Principles and mechanisms of regeneration in the mouse model for wound-induced hair follicle neogenesis

Abstract

Wound induced hair follicle neogenesis (WIHN) describes a regenerative phenomenon in adult mammalian skin, wherein fully functional hair follicles regenerate de novo in the center of large excisional wounds. Originally described in rats, rabbits, sheep, and humans in 1940-60, the WIHN phenomenon was reinvestigated in mice only recently. The process of de novo hair regeneration largely duplicates the morphological and signaling features of normal embryonic hair development. Similar to hair development, WIHN critically depends on the activation of canonical WNT signaling. However, unlike hair development, WNT activation in WIHN is dependent on Fgf9 signaling generated by the immune system's gamma delta (γδ) T cells. The cellular bases of WIHN remain to be fully characterized, however, the available evidence leaves open the possibility for a blastema-like mechanism, wherein epidermal and/or dermal wound cells undergo epigenetic reprogramming toward a more plastic, embryonic-like state. De novo hair follicles do not regenerate from preexisting hair-fated bulge stem cells. This suggests that hair neogenesis is not driven by preexisting lineage-restricted progenitors, as is the case for amputation-induced mouse digit tip regeneration, but rather may require a blastema-like mechanism. The WIHN model is characterized by several intriguing features, which await further explanation. These include: (i) minimum wound size requirement for activating neogenesis, (ii) restriction of hair neogenesis to the wound's center, (iii) imperfect patterning outcomes, both in terms of neogenic hair positioning within the wound and in terms of their orientation. Future inquires into the WIHN process, made possible by a wide array of the available skin-specific genetic tools, will undoubtedly expand our understanding of the regeneration mechanisms in adult mammals.

Figure 1

Injury types and regenerative responses by adult HFs. HFs can efficiently regenerate following micro‐injury, as well as partial and complete amputation. (A), (B) Micro‐injury of bulge stem cells in telogen HFs, such as by laser ablation, can be efficiently repaired from the neighboring epithelial progenitor populations in the hair germ and possibly isthmus. Genetic ablation of dermal papilla cells in anagen HFs can be restored from the surviving dermal papilla cells and/or via recruitment of the neighboring dermal sheath cells. (C), (D) Anagen vibrissa follicles efficiently regenerate following amputation of the lower third, which includes the entire dermal papilla. Midway (lower half) amputations can also regenerate; however, this requires transplantation of a new dermal papilla. (E), (F) HFs can regenerate de novo following large excisional skin wounding in adult mice. This regenerative phenomenon is known as wound‐induced hair follicle neogenesis (WIHN). WIHN does not occur in small excisional wounds.

Figure 2

Timeline of hair follicle regeneration in the WIHN model. (A) Hair neogenesis in mice occurs in large excisional wounds equal to, or larger than, 1 × 1 cm. (B) The wound epithelializes and granulation tissue forms during early PWD0−14. (C) De novo hair placodes start to form around PWD14 and continue until approximately PWD19. (D) Newly formed HFs achieve full differentiation over the next 14−15 days (until approximately PWD33−34). (E), (F) Following a transient telogen phase (PWD35), de novo follicles reenter second anagen at around PWD45. Similar to normal HFs in the unwounded skin, de novo follicles in the wound center contain bulge stem cells and can cycle repetitively.

Figure 3

Key features of regenerated excisional skin wounds. (A) Typically, de novo HFs occupy the center of the regenerated wound (neogenesis zone, green). The neogenesis zone is always separated from the unwounded skin (yellow) by a hairless scar (red). This way, de novo HFs can be positively identified as residing in hair‐bearing areas surrounded by the rim of hairless scar tissue. At early PWD time points, both less and more mature de novo HFs can be seen, reflecting partial asynchrony of hair neogenesis. (B) Mature anagen de novo HFs are present during the late post‐wounding time period, PWD28. While the majority of the de novo HFs lack pigmentation, occasionally a few pigmented follicles can regenerate. (C), (D) Hair neogenesis displays a notable degree of variability, ranging from just a few HFs (D) to several hundreds (C). Here, WNT pathway reporter Axin2‐LacZ (A) and BMP pathway reporter BRE‐gal (B−D) mice (Javier et al. 2012) were used to aid visualization of neogenic hairs as strongly lacZ‐positive. Size bar 1 mm.

Figure 4

Distribution and orientation of neogenic hairs in regenerated wounds. (A) Commonly, regenerated HFs form one large cluster (also see Fig. 3). One or a few small secondary cluster(s) can also be present. (B) Seldom, multiple small de novo HF clusters can form (eight clusters here). (C), (D) Orientation of de novo HFs can range from seemingly random (D, purple region; also see Fig. 3) to unidirectional. Commonly, the neogenesis zone can contain several sub‐clusters of HFs with distinct orientation (C). Hairs have similar direction within a sub‐cluster, but often opposite of that in the neighboring sub‐cluster (white vs. black in C and D). Here, WNT pathway reporter BAT‐gal (A−C) and BRE‐gal reporters (D) were used to aid visualization of neogenic hairs as strongly lacZ‐positive. Size bar 1 mm.