Background: Transplant tolerance is exquisitely sensitive to changes in secondary lymphoid organ structure and function. Costimulatory blockade plus donor-specific splenocyte transfusion (DST) induces long-term graft acceptance in mice. Tolerization induces regulatory T cells (Treg), with their generation occurring in conjunction with gross changes in lymph node (LN) morphology, and specifically the important structural fiber ER-TR7. We tested the hypothesis that targeting the ER-TR7 fibers involved in LN remodeling would prevent transplant tolerance.

Methods: Tolerance was induced in C57BL/6 recipients of BALB/c cardiac allografts with DST + anti-CD40L mAb. ER-TR7+ stromal fibers were targeted by treating tolerized recipients with anti-ER-TR7 mAb. Quantitative immunohistochemistry defined LN structure and morphology of ER-TR7, laminins, and desmin. Adoptively transferred alloantigen specific T cells were tracked with vital dyes.

Results: Tolerance induction caused an increase in the total amount of ER-TR7+ fibers in the LN and the branching of ER-TR7+ fibers away from the high endothelial venules (HEV), showing active LN remodeling. Treatment of tolerant recipients with anti-ER-TR7 mAb changed LN morphology to resemble the morphology of rejecting LN, with increased desmin, total laminin, and laminin-10/11 around the HEV. Anti-ER-TR7 mAb also increased the number of naive T cells and decreased the number of Treg associated with the HEV. Finally, treatment of allograft recipients with anti-ER-TR7 mAb prevented tolerization with DST + anti-CD40L. The resulting graft pathology showed patchy perivascular and interstitial monocytic infiltrates, parenchymal hemorrhage, and perivascular and interstitial fibrosis.

Conclusions: Tolerance induction requires gross and fine structural modifications within the LN, including changes in the total amounts and morphology of the structural elements ER-TR7, desmin, and laminin. Structural alterations in and around the HEV regulate naive and Treg entry into and localization within the LN. Anti-ER-TR7 mAb altered LN morphology, lymphocyte entry and lymphocyte localization around the HEV, so that the structures now resembled the rejecting phenotype, while at the same time preventing tolerization. Thus, tolerance requires a specialized and unique LN structure that dictates T cell migration, function, and fate, and inhibition of these structural changes results in reversal of transplant tolerance.

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