Biology subjects

Brusko, T.

Publications and source records attributed to Brusko, T..

6 recordsLinked to original sources

bioRxiv2026

Generation of hypoimmunogenic gastric insulin-secreting organoids

Gastric insulin-secreting organoids (GINS) represent a promising source of {beta}-like cells for type 1 diabetes (T1D) therapy. In same-donor comparisons with induced pluripotent stem cell-derived islets (iPSC-islets), GINS displayed robust glucose responsiveness and reduced expression of key T1D autoantigens. Importantly, GINS exhibited decreased susceptibility to cytotoxicity mediated by engineered HLA-matched preproinsulin-specific effector T cells (Avatar Teffs) and a distinct transcriptional profile enriched for immune-modulatory and stress-adaptive gene programs. To enhance immune evasion, we engineered gastric stem cells to overexpress Programmed Death Ligand 1 (PD-L1) in an inducible manner. PD-L1+ GINS maintained normal functionality, while exhibiting improved survival under allogeneic Avatar Teff challenge in a MHC class I-independent fashion. We evaluated PD-L1-mediated protection against autologous Avatar Teff attack using an endothelialized microfluidic platform recapitulating physiologic immune interactions. T cells show reduced infiltration into PD-L1 GINS, resulting in significantly higher organoid viability compared to control GINS. Together, these findings identify GINS as a functional and engineerable {beta}-like cell platform with intrinsic hypoimmunogenic features, and support PD-L1 engineering as a strategy to enhance immune protection for both allogeneic and autologous transplantation in T1D.

immunology↗

bioRxiv2025

BayesCNet: Bayesian inference for cell type-specific regulatory networks leveraging cell type hierarchy in single-cell data

Understanding gene regulatory networks (GRNs) is essential for deciphering biological processes and disease mechanisms. Single-cell multiome technologies now enable joint profiling of chromatin accessibility and gene expression, offering an powerful means to infer cell type-specific GRNs. However, existing methods analyze each cell type independently or aggregate data into pseudo-bulk profiles, limiting their ability to resolve rare populations and capture cellular heterogeneity. We introduce BayesCNet, a Bayesian hierarchical model that jointly infers enhancer-gene linkages across all cell types while leveraging their hierarchical relationships for information sharing. Through extensive simulations, BayesCNet consistently outperforms state-of-the-art methods, with the largest improvements in rare cell types. When applied to real datasets, BayesCNet identifies enhancer-gene linkages with higher accuracy validated by promoter-capture Hi-C data, and reconstructs cell type-specific GRNs that highlight key regulators, demonstrating its power to resolve gene regulatory programs across diverse cell types.

bioinformatics↗

bioRxiv2024

Inhibition of CD226 Co-Stimulation Suppresses Diabetes Development in the NOD Mouse by Augmenting Tregs and Diminishing Effector T Cell Function

Aims/hypothesisImmunotherapeutics targeting T cells are crucial for inhibiting autoimmune disease progression proximal to disease onset in type 1 diabetes. A growing number of T cell-directed therapeutics have demonstrated partial therapeutic efficacy, with anti-CD3 (-CD3) representing the only regulatory agency-approved drug capable of slowing disease progression through a mechanism involving the induction of partial T cell exhaustion. There is an outstanding need to augment the durability and effectiveness of T cell targeting by directly restraining proinflammatory T helper type 1 (Th1) and type 1 cytotoxic CD8+ T cell (Tc1) subsets, while simultaneously augmenting regulatory T cell (Treg) activity. Here, we present a novel strategy for reducing diabetes incidence in the NOD mouse model using a blocking monoclonal antibody targeting the type 1 diabetes-risk associated T cell co-stimulatory receptor, CD226. MethodsFemale NOD mice were treated with anti-CD226 between 7-8 weeks of age and then monitored for diabetes incidence and therapeutic mechanism of action. ResultsCompared to isotype-treated controls, anti-CD226 treated NOD mice showed reduced insulitis severity at 12 weeks and decreased disease incidence at 30 weeks. Flow cytometric analysis performed five weeks post-treatment demonstrated reduced proliferation of CD4+ and CD8+ effector memory T cells in spleens of anti-CD226 treated mice. Phenotyping of pancreatic Tregs revealed increased CD25 expression and STAT5 phosphorylation following anti-CD226, with splenic Tregs displaying augmented suppression of CD4+ T cell responders in vitro. Anti-CD226 treated mice exhibited reduced frequencies of islet-specific glucose-6-phosphatase catalytic subunit related protein (IGRP)-reactive CD8+ T cells in the pancreas, using both ex vivo tetramer staining and single-cell T cell receptor sequencing (scTCR-seq) approaches. 51Cr-release assays demonstrated reduced cell-mediated lysis of beta-cells by anti-CD226-treated autoreactive cytotoxic T lymphocytes. Conclusions/interpretationCD226 blockade reduces T cell cytotoxicity and improves Treg function, representing a targeted and rational approach for restoring immune regulation in type 1 diabetes. Research in ContextO_ST_ABSWhat is already known about this subject?C_ST_ABSO_LIThe co-stimulatory receptor CD226 is upregulated upon activation and is highly expressed on NK cell subsets, myeloid cells, and effector T cells. C_LIO_LIA single nucleotide polymorphism in CD226 (rs763361; C>T) results in a Gly307Ser missense mutation linked to genetic susceptibility for type 1 diabetes. C_LIO_LIGlobal knockout of Cd226 and conditional Cd226 knockout in FoxP3+ Tregs reduced insulitis severity and diabetes incidence in NOD mice, indicating a crucial role for CD226 in disease pathogenesis. C_LI What is the key question?O_LICan CD226 blockade reduce T cell cytotoxicity and improve Treg function to diminish diabetes incidence in NOD mice? C_LI What are the new findings?O_LIAnti-CD226 treatment reduced insulitis, decreased disease incidence, and inhibited splenic CD4+ and CD8+ effector memory T cell proliferation. C_LIO_LIPancreatic Tregs from anti-CD226 treated mice exhibited increased CD25 expression; splenic Tregs displayed augmented STAT5 phosphorylation and suppressive capacity in vitro. C_LIO_LIAnti-CD226 treatment reduced IGRP-specific pancreatic CD8+ T cell frequencies, and reduced autoreactive CD8+ T cell-mediated lysis of beta-cells in vitro. C_LI How might this impact on clinical practice in the foreseeable future?O_LICD226 blockade could reduce autoreactive T cell cytotoxicity, enhance Treg function, and slow disease progression in high-risk or recent-onset type 1 diabetes cases. C_LI

immunology↗

bioRxiv2022

Deletion of CD226 in Foxp3+ T cells Reduces Diabetes Incidence in Non-Obese Diabetic Mice by Improving Regulatory T Cell Stability and Function

Co-stimulation serves as a critical checkpoint for T cell development and activation, and several genetic variants affecting co-stimulatory pathways confer risk for autoimmune diseases. A single nucleotide polymorphism in CD226 (rs763361; G307S) has been shown to increase susceptibility to type 1 diabetes, multiple sclerosis, and rheumatoid arthritis. CD226 competes with the co-inhibitory receptor TIGIT (T cell immunoreceptor with Ig and ITIM domains) to bind CD155 to amplify TCR signaling. We previously found that Cd226 knockout protected non-obese diabetic (NOD) mice from disease, but the impact of CD226 signaling on individual immune subsets remained unclear. We focused on regulatory T cells (Tregs) as a population of interest, as prior reports demonstrated that human CD226+ Tregs exhibit reduced FOXP3+Helios+ purity and suppressive function following expansion. Hence, we hypothesized that global deletion of Cd226 would increase Treg stability and accordingly, Treg-specific Cd226 deletion would inhibit diabetes in NOD mice. Indeed, crossing the NOD.Cd226-/- and NOD.Foxp3-GFP-Cre.R26-loxP-STOP-loxP-YFP Treg-fate tracking strains resulted in increased Treg induction and decreased FoxP3-deficient "ex-Tregs" in the pancreatic lymph nodes. We generated a Treg-conditional knockout (Treg{Delta}Cd226) strain and found that female Treg{Delta}Cd226 mice had decreased insulitis and diabetes incidence compared to TregWT mice. Additionally, we observed increased TIGIT expression on Tregs and conventional CD4+ T cells within the pancreas of Treg{Delta}Cd226 versus TregWT mice. These findings demonstrate that an imbalance of CD226/TIGIT signaling may contribute to Treg destabilization in the NOD mouse and highlight the potential for therapeutic targeting of this pathway to prevent or reverse autoimmunity.

immunology↗

bioRxiv2021

3D-mapping of human lymph node and spleen reveals integrated neuronal, vascular, and ductal cell networks

The spleen and lymph node represent important hubs for both innate and adaptive immunity1,2. Herein, we map immune, endothelial, and neuronal cell networks within these tissues from "normal"/non-diseased organ donors, collected through the NIH Human BioMolecular Atlas Program (HuBMAP)3, using highly multiplexed CODEX (CO-Detection by indEXing) imaging and 3D light sheet microscopy of cleared tissues. Building on prior reports4-6, we observed the lymph node subcapsular sinus expressing podoplanin, smooth muscle actin, and LYVE1. In the spleen, LYVE1 was expressed by littoral cells lining venous sinusoids, whereas podoplanin was restricted to arteries and trabeculae. 3D visualization of perivascular innervation revealed a subset of axonal processes expressing choline acetyl transferase in both tissues, in contrast with prior literature on human spleen7. We further report our novel observations regarding the distinct localization of GAP43 and {beta}3-tubulin within the vascular anatomy of both lymph node and spleen, with Coronin-1A+ cells forming a dense cluster around {beta}3-tubulin positive GAP43 low/negative segments of large vessels in spleen. These data provide an unprecedented 2D and 3D visualization of cellular networks within secondary lymphoid tissues, laying the groundwork for future disease-specific and system-wide studies of neural regulation of immunity in human lymphatics.

immunology↗

bioRxiv2021

geneBasis: an iterative approach for unsupervised selection of targeted gene panels from scRNA-seq.

The problem of selecting targeted gene panels that capture maximum variability encoded in scRNA-sequencing data has become of great practical importance. scRNA-seq datasets are increasingly being used to identify gene panels that can be probed using alternative molecular technologies, such as spatial transcriptomics. In this context, the number of genes that can be probed is an important limiting factor, so choosing the best subset of genes is vital. Existing methods for this task are limited by either a reliance on pre-existing cell type labels or by difficulties in identifying markers of rare cell types. We resolve this by introducing an iterative approach, geneBasis, for selecting an optimal gene panel, where each newly added gene captures the maximum distance between the true manifold and the manifold constructed using the currently selected gene panel. We demonstrate, using a variety of metrics and diverse datasets, that our approach outperforms existing strategies, and can not only resolve cell types but also more subtle cell state differences. Our approach is available as an open source, easy-to-use, documented R package (https://github.com/MarioniLab/geneBasisR).

bioinformatics↗