Desmond J. Tobin

The cutaneous serotoninergic/melatoninergic system: securing a place under the sun

It was recently discovered that mammalian skin can produce serotonin and transform it into melatonin. Pathways for the biosynthesis and biodegradation of serotonin and melatonin have been characterized in human and rodent skin and in their major cellular populations. Moreover, receptors for serotonin and melatonin receptors are expressed in keratinocytes, melanocytes, and fibroblasts and these mediate phenotypic actions on cellular proliferation and differentiation. Melatonin exerts receptor‐independent effects, including activation of pathways protective of oxidative stress and the modification of cellular metabolism. While serotonin is known to have several roles in skin—e.g., pro‐edema, vasodilatory, proinflammatory, and pruritogenic—melatonin has been experimentally implicated in hair growth cycling, pigmentation physiology, and melanoma control. Thus, the widespread expression of a cutaneous seorotoninergic/melatoninergic system(s) indicates considerable selectivity of action to facilitate intra‐, auto‐, or paracrine mechanisms that define and influence skin function in a highly compartmentalized manner. Notably, the cutaneous melatoninergic system is organized to respond to continuous stimulation in contrast to the pineal gland, which (being insulated from the external environment) responds to discontinuous activation by the circadian clock. Overall, the cutaneous serotoninergic/melatoninergic system could counteract or buffer external (environmental) or internal stresses to preserve the biological integrity of the organ and to maintain its homeostasis.—Slominski, A. J., Wortsman, J., Tobin, D. J. The cutaneous serotoninergic/melatoninergic system: securing a place under the sun. FASEB J. 19, 176–194 (2005)

The human hair follicle immune system: cellular composition and immune privilege.

The immunology of the hair follicle, its relationship with the ‘skin immune system’ and its role in hair diseases remain biologically intriguing and clinically important. In this study, we analysed the immunoreactivity patterns of 15 immunodermatological markers to determine the cellular composition and immune privilege of the human hair follicle immune system in anagen VI (growth phase). The most prominent cells located in or around the hair follicle were Langerhans cells, CD4+ or CD8+ T cells, macrophages and mast cells, whereas B cells, natural killer cells and γδ T cells were found very rarely. Langerhans cells (CD1a+, major histocompatibility complex, MHC class II+), and T cells (CD4+ or CD8+) were predominantly distributed in the distal hair follicle epithelium, whereas macrophages (CD68+, MHC class II+) and mast cells (Giemsa+) were located in the perifollicular connective tissue sheath. Transmission electron microscopy confirmed low numbers of immune cells in the proximal hair follicle epithelium, and very few macrophages and Langerhans cells were seen in the dermal papilla. Melanophages were observed in the connective tissue sheath and dermal papilla. MHC class I (HLA‐A, ‐B, ‐C) and β2‐microglobulin immunoreactivity was found on most skin cells, but was substantially reduced on isthmus keratinocytes and virtually absent in the proximal hair follicle epithelium. Apart from the absence of Fas ligand immunoreactivity, the sharply reduced numbers of T cells and Langerhans cells, and the virtual absence of MHC class I expression all suggest that the anagen proximal hair follicle constitutes an area of immune privilege within the hair follicle immune system, whose collapse may be crucial for the pathogenesis of alopecia areata.

Serotoninergic and melatoninergic systems are fully expressed in human skin

We investigated the cutaneous expression of genes and enzymes responsible for the multistep conversion of tryptophan to serotonin and further to melatonin. Samples tested were human skin, normal and pathologic (basal cell carcinoma and melanoma), cultured normal epidermal and follicular melanocytes, melanoma cell lines, normal neonatal and adult epidermal and follicular keratinocytes, squamous cell carcinoma cells, and fibroblasts from dermis and follicular papilla. The majority of the samples showed simultaneous expression of the genes for tryptophan hydroxylase, arylalkylamine N‐acetyltransferase (AANAT), and hydroxyindole‐Omethyltransferase (HIOMT). The products of AANAT activity were identified by RP‐HPLC with fluorimetric detection in human skin and in cultured normal and malignant melanocytes and immortalized keratinocytes; HIOMT activity was detected in human skin, keratinocytes, and melanoma cells. N‐acetylserotonin (NAS) was detected by RP‐HPLC in human skin extracts. NAS identity was confirmed further by LC/MS in keratinocytes. In conclusion, we provide evidence that the human skin expresses intrinsic serotonin and melatonin biosynthetic pathways.

On the role of melatonin in skin physiology and pathology

Melatonin has been experimentally implicated in skin functions such as hair growth cycling, fur pigmentation, and melanoma control, and melatonin receptors are expressed in several skin cells including normal and malignant keratinocytes, melanocytes, and fibroblasts. Melatonin is also able to suppress ultraviolet (U)-induced damage to skin cells and shows strong antioxidant activity in Uexposed cells. Moreover, we recently uncovered expression in the skin of the biochemical machinery involved in the sequential transformation of l-tryptophan to serotonin and melatonin. Existence of the biosynthetic pathway was confirmed by detection of the corresponding genes and proteins with actual demonstration of enzymatic activities for tryptophan hydroxylase, serotonin N-acetyl-transferase, and hydroxyindole-O-methyltransferase in extracts from skin and skin cells. Initial evidence for in vivo synthesis of melatonin and its metabolism was obtained in hamster skin organ culture and in one melanoma line. Therefore, we propose that melatonin (synthesized locally or delivered topically)could counteract or buffer external (environmental)or internal stresses to preserve the biological integrity of the organ and to maintain its homeostasis. Furthermore, melatonin could have a role in protection against solar radiation or even in the management of skin diseases.

Functional activity of serotoninergic and melatoninergic systems expressed in the skin.

We tested the expression of genes coding receptors of a cutaneous serotoninergic/melatoninergic system in whole human skin and in normal and pathologic cultured skin cells. Evaluation of serotonin (5HT), melatonin (MT), and melatonin‐related receptors (MRR) showed expression of the isoforms 5HT2B, 5HT7, and MT1 genes in almost all the tested samples. Expression of other isoforms was less prevalent; 5HT2C, MRR, and MT2 were rarely detected. We also found novel isoforms for MT2, MRR, and 5HT2B and documented the process of RNA editing for 5HT2C. Testing for functional activity of these receptors with serotonin and melatonin (10−14 to 10−10 M) showed variable effects depending on cell type and culture conditions. Thus, serotonin stimulated proliferation of melanocytes in medium deprived of growth factors, while inhibiting cell growth in the presence of growth factors. Melatonin inhibited both apoptosis of HaCaT keratinocytes incubated in serum‐free media, and proliferation of cells cultured in medium supplemented with serum. Melatonin also increased the numbers of viable fibroblasts incubated in serum free medium. N‐acetylserotonin (NAS) and 5 methoxytryptamine (5MTT) were generally without effect on cell proliferation, with the exception of an inhibition of melanocyte proliferation at the higher 5MTT concentration of 10−10 M. Thus, skin cells represent a true target for the products of the serotoninergic/melatoninergic cutaneous pathway with their actions modulating cell proliferation or viability.

Melatonin in the skin: synthesis, metabolism and functions

Melatonin, a ubiquitous methoxyindole, is produced by and metabolized in the skin. Melatonin affects skin functions and structures through actions mediated by cell-surface and putative-nuclear receptors expressed in skin cells. Melatonin has both receptor-dependent and receptor-independent effects that protect against oxidative stress and can attenuate ultraviolet radiation-induced damage. The widespread expression and pleiotropic activity of the cutaneous melatoninergic system provides for a high level of cell-specific selectivity. Moreover, intra-, auto- and para-crine mechanisms equip this system with exquisite functional selectivity. The properties of endogenous melatonin suggest that this molecule is an important effector of stress responses in the skin. In this way, melatonin actions may counteract or buffer both environmental and endogenous stressors to maintain skin integrity.

Key Role of CRF in the Skin Stress Response System

The discovery of corticotropin-releasing factor (CRF) or CRH defining the upper regulatory arm of the hypothalamic-pituitary-adrenal (HPA) axis, along with the identification of the corresponding receptors (CRFRs 1 and 2), represents a milestone in our understanding of central mechanisms regulating body and local homeostasis. We focused on the CRF-led signaling systems in the skin and offer a model for regulation of peripheral homeostasis based on the interaction of CRF and the structurally related urocortins with corresponding receptors and the resulting direct or indirect phenotypic effects that include regulation of epidermal barrier function, skin immune, pigmentary, adnexal, and dermal functions necessary to maintain local and systemic homeostasis. The regulatory modes of action include the classical CRF-led cutaneous equivalent of the central HPA axis, the expression and function of CRF and related peptides, and the stimulation of pro-opiomelanocortin peptides or cytokines. The key regulatory role is assigned to the CRFR-1α receptor, with other isoforms having modulatory effects. CRF can be released from sensory nerves and immune cells in response to emotional and environmental stressors. The expression sequence of peptides includes urocortin/CRF→pro-opiomelanocortin→ACTH, MSH, and β-endorphin. Expression of these peptides and of CRFR-1α is environmentally regulated, and their dysfunction can lead to skin and systemic diseases. Environmentally stressed skin can activate both the central and local HPA axis through either sensory nerves or humoral factors to turn on homeostatic responses counteracting cutaneous and systemic environmental damage. CRF and CRFR-1 may constitute novel targets through the use of specific agonists or antagonists, especially for therapy of skin diseases that worsen with stress, such as atopic dermatitis and psoriasis.

Dilated cardiomyopathy in mice deficient for the lysosomal cysteine peptidase cathepsin L

Dilated cardiomyopathy is a frequent cause of heart failure and is associated with high mortality. Progressive remodeling of the myocardium leads to increased dimensions of heart chambers. The role of intracellular proteolysis in the progressive remodeling that underlies dilated cardiomyopathy has not received much attention yet. Here, we report that the lysosomal cysteine peptidase cathepsin L (CTSL) is critical for cardiac morphology and function. One-year-old CTSL-deficient mice show significant ventricular and atrial enlargement that is associated with a comparatively small increase in relative heart weight. Interstitial fibrosis and pleomorphic nuclei were found in the myocardium of the knockout mice. By electron microscopy, CTSL-deficient cardiomyocytes contained multiple large and apparently fused lysosomes characterized by storage of electron-dense heterogeneous material. Accordingly, the assessment of left ventricular function by echocardiography revealed severely impaired myocardial contraction in the CTSL-deficient mice. In addition, echocardiographic and electrocardiographic findings to some degree point to left ventricular hypertrophy that most likely represents an adaptive response to cardiac impairment. The histomorphological and functional alterations of CTSL-deficient hearts result in valve insufficiencies. Furthermore, abnormal heart rhythms, like supraventricular tachycardia, ventricular extrasystoles, and first-degree atrioventricular block, were detected in the CTSL-deficient mice.

What are melanocytes really doing all day long...

Abstract:  Everyone knows and seems to agree that melanocytes are there to generate melanin – an intriguing, but underestimated multipurpose molecule that is capable of doing far more than providing pigment and UV protection to skin ( 1 ). What about the cell that generates melanin, then? Is this dendritic, neural crest‐derived cell still serving useful (or even important) functions when no‐one looks at the pigmentation of our skin and its appendages and when there is essentially no UV exposure? In other words, what do epidermal and hair follicle melanocytes do in their spare time – at night, under your bedcover? How much of the full portfolio of physiological melanocyte functions in mammalian skin has really been elucidated already? Does the presence or absence of melanoctyes matter for normal epidermal and/or hair follicle functions (beyond pigmentation and UV protection), and for skin immune responses? Do melanocytes even deserve as much credit for UV protection as conventional wisdom attributes to them? In which interactions do these promiscuous cells engage with their immediate epithelial environment and who is controlling whom? What lessons might be distilled from looking at lower vertebrate melanophores and at extracutaneous melanocytes in the endeavour to reveal the ‘secret identity’ of melanocytes? The current Controversies feature explores these far too infrequently posed, biologically and clinically important questions. Complementing a companion viewpoint essay on malignant melanocytes ( 2 ), this critical re‐examination of melanocyte biology provides a cornucopia of old, but under‐appreciated concepts and novel ideas on the slowly emerging complexity of physiological melanocyte functions, and delineates important, thought‐provoking questions that remain to be definitively answered by future research.

NF-κB transmits Eda A1/EdaR signalling to activate Shh and cyclin D1 expression, and controls post-initiation hair placode down growth

A novel function of NF-κB in the development of most ectodermal appendages, including two types of murine pelage hair follicles, was detected in a mouse model with suppressed NF-κB activity (cIκBαΔN). However, the developmental processes regulated by NF-κB in hair follicles has remained unknown. Furthermore, the similarity between the phenotypes of cIκBAΔN mice and mice deficient in Eda A1 (tabby) or its receptor EdaR (downless) raised the issue of whether in vivo NF-κB regulates or is regulated by these novel TNF family members. We now demonstrate that epidermal NF-κB activity is first observed in placodes of primary guard hair follicles at day E14.5, and that in vivo NF-κB signalling is activated downstream of Eda A1 and EdaR. Importantly, ectopic signals which activate NF-κB can also stimulate guard hair placode formation, suggesting a crucial role for NF-κB in placode development. In downless and cIκBαΔN mice, placodes start to develop, but rapidly abort in the absence of EdaR/NF-κB signalling. We show that NF-κB activation is essential for induction of Shh and cyclin D1 expression and subsequent placode down growth. However, cyclin D1 induction appears to be indirectly regulated by NF-κB, probably via Shh and Wnt. The strongly decreased number of hair follicles observed in cIκBαΔN mice compared with tabby mice, indicates that additional signals, such as TROY, must regulate NF-κB activity in specific hair follicle subtypes.