p. injection, intradermal challenge with rmKC or rmLcn2 led to a stimulation of PMNs influx (Fig. 3C). In order to evaluate the kinetics of PMN mobilization from BM, we injected either rmLcn2 (200 nM) or solvent i.v. and measured granulocyte counts in the blood before injection and 1, 4, and 12 h after injection (Fig. 3D). Intriguingly, we observed a significant increase in the number of PMNs even 1 h after rmLcn2 administration (p = 0.023; Fig. 3D). PMNs counts in the periph-eral blood remained higher for the entire observation period of 12 h as compared to solvent treated animals (Fig. 3D). Because Lcn2−/− mice have reduced resistance against infections with certain gram-negative bacteria [7, 12, 14, 24-26],

we questioned whether part of this effect may be traced back to a reduced migratory potential of PMNs. Therefore, we first investigated the chemotactic activity

of blood PMNs from Lcn2−/− mice. Unexpectedly, the chemotaxis of granulocytes from C59 wnt molecular weight Lcn2−/− mice could not be stimulated upon addition of rmKC and rmLcn2 (Fig. 4A). Intriguingly, this impairment of PMN chemotaxis following addition selleck chemical of chemotactic stimuli was significant as compared to Lcn2+/+ PMNs for both, stimulation with rmKC (p = 0.022; Fig. 4C) and rmLcn2 (p = 0.029; Fig. 4D). These differences could not be explained by differences in Lcn2 receptor mRNA expression. While megalin was not expressed on PMNs of neither Lcn2+/+ or Lcn2−/− PMNs, we detected comparable mRNA expression signals of 24p3R in PMNs of Lcn2+/+ and Lcn2−/− mice. Considering the role of Lcn2 as a siderocalin, we were interested in the chemoattractive effect of Lcn2 toward PMN expression in the early course of inflammation. We thus analyzed the number and composition of white blood cells in the peritoneal cavity of thioglycolate or PBS-treated Lcn2+/+ and Lcn2−/− animals. While there was no difference in lymphocyte counts between the two genotypes (data not shown), the numbers of PMNs (p = 0.034) and monocytes (p = 0.034) were significantly lower in peritoneal cavity of thioglycolate-injected Lcn2−/− as compared to Lcn2+/+ mice (Fig. 5A and B).

Importantly, we did not observe a genotype specific difference (Lcn2+/+ versus Lcn2−/−) in the concentrations of other chemoattractants, KC and CXCL10, in the peritoneal lavage at 4 h of thioglycolate administration (details not shown). SPTLC1 To study leukocyte infiltration after a bacterial challenge, we then injected 500 CFU S. typhimurium intradermally into mice and examined the skin at site of injection 24 h later. As shown in Fig. 5C, the recruitment of immune effector cells was much lower in Lcn2−/− than in Lcn2+/+ mice (Fig. 5C). Interestingly, 48 h after infection there was no difference in abscess number or size (Supporting Information Fig. 4). We quantified S. typhimurium by immunofluorescence and detected significantly more bacteria in Lcn2−/− compared to Lcn2+/+ mice at 48 h after infection (Supporting Information Fig. 3).