A CD28 superagonistic antibody elicits 2 functionally distinct waves of T cell activation in rats
Nora Müller, Jens van den Brandt, Francesca Odoardi, Denise Tischner, Judith Herath, Alexander Flügel, Holger M. Reichardt
Nora Müller, Jens van den Brandt, Francesca Odoardi, Denise Tischner, Judith Herath, Alexander Flügel, Holger M. Reichardt
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Research Article

A CD28 superagonistic antibody elicits 2 functionally distinct waves of T cell activation in rats

Abstract

Administration of the CD28 superagonistic antibody JJ316 is an efficient means to treat autoimmune diseases in rats, but the humanized antibody TGN1412 caused devastating side effects in healthy volunteers during a clinical trial. Here we show that JJ316 treatment of rats induced a dramatic redistribution of T lymphocytes from the periphery to the secondary lymphoid organs, resulting in severe T lymphopenia. Live imaging of secondary lymphoid organs revealed that JJ316 administration almost instantaneously (<2 minutes) arrested T cells in situ. This reduction in T cell motility was accompanied by profound cytoskeletal rearrangements and increased cell size. In addition, surface expression of lymphocyte function–associated antigen-1 was enhanced, endothelial differentiation sphingolipid G protein–coupled receptor 1 and L selectin levels were downregulated, and the cells lost their responsiveness to sphingosine 1–phosphate–directed migration. These proadhesive alterations were accompanied by signs of strong activation, including upregulation of CD25, CD69, CD134, and proinflammatory mediators. However, this did not lead to a cytokine storm similar to the clinical trial. While most of the early changes disappeared within 48 hours, we observed that CD4+CD25+FoxP3+ regulatory T cells experienced a second phase of activation, which resulted in massive cell enlargement, extensive polarization, and increased motility. These data suggest that CD28 superagonists elicit 2 qualitatively distinct waves of activation.

Authors

Nora Müller, Jens van den Brandt, Francesca Odoardi, Denise Tischner, Judith Herath, Alexander Flügel, Holger M. Reichardt

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Figure 6

JJ316 induces proadhesive changes and interferes with S1P-mediated migration.

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T cells rapidly upregulate expression of membrane activation markers and...
(A) Lymph node cells were analyzed before and 10 hours after application of 1.0 mg JJ316 for surface expression of LFA-1 (αLβ2) and CD62L (L selectin) on CD4+ T cells by flow cytometry. (B) CD4+ T cells were purified from JJ316-treated rats at the indicated time points and analyzed for Edg1 mRNA expression by quantitative PCR. The values were normalized to β-actin (Actb) levels. n = 4; mean ± standard error of the mean. *P < 0.05. (C) The chemotactic response of CD4+ T cells isolated 12 hours after JJ316 or PBS injection toward different concentrations of S1P was studied using a transwell migration assay. The index refers to the relative number of cells that had transmigrated after 3 hours as compared with the number of cells that passed the filter in the absence of S1P. n = 4; mean ± standard error of the mean. *P < 0.05. (D) CD4+ T cells were purified from JJ316-treated rats at the indicated time points and analyzed for surface expression of CD69 by flow cytometry. Anti-mouse IgG1κ was used as an isotype control. (E) Blood samples from rats i.v. injected with 1.0 mg JJ316 or 0.1 mg FTY720 were taken at the indicated time points and analyzed for the percentages of circulating T cells (left panel) or granulocytes (right panel) by flow cytometry. n = 5; mean ± standard error of the mean. Statistical analysis was performed by comparing the 2 time courses using a 1-way ANOVA.