Modulating embryonic signaling pathways paves the way for regeneration in wound healing.

Epithelial tissues, including the skin, are highly proliferative tissues with the capability to constant renewal and regeneration, a feature that is essential for survival as the skin forms a protective barrier against external insults and water loss. In adult mammalian skin, every injury will lead to a scar. The scar tissue that is produced to seal the wound efficiently is usually rigid and lacks elasticity and the skin's original resilience to external impacts, but also secondary appendages such as hair follicles and sebaceous glands. While it was long thought that hair follicles develop solely during embryogenesis, it is becoming increasingly clear that hair follicles can also regenerate within a wound. The ability of the skin to induce hair neogenesis following injury however declines with age. As fetal and neonatal skin have the remarkable capacity to heal without scarring, the recapitulation of a neonatal state has been a primary target of recent regenerative research. In this review we highlight how modulating dermal signaling or the abundance of specific fibroblast subsets could be utilized to induce de novo hair follicles within the wound bed, and thus to shift wound repair with a scar to scarless regeneration.

Hedgehog Signaling in Papillary Fibroblasts Is Essential for Hair Follicle Regeneration during Wound Healing.

Patients suffering from large scars such as burn victims not only encounter aesthetic challenges but also ongoing itching or pain that substantially deteriorates their quality of life. Skin appendages such as hair follicles rarely regenerate within the healing wound. Because they are crucial for skin homeostasis and the lack thereof constitutes one of the main limitations to scarless wound healing, their regeneration represents a major objective for regenerative medicine. Fibroblasts, the main resident cell type of the skin dermis, mediate embryonic hair follicle morphogenesis and are particularly involved in wound healing because they orchestrate extracellular matrix remodeling and collagen deposition in the wound bed. Importantly, dermal fibroblasts originate from two distinct developmental lineages with unique functions that differently mediate the response to epidermal signals such as Hedgehog signaling. In this study, we show that Hedgehog signaling in the reticular fibroblast lineage promotes the initial phase of wound repair, possibly by modulating angiogenesis and fibroblast proliferation, whereas Hedgehog signaling in papillary fibroblasts is essential to induce de novo hair follicle formation within the healing wound.

Isolation of Papillary and Reticular Fibroblasts from Human Skin by Fluorescence-activated Cell Sorting.

Fibroblasts are a highly heterogeneous cell population implicated in the pathogenesis of many human diseases. In human skin dermis, fibroblasts have traditionally been attributed to the superficial papillary or lower reticular dermis according to their histological localization. In mouse dermis, papillary and reticular fibroblasts originate from two different lineages with diverging functions regarding physiological and pathological processes and a distinct cell surface marker expression profile by which they can be distinguished. Importantly, evidence from explant cultures from superficial and lower dermal layers suggest that at least two functionally distinct dermal fibroblasts lineages exist in human skin dermis as well. However, unlike for mouse skin, cell surface markers enabling the discrimination of different fibroblast subsets have not yet been established for human skin. We developed a novel protocol for the isolation of human papillary and reticular fibroblast populations via fluorescence-activated cell sorting (FACS) using the two cell surface markers Fibroblast Activation Protein (FAP) and Thymocyte antigen 1 (Thy1)/CD90. This method enables the isolation of pure fibroblast subsets without in vitro manipulation, which was shown to affect gene expression, thus permitting accurate functional analysis of human dermal fibroblast subsets in regard to tissue homeostasis or disease pathology.

Lineage Identity and Location within the Dermis Determine the Function of Papillary and Reticular Fibroblasts in Human Skin.

Human skin dermis is composed of the superficial papillary dermis and the reticular dermis in the lower layers, which can easily be distinguished histologically. In vitro analyses of fibroblasts from explant cultures from superficial and lower dermal layers suggest that human skin comprises at least two fibroblast lineages with distinct morphology, expression profiles, and functions. However, while for mouse skin cell surface markers have been identified, allowing the isolation of pure populations of one lineage or the other via FACS, this has not been achieved for human skin fibroblasts. We have now discovered two cell surface markers that discriminate between papillary and reticular fibroblasts. While FAP+CD90- cells display increased proliferative potential, express PDPN and NTN1, and cannot be differentiated into adipocytes, FAP-CD90+ fibroblasts express high levels of ACTA2, MGP, PPARγ, and CD36 and readily undergo adipogenic differentiation, a hallmark of reticular fibroblasts. Flow cytometric analysis of fibroblasts isolated from superficial and lower layers of human dermis showed that FAP+CD90- cells are enriched in the papillary dermis. Altogether, functional analysis and expression profiling confirms that FAP+CD90- cells represent papillary fibroblasts, whereas FAP-CD90+ fibroblasts derive from the reticular lineage. Although papillary and reticular fibroblasts are enriched in the upper or lower dermis, respectively, they are not spatially restricted, and the microenvironment seems to affect their function.

PI3K activity in dendritic cells exerts paradoxical effects during autoimmune inflammation.

The peripheral activation of autoreactive T cells and subsequent central nervous system (CNS) immune cell infiltration are key events relevant for experimental autoimmune encephalomyelitis (EAE), a commonly employed multiple sclerosis (MS) model, influenced by TH1 and TH17 mediated immunity. The phosphoinositide-3-kinase (PI3K)-AKT kinase pathway modulates outcome during EAE, with direct actions of PI3K on adaptive immunity implicated in deleterious and effects on antigen presenting cells involved in beneficial responses during EAE. Here, by genetically deleting the regulatory subunit of Class Ia PI3K, p85α, in selective myeloid cells, we aimed to resolve the impact of PI3K in EAE. While genetically deleting PI3K in LysM expressing cells exerted unremarkable effects, attenuating PI3K function in CD11c+ dendritic cells (DCs), promoted secretion of pathogenic EAE promoting cytokines, particularly skewing TH1 and TH17 immunity, while notably, improving health in EAE. Neutralizing IFN-γ activity using blocking antibodies revealed a prolonged TH1 response was critical for the decreased disease of these animals. Thus, PI3K-AKT signaling in DCs acts in a paradoxical manner. While attenuating EAE associated TH1 and TH17 responses, it impairs health during autoimmune inflammation.