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[Preprint]. 2023 Sep 20:2023.08.14.553236. Originally published 2023 Aug 16. [Version 2] doi: 10.1101/2023.08.14.553236

Figure 2. TLR2 is enriched in HFSCs and is upregulated during HFSC activation.

Figure 2.

TLR2-GFP reporter mouse skin sections were immunostained with anti-GFP to assess TLR2 expression in the hair follicles.

A. Representative confocal images of P21 first telogen hair follicle immunostained for TLR2-GFP, CD34 (bulge stem cells), P-cad (secondary hair germ (sHG)), and DAPI (nuclei). The green color in the surface rendering panel represents TLR2 expression, and other surfaces show co-localization between TLR2 and specific markers. TLR2 is present in bulge, sHG, and dermal papilla (DP) cells. P represents postnatal days. Scale bar is 10 μm.

B. TLR2-GFP in P28 anagen was co-immunostained with CD49f of basement membrane outlining the DP Scale bar is 10 μm.

C. TLR2 is co-localized to the sHG lineage (P-cad+ layers), DP, and outer root sheath (ORS) lineage. Scale bar is 20 μm.

D. TLR2-GFP in P28 anagen was co-immunostained with CD34 in old bulge (D) and Ker5 in ORS (E) revealing TLR2 localization to the old bulge, ORS, but not inner root sheath (IRS). Scale bars are 20 μm.

F. Co-immunostaining of TLR2-GFP in P38 catagen hair follicle with Ker5 in ORS lineage cells showing co-localization of TLR2 with ORS and bulge. Scale bar is 20 μm.

G. P41 late catagen hair follicle immunostained for TLR2 and CD34 showing co-localization of TLR2 to the old bulge, new bulge, sHG, and DP Scale bar is 20 μm.

H. P53 second telogen hair follicle immunostained for TLR2, CD34, and P-cad reveals co-localization of TLR2 to the bulge, sHG, and DP Scale bar is 20 μm.

I. Quantification of TLR2 fluorescent intensity in bulge cells at different phases showing TLR2 upregulation in anagen. N = 3 for each group.

J. QPCR analysis of Tlr2 mRNA expression in FACS-purified mouse HFSCs in anagen, telogen, and catagen. N = 3 or 4 per group.

K. QPCR analysis of Tlr2 mRNA expression in mouse epidermal cells and FACS-purified HFSCs showed significantly higher Tlr2 expression in HFSCs compared with raw epidermal cells. N = 6 mice per group.

All bar graphs are mean ± s.e.m. Two-tailed unpaired t-test (K) or Kruskal-Wallis test with Dunn’s post hoc test (I, J) was used to determine statistical difference.