Figure 6: iMo require FcRγ chain and Syk for migration into the peritoneal cavity after CLP.
(a,c) iMo from WT, AF251705−/−, Tyrobp−/− or Fcer1g−/− mice were incubated on human IgG1-precoated or VCAM-1-Fc-precoated plates in the presence or absence of LPS for 20 h, and adherent cells were counted by means of microscopy (n=3 per group). Scale bars, 100 μm. (b) iMo from WT or AF251705−/− mice were incubated in the presence or absence of LPS for 20 h, subjected to flow adhesion assays to VCAM-1-Fc or human IgG1-coated glass capillaries, and then adherent cells were counted by means of microscopy (n=3 per group). (d) CFSE-labelled iMo from WT, AF251705−/−, Tyrobp−/− or Fcer1g−/− mice were transferred intravenously into AF251705−/− mice 20 h after CLP. Migrated CFSE+ iMo in the peritoneal cavity were counted by using flow cytometry 20 h after CLP (n=7, 9 or 13 per group). (e) iMo from WT, AF251705−/−, Tyrobp−/− or Fcer1g−/− mice were transferred intravenously into AF251705−/− mice immediately before CLP. Mice were monitored every 6 h for 6 days to determine survival (n=6, 7, 3 or 5 per group). (f) iMo from WT or AF251705−/− mice were stimulated with LPS on VCAM-1-Fc-precoated plates in the presence or absence of piceatannol, a Syk inhibitor, for 20 h, and adherent cells were counted by means of microscopy (n=3 per group). (g) iMo from WT or Myd88−/− mice were left unstimulated or stimulated with LPS for 20 h, and the expression of Fcer1g and Tyrobp mRNA was analysed by using real-time reverse transcription–PCR (n=3 per group). (h) iMo from WT or Myd88−/− mice were incubated on VCAM-1-Fc-precoated plates in the presence or absence of LPS for 20 h, and adherent cells were counted by means of microscopy (n=3 per group). *P<0.05, **P<0.01 and ***P<0.001 (two-tailed Student’s t-test in (a–d,f–h)) and log-rank test in e). N.S., not significant. Error bars indicate s.d. Data are representative of two independent experiments. See also Supplementary Figs 9 and 10.
