6,393 results for University of Otago

  • The effect of the trafficking protein p11 on the epithelial sodium channel

    Arora, Nikhil (2017)

    Undergraduate thesis
    University of Otago

    Regulation of renal sodium (Na+) excretion is crucial for the maintenance of extracellular salt and volume homeostasis and thus for blood pressure control. The epithelial sodium channel (ENaC) is composed of three subunits; α, β and γ; each subunit contains two transmembrane domains where both C- and N-terminal domains are cytoplasmic and allow interaction with regulatory proteins. For its sodium regulating properties, ENaC is principally present in the kidney collecting duct, reabsorbing ions from the urine to prevent unnecessary loss of salt and hence, water. This makes the channel vital for maintaining ECF homeostasis and consequently blood pressure. Channel activity is highly dependent on the density at the apical membrane, where sodium current is proportional to channel number. Dysregulation of ENaC or its associated trafficking proteins can lead to an array of problems which disrupt sodium homeostasis, leading to hypo/hypertension. Although extensive research has gone into unravelling the downregulation of ENaC by endocytosis, there has been significantly less research into its exocytosis. The p11 protein is known to promote exocytosis of a number of other membrane channels. We hypothesized that addition of p11 would cause an increase ENaC trafficking, and subsequently increase the amiloride sensitive current in Xenopus laevis oocytes. Previous research at the University of Otago confirmed the presence of an interaction between p11 and ENaC, and also identified that p11 is expressed endogenously within epithelial cells. To confirm a functional consequence of this interaction electrophysiological experiments were conducted. First, Xenopus laevis oocytes were injected with mRNA coding for α, β and γ-ENaC alone or together with mRNA coding for p11. Two electrode voltage clamp was carried out to measure ENaC current. Results from my experiments showed an increase in the amiloride sensitive current in the presence of p11 at both 0.75ng (12%) and 1.50ng (17%) p11 concentrations, however the results were insignificant for both 0.75ng (p=0.46) and 1.5ng (p=0.24). Preliminary results from the Condliffe lab show increased amiloride sensitive current for oocytes co-expressing ENaC + p11 as compared to oocytes expressing ENaC alone, indicating, that the presence of p11 promotes ENaC membrane insertion. Proteins from the oocytes were also used for western blotting to identify p11 within the oocytes, however, inconclusive results were obtained. Second, we wanted to determine if the amiloride sensitive current would reverse upon silencing of p11. Fisher rat thyroid epithelia were transfected with plasmids encoding ENaC subunits + si-p11 RNA, and their resistance and amiloride sensitive currents recorded using an Ussing chamber apparatus. Results show a significant decrease (average of 75%) (p=0.04) in the amiloride sensitive current for ENaC + si-p11, when compared to control (ENaC + si-control). Overall, it is confirmed that p11 interacts with ENaC. Furthermore, it is highly likely that p11 plays a role in aiding the exocytosis of ENaC, as concluded from both the overexpression and knockdown experiments. A significant lack of any previous research on the interaction between ENaC and p11 contributed to the difficulty of this project, however, the resultant information significantly aids our understanding of the exocytic process of ENaC and the individual proteins involved, such as p11. The real-world applications of this information span across a wide spectrum including therapeutic approaches for both hyper and hypotension which are large contributors to mortality around the world.

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  • Identification of Transporters Involved in Drug-drug Interactions During Gout Treatment in Primary Rat Hepatocytes

    Nguyen, Khanh Ho Kim (2017)

    Undergraduate thesis
    University of Otago

    Gout is one of the most common form of inflammatory arthritis, with hyperuricemia as the major risk factor. Chronic hyperuricemia, or high level of serum uric acid (SUA), can lead to the formation of monosodium urate crystals (MSU) in the joints, which can result in acute flares in gout. Allopurinol is the gold standard therapy for gout, with its active metabolite, oxypurinol, acts to inhibit the enzyme responsible for uric acid synthesis, xanthine oxidase (XO), thus lowering SUA level. Moreover, ~ 70% of US adults with gout also has hypertension, which is often treated with diuretics such as furosemide. However, concomitant treatment with furosemide compromises the therapeutic effects of allopurinol. The molecular mechanisms underlying this adverse drug-drug interaction are unknown. Based on current knowledge of the transport of furosemide and allopurinol/oxypurinol by transporters in the kidney, and the fact that similar transporter setups exists in the liver, as well as evidence from clinical studies, we hypothesise that transporters known to translocate these drugs in the kidney are responsible for the drug-drug interactions in the liver, where allopurinol/oxypurinol act to lower SUA. Hence, the aim of this project was to mimic the in vivo situation in gout patients with our cell model by treating cultured primary rat hepatocytes with gout-associated drugs. The functional outputs were assessed by measuring extra- (EUA) and intracellular uric acid (IUA) level. First, we were able to successfully establish a protocol to extract primary hepatocytes via in situ perfusion method. These extracted cells had polygonal morphology, characteristic of primary hepatocytes described in the literature. We characterised the gene expression profile of urate transporters and its converting enzymes in these hepatocytes and in the rat liver tissue. In both the extracted hepatocytes (n=3) and the tissue (n=6), qPCR analysis confirmed expression of the following genes: AOX1, XO, Oat2, Oat3, Glut9, Mrp4, Npt1, Npt4, and Abcg2. Furthermore, immunoblotting was carried out to confirm protein expression of our main proteins of interest: XO, Oat2, Glut9, Mrp4, and Abcg2. However, a temporal profile of the gene expression of the hepatocytes found that, after 48h there was a downregulation of most of the genes, except for Npt4 and Mrp4, which seemed to have not changed, and Abcg2 seemed to be upregulated (n = 1). From functional studies, we treated the cells with combinations of 250 μM allopurinol, 250 μM oxypurinol, 1 mM furosemide, and 1 mM probenecid. 24h after treatment, the cells and media were harvested for analysis. Our results showed that there was a significant decrease in EUA in cells treated with probenecid, which was in agreement with our model. However, due to low sample numbers, we were not able to draw any conclusion from these functional results. Additionally, a knockdown study was performed to evaluate the contribution of Oat2 and Glut9 to the transport of allopurinol and oxypurinol. Results from one set of experiment suggest that Oat2 might play a bigger role in transport of these drugs than Glut9. Further experiments are required to confirm transport of allopurinol/oxypurinol by these two transporters.

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  • Geology of the Dusky Sound area, Fiordland, with emphasis on the structural-metamorphic development of some porphyroblastic staurolite pelites.

    Ward, Christopher Mark (1984)

    Post-doctoral thesis
    University of Otago

    Digital copy stored under Section 55 of the NZ Copyright Act.

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  • Novel grafting materials for sinus floor elevation in the sheep model

    Sheftel, Yevgeny (2017)

    Doctoral thesis
    University of Otago

    Introduction: Sinus floor elevation (SFE) may be required for implant placement in the severely resorbed posterior maxilla. Although often successful, autologous bone grafting requires a donor site and may lose substantial volume while remodelling. Bone replacement grafting (BRG) materials were developed to overcome these limitations. This study investigated three novel grafting materials: 1) equine collagen cone (CN), 2) equine collagen cone filled with biphasic calcium phosphate particles (CO), 3) deproteinized bovine bone particles coated with polylactic acid and poly ε -caprolactone copolymer (SB). These were compared with the most commonly-used bovine bone BRG, Geistlich Bio-Oss® (BO). Methods: The extra-oral access sinus grafting model from Haas et al. (1998) was used in 11 cross-bred female sheep. Two experimental sites on each side of the animal were prepared. CN, CO, SB, BO were each placed through separate 10 mm access window in the antral wall, under the elevated Schneiderian membrane. BO sites were covered with a porcine collagen membrane (Geistlich Bio-Gide®), while for CO, SB, BO sites the equine collagen membrane (RESORBA PARASORB®) from the manufacturer of these experimental materials was used. The animals were euthanised after 16 weeks. New bone, residual graft particles and connective tissue areas were measured on un-demineralised resin-embedded sections. Results: One sheep did not survive the surgery. All sites in remaining ten sheep healed uneventfully. The CN and SB grafting materials resorbed completely and failed to form new bone. BO and CO particles were bridged by the new bone, the new bone fraction was 10% (±9%) for BO and 4% (±5%) for CO. The differences were not statistically significant. Conclusions: CN and SB cannot be recommended for sinus grafting, based on this model. BO and CO demonstrated comparable histologic and histomorphometric outcomes.

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  • Plutonic and metamorphic rocks in the Victoria range segment of the Karamea batholith, Southwest Nelson, New Zealand.

    Tulloch, Andrew James (1979)

    Post-doctoral thesis
    University of Otago

    Digital copy stored under Section 55 of the NZ Copyright Act.

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  • The Poverty Bay massacre of 1868

    Black, Marjorie Edith Stuart (1935)

    Masters thesis
    University of Otago

    Digital copy stored under Section 55 of the NZ Copyright Act.

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  • Genetic factors associated with orthodontic pain in children and adolescents: a pilot study

    Sew Hoy, William Hugh (2017)

    Doctoral thesis
    University of Otago

    Introduction: Pain is often reported as being the worst aspect of orthodontic treatment. Nearly all patients experience pain and discomfort at their teeth at some point during orthodontic treatment. Little information exists on the severity of pain in the latter stages of orthodontic treatment. In addition, no studies have investigated the role of genetic factors on pain caused by fixed appliances. Objectives: To investigate whether demographic, clinical or genetic factors are associated with the severity of pain experienced following adjustment of fixed orthodontic appliances. Methods: Eighty-two participants undergoing fixed orthodontic treatment were recruited. Baseline DNA was collected via blood or saliva samples. Immediately after bond-up or an adjustment of the fixed appliances, the participants used a smartphone app to record regular pain scores at their teeth over the following three days. Results: Pain peaked approximately 19 hours after the orthodontic adjustment, then gradually returned toward baseline levels by day three. Pain on chewing was significantly greater than the resting pain at the teeth at all time points concerned. There was a significant difference in the total amount of pain at the teeth over the three days when comparing bond-ups to no arch wire changes (with or without bends placed). Gender, age, and time in treatment were not associated with the severity of pain experienced after an orthodontic adjustment. The rs931233 SNP of the HTR2A and the rs4646310 SNP of the COMT genes were significantly associated with pain severity. Haplotypes of the COMT gene also showed promising, although non-significant associations with pain severity. Conclusions: Pain on chewing is significantly more painful compared to resting pain at the teeth after adjustment of fixed appliances. SNPs of the HTR2A and COMT gene were associated with the severity of pain following adjustment of fixed appliances. Therefore, it seems that genetic factors have a modifying effect on orthodontic pain (as is the case with many other pain conditions such as TMD, fibromyalgia, and experimental pain). Larger samples are required to investigate these associations further.

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  • Exploring the Different CaMKII Isoforms in the Vasculature

    Jagau, Kevin (2017)

    Undergraduate thesis
    University of Otago

    Coronary artery disease continues to be the leading cause of mortality in the world and a major source of disability, particularly for the aged population. The presence of vascular diseases such as atherosclerosis is a predisposition to life threatening events such as acute myocardial infarctions and strokes. Whilst the current best treatment is the use of statins, they still hold a significant residual cardiovascular risk. 1 in 4 people on statins still die as a result of cardiovascular disease, therefore there is much potential for supplementary treatments. Previous work in the Heather Lab has shown that the inhibition of the enzyme calcium/calmodulin dependent protein kinase 2 (CaMKII) through the administration of KN-93 reduces the atherosclerotic lesion size in ApoE-/- mice. The results of this study show the involvement of CaMKII in atherosclerosis, and thereby a potential target for treatment. Pathologies occurring in the vasculature such as atherosclerosis are characterised by endothelial dysfunction and increased migration and proliferation of vascular smooth muscle cells (VSMC). Different isoforms of CaMKII has emerged to play a role in the regulation of vascular homeostasis, namely the delta and gamma isoforms. Studies in rats using balloon angioplasty have shown that the CaMKII delta isoform is associated with adverse migration and proliferation of VSMC, whilst CaMKII gamma isoform is associated with decreased VSMC migration and proliferation. Determining which CaMKII isoforms are present in ApoE-/- mice, and their expression pattern during the development of atherosclerosis remains an active field of research, and will lead to a better understanding of the mechanism of atherosclerosis. It was hypothesised that as atherosclerosis progresses, the CaMKII delta isoform would both increase at the mRNA and protein level, whilst the CaMKII gamma isoform would decrease in the mouse aorta. To test this hypothesis, 13, 16 and 20 week old ApoE-/- mice had their whole aorta extracted and analysed for CaMKII protein and mRNA expression. The expression of protein and mRNA of both CaMKII delta and CaMKII gamma was also explored in human umbilical vein endothelial cells (HUVEC), human coronary artery endothelial cells (HCAEC) and human coronary artery smooth muscle cells (HCASMC). It was hypothesised that there are differences in CaMKII isoform expression among the different human cell types of HUVEC, HCAEC and HCASMC.

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  • Determining the best combination of TLR Agonist and Tumour Peptide for Cancer Vaccination

    Gaskarth, Douglas (2017)

    Undergraduate thesis
    University of Otago

    Immunotherapy has revolutionised the treatment of cancer in recent years; significantly improving patient response and long-term survival. Though many immunotherapies focus on increasing the effector function of immune cells, the ability to generate and stimulate new tumour-specific immune cells has become an important topic for patients who do not respond to first line therapy. Previous work in our laboratory identified that intracellularly reversible conjugation of mode antigen ‘Ovalbumin’(OVA) to CpG B adjuvant improves the tumour specific response both in terms of immune cell activation, proliferation, and cytokine release, leading to complete tumour clearance in mice. This year I aimed to repeat this using the clinically significant melanoma antigen ‘gp100’ instead of OVA and to compare the use of CpG B adjuvant to CpG C adjuvant, which stimulates additional cytokine release, in the conjugate vaccine model. Using a combination of reversed phase high performance liquid chromatography (RP-HPLC) and cell culture, conjugates were produced and tested on their ability to activate Dendritic cells, induce their production of pro-inflammatory cytokines and induce T cell response in co-culture. Purification of gp100 conjugates was unsuccessful via RP-HPLC and testing reverted to the OVA model when comparing CpG conjugates. In antigen presenting cells, both conjugates induced similar levels of activation and antigen presentation but had unique cytokine profiles, with both conjugates trending towards higher levels of IL-1β and IL-12p70. Both CpG B-OVA and CpG C-OVA conjugates also induced a strong tumour-specific response with increased CD4+ and CD8+ T cell proliferation and significantly increased CD8+ T cell IFN-γ secretion. With these results in mind, both conjugates appear as strong candidates for therapeutic vaccination trials as either a monotherapy or a combined therapy with Checkpoint Blockade or Adoptive T Cell Therapy. In vivo testing using the CpG C construct is needed to assess its efficacy over CpG B.

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  • Seallagain: Gaelic Grammar at a Glance

    Parsons, Catrìona NicÌomhair (2016)

    Book
    University of Otago

    A native Gaelic speaker born in the Isle of Lewis and a graduate of Edinburgh University, Scotland, Catrìona NicÌomhair Parsons has been involved in the teaching of Gaelic language and song in North America for decades. For thirty summers, she taught Scottish Gaelic at the Gaelic College, St. Ann’s, Cape Breton, Nova Scotia, where she was commissioned to prepare Gàidhlig troimh Chòmhradh, a Gaelic course in three volumes with recorded text. For many years, she taught in the Celtic Studies Department of St. Francis Xavier University, Nova Scotia; after retiring, she spent six years working for the newly constituted Nova Scotia Office of Gaelic Affairs. She has written well over a hundred Gaelic-English articles for local newspapers. Her poetry has been published in Scottish Gaelic periodicals GAIRM and GATH, and she has produced her solo CD of Gaelic songs entitled “Eileanan mo Ghaoil” in tribute both to Cape Breton and Lewis. From Seattle, Washington, to Grandfather Mountain, North Carolina; from Toronto to Nova Scotia, Canada; from Sydney, Australia, to Dunedin, New Zealand, Catrìona has been privileged to share her beloved language and culture with motivated students, many of whom are now instructors themselves. This, her most recent work, is a synthesis of all of the grammatical insights garnered from decades of experience teaching Scottish Gaelic to learners around the world. It clearly demonstrates in easy-to-read chapters, tables, and examples how the Gaelic language is structured. Rules, forms, pronunciation, and a host of other issues are all logically and systematically explained. Furthermore, this book can act as a handy reference for either the beginner or native speaker.

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  • The effect of a diet moderately high in protein and fiber on insulin sensitivity measured using the Dynamic Insulin Sensitivity and Secretion Test (DISST)

    Te Morenga, Lisa; Docherty, Paul; Williams, Sheila; Mann, Jim (2017-11-08)

    Journal article
    University of Otago

    This paper is a version chapter from a PhD thesis (Te Morenga, L. A. (2010). The effects of altering macronutrient composition on diabetes risk (Thesis, Doctor of Philosophy). University of Otago. Retrieved from http://hdl.handle.net/10523/439) and as such has been externally reviewed by experts in the field. It is currently under peer review.

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  • Herr Daniel Bandmann and Shakespeare vs the World

    Warrington, Lisa (2016-12-27)

    Journal article
    University of Otago

    German actor Daniel Bandmann played his first Hamlet at the age of 20, and made his English language debut as Shylock in New York, 1863. In his prime, he performed extensively in America, Great Britain, Australia, and New Zealand, amongst other countries. Though he played roles which ranged from Narcisse and the Corsican twins to Jekyll and Hyde, he was perhaps most closely identified with a handful of Shakespearean roles: Hamlet, Shylock, Macbeth, Othello, Iago. His apparently ungovernable temper led to a love/hate relationship with the critics, played out in public through the newspapers. His responses to criticism open a window into his playing of these roles. This paper examines Bandmann’s acting in the role of Hamlet and the critical interchanges he engaged in around the world, as an exemplar of the interaction of theatre and the global media.

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  • Expression of the lysyl oxidase family in odontogenic lesions

    Abdul Rahman, Nawal Radhiah (2017)

    Doctoral thesis
    University of Otago

    Background: The lysyl oxidase family of enzymes consists of five members, namely lysyl oxidase (LOX) and lysyl oxidase-like (LOXL) 1-4. They are secreted enzymes whose function is to stabilise the extracellular matrix via crosslinking collagens and elastins. The enzymes are mostly known for their extracellular function but little is known about their intracellular function. Few studies have investigated LOX expression in the oral and maxillofacial region and those that exist are largely concerned with oral submucous fibrosis and oral squamous cell carcinoma. With regard to odontogenic lesions one study exists which showed increased expression of LOXL4 in the stromal tissue of odontogenic keratocyst (OKC). Objective: To determine the expression of LOX family proteins and genes in locally aggressive odontogenic lesions (ameloblastoma and OKC) in comparison with non-aggressive odontogenic lesions (dentigerous cyst (DC) and hyperplastic dental follicle (DF)) using immunohistochemistry (IHC) and quantitative reverse transcriptase real-time polymerase chain reaction (qRT2-PCR). Method: For IHC, formalin-fixed paraffin-embedded (FFPE) tissue samples of ameloblastoma (n = 10), OKC (n = 15), DC (n = 6) and DF (n = 9) were used with antibodies against LOX and LOXL1-4. Positive and negative controls were used for validation. Qualitative assessment of the pattern and distribution of staining was undertaken at varying magnifications. Automated quantitative assessment of digitised IHC images was performed using Fiji Software (Image J 1.51K). Specifically, the extent of positive reaction and intensity of staining were examined in three representative areas of the epithelium and connective tissue in each specimen at 400x magnification. One way ANOVA tests were performed using GraphPad Prism software (La Jolla California USA), and P values of 2 and p<0.05 for each analysis. Result: The LOX family proteins and genes showed differential patterns of expression in each lesion examined. Significant reduction of LOXL3 was observed in ameloblastoma at both protein and gene levels. LOXL4 protein was overexpressed in the epithelium, but underexpressed in the connective tissue of ameloblastoma and OKC. The expression of LOX family genes and proteins in DC showed a significant variation compared with ameloblastoma and OKC, whereas the protein expression patterns were similar between ameloblastoma and DF. Conclusion: 1) LOX family expression was different in ‘aggressive’ odontogenic lesions compared with ‘non-aggressive’ odontogenic lesions, dentigerous cyst in particular. 2) Variable LOX family expression between ameloblastoma and OKC may reflect their pathogenesis and biological behaviour. 3) The similarities of LOX family expression observed in ameloblastoma and hyperplastic dental follicle may reflect the embryonic dedifferentiation of ameloblastoma. 4) Dentigerous cyst serves as a proper ‘control’ tissue as opposed to hyperplastic dental follicle with regard to study of the LOX family.

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  • The value of hypnotism in the general practice of medicine

    Fenwick, Percival Clennell (1907)

    Post-doctoral thesis
    University of Otago

    Digital copy stored under Section 55 of the NZ Copyright Act.

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  • Floating Islanders: Pasifika Theatre in Aotearoa

    Warrington, Lisa; O'Donnell, David (2017)

    Book
    University of Otago

    ‘We float – we’re not based in one place – we’re floating Islanders. I always come back to theatre, theatre is my first home.’ – Makerita Urale This book celebrates 30 years of Pasifika theatre in Aotearoa/New Zealand. Pacific Underground, Pacific Theatre, The Laughing Samoans, The Conch, The Naked Samoans, Kila Kokonut Krew – the distinctive style and themes of Pasifika theatre have been developed by many individuals and theatre companies in New Zealand. Authors Lisa Warrington and David O’Donnell have interviewed over 30 theatre practitioners to tell the story of Pasifika theatre in Aotearoa from 1984 to 2015. This lively book showcases playwrights, directors and performers whose heritage lies in Samoa, Niue, Fiji, Tonga, Tokelau and the Cook Islands. Extracts from the interviews are threaded throughout the book, providing often entertaining insights into their history and creative practice. While the immigrant experience of living in two worlds is often seen as troubled, the authors suggest that this ‘in-between-ness’ has been turned to advantage in Pasifika theatre to create a unique and often subversive performance phenomenon. Not only is Pasifika theatre a success story within the performing arts in New Zealand, it is also an intriguing case study of migrant theatre that has international resonance.

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  • Lhx9 is required for urogenital ridge development and ovarian function

    Workman, Stephanie (2017)

    Undergraduate thesis
    University of Otago

    While there has been extensive research into the differentiation of sexually dimorphic gonads, there is much to learn about the bi-potential structure they arise from, the urogenital ridge (UGR). This gap in knowledge is imperative in the contexts of disorders of sex development and infertility, of which many cases have unknown aetiology. The transcription factor LIM Homeobox 9 (Lhx9) has been shown to have a functional role in the development of the UGR. Lhx9 -/- mice display gonadal agenesis and complete male to female sex reversal. Little is known about the regulation of Lhx9 gene expression in the UGR or its role in the greater genetic networks of reproductive development and beyond. We hypothesised that Lhx9 expression is regulated by Notch signalling in the UGR. To investigate the regulation of Lhx9 in the UGR in situ hybridisation was used to analyse the expression patterns of Lhx9, Notch (receptor), and Hes1 (Notch downstream effector) in the embryonic mouse gonad. Overlapping expression patterns in the UGR suggested a coregulatory interaction between the genes. This was further demonstrated by explant culture of embryonic gonads in the presence of the Notch pathway inhibitor DAPT. RT-qPCR revealed reduced Lhx9 expression in RNA extracted from the treated gonads, providing a strong case for a regulatory relationship between Notch and Lhx9. Due to declines in Lhx9 heterozygote (Lhx9+/-) fertility and embryo viability we hypothesised that a reduction in Lhx9 expression would result in impaired fertility in the mouse model. RNA extracted from the gonads of Lhx9+/- embryos was used for RT-qPCR to determine the relative expression of markers of key cell types in the developing gonad. Significant changes in the expression of markers of both male and female somatic and germ cells were found. This raised the question of whether Lhx9 was expressed in the adult ovary, and if so were the observed fertility declines due to reduced Lhx9 expression. In situ hybridisation revealed the novel discovery of Lhx9 expression localised to the follicles of the ovary, this was confirmed by immunohistochemistry. RT-qPCR of heterozygote ovaries revealed trends of reduced expression of critical ovarian fertility genes, a finding reflected in abnormalities seen in histological analysis. These results provide significant evidence for the role of Lhx9 in UGR development and the adult ovary, and offer direction for further investigation into its potential role in the underlying genetic networks DSD and infertility.

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  • Dietary patterns, physical fitness, and markers of cardiovascular health in 9-11 year-old Dunedin children

    Saeedi, Pouya (2017)

    Doctoral thesis
    University of Otago

    Chronic diseases such as cancer, diabetes, and cardiovascular diseases (CVD) are the main health concerns of the 21st century, with CVD as the number one cause of mortality in New Zealand and worldwide. Although CVD hard endpoints such as stroke or heart attacks do not usually occur in children, there is evidence that the manifestation of CVD risk factors begins in childhood, preceding clinical complications of CVD in adulthood. Several factors including biological, environmental, and behavioural factors are associated with the development and advancement of CVD complications. Of these, dietary intake is a modifiable risk factor that has been shown to make a substantial contribution to the risk of death from CVD. Health professionals have long recognised the importance of diets high in fruits and vegetables, wholegrain/high fibre bread and cereals, and limited intakes of sugar and sugar-sweetened beverages in reducing the risk of CVD in adults. However, there is a lack of research in the paediatric population. Thus, the aim of this thesis was to determine associations between dietary intake, particularly dietary patterns as a more global approach of assessing dietary intake and markers of cardiovascular health in 9-11 year-old children in Dunedin, New Zealand. The study was conducted in two phases. In the first phase, a short (28-item) non-quantitative food frequency questionnaire (FFQ) was developed and assessed for its reproducibility and relative validity. Fifty children (mean age±SD: 9.40±0.49 years old) from three Dunedin primary schools completed the FFQ twice, as well as a four-day estimated food diary (4DEFD) over a two-week period. Intraclass correlation coefficients (ICC) and Spearman’s correlation coefficients (SCC) were used to determine the reproducibility and relative validity of the FFQ, respectively. More than half of the food items/groups (52.2%) had an ICC ≥0.50. In relative validity analyses, 70% of food items/groups had a SCC ≥0.30. This FFQ has been used to rank children according to the frequency of consumption of specified food items/groups. The low respondent burden and relative simplicity of the FFQ make it suitable for use in large cohort studies in New Zealand children with similar characteristics. The second phase of the thesis used data from the ‘Physical activity, Exercise, Diet, And Lifestyle Study’ (PEDALS), conducted in 17 primary schools in Dunedin. Of the children who took part in PEDALS, the mean age±SD was 9.72±0.68 years old, 76% were of normal weight, 80% met the guidelines of 60 minutes of daily moderate-vigorous physical activity, and 99% were categorised as fit based on the FITNESSGRAM standards. The first objective of phase II was to identify dietary patterns using principal component analysis (PCA), using the FFQ validated in phase I. Two dietary patterns, namely ‘Snacks’ and ‘Fruit & Vegetables’ were identified. The mean ‘Snacks’ and ‘Fruit & Vegetables’ scores were -0.068±1.98 and -0.005±1.83, respectively. The two identified dietary patterns in PEDALS were similar to commonly identified dietary patterns in both international and national studies. The second objective of phase II was to determine associations between the two identified dietary patterns and components of physical fitness (i.e., cardiorespiratory fitness and handgrip strength). Cardiorespiratory fitness was measured as mean relative V ̇O2max obtained from a 20-metre shuttle run test (20msrt). A digital hand dynamometer was used to measure handgrip strength of both the dominant and non-dominant hands. Complete data was available for 398 participants. Mixed effects linear regression models with robust standard errors and school as a random effect were employed to assess relationships between dietary patterns and components of physical fitness. Mean relative V ̇O2max was 48.7±4.75 ml/kg/min. Handgrip strength of the dominant and non-dominant hand was 15.2±3.29 and 14.4±3.17 kg, respectively. There were no significant associations between the dietary pattern scores and cardiorespiratory fitness. However, fat mass index (FMI) was independently associated with cardiorespiratory fitness. Excess body fat is associated with poorer performance and consequently lower estimated V ̇O2max (ml/kg/min). Furthermore, PEDALS did not find clinically meaningful associations between dietary patterns and handgrip strength of the dominant or non-dominant hand, while sex and fat-free mass index were independent determinants of handgrip strength. Considering the important impact of muscular strength on current and future health status, sex-specific exercise training to improve children’s fat-free mass and muscular strength from as young as 9 years old should be promoted. The third objective of phase II was to investigate relationships between dietary patterns and indices of arterial stiffness (i.e., augmentation index (AIx) and pulse wave velocity (PWV)). Indices of arterial stiffness were assessed using the XCEL system. Data for AIx and PWV analyses were available for 337 and 389 participants, respectively. Mixed effects linear regression models were used to assess associations between dietary patterns and indices of arterial stiffness. Mean AIx and PWV were -2.14±14.1% and 5.78±0.79 m/s, respectively. There were no clinically significant relationships between the dietary pattern scores and AIx and PWV. Arterial stiffness is one of the earliest detectable measures of vascular damage and can be seen in the first decade of life. Although evidence has shown that obesity can accelerate the age-associated arterial stiffening process, the majority (76%) of PEDALS children were normal weight, which may explain the lack of an association. Overall, there were no significant associations between dietary patterns and markers of cardiovascular health in children who took part in PEDALS. The majority of the PEDALS population had a healthy weight status, were of New Zealand European ethnicity, and from families of middle/high socio-economic status. Further research is suggested in a cohort of 9-11 year-old children from families of low socio-economic status and minority ethnic groups such as Māori and Pacific children, using the established methodology of PEDALS. Comparison of the results from PEDALS with a similar study on a group of children with different socio-demographic characteristics would be useful to inform policy and provide further insights on the importance of designing appropriate prevention strategies in both the general and high-risk paediatric populations.

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  • Functional Characterisation of Direct Neural Inputs to Arcuate Nucleus Kisspeptin Neurons in the Mouse

    Schafer, Danielle (2017)

    Masters thesis
    University of Otago

    The pulsatile pattern of gonadotropin release is critical for puberty and fertility. Kisspeptin neurons in the arcuate nucleus (ARN) are thought to play an important role in generating pulsatile gonadotropin secretion. Previous studies have shown that kisspeptin neurons within the ARN project to gonadotropin-releasing hormone (GnRH) neurons and that the synchronous activation of kisspeptin neurons generates pulsatile LH secretion in vivo. While studies indicated that ARN kisspeptin neurons co-express and are regulated by dynorphin (DYN) and neurokinin B (NKB), the neurotransmitters that control these neurons have not yet been fully explored. Many different internal and external factors, such as changes in stress and metabolic state, modulate pulsatile gonadotropin secretion. I hypothesized the neurotransmitters mediating the effects of stress and metabolism within the brain may act directly on ARN kisspeptin neurons to ultimately regulate pulsatile LH secretion. As such, I aimed to characterise the direct effects of several neurotransmitters upon the ARN kisspeptin neurons. To identify effects, I recorded calcium activity from ARN kisspeptin neurons in acute brain slices prepared from transgenic intact male and female mice expressing the calcium indicator GCaMP6f selectively in ARN kisspeptin neurons. These experiments were conducted in the presence of a voltage-gated sodium channel blocker and ionotropic amino acid receptor antagonists to ensure that only direct effects were measured. The effects of neurotransmitters on intracellular calcium levels in ARN kisspeptin neurons were determined by measuring changes in fluorescence within individual neurons. In intact diestrous females, experiments demonstrated that NKB, corticotropin-releasing hormone, arginine vasopressin (AVP), oxytocin, noradrenaline (NA), serotonin (5-HT), dopamine, neuropeptide Y, vasoactive intestinal peptide (VIP), and pituitary adenylate cyclase-activating polypeptide (PACAP) all act directly on ARN kisspeptin neurons to increase intracellular calcium levels, whereas DYN and beta-endorphin directly inhibit the stimulatory effects of NKB on these cells. Alpha-melanocyte-stimulating hormone (α-MSH) and GnRH did not exhibit any direct effects on GCaMP6f fluorescence levels in ARN kisspeptin neurons. I also identified substantial regional and sex differences. Whereas, AVP, 5-HT, VIP, and PACAP all activated a significantly greater percentage of kisspeptin neurons in caudal ARN slices, NA exerted its most potent effects in the middle ARN region. Furthermore, the effect of AVP, 5-HT, VIP, and NA were pronounced in intact females, but significantly reduced, or even absent, in intact males. These results demonstrate that many of the neurotransmitters thought to be involved in mediating stress and metabolic actions within the brain also directly modulate the ARN kisspeptin neurons. As such, these neurons may provide an integration point through which different modalities regulate pulsatile LH secretion.

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  • A novel keratin-chitosan-tricalcium phosphate biocomposite as a potential scaffold for regenerative endodontics : an in vitro study

    Ramawarrier, Arunjith (2017)

    Doctoral thesis
    University of Otago

    The present study is the ‘first look’ at a novel biocomposite which may be used as a 3D implantable intracanal scaffold for regenerating dental pulp and periapical tissues. The main ingredient of the biocomposite was low molecular weight keratin protein extracted from sheep wool. Merino sheep wool offers a grand storehouse of keratin proteins which can extracted by many methods. This study used a novel chemical-free method using high temperature and pressure. The extracted proteins were of low molecular weight (3.5-15 kDa) and was water soluble. These proteins were used for the fabrication of the biocomposite along with other ingredients namely chitosan, tricalcium phosphate,barium sulphate and glycerol. This is, perhaps the first study that has explored the use of low molecular weight keratin in biomedical applications. Other constituents of the biocomposite were selected in order to provide specific properties to the composite. Keratin-chitosan formed a mechanically stable homogenous matrix. Chitosan also imparted an antimicrobial potential to the scaffold. Tricalcium phosphate acted as the filler and also a supplier of calcium ions. Barium sulphate provided radiopacity to the scaffold while glycerol was the plasticizer. The scaffold demonstrated many key characteristics relevant to tissue regeneration applications such as adequate porosity and degradation, as well as to endodontic applications such as moderate swelling and radiopacity. Assessment of cytocompatibility yielded promising results. The scaffold promoted proliferation of MDPC 23 (odontoblast like cells) and OCCM 30 cells (cementoblast like cells). The cells were able to grow and achieve functional differentiation when cultured after exposure to scaffold extracts as evidenced by ALP assay which detected elevated ALP levels in culture. AR-S staining detected calcium deposits which further confirmed cell differentiation. Furthermore, immunocytochemical analysis revealed expression of DSPP by MDPC 23 cells which was indicative of odontogenic differentiation. These cell reactions demonstrated the regenerative potential of the biocomposite scaffold. The population density of viable stem cells and their successful differentiation is an absolute prerequisite for successful regenerative pulp therapy, so is the effective disinfection of the root canal system. The antimicrobial potential of the scaffold was tested against S.mutans which was a representative organism for primary infection and E faecalis which represented secondary or re-infection of the root canal system. The scaffold was able to successfully inhibit growth of both the species. This potent antibacterial action of the scaffold eliminated the need for using highly potent antibiotics and other antimicrobials detrimental to host cell survival during endodontic regenerative procedures.

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  • Lymph nodes as a pre-metastatic niche for oral squamous cell carcinoma

    Al Kharusi, Adil (2017)

    Doctoral thesis
    University of Otago

    Background Pre-metastatic niche (PMN) is a new concept in the process of metastasis defined as tumour microenvironment at the future metastatic site which is established by the tumour as a preparation before the arrival of the disseminated tumour cells. Certain cells and cytokines have been reported to be a key factor in building these niches. To date, there is no single study that has investigated the PMN in the oral squamous carcinoma (OSCC). My hypothesis is that IL17, IL22, IL23 and STAT3 play part in the formation of PMN of metastatic OSCC. Aim: To compare the expression of STAT3 and cytokines (IL22, IL23 and IL17) between positive and negative lymph nodes from OSCC. Methods: A total of 36 formalin fixed paraffin embedded (FFPE) tissue specimens were obtained from the Malaysian Oral Cancer Database & Tissue Bank System (MOCDTBS). Sample were divided into two groups. Positive lymph nodes were those with histological evidence of metastatic OSCC while negative nodes were those with no sign of metastasis. Th expression of IL17, IL22, IL23 and STAT3 was investigated using immunohistochemistry (IHC). Gene expression was done using Real time polymerase chain reaction (RT-PCR) to validate the results. Image J was used to count the number of positively staining cells. SPSS was used to analyze the data. Results: IHC results shows that the expression of IL22, IL23 and STAT3 was significantly higher in the negative lymph nodes when compared with the positive group which proof our hypothesis. However, the difference in gene expression was not significant. Conclusion: My results suggest that negative lymph nodes can be a PMN for the OSCC. In addition, IL22, IL23 and STAT3 can be responsible at least partially for the formation of this PMN.

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