Table 1.
Mouse primers used.
Fig 1.
Intradermal administration of IL-31 results in an enhanced epidermal thickness.
(A) C57BL/6 mice were injected intradermally with saline and rIL-31 (20μg) daily for 14 days and a portion of dorsal skin was excised and processed for thin-sectioning. Haematoxylin/eosin staining was performed to analyze epidermal thickness. (B) Quantification of the epidermal thickness was performed using MetaMorph Image analysis software v6.2. Data are cumulative of three independent experiments, with 8–13 total numbers of mice in each group, and represented as mean ± SEM. Unpaired Student’s t-test was used to measure significant differences between the groups; **, P<0.01.
Fig 2.
IL-31 increases trans-epidermal water loss (TEWL) in skin.
Level of TEWL at Days 7 and 14 on dorsal skin was measured. Data are cumulative of three independent experiments with 8–13 total mice in each group and represented as mean ± SEM. Unpaired Student’s t-test was used to measure significant differences between the groups: ***, P<0.001; ****, P<0.0001.
Fig 3.
IL-31 increases epidermal cell proliferation in skin.
C57BL/6 mice were injected intradermally with saline or rIL-31 (20μg) daily for 14 days. A portion of dorsal skin was excised and processed for thin-sectioning and immunohistochemistry. (A) Skin sections from saline- and IL-31-treated mice were immunostained using Ki67 antibody. (B) The number of Ki67+ cells in saline- and rIL-31-treated skin were determined using MetaMorph Image analysis software and represented as mean ± SEM. Unpaired Student’s t-test was used to measure significant differences between the groups; *, P<0.05.
Fig 4.
IL-31 regulates expression of genes involved in skin damage.
(A) C57BL/6 mice were injected intradermally with saline or rIL-31 (20μg) daily for 14 days. A portion of dorsal skin was excised and RNA was isolated. RNA-Seq analysis was performed using next-generation sequencing. Heat map shows two clusters of differentially expressed genes that were either up or down regulated (indicated with color key) in rIL-31-treated mice compared to saline-treated controls. A total of 1,016 significant gene results was analyzed using a P value cut-off of 0.05; FDR<0.1 and greater than two-fold changes. (B) Network representation of the biological-function enrichment analysis of IL-31-regulated genes identified by RNA-Seq and top-gene function analysis. Solid lines depict the interactions between the genes and biological processes.
Fig 5.
IL-31 increases the expression of genes involved in cell proliferation and tissue remodeling.
C57BL/6 mice were injected intradermally with saline or rIL-31 (20μg) daily for 14 days and a portion of dorsal skin was excised. RNA was isolated and converted to c-DNA, and transcripts of genes identified to be regulated through IL-31 network were quantified using RT-PCR. (A-C) Proliferative markers—Il6, Il1β, Egr1; Remodelling markers—(D-F) Fn1, Mmp2, Mmp10, and (G) Trpv2. Numbers of mice in each group were 4–5. Data are represented as mean ± SEM, and unpaired Student’s t-test was used to measure significant differences between the groups: *, P<0.05; **, P<0.01; ***, P<0.001.
Fig 6.
IL-31 increases the expression of genes that alter the mechanical integrity of skin.
C57BL/6 mice were injected intradermally with saline or rIL-31 (20μg) daily for 14 days, and a portion of dorsal skin was excised. RNA was isolated and converted to c-DNA. Genes known to be involved in barrier function, inflammation, and mechanical integrity were quantified using qRT-PCR; (A-B) S100A8 and S100A9, (C) Saa1, (D) Krt1, and (E) Jup. Numbers of mice in each group were 4–5. Data are represented as mean ± SEM, and unpaired Student’s t-test was used to measure significant differences between the groups: *, P<0.05; **, P<0.01.