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Identifier

000379479

Title

Ex vivo visualization of megakaryocytic endomitosis and the role of cyclin E in this process

Alternative Title

Ex vivo απεικόνιση της ενδομιτωτικής διαδικασίας των μεγακαρυοκυττάρων και διερεύνηση του ρόλου της κυκλίνης Ε στη διαδικασία αυτή

Author

Παπαδαντωνάκης, Νικόλαος Ε

Reviewer

Παπαδάκη, Ελένη
Μπούμπας, Δημήτριος
Ηλιόπουλος, Αριστείδης

Abstract

The endomitotic cell cycle in Megakaryocytes, which is comprised by repeated cycles of abrogated mitosis is an integral part of Megakaryocytic development . By utilizing a unique transgenic mouse model where GFP-histone expression is limited to megakaryocytic lineage we demonstrated that endomitosis is different between Megakaryocytes of high and low ploidy. In the low ploidy Megakaryocytes the DNA is clearly segregated into two distinct regions before it rejoins into a single nucleus. In High ploidy Megakaryocytes distinct segregation of DNA is not observed prior to DNA content rejoining into a single nucleus. Our study was the first one to describe the chromosomal dynamics of endomitosis Ex vivo using live imaging. In order to further characterize the endomitotic process efforts to generate transgenic mice that their microtubule network could be visualized were pursued. Several transgenic lines were generated without success and a novel approach is also outlined that may overcome the problems of previous transgenic mice attempts. In addition, based on transgenic mouse model where human Cyclin E1 overexpression is restricted in the megakaryocytic lineage we demonstrate that a higher fraction of bone marrow transgenic Megakaryocytes are in the S phase compared to wild type Megakaryocytes indicating that Cyclin E overexpression is sufficient to drive Megakaryocytes to S phase. Furthermore, a method to rapidly identify homozygotes from heterozygotes in the founder lines of transgenic mice is also described. Polo Like Kinase 3 is a protein that is implicated in the modulation of cell cycle and we detected its expression in Megakaryocytes but experiments using Knock out mice did not reveal an impact in polyploidization. Lysyl Oxidase propeptide (LOX-PP) is a moiety that is is part of the Pre Lysyl oxidase protein and is liberated when mature Lysyl oxidase is produced. It has been shown in multiple studies to exert a powerful anti- proliferation effect in cell lines and here we describe its effect in Megakaryocytic physiology and ploidy. Namely the Lysyl Oxidase Propeptide was able to inhibit Megakaryocytic polyploidization in bone marrow cultures without affecting viability. Importantly, Megakaryocytes expressing human Cyclin E1 were not able the inhibitory effect of LOX-PP.

Language

Greek, English

Subject

Cell Physiological Phenomena

Cyclin

Endomitosis

Ex-vivo

Marrow

Megakaryocytes

Molecular Biology

PLK-3

Ploidy