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Primary cilia and cellular senescence 05 Sep 2017

Somatic cells undergo senescence after a finite number of divisions, indefinitely arresting their proliferation. The mechanisms of cellular senescence are not well understood, although DNA damage signalling is one major cause. We have found that senescent human fibroblasts have increased frequency and length of primary cilia, antenna-like structures that sense and transduce various extracellular signals, notably Hedgehog. Here we propose to test the hypothesis that primary ciliation contributes to cellular senescence. We will use genome editing in primary cells to ablate CEP164, which is required for primary ciliogenesis, and then follow the kinetics of cellular senescence in the knockout population. We also propose to examine cilium-controlled signalling pathways to determine how they are affected during the initiation of senescence programmes. We will perform competitive co-culture experiments between ciliated and non-ciliated populations to define how ciliation capacity directs senescence in mixed populations. Together, the proposed experiments will test a novel cellular mechanism of senescence, a process of great significance in human health and the normal aging process.

Amount: £51,168
Funder: The Wellcome Trust
Recipient: National University of Ireland Galway

Evaluating the combination of Oncotype DX and Neutrophil/Leukocyte Ratios as prognostic indicators of chemotherapy response in breast cancer. 27 Apr 2017

This research will study the effect of the individual’s immune system on their reoccurrence of breast cancer after chemotherapy. This would be a correlation between the Neutrophil to Lymphocyte Ratio (NLR) and the Oncotype Dx score. Previous research (unpublished) has demonstrated the significance of the NRL in predicting breast cancer outcomes in HER 2 overexpressing and Luminal B breast cancer (accounting for approx. 20% of all breast cancers). We will extend our previous research to include all four cancer subtypes (Luminal A and Triple negative breast cancer). We aim to compare the immune system’s reaction to chemotherapy across all four cancer subtypes. A key goal is to determine if NLR predicts outcome or survival in Luminal A or TNBC. A further key goal is to determine if there is any predictive value of combining the NLR with Oncotype scores (in relevant cases).

Disease-specific quality of life in children with suppurative otitis media, including children with Down Syndrome and Autistic Spectrum Disorder 27 Apr 2017

Acute otitis media (AOM) is a bacterial or viral infection of the middle ear which may accompany an upper respiratory tract infection. AOM can occur at any age, however, it is most common between 3 months and 3 years old. At this age, the Eustachian tube is structurally and functionally immature. Diagnosis of AOM is made by clinical and physical examination with an otoscope. Otoscopic examination can show a bulging, erythematous tympanic membrane with indistinct landmarks and displacement of the light reflex. The treatment for AOM are analgesics such as paracetamol or ibuprofen and many cases resolve spontaneously. Antibiotics are frequently given. Antibiotics relieve symptoms quicker and may reduce the chance of residual hearing loss and labyrinthine or intracranial sequelae. With the emergence of antibiotic resistant organisms, it is recommended that a round of antibiotics be administered only for children at highest risk or for those with recurrent acute otitis media(RAOM). RAOM is defined as four or more episodes of AOM in a 6 month period. The aim of this project is to investigate the impact on the quality of life of children that present with recurrent acute otitis media(RAOM).

Haemocompatibility and biocompatibility of a polyurethane-based polymer for use in prosthetic heart valves 27 Apr 2017

Developing valves with efficient function and long-term durability without the need for anticoagulation therapy, coupled with the ability to be accommodated in many different types of patient, are the principal requirements of replacement heart valves. The current treatment options available do not fit well with these critieria. The overall aim of the project is to develop a biomimetic polymer alternative to pericardium heart valve leaflets with a more repeatable, consistent and predictable production process. Main research objective: To investigate the haemocompatability of the polymer valve. That is, to measure the blood's response to contact with the valve by measuring parameters such as thrombogenicity, activation of coagulation, blood cell countsand platelet activation. Secondary research objective: To carry out a comprehensive collaborative literature review with my supervisor to deisign an experiment that helps us to understand the blood's response to the polymer in a dynamic environment i.e. simulating the in vivo scenario of blood being pumped across the valve and therefore coming in contact with the valve while under stress. Time allowing, we will then carry out this experiment. We hypothesize that the polyurethane-based material will have equivalent haemocompatibility to that of a commercially available aortic heart valve replacement.

Functional characterisation of excitatory neurons 27 Apr 2017

Autism spectrum disorder (ASD) is a complex, neurodevelopmental disorder characterised by deficits in social and communication skills, language delay and repetitive or stereotypical behaviour. A variety of copy number variations have been identified that play a role in ASD, including the Neurexin1 gene (NRXN1) which encodes a family of presynaptic cell adhesion transmembrane proteins, vital for neurotransmission and synapse differentiation. It is widely accepted that neurexin and mutations disturb the balance between excitatory (alpha- and beta-nrxn) and inhibitory (mainly alpha-isoform) synaptic activity that is thought to be critical in the pathology in ASD. The seminal discovery that somatic cells can be reprogrammed to produce induced pluripotent stem cells (IPSCs) has revolutionised biological research and medicine. The ability to differentiate these IPSCs into neural cells offers a tremendous opportunity to study disease modelling associated with impaired neuronal cell biology. In this study we will characterise the functional firing properties of IPSc derived glutaminergic neurons differentiated from normal controls and patients carrying a NRXN1 mutation. Functional properties will be assessed by patch clamp examining voltage gated sodium and potassium channels and action potential firing properties under current clamp conditions. This work will be essential to aiding our understanding of ASD.

Biomaterial-enhancement of an ex vivo model of Parkinson’s disease 27 Apr 2017

Parkinson’s disease (PD) is a disorder of the aged and efforts at finding a cure have so far failed. In order to develop new therapies that halt or reverse the decline in brain function that occurs, researchers must find ways to better model the disorder in the lab. One approach makes use of organotypic rat brain slice cultures that retain many of the 3D features and properties of brains in vivo, but are easily manipulated in vitro. However, cultured slices often do not survive well and neurons of the substantia nigra pars compacta (SNpc), the main brain region implicated in PD pathology, are particularly vulnerable to culture conditions. The key aim of the proposed project is therefore to test the viability-enhancing properties of medium supplements, synthetic (poly(dimethylsilane), poly(glycolic acid), poly(lactic acid) and poly(vinyl chloride), poly(lactic-co-glycolic acid) and natural (collagen, hyaluronic acid, laminin, fibronection, fibrin, gelatin, chitosan, other brain extracellular matrix proteins) polymers. Cultured tissue viability will be monitored using LDH, MTT, live-dead assays, as well as immunohistochemical detection of the dopamine neuron marker tyrosine hydroxylase. The project will be carried out in the FitzGerald lab, within the Centre for Research in Medical Devices (CÚRAM) at NUI Galway.

Effect of Cholinergic Neurotransmission Challenge on Emotion-Related Attention in Bipolar Disorder 27 Apr 2017

A low dose of the acetylcholinesterase inhibitor, physostigmine will be used to challenge the cholinergic system while performing a functional MRI task of emotion inhibition in psychiatrically healthy participants and euthymic subjects with bipolar disorder. Prior to and immediately following the infusion of physostigmine, subjects will perform behavioural tasks of attention as well as rate their mood using the profile of mood states (POMS) and visual analogue scale (VAS) rating scales. The proposed project will assess an effect of cholinergic system challenge on mood and cognition and will compare effects between control and bipolar groups as well as across the cholinergic muscarinic type-2 receptor genotype (CHRM2 rs324650). We hypothesize that 1) the dose involved will not demonstrate measurable effects on mood, 2) will have detectable effects on performance of the task of emotion inhibition, and 3) the latter effect will be more pronounced in those of the genotype (TT) associated with reduced autoreceptor (M2) concentrations (the brakes on cholinergic neurotransmission). The findings of the project will feed into a larger analysis of the activation patterns associated with attention and performance of the task of emotion inhibition and their changes following challenge of the cholinergic system.

Effect of NRXN1 gene deletion on neurite outgrowth and cell fate in human induced pluripotent stem cells derived from ASD patients 27 Apr 2017

Autism is a devastating neurodevelopmental disorder with unmet clinical needs. Multiple genetic factors are involved, and NRXN1 is one of the most common rare but significant factors. Professor Shen at NUIG has been granted a SFI Investigator’s award to investigate autism using induced pluripotent stems cells (iPSCs). They have derived iPSCs from controls and autistic patients with NRXN1 deletions, and differentiate iPSCs into astrocytes and neurons. RNA sequencing data from 100 days of differentiated cultures demonstrated a disturbed balance of neurons and astrocytes in patients' samples. This project will therefore investigate if NRXN1 deletion affects early birth of cell types. I will perform lineage differentiation protocol to directionally differentiate iPSCs into cortical lineage. I will compare proliferation in neural stem cells, and quantify their rates between patients and controls. I will investigate astrocyte/neuronal lineage determination using Tuj1 and GFAP markers. Autism symptoms appear early postnatally which correlates with development of neuronal network, and preliminary data showed dysregulation of MAPK and GSK3beta in patient neural stem cells, which play important roles in neurite outgrowth. I will therefore also investigate neuritogenesis during early neuronal differentiation. This internship is anticipated to provide evidence into how NRXN1 deletion may affect autism-related early brain development.

Analysis of H2AX protein abundance in human breast cancer cells 27 Apr 2017

The integrity and stability of the genetic information is essential to life, however, DNA is continuously being damaged, either as a result of normal cellular processes or via the environment. Each cell receives thousands of DNA lesions per day and if these lesions are not repaired, or are repaired incorrectly, they can lead to mutations that can threaten cell or organism viability and ultimately cause cancer. In order to detect and repair the various forms of DNA damage, cells have developed a mechanism known as the DNA damage response. This project aims to better understand the role of an important protein component of the DNA damage response, H2AX, in breast cancer. We aim to measure H2AX protein abundance in several breast cancer cell lines and to identify if there is a link between the abundance of H2AX in breast cancer patients with different disease subtypes, such as luminal or basal/triple-negative. H2AX expression and protein abundance is potentially an important biomarker for breast cancer and here we aim to optimise methodologies so that small samples taken at biopsy can be measured. Furthering our knowledge of the DNA damage response in cancer cells is important to understand disease progression and treatment.

Microanatomy of the thumb carpometacarpal (1CMC) joint. 27 Apr 2017

The 1CMCjoint is the joint in the hand most affected by osteoarthritis.However there is only limited information available on its microscopic structure, and the studies which have been performed are difficult to compare as they all utilise different techniques(CT, MRI, SEM).In work as yet unpublished (Wilkins) it demonstrated the feasibility of obtaining joints and imaging them by eSEM to give high resolution surface maps of the articular surfaces, comparable to those obtained in standard SEM.This project was commenced to attempt to integrate and correlate information from several of these techniques. The joints were then reprocessed for routine paraffin histology and the appearance of the joint surface correlated with the morphology of the articular cartilage and subchondral bone in histological sections.This is the first time that direct correlation of the joint surface with joint histology has been demonstrated.This project aims to extend these studies to include micro CT in the examination sequence.Radiographic and CT imaging are clinically useful in assessing joint disease. On successful completion this will enable direct correlation of clinically relevant imaging data with high resolution views of the joint surface and the cartilage bone interface.

Family Factors and Psychosis 27 Apr 2017

My research will focus on individuals that have been diagnosed with schizophrenia, bipolar disorder or schizo-affective disorder. The hypothesis which I will test in this research is that poorly functioning families, alongside families with high expressed emotion, are a prognostic factor in the course of schizophrenia, bipolar disorder and schizoaffective disorder and are linked with the development of psychotic episodes as a symptom of these disorders. I aim to investigate the correlation of the severity of the illness with the level of dysfunction within the family and establish what specific changes can be made in the family system to serve as protective factors and hence improve patient outcomes. I also hope to identify which family styles are protective to patients' mental health and are associated with the best patient outcomes and how the family perspective on mental health impacts on the health and recovery of patients. I will ask patients and families to fill in suitable questionnaires and I will analyse the data collected to come to evidence based conclusions.I will complete the project by undertaking a cohort study with baseline data collection of patients and family members.

Vitamin D in Irish Children with Type 1 Diabetes : A Cross-sectional Study 27 Apr 2017

Vitamin D can be found in the form of cholecalciferol1 (vitamin D3) or ergocalciferol (vitamin D2) which can be obtained from food or dietary supplements. Recent studies revealed that vitamin D plays an important role in the prevention of various chronic diseases including cardiovascular diseases2, cancers such as colorectal and prostate cancers and diabetes mellitus in children3. Low vitamin D levels in children are a widespread problem across the world. The prevalence reported in healthy European children varies widely between 8% and 95%4. Vitamin D deficiencies can be a result of several factors such as inadequate exposure to the sun and disorders that hampers vitamin D absorption to name a few. Vitamin D deficiencies has also been associated with risks of cancer, increased risk of preeclampsia in pregnant women and an increased risk of type 1 diabetes in children and adolescents5. Hence, the aim of this study is to report the vitamin D levels of a cohort of Irish children with Type 1 Diabetes attending a clinic in the west of Ireland, to establish the frequency of low vitamin D levels in that cohort (insufficiency and deficiency) and to compare them to recent study reports from other countries worldwide.

Knowledge, Attitude and Behaviours of Medical Students Towards Sexual Health 27 Apr 2017

Sexual health is a key component to a holistic approach to health frequently overlooked as quite a taboo subject. Medical students as future healthcare professionals and advocates of a healthy lifestyle, it is therefore critical that they understand the importance of their own sexual health and that of their future parents. As part of my research project I plan to investigate the knowledge attitudes and behaviours of undergraduate medical students around sexual health. Notably focussing on the areas of contraception, STI’s, sexual activity and how students feel their knowledge in these areas has been influenced by the medical curriculum. As well as this identifying where possible influential factors such as social, cultural, personal, economic and educational. This study will be carried out following extensive literary reviews in the areas, before compiling a questionnaire for distribution in a cross-sectional survey of undergraduate medical students. The results obtained will be entered into SPSS for cleaning and analyses. This data can then be used a comparison against both national and international based studies. Highlighting any new data. Finally dissemination of findings, via Wellcome Trust research report, conference presentation and peer reviewed journal article.

Investigation of COX2 Regulation of HERV-K in Triple Negative Breast Cancer 27 Apr 2017

HERV-K is the youngest family of human endogenous retroviruses which constitute 8% of our genome. HERV-K is capable of transcribing gag and env proteins, and the spliced accessory proteins Rec and Np9. My supervisor Dr. Sharon Glynn has previously demonstrated the HERV-K env protein is expressed in breast and prostate cancer, and is associated with lymph node metastasis and poor outcome in breast cancer patients. However the molecular mechanisms by which HERV-K is induced and impacts breast cancer progression are unknown. Currently research has established a role for estrogen and progesterone as inducers of HERV-K in breast cancer. However HERV-K is also expressed in hormone receptor negative-triple negative breast cancer. We hypothesise that inflammatory mediators including COX2 may play a role in HERV-K expression in triple negative breast cancer. Objectives: Determine if pro-inflammatory cytokines and prostaglandins are inducers of HERV-K mRNA in triple negative breast cancer. Determine if NSAIDs (e.g. ibuprofen and aspirins) inhibit HERV-K mRNA expression in triple negative breast cancer through inhibition of cyclooxygenase enzymatic activity. Examine the association of COX2 and HERV-K env expression in 20 cases of invasive ductal carcinoma of the triple negative breast cancer subtype, and correlation with pathological features and disease free survival.

Potential role of tumour-secreted exosomes in detection and monitoring of breast cancer 27 Apr 2017

MicroRNAs (miRNA) are short, non-coding sequences of RNA that have been shown to be dysregulated in the circulation of patients with a variety of cancers, including breast cancer. Numerous studies have shown that miRNA-containing exosomes are secreted into the circulation and their miRNA content has been shown to reflect that of secreting cells. As such, exosome-encapsulated miRNAs may represent an ideal biomarker for diseases at an early stage. This study aims to identify clinically relevant exosome-encapsulated microRNAs in the circulation of breast cancer patients that are reflective of disease characteristics. Recent initial studies of ExomiRs in the circulation of breast cancer patients have provided an important proof-of-principle, although have been limited in size and scope, and have not reflected the heterogeneity of the disease. The Discipline of Surgery Biobank currently contains >1200 serum/plasma samples, from which exosomes can be isolated. This is an invaluable resource and is linked to complete clinicopathological, and treatment details of patients. This work will determine the clinical relevance (in circulation) of exosome-encapsulated miRNAs recently identified to be secreted by breast tumours In Vitro and In Vivo by the host laboratory group.

Assessing Redundancy in DNA Replication Initiation 27 Apr 2017

CDC7 kinase is a key molecular switch for DNA replication. CDC7 kinase is a protein kinase which phosphorylates and activates the mini-chromosome maintenance (MCM) helicase thus triggering DNA replication origin firing. However, recent research has shown that CDC7 kinase inhibition with some ATP competitive inhibitors does not entirely inhibit DNA replication and cell cycle can still proceed, thus have marginal effects on cell proliferation. Importantly the FANCI protein, that is normally involved in DNA repair, has been shown to be involved for the firing of DNA replication origins during replication stress and to regulate CDC7 dependent activation of MCM helicase at "dormant origins" . We will test the hypothesis that CDC7 and FANCI cooperate in the activation origin during normal replication and that the activity of CDC7 inhibitors could be greatly enhanced in FANCI depleted cells. For this we will deplete FANCI from breast derived cells and traet them with different doses of the CDC7 inhibitor XL413. Key goals are 1) reproducible detection of FANCI by western blotting, 2) efficacious depletion of FANCI by siRNA and 3) implementation of DNA replication and cell survival assays.

Diabetes Mellitus, Human Uterine Contractility and Labour 27 Apr 2017

It is well recognised that both maternal obesity in pregnancy, and maternal gestational Diabetes Mellitus (DM) result in a much greater need for caesarean section for delivery of the infant (1-3). There is an emerging view that the human uterine smooth muscle is subjected to some type of metabolic dysregulation in these conditions, but the exact factors involved are unknown. The aim of this project is to examine the effects of a number of biomarkers associated with DM in obese women on the contractility of human pregnant myometrium in vitro. The effects of glucose, Proinsulin C-Peptide, and Insulin, on spontaneous, and agonist (oxytocin)-induced contractions of human myometrium in vitro, obtained in the third trimester of pregnancy, will be examined. The following parameters will be measured, and compared with simultaneously run control experiments: frequency and force of contractions, peak amplitude and integrals of contractile activity over 15-minute periods. The raw data measured will be compared using ANOVA, followed by Tukey's HSD test for post-hoc analysis, with significance taken as a P value

A study of the utility of a computerised version of the MBT in assessing cognition in a population with mixed neuropsychiatric presentations. 27 Apr 2017

This study aims to explore the usefulness of MBTc (computerised version of the months backwards test) at detecting delirium. A major challenge in accurately detecting delirium is the lack of a reliable and convenient tool that can be used in everyday practice (O’Hanlon et al 2013). Despite the ease of use of the verbal Months Backwards Test, there are inconsistencies in how it is administered e.g. how to deal with errors and the use of prompts. A novel computerised version of the months backwards test has been developed at the University of Limerick and its performance has been examined in a population of hospitalised elderly patients and cognitively intact young people. The MBTc test had a high correlation with the verbal version of the test and demonstrated excellent sensitivity for detecting cognitive impairment in elderly patients. It is thought the enhanced standardisation of administration of the computerised version of the test will improve reliability of clinical detection of these symptoms. Participants will also be asked to complete other psychiatry tests e.g. Serial Sevens and WORLD backwards. Patients’ perspectives of the various tests will be sought. The above investigations will form part of a standard Consultation Liaison Psychiatry assessment.

Investigation of the Signalling Pathways Underpinning the Enhanced Immunosuppressive Phenotype of Tumour Associated Mesenchymal Stromal Cells 27 Apr 2017

The aim of this project is to understand how stromal cells underlying the colonic crypts interact with the epithelial tumour cells, protecting them from immune mediated destruction. The research objective will be to identify if activation of transcription factors, such as NF-kappa-B and STAT are necessary for the heightened immunosuppressive phenotype observed in tumour conditioned stromal cells. Research will be carried out using syngenic models of colon cancer in vitro, including Balb/c metastatic colon cancer cells, Balb/c mesenchymal stromal cells and Balb/c CD3+T cells. The research, carried out over the course of an 8 week project will aim to; 1. Confirm the potentiated immunosuppressive ability of tumour conditioned stromal cells 2. Investigate if the enhanced immunosuppressive effect is associated with activation of NFkappaB (p65 phosphorylation) and STAT (STAT3 phosphorylation) using western blotting. The desired outcome of this research project, is to understand the signalling pathways that are activated in tumour associated stromal cells that influence their heightened immunosuppressive potential. As part of Dr Ryan's larger research group, ultimately it is envisaged that this research will identify ways in which these interactions can be targeted and blocked, in order to enhance the immune system's effect on the tumour.