Homeostatic Control of Sebaceous Glands by Innate Lymphoid Cells Regulates Commensal Bacteria Equilibrium.

Immune cells and epithelium form sophisticated barrier systems in symbiotic relationships with microbiota. Evidence suggests that immune cells can sense microbes through intact barriers, but regulation of microbial commensalism remain largely unexplored. Here, we uncovered spatial compartmentalization of skin-resident innate lymphoid cells (ILCs) and modulation of sebaceous glands by a subset of RORγt+ ILCs residing within hair follicles in close proximity to sebaceous glands. Their persistence in skin required IL-7 and thymic stromal lymphopoietin, and localization was dependent on the chemokine receptor CCR6. ILC subsets expressed TNF receptor ligands, which limited sebocyte growth by repressing Notch signaling pathway. Consequently, loss of ILCs resulted in sebaceous hyperplasia with increased production of antimicrobial lipids and restricted commensalism of Gram-positive bacterial communities. Thus, epithelia-derived signals maintain skin-resident ILCs that regulate microbial commensalism through sebaceous gland-mediated tuning of the barrier surface, highlighting an immune-epithelia circuitry that facilitates host-microbe symbiosis.

Thymic stromal lymphopoietin induces IL-4/IL-13 from T cells to promote sebum secretion and adipose loss.

BACKGROUND: The cytokine TSLP promotes type 2 immune responses and can induce adipose loss by stimulating lipid loss from the skin through sebum secretion by sebaceous glands, which enhances the skin barrier. However, the mechanism by which TSLP upregulates sebaceous gland function is unknown. OBJECTIVES: This study investigated the mechanism by which TSLP stimulates sebum secretion and adipose loss. METHODS: RNA-sequencing analysis was performed on sebaceous glands isolated by laser capture microdissection and single-cell RNA-sequencing analysis was performed on sorted skin T cells. Sebocyte function was analyzed by histological analysis and sebum secretion in vivo and by measuring lipogenesis and proliferation in vitro. RESULTS: This study found that TSLP sequentially stimulated the expression of lipogenesis genes followed by cell death genes in sebaceous glands to induce holocrine secretion of sebum. TSLP did not affect sebaceous gland activity directly. Rather, single-cell RNA-sequencing revealed that TSLP recruited distinct T-cell clusters that produce IL-4 and IL-13, which were necessary for TSLP-induced adipose loss and sebum secretion. Moreover, IL-13 was sufficient to cause sebum secretion and adipose loss in vivo and to induce lipogenesis and proliferation of a human sebocyte cell line in vitro. CONCLUSIONS: This study proposes that TSLP stimulates T cells to deliver IL-4 and IL-13 to sebaceous glands, which enhances sebaceous gland function, turnover, and subsequent adipose loss.

Short-Chain Fatty Acids from Cutibacterium acnes Activate Both a Canonical and Epigenetic Inflammatory Response in Human Sebocytes.

The regulation of cutaneous inflammatory processes is essential for the human skin to maintain homeostasis in the presence of the dense communities of resident microbes that normally populate this organ. Forming the hair follicle-associated sebaceous gland, sebocytes are specialized lipid-producing cells that can release inflammatory mediators. Cytokine and chemokine expression by pilosebaceous epithelial cells (i.e., sebocytes and follicular keratinocytes) has been proposed to contribute to the common human skin disease acne vulgaris. The underlying mechanisms that drive inflammatory gene expression in acne-involved pilosebaceous epithelial cells are still unknown because almost all sebaceous follicles contain dense concentrations of bacteria yet only some show an inflammatory reaction. In this study, we hypothesized that metabolites from the abundant skin-resident microbe Propionibacterium acnes can influence cytokine expression from human sebocytes. We show that short-chain fatty acids produced by P. acnes under environmental conditions that favor fermentation will drive inflammatory gene expression from sebocytes. These molecules are shown to influence sebocyte behavior through two distinct mechanisms: the inhibition of histone deacetylase (HDAC) activity and the activation of fatty acid receptors. Depletion of HDAC8 and HDAC9 in human sebocytes resulted in an enhanced cytokine response to TLR-2 activation that resembled the transcriptional profile of an acne lesion. These data provide a new insight into the regulation of inflammatory gene expression in the skin, further characterize the contribution of sebocytes to epidermal immunity, and demonstrate how changes in the metabolic state of the skin microbiome can promote inflammatory acne.

Homeostasis of the sebaceous gland and mechanisms of acne pathogenesis.

BACKGROUND: Sebaceous glands (SGs) are appendages of mammalian skin that produce a mixture of lipids known as sebum. Acne vulgaris is an exceptionally common skin condition, characterized by elevated sebum production, altered sebum composition, and the formation of infundibular cysts, called comedones. Comedo-associated SGs are atrophic, suggesting that comedo formation involves abnormal differentiation of progenitor cells that generate the SG and infundibulum: the 'comedo switch'. Understanding the biological processes that govern SG homeostasis promises to highlight potential aetiological mechanisms underlying acne and other SG-associated skin disorders. RESULTS: In this review, we discuss the clinical data, genetic mouse models and in vitro research that have highlighted major hormones, paracrine factors, transcription factors and signalling pathways that control SG homeostasis. These include, but are not limited to androgens, progestogens and oestrogens; retinoids; receptor tyrosine kinases such as ErbB family receptors, fibroblast growth factor receptor 2 and insulin/insulin-like growth factor 1 receptors; peroxisome proliferator-activated receptor γ; aryl hydrocarbon receptor; and the Wnt signalling pathway. Where possible, the cellular and molecular mechanisms by which these regulatory factors control SG biology are indicated, along with considerations as to how they might contribute to acne pathogenesis. CONCLUSIONS: Future research should seek to establish the relative importance, and causative relationships, of altered sebum production, sebum composition, inflammation and abnormal differentiation of sebaceous progenitors to the process of comedo formation in acne. Such an understanding will allow for therapeutic targeting of regulatory factors that control SG homeostasis, with the aim of treating acne.

Atrophic scar formation in patients with acne involves long-acting immune responses with plasma cells and alteration of sebaceous glands.

BACKGROUND: Possible outcomes of acne lesions are atrophic scars, which may cause serious psychological distress. Current treatments for postacne scarring often require invasive procedures. Pathophysiological studies on acne scarring have only investigated the first week of papule life. OBJECTIVES: To study the pathophysiology of atrophic scar formation to identify molecular and cellular pathways that can lead to new therapies for the prevention of acne scarring. METHODS: Large-scale gene expression profiling and immunohistochemistry analysis were performed on uninvolved skin and papules in both scar-prone (SP) and non-scar-prone (NSP) patients with acne, at different time points. RESULTS: Gene expression and immunohistochemistry analyses showed a very similar immune response in 48-h-old papules in SP and NSP populations, characterized by elevated numbers of T cells, neutrophils and macrophages. However, the immune response only persisted in SP patients in 3-week-old papules, and was characterized by an important B-cell infiltrate. Transient downmodulation of sebaceous gland markers related to lipid metabolism was observed in 48-h-old papules in NSP patients, followed by normalization after 3 weeks. In contrast, in SP patients a drastic reduction of these markers persisted in 3-week-old papules, suggesting an irreversible destruction of sebaceous gland structures after inflammatory remodelling in SP patients with acne. CONCLUSIONS: Long-lived acne papules are characterized by a B-cell infiltrate. A relationship exists between the duration and severity of inflammation and the alteration of sebaceous gland structures, leading to atrophic scar formation in acne.

Demodex mites modulate sebocyte immune reaction: possible role in the pathogenesis of rosacea.

BACKGROUND: Rosacea is a common facial skin disorder mainly affecting middle-aged adults. Its aetiology is unknown and pathogenesis uncertain. Activation of the host innate immune response has been identified as an important factor. The Demodex mite population in the skin of rosacea patients is significantly higher than in patients with normal skin, suggesting that they may be of aetiological importance in this disorder. OBJECTIVES: To determine the potential of Demodex mites to interact with the host immune system. METHODS: Live Demodex mites were extracted from normal facial skin of control subjects and used in cell stimulation experiments with the immortalized SZ95 sebocyte line. Time- and mite-dose-dependent experiments were performed. Direct effects of Demodex and effects of the medium in which Demodex had been cultured were evaluated on the Toll-like receptor (TLR) signalling pathway on both a gene and protein expression level. RESULTS: Mites modulated TLR signalling events on both mRNA and protein levels in SZ95 sebocytes. An initial trend towards downmodulation of genes in this pathway was observed. A subsequent switch to positive gene upregulation was recorded after 48 h of coculture. Demodex secreted bioactive molecules that affected TLR2 receptor expression by sebocytes. High numbers of Demodex induced proinflammatory cytokine secretion, whereas lower numbers did not. CONCLUSIONS: Demodex mites have the capacity to modulate the TLR signalling pathway of an immortalized human sebocyte line. Mites have the capacity to secrete bioactive molecules that affect the immune reactivity of sebocytes. Increasing mite numbers influenced interleukin-8 secretion by these cells.

Bilateral lacrimal caruncle lesions.

A 65-year-old man was referred to our hospital for the treatment of a lesion on the medial lacrimal canthus of both eyes. He had a history of perinuclear anti-neutrophil cytoplasmic antibodies, i.e., pANCA-positive interstitial pneumonia. Orbital magnetic resonance imaging excluded space occupying lesions, and laboratory testing excluded thyroid-related diseases. The masses were excised, and histopathological examinations showed sebaceous gland hyperplasia and inflammatory changes around the gland. In addition, the specimen from the left eye showed a retention cyst possibly caused by an infection. It was also possible that the use of steroid was involved in the development of the lesions. A relationship between the ANCA and the lesions was not completely eliminated.

Beyond acne: Current aspects of sebaceous gland biology and function.

The sebaceous gland is most commonly found in association with a hair follicle. Its traditional function is the holocrine production of sebum, a complex mixture of lipids, cell debris, and other rather poorly characterized substances. Due to the gland's central role in acne pathogenesis, early research had focused on its lipogenic activity. Less studied aspects of the sebaceous gland, such as stem cell biology, the regulation of cellular differentiation by transcription factors, the significance of specific lipid fractions, the endocrine and specially the neuroendocrine role of the sebaceous gland, and its contribution to the innate immunity, the detoxification of the skin, and skin aging have only recently attracted the attention of researchers from different disciplines. Here, we summarize recent multidisciplinary progress in sebaceous gland research and discuss how sebaceous gland research may stimulate the development of novel therapeutic strategies targeting specific molecular pathways of the pathogenesis of skin diseases.

Specificity of granular IgM deposition in folliculosebaceous units and sweat gland apparatus in direct immunofluorescence (DIF) of lupus erythematosus.

BACKGROUND: Diagnosis of lupus erythematosus (LE) in direct immunofluorescence testing is based on the finding of positive immunofluorescence at the dermoepidermal junction (DEJ). OBJECTIVES: We sought to evaluate the sensitivity of IgM deposition at the DEJ and adnexal structures in the diagnosis of lupus erythematosus. METHODS: We conducted a retrospective study of 100 previously diagnosed cases of lupus erythematosus and 158 cases of other immune-mediated dermatosis. Deposition of IgG, IgM, IgA, and C3 at the DEJ, follicular units, and sweat glands were recorded. Presence or absence of adnexal structures was documented. The immunoreactant deposition was documented as linear, coarse granular, or stippled. RESULTS: The most frequently deposited immunoreactant in lupus erythematosus cases was IgM along the DEJ and stromal-epithelial junction of hair follicles and sweat glands. IgM deposition along the stromal-epithelial junction of hair follicles and sweat glands was strongly associated with a diagnosis of lupus erythematosus compared with other immune-mediated diseases collectively (P value < .001). The pattern of IgM in lupus and dermatomyositis is granular, in contrast to the linear deposition in the other disorders evaluated. LIMITATIONS: This was a retrospective study of archived material. CONCLUSION: Granular IgM deposition at the stromal-epithelial junction of cutaneous adnexal structures suggests a diagnosis of lupus erythematosus or dermatomyositis.

Neutrophilic sebaceous adenitis with intralobular Demodex mites: a case report and review of the literature.

A 61-year-old white man presented with a 1-week history of an asymptomatic erythematous, annular plaque with minimal scale limited to the nasal bridge. Histological examination showed a mixed infiltrate of lymphocytes and neutrophils within sebaceous glands. The clinical and histopathological presentation was consistent with a diagnosis of neutrophilic sebaceous adenitis. Several Demodex brevis mites were present deep within the affected sebaceous lobules. Demodex brevis mites are uncommon inhabitants of sebaceous glands of the nose, presenting more commonly on other body sites. The cause of neutrophilic sebaceous adenitis is unknown, but the presence of D. brevis in affected sebaceous glands in this case suggests a possible association.

Inherited Alopecia Areata in Mice as a Result of Adoptive Transfer of In Vivo Mitogen-Activated Splenic Cells to Females During Early Period of Gestation.

Postnatal morphogenesis of the skin and its appendages were studied in the progeny of mice subjected during early gestation period to adoptive transfer of in vivo activated splenic cells. Reversible focal hair loss was observed on postnatal day 17 in the progeny. The number of hair follicles and sebaceous glands, the level of extracellular matrix, and the content of cells in the skin, including mast cells in the derma, decreased, while neutrophils prevailed. Alopecia areata was found in the subsequent generations after mating of male and female progeny. Hence, stimulation of the maternal immune system during early pregnancy led to disorders of the pre- and postnatal morphogenesis of the skin in the progeny and to development of alopecia areata that became an inherited sign.

Antibodies get under the skin.

By inhibiting receptor-ligand interactions in sebaceous glands, antibodies may be able to treat certain skin conditions.

TRPV3 promotes sebocyte inflammation via transcriptional modulating TLR2 in acne.

Acne is a common chronic inflammatory disease of the pilosebaceous unit. Transient receptor potential vanilloid 3 (TRPV3) is an ion channel that is involved in inflammatory dermatosis development. However, the involvement of TRPV3 in acne-related inflammation remains unclear. Here, we used acne-like mice and human sebocytes to examine the role of TRPV3 in the development of acne. We found that TRPV3 expression increased in the skin lesions of Propionibacterium acnes (P. acnes)-injected acne-like mice and the facial sebaceous glands (SGs) of acne patients. TRPV3 promoted inflammatory cytokines and chemokines secretion in human sebocytes and led to neutrophil infiltration surrounding the SGs in acne lesions, further exacerbating sebaceous inflammation and participating in acne development. Mechanistically, TRPV3 enhanced TLR2 level by promoting transcriptional factor phosphorylated-FOS-like antigen-1 (p-FOSL1) expression and its binding to the TLR2 promoter, leading to TLR2 upregulation and downstream NF-κB signaling activation. Genetic or pharmacological inhibition of TRPV3 both alleviated acne-like skin inflammation in mice via the TLR2-NF-κB axis. Thus, our study revealed the critical role of TRPV3 in sebaceous inflammation and indicated its potential as an acne therapeutic target.

PGD2 induces eotaxin-3 via PPARγ from sebocytes: a possible pathogenesis of eosinophilic pustular folliculitis.

BACKGROUND: Eosinophilic pustular folliculitis (EPF) is a chronic intractable pruritic dermatosis characterized by massive eosinophil infiltrates involving the pilosebaceous units. Recently, EPF has been regarded as an important clinical marker of HIV infection, and its prevalence is increasing in number. The precise mechanism by which eosinophils infiltrate into the pilosebaceous units remains largely unknown. Given that indomethacin, a COX inhibitor, can be successfully used to treat patients with EPF, we can assume that COX metabolites such as prostaglandins (PGs) are involved in the etiology of EPF. OBJECTIVE: To determine the involvement of PGs in the pathogenesis of EPF. METHODS: We performed immunostaining for PG synthases in EPF skin lesions. We examined the effect of PGD(2) on induction of eotaxin, a chemoattractant for eosinophils, in human keratinocytes, fibroblasts, and sebocytes and sought to identify its responsible receptor. RESULTS: Hematopoietic PGD synthase was detected mainly in infiltrating inflammatory cells in EPF lesions, implying that PGD(2) was produced in the lesions. In addition, PGD(2) and its immediate metabolite 15-deoxy-Δ 12,14-PGJ(2) (15d-PGJ(2)) induced sebocytes to produce eotaxin-3 via peroxisome proliferator-activated receptor gamma. Consistent with the above findings, eotaxin-3 expression was immunohistochemically intensified in sebaceous glands of the EPF lesions. CONCLUSION: The PGD(2)/PGJ(2)-peroxisome proliferator-activated receptor gamma pathway induces eotaxin production from sebocytes, which may explain the massive eosinophil infiltrates observed around pilosebaceous units in EPF.

The pilosebaceous unit--a phthalate-induced pathway to skin sensitization.

Allergic contact dermatitis (ACD) is caused by low-molecular weight compounds called haptens. It has been shown that the potency of haptens can depend on the formulation in which they are applied on the skin. Specifically the sensitization potency of isothiocyanates, a group of haptens which can be released from e.g. adhesive tapes and neoprene materials, increases with the presence of phthalates; however, the underlying mechanisms are not clear. A better understanding of the mechanisms governing the potency of haptens is important, e.g. to improve the risk assessment and the formulation of chemicals in consumer products. In this study we have explored phthalate-induced effects on the sensitization potency, skin distribution, and reactivity of fluorescent model isothiocyanate haptens using non-invasive two-photon microscopy to provide new insights regarding vehicle effects in ACD. The data presented in this paper indicate that the sensitization potency of isothiocyanates increases when applied in combination with dibutylphthalate due to a specific uptake via the pilosebaceous units. The results highlight the importance of shunt pathways when evaluating the bioavailability of skin sensitizers. The findings also indicate that vehicle-dependent hapten reactivity towards stratum corneum proteins regulates the bioavailability, and thus the potency, of skin sensitizers.

KdPT, a tripeptide derivative of alpha-melanocyte-stimulating hormone, suppresses IL-1 beta-mediated cytokine expression and signaling in human sebocytes.

Acne is the most common inflammatory skin disease in which IL-1 plays a central role. Although alpha-melanocyte-stimulating hormone has immunomodulatory effects, its usefulness as an anti-inflammatory agent in acne is hampered owing to its lipid- and pigment-inducing effects via activation of melanocortin receptors (MC-Rs). We used the immortalized human sebocyte line SZ95 as an in vitro model to investigate the anti-inflammatory potential of KdPT, a tripeptide derivative of the C-terminal end of alpha-melanocyte-stimulating hormone. KdPT potently suppressed IL-1beta-induced IL-6 and IL-8 expression. Mechanistically, KdPT decreased IL-1beta-mediated IkappaBalpha degradation, reduced nuclear accumulation of p65, and attenuated DNA binding of NF-kappaB. Moreover, KdPT reduced IL-1beta-mediated generation of intracellular reactive oxygen species, which contributed to IL-1beta-mediated cytokine induction. KdPT also reduced cell surface binding of fluorochrome-labeled IL-1beta in SZ95 sebocytes. Analysis of the crystal structure of the complex between IL-1beta/IL-1R type I (IL-1RI), followed by computer modeling of KdPT and subsequent modeling of the peptide receptor complex with the crystal structure of IL-1RI via manual docking, further predicted that the tripeptide, through several H-bonds and one hydrophobic bond, interacts with the IL-1RI. Importantly, KdPT did not bind to MC-1Rs, as demonstrated by blocking experiments with a peptide analog of Agouti signaling protein and by binding assays using MC-1R-expressing B16 melanoma cells. Accordingly, KdPT failed to induce melanogenesis. Our data demonstrate a promising anti-inflammatory potential of KdPT and point toward novel future directions in the treatment of acne-as well as of various other IL-1-mediated inflammatory diseases-with this small molecule.

Epidermal Growth Factor Modulates Palmitic Acid-Induced Inflammatory and Lipid Signaling Pathways in SZ95 Sebocytes.

Epidermal growth factor (EGF) acts as a paracrine and autocrine mediator of cell proliferation and differentiation in various types of epithelial cells, such as sebocytes, which produce the lipid-rich sebum to moisturize the skin. However, sebum lipids via direct contact and by penetrating through the epidermis may have regulatory roles on epidermal and dermal cells as well. As EGF receptor (EGFR) is expressed throughout the proliferating and the lipid-producing layers of sebaceous glands (SGs) in healthy and acne-involved skin, we investigated the effect of EGF on SZ95 sebocytes and how it may alter the changes induced by palmitic acid (PA), a major sebum component with bioactive roles. We found that EGF is not only a potent stimulator of sebocyte proliferation, but also induces the secretion of interleukin (IL)6 and down-regulates the expression of genes involved in steroid and retinoid metabolism. Importantly, when applied in combination with PA, the PA-induced lipid accumulation was decreased and the cells secreted increased IL6 levels. Functional clustering of the differentially regulated genes in SZ95 sebocytes treated with EGF, PA or co-treated with EGF+PA further confirmed that EGF may be a potent inducer of hyperproliferative/inflammatory pathways (IL1 signaling), an effect being more pronounced in the presence of PA. However, while a group of inflammatory genes was up-regulated significantly in EGF+PA co-treated sebocytes, PA treatment in the absence of EGF, regulated genes only related to cell homeostasis. Meta-analysis of the gene expression profiles of whole acne tissue samples and EGF- and EGF+PA -treated SZ95 sebocytes showed that the EGF+PA co-activation of sebocytes may also have implications in disease. Altogether, our results reveal that PA-induced lipid accumulation and inflammation can be modulated by EGF in sebocytes, which also highlights the need for system biological approaches to better understand sebaceous (immuno)biology.

miR-146a modulates TLR1/2 and 4 induced inflammation and links it with proliferation and lipid production via the indirect regulation of GNG7 in human SZ95 sebocytes.

Activation of Toll-like receptors (TLR) 1/2 and 4 are central in inducing inflammation in sebocytes by regulating the expression of protein coding mRNAs, however the microRNA (miRNA) profile in response to TLR activation and thus the possible role of miRNAs in modulating sebocyte functions has not been elucidated. In this work we identified miR-146a to have the highest induction in the TLR1/2 and 4 activated SZ95 sebocytes and found that its increased levels led to the down-regulation of IL-8 secretion, decreased the chemoattractant potential and stimulated the proliferation of sebocytes. Assessing the gene expression profile of SZ95 sebocytes treated with a miR-146a inhibitor, the induction of GNG7 was one of the highest, while when cells were treated with a miR-146a mimic, the expression of GNG7 was down-regulated. These findings correlated with our in situ hybridization results, that compared with control, miR-146a showed an increased, while GNG7 a decreased expression in sebaceous glands of acne samples. Further studies revealed, that when inhibiting the levels of GNG7 in SZ95 sebocytes, cells increased their lipid content and decreased their proliferation. Our findings suggest, that miR-146a could be a potential player in acne pathogenesis by regulating inflammation, inducing proliferation and, through the indirect down-regulation of GNG7, promoting the lipid production of sebocytes.

Resveratrol ameliorates lipid accumulation and inflammation in human SZ95 sebocytes via the AMPK signaling pathways in vitro.

BACKGROUND: Acne vulgaris is a prevalent skin disease lacking effective and well-tolerated treatment. An earlier study indicated that resveratrol (RVT) has therapeutic effects in acne patients through unknown mechanisms. OBJECTIVES: To evaluate the effects of RVT on linoleic acid (LA)-induced lipogenesis and peptidoglycan (PGN)-induced inflammation in cultured SZ95 sebocytes in vitro, and to investigate the underlying mechanisms. METHODS: RNA-sequencing was used to analyze the whole transcriptome. Nile red staining was used to detect intracellular neutral lipids, whereas lipidomics was used to investigate changes in the lipid profile in sebocytes. Interleukin (IL)-1ß and IL-6 mRNA and protein levels were assessed through quantitative real-time PCR and Enzyme-linked immunosorbent assay, respectively. Western blot was used to evaluate the expression of lipogenesis-related proteins, the inflammatory signaling pathway, and the AMP-activated protein kinase (AMPK) pathway. Further, specific small interfering RNA (siRNA) was used to knockdown sirtuin-1 (SIRT1) expression. RESULTS: RVT inhibited the lipogenesis-related pathway and nuclear factor-kappa B (NF-κB) signaling pathway in SZ95 sebocytes. It also downregulated LA-induced lipogenesis, the expression of lipid-related proteins, and the contents of unsaturated fatty acids. Besides, RVT promoted SIRT1 expression and deacetylation of the NF-κB p65 subunit, thereby lowering IL-1ß and IL-6 secretion under PGN induction. Furthermore, pretreatment with AMPK inhibitor Compound C abolished RVT-mediated sebosuppressive and anti-inflammation effects. Meanwhile, SIRT1 silencing abrogated the anti-inflammatory potential of RVT. CONCLUSION: In human SZ95 sebocytes, RVT exhibits sebosuppressive and anti-inflammatory effects partially through the AMPK pathway, which may justify the role of RVT treatment in acne vulgaris.