Massey Documents by Type

Permanent URI for this communityhttps://mro.massey.ac.nz/handle/10179/294

Browse

Subject: search.filters.subject.Biochemical markers

Search Results

Now showing 1 - 5 of 5
  • Thumbnail Image
    Item
    Mass spectrometry-based metabolomics as a tool for biomarker discovery and diagnosing metabolic health : a thesis presented in fulfilment of the requirements for the degree of Doctor of Philosophy in Nutritional Science, School of Health Science, Massey University, Palmerston North, New Zealand
    (Massey University, 2021) Zhanxuan, Wu
    Although obesity and prediabetes have been long-established risk factors for type 2 diabetes (T2D), excess fat deposition in visceral and ectopic organ sites (i.e. the “risky” fat depot) has been increasingly recognised as key conduits for T2D. However, quantification of these fat depots relies on expensive and time-consuming imaging techniques. There is a need for identification of biomarkers predictive of risky fat depot levels. Metabolomics is a promising tool for discovering novel markers and generating mechanistic speculation. This PhD project aims to identify and understand plasma metabolite markers associated with metabolic risk factors including elevated fasting plasma glucose (FPG), fat deposition in visceral (VAT) and ectopic organ sites (liver and pancreas) in non-T2D human. To achieve this, a workflow to select the optimal injected concentration of different sample types for two complementary LC-MS untargeted analyses (polar metabolites and lipidomics) was established and applied to examine the value of measuring the plasma metabolome as a proxy for metabolite concentrations in various tissue sites including adipose tissue, muscle and the liver. Lastly, plasma metabolomic signatures for elevated FPG and VAT were characterised and metabolite markers predictive of VAT and ectopic fat deposition in the liver and pancreas were identified. This PhD study highlighted the critical importance in optimising injected concentration for LC-MS analysis of different sample types to ensure the maximal number of linear features were obtained, and for the first time showed the plasma metabolomics profile was more reflective of the liver profile than muscle or adipose tissue. Subsequent metabolomics characterisation of clinical plasma samples reported profound associations between FPG or VAT with changes in several glycerolipid species independent of gender, ethnicity, age and body mass index (BMI). VAT was additionally associated with changes in phospholipid, ether-linked phospholipid and sphingolipid species independent of covariates. Liver fat deposition was predicted by a number of glycerolipid, phosphatidylethanolamine and dihydroceramide lipid species whose plasma concentrations were linearly correlated with the liver counterparts. A novel marker, sulfolithocholic acid, for the prediction of pancreatic fat independent of age, BMI and visceral adiposity was also identified. Finally, the study also reported an improved prediction of ectopic fat deposition by utilising a panel of metabolite markers compared to clinical measurements, and demonstrated the usefulness of the metabolomic signature to identify a subset of normoglycaemic individuals with a worse cardiometabolic profile. Findings from this PhD study highlighted the value of metabolomics as a promising tool to capture metabolic risk, and that candidate markers identified by metabolomics may offer opportunities for improved risk prediction and stratification, disease progression monitoring and to develop alternative means for the measurement of the effectiveness of dietary interventions.
  • Thumbnail Image
    Item
    Biomarker development to assess bone health : a thesis presented in partial fulfilment of the requirements for the degree of Doctor of Philosophy in Nutritional Science at Massey University, Palmerston North, New Zealand
    (Massey University, 2019) Cabrera Amaro, Diana Leticia
    Postmenopausal women experience an accelerated bone loss with increased fracture risk caused by oestrogen deficiency. Biomarkers of bone turnover assess the changes of bone metabolism in postmenopausal women; however, prediction of bone loss with these common biomarkers cannot be achieved because bone biomarkers might not reflect the bone microenvironment status. Thus, there is a need for discovering new bone biomarkers that can efficiently predict bone loss in postmenopausal women. Previous studies suggest that the ovariectomised sheep in combination with injected glucocorticoids may be a reliable model to evaluate the biological response to oestrogen withdrawal as well as the bone remodelling process. The purpose of this research programme was to test the following hypotheses: 1) ovariectomising sheep in combination with monthly injections of glucocorticoids would result in decreased bone mineral density (BMD) and increased plasma bone remodelling marker concentration over a shorter period of time; 2) the plasma metabolome and lipidome of ovariectomised sheep would be different, and the biochemical changes in plasma and bone remodelling would be associated with bone loss; 3) and finally, there would also be a difference in the plasma metabolome and lipidome of Singaporean–Chinese postmenopausal women according to their bone mineral density status. The hypotheses were evaluated using the OVX sheep in combination with glucocorticoids as a large animal model for postmenopausal osteoporosis, as well as comprehensive LC–MS-based metabolomics as a diagnostic approach to identify lipids and metabolites associated with bone loss in postmenopausal women. The OVX sheep model was successfully validated over five months of this study period, and bone mineral density was decreased and bone biomarkers increased after five months. Then, plasma samples from this animal model were analysed to measure the metabolome and lipidome of the OVX sheep. In the OVX sheep, metabolite and lipid alterations associated with bone loss included methionine, glutaric acid, tryptophan, 5-methoxytryptophan, CL and CerP, and these correlated with OC, CTx-1, femoral BMD and lumbar spine BMDThese studies revealed dynamic changes of the metabolite and lipid profiles from affected sheep, such as perturbation in multiple amino acids, metabolites, and fatty acid β-oxidation. Additionally, the results from the Singaporean–Chinese postmenopausal women showed alterations in proline, threonine, methionine, 4-aminobutyric acid, aminopropionitrile, phosphatidic acid, diacylglycerol, CerP and phosphatidylinositol correlated with low femoral neck BMD. Methionine and CerP were the common compounds altered in OVX sheep and SC women with low BMD when compared with healthy groups. Those compounds, which are known to be involved in bone remodelling, have the potential for studying early bone loss in postmenopausal women, where identifying new bone-specific biomarkers may aid in clarifying novel molecular mechanisms of bone loss.
  • Thumbnail Image
    Item
    Development of the [beta]-glucuronidase reporter gene system to study Acremonium endophyte interactions with perennial ryegrass : a thesis presented in fulfilment of the requirements for the degree of Master of Science in Genetics at Massey University
    (Massey University, 1997) Saunders, Karyn
    A transformant of the fungal endophyte Acremonium lolii, strain Lp19, containing the gusA gene under the control of the constitutive Pgpd promoter was generated, and assigned the name KS1. Analytical digests and Southern hybridisation showed that this transformant contained a single chromosomally integrated copy of the gusA gene. The transformation frequency of Lp19 was found to be very low, and attempts to increase the transformation frequency were unsuccessful. KS1 was used to artificially infect seedlings of several different genotypes of Lolium perenne, all of a single cultivar, 'Nui'. These seedlings were grown into mature plants, and the endophytically produced GUS enzyme was extracted from individual plant tissues. Assays were performed on the enzyme extracts, and the levels determined were used as a measure of endophyte metabolic activity. Alterations of the gusA gene in some plants was detected by Southern hybridisation. One alteration was found to result in loss of GUS activity, the other did not appear to alter gusA expression. Levels of transformed endophyte GUS activity were initially compared between clonal plant material of a single genotype. Statistical analysis revealed that no significant differences were detectable for a particular tissue between the different plants. This showed that plant material of identical genotype could be pooled for analysis without the pooling of the individual plants having an affect on the outcome of the analysis. Next, levels of the transformed endophyte GUS activity were compared between genetically diverse perennial ryegrass plants of cultivar 'Nui'. Significant differences in GUS activity were detected in most tissues tested between the different genotypes, with only the most mature tissue displaying no detectable differences. Finally, a single plant of each of two individual genotypes was divided into several clonal plants, and the resulting mature plants were pooled in their genotypes for analysis of GUS, peramine, ergovaline and lolitrem B levels. The F test was not particularly sensitive in this experiment, and only one major difference between genotypes could be detected. Despite this, some trends emerged which were found to be consistent with those found in other studies. Metabolic activity and peramine levels were shown to be highest in the leaf sheath tissue, with levels generally decreasing with increasing tissue age. Lolitrem B was found to be highest in leaf sheath tissue also, but with levels increasing in general with tissue age. Ergovaline levels were very low in all tissues. The results presented show the potential of the use of the GUS reporter gene system to study endophyte gene expression in planta, and pooling of plants can be carried out to allow simultaneous study of toxin expression.
  • Thumbnail Image
    Item
    Development of assays for biomarkers of oxidative damage to assess the efficacy of fruit-derived antioxidants : a thesis presented in partial fulfilment of the requirements for the degree of Master of Science in Biochemistry at Massey University
    (Massey University, 2003) Barnett, Laura Evelyn
    The diet is a very important part of maintaining a healthy lifestyle. Increased consumption of fruits and vegetables is one practice postulated to decrease the incidence of diseases such as cancer, cardiovascular disease and other disorders. Although there are a number of possible beneficial compounds in fruit, it is believed that the antioxidant components found in these foods may decrease the oxidative damage that could lead to such diseases. Oxidative damage to cellular proteins, lipids and DNA is considered to result from an increase in the production of free radicals, which overwhelm the body's defence system. This research investigated fruit-derived antioxidants, and developed biomarker assays to measure the potential health benefits they may offer. To determine the in vivo antioxidant efficacy of berry fruit anthocyanins, oxidative damage to proteins, lipids and DNA was measured in rats fed several combinations of natural and synthetic diets. Mild oxidative damage was induced by the inclusion of fish oil in these diets. DNA oxidation was determined by measuring urinary 8-hydroxy-2'-deoxyguanosine using reversed-phase high performance liquid chromatography with electrochemical detection. ELISA and colorimetric techniques were used to measure protein carbonyl content of plasma as a reflection of protein oxidation. Oxidation to lipids was assessed by measuring malondialdehyde, which results from lipid peroxidation. Supplementation with fish oil induced a mild form of dietary oxidative damage, as shown by an increase in lipid and protein oxidation. In most cases the berry fruit extracts had little effect on the level of fish oil-induced oxidative damage, however, boysenberry anthocyanin extract significantly reduced protein oxidation when used in combination with the natural diet. Taken together the results suggest that oxidative damage to biomacromolecules may occur by different pathways of oxidative stress, which selectively target either DNA, protein or lipids at varying levels, and the antioxidant is effective only with selected mechanisms of oxidative damage.
  • Thumbnail Image
    Item
    The search for biomarkers of facial eczema, following a sporidesmin challenge in dairy cows, using mass spectrometry and nuclear magnetic resonance of serum, urine, and milk : a thesis presented in partial fulfilment of the requirements for the degree of Doctor of Philosophy in Sciences at Massey University, Palmerston North, Manawatu, New Zealand
    (Massey University, 2015) Matthews, Zoe Maree
    Facial eczema (FE) is a secondary photosensitisation disease of ruminants that is significant in terms of both its economic importance to New Zealand and its impact on animal welfare. The clinical photosensitivity signs, caused by the retention of phytoporphyrin, occur secondarily to hepatobiliary damage caused by the mycotoxin sporidesmin. Currently it is difficult to diagnose subclinical animals and those in the early stages of the disease. The project was aimed at applying new analytical and statistical techniques, to attempt the early diagnosis of FE in dairy cows following the administration of a single oral dose (0.24 mg/kg) of sporidesmin. Well-established traditional techniques including production parameters, liver enzyme (GGT, GDH) activity measurements, as well as measurements of phytoporphyrin by fluorescence spectroscopy were made for comparison. Serum, urine, and milk were analysed using 1H Nuclear Magnetic Resonance (NMR), multivariate analysis (MVA), and time series statistics. Urine and milk did not prove useful for identification of sporidesmin intoxication. Serum metabolites differed between treated cows before and after administration of the toxin, and could distinguish samples belonging to the clinical group. The metabolites that were identified as being relevant to this classification were a mixture of glycoproteins, carboxylic acids, ketone bodies, amino-acids, glutamate, and glycerol, which were elevated for treated cattle, and acetate, choline, isoleucine, trimethylamine N-oxide, lipids, lipoproteins, cholesterol, and -glucose, which showed decreased concentrations. Citrate was found to be at higher concentration in non-responders and subclinicals only. When serum was analysed using ultra performance liquid chromatography electrospray ionisation mass spectrometry (UPLC/ESI-MS) and UPLC tandem MS (MS/MS), only samples from clinical cows could be discriminated. The molecular ions involved could be tentatively identified as a combination of taurine- and glycine-conjugated bile acids. These bile acids all became elevated. This study confirmed that liver enzyme activities (GGT, GDH) and phytoporphyrin concentrations are not effective as markers of early stage sporidesmin damage. Additionally, the new techniques were unable to detect early stage FE. However, some markers of treated cows were identified. The research does provide a strong foundation for future applications of metabolomics analysis, with MVA and time series statistics, for early stage FE diagnosis.