Fig. 6

Contralateral SC MSC injection does not inhibit LN neutrophil TNF response to LPS re-exposure. (A) C57BL/6 mice were SC-injected with 30 ng LPS in 50 µl PBS in the left hock, and 4 h later the skin around the LPS injection site (LPS inflamed) and on the contralateral hock (LPS non-inflamed) was excised and homogenised for cytokine analysis. Data expressed as mean± SEM; n = 10 mice per group from 2 independent experiments. One-way ANOVA, Tukey’s multiple comparison, * p < 0.05, ** p < 0.01, *** p < 0.001, ns, not significant. (B) Bioluminescent imaging of mice that received SC injection of 30 ng LPS in 50 µl PBS in the left hock, and 1 h later 1 × 10⁶ luciferase-expressing MSCs in the same hock, or (C) in the non-inflamed hock on the contralateral side. (D) Bar graphs represent changes in radiance over time. n = 3 mice per group from one experiment. (E) C57BL/6 mice were SC-injected with 30 ng LPS in 50 µl PBS in the left hock. 1 h after LPS injection, mice received a SC injection of 1 × 10⁶ human adipose MSCs (MSC-c) or control PBS treatment (PBS-c) in the contralateral (non-inflamed) hock. The popliteal LNs draining the LPS-injected and the contralateral skin were harvested 4 h after LPS injection (i.e. 3 h after MSC-c/PBS-c treatment) and restimulated with 10 µg/ml LPS for 4 h. Intracellular TNF production was used as an indicator of the pro-inflammatory response to the LPS rechallenge. (F) Number of TNF⁺ neutrophils after LPS restimulation of LN cells from mice that received SC injection of MSC or control PBS on the contralateral hock. Data expressed as mean ± SEM; n = 5 mice per group. One-way ANOVA, Dunnett’s multiple comparison; * p < 0.05, ** p < 0.01, *** p < 0.001, ns, not significant. (G) Number of TNF⁺ monocytes after LPS restimulation of LN cells from mice that received SC injection of MSC or control PBS on the contralateral hock. Data expressed as mean ± SEM; n = 5 mice per group. One-way ANOVA, Dunnett’s multiple comparison; * p < 0.05, ** p < 0.01, *** p < 0.001, ns, not significant.