Delivery of costimulatory blockade to lymph nodes promotes transplant acceptance in mice
Jing Zhao, Sungwook Jung, Xiaofei Li, Lushen Li, Vivek Kasinath, Hengcheng Zhang, Said N. Movahedi, Ahmad Mardini, Gianmarco Sabiu, Yoonha Hwang, Vikas Saxena, Yang Song, Bing Ma, Sophie E. Acton, Pilhan Kim, Joren C. Madsen, Peter T. Sage, Stefan G. Tullius, George C. Tsokos, Jonathan S. Bromberg, Reza Abdi
Jing Zhao, Sungwook Jung, Xiaofei Li, Lushen Li, Vivek Kasinath, Hengcheng Zhang, Said N. Movahedi, Ahmad Mardini, Gianmarco Sabiu, Yoonha Hwang, Vikas Saxena, Yang Song, Bing Ma, Sophie E. Acton, Pilhan Kim, Joren C. Madsen, Peter T. Sage, Stefan G. Tullius, George C. Tsokos, Jonathan S. Bromberg, Reza Abdi
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Research Article

Delivery of costimulatory blockade to lymph nodes promotes transplant acceptance in mice

Abstract

The lymph node (LN) is the primary site of alloimmunity activation and regulation during transplantation. Here, we investigated how fibroblastic reticular cells (FRCs) facilitate the tolerance induced by anti-CD40L in a murine model of heart transplantation. We found that both the absence of LNs and FRC depletion abrogated the effect of anti-CD40L in prolonging murine heart allograft survival. Depletion of FRCs impaired homing of T cells across the high endothelial venules (HEVs) and promoted formation of alloreactive T cells in the LNs in heart-transplanted mice treated with anti-CD40L. Single-cell RNA sequencing of the LNs showed that anti-CD40L promotes a Madcam1+ FRC subset. FRCs also promoted the formation of regulatory T cells (Tregs) in vitro. Nanoparticles (NPs) containing anti-CD40L were selectively delivered to the LNs by coating them with MECA-79, which binds to peripheral node addressin (PNAd) glycoproteins expressed exclusively by HEVs. Treatment with these MECA-79–anti-CD40L-NPs markedly delayed the onset of heart allograft rejection and increased the presence of Tregs. Finally, combined MECA-79–anti-CD40L-NPs and rapamycin treatment resulted in markedly longer allograft survival than soluble anti-CD40L and rapamycin. These data demonstrate that FRCs are critical to facilitating costimulatory blockade. LN-targeted nanodelivery of anti-CD40L could effectively promote heart allograft acceptance.

Authors

Jing Zhao, Sungwook Jung, Xiaofei Li, Lushen Li, Vivek Kasinath, Hengcheng Zhang, Said N. Movahedi, Ahmad Mardini, Gianmarco Sabiu, Yoonha Hwang, Vikas Saxena, Yang Song, Bing Ma, Sophie E. Acton, Pilhan Kim, Joren C. Madsen, Peter T. Sage, Stefan G. Tullius, George C. Tsokos, Jonathan S. Bromberg, Reza Abdi

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Figure 1

FRCs are critical for anti-CD40L–induced long-term heart allograft survival.

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FRCs are critical for anti-CD40L–induced long-term heart allograft survi...
(A) Comparison of heart allograft survival between WT (n = 5 mice/group, MST > 100 days) and CCL19/DTR recipients (n = 6 mice/group, MST = 34.5 days) of BALB/c (B/c) hearts treated with high-dose anti-CD40L and DT. Log-rank test for graft survival. (B) Representative light micrographs of H&E-stained heart allograft sections from WT and CCL19/DTR recipients on day 50 after heart transplantation. Scale bars: 100 μm. (C) Comparison of cellular infiltration and vascular damage of heart allografts in WT and CCL19/DTR recipients (n = 4 mice/group). (D) Comparison of MFI of CD3+ cells, CD11b+ cells, and collagen I+ cells in heart allografts from WT and CCL19/DTR recipients (n = 4 mice/group). (E) Comparison of Treg numbers in DLNs from WT and CCL19/DTR recipients by flow cytometry (n = 4–5 mice/group). (F) Comparison between percentages of CD4+ Teffs, CD4+TNF-α+, CD4+IFN-γ+, CD4+IL-17+, CD8+ Teffs, CD8+TNF-α+, CD8+IFN-γ+, and CD8+IL-17+ cells in the DLNs of WT and CCL19/DTR recipients by flow cytometry (n = 4–5 mice/group). (G) Intravital imaging showed GFP+ T cells migrating around the HEVs in the DLNs of CCL19/DTR and WT skin allograft recipients. Scale bars: 50 μm. (H) Comparison of average velocity of T cells in the DLNs from WT and CCL19/DTR mice. (I) Comparison between numbers of type I conventional DCs (cDC1), type II conventional DCs (cDC2), and peripheral DCs (pDC) and percentages of CD80+ cDC1, CD86+ cDC1, CD80+ cDC2, CD86+ cDC2, MHC II+ cDC2, CD80+ pDC2, CD86+ pDC2, and MHC II+ pDC2 in the DLNs of WT and CCL19/DTR recipients by flow cytometry (n = 4–5 mice/group). (J) Comparison between numbers of CD11c-GFP cells in DLNs from WT and CCL19/DTR mice 2 hours after i.v. or s.c. injection (n = 3 mice/group). Student’s t test for 2-group comparisons. Data presented as mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.