Saudi Cultural Missions Theses & Dissertations
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Item Restricted Characterization of peripheral immune dysregulation and brain neuroimmune pathophysiology in the Df(16)A+/- Mouse Model of 22q11.2 Deletion Syndrome, a high genetic risk factor for schizophrenia(Saudi Digital Library, 2025) AlFadhel, Hassan Ali H; Jonathan, CraigSchizophrenia (SCZ) is a severe and chronic neurodevelopmental disorder that imposes a profound global health burden. Among its complex genetic underpinnings, the 22q11.2 Deletion Syndrome (22q11.2DS) stands out as the highest-known genetic risk factor, with approximately 25-30% of affected individuals developing a psychotic disorder. Recent research in SCZ has undergone a paradigm shift, moving beyond purely neurocentric models to embrace growing evidence that implicates immune dysregulation and neuroinflammation as critical etiological processes. While 22q11.2DS is known to confer a complex immunological phenotype, including foundational T-cell deficits and a high incidence of autoimmune disorders, the functional consequences of this genetic 'first hit' on immune reactivity and central nervous system (CNS) vulnerability is not well understood. Therefore, the primary objective of this thesis was to systematically characterize immune dysregulation in the Df(16)A+/- mouse model of 22q11.2DS to test the hypothesis that the genetic deletion programs a dysfunctional response to subsequent inflammatory challenges, thereby providing a plausible biological substrate for neuropsychiatric risk. This investigation utilized the Df(16)A+/- mouse model, which carries a chromosomal deletion syntenic to the human 22q11.2DS critical region. Our experimental strategy involved a comprehensive profiling of the peripheral and central immune systems at a homeostatic baseline and following provocation with distinct immune stimuli, including a sustained Toll-like receptor 7 (TLR7) agonist (Aldara) and an acute Toll-like receptor 3 (TLR3) agonist (LMW-Poly(I:C)). Given the limited characterization of the Df(16)A+/- immune profile and the incompletely elucidated mechanisms linking 22q11.2DS to psychiatric risk, this thesis employed both hypothesis-driven and exploratory analytical frameworks to systematically characterize the immune phenotype. Primary analytical techniques included multi-plex immunoassays (Luminex), multi-parameter flow cytometry, and quantitative immunohistochemistry. This central analysis was supported by the development of a high-throughput, objective morphometric pipeline to robustly quantify glial reactivity and neuronal cytoarchitecture within the hippocampus. Our results demonstrate that the Df(16)A+/- genotype is not immunologically silent but confers a "first hit" of subtle, multi-system dysregulation. Peripherally, this is defined by a "central deficit, peripheral compensation" model, with foundational T-cell developmental deficits in the bone marrow and thymus that are compensated for in circulating blood and spleen populations. This cellular alteration is mirrored by a rewired baseline molecular milieu, characterized by increased inter-animal heterogeneity and a less interconnected systemic cytokine network. Centrally, this state corresponds to a latent glial vulnerability, defined by a subtle but significant reduction in the morphological complexity of hippocampal microglia, anatomically co-localized to the dorsal CA1 and dentate gyrus. Subsequent immune challenges revealed this vulnerable baseline dictates a "second hit" of a profoundly dysfunctional and uncoupled neuro-immune response. The systemic reaction to the TLR7 challenge was both attenuated in magnitude, with blunted splenomegaly and pro-inflammatory cytokine upregulation, and qualitatively dysregulated, exemplified by the paradoxical downregulation of IFN-alpha and the collapse of the organized inflammatory network. Critically, this peripheral impairment was associated with a complete uncoupling of the glial inflammatory cascade in the dorsal hippocampus: wildtype mice mounted a coordinated neuroinflammatory response, whereas Df(16)A+/- mice exhibited a dissociated cascade where microglia adopted a morphologically reactive state in the complete absence of a canonical astrocyte (GFAP+) response. This microglial response was itself dysfunctional, occurring without the expected upregulation of the functional protein Iba1. This glial uncoupling was directly associated with the neuronal findings: Df(16)A+/- mice exhibited an altered parvalbumin-positive (PV+) interneuron response, lacking the significant change in cell counts observed in wildtype animals following the challenge. Findings from an acute TLR3 challenge, qualified by a severe physiological confound of hypothermia, also pointed towards a rewired inflammatory program by revealing a paradoxical central myeloid response. The central conclusion of this thesis is that the 22q11.2DS-relevant genetic deletion establishes a "first hit" of homeostatic instability that dictates a functionally impaired "second hit" response to immune challenges. This work links this high-impact genetic risk factor to a specific CNS vulnerability, defined by an uncoupled neuroinflammatory cascade, where microglia activate but astrocytes fail to respond, and a corresponding genotype-specific PV+ interneuron response, where the significant change in cell counts seen in wildtype animals was absent in the Df(16)A+/- mice. This integrated phenotype provides a novel and plausible biological substrate for the increased neuropsychiatric risk observed in the human 22q11.2DS population.35 0Item Restricted IMPACT OF DIABETES ON ACUTE LUNG INJURY: UNRAVELLING MECHANISMS TO IMPROVE OUTCOMES(University of Georgia, 2025) Alanazi, Abdulaziz Hammad; Shenoy, SomanathDiabetes mellitus (DM) is a common comorbidity in acute lung injury (ALI) patients. Despite its well-established prevalence and known complications affecting many organs, our understanding of the lung as a target organ in DM remains limited, relying heavily on inconsistent findings from observational data. This thesis aims to investigate the effects of DM on lung health and ALI development and outcomes using a combinatorial approach incorporating clinical, cellular, and pre-clinical data. The Fluids and Catheters Treatment Trial (FACTT) dataset and plasma samples were analyzed to assess the impact of pre-existing diabetes on ALI patients. Additionally, the molecular effects of advanced glycation end-products (AGEs) were examined in human microvascular lung endothelial (HMLE) and epithelial (A549) cells, while the influence of DM on lung health and ALI severity was evaluated in 12-week-old type 1 DM mice with or without sepsis. Clinically, we found that ARDS patients with DM had lower survival rates and longer hospital and ICU stays compared to those without DM. Mechanistically, AGE treatment (50 µg/ml) modulated tight junction proteins and impaired barrier function. AGE exposure also activated key inflammatory pathways (AKT and P38 MAPK) and elevated the expression of inflammatory cytokines (TNF-α, IL-1β, IL-10, and IL-6). Experimentally, mouse DM lungs exhibited significant inflammation and edema compared to non-DM lungs. RNA sequencing identified genes involved in inflammation and endothelial barrier dysfunction, supporting the link between DM and lung injury. Furthermore, DM increased the risk of worsening ALI in sepsis mice compared to on-DM sepsis. Overall, our findings indicate that DM is a risk factor for lung inflammation and worsened ALI progression. This study highlights the potential for improving ALI outcomes by targeting diabetes-associated inflammatory and metabolic pathways, offering new insights for therapeutic interventions in diabetic patients at risk for ALI.24 0Item Restricted INVESTIGATING LINKS BETWEEN THE IMMUNE RESPONSE AND HELICOBACTER PYLORI-MEDIATED DISEASE OUTCOMES.(University of Nottingham, 2024) Alzahrani, shatha; Karen, RobisonBackground: Helicobacter pylori is the primary cause of chronic gastritis and peptic ulcer disease and a significant risk factor for gastric adenocarcinoma. Numerous studies have reported the role of cytokines in the pathogenesis of gastric inflammation during H. pylori infection and disease outcomes are more likely with virulent strains that carry the Cytotoxin Associated Gene Pathogenicity Island (cag PAI). Interleukin-16 (IL-16) is a multifunctional cytokine implicated in various chronic inflammatory conditions, such as inflammatory bowel disease, asthma, and rheumatoid arthritis. Elevated IL-16 levels have previously been observed in the serum of gastric cancer patients compared to healthy controls. However, the connection between IL-16 production and H. pylori infection remained unclear. A comprehensive study profiling plasma cytokine levels across different stages of gastric cancer prognosis, including non-atrophic gastritis, atrophic gastritis, intestinal metaplasia, and gastric cancer could provide insights into the correlation between cytokines and disease progression and their potential as biomarkers for gastric cancer. Previous research demonstrated H. pylori's ability to induce IL-16 production in gastric epithelial cell lines. Since monocytes and dendritic cells are among the first cells to encounter H. pylori after it breaches the gastric epithelial barrier, the effect of H. pylori and its virulence factors on IL-16 production in these cells has not been explored. Aims; A key objective of this thesis was to measure IL-16 concentrations in plasma samples from H. pylori-positive and negative patients and to investigate potential links between IL-16 and H. pylori-mediated gastro-duodenal disease. The study explored relationships between serum IL-16 and colonization by more virulent cagA-positive H. pylori strains, associations with gender and smoking status, the presence and severity of histopathological changes in the gastric mucosa, oesophageal diseases, and the impact of H. pylori eradication therapy and other co-expressed inflammatory cytokines. Additionally, the thesis aimed to explore the effects of H. pylori infection and its virulence factors, and bile metabolites, on cytokine production by monocytes and dendritic cells. Another significant aim was to understand differential cytokine expression concerning gastric cancer progression and their potential as biomarkers for the disease. Materials and Methods; Gastric biopsy and plasma samples were collected from H. pylori-infected and non-infected patients, including those who had received successful eradication therapy for the infection. IL16 mRNA and plasma IL-16 levels were measured using RT-qPCR and a commercial ELISA kit respectively. cag PAI-negative H. pylori, cag PAI-positive H. pylori, and isogenic virulence factor mutants co-cultured with human peripheral blood monocytes, monocyte-derived dendritic cells, and THP-1 and KG-1 cell lines for 24 hours. IL-6, IL-10 and IL-16 cytokines were measured by ELISA and flow cytometry. 320 plasma samples were collected from H. pylori-infected and uninfected patients with various gastric conditions, including non-atrophic gastritis, atrophic gastritis, intestinal metaplasia, and gastric cancer. Concentrations of cytokines and chemokines (including IL-1β, IL-4, IL-6, IL-8, IL-10, IL-12p70, IL-13, IL-16, IL-17, IL-18, IL-23, CCL2, CCL3, CXCL10, IFN-γ, and TNF-α) were measured using multiplex assays from Mesoscale Discovery (MSD). Results: IL-16 was detected in all plasma samples, from both H. pylori-infected and uninfected patients, with wide variation in both groups, and there were no significant differences. IL-16 levels were not significantly associated with gender, age, or smoking status, nor did they differ based on whether the colonizing strain was cagA-positive or negative. No link was found between IL-16 concentration and histopathological changes in the gastric mucosa, or oesophageal diseases, and there were no significant differences in plasma IL-16 levels before and after H. pylori eradication. The results showed that H. pylori virulence factors did not alter cytokine production by monocytes and dendritic cells. Interestingly, GC patients exhibited a significant reduction in IL-16 plasma levels compared to individuals with gastritis, peptic ulcer disease, and healthy controls. This outcome contradicts a previous study that reported elevated IL-16 serum concentrations in GC patients relative to healthy controls, likely due to different patient recruitment criteria. This finding's reliability was further substantiated by similar results from another ethnic group, a Mexican patient population, which showcased a reduction in both IL-16 and IL-18 plasma levels in GC patients compared to those with non-atrophic gastritis, atrophic gastritis, and intestinal metaplasia. Conclusion: This comprehensive study analyzed various sample types, including plasma, gastric tissue, cell lines, and peripheral blood monocytes, utilizing techniques such as ELISA, MSD, RT-qPCR, and Flow Cytometry. Participants were categorized as healthy controls, H. pylori-infected and uninfected patients, individuals with different gastric conditions, and those from diverse ethnic backgrounds. The study considered multiple parameters potentially influencing plasma IL-16 levels, including age, gender, smoking status, cagA status, and oesophageal disorders. The study concluded that H. pylori infection does not affect IL-16 cytokine levels. A suggested correlation between IL-16 and IL-18 cytokines in the context of gastric cancer presents a novel research avenue. Both interleukins could be components of a larger immune regulatory network. Disruption in this network might lead to inadequate tumor control. Further research should be conducted to explore how IL-16 and IL-18 interact and influence each other in the context of cancer immunology. Investigating the combined roles of IL-16 and IL-18 can provide deeper insights into their contributions to immune response modulation.17 0Item Restricted The Modulation Effect of Inflammatory Cytokines on T cell Proliferation in Hypertension(University of Glasgow, 2024-05) Alsheikh, Eman; Marta, Czesnikiewicz-Guzik; Tomasz, GuzikHypertension is a common medical condition with very serious target organ consequences, increasing the risk of heart disease, stroke, and severe health complications. Despite the identification of various mechanisms (vascular, renal, and central mechanism) contributing to the pathogenesis of hypertension, the majority of cases lack a clear aetiology. Emerging evidence has established a significant association between hypertension and immune responses, particularly involving adaptive immune cells and inflammatory cytokines. Immunosuppressive drugs and cytokine inhibitors have shown potential in mitigating hypertension, suggesting a crucial role of the immune system in this condition. Given the central role of T lymphocytes in the adaptive immune response, this study hypothesises that, in the context of hypertension, inflammatory cytokines can modulate T cell activation independently of antigen stimulation. To test this hypothesis, total T cells were isolated from the spleens and PBMCs of normotensive and hypertensive mice and exposed to a range of cytokines, including TNF-α, IL-6, IL-15, IFN-γ, IL-7, IL-1β, IL-17A, IL-2, and IL-12, using different stimulation protocols. Aiming to understand the effects of these cytokines on T cell proliferation, differentiation, and the expression of activation markers such as CD69. Our findings highlight the varying abilities of cytokines to sustain T cell viability, with IL 7, IL-15, and IL-6 demonstrating a tendency for greater efficacy compared to other cytokines. In addition, IL-7 and IL-15 significantly impact T cell proliferation, notably affecting the CD8+ T cell population. However, despite these effects, no significant difference was detected between normotensive and hypertensive T cells in response to IL-7 and IL-15. This suggests that while these cytokines are potent in driving T cell proliferation, their influence is not specifically heightened in the context of hypertension. In GSEA and KEGG analyses, the Ca2+ signalling pathway was distinctively activated in response to IL-7 and IL-15 in Ang II induced hypertension. Conclusion: These data imply that most studied cytokines linked to hypertension pathology do not substantially affect normotensive or hypertensive T cells in a murine model. However, T cell proliferation was elevated in both Sham and Ang II mice in response to IL-15 and IL-7. Together, the data presented in this thesis warrant further investigations into the role of cytokines in hypertension and may point to IL-15 or IL-7 as biological targets for antihypertension therapy.49 0Item Restricted Investigating the mechanisms underpinning antipsychotic-mediated pneumonia(Saudi Digital Library, 2023-11-01) Howsawi, Alanood; Caroline, Copeland; Athanasios, SekerisBackground: The mechanisms behind the association between antipsychotics and pneumonia risk remain inadequately understood. Existing studies propose that antipsychotics might attenuate the immune response through cytokine modulation. The present study aimed to explore the impact of olanzapine and quetiapine on surface markers expression and cytokines released by CD4+ lymphocytes. Methods: Peripheral blood mononuclear cells (PBMCs) were isolated from four healthy volunteers. Olanzapine doses (6, 20, and 60 ng) and quetiapine doses (15, 50, 150 ng) were employed. The cells were stimulated with Staphylococcus aureus (SA) and Pseudomonas aeruginosa (PA). Stimulated cells were treated with olanzapine or quetiapine and compared to control cells. Flow cytometry was used to analyse CD25, CD28, CD127 (IL-7R-a), CD152 (CTLA4), CD274 (PD-L1), CD279 (PD-1) markers, and cytokines IL-2, IL-10, and IFN-Y. Statistical analyses were performed with GraphPad Prism 9.0, employing one way- analysis of variance (ANOVA) and Friedman tests (P < 0.05 for significance). Results: Olanzapine and quetiapine increased CD25 expression, with olanzapine 20 ng reached statistical significance in PA-stimulated cells. PD-L1 expression exhibited an upward trend in SA-stimulated cells treated with both drugs, with quetiapine 15 ng being significant. Both drugs significantly increased CD28 expression in the stimulated cells. CD152 decreased slightly with therapeutic doses of both drugs in PA-stimulated cells. CD127 and PD-1 expressions remained unchanged. Regarding cytokines, olanzapine 6 ng significantly reduced IL-2 release in stimulated cells, while quetiapine caused a mild IL-2 release in PA-stimulated cells. IFN-Y release slightly dropped in PA-stimulated cells in both drugs. Quetiapine slightly lowered IFN-Y release in SA-stimulated cells, whereas olanzapine left IFN-Y unchanged. IL-10 release decreased significantly with olanzapine 6 ng and 20 ng in PA and SA-stimulated cells, and quetiapine led to a slight reduction in IL-10 release. Conclusions: This in vitro study revealed how olanzapine and quetiapine affect CD4+ markers and cytokines. It emphasizes immune reduction via elevated CD25 and PD L-1 markers, known to decrease activated T-cells. Reduced immune response is evidenced by lower cytokine levels from activated T-cells, possibly increasing infection susceptibility, including pneumonia. Immune stimulation via CD28 and CD152 modulation might induce immunosuppression over time, exacerbating infection risk. Further studies are warranted to investigate the impact of antipsychotics on immune cells surface markers to better understand their immunomodulatory effects.6 0