Biology subjects

Braciale, T. J.

Publications and source records attributed to Braciale, T. J..

5 recordsLinked to original sources

bioRxiv2025

Initiation of primary T cell--B cell interactions and extrafollicular antibody responses in an organized microphysiological model of the human lymph node

In vitro microphysiological systems (MPS) are needed to replicate events in the lymph node (LN) leading to humoral immunity against new immune threats, but current lymphoid MPS focus largely on recall responses from memory lymphocytes. Here, an LN MPS was developed from primary, naive human lymphocytes in microfluidic 3D culture to model interactions and antibody production at the LN T cell--B cell border. Naive CD4+ T cells exhibited CCL21-dependent chemotaxis, chemokinesis, and activation in the MPS, and were skewed to a T follicular helper (pre-Tfh) phenotype. IgM secretion was induced in co-culture with activated B cells in the presence of a superantigen, staphylococcal enterotoxin B (SEB); micropatterning confirmed that the interaction required physical proximity. SEB-dependence of IgM secretion was greatest at a 1:5 T:B ratio, while seeding more pre-Tfh cells accelerated plasmablast differentiation and clustering. On-chip co-cultures at a 1:5 T:B ratio developed large lymphoid clusters containing CD38+ plasmablasts and CD138+ plasma cells after 15 days, with response varying between donors. Significant plasmablast induction in T-B co-cultures did not require the pre-Tfh phenotype, but pre-Tfh cells were required for inducing IgM secretion. We envision that this LN MPS will enable predictions and mechanistic analyses of human humoral immunity in vitro.

bioengineering↗

bioRxiv2024

Engineered human lymph node stroma model for examining interstitial fluid flow and T cell egress

The lymph node (LN) performs essential roles in immunosurveillance throughout the body. Developing in vitro models of this key tissue is of great importance to enhancing physiological relevance in immunoengineering. The LN consists of stromal populations and immune cells, which are highly organized and bathed in constant interstitial flow. The stroma, notably the fibroblastic reticular cells (FRCs) and the lymphatic endothelial cells (LECs), play crucial roles in guiding T cell migration and are known to be sensitive to fluid flow. During inflammation, interstitial fluid flow rates drastically increase in the LN. It is unknown how these altered flow rates impact crosstalk and cell behavior in the LN, and most existing in vitro models focus on the interactions between T cells, B cells, and dendritic cells rather than with the stroma. To address this gap, we developed a human engineered model of the LN stroma consisting of FRC-laden hydrogel above a monolayer of LECs in a tissue culture insert with gravity-driven interstitial flow. We found that FRCs had enhanced coverage and proliferation in response to high flow rates, while LECs experienced decreased barrier integrity. We added CD4+ and CD8+ T cells and found that their egress was significantly decreased in the presence of interstitial flow, regardless of magnitude. Interestingly, 3.0 {micro}m/s flow, but not 0.8 {micro}m/s flow, correlated with enhanced inflammatory cytokine secretion in the LN stroma. Overall, we demonstrate that interstitial flow is an essential consideration in the lymph node for modulating LN stroma morphology, T cell migration, and inflammation.

bioengineering↗

bioRxiv2024

Mitigating reactive oxygen species production and increasing gel porosity improves lymphocyte motility and fibroblast spreading in photocrosslinked gelatin-thiol hydrogels

On-chip 3D culture systems that incorporate immune cells such as lymphocytes and stromal cells are needed to model immune organs in engineered systems such as organs-on-chip. Photocrosslinking is a useful tool for creating such immune-competent hydrogel cultures with spatial cell organization. However, loss of viability and motility in photocrosslinked gels can limit its utility, especially when working with fragile primary cells. We hypothesized that optimizing photoexposure-induced ROS production, hydrogel porosity or a combination of both factors was necessary to sustain cell viability and motility during culture in photocrosslinked gelatin-thiol (GelSH) hydrogels. Jurkat T cells, primary human CD4+ T cells and human lymphatic fibroblasts were selected as representative lymphoid immune cells to test this hypothesis. Direct exposure of these cells to 385 nm light and LAP photoinitiator dramatically increased ROS levels. Pretreatment with an antioxidant, ascorbic acid (AA), protected the cells from light + LAP-induced ROS and was non-toxic at optimized doses. Furthermore, scanning electron microscopy showed that native GelSH hydrogels had limited porosity, and that adding collagen to GelSH precursor before crosslinking markedly increased gel porosity. Next, we tested the impact of AA pretreatment and increasing gel porosity, alone or in combination, on cell viability and function in 3D GelSH hydrogel cultures. Increasing gel porosity, rather than AA pretreatment, was more critical for rescuing viability of Jurkat T cells and spreading of human lymphatic fibroblasts in GelSH-based gels, but both factors improved the motility of primary human CD4+ T cells. Increased porosity enabled formation of spatially organized co-cultures of primary human CD4+ T cells and human lymphatic fibroblasts in photo-crosslinked gels in a multi-lane microfluidic chip, towards modeling the lymphoid organ microenvironment. Some optimization is still needed to improve homogeneity between regions on the chip. These findings will enable researchers utilizing photocrosslinking methods to develop immunocompetent 3D culture models that support viability and function of sensitive lymphoid cells.

bioengineering↗

bioRxiv2022

Single cell RNA sequencing reveals mechanisms underlying a senescence-like phenotype of Alveolar Macrophages during Aging

Alveolar Macrophages (AMs) are unique innate immune cells that reside in the alveolar space and accommodate the ever-changing needs of the lungs against internal and external challenges. During homeostasis, AMs maintain themselves through self-renewal without input from adult hematopoietic stem cells. Currently, little is known regarding the influence of aging on AM dynamics, heterogeneity and transcriptional profiles. Here, we identified CBF{beta} as an indispensable transcription factor that ensures AM self-renewal. Deficiency in CBF{beta} led to decreased proliferation and self-renewal ability of AMs. Moreover, with single cell RNA sequencing analysis of AMs from young and aged mice, we discovered that despite similarities in the transcriptome of proliferating cells, AMs from the aged mice exhibited reduced embryotic stem cell-like features. Aged AMs also showed diminished capacity for DNA repair, potentially contributing to impaired cell cycle progression and elevation of senescence markers. In accordance with the analysis, we observed reduced number of AMs in aged mice, which had defective self-renewal ability and were more sensitive to the reduction of GM-CSF. Interestingly, decreased CBF{beta} was observed in the cytosol of AMs from aged mice. A similar senescence-like phenotype was also found in human AMs. Taken together, we conclude that AMs in the aged host harbor a senescence-like phenotype, potentially mediated by the activity of CBF{beta}. HighlightsO_LIscRNAseq revealed Alveolar Macrophage (AM) heterogeneity and self-renewal C_LIO_LICBF{beta} is associated with AM cell cycle and facilitate AM self-renewal C_LIO_LIAMs displayed a senescence-like phenotype during physiological Aging C_LIO_LIAging impairs CBF{beta} expression in mouse and human AMs C_LI

immunology↗

bioRxiv2018

Targeting ion channel TRPM7 promotes the thymic development of Regulatory T cells by increasing IL-2-dependent STAT5 activation

In BriefGenetic deletion of Trpm7 in T-cells or pharmacological inhibition of TRPM7 channel promotes the development of fully functional Treg cells by increasing IL-2R and STAT5-dependent FOXP3 expression in the developing thymocytes. The study identifies the ion channel TRPM7 as a putative drug target to increase Treg numbers in vivo and induce immunotolerance.\n\nHIGHLIGHTSO_LIIon channel TRPM7 controls Treg development\nC_LIO_LIThe deletion of Trpm7 in the T-cell lineage increases fully functional Treg cells in the periphery\nC_LIO_LITRPM7 negatively regulates Foxp3 expression by restraining IL-2-dependent STAT5 activation\nC_LIO_LIInhibition of TRPM7 channel by FTY720 promotes the development of functional Treg cells\nC_LI\n\nSUMMARYThe thymic development of regulatory T cells (Treg), the crucial suppressors of the effector T cells (Teff), is governed by the transcription factor FOXP3. Despite the clinical significance of Treg cells, there is a dearth of druggable molecular targets capable of increasing Treg numbers in vivo. We report a surprising discovery that TRPM7 restrains Treg development by negatively regulating STAT5-dependent Foxp3 expression. The deletion of Trpm7 potentiates the thymic development of Treg cells, leads to a significantly higher frequency of functional Treg cells in the periphery and renders the mice highly resistant to T cell-dependent hepatitis. The deletion of Trpm7 or the inhibition of TRPM7 channel activity by the FDA-approved prodrug FTY720, increases IL-2 sensitivity through a feed forward positive feedback loop involving high IL-2R expression and STAT5 activation. Enhanced IL-2 signaling increases the expression of Foxp3 in thymocytes and promotes the development of Treg cells. Thus, TRPM7 emerges as the first ion channel that can be drugged to increase Treg numbers, revealing a novel pharmacological path toward the induction of immune tolerance.

immunology↗