Biology subjects

Gallon, L.

Publications and source records attributed to Gallon, L..

5 recordsLinked to original sources

bioRxiv2025

Cellular and Molecular Resolution of Focal Segmental Glomerulosclerosis Recurrence in Human Allografts

Primary Focal Segmental Glomerulosclerosis (FSGS) is an important cause of end-stage renal disease (ESRD). Primary FSGS recurrence rates in transplanted kidneys are high, with 25-50% in first transplants and up to 80% in second transplants, often leading to graft loss. To investigate the molecular and cellular events underlying recurrent primary FSGS (reFSGS), we performed single-nucleus RNA sequencing (snRNA-seq) on kidney transplant biopsies from patients with reFSGS and controls with normal allograft function. Our analysis revealed that podocyte loss in reFSGS is driven by metabolic and structural dysregulation rather than apoptosis. Overexpression of vascular endothelial growth factor (VEGF)-A by podocytes was observed, potentially disrupting glomerular endothelial cell growth and permeability. Parietal epithelial cells (PECs) exhibited dedifferentiation towards a podocyte-like state, potentially compensating for podocyte loss, but this was associated with increased collagen deposition and glomerular sclerosis. Ligand-receptor interactions between glomerular cells and B cells further promoted extracellular matrix deposition and fibrosis. Additionally, tubular cells demonstrated evidence of tubular sclerosis and impaired regenerative potential, accompanied by increased interactions with T cells. These findings provide novel insights into the pathogenesis of reFSGS and identify potential therapeutic targets. This study establishes a foundation for future research to further investigate cell-type-specific interventions in recurrent FSGS.

molecular biology↗

bioRxiv2024

Evaluation of Methodologies in Anti-nephrin Autoantibody Detection

Recent studies discovered the prominent presence of anti-nephrin autoantibodies in minimal change disease, steroid-sensitive nephrotic syndrome and/or post-transplant recurrent focal segmental glomerulosclerosis (FSGS). However, widely different, and often unconventional autoantibody detection methods were used among these studies, making it challenging to assess the pathogenic role for the antibodies. Here we examined methods of conventional ELISA, magnetic on-beads ELISA, immunoprecipitation-immunoblotting (IP-IB), and cell- and tissue-based antibody assays with 127 plasma samples of kidney and non-kidney diseases. On the antigen side, we compared commercially available recombinant human nephrin extracelluar domain (ECD) produced from human or mouse cell lines, as well as lab-made full length, ECD, and series of ECD truncates for measuring autoantibody reactivity and specificity. Surprisingly, different assay methods and different antigen preparations led to observation of assay-specific false-positive and false-negative results. In general, a set of tests that combines magnetic beads-enhanced ELISA, followed by IP-IB, and epitope mapping showed the most robust results for anti-nephrin autoantibodies, detected in two primary FSGS patients among all cases tested. It is interesting to note that cell/tissue-based results, also supported by antigen truncation studies, clearly suggest steric hindrance of reactive epitopes, as in full length nephrin that forms compact self-associated complexes. In conclusion, anti-nephrin positivity is rare among the tested patients (2/127), including those with FSGS (2/42), and autoantibody results can be affected by the choice of detection methods.

biochemistry↗

bioRxiv2024

Genetic polymorphisms of Leukocyte Immunoglobulin-Like Receptor B3 (LILRB3) gene in African American kidney transplant recipients are associated with post-transplant graft failure

BackgroundAfrican American (AA) kidney transplant recipients exhibit a higher rate of graft loss compared to other racial and ethnic populations, highlighting the need to identify causative factors underlying this disparity. MethodWe analyzed RNA sequences of pretransplant whole blood from subjects followed in three kidney transplant cohorts to identify single nucleotide polymorphisms (SNPs) associated with death censored graft loss (DCGL). We employed a meta-analysis to uncover key transcriptional signatures and pathways associated with the identified SNPs and used single cell RNA to define cellular specificity. We characterized SNP functions using in vitro immunological and survival assays and tested for associations between the identified SNPs and other immune-related diseases using a [~]30,100 subject, electronic health record (EHR)-linked database. ResultsWe uncovered a cluster of four consecutive missense SNPs in the Leukocyte Immunoglobulin-Like Receptor B3 (LILRB3, a negative immune response regulator) gene that strongly associated with DCGL. This LILRB3-4SNPs cluster encodes missense mutations at amino acids 617-618 proximal to a SHP-1/2 phosphatase-binding ITIM motif. LILRB3-4SNPs is specifically enriched within subjects of AA ancestry (8.6% prevalence vs 2.3% in Hispanic and 0.1% in European populations), is not linked to APOL1 G1/G2 alleles, and exhibited a strong association with DCGL. Analysis of PBMC and transplant biopsies from recipients with LILRB3-4SNPs showed evidence of enhanced adaptive immune responsiveness and ferroptosis-associated death in monocytes. Overexpression of the variant allele in THP-1 cells (macrophage line) induced augmented inflammation and ferroptosis, which were attenuated by a ferroptosis inhibitor, verifying a causal link. The LILRB3-4SNPs also associated with multiple systemic and organ-specific immune-related diseases in AAs, consistent with conferring a broadly relevant immune function. Conclusionthe LILRB3-4SNPs represent a functionally important, distinct genetic risk factor for kidney transplant outcome and development/severity of other immune-related diseases in patients of AA ancestry. Pharmacological targeting of ferroptosis should be tested to prevent or treat these disease processes in AA recipients carrying LILRB3-4SNPs.

genetics↗

bioRxiv2023

Deep-learning Based Pathological Assessment of Frozen Procurement Kidney Biopsies Predicts Graft Loss and Guides Organ Utilization: A Large-scale Retrospective Study

BackgroundLesion scores on procurement donor biopsies are commonly used to guide organ utilization. However, frozen sections present challenges for histological scoring, leading to inter- and intra-observer variability and inappropriate discard. MethodsWe constructed deep-learning based models to recognize kidney tissue compartments in H&E stained sections from procurement biopsies performed at 583 hospitals nationwide in year 2011-2020. The models were trained and tested respectively on 11473 and 3986 images sliced from 100 slides. We then extracted whole-slide abnormality features from 2431 kidneys, and correlated with pathologists scores and transplant outcomes. Finally, a Kidney Donor Quality Score (KDQS) incorporating digital features and the Kidney Donor Profile Index (KDPI) was derived and used in combination with recipient demographic and peri-transplant characteristics to predict graft loss or assist organ utilization. ResultsOur model accurately identified 96% and 91% of normal/sclerotic glomeruli respectively; 94% of arteries/arterial intimal fibrosis regions; 90% of tubules. Three whole-slide features (Sclerotic Glomeruli%, Arterial Intimal Fibrosis%, and Interstitial Fibrosis%) demonstrated strong correlations with corresponding pathologists scores (n=2431), but had superior associations with post-transplant eGFR (n=2033) and graft loss (n=1560). The combination of KDQS and other factors predicted 1- and 4-year graft loss (discovery: n=520, validation: n=1040). Finally, by matching 398 discarded kidneys due to "biopsy findings" to transplanted population, the matched transplants from discarded KDQS<4 group (110/398, 27.6%) showed similar graft survival rate to unmatched transplanted kidneys (2-, 5-year survival rate: 97%, 86%). KDQS [≥] 7 (37/398, 9.3%) and 1-year survival model score [≥] 0.55 were determined to identify possible discards (PPV=0.92). ConclusionThis deep-learning based approach provides automatic and reliable pathological assessment of procurement kidney biopsies, which could facilitate graft loss risk stratification and organ utilization. Translational StatementThis deep-learning based approach provides rapid but more objective, sensitive and reliable assessment of deceased-donor kidneys before transplantation, and improves the prognostic value of procurement biopsies, thus could potentially reduce inappropriate discard and stratify patients needing monitoring or preventative measures after transplantation. The pipeline can be integrated into various types of scanners and conveniently generates report after slide scanning. Such report can be used in conjunction with pathologists report or independently for centers lacking renal pathologists.

bioinformatics↗

bioRxiv2022

Multiscale genetic architecture of donor-recipient differences reveals intronic LIMS1 locus mismatches associated with long-term renal transplant survival

BackgroundLong-term kidney allograft survival remains suboptimal. Emerging evidence indicates donor-recipient (D-R) mismatches outside of human leukocyte antigens (HLA) contribute to graft survival but mechanisms remain unclear, specifically for those mismatches within intronic regions. MethodsWe analyzed genome-wide SNP data of D-R pairs from two well-phenotyped kidney transplant cohorts (median follow-up ~1800 days), Genomics of Chronic Allograft Rejection (GoCAR; n=385) and Clinical Trials in Organ Transplantation 1/17 (CTOT1/17; n=146), quantifying genetic mismatches outside of HLA for every D-R pair at variant, gene, and genome-wide scales. ResultsUnbiased genome-wide screen of GoCAR D-R pairs uncovered the LIMS1 locus as the topranked candidate where D-R mismatches associated with death censored graft loss (DCGL). Independent of HLA, a previously unreported relationship between mismatches at a highly linked, intronic haplotype of 30 SNPs was seen as associated with DCGL, with confirmatory association in intra-ancestry D-Rs. Validation testing within the CTOT-01/17 showed similar associations with DCGL. Haplotype D-R mismatches showed a dosage effect, and the introduction of minor alleles in the donor to major allele-carrying recipients showed a greater risk of DCGL. Both the new LIMS1 haplotype and the previously reported LIMS1 SNP rs893403 are expression quantitative trait loci (eQTL) for the gene GCC2 in recipient immune cells, without alterations in GCC2 or LIMS1 coding sequences. Transcriptome enrichment analyses performed on whole blood and within multiple T cell subsets demonstrated significant associations of GCC2 gene, and of either allelic locus, with regulation of TGF-beta-SMAD signaling, implying a role in Treg function and association with rejection. ConclusionsOur analysis unravels intronic non-HLA SNP mismatches within LIMS1 that do not induce protein sequence variation but associate with DCGL. By acting as cis-eQTLs for GCC2 expression, these SNPs modulates TGF-beta signaling and T cell function, associating with immune events and graft outcomes. The findings have clinical implications for risk assessment and individualized therapy in kidney transplant recipients.

genetics↗