Biology subjects

Wagner, A. K.

Publications and source records attributed to Wagner, A. K..

7 recordsLinked to original sources

bioRxiv2026

The solid tumor microenvironment changes the hierarchy of CD155 and CD112 receptors, shaping checkpoint blockade outcome

Reproducing a physiologically relevant tumor microenvironment in vitro is essential for developing effective immunotherapeutic treatments. By integrating the use of combinatorial receptor blockade and organoid models we provide a deep functional understanding of CD155 and CD112 receptors in solid tumors and their impact on cellular immunotherapy and infiltration. CD226 showed plasticity in response to the environment, being able to switch between CD155 and CD112 depending on the ligand availability. In addition, CD226 drove NK cell infiltration into tumor tissues via CD155 and CD112 ligation, with CD226-CD112 interaction specifically promoting migration from the periphery to the core. Downregulation of CD155 and TIGIT induced by the tumor microenvironment and previous drug exposure reduced the long-term efficacy of TIGIT blockade. Taken together, our findings point towards using CD112R blockade in primary tumors to simultaneously enhance NK cell killing activity and promote infiltration into the tumor core via CD226 and CD112 interaction. ONE SENTENCE SUMMARYTumors shape the hierarchy of CD155-CD112 receptors, reducing TIGIT blockade efficacy, while CD226 drives NK infiltration and shows binding plasticity

immunology↗

bioRxiv2026

Intratumoral expression of JAML on NK cells is controlled by tumor microenvironment and MHC class I interaction

Junctional adhesion molecule-like (JAML) is an adhesion molecule known to promote T cell activation and T cell-mediated tumor rejection. In the current study, we show that JAML expression is enriched on mouse intratumoral NK cells compared with splenic NK cells. JAML+ NK cells were associated with tissue residency and co-expressed the immune checkpoints PD-1 and LAG3. JAML expression could be induced on splenic NK cells by IL-2 and further enhanced by IL-21. JAML levels were inversely correlated with inhibitory signaling, as NK cells expressing self-recognizing Ly49 receptors had reduced JAML expression, suggesting regulation of JAML expression by MHC class I molecules. Interaction with the JAML ligand CXADR also reduced JAML surface expression, indicating that tumor-mediated membrane stripping may represent a mechanism of immunoediting. Although JAML RNA transcripts were detectable in human NK cells, JAML protein was found only intracellularly. Together, these findings identify the JAML-CXADR interaction as a potential regulatory pathway in NK cell-mediated killing of tumors.

immunology↗

bioRxiv2024

A thermoplastic chip for correlative assays combining screening and high-resolution imaging of immune cell responses

Single-cell immune assays are developed for the identification and characterization of individual immune cell responses. Some methods provide snapshots of the phenotype of the cell, such as flow cytometry and single-cell RNA sequencing, whereas others, almost exclusively microscopy-based, can be used for longitudinal studies of individual cells. However, obtaining correlative data on cell dynamics and phenotype of individual immune cells is challenging but can provide more nuanced information of heterogeneous immune cell responses. In this work, we have addressed this challenge by developing an easy-to-use, disposable, thermoplastic microwell chip, designed to support screening and high-resolution imaging of single-cell behavior in two-and three-dimensional cell cultures. We show that the chip has excellent optical properties and we provide simple protocols for efficient long-term cell culture of suspension and adherent cells, the latter grown either as monolayers or as hundreds of single, uniformly-sized spheroids. We demonstrate the applicability of the system for single-cell analysis by correlating the dynamic cytotoxic response of single immune cells grown under different metabolic conditions to their intracellular cytolytic load at the end of the assay. Additionally, we illustrate highly multiplex cytotoxicity screening of tumor spheroids in the chip, comparing the effect of environment cues characteristic of the tumor microenvironment on natural killer (NK) cell-induced killing. Following the functional screening, we perform high-resolution 3D immunofluorescent imaging of infiltrating NK cells within the spheroid volumes.

immunology↗

bioRxiv2022

Jagged1 overexpression on T cells induces thymic regulatory T cells leading to thymic involution

We previously described a mouse model, tg66, with a severe defect resulting in diminished thymic size and complete involution by early adulthood. In the current study, we identified overexpression of Jagged1 as a mechanism for the alterations in thymic development in tg66 mice. T cells in the tg66 thymus were skewed towards CD8+ T cells, and within the CD4+ T cell compartment there was an over-representation of Foxp3+ cells. Regulatory Foxp3+ T cells (Tregs) isolated from tg66 mice had increased ST2 and CD103 expression. These Tregs could suppress proliferation to the same extent as conventional Tregs. Corroborating these results, tg66 mice were resistant to experimental induction of neuroinflammation in a common animal model for multiple sclerosis (EAE). Using bone marrow chimeras, we recorded a stark reduction in the number of thymocytes and a corresponding increase in Tregs in the thymus of mice receiving tg66 bone marrow. Conversely, through blocking Jagged1, the number of thymocytes was significantly increased, being concomitantly associated with a drop in the frequency of Tregs. We conclude that Tregs may play a role in thymic involution and could explain early thymic involution or loss as it is observed in diseases of thymic atrophy such as Down syndrome.

immunology↗

bioRxiv2022

Control of NK cell tolerance in MHC class I-deficiency by regulated SHP-1 localization to the activating immune synapse

Signaling via inhibitory KIR/Ly49 receptors preserves natural killer (NK) cell self-tolerance but also conveys NK cell reactivity towards MHC class-I low target cells in an education process. Here, we demonstrate that mouse NK cell education by H-2Dd regulates transcription of several genes in Ly49A+ NK cells including Ptpn6, encoding the phosphatase SHP-1. SHP-1 was highly expressed in uneducated NK cells, in which knock-out of Ptpn6 increased responsiveness. Following NKp46 triggering of uneducated NK cells, a higher synaptic abundance of phosphorylated SHP-1 was found relative to educated NK cells, concomitant with reduced phosphorylation of several signaling molecules, including PLC-g2, SLP-76, ZAP70/Syk and ERK1/2. SHP-1 overlapped extensively with F-actin and SLP-76 in the uneducated activating synapse of Ly49A+ NK cells, whereas a greater association between Ly49A and SHP-1 was observed in educated NK cells. Thus, our results indicate that in addition to transcriptional regulation, a distinct SHP-1 patterning in NK cell activating synapses can determine their tolerance.

immunology↗

bioRxiv2021

PD-1 expression on NK cells can be related to cytokine stimulation and tissue residency

Although PD-1 was shown to be a hallmark of T cells exhaustion, controversial studies have been reported on the role of PD-1 on NK cells. Here, we found by flow cytometry and single cell RNA sequencing analysis that PD-1 can be expressed on MHC class I-deficient tumor-infiltrating NK cells in vivo. We also demonstrate distinct alterations in the phenotype of PD-1-deficient NK cells which in part could be attributed to a decrease in tumor-infiltrating NK cells in PD-1-deficient mice. NK cells from PD-1-deficient mice exhibited a more mature phenotype which might reduce their capacity to migrate and kill in vivo. Finally, our results demonstrate that PD-L1 molecules in membranes of PD-1-deficient NK cells migrate faster than in NK cells from wildtype mice, suggesting that PD-1 and PD-L1 form cis interactions with each other on NK cells.

immunology↗

bioRxiv2021

GABAergic signaling in human and murine NK cells upon challenge with Toxoplasma gondii

Protective cytotoxic and proinflammatory cytokine responses by natural killer (NK) cells impact the outcome of infections by Toxoplasma gondii, a common parasite in humans and other vertebrates. However, T. gondii can also sequester within NK cells and downmodulate their effector functions. Recently, the implication of {gamma}-aminobutyric acid (GABA) signaling in infection and inflammation-related responses of mononuclear phagocytes and T cells has become evident. Yet, the role of GABAergic signaling in NK cells has remained unknown. Here, we report that human and murine NK cells synthesize and secrete GABA in response to infection challenge. Parasitized NK cells secreted GABA while activation stimuli, such as IL-12/IL-18 or parasite lysates, failed to induce GABA secretion. GABA secretion by NK cells was associated to a transcriptional upregulation of GABA synthesis enzymes (GAD65/67) and was abrogated by GAD-inhibition. Further, NK cells expressed GABA-A receptor subunits and GABA signaling regulators, with transcriptional modulations taking place upon challenge with T. gondii. Exogenous GABA and GABA-containing supernatants from parasitized dendritic cells (DCs) impacted NK cell function by reducing the degranulation and cytotoxicity of NK cells. Conversely, GABA-containing supernatants from NK cells enhanced the migratory responses of parasitized DCs. This enhanced DC migration was abolished by GABA-A receptor antagonism or GAD-inhibition and was reconstituted by exogenous GABA. Jointly, the data show that NK cells are GABAergic cells and that GABA hampers NK cell cytotoxicity in vitro. We hypothesize that GABA secreted by parasitized immune cells modulates the immune responses to T. gondii infection. Summary sentenceIn response to infection challenge, NK cells synthesize and secrete{gamma} -aminobutyric acid (GABA), which impacts NK cell and dendritic cell functions via GABA-A receptors.

immunology↗