Biology subjects

Khatri, R.

Publications and source records attributed to Khatri, R..

12 recordsLinked to original sources

bioRxiv2025

Spatial Analysis of T Cell Clonality in Autoimmune Kidney Disease Using TRV Probes

Hypothesizing that the localization of T cell clones correlates with immune function, our goal was to develop an unbiased method to study the spatial distribution of T cells in tissues. We created an in situ hybridization panel with 248 probes that identify immune and tissue cell types, and 132 probes for all variable TRAV, TRBV, TRGV, and TRDV gene segments. Applying this approach to analyze renal biopsies from patients with autoimmune kidney disease, combinations of TRV segments provided spatial information about T cell clonality. Confined clusters of clonally related {beta} T cells were found in proximity to increased numbers of antigen-presenting cells, B cells, and other T cells, reflecting local immune cell interactions. {gamma}{delta} T cells were more frequently located outside or at their periphery of T cell infiltration areas. In conclusion, integrating spatial information with TCR clonotype analysis provided new insights into the organization of immune responses at the tissue level.

immunology↗

bioRxiv2025

SCALE: Unsupervised Multi-Scale Domain Identification in Spatial Omics Data

Single-cell spatial transcriptomics enables precise mapping of cellular states and functional domains within their native tissue environment. These functional domains often exist at multiple spatial scales, with larger domains encompassing smaller ones, reflecting the hierarchical organization of biological systems. However, the identification of these functional domain hierarchies has been hardly explored due to the lack of appropriate computational methods. In this work, we present SCALE, an unsupervised algorithm for multi-scale domain identification in spatial transcriptomics data. SCALE combines neural graph representation learning with an entropy-based search algorithm to detect functional domains at different scales. It reaches state-of-the-art performance in single- and multi-scale domain detection on simulated and murine brain Xenium and MERFISH data, as well as patient-derived kidney tissue, highlighting its robustness and scalability across diverse tissue types and platforms. SCALEs ease of use makes it a powerful aid for advancing our understanding of tissue organization and function in health and disease.

bioinformatics↗

bioRxiv2024

Spatio-temporal interaction of immune and renal cells determines glomerular crescent formation in autoimmune kidney disease

Rapidly progressive glomerulonephritis (RPGN) is the most aggressive group of autoimmune kidney disease with the worst prognosis. Anti-neutrophil cytoplasmic antibody (ANCA) associated vasculitis, anti-glomerular basement membrane (anti-GBM) and lupus nephritis are the most common causes of RPGN and are characterized by the formation of glomerular crescents and infiltration of leukocytes that eventually lead to glomerulosclerosis and kidney failure. In this work, we used high-resolution spatial transcriptomics of 32 ANCA, 19 lupus nephritis, 6 anti-GBM, and 6 control patients to understand how intercellular signaling between immune and renal tissue cells leads to renal inflammation and glomerular injury. Using 3,218,210 immune and kidney cells, we observed that the biological pathways involved in the sequence of glomerular crescent formation are similar across the diseases. While innate immune cells infiltrated the glomerular compartment relatively early, later increases in adaptive immune cells were largely restricted to the periglomerular regions. These changes in immune cells temporally correlated with increases in glomerular parietal epithelial (PEC) and fibrotic mesangial cells, suggesting disease-relevant functional signaling between these immune and renal cells. Cell communication analysis revealed early disease PDGF signaling from epithelial and mesangial cells to PECs, causing their activation and proliferation. At later stages, TGF-{beta} signaling from macrophages, T cells, epithelial cells, and mesangial cells to PECs triggered the expression of extracellular matrix components resulting in glomerulosclerosis. Our results highlight a spatio-temporally conserved progression into glomerular crescents and sclerosis for ANCA, lupus nephritis, and anti-GBM disease, which is driven by consecutive PDGF and TGF-{beta} signaling to PECs.

immunology↗

bioRxiv2024

PCA-based spatial domain identification with state-of-the-art performance

The identification of biologically meaningful domains is a central step in the analysis of spatial transcriptomic data. Following Occams razor, we show that a simple PCA-based algorithm for spatial domain identification rivals the performance of ten competing state-of-the-art methods across six single-cell spatial transcriptomic datasets. Our reductionist approach, NichePCA, provides researchers with intuitive domain interpretation and excels in execution speed, robustness, and scalability.

bioinformatics↗

bioRxiv2023

A universal preservation protocol for multi-omic and histological analysis of kidney tissue

Biobanking of tissue from clinically obtained kidney biopsies for later use with multi-omic and imaging techniques is an inevitable step to overcome the need of disease model systems and towards translational medicine. Hence, collection protocols ensuring integration into daily clinical routines using preservation media not requiring liquid nitrogen but instantly preserving kidney tissue for clinical and scientific analyses are of paramount importance. Thus, we modified a robust single nucleus dissociation protocol for kidney tissue stored snap frozen or in the preservation media RNAlater and CellCover. Using porcine kidney tissue as surrogate for human kidney tissue, we conducted single nucleus RNA sequencing with the Chromium 10X Genomics platform. The resulting data sets from each storage condition were analyzed to identify any potential variations in transcriptomic profiles. Furthermore, we assessed the suitability of the preservation media for additional analysis techniques (proteomics, metabolomics) and the preservation of tissue architecture for histopathological examination including immunofluorescence staining. In this study, we show that in daily clinical routines the RNAlater facilitates the collection of highly preserved kidney biopsies and enables further analysis with cutting-edge techniques like single nucleus RNA sequencing, proteomics, and histopathological evaluation. Only metabolome analysis is currently restricted to snap frozen tissue. This work will contribute to build tissue biobanks with well-defined cohorts of the respective kidney disease that can be deeply molecularly characterized, opening new horizons for the identification of unique cells, pathways and biomarkers for the prevention, early identification, and targeted therapy of kidney diseases.

genomics↗

bioRxiv2023

Multiomic ALS signatures highlight sex differences and molecular subclusters and identify the MAPK pathway as therapeutic target

Amyotrophic lateral sclerosis (ALS) is the most common motor neuron disease and lacks effective disease-modifying treatments. Here, we performed a multiomic analysis of the prefrontal cortex of 51 patients with sporadic ALS and 50 control subjects, as well as four transgenic mouse models of C9orf72-, SOD1-, TDP-43-, and FUS-ALS to characterize early and sex-specific disease mechanisms in ALS. Integrated analyses of transcriptomes, (phospho)proteomes, and miRNAomes revealed more pronounced changes in males. We identified transcriptome-based human ALS subclusters driven by the immune response, ECM, mitochondrial respiration, and RNA metabolism. The molecular signatures of human subclusters were reflected in specific mouse models. Individual and integrative multiomics analysis highlighted the mitogen-activated protein kinase pathway as an early disease-relevant mechanism. Its modulation by trametinib in vitro and in vivo validated that mitogen-activated protein kinase kinase 2 is a promising therapeutic target with beneficial effects in females.

neuroscience↗

bioRxiv2023

Epigenetic neural glioblastoma enhances synaptic integration and predicts therapeutic vulnerability

Neural-tumor interactions drive glioma growth as evidenced in preclinical models, but clinical validation is nascent. We present an epigenetically defined neural signature of glioblastoma that independently affects patients survival. We use reference signatures of neural cells to deconvolve tumor DNA and classify samples into low- or high-neural tumors. High-neural glioblastomas exhibit hypomethylated CpG sites and upregulation of genes associated with synaptic integration. Single-cell transcriptomic analysis reveals high abundance of stem cell-like malignant cells classified as oligodendrocyte precursor and neural precursor cell-like in high-neural glioblastoma. High-neural glioblastoma cells engender neuron-to-glioma synapse formation in vitro and in vivo and show an unfavorable survival after xenografting. In patients, a high-neural signature associates with decreased survival as well as increased functional connectivity and can be detected via DNA analytes and brain-derived neurotrophic factor in plasma. Our study presents an epigenetically defined malignant neural signature in high-grade gliomas that is prognostically relevant.

neuroscience↗

bioRxiv2022

Genomic screening reveals UBA1 as a potent and druggable target in c-MYC-high TNBC models

Triple negative breast cancer (TNBC) accounts for over 30% of all breast cancer-related deaths, despite accounting for only 10%-15% of total breast cancer cases. Targeted therapy development has largely stalled in TNBC, underlined by a lack of traditionally druggable addictions like receptor tyrosine kinases (RTKs). Here, through full genome CRISPR/Cas9 screening of TNBC models, we have uncovered the sensitivity of TNBCs to the depletion of the Ubiquitin-Like Modifier Activating Enzyme 1 (UBA1). Targeting UBA1 with the first in-class UBA1 inhibitor TAK-243 induced unresolvable ER-stress and activating transcription factor 4 (ATF4)-mediated upregulation of pro-apoptotic NOXA, leading to cell death. In five patient derived xenograft models (PDXs) of TNBC, TAK-243 therapy led to tumor inhibition or frank tumor regression. In an intracardiac metastatic model of TNBC, TAK-243 markedly reduced metastatic burden. Importantly, there was an order of magnitude greater sensitivity of TNBC lines to TAK-243 compared to normal tissue-derived cells. Lastly, c-MYC expression correlates with TAK-243 sensitivity and cooperates with TAK-243 to induce a stress response and cell death. We posit UBA1 is an important new target in TNBC expressing high levels of c-MYC. SignificanceGenomic screening of TNBC cell lines revealed broad sensitivity to depletion of the E1 ubiquitin enzyme, UBA1. Disrupting UBA1 with the first in-class inhibitor TAK-243 in TNBC models induces ER-stress through an ATF4-NOXA axis that is dependent on c-MYC, leading to apoptosis, in vitro and in vivo, primary tumor growth inhibition and metastatic inhibition.

cancer biology↗

bioRxiv2022

SARS-CoV-2 and its variants, but not Omicron, induces thymic atrophy and impaired T cell development

Pathogenic infections cause thymic atrophy, perturb thymic-T cell development and alter immunological response. Previous studies reported dysregulated T cell function and lymphopenia in coronavirus disease-19 (COVID-19) patients. However, immune-pathological changes, in the thymus, post severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection have not been elucidated. Here, we report SARS-CoV-2 infects thymocytes, depletes CD4+CD8+ (double positive; DP) T cell population associated with an increased apoptosis of thymocytes, which leads to severe thymic atrophy in K18-hACE2-Tg mice. CD44+CD25-T cells were found to be enriched in infected thymus, indicating an early arrest in the T cell developmental pathway. Further, Interferon gamma (IFN-{gamma}) was crucial for thymic atrophy, as anti-IFN-{gamma} antibody neutralization rescued the loss of thymic involution. Therapeutic use of remdesivir (prototype anti-viral drug) was also able to rescue thymic atrophy. While Omicron variant of SARS-CoV2 caused marginal thymic atrophy, delta variant of SARS-CoV-2 exhibited most profound thymic atrophy characterized by severely depleted DP T cells. Recently characterized broadly SARS-CoV-2 neutralizing monoclonal antibody P4A2 was able to rescue thymic atrophy and restore thymic developmental pathway of T cells. Together, we provide the first report of SARS-CoV-2 associated thymic atrophy resulting from impaired T cell developmental pathway and also explains dysregulated T cell function in COVID-19.

immunology↗

bioRxiv2022

Immunopharmacological evaluation of adjuvant efficacy of Monophosphoryl lipid-A and CpG ODN with SARS-CoV-2 RBD antigen

SARS-CoV-2 infection has made the mankind to witness most sever and serious pandemic situation in the history. Millions of people have suffered and are still suffering with this infection which has caused a mass mortality in the past three years. Development of an effective vaccine to control the spread of infection and to prevent this viral infection is need of the hour. Adjuvanted vaccines have proven their efficacy in controlling many other viral infections like flu, keeping this context in view we have evaluated the immunopharmacological efficacy of two adjuvants MPL-A and CpG ODN in combination with MF59 emulsion against SARS-CoV-2 antigen. From the data obtained we can infer that both the adjuvants were capable of eliciting a potent antibody response against antigen alone and MF59 groups. Comparatively MPL-A was eliciting a Th1 polarized response in terms of IgG2a and cytokine production. Both the adjuvants were capable of enhancing the CD 4, 8 and 19 cell populations. Overall the pre clinical evaluation has given a clue of the effectiveness of MPL-A and CpG adjuvants against SARS-CoV-2 antigen.

immunology↗

bioRxiv2022

DiSCERN - Deep Single Cell Expression ReconstructioN for improved cell clustering and cell subtype and state detection

Single cell sequencing provides detailed insights into biological processes including cell differentiation and identity. While providing deep cell-specific information, the method suffers from technical constraints, most notably a limited number of expressed genes per cell, which leads to suboptimal clustering and cell type identification. Here we present DISCERN, a novel deep generative network that reconstructs missing single cell gene expression using a reference dataset. DISCERN outperforms competing algorithms in expression inference resulting in greatly improved cell clustering, cell type and activity detection, and insights into the cellular regulation of disease. We used DISCERN to detect two unseen COVID-19-associated T cell types, cytotoxic CD4+ and CD8+ Tc2 T helper cells, with a potential role in adverse disease outcome. We utilized T cell fraction information of patient blood to classify mild or severe COVID-19 with an AUROC of 81% that can serve as a biomarker of disease stage. DISCERN can be easily integrated into existing single cell sequencing workflows and readily adapted to enhance various other biomedical data types.

bioinformatics↗

bioRxiv2018

Increased association between Epstein-Barr virus EBNA2 from type 2 strains and the transcriptional repressor BS69 restricts B cell growth

Natural variation separates Epstein-Barr virus (EBV) into type 1 and type 2 strains. Type 2 EBV is less transforming in vitro due to sequence differences in the EBV transcription factor EBNA2. This correlates with reduced activation of the EBV oncogene LMP1 and some cell genes. Transcriptional activation by type 1 EBNA2 can be suppressed through the binding of two PXLXP motifs in its transactivation domain (TAD) to the dimeric coiled-coil MYND domain (CC-MYND) of the BS69 repressor protein (ZMYND11). We identified a third conserved PXLXP motif in type 2 EBNA2. We found that type 2 EBNA2 peptides containing this motif bound BS69CC-MYND efficiently and that the type 2 EBNA2TAD bound an additional BS69CC-MYND molecule. Full-length type 2 EBNA2 also bound BS69 more efficiently in pull-down assays. Molecular weight analysis and low-resolution structures obtained using small-angle X-ray scattering showed that three BS69CC-MYND dimers bound two molecules of type 2 EBNA2TAD, in line with the dimeric state of full-length EBNA2 in vivo. Importantly, mutation of the third BS69 binding motif in type 2 EBNA2 improved B-cell growth maintenance. Our data indicate that increased association with BS69 restricts growth promotion by EBNA2 and may contribute to reduced B-cell transformation by type 2 EBV.\n\nAuthor summaryEpstein-Barr virus (EBV) drives the development of many human cancers worldwide including specific types of lymphoma and carcinoma. EBV infects B lymphocytes and immortalises them, thus contributing to lymphoma development. The virus promotes B lymphocyte growth and survival by altering the level at which hundreds of genes are expressed. The EBV protein EBNA2 is known to activate many growth-promoting genes. Natural variation in the sequence of EBNA2 defines the two main EBV strains: type 1 and type 2. Type 2 strains immortalise B lymphocytes less efficiency and activate some growth genes poorly, although the mechanism of this difference is unclear. We now show that sequence variation in type 2 EBNA2 creates a third site of interaction for the repressor protein (BS69, ZMYND11). We have characterised the complex formed between type 2 EBNA2 and BS69 and show that three dimers of BS69 form a bridged complex with two molecules of type 2 EBNA2. We demonstrate that mutation of the additional BS69 interaction site in type 2 EBNA2 improves its growth-promoting function. Our results therefore provide a molecular explanation for the different B lymphocyte growth promoting activities of type 1 and type 2 EBV. This aids our understanding of immortalisation by EBV.

microbiology↗