotype and homeostasis as well as its effects in vivo, in a myocardial injury context. CSP cells from WT, Abcg2-KO and Mdr1a/b-KO mice from different developmental stages were analyzed by flow cytometry to examine the relative contribution of each 10 transporter. The effects of Abcg2 on CSP cell proliferation and survival were determined through various methods. Gain- and loss-of-function approaches were utilized to confirm the results. Site-mutagenesis allowed the assessment of the role of Abcg2 efflux capacity in CSP proliferation. Cell cycle marker staining, lentiviral cell cycle indicators and RT-PCR based gene arrays were used in combination with live cell imaging, to delineate the cell cycle profile of WT and Abcg2-deficient CSP cells. Immuno-cytochemical staining for cell fate determinants and cardiac markers, revealed the effects of Abcg2 on CSP asymmetric division and cardiomyogenic differentiation. Moreover, ischemia-reperfusion and myocardial infarction were used as myocardial injury models to investigate the in vivo role of Abcg2. Lastly, the surface marker expression, proliferation and differentiation capacity of CSP cells from different developmental stages were analyzed. My work reveals for the first time that Abcg2 has an age-dependent contribution to the CSP phenotype. Moreover, Abcg2 was found to promote CSP cell cycle progression and survival while inhibiting their asymmetric division and differentiation. Finally, my work provides in vivo evidence supporting that Abcg2 plays a protective role following myocardial injury.

Language

Greek

Subject

Asymmetricdivision

Cardiac side population cells (CSP)

Cardiovascular diseases

Cell cycle

Membrane transporter ABCG2

Myocardial infarction

Stem/progenitor cells

cardiac regeneration

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