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An old drug with new tricks; examining the anti-cancer potential of metformin in mouse and cell models of EML4- ALK+ non-small cell lung cancer
No full textLung cancer is an aggressive disease producing the highest cancer worldwide. Non-small cell lung cancer (NSCLC) accounts for the majority of all lung cancer cases. The discovery of oncogenic receptor tyrosine kinases (RTKs) has led to the development of targeted drugs, improving the survival of many patients. One such RTK occurs when a chromosomal rearrangement fuses the echinoderm microtubule-associated protein-like 4 (EML4) with the anaplastic lymphoma kinase receptor (ALK) producing the constitutively active EML4-ALK. Crizotinib is a first-generation tyrosine kinase inhibitor (TKI) which targets ALK fusions, preventing cancer cell growth, and increasing the progression free survival of patients. However, the effect of crizotinib and other TKIs is limited by the development of acquired resistance. To overcome this, various strategies are being explored, including repurposing existing drugs which may possess anti-cancer mechanisms. Metformin, a hypoglycaemic agent, has promising epidemiological evidence in reducing the risk of several cancers. This thesis aimed to examine if metformin is active against EML4-ALK+ lung cancer and if it provides synergistic benefit to crizotinib.
The first section of this thesis examines if a combination of metformin and crizotinib produces toxicity, which was followed by an in vivo xenograft model study. The drugs in combination showed no liver or kidney toxicity. Metformin did not affect CYP3A activity or polypeptide levels, the main metabolising enzyme of crizotinib. Crizotinib, metformin and the combination slowed the tumour growth rate in an EML4-ALK+ lung cancer xenograft model. However, the combination was not significantly different to the drugs alone and therefore, the combination was not examined further.
The second section of this thesis explored the potential anti-cancer effect of metformin by examining known targets using in vitro techniques. Metformin did not inhibit ALK phosphorylation in EML4-ALK+ cells, confirming that it has a different mechanism of action to that of crizotinib. Nor did metformin affect mTOR or AMPK, however, it did induce apoptosis and a G1 phase cell cycle arrest. Interestingly the mitochondrial membrane potential (Δψm) was reduced with metformin treatment, indicating that metformin is likely inducing mitochondrial dysfunction in cancer cells. An analogue of metformin, mito-metformin10, was shown to be far more cytotoxic and also reduced the Δψm at concentrations far less than that of metformin. To examine if crizotinib was limiting the entry of metformin into the cell, mass spectrometry was conducted. However, this experiment could not conclude whether the
intracellular levels of metformin were reduced with crizotinib treatment, as the within experimental group variation was greater than that of the between group variation.
This thesis demonstrated that metformin can produce anti-cancer effects in EML4-ALK+ lung cancer. Further experimentation should be conducted to confirm the new proposed hypothesis that metformin induces energetic stress in cancer cells due to an inhibition of mitochondrial complex I and reducing hepatogluconeogenesis, preventing glycolysis-induced ATP production. Nevertheless, this thesis supports the hypothesis that metformin may be a beneficial anti-cancer compound in EML4-ALK+ lung cancer
Screening and treating pre-sarcopenia in middle-aged adults
No full textDespite the presence of pre-sarcopenia (stage of sarcopenia in which muscle mass deteriorates, but muscle strength is still intact) in middle-aged adults, previous research has focussed on older individuals. A better understanding of early markers of sarcopenia, age-appropriate screening models, and exercise focused prevention-based interventions, will assist to counteract the surging prevalence of sarcopenia in the ageing population.
The first part of this thesis used the Dunedin Multidisciplinary Health and Development Study data (n = 899) to classify pre-sarcopenic participants based on appendicular lean soft tissue mass index (ALMI) and grip strength. The relationship between pre-sarcopenia at age 45, and pro-dromal markers at ages 26, 32 and 38, was assessed by multivariate logistic regression. Regression models were internally validated, and overall performance, calibration and discrimination of the models were estimated. These models were confirmed using logistic least absolute selection and shrinkage operator regression. Furthermore, female age 38 models were externally validated in a separate sample of adults (n = 99). The second part of this thesis consisted of a 40-week, single-blinded, randomised controlled trial (RCT), conducted in eighty-two middle-aged adults with low ALMI. Participants were randomly allocated to group-based, 20-week high intensity aerobic and resistance training (HIART) (n = 41), or a single education session (n = 41). The initial 20-weeks were followed by a 20-week HIART home program for both groups. Outcome measures included body composition, physical function, and physical fitness. The primary outcome measure, lean soft tissue mass, was assessed at baseline, 20-weeks, and 40-weeks. Analyses were performed using intention to treat principles and linear mixed models.
The first part of this thesis indicated that, in females, protective factors included high BMI from age 26 onward, and high grip strength at age 38. In males, low self-perceived fitness level at age 38 was a risk factor of pre-sarcopenia, whereas significant protective factors included a high BMI from age 26 onward and high grip strength at age 38. The sex-specific regression models revealed an explained variance that increased from age 26 to age 38 and showed good accuracy of the models. External validation in females demonstrated good discrimination and calibration, as well as a high level of sensitivity (0.92) and specificity (0.76). For the second part of this thesis, the sample for the RCT included 82 participants (mean age 45.1 years; BMI 25.8 kg/m2) of which 85% were female. Significant group differences were observed in favour of the intervention group, after 20-weeks for measures of body composition, physical function, and physical fitness. Furthermore, lean soft tissue mass and the 30-seconds sit-to-stand remained significant at 40-weeks.
This thesis highlighted the association of pre-sarcopenia with modifiable markers in middle-aged adults. Screening for these markers predicted sex-specific risks of pre-sarcopenia in the targeted population, which was internally and externally validated. Furthermore, HIART was shown to be a safe, tolerable, and effective exercise intervention to mitigate the loss of lean soft tissue mass, in middle-aged adults with low ALMI
Views of People with Mental Illness on Assessment of, and Support During, Diminished Capacity.
Get PDFDefinitions of incapacity are socially constructed, resulting in inconsistency in judicial approaches to supporting people with mental illness with diminished capacity in New Zealand. Debates on the nature of incapacity and decision-making during diminished capacity have largely occurred in academic, legal and professional arenas, yet people with mental illness are uniquely able use their experience of diminished capacity to contribute to these debates.
This mixed-methods, exploratory study sought to ascertain the range of views held by people with mental illness on: 1. factors deemed relevant to capacity assessment, 2. who should be involved in capacity assessment and decision-making, and 3. how decisions should be made during diminished capacity. Twenty-eight people participated in qualitative focus groups and / or individual interviews in Dunedin. Constant comparison analysis, content analysis, keyword-in-context and quantitative statistical analysis were used to analyse data.
There was no significant consensus in response to the research questions. Instead, participants argued for holistic, individualised assessment and decision-making that accounts for their individuality, their context and their experience. Participants wanted choice in who was involved, often preferred more than one person, and wanted these people to have knowledge of them, their illness and their sense of ‘control’. Significant barriers to involving others were identified, often culminating in stigma and social isolation. Participants endorsed their own involvement in decision-making and supported decision-making in the first instance. Beyond this first preference, participants favoured an individualised menu of decision-making options. Several requested decisions be overridden should these prove ‘risky.’ A spontaneous finding highlighted the importance of relational autonomy in facilitating capacity, whether retained or diminished. It is recommended that current legislation be reviewed to allow flexibility and choice in capacity assessment, decision-making, and involving others, and that people with mental illness be encouraged to engage in discussion on principles guiding decision-making should their capacity be diminished
Essentiality of succinate metabolism in Mycobacterium tuberculosis
No full textNew drugs are urgently required to combat the tuberculosis epidemic that claims over 1.5 million lives annually. The identification of bedaquiline (BDQ) as an inhibitor of the mycobacterial F1Fo ATP synthase and its subsequent clinical success has validated mycobacterial energy generation as a high priority target space for antitubercular drug development. However, continued development of this promising target space requires an increased understanding of the mechanisms Mycobacterium tuberculosis uses to generate energy, as well as the identification of novel drugs and drug targets within the mycobacterial respiratory chain.
Succinate dehydrogenase (SDH), or complex II of the electron transport chain, couples the two-electron oxidation of succinate, a TCA cycle metabolite, to the reduction of quinone (succinate + quinone ↔ fumarate + quinol), thereby directly linking central carbon metabolism to the respiratory chain. M. tuberculosis encodes multiple enzymes predicted to be capable of catalyzing succinate oxidation including two different succinate dehydrogenase enzymes, Sdh1 and Sdh2, as well as a separate fumarate reductase (Frd) with possible bi-directional behavior. The dual role of SDH within both the TCA cycle and respiratory chain of M. tuberculosis makes it a particularly attractive drug target. However, the roles and essentiality of the succinate dehydrogenase and fumarate reductase enzymes encoded by M. tuberculosis are poorly defined, and no candidate inhibitors of the mycobacterial succinate dehydrogenase have been identified.
Previous investigations into the roles and essentiality of three SDH / FRD enzymes encoded by M. tuberculosis have been limited by the use of single deletion mutants, which are unable to overcome the functional redundancy in SDH catalysis. We address this by utilizing a recently developed mycobacterial CRISPR Interference system to create single, double, and triple transcriptional knockdowns of sdh1, sdh2, and frd in M. tuberculosis. We show that succinate oxidation can be catalyzed by either Sdh1 or Sdh2, but not Frd, and demonstrate that impaired succinate oxidation prevents growth and dysregulates respiration in M. tuberculosis. Moreover, our results suggest a requirement for succinate oxidation in resistance to isoniazid and pretomanid.
Furthermore, we identify and characterize the mechanism of action of a putative succinate dehydrogenase inhibitor; BB2-50F. We demonstrate that BB2-50F has rapid bactericidal activity against M. tuberculosis and show that it binds to the catalytic subunit of SDH. Using metabolomic and transcriptomic profiling of BB2-50F-treated M. tuberculosis and M. smegmatis, we propose that the mechanism of action of BB2-50F involves the dysregulation of succinate metabolism and the production of reactive oxygen species.
Overall, this thesis provides fundamental insights into the essentiality of succinate metabolism in mycobacteria and validates succinate dehydrogenase as a potential drug target in M. tuberculosis
∆122p53 mice present persistent colitis following DSS treatment
No full textInflammatory bowel disease (IBD) is a leading health concern in New Zealand as it a
lifelong condition that may cycle between periods of remission and relapse, often
requiring extensive treatments and medical interventions. The two typical forms of
this disease are Crohn’s disease and ulcerative colitis, both involving chronic
inflammation colon caused by a dysregulated immune response. How the immune
response is regulated in IBD in uncertain. Investigations into cancer using transgenic
mice that express Δ122p53, a mimic of the human isoform of the tumour protein 53
gene (Δ133p53) display a proinflammatory phenotype with increased serum
cytokine concentrations.
To investigate a role for Δ133p53 in IBD, we employed the dextran sulfate sodium
(DSS)-induced colitis model using Δ122p53 mice. Mice heterozygous for Δ122p53
and wild-type p53 (Δ122p53), and control mice heterozygous for wild-type p53 and
a p53 null allele (p53±) were administered DSS for seven days and euthanised at 0,
7, or 14 days after. Colon tissue was taken and investigated for colitis, recovery, and
cytokine production using routine histology and RNAscope assays.
Analysis of histology sections show that Δ122p53 mice have about twice as much
(20.5% versus 10%, p=0.042) of mucosal colitis when compared to p53± mice,
immediately after DSS treatment. The DSS-induced colitis persisted for longer but
was able to recover in Δ122p53 compared with p53± mice. Seven-day recovery mice
had 15.5% and 5.3% of colon mucosa affected by colitis for Δ122p53 mice and p53±
mice (p = 0.002). Fourteen-day recovery had no difference in the amount of mucosa
affected by inflammation 1.9% versus 1.7% for Δ122p53 and p53± mice,
respectively (p = 0.88). Mucosal mRNA expression of various cytokines was also
measured in-situ, using RNAscope® analysis (CCL11, CSF2, IL-6, CCL3, IL12B, and
IL12RB1). Only IL-6 mRNA had a significant difference in expression between
Δ122p53 and p53± mice, in areas of colitis at 7- and 14-days post DSS treatment.
This suggests that once Δ122p53 had recovering mucosal architecture at 14 days,
there was still an increased inflammatory response with increased IL-6 production.
The findings from this study suggest Δ122p53 increases colitis. Future directions
include investigating how Δ122p53 facilitates inflammation in colitis and if Δ133p53
is associated with IBD in patient
The toheroa, Paphies ventricosa, the feeding and diet of, with morphology notes
Get PDFThe toheroa, Paphies ventricosa, is a species with a rich history of exploitation but in the last 30 odd years harvesting has been drastically curtailed in attempts to stabilise population numbers. Only two fishing seasons have occurred since 1981, both for one day only and both located at a single beach, Oreti Beach. During these open days each person who wished to try was allowed to collect five toheroa that measured over 10cm in length. For both open days over 10,000 people turned out trying to get their quota, some carted off the beach suffering from hypothermia. The dedication and numbers turning up to open days suggest a still high regard for the protected shellfish. Aquaculture operations, whether for conservation or economic endeavours, have been suggested for the toheroa for these reasons. The suitability of the toheroa, Paphies ventricosa, for aquaculture is therefore examined, with specific investigations into the diet, sedimentological habitat, morphology and feeding ability conducted for toheroa
from the Oreti Beach population.
Diet was examined in two ways, by examination of gut contents and water sample counts. The gut content analysis was an abysmal failure with no species recognisable once the samples were returned to the laboratory and dissected and preserved. Water samples proved more successful with five species sharing dominance throughout the year. Two of the five species have been reported as dominant at other toheroa beaches, but cell counts observed were highly variable. No samples coincided with the renowned phytoplankton blooms common to toheroa beaches, so cell counts never reached levels reported in Rapson (1954) or Cassie and Cassie (1960). Organic concentration of the water was also tested and produced levels that were fairly consistent throughout the year with mean organic concentration for the year recorded as around 14mg l-1. This level compares favourably with levels reported for other beaches, but the lower temperatures at the southern beach might inhibit feeding rates and growth to a degree not determined.
Sedimentology data from Oreti Beach revealed that the southern beach has a significantly smaller mean grain size than the northern most beaches. Samples collected at Oreti contained on average around 30% of particles smaller than 0.125mm compared to less than 2% for areas of Dargaville Beach that had good beds of toheroa. Sediment to a depth of at least 10cm was highly sorted, indicating why the toheroa is required to maintain the high burrowing ability reported by Kondo and Stace (1995). Sediment found within the toheroa beds and located higher up the beach profile were statistically similar to a depth of 5cm, below this the sediment located higher up the beach became significantly smaller. Within a sample stratum the mean sediment size and sorting remained statistically similar.
Shell morphology was examined and the results from two methods of measuring length were examined, one used by Redfearn (1974) in examining the toheroa, the other suggested as a standard method for measuring shells between bivalve species by Stanley (1970). The two methods produced results that although different, were easily reconciled using a simple relationship. Length, height and width data were also used to reveal that the toheroa joins a select list of relatively thick shelled species that inhabit hostile environments and have a high burrowing ability. Other relationships length has with other measurable quantities were also reported.
Morphological measurements are also used to establish a method to distinguish between the toheroa and the southern tuatua, Paphies donacina. A score based system similar to that used by Richardson et al. (1982) is devised using ratios commonly used to describe shell shape. Only 4% of southern tuatua tested were misclassified using this system, and no toheroa. While establishing this test for species, it was noted that the score equation reported in Richardson et al. (1982) had an error in it. This equation, used to distinguish between the two species of tuatua, was length biased. Once specimens smaller than 5cm were tested it was highly likely they were misclassified as the northern tuatua, Paphies subtriangulata.
Feeding experiments that used a specialised feeding chamber designed specifically to allow for intertidal simulation for infaunal animals were conducted. Results showed the toheroa to have a slow rate of clearance for the concentrations tested. The clearance rate was related to the size of the individual, with smaller individuals clearing less than larger ones. Clearance rate followed the general formula observed for bivalve filter feeders: CR= aDWb with the b value calculated to be around 1.07 for all three dietary regimes tested. The a value differed significantly between dietary regimes indicating that toheroa regulate their clearance rate, clearing larger volumes in waters with a low phytoplankton concentration. High b values indirectly support Rapson's (1954) observation that toheroa have a high metabolic rate requiring significant amounts of oxygen.
Ingestion rates displayed a significant difference between all dietary levels tested. Finding the relationship between the ingestion rate and phytoplankton cell concentration suggested that at levels below around 350,000 cells l-1 toheroa would not actively feed. This was supported by significantly different levels of clearance for samples that were collected when cell concentration were only 100,000 cells l-1. It thus appears that at low phytoplankton concentrations toheroa may pump water across gills for respiratory purposes mainly, but will ingest what food comes its way at the same time
Social Media Influencers in Neoliberal Networked Publics and their Presentation of Contraceptive Apps
No full textSocial media influencers are becoming increasingly wide-reaching and influential, and many specialise in health and well-being, promoting products and apps that are sold as ways to improve health outcomes. Using a critical neoliberal lens, this thesis will focus on social media influencers’ promotion of a specific reproductive health app, Daysy. Daysy is advertised as a reliable and trustworthy form of safe contraception, despite its effectiveness being called into question.
Social media influencers self-brand and self-objectify as they quantify the self through tracking technology, which is often viewed as a neoliberal understanding of health. The internet has provided a space for community and connection for those concerned with health, with lay people embodying the role of expert in response to being disillusioned with medical authority. These online experts promote certain methods towards health and wellbeing, often shifting into a naturalness bias and focus on the individual through fitness and diet. Using online platforms, individuals present themselves as authentic, and develop trust with their audience over time.
Netnography was the method used to gather data, a form of online ethnography developed in 2010. I collected public data from three key social media influencers who work in Aotearoa New Zealand and promote the Daysy app. The data is richly varied, including field notes and observations collected in the form of screen shots. Data was gathered from profiles, images, text on posts, videos, vlogs and blogs. I examined the relationships and reciprocations between social media influencers, products and followers.
In my analysis, I begin by outlining this particular group of connected influencers as a networked public, highlighting the features, affordances and dynamics all present within this community. I then conduct a reflexive thematic analysis of my case study data and present four themes. The first theme addresses how social media influencers perform authenticity for their audience through displaying the highlights and lowlights of their lives publicly, while also commodifying those experiences. The second theme examines how social media influencers within this networked public express themselves as self-made experts on a variety of topics, including reproductive health, presenting neoliberal citizenship and consumerism as desirable. The third theme covers how these influencers emphasise achieving a docile body through the use of tracking technology and the human to non-human interactions afforded by this technology. The fourth theme addresses how technology becomes imbued with naturalness, and becomes an augment of the human body, a tool for translation between the mind and the body.
Overall, influencers within this networked public promote the consumption and use of reproductive health apps as a form of liberation and freedom from hormonal contraception. Influencers use their platform to display ideal neoliberal citizenship by both actively participating in and promoting neoliberal capitalist ideology. Participation in neoliberal consumption, alongside the rejection of medical and scientific authority, is framed as a revolutionary liberation from oppression. Despite using language suggesting collective action and protest, the method of protest is centred solely on individual purchasing and self-control
A homebrewers guide to upcycle unwanted bread in to beer
Get PDFUpcycling’ some of the 29 million loaves of bread wasted annually in New Zealand into beer, is a fun concept that can help to raise awareness of food waste and divert unused bread away from landfills. Working with Citizen Collective Food Scientists at the University of Otago set out to explore the maximum amount of bread that could be substituted into a good tasting beer by developing a unique mashing regime.
This infographic provides step-by-step instructions on how you can become a food waste conscious brewer. From a simple ‘how-to-guide’ through to a section on ‘Extra for Experts’, this resource guide contains everything you need to know about brewing beer from bread
The long-term impacts of opioid use before and after joint replacement surgery: Report on finding the optimal matching model
No full textEstablishment of woody species in the gorse-forest succession
Get PDFAn understanding of underlying causes assists in the prediction of outcomes and management of plant successions. The purpose of this study was to investigate the causes of the succession from gorse to broad-leaved forest by experiments under controlled conditions and manipulative experiments in the field
In the controlled experiments, the effect of different light levels on germination, survival and growth was investigated for some of the major species in the succession (principally, Ulex europaeus (gorse), Kunzea ericoides, Coprosma robusta, Pittosporum eugenioides and Melicytus ramiflorus). Species performance was matched against the generalised behaviour of typical early- and late-successional
species.
While seed characteristics and germination responses were not consistent with those predicted on the basis of successional status, the study species had growth rates and biomass allocations consistent with species of similar successional status. Gorse had significantly higher growth rates than the native woody species at high light levels but not at lower light levels. Differences in growth rates with respect to light levels explained the early dominance of gorse and the later inclusion of native woody species in the canopy. There was little evidence of trade-offs between high potential growth rates and high growth rates and survival at low light levels.
Seed bank studies investigated seed availability in the field. Seasonal patterns of germination and the effect of canopy clearance and litter removal were examined over two years. Survival of emerging seedlings and the growth and survival of transplanted seedlings was monitored in clearings cut in the gorse, under a gorse canopy and under a native forest canopy.
Seed availability of native woody species was low in young gorse stands and higher in older sites closer to seed sources of native forest species. Gorse seed dominated the seed bank of all sites except those within native forest vegetation where gorse had not previously grown. Gorse seed germinated in every month of the study, with peaks in both summers and the first autumn of the study, whereas seeds of native woody species germinated only in spring. The germination of gorse seed tended to be promoted by canopy clearance whereas that of Kunzea ericoides was unchanged and that of Melicytus ramiflorus tended to be inhibited. Litter removal from under an intact canopy promoted the germination of all species. Survival of transplanted seedlings of all species was low under dense native canopy and in unfenced plots. Gorse had the highest growth rates of the study species in the clearing but there was little difference between growth rates under the gorse canopy. Artificial shading in the field had a similar effect on growth rates to the gorse canopy.
Differences in growth rates in the field were again sufficient to explain species replacements early in the succession, but differences in life span and stature were required to explain subsequent exclusion of gorse. Seed availability and germination would influence the starting point of the succession and could result in a longer phase of gorse dominance than that predicted from growth rates alone.
Findings were consistent with the hypothesis that differences in competitive ability along a changing gradient of light availability were one of the major causes of the succession from gorse to broad-leaved forest. However there was little evidence for the existence of the trade-offs that have been proposed as the cause of these changes in competitive ability.
This study allowed options for active intervention in the management of gorse-covered land that might promote the succession to broad-leaved forest to be proposed. These include addition of seed, removal of litter, planting of seedlings and limiting any disturbance of the gorse canopy to winter in order to minimise the effects of clearance on gorse germination