Biology subjects

Mahdy, A. K. H.

Publications and source records attributed to Mahdy, A. K. H..

6 recordsLinked to original sources

bioRxiv2025

High throughput profiling of the B cell repertoire identifies systematic changes in the repertoire of individuals with Crohn's disease

The B cell repertoire contains the recombined DNA sequences that encodes the entire antibody repertoire of an individual. The repertoire is made from three antigenic binding chains, namely the immunoglobulin heavy chain (IGH) and two immunoglobulin light chains, {kappa} (IGK) and {lambda} (IGL). Compared to the T cell repertoire, the B cell repertoire is understudied in inflammatory bowel diseases (IBD) even though different antibodies such as ASCA and ANCA have been shown to be elevated in individuals with IBD. Furthermore, most IBD B cell repertoire studies have profiled the repertoire of treated individuals, thus capturing the combined effect of treatment and disease on the repertoire. To address this limitation, we profiled the repertoire of 24 treatment-naive individuals with CD with matching 24 symptomatic controls. The repertoire of individuals with CD showed a significant reduction in diversity and an increase in clonality, suggesting an antigen-driven expansion of clonotypes that might be driving the disease. Furthermore, we observed a significant reduction in the expansion of IgM and IgD and an expansion of IgA2 and IgG2 clonotypes in individuals with CD relative to controls. Lastly, we observed a reduction in the somatic hypermutation rate in the IGH J gene, particularly in IgM and IgA1 clonotypes, among individuals with CD relative to controls. Thus, despite the small sample size, we identified multiple alterations in the B cell repertoire of individuals with CD, highlighting the potential of the B cell repertoire in identifying antigenic exposures implicated in the diseases, demanding now larger international studies ideally including also treatment-naive and pre-clinical cases.

immunology↗

bioRxiv2025

Quantifying The Impact of Bulk TCR-Seq Methodological Choices on The Profiled T Cell Repertoire

Bulk T cell repertoire profiling using sequencing (TCR-Seq) is a powerful method to investigate T cell responses to natural infections, vaccines, cancers, and autoimmune diseases. This assay can be conducted using various techniques, such as multiplex PCR or 5-RACE. However, each methods introduces systematic biases that can result in different pictures of the underlying T cell repertoire. Furthermore, the impact of technical variables on the accuracy of these methods remains understudied. Thus, in this study, we systematically characterized different multiplex PCR-based protocols, focusing on the quality and quantity of the utilized RNA/DNA, extraction methods, amplification programs, variations between production batches, and technical handling of samples. Our findings highlight the important role of RNA/DNA quality in shaping the profiling results of T cell repertoires. Whereas low RNA/DNA quantities can be partially compensated for by increasing the number of PCR cycles, this is partially not possible with lower quality. In conclusion, our results highlight the influence of different technical choices on the biological conclusions drawn from TCR-Seq data and provide practical guidelines to finetune these variables to ensure consistent and reliable results under diverse experimental constraints.

immunology↗

bioRxiv2025

Analyzing the T cell receptor repertoire of 2,804 individuals with inflammatory bowel disease identifies public T cell responses involved in the pathogenesis

Whereas altered immune processes have been identified in individuals with inflammatory bowel disease (IBD), potentially causative antigens remain to be identified. By interrogating the immune repertoire of individuals with IBD, an identification of common antigenic exposures associated with the disease can be obtained. We analyzed the T cell receptor beta (TRB) chain repertoire of 1,890 individuals with Crohns disease (CD) and 914 individuals with ulcerative colitis (UC), enabling the identification of 327 TRB clonotypes associated with CD and 130 with UC. We validated the expansion of these clonotypes in a cohort of treatment-naive individuals with either CD, UC or symptomatic control (n=855). These disease-associated clonotypes were restricted to disease-associated risk HLA alleles and their expansion correlated with disease-severity but not with surgery or treatment trajectory. In conclusion, we identified and validated TRB clonotypes that are associated with either CD or UC, these clonotypes are a novel therapeutic target in IBD.

immunology↗

bioRxiv2025

Multi-centered T cell repertoire profiling identifies novel alterations in the immune repertoire of individuals with inflammatory bowel disease and validates previous findings

IntroductionIBD is an incurable immune-mediated inflammatory disease (IMID), affecting the gut with a high rate of primary- and secondary-loss-of-response to therapy. By investigating the T cell receptor repertoire of individuals with IBD, novel therapeutic and preventive strategies can be identified, and a better understanding of IBD can be obtained. MethodsWhereas most studies have so far focused on the more diverse T cell receptor beta (TRB) repertoire, we here profiled the alpha (TRA) repertoire of three cohorts containing treatment-naive and treated individuals in addition to individuals living with the disease for >20 years, resulting in an exhaustive dataset containing the TRA repertoire of 2,151 individuals. ResultsUsing the generated datasets, we were able to replicate previous findings describing the expansion of Crohns-associated invariant T (CAIT) cells in individuals with Crohns disease (CD) in the three cohorts. Using a hypothesis-free statistical testing framework, we identified clonotypes that were associated with the disease at its different stages, e.g., at the time of diagnosis and decades post-diagnosis. By conducting a meta-analysis across the three cohorts, we were able to identify a set of clonotypes that were associated with the disease regardless of its stage. We validated our findings in a previously published independent test dataset from a German cohort, showing the robustness of the identified sets of clonotypes. ConclusionThe identified clonotypes are potential novel therapeutic targets to treat IBD, e.g., through targeted depletion. These clonotypes are also of major interest as they can be investigated in a targeted fashion to identify culprit antigen(s) in IBD.

immunology↗

bioRxiv2025

Mapping adaptive immune responses toward fungal antigens in inflammatory bowel disease using T cell repertoire sequencing and phage-immunoprecipitation sequencing

Inflammatory bowel disease (IBD) is an idiopathic, immune-mediated chronic inflammatory disease of the gut with two primary clinical forms, Crohns disease (CD) and ulcerative colitis (UC). Several genetic susceptibility variants have been associated with IBD, such as ATG16L, NOD2, and several human leukocyte antigen (HLA) alleles, nonetheless, the actual disease causes remain unknown. Whereas previous findings have shown elevated responses toward fungal antigens in individuals with IBD, e.g., elevated anti-Saccharomyces cerevisiae antibody (ASCA) levels, an exhaustive mapping of immune responses toward fungal antigens remains incomplete. Thus, we analyzed the fungal mycobiome profiled using internal transcribed spacer 2 (ITS2)-sequencing simultaneously with the T cell repertoire of 637 individuals with IBD from the SPARC IBD cohort, which enabled us to identify 31 T cell clonotypes targeting several prevalent members of the gut mycobiome. Subsequently, we developed a novel phage-immunoprecipitation sequencing (PhIP-Seq) library covering 12,000 potential antigens from the proteome of S. cerevisiae and screened for antibody responses in 100 individuals with CD and 60 healthy controls with known ASCA status, enabling us to identify public and private antibody responses against several S. cerevisiae proteins. In conclusion, we corroborated previous findings showing elevated T cell responses against fungal antigens in individuals with IBD and identified multiple antigenic proteins from the proteome of S. cerevisiae that are targeted by the immune system of individuals with and without CD.

immunology↗

bioRxiv2024

Simultaneous Profiling of the Blood and Gut T and B Cell Repertoires in Crohn's Disease and Symptomatic Controls Illustrates Tissue-specific Alterations in the Immune Repertoire of Crohn's Disease Patients

Crohns disease (CD) is a clinical subset of inflammatory bowel disease that is characterized by patchy transmural inflammation across the gastrointestinal tract. Although the exact etiology remains unknown, recent findings suggest that it is a complex multifactorial disease with contributions from the host genetics and environmental factors such as the microbiome. We have shown that the T cell repertoire of CD patients harbors a group of highly expanded T cells which hints toward an antigen-mediated pathology. To profile the immunological signature of CD at a high-resolution we simultaneously profiled the {beta} and {gamma}{delta} T cell repertoire in addition to the B cell repertoire of both the blood and the colonic mucosa for 27 treatment-naive CD patients and 27 age-matched symptomatic controls. Regardless of disease, we observed multiple physiological differences between the immune repertoire of blood and colonic mucosa. Additionally, by comparing the repertoire of CD patients relative to controls, we observed different alterations that were only detected in the blood or colonic mucosa. These include a depletion of mucosal-associated invariant T (MAIT) cells in the blood repertoire, an expansion of TRAV29/DV5-TRAJ5+ clonotypes and a significant depletion of multiple IGHV3-33-IGHJ4+and IGHV3-33-IGHJ6+ clonotypes in the blood and gut IGH repertoire of CD patients. In conclusion, our findings highlight the importance of studying the immune repertoire in a tissue-specific manner and the need to profile the T and B cell immune repertoire of gut tissues.

immunology↗