*EPSRC* Analysis of mechanical forces in embryonic development of the haematopoietic system

Haematopoietic stem cells (HSCs) lie at the foundation of the adult haematopoietic hierarchy and, through a fine balance between self-renewal and differentiation, maintain haematopoiesis throughout an animal’s lifespan. Elucidating mechanisms underlying HSC development is of fundamental significance with clinically relevant implications. The regulatory interactions between stem cells and their niches is a key issue in stem cell biology and physical forces are an important factor known to be involved in stem cell regulation. We have shown previously that HSCs emerge in the ventral domain of the embryonic dorsal aorta within the area called the aorta-gonad-mesonephros (AGM) region. The AGM region is subjected to compression and stretching forces during morphogenesis. Notably, physical stiffness is an important factor playing a role in HSC physiology in adult bone marrow. Using enforced reaggregation of AGM cells, we have been able to recapitulate HSC development in vitro, which supports our hypothesis that physical forces play a role in this process. We have also revealed a complex spatially polarised signalling milieu in the AGM region which is important for HSC development. We hypothesise that uneven physical tensions play a role in patterning expression of genes which underlie HSC development in the AGM niche.


The main objective of this project is to establish whether physical forces generated during embryogenesis play a role in patterning polarized gene expression in the AGM niche. Specific aims are: Aim 1) to spatially and temporally map AGM stiffness using bespoke high resolution optical coherence elastography tomography (OCT) microscope. Briefly, the low coherent light from a superluminescent diode is collimated into a custom scanning head and optics into an inverted microscope. The back propagating spectral interferences will then be resolved on a spectrometer to retrieve in-depth microstructural information. In the elastography mode, images will be acquired at the same location to record change in displacement (phase) to subsequently compute a strain map indicative for tissue stiffness. OCT method can uniquely map tissue elasticity with high resolution (5 μm) at 2mm depth. This will allow us to image and map mechanical properties of the AGM region for the first time in intact embryos, avoiding potential artefacts associated with dissection. Stiffness conditions will be then modelled in culture and effects studied at gene expression levels. Aim2) to determine impact of cell compression on gene expression. Tissue compression will be studied using a piezoelectric loading system which will then be used to replicate similar mechanical conditioning in vitro, e.g. by compression of cell-seeded gels. We will study effects of compression and stretching on various cell populations from the AGM niche to monitor modulation of expression of transcription factors and secreted molecules involved in HSC development. Bioinformatics analysis will be employed to process the data.

During PhD study, the student will be trained in analysis of physical properties of biological tissues and cell deformation complemented by methods of biological analysis such as flow cytometry cell sorting, cell culture, RT-PCR, RNA-Seq and confocal microscopy.

This is a collaborative cross-college project between two laboratories with complementary expertise necessary for this project:

1st supervisor: stem cell biologist, Prof. A. Medvinsky is an expert in embryonic development of HSC:

http://www.crm.ed.ac.uk/research/group/ontogeny-haematopoietic-stem-cells

2nd supervisor: physicist Dr. P. Bagnaninchi, expert in non-invasive analysis of tissue mechanics http://www.crm.ed.ac.uk/research/associate/cell-and-tissue-monitoring

He is the recipient of EPSRC EP/P031250/1 grant that provides essential equipment for this PhD project.