Background: Tolerance and immunity are associated with differential expression of lymph node (LN) stromal laminins α4β1γ1 (411) and α5β1γ1 (511), respectively, which determine how alloreactive T cells enter high endothelial venules and traffic within LN. We tested the hypothesis that laminins differentially stimulated CD4 T cell surface receptors and intracellular signaling pathways, thus regulating the balance of the Treg/Th17 axis.

Methods: CD4 T cells isolated from C57BL/6 mice were activated in pro-Th17 (IL6, TGFβ, anti-IL4, anti-IFNγ) or pro-Treg cultures (TGFβ, IL2). Expression of Foxp3, IL-17 and phosphorylated ERK1/2, AKT, STAT5 and p38 were measured by flow cytometry. T cell receptor transgenic CD4 T cells from TEa mice, recognizing donor Ea alloantigen presented by recipient I-A^b, were transferred into Ea immunized mice and assayed for Th17 differentiation. C57BL/6 mice, and laminin α5 floxed x Pdgfrb-Cre mice that lack laminin α5 expression in LN, received cardiac transplants from BALB/c donors. Recipients were treated with blocking mAbs to the laminin 511 receptors α6 integrin or α-dystroglycan.

Results: In pro-Treg cultures, laminin 511 inhibited Treg induction while upregulating P-STAT5, P-ERK1/2, P-AKT, and P-p38. The antibody against α6 integrin abrogated these effects. In contrast, laminin 411 increased Treg and inhibited transcription factor phosphorylation as well as laminin 511 activity. In pro-Th17 cultures, laminin 511 promoted Th17 induction, which was abrogated by anti-α-dystroglycan. Laminin 411 reduced the proportion of Th17 cells and partially reversed the laminin 511 effect. Laminin 411 inhibited P-STAT5 while laminin 511 increased it. In vivo blockade of α-dystroglycan decreased alloantigen specific Th17 induction in Ea peptide immunized recipients. Adding anti-α6 integrin to anti-CD40L mAb resulted in prolongation of allograft survival (mean survival time 53 days to 167 days (P=0.0018)). Laminin α5 deficient mice treated with anti-CD40L mAb also had a marked increase in allograft survival compared to controls (109 vs 87 days).

Conclusions: Acting as pro-suppressive and pro-inflammatory ligands, respectively, laminins 411 and 511 directly regulate the Treg/Th17 axis. T cells recognize laminin 511 through α6 integrin and α-dystroglycan receptors. These act as costimulatory receptors both in vitro and in vivo. Laminins are not only LN structural molecules, but directly modulate the Treg/Th17 axis and act as molecular switches for tolerance and immunity.

CITATION INFORMATION: Li L., Simon T., Piao W., Xiong Y., Saxena V., Zhang T., Wagner C., Bromberg J. Lymph Node Stromal Laminins Modulate the Dynamic Balance of the Treg/Th17 Axis Am J Transplant. 2017;17 (suppl 3).

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