2020 Student Cohort

Skin constitutes the first line of defence against disease-causing organisms, but is susceptible to injury such as burns. Several hundred thousand people in the UK sustain burns every year, and hundreds are fatal. Most dangerously, the loss of the barrier to pathogens means that patients can succumb to overwhelming infection – sepsis – within a few weeks after injury.
The gold standard for treatment is to remove the dead tissue surgically and resurface with new skin. Full-thickness skin grafts are rarely used because there are few donor sites on the body surface, and this thicker skin is less likely to pick up a blood supply at its recipient site. There are several commercially available skin substitutes to replace the epidermis/dermis, or both; however, these options are expensive and have not yielded any acceptable long-term clinical result yet.
Recent advances utilise cell-based techniques – extracting biopsies from the patient’s skin, isolating keratinocytes and expanding them in laboratory culture. There are important limitations; time constraints, difficulties in achieving sufficient numbers for clinical application, and the possibility of instigating neoplastic change due to components added during the cell culture process.
This project aims to overcome some of these challenges by implementing novel cell culture techniques in specialised bioreactors. The key aim is to advance culture techniques that allow an optimal, accelerated growth of keratinocytes from an autograft. These expanded keratinocytes will potentially be applied to seal burn wounds, reducing morbidity and mortality after large injuries.

The ability to maintain haematopoietic stem cell (HSC) populations in vitro would provide large societal benefit. Haematopoietic cancers, such as leukaemia, arise from genetic alterations in HSCs. The current approach is to kill malignant HSCs and then use bone marrow transplantation to provide long-term reconstituting (LTR) HSC populations to regenerate the blood system. Bone marrow transplantation is, however, a one donor – one patient therapy and donors are in urgent demand. There have been many attempts to maintain LTR-HSCs in vitro, out of the bone marrow niche. However, out of their niche HSCs either die or expand rapidly losing long-term reconstituting phenotype as they grow; these LTR HSCs are critical to provide to patients as they are the cells required to engraft and to repopulate the marrow to produce new blood cells.
Working with Manchester BIogel’s synthetic Peptigel® hydrogels, this project will focus on bioengineering in vitro HSC niches. Using physical cues such as stiffness and solid-phase growth factor presentation, and biological cues such as other cells, we will develop microenvironments where LTR-HSC populations can be maintained in the laboratory. Being able to conserve LTR-HSC number in vitro is important as it allows us to study and manipulate the cells. For example, in this new project, we will develop CRISPR approaches to edit the stem cells. This is important as e.g. chronic myloid leukaemia is typified by the BCR-ABL mutation. If we can edit out and correct such mutations, then we can envisage ways to repair patients’ cells ex vivo and then provide them back to the patients – removing the cancer and regenerating a disease free blood system with their own cells.

The motor functions of the intestine including rhythmic contractions and relaxations allow transferring and processing of chyme. Each of these functions generates a shear stress on the epithelium that varies in magnitude. How such shear stress affects epithelial processes such as crypt fission and fusion, and tissue compartmentalization as well as the mechanical role of mucus is not well understood. To address these questions, here we developed a computational fluid dynamics model that integrates intestinal mucus, chyme, intraluminal pressure, and crypt geometry to predict the time-space mosaic of shear stress. We combine this model with an organ-on-a-chip device that could allow traction force microscopy to map cellular forces. Our computational data show that the intestinal mucus may significantly reduce the amount of shear stress applied to the crypts with significant apico-basal variation and increase chyme velocity. Using our on-chip model we aim to explore the interplay among shear stress, crypt dynamics, and mucus.

Flow cytometry is a cell analysis technique which can make measurements of cells in solution as they pass by the instrument’s laser at rates of 10,000 cells per second (or more). Because of its speed and ability to scrutinize at the single-cell level, flow cytometry offers the cell biologist the statistical power to rapidly analyze and characterize millions of cells. An extension of flow cytometry is the sorting module, allowing physical separation of the cells based on a real-time reading in the analysis module, leading to the effective isolation of sub-populations based on the extent and presence of a specific cellular marker. The current method of single cell sorting are based on fluorescent tags, directed against specific molecular targets. While this approach provides high selectivity and specificity, it is not always possible to find a unique label for a specific biological condition. Recently this limitation has been tackled by means of label-free cytometers, measuring physical parameters of the cells such as the dry mass or the elasticity. The proposed project aims at leveraging the full power of this concept, integrating the analysis with the sorting, and addressing single cell label-free sorting. The new technology will be applied to the isolation of stem cells for regenerative medicine and to the identification of Leishmania infections in the blood. Development of label free techniques will increase the power of the drug discovery pipeline for many conditions from infection, to stem cell therapies to cancer treatments.

Tuberculosis (TB) is a disease caused by the organism Mycobacterium tuberculosis which kills 1.5 million people each year and is thought to infect one quarter of the world’s population. Treatment of TB is becoming ever more difficult with the emergence of antibiotic resistance, and this has propagated an urgent unmet need to discover new antibiotics with novel mechanisms of action. The development of high throughput microfluidics has revolutionized the way in which we can discover new antibiotics. In this collaborative, multidisciplinary project, we aim to optimize one such platform for use in screening for antibiotics with activity against M. tuberculosis.
The successful candidate will be trained by and work with Sphere Fluidics to engineer a platform that is optimized for Mycobacterial drug discovery, controlling various aspects of bacterial culture to produce a platform that is physiologically relevant and suitable for high throughput drug discovery. Subsequently, they will transfer this platform to Aston University to the Mycobacterial Research Laboratory for use in screening a highly valuable set of anti-TB drugs, to discover their activity and investigate their mechanisms of action. The selected student will benefit from cross-disciplinary training in both engineering and biological sciences and have the opportunity to apply that training to develop technology that improves the discovery and development of novel antibiotics.

The inflammatory response following traumatic spinal cord injury induces formation of a fibrotic scar at the lesion site. Many questions remain regarding the extracellular matrix deposited after spinal cord injury and the inhibitory molecules that are upregulated. This project will focus on matrix deposition by glial cells and leptomeningeal fibroblasts. A novel culture system will be utilized and combined with anti-fibrotic compounds for spinal cord repair. An important focus of this project will involve analysis of cell glycosylation, as glycosylation has been shown to play an important role in many biological processes including tissue repair.

This research aims to establish an in vitro immune model of the human T-cell response to functionalised osteogenic biomaterials which present surface restricted growth factors to mesenchymal stem cells (MSCs), thereby inducing osteoblast differentiation. The biomaterial has proven capable of promoting bone regeneration across critical defects in murine models, however human tolerance remains unknown. MSCs have been widely used in this field to cellularise biomaterials, due their perceived lack of immunogenic properties. However, over longer time periods, the allogenic MSCs within this biomaterial differentiate into mature osteoblasts, which express surface markers capable of triggering adaptive immune responses in humans. Concern therefore exists about the potential for adverse adaptive immunity to this mature, differentiated construct, negatively impacting its tissue integration and regenerative profile. This research will characterise and define the human adaptive immune response to this biomaterial and the model will then be used to trial different therapeutic adaptations to promote favourable immune interaction. The ultimate goal is to have a functionalised biomaterial that promotes a pro-repair/ pro-tolerance environment, in addition to its bone regeneration properties, for use in humans at sites of bone injury.

Chronic intestinal pseudo-obstruction (CIPO) indicate a class of rare gastrointestinal disorders sharing the same symptoms (mainly abnormalities affecting the involuntary, coordinated muscular contractions) but for which a clear and definite genetic marker has not been identified yet. Mistreatment associate to lack of diagnosis is a burden that might hardly influence the development of the disease. Nevertheless, cells extracted from patients appear to share similar phenotype and the proposed study aims at characterizing the physical phenotype (mechanics, morphology) with high throughput to identify a label-free marker that might be prognostic for the disease.

Globally, >295,000 women are diagnosed with ovarian cancer (OC) each year and the 5-year survival rate is unacceptably high at 30-50%. There are three main reasons for this high mortality: late detection, resistance to platinum-based therapies and insufficient concentration of the therapeutic at the target. This project aims to address the latter two of these.
In this project we focus on cell-based immunotherapy for OC. Natural Killer (NK) cells have been shown to kill cancer cells and reduce tumor burden by our group and others in pre-clinical studies of OC, and furthermore have the potential to be used as an off-the-shelf therapeutic. Production of Chimeric Antigen Receptor (CAR) NK cells, targeted to receptors commonly expressed on the surface of OC cells, is hypothesised to further increase this cytotoxicity. Despite a hypothesised enhancement in cytotoxicity modified NK cells still need to concentrate in the peritoneum to exert a potent therapeutic effect. We propose an implantable device for repeated site-specific targeted delivery of immunotherapy. This strategy aims to prolong and evenly distribute the therapeutic agent to the target region while also considering repeated minimally invasive delivery of therapy, ease of placement/insertion, patient comfort and effect on the therapy. The PhD student will develop this concept by designing, testing, optimising and manufacturing CAR-NK delivery devices for both mouse and human/porcine implantation with a focus on translation to the clinic.

Research Need: 3D spheroid models greatly enhance our ability to develop ex vivo models of both disease and normal physiology. Much advancement has been made with the use of scaffolds and hydrogels. Once of the next challenges is to create a 3D spatial atlas of the cell-cell interactions that occur with spheroids and in turn human tissues. This will allow is to visualize in 3D the signals that are activated within cells as influenced by their proximity to different cell types, hypoxic regions and nutritional availability and/or drug responses. Suguna’s PhD project will use 3D tumour epithelium-stromal versus 3D normal epithelium-stromal spheroid models to investigate (1) the impact of stromal cells on tumour metastasis, and (2) to develop new methods to visualize protein expression, hypoxic regions, cell morphology while maintaining tissue architecture so that a 3D protein-cell map of the spheroid can be built. This will greatly enhance our ability to confirm the relevance of 3D models to real patient pathological conditions. Research from project supervisor Dr. Sharon Glynn’s laboratory have shown that bone metastatic prostate cancer cells preferentially induce differentiation of healthy MSC to a pro-inflammatory stromal cell that promotes PrCa proliferation and invasion (Ridge et al, 2018). In collaboration with the NIH we are developing CODEX based spatial digital pathology to assess the protein patterns and cellular milieu in triple negative breast cancer patient specimens which we will apply to our tumour-stromal 3D models. Using these models Suguna’s PhD will address the following aims:
• Establishment of 3D spheroid models of normal/tumour – stromal cell co-cultures
• Development of multiplexing imaging system that preserves the 3D cellular orientation.
• Investigation of cellular dynamics of response to therapy in 3D spheroid models
• Explore the exploitation of the immunosuppressive properties of MSCs by tumour cells for immune tolerance and immune suppression during tumour metastasis & chemotherapeutic response.

Cells need three dimensional environments to grow and to differentiate, and thus efforts have been made to engineer materials that can display adhesion ligands, growth factors and that have controlled properties. ECM-derived matrices, such as Matrigel, are currently widely used and support the growth and function of a wide variety of cell types in vitro. However, Matrigel is an undefined mixture of ECM proteins and growth factors (GFs), with undefined composition undefined, lack of control of mechanical properties and to lot-to-lot variability. Matrigel is widely used as it contains GFs that provide biological activity not yet achieved by synthetic systems. Given this, there is a pressing need to design synthetic matrices that can fulfil the roles of the ECM.
This project will provide further functionality to Manchester Biogel’s synthetic Peptigel® hydrogels (MBG) by incorporating solid-phase presentation of growth factors into the hydrogels. This will enhance bioactivity of MBG gels to target (a) stem cell differentiation (e.g. BMP-2 to promote osteogenesis) and maintenance of stem cell phenotypes. The PhD project will work to engineer this new family of self-assembling hydrogels that have the potential to recruit and present growth factors. We will also show proof of concept of biological functionality by triggering stem cell differentiation and maintenance of phenotypes.

When producing in vitro models, animal derived matrices such as collagen, lack reproducibility and have a high to batch-to-batch variation. Synthetic peptide hydrogels may offer an effective alternative. These hydrogels are relatively cheap to manufacture, easy to use, and have a defined chemical composition making them highly reproducible. This project focuses on developing novel peptide hydrogels for cell culture. Cell Guidance systems have developed PODs technology which can be attached to the cell scaffolds to allow sustained release of growth factors. This project will be highly multidisciplinary; involving aspects of synthesis, gel formation, characterisation, and using the gels as cell scaffolds.

The project will include an industry placement with the Cell and Gene Therapy Catapult at Guys Hospital, London Bridge, London. The placement will enable the student to gain an understanding of the roles of process development and analytic/data analysis teams. They will apply their knowledge and the technology generated to a range of scalable bioreactor cell cultures and to gain a deeper understanding of typical industry workflows, by working with one of the leading translational cell and gene therapy institutes in the world. It will also enable the student to understand the wider commercial and R&D ecosystem that underpins the regenerative medicine sector.

Droplet microfluidics have shown exceptional promise for cell and tissue engineering, single cell study, cell testing and sorting. This typically involves complex experiments, which require a microfluidic platform to be able to perform multiple functions on each droplet. In this project, we will focus on different cell types and examine their behavior when exposed to different environments in order to understand how cells can be influenced and how to cause desired differentiations. This application if highly relevant for cell engineering, as well as medical exposure studies.
The student will exploit state-of-art microfluidic techniques to design and fabricate new prototypes and devices to probe the response of multiple cell types to different environments. This encompasses the use of established methods like soft-lithography and molding but also requires gaining expertise in more advanced techniques combining microfluidics with optical, dielectrophoretic and acoustical setups. Droplet based microfluidics is essential and the key and the student will become an expert in all aspects of this technology.

This project will allow us to both probe cell behavior in controlled environments and to look for beneficial cellular responses.