Figure 3. TNFα instruct the neutrophils to migrate into the lymphatic system upon antigen sensitisation.

Neutrophil migration into the lymphatic system of the cremaster muscle following antigen sensitisation with complete Freund’s adjuvant (CFA+Ag) was induced in WT and TNFRdbKO animals as well as in chimeric animals exhibiting neutrophils deficient in TNFRs. (a) Time course of TNFα release in the cremaster muscles of WT mice following intra-scrotal injection of CFA+Ag and as quantified by ELISA. (b) TNFα release in mice subjected to clodronate liposome-induced macrophage depletion. (c) Number of extravasated neutrophils in cremaster muscles of WT and TNFRdbKO mice at 16 hrs post-CFA+Ag-stimulation as quantified by confocal microscopy. (d) Number of neutrophils within cremaster lymphatic vessels of WT and TNFRdbKO mice at 16 hrs post-CFA+Ag-stimulation as quantified by confocal microscopy. (e) Percentage of neutrophils in dLNs of WT and TNFRdbKO mice at 16 hrs post-CFA+Ag-stimulation as quantified by flow cytometry. (f) Number of extravasated neutrophils in cremaster muscles at 16 hrs post-CFA+Ag-stimulation from chimeric animals receiving bone marrow transplant from WT or TNFRdbKO donor mice and as quantified by confocal microscopy. (g) Number of neutrophils within cremaster lymphatic vessels at 16 hrs post-CFA+Ag-stimulation from chimeric animals receiving bone marrow transplant from WT or TNFRdbKO donor mice and as quantified by confocal microscopy. (h) Number of neutrophils found in the dLNs of chimeric animals receiving bone marrow transplant from WT or TNFRdbKO donor mice as quantified by confocal microscopy 16 hrs post-CFA+Ag-stimulation. Data are expressed as mean ± SEM of N = 5–12 animals per group from at least 5–10 experiments. Statistically significant differences between stimulated and control groups or between WT and TNFRdbKO mice are indicated by asterisks: *P < 0.05; **P < 0.01; ****P < 0.0001.