Data on importance of hematopoietic cell derived Lipocalin 2 against gut inflammation

The data herein is related to the research article entitled “Microbiota-inducible innate immune siderophore binding protein Lipocalin 2 is critical for intestinal homeostasis” (Singh et al., 2016) [1]. In the present article, we monitored dextran sodium sulfate (DSS)-induced colitis development upon Lipocalin 2 (Lcn2) neutralization, and examined the survival of Lcn2 deficient (Lcn2KO) mice and their WT littermates upon DSS challenge. To dissect the relative contribution of immune and non-immune cells-derived Lcn2 in mediating protection against gut inflammation, we generated respective bone marrow chimera and evaluated their susceptibility to IL-10 receptor neutralization-induced chronic colitis. Neutralization of Lcn2 in WT mice resulted in exacerbated DSS-induced colitis. Notably, mice lacking Lcn2 exhibited 100% mortality whereas only 20% mortality was observed in WT mice upon DSS challenge. Further, data from bone marrow chimera showed that immune cell-derived Lcn2 is the major contributor in conferring protection against colitis.


a b s t r a c t
The data herein is related to the research article entitled "Microbiota-inducible innate immune siderophore binding protein Lipocalin 2 is critical for intestinal homeostasis"   [1]. In the present article, we monitored dextran sodium sulfate (DSS)induced colitis development upon Lipocalin 2 (Lcn2) neutralization, and examined the survival of Lcn2 deficient (Lcn2KO) mice and their WT littermates upon DSS challenge. To dissect the relative contribution of immune and non-immune cells-derived Lcn2 in mediating protection against gut inflammation, we generated respective bone marrow chimera and evaluated their susceptibility to IL-10 receptor neutralization-induced chronic colitis.
Neutralization of Lcn2 in WT mice resulted in exacerbated DSSinduced colitis. Notably, mice lacking Lcn2 exhibited 100% mortality whereas only 20% mortality was observed in WT mice upon DSS challenge. Further, data from bone marrow chimera showed that immune cell-derived Lcn2 is the major contributor in conferring protection against colitis.
& The data provide information on protective roles of immune vs non-immune cell derived Lcn2 during gut inflammation.
The data support future studies in delineating the role of Lcn2 in mucoprotection.

Data
In the data, we presented the higher mortality rate of Lcn2KO mice in DSS-induced acute colitis when compared to their WT littermates (Fig. 1). Further, neutralization of Lcn2 in DSS-treated WT mice resulted in exacerbated gut inflammation as indicated by greater body weight loss, splenomegaly, colon thickening, and serum KC (Fig. 2). Experiments with bone marrow chimeric mice revealed that immune cells-derived Lcn2 are the major contributor in mediating protection against IL-10R neutralization-induced chronic colitis (Fig. 3) indicated by reduction in splenomegaly, colomegaly, shrunken ceca, MPO protein levels and activity.

DSS induced mortality
Eight weeks old female C57BL/6 WT and Lcn2KO mice (n ¼5) were administered 1.5% DSS (MP Biomedicals Inc.) in drinking water for 7 days. Colonic inflammation was examined by fecal occult blood, diarrhea, and loss of body weight. After 7 days, mice were switched to regular drinking and subsequently monitored for mortality up to 21 days. Fig. 2. Data on effect of Lcn2-neutralization on severity of colitis. WT mice (female, 8 weeks old, n¼ 5) were administered 1.5% DSS for 7 days. On day 3 onwards up to day 7, colitic mice were administered once/day with either anti-mouse Lcn2 mAb (mLcn2, 20 mg/mouse/day, intraperitoneally) or isotype antibody (IgG1). After 7 days, mice were euthanized and analyzed for standard colitis parameters. (A) Body weight, (B) spleen weight, (C) colon thickening and (D) serum KC. Data are expressed as mean 7SEM. Unpaired t-test was used to compare the difference between groups. *p o 0.05 was considered statistically significant.

Lipocalin 2 (Lcn2) neutralization in DSS-induced colitis model
Female WT mice (8 weeks old; n ¼5) were administered 1.5% DSS in drinking water for 7 days. On day 3 onwards up to day 7, mice were administered once daily with anti-mouse Lcn2 monoclonal antibody (20 mg/mouse/day, intraperitoneally; R&D Systems). Control mice received isotype (IgG1) control. On day 7, mice were euthanized by CO 2 asphyxiation and blood samples were collected in a serum separating tube (Becton Dickinson). Upon centrifugation, the hemolysis-free serum were collected and stored at À 80°C until further analysis. Standard colitis parameters (body weight, spleen weight, occult blood and colon shortening) were assessed. Serum KC were analyzed by ELISA according to the manufacturer's protocol [2].
After 8 weeks post-transplant, Lcn2 bone marrow chimeric mice (n ¼4) were administered antimouse α-IL10R mAb (1.0 mg/mouse, intraperitoneally; BioXcell) once weekly for total of 4 weeks to induce chronic colitis as described earlier [1]. Control mice received isotype (IgG1) control. Mice were regularly monitored for fecal occult blood and change in body weight. One week after the fourth injection, the mice were euthanized and analyzed for standard colitis parameters [5]. MPO levels in the serum and colon lysate were measured by ELISA according to the manufacturer's protocol.

Statistical analysis
Data are expressed as mean 7SEM. The significance of difference between two groups was determined by unpaired student t-test. The statistical significance between 3 or more groups were analyzed by one-way ANOVA with Tukey's post-hoc test. p value less than 0.05 were considered statistically significant. Statistical significance was calculated using Graph Pad Prism version 6.0 (Graph Pad Software Inc.).