Spatial Distribution and Functional Impact of Human Scalp Hair Follicle Microbiota.

Human hair follicles (HFs) constitute a unique microbiota habitat that differs substantially from the skin surface. Traditional HF sampling methods fail to eliminate skin microbiota contaminants or assess the HF microbiota incompletely, and microbiota functions in human HF physiology remain ill explored. Therefore, we used laser-capture microdissection, metagenomic shotgun sequencing, and FISH to characterize the human scalp HF microbiota in defined anatomical compartments. This revealed significant compartment-, tissue lineage-, and donor age-dependent variations in microbiota composition. Greatest abundance variations between HF compartments were observed for viruses, archaea, Staphylococcus epidermidis, Cutibacterium acnes, and Malassezia restricta, with the latter 2 being the most abundant viable HF colonizers (as tested by propidium monoazide assay) and, surprisingly, most abundant in the HF mesenchyme. Transfection of organ-cultured human scalp HFs with S. epidermidis-specific lytic bacteriophages ex vivo downregulated transcription of genes known to regulate HF growth and development, metabolism, and melanogenesis, suggesting that selected microbial products may modulate HF functions. Indeed, HF treatment with butyrate, a metabolite of S. epidermidis and other HF microbiota, delayed catagen and promoted autophagy, mitochondrial activity, and gp100 and dermcidin expression ex vivo. Thus, human HF microbiota show spatial variations in abundance and modulate the physiology of their host, which invites therapeutic targeting.

Management of the human hair follicle microbiome by a synthetic odorant.

BACKGROUND: Human scalp hair follicles (HFs) engage in olfactory receptor (OR)-dependent chemosensation. Activation of olfactory receptor family 2 subfamily AT member 4 (OR2AT4) by the synthetic, sandalwood-like odorant Sandalore® up-regulated HF antimicrobial peptide expression of dermcidin (DCD), which had previously been thought to be produced exclusively by sweat and sebaceous glands. OBJECTIVES: To understand if intrafollicular DCD production can be stimulated by a commonly used cosmetic odorant, thus altering human HF microbiome composition in a clinically beneficial manner. METHODS: DCD expression was compared between fresh-frozen scalp biopsies and microdissected, full-length scalp HFs, organ-cultured in the presence/absence of the OR2AT4 agonist, Sandalore® and/or antibiotics and/or the competitive OR2AT4 antagonist, Phenirat®. Amplicon-based sequencing and microbial growth assays were performed to assess how this treatment affected the HF microbiome. RESULTS: Synthetic odorant treatment upregulated epithelial DCD expression and exerted antimicrobial activity in human HFs ex vivo. Combined antibiotic and odorant treatment, during an ex vivo dysbiosis event, prevented HF tissue damage and favoured a more physiological microbiome composition. Sandalore®-conditioned medium, containing higher DCD content, favoured Staphylococcus epidermidis and Malassezia restricta over S. aureus and M. globosa, while exhibiting antimicrobial activity against Cutibacterium acnes. These effects were reversed by co-administration of Phenirat®. CONCLUSIONS: We provide the first proof-of-principle that a cosmetic odorant impacts the human HF microbiome by up-regulating antimicrobial peptide production in an olfactory receptor-dependent manner. Specifically, a synthetic sandalwood-like odorant stimulates intrafollicular DCD production, likely via OR2AT4, and thereby controls microbial overgrowth. Thus, deserving further exploration as an adjuvant therapeutic principle in the management of folliculitis and dysbiosis-associated hair diseases.

Laser capture microdissection as a method for investigating the human hair follicle microbiome reveals region-specific differences in the bacteriome profile.

OBJECTIVE: Human hair follicles (HFs) are populated by a rich and diverse microbiome, traditionally evaluated by methods that inadvertently sample the skin microbiome and/or miss microbiota located in deeper HF regions. Thereby, these methods capture the human HF microbiome in a skewed and incomplete manner. This pilot study aimed to use laser-capture microdissection of human scalp HFs, coupled with 16S rRNA gene sequencing to sample the HF microbiome and overcome these methodological limitations. RESULTS: HFs were laser-capture microdissected (LCM) into three anatomically distinct regions. All main known core HF bacterial colonisers, including Cutibacterium, Corynebacterium and Staphylococcus, were identified, in all three HF regions. Interestingly, region-specific variations in α-diversity and microbial abundance of the core microbiome genera and Reyranella were identified, suggestive of variations in microbiologically relevant microenvironment characteristics. This pilot study therefore shows that LCM-coupled with metagenomics is a powerful tool for analysing the microbiome of defined biological niches. Refining and complementing this method with broader metagenomic techniques will facilitate the mapping of dysbiotic events associated with HF diseases and targeted therapeutic interventions.

Application of Topical Sandalore® Increases Epidermal Dermcidin Synthesis in Organ-Cultured Human Skin ex vivo.

INTRODUCTION: Several olfactory receptors (ORs) are expressed in human skin, where they regulate skin pigmentation, barrier function, wound healing, and hair growth. Previously, we found that the selective activation of OR family 2 subfamily AT member 4 (OR2AT4) by the synthetic, sandalwood-like odorant Sandalore® differentially stimulates the expression of antimicrobial peptides (AMPs) in human scalp hair follicle epithelium ex vivo. As OR2AT4 is also expressed by epidermal keratinocytes, we hypothesized that it may modulate intraepidermal AMP synthesis, thereby contributing to skin microbiome management. METHODS: We investigated this hypothesis in organ-cultured human skin in the presence of Sandalore® and antibiotics and evaluated epidermal production of two AMPs, LL37 (cathelicidin) and dermcidin (DCD), as well as OR2AT4, by quantitative immunohistomorphometry. Moreover, we quantified DCD secretion into the culture medium by ELISA and studied the effect of culture medium on selected bacterial and fungal strains. RESULTS: Topical application of Sandalore®to organ-cultured human skin increased OR2AT4 protein expression, the number of DCD-positive intraepidermal cells, and DCD secretion into culture media, without significantly affecting epidermal LL37 expression. In line with the significantly increased secretion of DCD into the culture medium, we demonstrated, in a spectrophotometric assay, that application of conditioned media from Sandalore®-treated skin promotes Staphylococcus epidermidis, Malassezia restricta, and, minimally, Cutibacterium acnes and inhibits Staphylococcus aureus growth. CONCLUSION: In addition to demonstrating for the first time that DCD can be expressed by epidermal keratinocytes, our pilot study suggests that topical treatment of human skin with a cosmetic odorant (Sandalore®) has the potential to alter the composition of the human skin microbiome through the selective upregulation of DCD. If confirmed, Sandalore® could become an attractive adjuvant, nondrug treatment for dermatoses characterized by dysbiosis due to overgrowth of S. aureus and Malassezia, such as atopic dermatitis and seborrheic dermatitis.

Hydra and the hair follicle - An unconventional comparative biology approach to exploring the human holobiont.

The microbiome of human hair follicles (HFs) has emerged as an important player in different HF and skin pathologies, yet awaits in-depth exploration. This raises questions regarding the tightly linked interactions between host environment, nutrient dependency of host-associated microbes, microbial metabolism, microbe-microbe interactions and host immunity. The use of simple model systems facilitates addressing generally important questions and testing overarching, therapeutically relevant principles that likely transcend obvious interspecies differences. Here, we evaluate the potential of the freshwater polyp Hydra, to dissect fundamental principles of microbiome regulation by the host, that is the human HF. In particular, we focus on therapeutically targetable host-microbiome interactions, such as nutrient dependency, microbial interactions and host defence. Offering a new lens into the study of HF - microbiota interactions, we argue that general principles of how Hydra manages its microbiota can inform the development of novel, microbiome-targeting therapeutic interventions in human skin disease.