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Reference : UMR8227-STEEGE-002
Workplace : ROSCOFF
Date of publication : Thursday, July 23, 2020
Scientific Responsible name : Stéphane EGEE
Type of Contract : PhD Student contract / Thesis offer
Contract Period : 36 months
Start date of the thesis : 1 November 2020
Proportion of work : Full time
Remuneration : 2 135,00 € gross monthly
Description of the thesis topic
The hydration state of RBCs as an important parameter that ensures the functions of the cell in the blood
This project will aim at understanding the functions of ion channels within the RBC membrane, particularly in the context of RBC circulation and end of life. It will rely on a diversity of techniques and approaches (from the diversity of channel expression in progenitors to their involvement in aging and deformability tests) and also include RBC membrane permeability modelling to decipher the involvement of ion channels at these crucial life steps. It will bring implications useful for a better comprehension of pathologies linked to cation permeability, for storage lesions and for the production of cultured RBCs in Bioreactors.
State of the art:
Erythrocytes are highly specialized and atypical cells, as evolution has led to the complete loss of intracellular organelles in mature red cells to optimize respiratory functions. Nonetheless, human erythrocytes are far from being a simple bag of hemoglobin, and have to maintain their deformability, volume and intern homeostasis to accomplish with the best efficiency their function of gas transporters.
Indeed, RBCs are in continuous flow, with high and rapid variations of their environment and frequent deformability stresses. This deformability is linked to their unique cytoskeleton and surface / volume ratio, and to a maintenance of the volume; it is tested very regularly in the spleen. During their 120 days
of life, RBCs undergo an increase in cell density, reduction of cell volume and are eventually removed from the circulation in the spleen and recycled by macrophages. Membrane ion permeability is a key parameter for the hydration and volume preservation, and mammalian red cells have long been a paradigm to study membrane transport. Erythrocyte membranes are characterized by an important anionic permeability, allowing the equilibrium for chloride ions and facilitating the transport of CO2 via the Jacobs-Stewart cycle. In contrast, cationic permeability represents a threat linked to the colloido-osmotic pressure exerted by the high protein content of the cell.
Surprisingly erythrocytes possess not only anionic channels, but also a repertoire of cationic channels whose full description is still lacking, including notably the Gárdos channel (KCNN4, hSK4) (1), selective for K+ ions, PIEZO-1 (2), a mechanosensitive cation channel or the NMDA Receptor (3). The complete repertoire of ion channel is still under investigation, and so is it for their physiological roles (4, 5). A certitude is that dysregulations of cation channel activity (linked to mutations either on the channels or on partner proteins) are involved in multiple pathologies of the RBC (6, 7) and that these channels are linked to senescence of RBCs (8). The roles of these channels has almost always been studied in static conditions, using electrophysiological techniques.
This project aims to:
1) Fully describe the repertoire of ionic channels from early stages of terminal differentiation to mature RBCs.
2) Understand their physiological role in mature RBCs once they are released within circulation.
3) Unravel their involvement in senescence processes in health and disease conditions.
4) Ultimately all the data will be used to feed a quantitative model (already written and yet improved (9, 10)) for diagnostic predictive purposes, for blood preservation in vitro and for the upscaling of cultured RBC production in bioreactors.
- To determine the repertoire of ion channels in RBCs, we have started a study focusing on the expression of channel proteins in terminal erythropoiesis RBC. All tools are now ready and we have already acquired first results. This part of the project includes erythropoiesis ex vivo, qPCR, Western Blot, and immunofluorescence assays.
- Monitoring of ion channel activity linked to cell flow: We will perform patch clamp studies / CCCP monitoring of membrane potential before and after several stresses implying cell deformation: microsphiltration (11), Shear flow, spleen-on-chip or microfluidic in collaboration with partners from the Evidence ITN.
- model of membrane permeability: the model was originally developed by Lew and Bookchin (9), and was recently update
The EVIDENCE ITN is an EU funded international consortium of academic research centres, diagnostic labs, blood supply centres, and small companies that combines basic and translational research to improve diagnostic and therapeutic approaches on erythrocytes and during erythropoiesis. More detailsabout the consortium can be found at the EVIDENCE website https://evidence.eurice.eu/.
The MSC ITN programme offers highly competitive and attractive salary. The successful candidate will receive a salary in accordance with the regulations for MSC Fellows/early stage researchers (ESRs).
Exact salary will be confirmed upon appointment but will involve a generous Living Allowance and monthly mobility allowance depending on the successful candidate's personal family situation.
This PhD position will likely provide experience of erythroid culture, molecular and cell biology, imaging, flow cytometry, microfluidics and electrophysiology. In addition to their individual scientific projects, all fellows benefit from further continuing education, which includes internships and secondments, a variety of training modules as well as transferable skills courses and active participation in workshops and conferences.
Constraints and risks
Applicants should provide their complete applications in one single PDF
Each application should provide a cover letter, a CV detailing scientific experience to date and copy of qualification certificates and provide a contact information of appropriate 2 references. For further details of ESR6 or other projects, please refer to the website: https://evidence.eurice.eu/.
The application closing date is 15th September 2020. Candidates who have been selected for interview will be contacted for Skype interviews by the end of September 2020. PhD have to start before 15th December 2020.
The candidate must have a Master of Science and not have resided or carried out his main activity (e.g. work, studies) in France for more than 12 months in the 3 years immediately before the recruitment date.
The doctoral student will work in close collaboration with two engineers from the team and another doctoral student likely to be recruited in a year's time. The thesis writing expectations will be a point of vigilance during recruitment.
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