Exploring neutrophil heterogeneity in health and cancer

Interactions between cancer cells and their microenvironment are critical for tumour initiation, growth and metastasis. Consequently, a greater understanding of these interactions may offer novel opportunities for therapy. Neutrophils are innate immune cells that form a component of the tumour microenvironment. Both pro- and anti-tumour effects of neutrophils are reported in the literature, suggesting context-dependent complexity in neutrophil-tumour interactions. This complexity is further deepened by recent reports demonstrating that neutrophil populations are more heterogeneous than previously appreciated, but it is unclear what significance this may have in a cancer context. This thesis therefore aimed to investigate how tumours perturb neutrophil heterogeneity and the potential functional consequences of this perturbation for neutrophiltumour interactions. I first used mass cytometry to perform high-dimensional analysis of mouse neutrophil surface marker expression across tissues and control and tumour-bearing contexts, using the MMTV-PyMT mouse model of breast cancer. This revealed a number of neutrophil populations defined by their differential expression of surface markers, with shifts in the populations observed between control and tumour-bearing mice. In particular, a population of neutrophils defined by low expression of L-Selectin (CD62L- ) expands in the bone marrow, circulation and periphery of tumour-bearing mice. This phenotype was observed in a number of other mouse cancer models. Analysis of nuclear morphology found that CD62Lneutrophils have a more hyper-segmented nucleus than CD62L+ neutrophils, suggesting potential functional differences between these populations. However, a range of functional assays did not identify differences between the two populations. Instead, comparison of the total neutrophil population from control mice and tumour-bearing mice discovered differences in functionality. Most significantly, when comparing neutrophils from control and tumour-bearing mice I found changes in kinase activity, reactive oxygen species and in vitro interactions with cancer cells, indicating broad shifts in neutrophil phenotype during the switch from a healthy state to cancer

Current standards for the storage of human samples in biobanks

Biobanks are diverse in their design and purpose; the idea of fully harmonizing historical and future biobanks is unaffordable and unfeasible. Biobanks should focus their efforts instead on developing and maintaining high-quality collections of samples capable of providing a wide range of biological information using processes that minimize introduced variability. A full data audit trail on sample processing, archiving, and quality control procedures should also be provided. This should enable the data derived from biobanks to contribute as part of wider collaborative efforts with other similar resources

Comparison of DNA quantification methodology used in the DNA extraction protocol for the UK Biobank cohort

Correlation between UV/Vis (Trinean) and PicoGreen® methods of DNA quantification for DNA concentration < 60 ng/μL. Correlation plot showing all data points (442,859 samples) where DNA concentration measured via UV/Vis is < 60 ng/μL. Samples quantified via one method < 2 ng/ μL and with another method > 10 ng/μL are highlighted with a black cross. (PDF 166 kb

S-Adenosyl homocysteine and DNA ends stimulate promiscuous nuclease activities in the Type III restriction endonuclease EcoPI

In the absence of the methyl donor S-adenosyl methionine and under certain permissive reaction conditions, EcoPI shows non-specific endonuclease activity. We show here that the cofactor analogue S-adenosyl homocysteine promotes this promiscuous DNA cleavage. Additionally, an extensive exonuclease-like processing of the DNA is also observed that can even result in digestion of non-specific DNA in trans. We suggest a model for how DNA communication events initiating from non-specific sites, and in particular free DNA ends, could produce the observed cleavage patterns

Maintaining a sense of direction during long-range communication on DNA

Many biological processes rely on the interaction of proteins with multiple DNA sites separated by thousands of base pairs. These long-range communication events can be driven by both the thermal motions of proteins and DNA, and directional protein motions that are rectified by ATP hydrolysis. The present review describes conflicting experiments that have sought to explain how the ATP-dependent Type III restriction–modification enzymes can cut DNA with two sites in an inverted repeat, but not DNA with two sites in direct repeat. We suggest that an ATPase activity may not automatically indicate a DNA translocase, but can alternatively indicate a molecular switch that triggers communication by thermally driven DNA sliding. The generality of this mechanism to other ATP-dependent communication processes such as mismatch repair is also discussed

The Potential of Pressurised Water Reactors to Provide Flexible Response in Future Electricity Grids

The electricity market is undergoing significant change with the increasing deployment of Variable Renewable Energy Sources (VRES) and the adoption of policies to electrify transport, heating and industry, which will continue to increase demands on all conventional power plants including nuclear. The increase in VRES also puts additional emphasis on services such as inertia and frequency response that only conventional plants, including nuclear, are readily able to meet. This study discusses what factors limit the ability of nuclear power plants to provide flexible response and how the UK nuclear power plants might be affected by the changes in future demand profiles. The study focuses on what impact there will be on current Pressurised Water Reactor (PWR) plants, though it also considers Small Modular Pressurised Water Reactor plants which might offer benefits with respect to improved power manoeuvrability. The main finding is that the most important attribute is the minimum power level for long-term operation, followed by the speed at which the plants can be brought online (that is, both start-up rate and ramp rate during power operation). With respect to both of these attributes, new build future PWR plants could potentially achieve large and rapid power changes by dumping part of the steam directly into the condenser, bypassing the steam turbine. Discussions with plant operators highlighted that there is currently limited demand for flexible operation in the UK from nuclear plants when other power plants are readily available to partake in flexible operation. The lack of any requirement for nuclear plants to operate flexibly means that the UK lags behind France, for example, which has much more experience in nonstationary operation of nuclear power plants. The paper also draws attention to the fact that with increasing VRES, there will be fewer plants able to provide rotational inertia and therefore more emphasis on the role the remaining plants (which include nuclear) can play in maintaining grid stability.</jats:p