Differential fMet-Leu-Phe- and Platelet-activating Factor-induced Signaling Toward Ral Activation in Primary Human Neutrophils

We have measured the activation of the small GTPase Ral in human neutrophils after stimulation with fMet- Leu-Phe (fMLP), platelet activating factor (PAF), and granulocyte macrophage-colony stimulating factor and compared it with the activation of two other small GTPases, Ras and Rap1. We found that fMLP and PAF, but not granulocyte macrophage-colony stimulating factor, induce Ral activation. All three stimuli induce the activation of both Ras and Rap1. Utilizing specific inhibitors we demonstrate that fMLP-induced Ral activation is mediated by pertussis toxin-sensitive G-proteins and partially by Src-like kinases, whereas fMLP-induced Ras activation is independent of Src-like kinases. PAFinduced Ral activation is mediated by pertussis toxininsensitive proteins, Src-like kinases and phosphatidylinositol 3-kinase. Phosphatidylinositol 3-kinase is not involved in PAF-induced Ras activation. The calcium ionophore ionomycin activates Ral, but calcium depletion partially inhibits fMLP- and PAF-induced Ral activation, whereas Ras activation was not affected. In addition, 12-O-tetradecanoylphorbol-13-acetate-induced activation of Ral is completely abolished by inhibitors of protein kinase C, whereas 12-O-tetradecanoylphorbol- 13-acetate-induced Ras activation is largely insensitive. We conclude that in neutrophils Ral activation is mediated by multiple pathways, and that fMLP and PAF induce Ral activation differently

Early detection of cardiovascular disease:From identifying those at risk to making a timely diagnosis

Cardiovascular disease (CVD) is often diagnosed late: at the time of diagnosis, serious and permanent damage has often already occurred. Since evidence-based therapies exist for heart failure (HF), atrial fibrillation (AF) and coronary artery disease (CAD), early detection of these diseases is crucial to ensure that patients receive timely treatment to prevent severe disease.In Part I of this thesis, we studied who is at risk of which types of CVD: we demonstrated that patients with diabetes, COPD and screen-detected AF are at risk of developing various types of CVD, particularly HF.In Part II, we developed a brief questionnaire about symptoms of CVD that may help identify patients with previously unrecognized CVD. This questionnaire was included in the RED-CVD trial, in which we demonstrated that a proactive, stepwise diagnostic strategy increased the number of new diagnoses of HF, AF and CAD in primary care patients with diabetes and/or COPD. In Part III, we demonstrated that relative fat mass (RFM) is a suitable marker of cardiometabolic risk that that performs at least similarly, yet has several important advantages compared to established obesity measures such as BMI and waist circumference. This thesis highlights the importance of prevention and early detection of CVD – particularly HF – in high-risk populations, including patients with obesity, diabetes, COPD and AF. Although effects on patient outcomes remain to be studied, our results demonstrate that proactive diagnostic strategies are feasible and can improve early detection of CVD in primary care

Crosstalk between cAMP and MAP Kinase Signaling in the Regulation of Cell Proliferation

Hormonal stimulation of cyclic adenosine monophosphate (cAMP) and the cAMP-dependent protein kinase PKA regulates cell growth by multiple mechanisms. A hallmark of cAMP is its ability to stimulate cell growth in many cell types while inhibiting cell growth in others. In this review, the cell type-specific effects of cAMP on the mitogen-activated protein (MAP) kinase (also called extracellular signal-regulated kinase, or ERK) cascade and cell proliferation are examined. Two basic themes are discussed. First, the capacity of cAMP for either positive or negative regulation of the ERK cascade accounts for many of the cell type-specific actions of cAMP on cell proliferation. Second, there are several specific mechanisms involved in the inhibition or activation of ERKs by cAMP. Emerging new data suggest that one of these mechanisms might involve the activation of the GTPase Rap1, which can activate or inhibit ERK signaling in a cell-specific manner

Broken biosecurity?: veterinarians’ framing of biosecurity on dairy farms in England

There is seen to be a need for better biosecurity – the control of disease spread on and off farm – in the dairy sector. Veterinarians play a key role in communicating and implementing biosecurity measures on farm, and little research has been carried out on how veterinarians see their own and farmers’ roles in improving biosecurity. In order to help address this gap, qualitative interviews were carried out with 28 veterinarians from Royal College of Veterinary Surgeon farm accredited practices in England. The results were analysed using a social ecology framework and frame analysis to explore not only what barriers vets identified, but also how vets saw the problem of inadequate biosecurity as being located. Veterinarians’ frames of biosecurity were analysed at the individual, interpersonal and contextual scales, following the social ecology framework, which see the problem in different ways with different solutions. Farmers and veterinarians were both framed by veterinarians as individualised groups lacking consistency. This means that best practice is not spread and veterinarians are finding it difficult to work as a group to move towards a “predict and prevent” model of veterinary intervention. But diversity and individualism were also framed as positive and necessary among veterinarians to the extent that they can tailor advice to individual farmers. Veterinarians saw their role in educating the farmer as not only being about giving advice to farmers, but trying to convince the farmer of their perspective and values on disease problems. Vets felt they were meeting with limited success because vets and farmers may be emphasising different framings of biosecurity. Vets emphasise the individual and interpersonal frames that disease problems are a problem on farm that can and should be controlled by individual farmers working with vets. According to vets, farmers may emphasise the contextual frame that biosecurity is largely outside of their control on dairy farms because of logistical, economic and geographical factors, and so some level of disease on dairy farms is not entirely unexpected or controllable. There needs to be a step back within the vet-farmer relationship to realise that there may be different perspectives at play, and within the wider debate to explore the question of what a biosecure dairy sector would look like within a rapidly changing agricultural landscape