Masters Degrees (Physiological Sciences)
Permanent URI for this collection
Browse
Browsing Masters Degrees (Physiological Sciences) by browse.metadata.advisor "Joseph, Danzil"
Now showing 1 - 5 of 5
Results Per Page
Sort Options
- ItemCortisol, glucose, and oxidative stress: a unified hypothesis of psychological stress and neurological dysfunction(Stellenbosch : Stellenbosch University, 2021-12) Geddie, Hannah; Essop, M. Faadiel; Joseph, Danzil; Stellenbosch University. Faculty of Science. Dept. of Physiological Sciences.ENGLISH ABSTRACT: Chronic stress is a global pandemic implicated in the onset and progression of numerous non- communicable diseases. Although promising hypotheses regarding the relationship between stress and adverse health outcomes do exist, there are relatively limited insights regarding the underlying mechanisms driving this phenomenon. Related literature describes glucose metabolic perturbations mediated by chronic stress and their putative impact, particularly in terms of long-term functional defects observed in stress-related disorders. Furthermore, prior research established that oxidative stress-induced shunting of glycolytic intermediates into nonoxidative glucose pathways (NOGPs), such as the hexosamine biosynthetic pathway (HBP), is linked to pathophysiologic outcomes. Although changes in glucose metabolism and the induction of oxidative stress in the brain are key features of chronic stress, the mechanisms underlying such alterations and the related interplay between mediators are poorly understood. This review will therefore begin by establishing what is currently known about glucose metabolism and oxidative stress as primary, individual contributors to stress-induced pathology. Hereafter the two will be explored in tandem, culminating in a novel hypothesis whereby perturbed glucose metabolism and oxidative stress converge to promote and exacerbate pathological remodeling of the brain. We propose that prolonged activation of the stress response leads to increased brain HBP flux that impacts cerebral redox homeostasis and thereby results in neurological dysfunction.
- ItemThe detection of stress-related diseases: establishment of two unique methods to discover circulatory phospho- and glycoproteins(Stellenbosch : Stellenbosch University, 2023-12) Smith, Logan Jason; Essop, M. Faadiel ; Joseph, Danzil; Stellenbosch University. Faculty of Science. Dept. of Physiological Sciences.ENGLISH ABSTRACT: Introduction: Psychosocial stress has strong links to numerous chronic diseases related to the dysregulated activation of the physiological stress system. This heightens the burden of mortality as there is a robust relationship between chronic psychosocial stress and non- communicable diseases. Hence there is a robust impetus for the identification of novel, circulating biomarkers to earlier detect stress-related chronic diseases. Although protein post-translational modifications such as glycosylation and phosphorylation can act as putative markers of pathophysiology, their relatively low abundance complicates extraction and identification from samples with a high dynamic range. The main aim of this study was therefore to establish two unique enrichment methods for circulatory glycoprotein and phosphoprotein extraction that would then be applied in a preclinical model of chronic psychosocial stress. Methods: Phosphoprotein enrichment was performed using functionalized magnetic particles while glycoprotein enrichment occurred using lectin-bound magnetic particles. Both these methods were tested using a known purified phosphorylated and glycosylated protein and compared to bottom-up proteomics methodology using rat serum. The latter was obtained from a rat model of chronic stress that is well-established in our laboratory (n = 16 controls versus n = 16 stressed rats). These were randomly selected for proteomics analysis to assess the efficiency of retrieval in enriched versus unenriched samples. Fractions were analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and proteins visualized using Coomassie and specific fluorescent staining. Here, the relevant Pro-Q stains were employed for the identification of glycosylation and phosphorylation, respectively. The coupling of such enrichment methods to LC – tandem mass spectrometry (MS/MS) was enabled by employing various preparation steps such as deglycosylation and digestion. An exogenous protein was also included as part of the sample preparation to ensure quality control analysis of the LC-MS/MS experiment. Results: SDS-PAGE analyses and staining methods revealed non-specific enrichment with regards to intact protein retrieval. In addition, LC-MS/MS data demonstrated that enrichment using the current set of affinity materials was inadequate for glycopeptide and phosphopeptide retrieval in serum. Conclusion: A lack of enrichment indicates that stringent sample preparation is needed for biological materials with a high dynamic range. This may also be due to the porous nature of both materials employed for phospho- and glycoprotein/peptide enrichment respectively. A combination of enrichment and/or depletion methods may therefore be beneficial for deeper analysis of the blood proteome. These enrichment techniques and the subsequent sample preparation still require further optimization to derive more definitive conclusions in the chronic stress context.
- ItemEstablishing and validating an in vivo rodent model of chronic restraint stress(Stellenbosch : Stellenbosch University, 2022-12) Van Wyk, Minette; Essop, M. Faadiel; Joseph, Danzil; Stellenbosch University. Faculty of Science. Dept. of Physiological Sciences.ENGLISH ABSTRACT: Introduction. Psychological stress has emerged as one of the health epidemics of the 21st century and provides an impetus for increased investigation into the effects of a dysregulated stress response on whole body physiology. Although previous studies helped to clarify the association between chronic psychological stress and the onset and progression of cardiovascular diseases, a paucity of mechanistic insights underlying this association remain. Considering the complex nature of the stress system and the similarities that exist between humans and animals, it is therefore ideal to use rodent models to investigate stress-related disorders. Although the incidence and onset of various disorders in humans are gender-specific, clinical, and preclinical research using male subjects still far outnumber those using females. This study therefore aimed to establish and validate an in vivo model of chronic restraint stress in male and female Wistar rats. Materials and Methods. Male and female Wistar rats were subjected to a 4-week restraint stress protocol versus matched controls. Following this, behavioral tests (elevated plus maze [EPM] and tail flick task) were performed together with an assessment of body weight changes and biochemical biomarkers to ascertain whether the model was successfully established. Results & Findings. Our data revealed that male stressed rats displayed a decreased percentage change in body weight over time versus controls (p<0.01). Furthermore, the male stressed group exhibited increased plasma corticosterone levels compared to controls (p<0.01), while no significant differences were detected for plasma adrenocorticotropic hormone (ACTH) concentrations. Male brain-derived neurotrophic factor levels (biomarker for neuronal survival and growth) were lower in the stress group versus controls (p<0.05). Stressed males also displayed a reduced number of attempts into the open arms of the EPM versus controls (p<0.05). There were no significant weight changes for female rats. However, stressed females exhibited lowered plasma corticosterone levels versus controls (p<0.05), while also displaying higher plasma ACTH concentrations compared to the control group (p<0.05). Stressed females also displayed increased rears (as assessed by EPM test) versus matched controls (p<0.01). Our findings reveal intriguing sex-based differences in response to a chronic restraint stress protocol, with males displaying a depressive-type phenotype while females exhibited a post-traumatic stress disorder phenotype. Sex-specific preclinical research can provide unique insights into the various mechanisms driving stress-related diseases and should eventually lead to the identification of novel diagnostic and therapeutic targets.
- ItemExploring mechanisms underlying stress-related cardiometabolic complications(Stellenbosch : Stellenbosch University, 2022-03) Cairns, Megan Louise; Essop, M. Faadiel; Joseph, Danzil; Stellenbosch University. Faculty of Science. Dept. of Physiological Sciences.ENGLISH ABSTRACT: Introduction: Cardiometabolic diseases (CMD) constitute a major, global burden of disease. Of concern, psychosocial stress is an emerging risk factor for cardiovascular diseases onset. Although associations between psychological complications and CMD are well established, the underlying mechanisms driving this process require further investigation. Oxidative stress is linked to both chronic stress and CMD progression and can elicit detrimental sequelae. For example, it can augment PARP activity and thereby shunt glycolytic intermediates into the hexosamine biosynthetic pathway (HBP). Increased HBP flux can in turn lead to dysregulated O-GlcNAcylation of target proteins, thereby potentiating cardiometabolic complications. As excessive HBP flux is observed in a range of CMD, we hypothesized that increased oxidative stress and HBP activation play a key role in stress-mediated CMD onset and progression. Our main objective was to assess the degree of total protein O-GlcNAcylation and redox status of cardiac tissues isolated from stressed versus control rats. Methods: This preclinical study exposed male Wistar rats (n = 14 per group) to 9.5 weeks of unpredictable chronic mild stressors versus non-stressed controls. Behavioral tests were initially conducted to assess the presence of depression and anxiety. Post- euthanasia, plasma corticosterone, and epinephrine levels were evaluated, while myocardial redox state, glucocorticoid receptor expression, and activity, as well as HBP activation, were also determined. Results: Stressed rats displayed an anxious phenotype, with lowered plasma corticosterone levels (p = 0.0394 vs. controls) and higher plasma epinephrine concentrations (p = 0.0284 vs. controls). Our data revealed increased cardiac lipid peroxidation (p = 0.0421 vs. controls) but without any alterations in antioxidant defense systems (catalase, total glutathione, and oxygen radical absorbance capacity). Cardiac HBP activation remained unchanged between the experimental groups, with no significant alterations to GFAT1 or O-GlcNAc expression. Conclusion: These data show that the stress protocol triggered an anxious phenotype together with increased myocardial oxidative stress. The elevated oxidative stress may likely occur as a result of increased reactive oxygen species production instead of an impaired antioxidant system. The HBP data suggest that the mild degree of oxidative damage in the heart was insufficient to alter normal glucose metabolism.
- ItemTKTL1 plays a critical role in ameliorating intracellular oxidative stress under hyperglycemic conditions(Stellenbosch : Stellenbosch University, 2018-03) Reuter, Sulandy; Essop, M. Faadiel; Joseph, Danzil; Stellenbosch University. Faculty of Science. Dept. of Physiological Sciences.ENGLISH ABSTRACT: Diabetes is a growing, global concern, reaching a prevalence far beyond the projected statistics. Hyperglycemia is a common denominator of type 1 and type 2 diabetes mellitus that if left untreated, results in detrimental outcomes, e.g. oxidative damage. The pentose phosphate pathway (glucose-6-phosphate dehydrogenase [G6PD] and transketolase [TKT] as rate-limiting enzymes) offers an alternate ‟safer” route for excess metabolic fuels, while also replenishing anti-oxidant defenses and thus potentially offers value as a novel therapeutic target. As TKT-like 1 (TKTL1), a relatively novel member of the transketolase family, has been overlooked in this regard, we employed a TKTL1-knock-out (KO) mouse model as a targeted approach to evaluating TKTL1 function. Multiple low doses of streptozotocin (STZ) (40 mg/kg, intraperitoneal injections on five consecutive days) were used to induce hyperglycemia, a pathophysiologic state known to culminate in oxidative stress and damaging outcomes. Hyperglycemia incidence was 40% and 90% in wild-type (WT) and TKTL1-KO STZ-injected male groups, respectively, while only the TKTL1-KO STZ group displayed significantly higher blood glucose levels compared to the WT control (p<0.001) and the TKTL1-KO control group (p<0.001). The male TKTL1-KO STZ-treated group displayed larger relative liver weights (p<0.001) and kidney weights (p<0.05), respectively, compared to both WT and TKTL1-KO controls. Gastrocnemii weight of the TKTL1-KO STZ-treated group was lower than that of the WT control group (p<0.01). There was a significant, negative correlation between liver G6PD activity and blood glucose levels (Spearman’s correlation coefficient [r]=-0.37, p=0.03) while this correlation was not observed in cardiac tissue. In the latter, both TKTL1-KO groups displayed ~50% diminished transketolase activity compared to the WT control (p<0.001). No significant differences were noted in any of the oxidative stress and damage assessments of blood, heart, or liver samples. However, several adaptations were revealed in hepatic anti oxidant defenses: STZ-treated TKTL1-KO mice displayed increased catalase activity (p<0.05), with catalase activity showing a significant, positive correlation with blood glucose (r=0.39, p=0.02); ferric reducing anti-oxidant power (FRAP) concentrations as well as the reduced glutathione (GSH):to oxidized glutathione (GSSG) ratio were, significantly decreased in STZ-treated TKTL1-KO group (p<0.05) while both of these measurements were negatively correlation with blood glucose levels (r=0.45, p<0.01 for FRAP, r=-0.36, p=0.03 for GSH:GSSG ratio). Our data reveal that TKTL1 may play an anti-oxidant role in the liver to protect against hyperglycemia and to maintain overall glucose homeostasis. Thus our study demonstrates novel insights into TKTL1 function and shows that it remains a feasible therapeutic target for the treatment of diabetes.