(A–E). CD31 images using widefield fluorescence microscopy, with optical Z-stacks and deconvolution from 70 µm thick skin sections, shown as maximal projection images. Yellow-dotted line indicates …
(A–D) Wide field microscopy images of 10 µm skin sections from wild type C57/Bl6 mice at ages and hair cycle stages indicated, stained for endothelial marker CD31 and DAPI for nuclei. Ep, epidermis; …
(A) Quantification of total CD31+ area per skin field at different hair cycle stages (as shown in Figure 1) shows significant increase of vasculature at anagen. The total CD31+ area is defined by …
Spreadsheet of original quantification of CD31+ vasculature in total skin (for Figure 2A).
Spreadsheet of original quantification of CD31+ vasculature in HPuHG region (for Figure 2B).
Spreadsheet of original quantification of angles of CD31+ vasculature (for Figure 2C).
(A–F) Skin sections from mice in Figure 1 at late catagen (PD19), telogen (PD20), early anagen (PD21), and anagen (PD28) stained for CD31 and Ki67 or activated Caspase 3, imaged using confocal …
Number of Ki67+ CD31+ double positive cells during hair cycle (for Figure 3F).
(A) Schematic of tamoxifen (TM) inductions using the endothelial-specific Cdh5-CreERT2, with hair cycle stages indicated at postnatal days. M, morphogenesis; C, catagen, T, telogen, A, anagen. (B) …
(A) Images of skin sections from Alk1 EndKO and CT mice induced with TM at PD21 show anagen induction by PD31 (see also Table 1). (B) Quantification of Pcadhigh cells of stage-matched Alk1 EndKO and …
(A) Skin sections from Alk1flox/flox mice with Cdh5-CreERT2 (EndKO) or without (CT) were injected with tamoxifen (TM) at PD17 and sacrificed at different ages between ~PD 22-PD28, were stage matched …
Spreadsheet of original quantification of HPuHG CD31+ vasculature in Alk1 EndKO and CT (for Figure 5B).
(A) Example of Cdh5-CreERT2 x Alk1flox/flox mouse skin (Alk1 EndKO) with regions of interest analyzed in Figure 5. Blue line shows how the CD31+ vasculature in the HPuHG was delineated, dotted …
(A) Representative images of skin stained with BMP4 antibodies show prominent expression in the HPuHG region in vessel-like structure. Note increased BMP4 in the HPuHG region in the Alk1 EndKO skin. …
Spreadsheet of original quantification of BMP4 in Alk1 EndKO skin (for Figures 6B, C and D).
(A) Example of Cdh5-CreERT2 x Alk1flox/flox mouse skin (Alk1 EndKO) from mice injected with TM at PD17 and sacrificed at PD28 stage matched at Tel-Ana I, with regions of interest analyzed in Figure 6…
(A) Schematics of Runx1 mutant inductions in genetic mouse models to create varying levels of Runx1 in the hair follicle (HF). Runx1 expression in HF is shown in red in wild type follicles (top) and …
(A–B) Runx1flox/flox mice with Krt14-CreERT2 (EpiKO) or without (CT) were injected with tamoxifen (TM) at PD17, sacrificed at ~PD20. Skin sections were DAPI stained, inspected for hair morphology …
Spreadsheet of original quantification on Runx1 EpiKO skin (Figure 8C and D).
Spreadsheet of original quantification of BMP4 in Runx1 EpiKO skin (for Figure 8E and F).
(A) Examples of images from Krt14-CreERT2 x Runx1flox/flox mice (Runx1 EpiKO) and littermate control mice without Cre allele (CT) that were analyzed in Figure 7. Light blue line shows how the CD31+ …
Left side: During late stages of HFSC quiescence (late catagen/telogen) remodeling of skin vasculature results in assembly of the horizontal plexus under hair germ HPuHG. This results in high …
Mouse ID | Genotype | Sex | Hair cycle stage |
---|---|---|---|
PD17-20 | |||
F520.01 | CT | M | Tel |
F520.04 | CT | F | Tel |
F520.05 | CT | M | Tel |
F520.06 | CT | F | Tel |
PD17-22 | |||
F167.02 | CT | M | Ana I |
F64.02 | CT | F | Tel - AnaI |
F64.01 | KO | F | Tel |
PD17-25 | |||
F81.06 | CT | F | Tel- Ana I |
F146.05 | CT | M | Ana I - IIa |
F167.05 | CT | M | Ana I - IIa |
F433.03 | CT | F | Tel- Ana I |
F433.04 | CT | F | Tel- Ana I |
F81.07 | KO | F | Tel |
F86.02 | KO | F | Ana I |
F86.04 | KO | M | Tel |
F99.01 | KO | M | Tel |
F433.06 | KO | F | Tel |
PD17-28 | |||
E963.02 | CT | M | Ana IIIc |
F80.04 | CT | F | Tel - Ana I |
F90.01 | CT | M | Ana I - Ana IIa |
F146.02 | CT | F | Ana IIa-c |
F333.01 | CT | M | Ana I - IIa |
F333.03 | CT | M | Tel - Ana I |
F333.04 | CT | M | Tel - Ana I |
F11.04 | KO | F | Tel - Ana IIa |
F89.01 | KO | M | Ana I-IIa |
F80.03 | KO | M | Tel - Ana I |
F90.02 | KO | F | Tel - Ana I |
F333.02 | KO | F | Tel - Ana I |
PD17-31 | |||
F3.01 | CT | M | Ana II - Ana III |
F3.03 | CT | M | Ana II |
F90.05 | CT | M | Ana II-III |
F146.06 | CT | M | Ana III |
E906.01 | KO | M | Tel-Ana I |
F90.07 | KO | F | Tel-Ana I |
PD17-34/35 | |||
E928.03 | CT | F | Ana III |
E928.05 | CT | F | Ana IV |
E960.01 | KO | F | Ana I - Ana II |
E928.01 | KO | F | Ana I-Ana II |
PD21-26 | |||
F218.01 | CT | M | Ana I - Ana III |
F218.05 | KO | M | Ana III |
PD21-31 | |||
F218.02 | CT | F | Ana I - Ana II |
F218.04 | KO | F | Tel - Ana I |
PD23-26 | |||
F273.02 | CT | F | Ana III |
F273.01 | KO | M | Ana III |
Gene abbreviation | Full gene name | Known function | Fold change bu Runx1 TG/WT | Fold change in WT HG vs bu |
---|---|---|---|---|
Figf | c-fos induced growth factor | Agiogenesis, VEGF signaling | 2x | 16x |
Col5a1 | Colagen five type a1 | Ehlers‐Danlos syndrome with lethal arterial events | −11x | −2.7x |
Adamts1 | A disintegrin-like and metallopeptidase thrombospondin type 1 | Negative regulation of angiogenesis | 2x | 2x |
Cxcl1 | chemokine (C-X-C motif) ligand1 | Inflammatory response | 2x | 3x |
Cxcl10 | chemokine (C-X-C motif) ligand10 | Negative regulation of angiogenesis | 6x | 16x |
Edn1 | endothelin 1 | Patterning of blood vessels | 2x | 2x |
Cxcl12 | chemokine (C-X-C motif) ligand 12 | Patterning of blood vessels | 2x | 2x |
Cx3cl1 | chemokine (C-X3-C motif) ligand1 | Angiogenesis | 2x | 4x |
Cxcl9 | chemokine (C-X-C motif) ligand9 | Inflammatory response | 7x | 10x |
Pla2g7 | phospholipase A2, group VII (platelet-activating factor acetylhydrolase, plasma) | Inflammatory response | 10x | 6x |
Sema3a | semaphorin 3A | Blood vessel patterning | 9x | 3x |
Reagent type (species) or resource | Designation | Source or reference | Identifiers | Additional information |
---|---|---|---|---|
Genetic reagent (M. musculus) | Krt14-CreERT2 | PMID: 11034212 | RRID: MGI:2177426 | |
Genetic reagent (M. musculus) | Cdh5-CreERT2 | PMID: 20445537 | RRID: MGI:3848982 | Dr. Anne Eichmann (Yale University) |
Genetic reagent (M. musculus) | Alk1flox | PMID: 17911384 | RRID: MGI:4398901 | Dr. Anne Eichmann (Yale University) |
Genetic reagent (M. musculus) | Runx1flox | PMID: 15784726 | RRID: MGI:3043614 | |
Genetic reagent (M. musculus) | Rosa26-tdTomato | Jackson Laboratory | RRID: MGI:3809523 | |
Genetic reagent (M. musculus) | C57BL/6 | Jackson Laboratory | RRID:MGI:3715241 | |
Antibody | Anti-mouse-CD31 (Purified rat monoclonal) | BD Biosciences | BD Cat#: 550274, RRID: AB_393571 | 1:100 |
Antibody | Anti-Ki67 (Rabbit polyclonal) | Abcam | Abcam Cat# ab15580, RRID:AB_443209 | 1:1000 |
Antibody | Anti-Caspase-3 (Rabbit polyclonal) | R and D Systems | R and D Cat# AF835, RRID:AB_2243952 | 1:300 |
Antibody | Anti-neurofilament-1 (Chicken polyclonal) | Millipore | Millipore Cat# AB5539, RRID:AB_11212161 | 1:1000 |
Antibody | Anti-BMP4 (Rabbit polyclonal) | Abcam | Abcam Cat# ab39973, RRID:AB_2063523 | 1:1000 |
Antibody | Anti-EphrinB2 (Rabbit monoclonal) | Abcam | Abcam Cat #: ab201512, RRID:AB_2810831 | 1:500 |
Antibody | Anti-P-Cadherin (Rat monoclonal) | Thermo Fisher Scientific | Thermo Fisher Scientific Cat# 13-2000Z, RRID:AB_2533006 | 1:200 |
Chemical compound, drug | Tamoxifen | Millipore | Millipore Cat#: T5648 | |
Software, algorithm | ImageJ | ImageJ | RRID:SCR_003070 | |
Software, algorithm | JMP Pro | SAS | RRID: SCR_014242 |