Hair morphology in androgenetic alopecia: sonographic and electron microscopic studies.

OBJECTIVES: To assess hair morphology in androgenetic alopecia on sonography and electron microscopy. METHODS: A prospective study was performed in 33 patients with androgenetic alopecia and 10 unaffected control participants. In vivo sonography of the hair follicles of the scalp and in vitro sonography and electron microscopy of the hair shafts were performed according to a standardized protocol that included analysis of the right frontal and occipital regions. The upper frequency limit of the ultrasound probes ranged between 15 and 18 MHz. RESULTS: Scalp hair follicles and hair shafts were recognizable on sonography in all cases. Hair follicles in alopecia cases had significantly lower depths (P < .05). The hair shafts in alopecia also had a different distribution of their laminar pattern on in vitro sonography, with a greater presence of mixed (trilaminar and bilaminar) and solely bilaminar tracts in comparison with the controls (mostly trilaminar). On electron microscopy, the alopecia hair tracts showed irregularities and commonly a "melted candle" appearance of the cuticle. CONCLUSIONS: Sonography and electron microscopy uncover distinct abnormalities in the morphology of hair in androgenetic alopecia, which may potentially support the diagnosis and management of this common condition.

Sonography of acne vulgaris.

OBJECTIVES: The purpose of this study was to assess the sonographic morphology of the clinical and subclinical pathology of facial acne vulgaris. METHODS: We studied patients with facial acne vulgaris diagnosed by certified dermatologists, and using a standardized protocol for sonographic examinations, we sequentially described the sonographic pathomorphologic characteristics. Lesions of particular interest to the referring clinician were also analyzed separately. Additionally, acne involvement was staged clinically and sonographically (SOS-Acne) using morphologic definitions of the relevant lesions and predefined scoring systems for gradation of the severity of acne lesions. RESULTS: A total of 245 acne lesions in 20 consecutive patients were studied. Sonographic abnormalities consisted of pseudocysts, folliculitis, fistulas, and calcinosis. Most conditions were subclinical and mostly due to lesion extensions deep into the dermis and hypodermis (52% of pseudocysts and 68% of fistulas). The statistical concordance between acne severity scores assigned by two separate clinicians was strong (κ = 0.8020), but the corresponding sonographic scores generally showed more severe and clinically occult involvement. CONCLUSIONS: Facial acne vulgaris often involves deeper tissues, beyond the reach of the spatially restricted clinical examination; these subclinical conditions can be detected and defined with sonography. Additionally, acne vulgaris is amenable to sonographic scoring.

Ultrasound in-depth characterization and staging of hidradenitis suppurativa.

BACKGROUND: The clinical diagnosis of fistulous tracts and recurrent fluid collections in hidradenitis suppurativa (HS) may be complex. Information on subclinical involvement and grading of severity may improve management. OBJECTIVE: To study HS lesions and evaluate the relevance of adding ultrasound (US) to the clinical examination. METHODS AND MATERIALS: We reviewed the sonograms of consecutive patients with HS from January 1, 2010 to May 31, 2012. The abnormalities observed in the US examinations were organized, classified, and integrated into a clinical-sonographic scoring system (SOS-HS), to stage the disease. RESULTS: Thirty-four patients with HS with a total of 142 lesional areas were evaluated. US findings included subclinical fluid collections in 76.4% of the patients, fistulous tracts in 29.4%, dermal pseudocysts in 70.6%, and widening of the hair follicles in 100%. Concordance with the clinical HS scoring performed by dermatologists showed a significant fair agreement (k = 0.30; p < .001); concordance of SOS-HS with clinical scores was acceptable but significantly lower (k = 0.27; p = .02) because clinical scores consistently underestimated disease stage and severity. CONCLUSION: US examination of HS lesions provides anatomic information that is clinically unavailable. HS is possible to stage sonographically.

Sonographic outcomes of cosmetic procedures.

OBJECTIVE: The purpose of this article is to review the sonographic outcomes of common cosmetic and plastic surgery procedures, taking advantage of recent developments in the field of ultrasound that opened its broad application to dermatologic practice. CONCLUSION: Because cosmetic procedures have increased dramatically in frequency, some procedures are being performed by unauthorized personnel and some agents are being used that are not approved by certifying institutions, leading to complications. Thus, documentation of these procedures is an important proposition.

Activity assessment in morphea using color Doppler ultrasound.

BACKGROUND: Morphea (circumcripted cutaneous scleroderma) can be difficult to assess for lesion activity. Because variable-frequency ultrasound with color Doppler provides details of skin morphology and function, it may help in the categorization of morphea. OBJECTIVE: We sought to evaluate color Doppler ultrasound as a probing tool for assessing activity in morphea lesions. METHODS: Consecutive patients with cutaneous morphea referred by dermatologists were studied with color Doppler ultrasound, and the assessment of lesion activity was compared with histologic findings. Normal skin controls were obtained by performing ultrasound scans of healthy subjects or of unaffected areas of the patients themselves. Measurements included cutaneous layer thickness, relative echogenicity, and blood flow with peak systolic velocity. Ultrasound sensitivity and specificity were determined for each phase of morphea activity and the results correlated with histology. RESULTS: Fifty-one patients had a total of 104 morphea lesions. Of the lesions, 20% were active, 22% were atrophic, and 58% were inactive. Five of the patients had the Parry-Romberg syndrome with ipsilateral parotid gland inflammatory involvement, and one had an asymptomatic but sonographically active morphea lesion. Sensitivity and specificity for ultrasound diagnosis were 100% and 98.8%, respectively. The most accurate sonographic signs of lesion activity were increased subcutaneous tissue echogenicity and increased cutaneous blood flow (sensitivity and specificity 100% and 100% for each one). LIMITATIONS: Ultrasound cannot define lesions less than 0.1-mm deep. CONCLUSIONS: The morphologic and functional data obtained noninvasively and in real time with color Doppler ultrasound provide new insight into the pathogenesis of morphea. The technique represents a useful counterpart to histologic examination for the assessment of lesion activity.

Clinical usefulness of variable-frequency ultrasound in localized lesions of the skin.

BACKGROUND: High variable-frequency ultrasound is a recently available technique capable of clearly defining skin layers and deeper structures that also provides local perfusion patterns obtained in real time. OBJECTIVES: The aim of the study was to assess the performance of variable-frequency ultrasound in the evaluation of skin lesions. METHODS: We performed a retrospective study of 4338 skin ultrasound examinations in predominantly localized skin lesions, and in a group of 130 healthy controls. We determined ultrasound sensitivity, specificity, and statistical level of certainty, and compared ultrasound diagnoses with clinical diagnoses. RESULTS: Referring diagnosis was correct in 73% of the lesions, and addition of ultrasound increased correctness to 97% (P < .001 for the difference). Ultrasound overall sensitivity was 99%, specificity was 100%, and statistical diagnostic certainty was 99% LIMITATIONS: Ultrasound in its current version cannot detect lesions that are epidermal only or that measure less than 0.1 mm in depth. CONCLUSIONS: Ultrasound is a reliable adjuvant for the accurate and precise diagnosis of skin lesions.

Benign tumors and pseudotumors of the nail: a novel application of sonography.

OBJECTIVE: The purpose of this study was to determine the scope of high-resolution sonography in the detection of benign tumors and pseudotumors of the nail unit. METHODS: We performed a retrospective study of the sonographic findings in 103 consecutive patients with benign tumors and pseudotumors of the nail that were medically derived and confirmed histologically. Statistical analysis (Student t test) was performed comparing clinical and sonographic diagnoses. RESULTS: Common benign tumors and pseudotumors of the nail can be detected on sonography, and they present different sonographic morphologic characteristics. According to origin, the lesions were considered ungual in 73% (n = 75) and periungual in 27% (n = 28) of the cases. Sonography showed their nature (solid or cystic), location, and extension as well as regional blood flow. In 35% of the cases, the use of sonography modified the clinical diagnosis, although the detailed anatomic information provided by sonography was useful in the planning of surgery in all cases. The addition of sonography was significant (P < .001) for the diagnosis of subungual exostosis and granulomas in comparison to clinical diagnosis. CONCLUSIONS: Sonography is a noninvasive imaging method that can reliably detect common benign tumors and pseudotumors of the nail and provide precise data about their characteristics. This imaging modality can support diagnosis and surgery and can allow a better definition and improvement of the cosmetic outcome of the treatment.

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.

Metabolism of melatonin by cytochrome P450s in rat liver mitochondria and microsomes.

In the present study we provide direct evidence for the involvement of rat microsomal cytochrome P450s in melatonin O-demethylation and hydroxylation at two different positions: 2 and 6, as well as generation of N(1)-acetyl-N(2)-formyl-5-methoxy-kynuramine (AFMK) and two unknown products. Moreover, we found that mitochondrial cytochrome P450s also converts melatonin into AFMK, N-acetylserotonin, 2-hydroxymelatonin, 6-hydroxymelatonin and the same two unknown products. Eadie-Hofstee plots for 6-hydroxylation and O-demethylation reactions were curvilinear for all tested fractions, suggestive of involvement of at least two components, one with a high affinity and low capacity, and another with a low affinity and high capacity. Mitochondrial cytochrome P450s exhibited higher affinity (suggesting lower K(m) value) and higher V(max) for melatonin 6-hydroxylation and O-demethylation for both high-affinity and low-affinity components as compared with microsomal enzymes. The intrinsic clearance for melatonin hydroxylation by high- and low-affinity components displayed the highest values in all tested fractions, indicating that both mitochondrial and microsomal cytochrome P450s metabolize melatonin principally by 6-hydroxylation, with O-demethylation representing a minor metabolic pathway.

Skin as an endocrine organ: implications for its function.

Described as the body's largest organ, the skin is strategically located at the interface with the external environment where it has evolved to detect, integrate and respond to a diverse range of stressors. A flurry of recent findings has established the skin as an important peripheral (neuro)endocrine organ that is tightly networked to central stress axes. This capability is contributing to the maintenance of body homeostasis, and in this way could be harnessed for therapeutic strategies.

Molecular cloning and initial characterization of African green monkey (Cercopithecus aethiops) corticotropin releasing factor receptor type 1 (CRF1) from COS-7 cells.

We report the expression of endogenous CRF1 in COS-7 cells (African green monkey origin). Cloning of the coding region of CRF1 gene identified three alternatively spliced isoforms with nucleotide and predicted amino acid sequences corresponding to the membrane bound alpha and c and soluble e isoforms. DNA sequencing of the main isoform CRF1alpha showed homologies of 99%, 97% and 91% with the rhesus monkey, human and rodent genes, respectively; the deduced protein sequence differed in only one amino acid with rhesus monkey and human. Western blot analysis with antibodies against human CRF1 demonstrated immunoreactive proteins with MW of 37, 52, 70 and 80-85 in crude membrane or cytoplasm preparation; two additional species of 40 and 60 kDa were detected only in the cytoplasmic fraction. On immunocytochemistry CRF1 was localized to both the cell surface and intracellularly. The receptor was functional, e.g., addition of CRF to COS-7 cells inhibited cell proliferation and stimulated release of arachidonic acid; nevertheless, it was poorly coupled to cAMP production (its stimulation was minimal in native cells). In conclusion, COS cells that are routinely used for the study of transfected CRF receptors do express endogenous CRF1 mRNA with splicing behavior similar to that reported in human and rodent cells, and translated into functional CRF1 receptors.

Differential expression of HPA axis homolog in the skin.

Human skin expresses elements of the hypothalamo-pituitary-adrenal (HPA) axis including pro-opiomelanocortin (POMC), corticotropin releasing hormone (CRH), the CRH receptor-1 (CRH-R1), key enzymes of corticosteroid synthesis and synthesizes glucocorticoids. Expression of these elements is organized in functional, cell type-specific regulatory loops, which imitate the signaling hierarchy of the HPA axis. In melanocytes and fibroblasts CRH-induced CRH-R1 stimulation upregulates POMC expression and production of ACTH through activation of cAMP dependent pathway(s). Melanocytes respond with enhanced production of cortisol and corticosterone, which is dependent on POMC activity. Fibroblasts respond to CRH and ACTH with enhanced production of corticosterone, but not cortisol, which is produced constitutively. Organ-cultured human scalp hair follicles also show a fully functional HPA axis equivalent, including cortisol synthesis and secretion and negative feedback regulation by cortisol on CRH expression. Thus, differential, CRH-driven responses of defined cutaneous cell populations reproduce key features of the central HPA axis at the tissue/single cell levels.

Constitutive and UV-induced metabolism of melatonin in keratinocytes and cell-free systems.

Melatonin, which can be produced in the skin, exerts a protective effect against damage induced by UV radiation (UVR). We have investigated the effect of UVB, the most damaging component of UVR, on melatonin metabolism in HaCaT keratinocytes and in a cell-free system. Four metabolites were identified by HPLC and LC-MS: 6-hydroxymelatonin, N1-acetyl-N2-formyl-5-methoxykynuramine (AFMK), 2-hydroxymelatonin (the main intermediate between melatonin and AFMK), and 4-hydroxymelatonin. Concentrations of these photoproducts were directly proportional to UVR-dose and to melatonin substrate content, and their accumulation was time-dependent. The UVR-dependent increase of AFMK and 2-hydroxymelatonin was also detected in keratinocytes, where it was accompanied by simultaneous consumption of intracellular melatonin. Of note, melatonin and its two major metabolites, 2-hydroxymelatonin and AFMK, were also detected in untreated keratinocytes, neither irradiated nor preincubated with melatonin. Thus, intracellular melatonin metabolism is enhanced under exposure to UVR. The additional biological activity of these individual melatonin metabolites increases the spectrum of potential actions of the recently identified cutaneous melatoninergic system.

Characterization of a ultraviolet B-induced corticotropin-releasing hormone-proopiomelanocortin system in human melanocytes.

CRH, the main regulator of the systemic response to stress, is also expressed in the skin where it is incorporated into a local homolog of the hypothalamic-pituitary-adrenal axis. To investigate the mechanisms of the induction of the CRH-proopiomelanocortin (POMC) response in human melanocytes, we used UVB as an epidermal-specific stressor. Human normal melanocytes cultured in vitro were irradiated with graded doses of UVB, and the CRH-POMC responses were measured in cell extracts and/or supernatants. UVB stimulated the CRH promoter, the CRH mRNA expression, and peptide release. The UVB-induced stimulation of the CRH promoter was suppressed by pharmacological inhibitors of protein kinase A or by plasmid overexpressing a dominant mutant cAMP response element (CRE)-binding protein (CREB). UVB also stimulated phosphorylation of CREB, binding of phosphorylated CREB to CRE sites in the CRH promoter, and activity of the reporter gene construct driven by consensus CRE sites. Mutation in the CRE site in the CRH promoter rendered the corresponding reporter gene construct less responsive to UVB in both normal and malignant melanocytes. In addition to CRH effects, UVB activated the POMC promoter, POMC mRNA expression, and ACTH release, whereas an antagonist of the CRH receptor 1 abrogated the UVB-stimulated induction of POMC. In conclusion, UVB induces CRH production in human melanocytes through stimulation of the protein kinase A pathway, with sequential involvement of CRH-CRH receptor 1 in the stimulation of POMC expression.

An alternative pathway of vitamin D metabolism. Cytochrome P450scc (CYP11A1)-mediated conversion to 20-hydroxyvitamin D2 and 17,20-dihydroxyvitamin D2.

We report an alternative, hydroxylating pathway for the metabolism of vitamin D2 in a cytochrome P450 side chain cleavage (P450scc; CYP11A1) reconstituted system. NMR analyses identified solely 20-hydroxyvitamin D2 and 17,20-dihydroxyvitamin D2 derivatives. 20-Hydroxyvitamin D2 was produced at a rate of 0.34 mol x min(-1) x mol(-1) P450scc, and 17,20-dihydroxyvitamin D2 was produced at a rate of 0.13 mol x min(-1) x mol(-1). In adrenal mitochondria, vitamin D2 was metabolized to six monohydroxy products. Nevertheless, aminoglutethimide (a P450scc inhibitor) inhibited this adrenal metabolite formation. Initial testing of metabolites for biological activity showed that, similar to vitamin D2, 20-hydroxyvitamin D2 and 17,20-dihydroxyvitamin D2 inhibited DNA synthesis in human epidermal HaCaT keratinocytes, although to a greater degree. 17,20-Dihydroxyvitamin D2 stimulated transcriptional activity of the involucrin promoter, again to a significantly greater extent than vitamin D2, while the effect of 20-hydroxyvitamin D2 was statistically insignificant. Thus, P450scc can metabolize vitamin D2 to generate novel products, with intrinsic biological activity (at least in keratinocytes).

Corticotropin releasing hormone and the skin.

Cotricotropin-releasing hormone (CRH) and related peptides are produced in skin that is dependent on species and anatomical location. Local peptide production is regulated by ultraviolet radiation (UVR), glucocorticoids and phase of the hair cycle. The skin also expresses the corresponding receptors (CRH-R1 and CRH-R2), with CRH-R1 being the major receptor in humans. CRH-R1 is expressed in epidermal and dermal compartments, and CRH-R2 predominantly in dermal structures. The gene coding for CRH-R1 generates multiple isoforms through a process modulated by UVR, cyclic adenosine monophosphate (cAMP) and phorbol 12-myristate 13-acetate. The phenotypic effects of CRH in human skin cells are largely mediated by CRH-R1alpha through increases in concentrations of cAMP, inositol triphosphate (IP3), or Ca2+ with subsequent activation of protein kinases A (PKA) and C (PKC) dependent pathways. CRH also modulates the activity of nuclear factor of kappa light polypeptide gene enhancer in B-cells (NF-kappaB), activator protein 1 (AP-1) and cAMP responsive element binding protein (CREB). The cellular functions affected by CRH depend on cell type and nutritional status and include modulation of differentiation program(s), proliferation, viability and immune activity. The accumulated evidence indicates that cutaneous CRH is also a component of a local structure organized similarly to the hypothalamo-pituitary-adrenal axis.