Afferent lymph–derived T cells and DCs use different chemokine receptor CCR7–dependent routes for entry into the lymph node and intranodal migration

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Abstract

Little is known about the molecular mechanisms that determine the entry into the lymph node and intranodal positioning of lymph-derived cells. By injecting cells directly into afferent lymph vessels of popliteal lymph nodes, we demonstrate that lymph-derived T cells entered lymph-node parenchyma mainly from peripheral medullary sinuses, whereas dendritic cells (DCs) transmigrated through the floor of the subcapsular sinus on the afferent side. Transmigrating DCs induced local changes that allowed the concomitant entry of T cells at these sites. Signals mediated by the chemokine receptor CCR7 were absolutely required for the directional migration of both DCs and T cells into the T cell zone but were dispensable for the parenchymal entry of lymph-derived T cells and dendrite probing of DCs. Our findings provide insight into the molecular and structural requirements for the entry into lymph nodes and intranodal migration of lymph-derived cells of the immune system.

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Figure 1: Ccr7−/− BMDCs do not reach the paracortical TCZ after intralymphatic injection.
Figure 2: Intranodal migratory activity of intralymphatically injected DCs in popliteal lymph nodes, as visualized by two-photon microscopy.
Figure 3: Intranodal positioning of afferent lymph–derived DCs contributes to T cell proliferation but not lymph-node shutdown.
Figure 4: CD4+ T cells enter lymph-node parenchyma in peripheral medullary areas after intralymphatic injection and require CCR7 signals for migration into the paracortical TCZ.
Figure 5: Two-photon imaging of the entry into the lymph node of intralymphatically injected CD4+ T cells in peripheral medullary areas.
Figure 6: CCR7-mediated signals are required for the progression of T cells from peripheral medullary areas into the paracortical TCZ.
Figure 7: Transmigration of activated DCs changes the morphology of the SCS floor on the afferent side and allows direct entry of coinjected T lymphocytes into lymph-node parenchyma.

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Acknowledgements

We thank M. Herberg for animal care; E. Kremmer (Helmholtz Zentrum München) for antibodies; and G. Bernhardt for discussions and comments on the manuscript. Supported by Deutsche Forschungsgemeinschaft (SFB566-A14, SFB587-B3 and EXC62, 'Rebirth', to R.F.) and Boehringer Ingelheim Fonds (A.B.).

Author information

R.F., A.B. and T.W. designed experiments, analyzed data and wrote the paper; A.B. did experiments, including intralymphatic injection; T.W. contributed to Figure 1, two-photon imaging and data analysis; S.H. contributed to Figure 7 and read and commented on the manuscript; G.L.M. contributed to Supplementary Figure 2 and read and commented on manuscript; and A.M., K.H. and J.B. did histology, flow cytometry staining and DC cultures.

Correspondence to Reinhold Förster.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–5 (PDF 444 kb)

Supplementary Video 1

Ccr7-/- BMDCs do not reach the paracortical TCZ after intralymphatic application. (MOV 6411 kb)

Supplementary Video 2

Long-term two-photon microscopical imaging of intranodal DC migration behavior. (MOV 9414 kb)

Supplementary Video 3

Directional displacement and uropod formation of wt BMDCs is not reflected in Ccr7-/- BMDCs. (MOV 3477 kb)

Supplementary Video 4

Close-up view of cellular morphology during intranodal migration. (MOV 1190 kb)

Supplementary Video 5

Wt and Ccr7-/- BMDCs exhibit comparable dendrite movement and sweeping behavior. (MOV 2993 kb)

Supplementary Video 6

After i.l. application, primary skin-derived wt DC display an intranodal migration comparable to wt BMDCs. (MOV 9439 kb)

Supplementary Video 7

Transmigration of wt CD4 through lymphatic endothelium of the capsule-lining sinus floor. (MOV 6430 kb)

Supplementary Video 8

Transmigration of wt CD4 through medullar sinus endothelium. (MOV 1676 kb)

Supplementary Video 9

Upon exiting the capsule-lining sinus, wt CD4 preferably migrate within peripheral MC. (MOV 10025 kb)

Supplementary Video 10

Intralymphatically administered CD4 T cells enter LN parenchyma in a CCR7-dependent manner primarily within peripheral medullary areas. (MOV 3958 kb)

Supplementary Video 11

Immigration of DC allows CD4 T cells to enter the LN at SCS. (MOV 7426 kb)

Supplementary Video 12

LPS-treated CD4 cells fail to enter popLN via SCS. (MOV 1003 kb)

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