T cell activation and effector function in the human Jurkat T cell model

Carrasco-Padilla, Carlos; Aguilar-Sopeña, Óscar; Gómez-Morón, Álvaro; Alegre-Gómez, Sergio; Sánchez‐Madrid, Francisco; Martín-Cófreces, Noa B.; Roda-Navarro, Pedro

doi:10.1016/bs.mcb.2022.09.012

Cited by 6 publications

(5 citation statements)

References 34 publications

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“…The study of the immunosuppressive activity of MSCs and prMSCs was carried out on Jurkat cells, a human acute T lymphocyte leukemia cell line. Jurkat cells modulate T lymphocyte functions and are widely used in in vitro models [ 32 ]. The prMSCs were obtained by treatment with 10 μg/mL poly(I:C) for 3 h and incubated for 24 h in growth medium before coculture.…”

Section: Resultsmentioning

confidence: 99%

The effect of TLR3 priming conditions on MSC immunosuppressive properties

Tolstova,

Dotsenko,

Kozhin

et al. 2023

Stem Cell Res Ther

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Background Mesenchymal stromal cells (MSCs) have regenerative and immunomodulatory properties, making them suitable for cell therapy. Toll-like receptors (TLRs) in MSCs respond to viral load by secreting immunosuppressive or proinflammatory molecules. The expression of anti-inflammatory molecules in MSCs can be altered by the concentration and duration of exposure to the TLR3 ligand polyinosinic-polycytidylic acid (poly(I:C)). This study aimed to optimize the preconditioning of MSCs with poly(I:C) to increase immunosuppressive effects and to identify MSCs with activated TLR3 (prMSCs). Methods Flow cytometry and histochemical staining were used to analyze MSCs for immunophenotype and differentiation potential. MSCs were exposed to poly(I:C) at 1 and 10 μg/mL for 1, 3, and 24 h, followed by determination of the expression of IDO1, WARS1, PD-L1, TSG-6, and PTGES2 and PGE2 secretion. MSCs and prMSCs were cocultured with intact (J−) and activated (J+) Jurkat T cells. The proportion of proliferating and apoptotic J+ and J− cells, IL-10 secretion, and IL-2 production after cocultivation with MSCs and prMSCs were measured. Liquid chromatography–mass spectrometry and bioinformatics analysis identified proteins linked to TLR3 activation in MSCs. Results Poly(I:C) at 10 μg/mL during a 3-h incubation caused the highest expression of immunosuppression markers in MSCs. Activation of prMSCs caused a 18% decrease in proliferation and a one-third increase in apoptotic J+ cells compared to intact MSCs. Cocultures of prMSCs and Jurkat cells had increased IL-10 and decreased IL-2 in the conditioned medium. A proteomic study of MSCs and prMSCs identified 53 proteins with altered expression. Filtering the dataset with Gene Ontology and Reactome Pathway revealed that poly(I:C)-induced proteins activate the antiviral response. Protein‒protein interactions by String in prMSCs revealed that the antiviral response and IFN I signaling circuits were more active than in native MSCs. prMSCs expressed more cell adhesion proteins (ICAM-I and Galectin-3), PARP14, PSMB8, USP18, and GBP4, which may explain their anti-inflammatory effects on Jurkat cells. Conclusions TLR3 activation in MSCs is dependent on exposure time and poly(I:C) concentration. The maximum expression of immunosuppressive molecules was observed with 10 µg/mL poly(I:C) for 3-h preconditioning. This priming protocol for MSCs enhances the immunosuppressive effects of prMSCs on T cells.

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Section: Resultsmentioning

confidence: 99%

The effect of TLR3 priming conditions on MSC immunosuppressive properties

Tolstova,

Dotsenko,

Kozhin

et al. 2023

Stem Cell Res Ther

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“…Samples were processed as described ( 17 , 51 – 53 ). In brief, Raji cells were stained with the CMAC cell tracker (10 μM; Molecular Probes, C2110) and pulsed with SEE (0.5μg/mL; Toxin Technologies, PE404) for 30 min at 37°C.…”

Section: Methodsmentioning

confidence: 99%

End-binding protein 1 regulates the metabolic fate of CD4+ T lymphoblasts and Jurkat T cells and the organization of the mitochondrial network

Gómez-Morón,

Requena,

Pertusa

et al. 2023

Front. Immunol.

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The organization of the mitochondrial network is relevant for the metabolic fate of T cells and their ability to respond to TCR stimulation. This arrangement depends on cytoskeleton dynamics in response to TCR and CD28 activation, which allows the polarization of the mitochondria through their change in shape, and their movement along the microtubules towards the immune synapse. This work focus on the role of End-binding protein 1 (EB1), a protein that regulates tubulin polymerization and has been previously identified as a regulator of intracellular transport of CD3-enriched vesicles. EB1-interferred cells showed defective intracellular organization and metabolic strength in activated T cells, pointing to a relevant connection of the cytoskeleton and metabolism in response to TCR stimulation, which leads to increased AICD. By unifying the organization of the tubulin cytoskeleton and mitochondria during CD4+ T cell activation, this work highlights the importance of this connection for critical cell asymmetry together with metabolic functions such as glycolysis, mitochondria respiration, and cell viability.

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“…Supernatants of Jurkat cells conjugated with Raji for 18h as described [24] or primary CD4 T cells activated with αCD3αCD28 Immunocult for 18 h were collected and diluted 1:5 or 1:10 respectively and stored at -80 ºC until analysis. IL-2 detection was performed by ELISA (Human IL-2 ELISA Kit, Diaclone, 851500010) according to manufacturer's instructions.…”

Section: Elisamentioning

confidence: 99%

“…Samples were processed as described [22,24]. Briefly, Raji B cells were washed once with HBSS and loaded with the CMAC cell tracker (10 μM; Molecular Probes, C2110) and with SEE (0.5μg/mL; Toxin Technologies, PE404) for 1 hour at 37 °C.…”

Section: Cell Conjugate Formation and Immunofluorescence Experimentsmentioning

confidence: 99%

“…mCherry in the samples was excited with the 560 nm laser. The elapsed time of videos was 90 ms. To estimate the speed of actin in TIRFm images, kymographs were created from the radius of the cell, and a line was drawn, following the signal of interest signal, to measure its angle and length [24]. The speed of actin was calculated using the following trigonometric function, where the hypotenuse (H) is the length of the drawn line, the catheti are the distance (Catd) and time (Catt), and the measured angle is the angle between the hypotenuse and Catd (θ):…”

Section: Tirfm and Image Analysismentioning

confidence: 99%

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T-cell receptor triggering requires inactivation of Lim kinase-1 by Slingshot-1 phosphatase

Roda-Navarro,

Gómez-Morón,

Alegre-Gómez

et al. 2024

Preprint

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Actin dynamics control early T cell receptor signalling during T cell activation. However, the precise regulation of initial actin rearrangements is not completely understood. Here, we have investigated the regulatory role of the phosphatase Slingshot-1 (SSH1) in this process. Our data show that SSH1 rapidly polarises to nascent cognate synaptic contacts and later relocalises to peripheral F-actin networks organised at the mature synapses. Knockdown of SSH1 expression by CRISPR/Cas9-mediated genome editing or small interfering RNA revealed a regulatory role for SSH1 in CD3ε conformational change, allowing Nck binding and proper downstream signalling and immunological synapse organisation. TCR triggering induced the SSH1-mediated activation of actin dynamics through a mechanism mediated by Limk1 inactivation. These data suggest that during early TCR activation, SSH1 is required for early F-actin rearrangements that mediate initial conformational changes of the TCR, integrin organisation and proximal signalling events for proper synapse organisation.

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T cell activation and effector function in the human Jurkat T cell model

Cited by 6 publications

References 34 publications

The effect of TLR3 priming conditions on MSC immunosuppressive properties

The effect of TLR3 priming conditions on MSC immunosuppressive properties

End-binding protein 1 regulates the metabolic fate of CD4+ T lymphoblasts and Jurkat T cells and the organization of the mitochondrial network

T-cell receptor triggering requires inactivation of Lim kinase-1 by Slingshot-1 phosphatase

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