Emergence of haematopoietic stem cells and cancer

 

kissa karima2

KISSA Karima

Team manager

CR1 INSERM

ATIP-Avenir 2014

karima.kissa
arobase univ-montp2.fr

lelievre e2

LELIEVRE Etienne
CR1, INSERM

 

etienne.lelievre

arobase univ-montp2.fr

 

 

fontenille l2

FONTENILLE Laura

Start-up Manager AZELEAD

 

laura.fontenille

arobase univ-montp2.fr

 

 

 

 

Past achievement:

    The zebrafish embryo, thanks to its transparency and the recent development of molecular tools and transgenic fluorescent reporter lines, is a relevant in vivo model to study the different steps lead-ing to definitive haematopoiesis both in terms of regulatory pathways and in term of cellular journey in the different compartments of the embryo.

Imaging of live zebrafish embryos during the first week of development showed the presence of in-creasing numbers of round cells displaying the morphology of haematopoietic precursors in the ven-tral part of the tail, around the caudal vein (CV) plexus. This haematopoietic population expanded dur-ing the development of this CV plexus. By 48 hpf, this tissue had extended largely beyond the ventral limit of the somitic muscles, and became entirely covered by them on either side within the next 24 hrs. Small cell clusters then started to develop at 3 dpf, and formed larger groups by 5–7 dpf.
We have called this CV plexus the Caudal Haematopoïetic Tissue (CHT) and we have shown that it is the functional homologue of the mammal foetal liver (Murayama et al, Immunity, 2006).


In mammals, the CD41 surface protein that is strongly expressed at the surface of platelets is also expressed, but to a lesser extent, by embryonic HSCs. Using the CD41:GFP transgenic zebrafish line, it has been shown that the CD41 surface protein is also strongly expressed in thrombocytes (homologous of mouse megakaryocytes). Our studies have shown that the first CD41:GFP+ cells appear in the fish AGM from 30 hpf. Then, they migrate to the CHT and later to the thymus and kidney. We then conducted a 4D confocal imaging experiment to investigate in vivo, for the first time, and in a vertebrate, the dynamics of multipotent haematopoietic precursors from the AGM to the developing thymus (Movie 1, Kissa et al, Blood, 2008).


Movie 1. Routes of the first hematopoietic immigrants to the nascent thymus. Kissa et al, Blood, 2008.


Finally, in order to image the emergence of HSCs from the aorta, we undertook a 4D confocal micros-copy analysis of the developing aorta in the trunk region between 18 and 72 hpf. We used the KDRl:GFP+ zebrafish transgenic line whose aortic endothelial cells express GFP under the control of the KDRl promoter (ortholog of Flk1/VEGF receptor 2). A strong GFP expression was detected at the surface of the arterial endothelium in this transgenic line. We systematically tracked each aortic cell in live embryos. Our results showed that multipotent precursors emerged directly from the aorta floor through a stereotyped process that involves a strong bending then egress of single endothelial cells from the aortic ventral wall into the sub-aortic space, and their concomitant transformation into haematopoietic cells. We named this process the endothelial haematopoietic transition (EHT, Kissa & Herbomel, 2010).
Thus, we have described that HSCs emerge from the ventral wall of the aorta, but the molecular and cellular events driving this process still have to be addressed. Moreover, the characterization and the long term fate of HSCs emerging from the ventral part of the aorta remain to be elucidated.

 


Research project:


Our research project is based on this recently published work, and is split into three parts.
Aim 1: Characterising the identity of aorta-emerging HSCs
We developed different tools allowing the long term fate of cells: live imaging, cell transplantation, cell tracing and parabiosis (see movie 2).


Movie 2. Cross-colonisation by primitive myeloid and erythroid cells in a gata1:dsred (right) // pu1:gfp (left) parabiote pair from 18.5–52 h.p.f. Demy et al, Nature Methods, 2014.


Aim 2: Regulatory pathways that govern the EHT process
The second part focuses on deciphering the regulatory pathways that govern the EHT process.

Aim 3: Aortic mechanical deformation and the EHT process
In collaboration with Andrea Parmeggiani (Biological Physics and Systems Biology, DIMNP - UMR 5235 CNRS/UM2/UM1 and Complex Systems & Nonlinear Phenomena, Laboratoire Charles Coulomb, UMR5221 CNRS/UM2, Montpellier II University), we are studying the relationship between aortic mechanical deformation and the EHT process (see movie 3).


Movie 3. Time-lapse confocal imaging of a zebrafish embryo from 1 to 4 days post fertilization, showing the correlation between aorta radial expansion then constriction and the period of endothelial to hematopoietic transition (EHT) of aorta floor cells Kissa & Herbomel, Nature, 2010.

 

Main publications

Primitive macrophages control HSPC mobilization and definitive haematopoiesis. Travnickova J, Tran Chau V, Julien E, Mateos-Langerak J, Gonzalez C, Lelièvre E, Lutfalla G, Tavian M, Kissa K.
Nat Commun. 2015 Feb 17;6:6227.

Choorapoikayil S, Kers R, Herbomel P, Kissa K, den Hertog J.
Pivotal role of Pten in the balance between proliferation and differentiation of hematopoietic stem cells in zebrafish.
Blood. 2014 Jan 9;123(2):184-90.

Faucherre A, Kissa K, Nargeot J, Mangoni M, Jopling C.
Piezo1 plays a role in erythrocyte volume homeostasis.
Haematologica. 2013 Jul 19.

Demy DL, Ranta Z, Giorgi JM, Gonzalez M, Herbomel P, Kissa K.
Generating parabiotic zebrafish embryos for cell migration and homing studies.
Nat Methods. 2013 Mar;10(3):256-8.

Renaud O, Herbomel P, Kissa K.
Studying cell behavior in whole zebrafish embryos by confocal live imaging: application to hematopoi-etic stem cells.
Nat Protoc. 2011 Nov 10;6(12):1897-904.

Kissa K, Herbomel P.
Blood stem cells emerge from aortic endothelium by a novel type of cell transition.
Nature. 2010 Mar 4;464(7285):112-5.

Levraud JP, Disson O, Kissa K, Bonne I, Cossart P, Herbomel P, Lecuit M.
Real-time observation of listeria monocytogenes-phagocyte interactions in living zebrafish larvae.
Infect Immun. 2009 Sep;77(9):3651-60.

Kissa K, Murayama E, Zapata A, Cortés A, Perret E, Machu C, Herbomel P.
Live imaging of emerging hematopoietic stem cells and early thymus colonization.
Blood. 2008 Feb 1;111(3):1147-56.

Le Guyader D, Redd MJ, Colucci-Guyon E, Murayama E, Kissa K, Briolat V, Mordelet E, Zapata A, Shinomiya H, Herbomel P.
Origins and unconventional behavior of neutrophils in developing zebrafish.
Blood. 2008 Jan 1;111(1):132-41.

Murayama E, Kissa K, Zapata A, Mordelet E, Briolat V, Lin HF, Handin RI, Herbomel P.
Tracing hematopoietic precursor migration to successive hematopoietic organs during zebrafish de-velopment.
Immunity. 2006 Dec;25(6):963-75.