Electronic Theses and Dissertations (PhDs)
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Item Identification of (Novel) Immune Targets with Potential Roles in the Progression of Pancreatic Ductal Adenocarcinoma (PDAC)(University of the Witwatersrand, Johannesburg, 2024) Nsingwane, Zanele; Nweke, EkeneBackground: Pancreatic ductal adenocarcinoma (PDAC) is a lethal cancer with a growing incidence and mortality despite novel therapeutic strategies. Its aggressiveness and difficulty to treat suggest the need for a better understanding of associated molecular mechanisms which could be targeted for treatment. The complement signalling pathway may play diverse roles in PDAC by eliciting an immune response, inducing inflammatory responses, and may elevate pathways linked to chemoresistance. However, their role in the progression of PDAC is not fully understood. This study aimed to identify potential immune response-related targets in a group of patients. Methods: In this study, 30 tissue samples (tumours and corresponding normal tissues) were obtained from 15 PDAC patients, 34 plasma samples were obtained from 25 PDAC patients, 6 patients with chronic pancreatitis, and 3 healthy control participants. Targeted pathway-specific PCR analysis was conducted to determine the gene expression profiles of immune-response-related genes. The circulating levels of complement proteins C3 and C5 were further investigated. Pharmacological inhibition of the complement pathway in MIA PaCa-2 pancreatic cancer cell lines was performed and the effect on cells was assessed by cell proliferation, cell migration, and cell cycle assays. Finally, SWATH-mass spectrometry was performed to identify potential molecular mechanisms during inhibition. Results: The results identified C3 to be overly expressed in early PDAC compared to later stages in plasma (p=0.047). Pharmacological inhibition of the complement pathway led to increased cell growth (p<0.0001), proliferation (p=0.001) and migration (p=0.002) in vitro. Proteomic analysis implicated several proteins such as the mitochondrial and histone proteins, that could play a role in inducing this phenotype. Conclusion: Both Complement C3 and C5 are elevated in PDAC samples compared to healthy ones. Furthermore, the inhibition of the complement pathway was shown in vitro to result in a more aggressive phenotype by stimulating cellular growth, proliferation, and migration, indicating the involvement of complement C3 and C5 in tumour progression. This study helps to further delineate the role of the complement pathway in PDAC progression.Item An in-silico analysis of the glycosylation inhibitors Brefeldin A and Tunicamycin C in colorectal cancer; characterization of novel targets(University of the Witwatersrand, Johannesburg, 2024) Naidoo, VivashColorectal cancer (CRC), a prevalent malignancy in South Africa, is significantly influenced by posttranslational modifications such as glycosylation. This study investigates the complex interactions between genes, signalling pathways, and cellular processes involved in CRC progression and glycosylation. The glycosylation inhibitors, Tunicamycin and Brefeldin A, are known to hinder colon cancer cell proliferation, migration, and invasion, making them potential therapeutic agents. We used Swiss Target Prediction Software to identify target proteins for both compounds and revealed that Protein Kinase C Alpha (PRKCA), Peroxisome Proliferator- Activated Receptor Gamma (PPARG), and Mitogen-Activated Protein Kinase 1 (MAP2K1) are specific for Brefeldin A, and TK1 and PRKCA for Tunicamycin, respectively. These proteins were selected based on their potential role in the glycosylation process and their role in CRC-related pathways. Further, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis disclosed significantly enriched pathways, including Epstein-Barr virus infection, cellular senescence, and cancer pathways. The 3D-crystallographic structures of PrKC1 (PDB ID 6ar4), TK1 (PDB ID 1w4r), PrCK1 (PDB ID: 6ar4) and MAPK (PDB ID: 3eqc) were retrieved from RCSB Protein Data Bank. The compounds BSP and EGCG were downloaded from PubChem. All non-relevant co-crystallized molecules, including ions, crystallographic water, and others, were removed. Missing residues in the proteins were filled in using the MODELLER algorithm on the UCSF Chimera Graphic User Interface. Molecular docking of Tunicamycin C and Brefeldin A was performed with UCSF Chimera, and the docked conformations were visualised in Maestro and Chimera. The complexes with the top docking scores were selected and prepared for molecular dynamics simulation studies to offer structural and dynamic perspectives on the inhibitory potential of the compounds against the target proteins. The Origin Lab software tool was used to post- analyze the docking conformations. Molecular Dynamics simulation was conducted using Graphics Processing Units version of the Particle Mesh Ewald Molecular Dynamics engine in the AMBER18 suite. Our investigation into the dynamic events leading to the proximal binding of Tunicamycin at the pockets of TK1 and PrKC1 suggested that the binding of Tunicamycin induced a conformational perturbation of the 3D structures of these proteins, resulting in a structural deviation that inhibited their activity. Tunicamycin's time-based dynamics indicated a stable pattern, leading to optimal interaction and maximal stabilization in the hydrophobic pockets of TK1 and PrKC1. Binding energy calculations showed a high-affinity interaction of Tunicamycin with these proteins. Similarly, the structural investigation revealed that the binding of Brefeldin A to Mitogen- Activated Protein Kinase (MAPK) and Protein Kinase C (PrKC1) inhibited their activity. A detailed analysis of active site residues revealed crucial residues that contributed to the binding stabilization of Brefeldin A. It was noted that the Brefeldin A/MAPK complex produced a binding energy of -22.18±4.50Kcal/mol while the Brefeldin A/PrCK1 complex produced a binding energy of -23.90±5.36Kcal/mol. These findings provide crucial insights into designing novel inhibitors of TK1 and PrKC1, potentially blocking glycosylation progression in cancer treatment. This study underscores the potential for exploiting glycosylation inhibition as a therapeutic strategy against CRC, opening avenues to mitigate cancer progressionItem Gene expression patterns of signalling pathways in PDAC: towards inhibiting metastases(University of the Witwatersrand, Johannesburg, 2024) Xelwa, Ntombikayise Hendrietta MarciaPDAC has a poor prognosis, with its prevalence varying by geographical location. In South Africa, PDAC ranked seventh among all cancer-related deaths in 2020 for both sexes. Specifically, it was the seventh leading cause of cancer death among males and the sixth among females. In 2020, an estimated 1,982 cancer deaths in South Africa were attributed to pancreatic cancer, with 1,006 occurring in males and 976 in females. However, the annual reported number of PDAC deaths in South Africa varies. This study aimed to identify potential novel therapeutic targets for PDAC in patients from the African population. Following ethical approval, tissue from fifteen patients (15 tumour and 15 corresponding normal tissues) were obtained during Whipple procedures from PDAC patients who consented to the study. Despite the development of new treatment strategies, patient outcomes have not significantly improved underscoring the necessity for extensive research to identify novel treatment options. A discovery study was conducted to determine the gene expression profile of signalling pathway-related genes using PDAC tissue samples. Top upregulated pathways included those involved in cytokine signalling, receptor kinase signalling, and PI3/Akt signalling. SPP1 was one of the most highly expressed genes identified in PDAC patients compared to normal corresponding tissues suggesting its potential role in the progression of the disease. To investigate SPP1's role in PDAC, RNA interference was employed to knockdown SPP1 in a PDAC cell line, MIA PaCa-2. Knockdown was confirmed by a significant reduction in SPP1 expression at the mRNA level. Combining SPP1 knockdown with conventional chemotherapy used for PDAC, gemcitabine, resulted in a synergistic effect, leading to an enhanced early apoptotic response. The study also examined the migratory and invasive capabilities of MIA PaCa-2 cells, revealing a noticeable decline in these abilities upon reduction in SPP1 expression with gemcitabine treatment. Furthermore, proteomic analyses uncovered the complex network of cellular processes influenced by the downregulation of SPP1 and the synergistic effects of combination therapy. Altogether, the findings from this study demonstrate the role of SPP1 in PDAC indicating that it could serve as a promising therapeutic target. The synergistic effects observed when SPP1 knockdown was combined with gemcitabine treatment suggest a potential avenue for developing more effective treatments for PDAC while exploring tumour cell adaptation for survival.Item Evaluation of the postgraduate family medicine decentralised training programme at the university of Witwatersrand, South Africa, using the logic model(University of the Witwatersrand, Johannesburg, 2024) Erumeda, Neetha Joe; George, Ann ZetaPostgraduate family medicine decentralised training programmes were implemented in South Africa about 15 years ago, but the University of the Witwatersrand’s programme has not been comprehensively evaluated. This study evaluated the programme using a complex programme evaluation logic model based on linearity theory. This theory assumes ideal inputs and processes produce good programmatic outputs and outcomes. Resources and support were evaluated as inputs, postgraduate supervision and workplace-based learning as processes, supervisory feedback as outputs and workplace-based assessments as outcomes. A parallel convergent mixed-methods instrumental case study was conducted with purposively-sampled family physicians (n=11) and trainees (n=11) from five decentralised training sites. Semi-structured interviews were audio recorded, transcribed verbatim, and analysed inductively using MAXQDA 2020 software. Descriptive statistical analysis was conducted on components of registrars’ learning portfolios (scores, supervisory feedback, and skills competence) and examination results using Stata 14.2 software. An integrative analysis involving transforming the quantitative results to qualitative findings and drawing meta-inferences was conducted. The integrated findings were used to modify the initial logic model and identify key recommendations to optimise the programme. The integrative analysis identified the need for more material and human resources, university and district management support, and standardised resources, supervision, and learning practices. Supervisors’ knowledge, skills, and behaviours varied across sites and their feedback was insufficient regarding soft skills like clinical reasoning and patient negotiation. Workplace-based assessments did not meet the required standards across training years and districts. Interpersonal interactions with patients, peers, supervisors and other professionals, engagement in district activities, promoted learning. Registrars’ professionalism and self-learning need improvement. The key recommendations include more explicit national guidelines, sufficient support from the provincial department, university, and district management, well maintained infrastructure, sufficient skilled supervisors, more professional development training for supervisors, protected time for registrar learning, and better use of self-learning and reflection. Emulating successful contextual adaptations while addressing challenges across sites contributes to thriving decentralised training programmes in district health systems. An improved understanding of the concepts and their interrelationships in training programmes could be translated to similar decentralised training platforms across medical disciplines of sub-Saharan Africa or low-middle income countriesItem Relationship of diet and physical activity with genetic susceptibility to obesity: a longitudinal analysis in adults in South Africa(University of the Witwatersrand, Johannesburg, 2024) Muti, Monica; Chikowore, Tinashe; Ware, LisaBackground Obesity-related disease conditions are a major public health concern in South Africa, exerting a healthcare cost of between ZAR 30 million and ZAR 36 million, the bulk of which is due to hypertension and type 2 diabetes. Moreover, evidence reveals that women in South Africa have higher BMI compared to men, yet men exhibit less insulin sensitivity and reduced beta cell function as well as stronger associations of adiposity with type 2 diabetes compared to women. The mechanisms underlying these sex differences are not known. BMI is highly polygenic in nature; however, genetic prediction of BMI has mostly been conducted using data from European ancestry populations that have poor predictive capacity in African ancestry populations. Moreover, the relationship of polygenic risks and proteomic profiles with regards to susceptibility to obesity and related cardiometabolic traits is yet to be explored in African populations. It has also been reported that using variants associated with the statistical variance of quantitative traits (vQTLs) like BMI aids in the depiction of components of BMI genetic susceptibility, which interacts with environmental factors such as diet and exercise. However, such studies are limited in continental Africans. Aim This thesis, sought to determine the interplay of diet and physical activity with BMI genetic susceptibility. The specific objectives were: 1. To determine the association of physical activity with BMI in middle-aged black South African men and women. 2. To develop a highly predictive genetic risk score for BMI and test its longitudinal predictive ability in middle-aged black South African men and women. 3. To determine gene x lifestyle (GXE) interactions that influence BMI in Black South African adult men and women. Methods Data from 11853 adult men and women in the African-Wits-INDEPTH partnership for Genomic studies (AWI-Gen) Cohort was used to fulfil objective 1. To fulfil objectives 2 and 3, data from 5921 AWI-Gen cohort participants in the three South African (SA) sites and a sub-study of AWI-Gen focusing on the factors influencing the risk of type 2 diabetes mellitus among middle-aged black South African men and women (GSK) was used. For objective 1, a sex-stratified meta-analysis of cross-sectional data from the study participants was used to assess the association of physical activity with BMI. The PRS-CSx method was used to develop a multi-ancestry PRS for BMI and evaluate its longitudinal prediction of severe obesity to meet objective 2. For objective 3, the Levene’s test, implemented in the OCSA Package, was used to determine candidate gene-interacting variants that exhibited trait variance heterogeneity in the study population. Detailed methods are in the relevant sections for each objective. Results Meeting the recommended weekly physical activity levels of at least 150 minutes was associated with a BMI that was 0.80kg/m2 lower in men (95% CI = -1.14; -0.47) and 0.68kg/m2 lower in women (95% Ci = -1.03; -0.33). Sex and site-specific differences were also observed in domains of physical activity with an inverse relationship between transport-related physical activity and BMI being observed among men in Agincourt (beta = -1.15 kg/m2, 95% CI = -2.26; -0.04) and Nanoro (beta = -0.79 kg/m2, 95%CI = -1.25; -0.33). Work related physical activity was associated with lower BMI in Navrongo men (beta = -0.76 kg/m2, 95% CI=-1.25; -0.27) and Nanoro women (beta = -0.90 kg/m2, 95%CI = -1.44; -0.36). The multi-ancestry PRS demonstrated superior predictive ability, explaining approximately 1.9% of variance in BMI compared to 0.7% and 1.2% explained by two scores developed using single ancestry methods. In addition, over a period of ten years, the multi-ancestry PRS was associated with repeated measures of BMI (β = 1.51 p = < 0.001) and there was significant longitudinal PRS * sex interaction (Pinteraction = 0.029), prompting subsequent sex-stratified analysis. In the combined analysis of men and women, being in the top 20% of the PRS distribution (top 20) was associated with three times greater hazard of severe obesity (hazard ratio = 2.98, 95% CI = 1.75 - 5.07, p = 5.33e-05) compared to being in the bottom 20% of the PRS distribution (bottom 20). This observation was shown to be driven by women, where being a woman in the top 20 was associated with 3.5 greater hazard of severe obesity (hazard ratio 3.48, 95% CI = 1.96 – 6.16, p = 1.94e-05) compared to being in the bottom 20 while the associations were not significant in men (hazard ratio = 1.13, 95% CI = 0.24 – 5.37, p = 0.878). Comparison of the associations of dysglycaemia with PRS, BMI and the proteomic score revealed no apparent sex differences in the association between BMI PRS and dysglycaemia for most of the glycaemic markers except for Matsuda Index though men exhibited lower insulin sensitivity compared to women. The proteomic score predicted higher insulin resistance in women than in men. Gene x lifestyle interaction analysis revealed novel interactions between three genetic variants with diet and lifestyle factors. The effect of the rs557505940 variant on BMI was accentuated by higher fruit intake (betainteraction = 0.03, Pinteraction = 0.04) in the combined analysis of men and women while higher SES, carbohydrate intake and self- reported physical activity attenuated the effect of rs527747185 (betainteraction = -0.349, Pinteraction = 0.037), rs3016751 (betainteraction = -0.056, Pinteraction = 0.035) and rs188275749 (betainteraction = -0.048, Pinteraction = 0.0001) respectively on BMI in men. Conclusions Sex and geographical differences exist in associations between domains of physical activity and BMI. In addition, genetic risk better predicts incident severe obesity in women than in men while proteomic profiles have a weak correlation with PRS and show heterogenous associations with dysglycaemia, fat distribution, nutrient patterns and physical activity between men and women. Novel GXE interactions were also observed. These results underscore the need for further inquiry into the sex differences in genetic risk and environmental factors associated with BMI. Furthermore, a precision approach to obesity prevention and control, paying attention to the sex differences and contextual factors may be more efficient.Item The effects of indigenous South African plant extracts (cotyledon c. orbiculata and tulbaghia. violacea) on triple negative breast cancer cells(University of the Witwatersrand, Johannesburg, 2024) Alaouna, Mohammed; Dlamini, ZodwaThis dissertation explored the potential therapeutic applications of water and methanol extracts of C. orbiculata and Tulbaghia violacea, indigenous to Southern Africa, targeting triple-negative breast cancer (TNBC). TNBC, a significant subset of breast cancer cases, is notably challenging because of the absence of key hormone receptors, often leading to less favourable patient outcomes and a high relapse rate within five years. The research approach was both thorough and meticulous, utilising two cell lines: one representing normal breast tissue and the other representing TNBC. Extensive cytotoxicity assays were conducted to determine the IC50 values for TNBC cells, which is critical for understanding how plant extracts affect cellular activities such as migration, invasion, adhesion, cell cycle regulation, and apoptosis induction. Additionally, the antioxidant properties of these extracts were examined, which showed significant effects, especially in the aqueous extract of Tulbaghia violacea, on TNBC cellular dynamics. This study employed a comprehensive array of analytical techniques, including Fourier transform infrared spectroscopy (FTIR), gas chromatography-mass spectrometry (GC-MS), and nuclear magnetic resonance (NMR) spectroscopy, to identify the specific molecular constituents of these extracts. Computational docking studies have focused on the interactions between these molecules and the anti-apoptotic protein, COX2. Whole transcriptome sequencing of RNA from both TNBC and normal breast cells treated with T. violacea extract provided valuable insights into the affected signaling pathways. An antibody array assay further elucidated protein changes in the receptor tyrosine kinase (RTK) pathway. The half-maximal inhibitory concentration (IC50) values were determined for the aqueous and methanol extracts of T. violacea at 400 μg/mL and 820 μg/mL, respectively, and for C. orbiculata at 830 μg/mL and 700 μg/mL, respectively. Exposure to the water-soluble extract of T. violacea resulted in a marked increase in apoptosis in TNBC cells, with approximately 82% undergoing programmed cell death, compared to 32% in normal breast cells. Chemical profiling identified a range of compounds, including 36 distinct compounds identified through GC-MS and 61 identified through NMR, many of which bear structural similarities to known anti-cancer agents. Notably, five compounds demonstrated a high affinity forbinding to COX2, with d-glycero-d-galacto-heptose achieving an impressive docking score, surpassing several established COX2 inhibitors. This study highlights the therapeutic potential of T. violacea compounds and lays the groundwork for further exploration of their mechanisms of action and potential applications in cancer treatment. This emphasises the importance of investigating natural plant extracts as a source for the development of new and effective treatments for TNBC, which is an area of urgent need in oncologyItem An ethico-legal analysis of broad consent for biobank research in South Africa: Towards an enabling framework(University of the Witwatersrand, Johannesburg, 2024) Maseme, Mantombi RebeccaBiobanks preserve collections of human biological material and data for the benefit of medical research. Using and transferring human biological data and materials both inside and outside of South Africa is often a requirement of biobank research. Broad consent is allowed by the South African National Department of Health Ethics Guidelines but appears to be prohibited by section 13(1) of the Protection of Personal Information Act 4 of 2013. Additionally, the Act mandates that all personal data (including biobank sample data) be collected for legitimate, definite, and clearly stated purposes. There is room for several interpretations of the Act because of this discord between the two instruments. Given the connection between the transfer of samples and data, the long-term nature of biobanking, which makes it impractical to provide too much or adequate information because it is simply not available at the time of sample collection, and the various ways that the Protection of Personal Information Act 4 of 2013 have been interpreted, I aim to respond to the following question: How should South Africa’s current regulatory framework appropriately permit broad consent use for biobank research where the transfer of samples and their associated data are contemplated? The research question is addressed by applying ethical principles and theories, as well as analysing and evaluating relevant ethico-legal frameworks and literature. The study involves no research participants and no collection or analysis of any new data. Arguments for and against using broad consent for biobank research are discussed by demonstrating the potential for biobank research to do a great deal of good for humanity; the ambiguity in the current regulatory framework regarding whether broad consent is permissible for personal information/data; and the ethical justifiability of broad consent. In summary, the proposed regulatory framework amendments are those that would be required to allow for ethically justifiable biobank research broad consent use. These include removing regulatory ambiguity regarding broad consent use, ensuring adequate safeguards for research participants by specifying rules for data access and personal information processing, and incorporating consent form information requirements into the national Consent Template as specified in the National Department of Health Ethics GuidelinesItem Adverse childhood experiences and social and health outcomes in later life(2024) Naicker, Sara N.Background: Well-established literature points to early life experiences and childhood adversities setting the foundation for health and development and influencing life trajectories. Nurturing, responsive caregiving in a safe and stable environment is associated with healthy, productive lives throughout adulthood. On the other hand, adverse experiences in childhood are associated with poor health and wellbeing, risky behaviour and reduced human capital. How this adversity is measured and the context in which it is measured may provide insight into the relationship between adversity and outcomes over and above what has been found in high income countries. Aim: The overall aim of this study is to examine adverse childhood experiences (ACEs) in a South African birth cohort. Specific objectives of the study include: a) developing prospective and retrospective profiles of ACEs in the sample, b) establishing levels of agreement between these two profiles of ACEs, c) estimating the prevalence and clustering of ACEs in this population-based urban sample, d) examining the associations between exposure to ACEs and a range of physical and mental health and social outcomes, and e) understanding the role that recent stress plays in the relationship between exposure to ACEs and poor outcomes. Methods: This study uses a secondary analysis design using data from the longitudinal Birth to Thirty cohort. The cohort began in 1990 with the enrolment of 3,273 pregnant mothers and has followed the children born to these women for more than thirty years. The 10-item ACE Index developed by the CDC-Kaiser’s ACEs Study was expanded to include five additional ACEs common in the South African context – chronic unemployment, violence in the community, household death, parent death, and separation from parents. Prospective profiles of ACEs were collated from data collected over the first 18 years of the child’s life, initially reported by primary caregivers until age 11, then self-reported from ages 11 to 18. Retrospective profiles of ACEs were collected in young adulthood when the participants were 22 years old, along with an index of recent stressors. A series of human capital outcomes – those encompassing physical and mental health and psychosocial adjustment, were assessed at age 28. ACEs in the sample were conceptualized in three ways ‒ as single adversities, such as physical or sexual abuse, cumulative adversity in the form of the ACE score, and clusters of adversity determined by their patterning. Cohen’s kappa statistics and concordance rates were generated to establish the levels of agreement and consistency between prospective and retrospective reports of ACEs (timing) and between reports given by caregivers and children at age 11 (source). Descriptive statistics and latent class analysis were used to estimate the prevalence of ACEs and to explore the patterning of ACEs among participants. Logistic regression analysis explored associations between all three conceptualizations of ACEs and outcomes, disaggregated by sex. Mediation and moderation analyses were conducted to examine the influence of recent stress on mental health outcomes. Findings: Comparisons between prospective and retrospective reports of ACEs show that there is relatively low-to-moderate agreement between timing and sources of reports of ACEs. Agreement varies depending on the adversity in question – with greater levels for objective Naicker, S.N. 2023. Adverse Childhood Experiences and Social and Health Outcomes in Later Life experiences such as parental death and lower levels for subjective experiences such as chronic unemployment. Differences in agreement were partly due to prospective and retrospective reports identifying largely different groups of people; those who only report high exposure prospectively, those who only report high exposure retrospectively and those that overlap. Using either prospective or retrospective reports, the prevalence of ACEs in this sample were high, although there were significant decreases in prevalence from prospective reporting to retrospective reporting. ACEs tended to co-occur, and where one ACE was reported, the likelihood of others increased. Clusters of ACEs split distinctively into high-low:dysfunction abuse categories; with one group likely to have low exposure, another with high generalized exposure to all ACEs, a third with moderate exposure characterized by household dysfunction and a fourth with moderate exposure driven by emotional abuse and/or neglect. All three conceptualizations of ACEs were significantly associated with poorer outcomes. Single ACEs such as physical, sexual and emotional abuse, and exposure to intimate partner violence, were independently and strongly associated with poorer outcomes in adulthood. Increased exposure to ACEs, or cumulative adversity, was also linked to poorer outcomes in a graded manner, with the likelihood of experiencing poor outcomes increasing along with exposure. The clusters with high levels of exposure to ACEs and moderate levels of exposure driven by emotional abuse were most at risk for poor outcomes. There were significant differences in exposure to ACEs, outcomes and the associations between the two by sex. Associations also differed for prospective and retrospective reporting with the strength of association varying depending on the outcome in question. Recent stressors were found to play a confounding role in the relationship between ACEs exposure and poor outcomes. Although recent stressors had a different impact on those who reported high ACEs exposure prospectively versus those who reported high ACEs exposure retrospectively. The influence of recent stressors on the mental health of those who reported high exposure to ACEs prospectively supported a sensitization model. In contrast, the role of recent stressors on the mental health of those who reported high exposure to ACEs retrospectively supported a stress inoculation model. This suggests two potential pathways for risk. Conclusion: In combination and accumulation, it is demonstrated here that adverse experiences in childhood have an impact on health and wellbeing in adulthood. Specific individual ACEs can be teased out for their independent effect on outcomes, but the additive effects of multiple adversities lead to almost exponential increases in the risk for a myriad of negative physical and mental health and social outcomes. These findings provide important links from South Africa’s context of high levels of violence in all forms and multiple hardships that families with large burdens of care endure, with little support, to many of the human capital outcomes on which productive, healthy and happy lives depend. Born at the dawn of democracy, with anticipation for opportunity, many of the children in this cohort were raised in contexts of adversity that may have been experienced as normative in those settings. Regardless of whether these experiences leave enough of a mark to be recalled later in life, the strain of cumulative adversity has had persistent and serious effects on their mental health, their ability to finish school, find a job and stay out of trouble.Item Integrated biological and behavioural assessment of human immunodeficiency virus and sexually transmitted infections among tertiary student men who have sex with men in Nairobi, Kenya: project bespoke(2024) Mwaniki, Samuel WaweruAims: The aims of this study were to: assess the appropriateness and acceptability of using respondent-driven sampling (RDS) as a strategy for recruiting tertiary student MSM (TSMSM) in a HIV/STIs bio-behavioural survey, estimate HIV/STIs prevalence and associated risk factors among TSMSM, explore experiences of TSMSM with access and use of health services, and assess healthcare providers’ (HCPs’) attitudes and perspectives towards care for TSMSM. Methods: The study was done in Nairobi, Kenya. During the first phase in September and October 2020, formative in-depth qualitative interviews were held with key personnel working in MSMfriendly health facilities (n=3), and TSMSM peer leaders (n=13), to assess the appropriateness and acceptability of using RDS to recruit TSMSM in a bio-behavioural survey. Subsequently, during the second phase in February and March 2021, six TSMSM selected from the 13 in the first phase, started off the RDS recruitment of another 242 TSMSM who participated in a cross-sectional biobehavioural survey to estimate HIV/STIs prevalence and associated risk factors. The survey was digitally self-administered on REDCap® platform. Participants received serological testing for HIV and Treponema pallidum, and pooled molecular testing for Chlamydia trachomatis, Mycoplasma genitalium, Neisseria gonorrhoeae, and Trichomonas vaginalis using urethral, anorectal and oropharyngeal samples. The third phase in September 2021 involved qualitative work to assess health access and delivery for TSMSM. In-depth interviews were held with TSMSM (n=22) purposely selected from the TSMSM (n=248) who participated in the biobehavioural survey. The interviews explored experiences of TSMSM with access and use of health services. During the same month, HCPs (n=36) took part in six focus group discussions to assess their attitudes and perspectives towards care for TSMSM. Qualitative data was analysed 2 thematically using NVivo v.11 (first phase) and v.12 (third phase), and quantitative data was analysed using Stata v.15 and RDS-Analyst v0.72 (second phase). Results: Formative qualitative work demonstrated that RDS was both appropriate and acceptable for recruiting TSMSM in the bio-behavioural survey. The median age of TSMSM who participated in the bio-behavioural survey was 21 years (interquartile range 20-22 years). RDS-adjusted prevalence of HIV, at least one of the five STIs, chlamydia, gonorrhea, Mycoplasma genitalium infection, trichomoniasis and latent syphilis were: 3.6%, 58.8%, 51.0%, 11.3%, 6.0%, 1.5% and 0.7%, respectively. Higher risk of HIV infection was independently associated with studying in private tertiary institutions, preferring a sex partner of any age, last sex partner being >25 years, meeting the last sex partner online and prevalent gonorrhea infection. Inconsistent condom use, and the last sex partner being a regular partner were independently associated with testing positive for at least one of the five STIs. From the qualitative work in the last phase, TSMSM vocalized experiences of prejudice, stigma and discrimination in public and institution-based health facilities, but felt they were equitably handled in community pharmacies, private and MSM-friendly health facilities. A majority of HCPs articulated positive attitudes towards care for TSMSM, while a minority expressed discomfort and displayed attitudes that likely reflected on their lived biases as it related to offering care and services to TSMSM. Conclusion: The demonstrated high HIV prevalence among TSMSM in Nairobi reflects the urgent need for tailored structural, biomedical and behavioural prevention interventions for this young key population. Structural interventions are required to address the environmental, social and economic factors that influence individual risk and protective behaviours in relation to HIV infection. Biomedical interventions such as pre-exposure prophylaxis are necessary to reduce the chances of transmission of HIV. The observed high prevalence of curable STIs calls for interventions to improve prevention, as well as prompt detection and treatment of these STIs. This is important because untreated STIs biologically potentiate the transmission and acquisition of HIV, and cause considerable morbidity on their own. Furthermore, there is a need for interventions that foster inclusive attitudes among, and improve the knowledge/skills of HCPs in tertiary institution-based health facilities, so as to make services more culturally competent, equitable and accessible for TSMSM.Item Potential protein biomarker discovery for gallbladder cancer in a South African cohort(2024) Baichan, PavanGallbladder cancer (GBC) has a poor prognosis with the prevalence of GBC varying according to geographical location. The prevalence of GBC in South Africa is poorly tracked, and the molecular mechanisms associated with GBC in African patients are inadequately understood. This study aimed to determine dysregulated proteins in tissue and blood plasma in South African GBC patients to identify potential molecular mechanisms of disease progression and plausible biomarkers. Following ethical approval, tissue from 27 GBC, 13 gallstone disease (GD), and five normal tissues were obtained. Blood plasma was collected from 54 GBC and 73 benign biliary pathology (BBP) patients who consented to the study. A bottom-up proteomics approach was undertaken, using PAC and HILIC digestion methods, and SWATHMS for quantitative proteomic profiling. Hierarchical cluster analysis, PCA analysis, and Spearman’s rank correlation analysis were performed. Furthermore, pathway and network analyses were conducted. There were 62, 194, and 105 dysregulated proteins in the GBC/Normal, GBC/GD, and GD/Normal group comparisons, respectively, and 33 dysregulated proteins in the GBC/BBP plasma comparison. The dysregulated proteins in GBC patients enriched pathways involved in smooth muscle contraction, metabolism, extracellular matrix organisation and interactions, innate immunity, and platelet and neutrophil degranulation. Further analysis showed that S100A8 and S100A9 were downregulated in GBC plasma patients with GD history compared to those with no GD history. Additionally, APOE and ITIH3 were elevated in non-metastatic staging GBC patients. Seven proteins were found to be commonly dysregulated in GBC/GD and GBC/BBP comparisons and another two proteins were commonly dysregulated in the GBC/Normal, GBC/GD, and GBC/BBP comparisons, termed “Commonly dysregulated proteins (CDPs)”. Quality control assessment of the MS2 fragment ion chromatograms of the CDPs indicated strong signal-to-noise ratios and correct fragment-to-precursor matching. The CDPs could distinguish between GBC and controls and the Spearman’s rank correlation test showed significant correlations between the expression of the CDPs. The identified dysregulated proteins aid in further understanding the molecular mechanisms associated with GBC in patients with African ancestry. The alteration of specific proteins in tissue and plasma samples suggests their potential use as biomarkers for GBC patients in this sample cohort.