Prostaglandin D2 inhibits hair growth and is elevated in bald scalp of men with androgenetic alopecia

Abstract

Testosterone is necessary for the development of male pattern baldness, known as androgenetic alopecia (AGA); yet, the mechanisms for decreased hair growth in this disorder are unclear. We show that prostaglandin D(2) synthase (PTGDS) is elevated at the mRNA and protein levels in bald scalp compared to haired scalp of men with AGA. The product of PTGDS enzyme activity, prostaglandin D(2) (PGD(2)), is similarly elevated in bald scalp. During normal follicle cycling in mice, Ptgds and PGD(2) levels increase immediately preceding the regression phase, suggesting an inhibitory effect on hair growth. We show that PGD(2) inhibits hair growth in explanted human hair follicles and when applied topically to mice. Hair growth inhibition requires the PGD(2) receptor G protein (heterotrimeric guanine nucleotide)-coupled receptor 44 (GPR44), but not the PGD(2) receptor 1 (PTGDR). Furthermore, we find that a transgenic mouse, K14-Ptgs2, which targets prostaglandin-endoperoxide synthase 2 expression to the skin, demonstrates elevated levels of PGD(2) in the skin and develops alopecia, follicular miniaturization, and sebaceous gland hyperplasia, which are all hallmarks of human AGA. These results define PGD(2) as an inhibitor of hair growth in AGA and suggest the PGD(2)-GPR44 pathway as a potential target for treatment.

Fig. 1

Summary of gene expression profiles of haired versus bald scalp from five men with AGA. (A) Correlation coefficients to compare the degree of difference between all haired samples, between all bald samples, or between haired versus bald samples within individuals. Data are means ± SEM (n = 5). ***P < 0.001 compared to both hair only and bald only. (B) Gene clustering algorithm of 250 significant genes in haired and bald scalps in five men labeled A to E. (C) Discontinuous graphs with gene expression level on the y axis and samples grouped as pairs along the x axis for the 250 most significant genes divided into those higher in the haired (H) scalp (169; left) and those higher in the bald (B) scalp (81; right). In each pair, the first sample (beginning of line) is the haired scalp and the second sample (end of line) is the bald scalp. (D) Gene ontology categories in haired scalp (left) versus balding scalp (right). Percentages listed are numbers of transcripts in each category out of total number of unique gene transcripts. P values are modified Fisher exact EASE (Expression Analysis Systematic Explorer) score for the significant enrichment of each category of function. (E) Fold change in PTGDS expression in bald scalp compared to haired scalp. Data are means ± SEM (n = 5). ***P < 0.0001 between haired and bald scalp for each probe set, which covers distinct areas within the PTGDS gene. (F) Schematic of the PGD₂ pathway.

Fig. 2

Increased PGD₂ pathway activity in balding scalp of men with AGA. (A) Expression of lipocalin-type PTGDS mRNA in bald versus haired scalp, as tested by qPCR. Data are means ± SEM (n = 4). (B and C) The amount of PTGDS protein in paired bald (B) and haired (H) scalps (n = 4), as shown by Western blotting with Ponceau stain used for verification of equal loading (B) and its quantitation as normalized to haired scalp (C). Data are means ± SEM. (D) PGD₂ production in bald scalp, as tested by ELISA. Data are means ± SEM (n = 3). (E and F) Fold change in PGD₂ (n = 17), 15-dPGJ₂) (n = 7), and PGE₂ (n = 17) expression in bald scalp compared to haired scalp (E). Total prostaglandin content is quantified in (F). Data are means ± SEM. *P < 0.05; **P < 0.01. In (F), P value compares haired versus bald samples for each prostaglandin.

Fig. 3

An increase in Ptgds expression precedes elevation of PGD₂ levels in the murine hair follicle during catagen. (A and B) At PN18 to PN53, Ptgds (n = 4), Ptgfr (n = 4), and Fgf5 (n = 4) mRNA (A and B) and PGD₂ lipid (n = 3) (B) were measured at different phases of hair follicle cycling. Letters correspond to hair cycle stages: A, anagen; C, catagen; T, telogen. Data are means ± SEM. In (A), **P < 0.01 for Ptgds and Fgf5 late anagen versus first telogen and for Ptgfr first telogen versus second telogen. In (B), *P < 0.05 for PGD₂ catagen versus early anagen. (C) Expression of Ptgds (n = 3) and production of PGD₂ (n = 3) in murine skin during the depilation-induced hair follicle cycle. Data are means ± SEM. *P < 0.05 for Ptgds on day 17 versus day 0; **P < 0.01 for PGD₂ day 19.5 versus day 37. (D) Murine outer root sheath keratinocytes below the stem cell–rich bulge area were stained for Ptgds (brown) 17 days after depilation. Dotted line delineates a full hair follicle. Scale bar, 100 μm. (E to G) Krt15 (red) and Ptgds (green) in permanent and non-permanent compartments of the hair follicle on day 0 (anagen) (E), day 17 (late anagen) (F), and day 19 (catagen) (G) after depilation. Scale bars, 100 μm.

Fig. 4

PTGDS is expressed in the nonpermanent portion of select human hair follicles in haired scalp, and more variably in bald scalp. (A to C) PTGDS immunostaining (brown) with blue nuclear counterstain in normal human hair follicles (A and B) and bald scalp hair follicle (C). (A) A single follicle in haired scalp with staining for PTGDS inferior to the isthmus. (B) Most anagen follicles of haired scalp have little staining for PTGDS. (C) Miniaturized hair follicle with PTGDS staining in sebocytes and hair follicle keratinocytes. Scale bars, 100 μm. (D to F) Immunofluorescence staining for PTGDS (green), nuclei (blue), and either KRT15 (red) (D) or tryptase (red) (E and F) in the hair follicle and adjoining sebaceous gland in bald scalp. Catagen follicle with some PTGDS-positive cells also expressed the mast cell marker tryptase (E). Perifollicular cells show distinct populations of PTGDS- and tryptase-positive cells with overlapping expression (F). Scale bars, 100 μm.

Fig. 5

K14-Ptgs2 transgenic mouse skin phenocopies AGA with alopecia, sebaceous gland hyperplasia, and elevated PGD₂ levels in skin. (A and B) K14-Ptgs2 animals develop alopecia (B) compared to wild-type (WT) controls (A). (C and D) Hematoxylin and eosin–stained skin from WT (C) and K14-Ptgs2 animals (D) shows sebaceous gland (yellow arrow) and hair follicle (black arrow) morphology. Scale bars, 100 μm. (E) Amount of different prostaglandins present in skin tissue from WT mice (n = 5) and K14-Ptgs2 transgenic mice (n = 3). Data are means ± SEM. **P < 0.01 comparing WT to K14-Ptgs2.

Fig. 6

PGD₂ inhibits mouse and human hair growth through GPR44. (A) Hair growth over the course of 16 days after depilation. 15-dPGJ₂ (10 μg) or vehicle was applied topically to mouse skin starting on day 8 after depilation. Data are means ± SEM (n = 3 per treatment group). *P < 0.05 compared to control. (B) Hair length 10 days after topical PGD₂ (1 μg; n = 3), 15-dPGJ₂ (1 μg; n = 3), or vehicle (n = 3) treatment. Data are means ± SEM. *P < 0.05; **P < 0.01. (C) Hair growth in Ptgdr (n = 8), Ptgds (n = 6), and Gpr44 (n = 11) knockout (KO) mice in response to PGD₂. Data are means ± SEM and are representative of three independent experiments per mouse. *P < 0.05 compared to vehicle. (D) In vitro growth of explanted human hair follicles over the course of 7 days in culture with PGD₂ (n = 3), 15-dPGJ₂ (n = 3), or vehicle (n = 3). Data are means ± SEM. ***P < 0.001 compared to vehicle.