Modelling Human Hair Follicles-Lessons from Animal Models and Beyond.

The hair follicle is a specialized appendage of the skin that is critical for multiple functions, including thermoregulation, immune surveillance, and sebum production. Mammals are born with a fixed number of hair follicles that develop embryonically. Postnatally, these hair follicles undergo regenerative cycles of regression and growth that recapitulate many of the embryonic signaling pathways. Furthermore, hair cycles have a direct impact on skin regeneration in homeostasis, cutaneous wound healing, and disease conditions such as alopecia. Here, we review the current knowledge of hair follicle formation during embryonic development and the post-natal hair cycle, with an emphasis on the molecular signaling pathways underlying these processes. We then discuss efforts to capitalize on the field's understanding of in vivo mechanisms to bioengineer hair follicles or hair-bearing skin in vitro and how such models may be further improved to develop strategies for hair regeneration.

Divergent functions of histone acetyltransferases KAT2A and KAT2B in keratinocyte self-renewal and differentiation.

The mammalian epidermis undergoes constant renewal, replenished by a pool of stem cells and terminal differentiation of their progeny. This is accompanied by changes in gene expression and morphology that are orchestrated, in part, by epigenetic modifiers. Here, we define the role of the histone acetyltransferase KAT2A in epidermal homeostasis and provide a comparative analysis that reveals key functional divergence with its paralog KAT2B. In contrast to the reported function of KAT2B in epidermal differentiation, KAT2A supports the undifferentiated state in keratinocytes. RNA-seq analysis of KAT2A- and KAT2B- depleted keratinocytes revealed dysregulated epidermal differentiation. Depletion of KAT2A led to premature expression of epidermal differentiation genes in the absence of inductive signals, whereas loss of KAT2B delayed differentiation. KAT2A acetyltransferase activity was indispensable in regulating epidermal differentiation gene expression. The metazoan-specific N terminus of KAT2A was also required to support its function in keratinocytes. We further showed that the interplay between KAT2A- and KAT2B-mediated regulation was important for normal cutaneous wound healing in vivo. Overall, these findings reveal a distinct mechanism in which keratinocytes use a pair of highly homologous histone acetyltransferases to support divergent functions in self-renewal and differentiation processes.

Generation and integration of hair follicle-primed spheroids in bioengineered skin constructs.

Skin is a complex organ made up of different cell layers, appendages, connective tissues, and immune repertoires. These different components interact extensively to maintain the overall functions of the integumentary system. In particular, appendages such as hair follicles critically contribute to the skin's function in thermoregulation, sensory perception, and homeostatic regeneration. Despite a strong need for better skin regenerative therapeutics, efforts to bio-engineer highly functional appendage-containing human reconstituted skinin vitrohave not yielded much success. Here, we report methods in generating and incorporating hair follicle-primed heterotypic spheroids into epidermal-dermal skin constructs that induced invaginating outgrowths with follicle-like organization and lineage gene expression. By co-culturing epithelial keratinocytes (KCs) with dermal papilla (DP) cells in low attachment plates, we established the media and culture conditions that best supported the viability, signalling and remodelling of the cell aggregates to form 3D KC-DP spheroids with the expression of both DP inductiveness and hair follicle lineage genes. We show that long-term growth and maturation of KC cells in these spheroids was supported by incorporation into epidermal-dermal constructs but not in scaffold-less media. When cultured, the bio-fabricated constructs developed invaginations from the integrated spheroids with follicle-forming potential. The generation of these constructs is a step towards the development of functional hair-bearing skin mimetics.

Monitoring Wound Healing with Topically Applied Optical NanoFlare mRNA Nanosensors.

An effective wound management strategy needs accurate assessment of wound status throughout the whole healing process. This can be achieved by examining molecular biomarkers including proteins, DNAs, and RNAs. However, existing methods for quantifying these biomarkers such as immunohistochemistry and quantitative polymerase chain reaction are usually laborious, resource-intensive, and disruptive. This article reports the development and utilization of mRNA nanosensors (i.e., NanoFlare) that are topically applied on cutaneous wounds to reveal the healing status through targeted and semi-quantitative examination of the mRNA biomarkers in skin cells. In 2D and 3D in vitro models, the efficacy and efficiency of these nanosensors are demonstrated in revealing the dynamic changes of mRNA biomarkers for different stages of wound development. In mouse models, this platform permits the tracking and identification of wound healing stages and a normal and diabetic wound healing process by wound healing index in real time.

A peptidoglycan storm caused by ß-lactam antibiotic's action on host microbiota drives Candida albicans infection.

The commensal fungus Candida albicans often causes life-threatening infections in patients who are immunocompromised with high mortality. A prominent but poorly understood risk factor for the C. albicans commensal‒pathogen transition is the use of broad-spectrum antibiotics. Here, we report that ß-lactam antibiotics cause bacteria to release significant quantities of peptidoglycan fragments that potently induce the invasive hyphal growth of C. albicans. We identify several active peptidoglycan subunits, including tracheal cytotoxin, a molecule produced by many Gram-negative bacteria, and fragments purified from the cell wall of Gram-positive Staphylococcus aureus. Feeding mice with ß-lactam antibiotics causes a peptidoglycan storm that transforms the gut from a niche usually restraining C. albicans in the commensal state to promoting invasive growth, leading to systemic dissemination. Our findings reveal a mechanism underlying a significant risk factor for C. albicans infection, which could inform clinicians regarding future antibiotic selection to minimize this deadly disease incidence.

Application of 3D Bioprinting Technologies to the Management and Treatment of Diabetic Foot Ulcers.

Diabetes mellitus (DM) is a chronic metabolic disease with increasing prevalence worldwide. Diabetic foot ulcers (DFUs) are a serious complication of DM. It is estimated that 15-25% of DM patients develop DFU at least once in their lifetime. The lack of effective wound dressings and targeted therapy for DFUs often results in prolonged hospitalization and amputations. As the incidence of DM is projected to rise, the demand for specialized DFU wound management will continue to increase. Hence, it is of great interest to improve and develop effective DFU-specific wound dressings and therapies. In the last decade, 3D bioprinting technology has made a great contribution to the healthcare sector, with the development of personalized prosthetics, implants, and bioengineered tissues. In this review, we discuss the challenges faced in DFU wound management and how 3D bioprinting technology can be applied to advance current treatment methods, such as biomanufacturing of composite 3D human skin substitutes for skin grafting and the development of DFU-appropriate wound dressings. Future co-development of 3D bioprinting technologies with novel treatment approaches to mitigate DFU-specific pathophysiological challenges will be key to limiting the healthcare burden associated with the increasing prevalence of DM.