Background: Immunity and tolerance are regulated by changes in secondary lymphoid organ (SLO) structure and function. SLO structural elements include the fibers ER-TR7, desmin, and laminin. Laminins are a stromal fiber family with multiple members possessing distinct functions and are remodeled by matrix metalloproteinases (MMP). CD40-CD40L blockade + donor-specific splenocyte transfusion (DST) induces long-term graft acceptance, yet the mechanisms by which acceptance occurs remain incompletely defined. This study investigated the hypothesis that tolerance induction results in unique tolerogenic structural changes within the lymph node (LN).

Methods: Tolerance was induced in C57BL/6 mice with BALB/c DST + anti-CD40L mAb. Quantitative immunohistochemistry of LN was used to define ER-TR7, laminin, and desmin. Adoptively transferred alloantigen specific T cells were tracked with vital dyes. Laminin remodeling was blocked with MMP inhibitors (MMI).

Results: There were immediate changes in LN structure and T cell migration occurring within hours of tolerogen treatment. There was an increase in ER-TR7 and desmin fiber expression throughout the LN, particularly in the regions of the cortical ridge and high endothelial venules (HEV). ER-TR7 branching complexity increased around tolerant HEV. In contrast, immunity caused a global increase in LN laminins, particularly laminin-10/11, which has costimulatory properties. In tolerant LN there was a relative decrease in laminin-10/11 and an increase in laminin-8, which promotes T cell egress from vessels. Concurrently, the number of total and antigen specific T cells entering the HEV increased as early as 4 hours following tolerance induction. Inhibition of laminin processing with MMI altered LN morphology and lymphocyte trafficking from tolerogenic to immunogenic in character, increasing the laminin-10/11 present in the LN and surrounding the HEV.

Conclusions: Tolerance induction results in acute and unique morphologic changes in the LN, showing LN scaffolding as a plastic, malleable structure. These changes occur both globally and on the micro-environmental scale. Structural changes to the HEV indicate an active and permissive environment for T cell entry, migration, and differentiation. Interference with these structural changes affects antigen specific and Treg trafficking through the HEV as they enter the LN. These findings provide novel evidence that the physical LN environment changes following tolerance induction and critically affects the trafficking of lymphocytes as they enter the LN.

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