bluuebunny/crossref_metadata_2025 huggingface dataset:
Substance P‐induction of endothelial nitric oxide synthase expression on human endothelial cells: modulation of this neurogenic inflammation by an Avena Rhealba® oatmeal extract
Authors: Aries M. F. , Vaissière C. , Charvéron M.
Published: August 2010

During cutaneous inflammatory diseases, the neuro‐immuno‐cutaneous system is impaired. In atopic dermatitis pathology, a perturbation of neuromediators release such as substance P (SP), calcitonin gene‐related peptide (CGRP) and bradykinin (BK) could be in part involved in the neurogenic inflammatory state; both of these peptides identified in human skin are potent inducers of vasodilatation, may induce pruritus and could mediate their effects via nitric oxide (NO). NO, considered as a major intra/intercellular messenger, is generated by NO synthase (NOS) enzymes identified in several cell types in the skin. NO displaying vasodilator properties are constitutively released and can also be synthesized in response to inflammatory mediators such as SP neuropeptide. The aim of the present study was first to determine whether SP, CGRP and BK were able to stimulate NO release from human endothelial cells (HECs); the second objective was to induce neurogenic inflammation on HECs with SP (10–100 pm) and to evaluate the activity of Avena Rhealba® oatmeal – Roasting extract – (0.001–0.005%) on endothelial NOS (eNOS) mRNA expression by RT‐PCR. Avena Rhealba® significantly inhibits endothelial cells substance P‐induced expression of eNOS. Our results demonstrate the regulator properties of Avena Rhealba® with respect to neurogenic inflammatory response showing therefore a real interest of Avena Rhealba® in atopic dermatitis inflammatory pathology.
Regulation of galanin gene expression in human keratinocytes
Authors: Moritz K. , Voglas E. , Sperl W. , Bauer J. W. , Kofler B.
Published: August 2010

Recently, we have detected abundant expression of specific immunoreactivity for galanin (GAL), a 29–30 amino acid neuropeptide, in the epidermis and in sweat glands in a non‐neuronal distribution and to a lesser extent in nerve fibre bundles and smooth muscle of blood vessels. In order to uncover a possible function of GAL in human epidermis, we investigated the regulation of GAL mRNA expression in cultured primary human keratinocytes (KCs). First, we determined whether PMA, a protein kinase C activator which is known to be a potent upregulator of GAL gene expression in chromaffin cells, has similar effects in KCs. Stimulation of human cultured primary KC with PMA resulted in a sixfold increase of GAL mRNA. Furthermore, we exposed KCs to a combination of the proinflammatory cytokines IL‐1β and TNF‐α which resulted in a twofold induction of GAL mRNA in comparison with a sixfold induction of ICAM‐1. Most interestingly, capsaicine, which usually leads to a secretion of neuropeptides from nerves endings, did not induce the secretion of GAL of KCs. In contrast, constitutive GAL mRNA expression and peptide secretion were reduced twofold upon treatment of KC with 10 µm capsaicine for 20 min and 24 h, respectively. To our knowledge, this is the first report on a direct regulatory effect by capsaicine on neuropeptide mRNA expression in non‐neuronal cells.
The neuropeptide PACAP upregulates expression and release of cytokines and cell adhesion molecules in human microvascular endothelial cells via VPAC type 1 receptor
Authors: Steinhoff A. , Grevelhörster A. , Schmidt W. E. , Luger T. A. , Steinhoff M.
Published: August 2010

Pituitary adenylate cyclase‐activating peptide (PACAP) and vasoactive intestinal peptide (VIP) belong to the same superfamily of neuropeptides which exert their effects by activating G‐protein‐coupled receptors defined as PACAP. So far, three receptor subtypes exist (PAC1R, VPACR‐1 and VPACR‐2). Because, PACAP appears to play a crucial role in cutaneous inflammation and vasoregulation, we examined the expression and biological effects of this peptide in primary human dermal microvascular endothelial cells (HDMECs). We detected the expression of PACAP and VPAC type 1 receptor at RNA and protein level by RT‐PCR and immunohistochemistry, indicating an autocrine regulatory mechanism. cAMP assays revealed VPAC1R to be functional in these cells. RT‐PCR showed upregulation of IL‐8 in a time‐ and dose‐dependent manner. ELISA experiments confirmed release of IL‐8 by HDMEC cells. We also investigated cell adhesion molecule expression after stimulation with PACAP. ICAM‐1 mRNA was upregulated at 3 and 6 h after treatment with PACAP, while VCAM was only upregulated maximally at 6 h after PACAP stimulation, indicating the regulation of cell adhesion molecule expression in human dermal endothelial cells via VPAC1R. Immunoreactivity for VPAC‐1R was enhanced in microvascular endothelial cells of patients with atopic dermatitis and urticaria, indicating upregulation of this receptor in endothelial cells during cutaneous inflammation. In summary, VIP and PACAP may play an important role in cutaneous neurogenic inflammation by activating VPAC‐1R on dermal microvascular endothelial cells.
Identification of novel genes regulated by α‐melanocyte‐stimulating hormone in murine bone marrow‐derived dendritic cells
Authors: Brzoska T. , Ehrchen J. , Li Z. , Luger T. A. , Böhm M.
Published: August 2010

Many strains of evidence indicate that α‐melanocyte‐stimulating hormone (α‐MSH) elicits its immunomodulatory activity via binding to melanocortin receptors (MC‐Rs) expressed on monocytes and dendritic cells. In order to identify novel target genes regulated by α‐MSH in these cells, we prepared bone marrow‐derived dendritic cell precursors from BALB/c mice and treated them with GM‐CSF and IL‐4 for 6 days. The MC‐R profile on these immature dendritic cells was first determined by quantitative RT‐PCR. Both transcripts for MC‐1R and MC‐5R were detected in these cells. Cells were subsequently stimulated with dinitrobenzene sulfonic acid (DNBS), α‐MSH or both substances for 2 or 16 h. After RNA preparation, cDNA synthesis and in vitro transcripton hybridization of biotinylated cRNA samples was performed on MG U74A Affymetrix gene chips. Data evaluation, cleansing, extraction and analysis of the more than 12 000 cloned genes and expressed sequence tags were performed using the GENE DATA ANALYST vs. 1 Expressionist software. Filter criteria included a minimum threshold of 100, normalization by the logarithmic mean and a quality setting of P < 0.04. Changes with a change factor of >2 were regarded as significant. As expected, stimulation with DNBS resulted in induction or upregulation of genes encoding proinflammatory cytokines, growth factors, signal transduction intermediates and transcription factors. Treatment with α‐MSH blocked the DNBS‐driven upregulation of several known genes such as IL‐1 or CD86. On the other hand, α‐MSH modulated the expression of several novel genes implicated in immunomodulation, e.g. IL‐1β converting enzyme, IFN‐γ receptor, FK506‐binding proteins or several neuropeptides and their receptors. These data indicate novel molecular targets by which α‐MSH exerts its immunomodulatory activities in immunocompetent cells.
The Langerhans' cell‐like cell lines XS52 and XS106 express mRNA for ciliary neurotrophic factor and neurotrophic factor 4/5
Authors: Seiffert K. , Wagner J. A. , Granstein R. D.
Published: August 2010

Neurotrophins are responsible for the survival and outgrowth of nerves within the peripheral and central nervous systems. These factors include brain‐derived neurotrophic factor (BDNF), CNTF, NT 3, and NT4/5. We have previously shown that LCs lie in close proximity to nerves and that several neuropeptides regulate LC function, implying that nerves send regulatory signals to LCs. To evaluate the possibility that LC signal nerves by release of neurotrophins, we examined LC expression of neurotrophins by RT‐PCR. To eliminate the possibility of contaminating keratinocytes in highly enriched LC preparations, we utilized the LC‐like cell lines XS52 (BALB/c derived) and XS106 (A/J derived) for initial experiments. The RNA obtained was digested with DNase to ensure complete absence of genomic DNA. Several independent RT‐PCRs revealed expression of bands of the expected size for CTNF and NT4/5, but not for BDNF and NT3 in XS106 and XS52 cells. In contrast, the transformed keratinocyte cell line PAM212 expressed BDNF, as well as CTNF and NT4/5. Preliminary experiments with purified LC confirm the expression of CTNF and NT4/5 and also show the expression of BDNF. However, we cannot be sure that BDNF expression is not due to keratinocyte contamination. We conclude that LCs may regulate nerve cells by the release of neurotrophic factors.
Functional absence of neutral endopeptidase in mice promotes hapten uptake, maturation and function of bone marrow‐derived dendritic cells
Authors: Scholzen T. E. , Fastrich M. , Brzoska T. , Armstrong C. A. , Ansel J. C. , Luger T. A.
Published: August 2010

The bioavailability of neuropeptides such as substance P (SP) released from sensory nerves or immune cells during skin inflammation is effectively controlled by proteolytic peptidases. Acute inhibition or genomic deletion of neutral endopeptidase (NEP) results in a SP‐dependent augmentation of murine allergic contact dermatitis (ACD) by affecting sensitization and elicitation phase. In this study, we address the hypothesis that absence of NEP may modulate ACD responses by affecting bone marrow‐derived dendritic cell (BmDC) maturation and function. BmDCs were generated from NEP‐deficient mice (C57BL/6J‐NEP–/–) or wild‐type controls (C57BL/6J). FACS analysis revealed that d3, d6 and d7 NEP–/– BmDCs expressed significantly more DC cell‐surface markers and costimulatory molecules compared to NEP+/+ mice BmDCs, in particular after BmDC maturation with LPS. In MLR utilizing d8 BmDCs pulsed 3 h in vitro with DNBS and T cells from in vivo DNFB‐haptenized NEP–/– and NEP+/+ mice, BmDCs from NEP–/– animals promoted proliferation of T cells with higher efficacy compared to wild‐type mice BmDCs. Likewise, T cells from NEP–/– mice demonstrated a higher proliferative response to Concavalin A stimulation or CD3/CD28 ligation compared to NEP+/+ mice. In addition, acute systemic NEP inhibition in NEP+/+ mice prior to sensitization with fluorescein isothiocyanate (FITC) after 24 h significantly augmented uptake of FITC in the CD11c‐positive DC fraction from regional lymph nodes but not from spleen compared to cells obtained from mice not treated with the NEP inhibitor. These data indicate that functional absence of NEP may significantly control cutaneous ACD inflammatory responses by promoting hapten uptake, DC maturation and T‐cell stimulation.
Modelization of skin endothelium reactivity, modulation by neuro‐peptides
Authors: Franchi J. , Crola C. , Marteau C. , Mitterrand M. , Schnebert S. , Mahé C. , André P. , Kieda C.
Published: August 2010

The endothelial cells which constitute the vascular wall are organo‐specific and reflect the biological reaction taking place in the tissue underlying the endothelial layer. This is the key for reparation after local injury, as, for example, response to UV induced production of neuromediators by neurosensory cells and keratinocytes. Endothelial cells react to the injury, mainly by the production of adhesion molecules to recruit competent cells which are selected to adhere extravasate and can operate local anti‐inflammatory/immune reaction. We demonstrate that the skin model constituted here by the cooperation of keratinocytes and skin‐derived microvascular endothelial cells is specific to the dermal selective processes as compared with the reaction of other organ microvasculary endothelial cells. Consequently, our data confirm that in order to reproduce in vitro local biological reactions it is indeed crucial to use the proper and representative endothelial cells. This is especially illustrated in this work by the conjugated effects produced by the UV of mediators production by keratinocytes and the modulation of their subsequent effect on activation of endothelial cell adhesion capacity.
Evidence for epidermal acetylcholine accumulation after oxidative stress by H2O2 and possible implications
Authors: Elwary S. , Gibbons N. , Rokos H. , Wood J. M. , Schallreuter K. U.
Published: August 2010

Acetylcholine (Ach) has been shown to be synthesized de novo and degraded in the human skin (1), while the presence of acetylcholinesterase (AchE), the degrading enzyme, was first mentioned in 1989 (2). It is now accepted that H2O2‐related oxidative stress is a major player in the development/acceleration of vitiligo (3). This oxidative stress affects the recycling of the essential cofactor (6R)‐l‐erythro 5,6,7,8 tetrahydrobiopterin (6BH4) via H2O2‐mediated oxidation of Trp and Met residues in the structure of pterin‐4a carbinolamine dehydratase (PCD) and dihydropteridine reductase (DHPR) (3). Only very recently, it was recognized that AchE is also affected and deactivated by 10−3 m H2O2 causing the accumulation of Ach in the epidermis of patients with vitiligo (4). One of the implications of oxidative stress in vitiligo includes pruritus, which was for a long time attributed to the presence of epidermal H2O2 in the 10−3 m range. In this context, a role for Ach has been suggested in association with pruritus, and therefore, we would like to suggest that Ach accumulation caused by H2O2‐mediated deactivation of AchE may well initiate pruritus (5). The Ach accumulation can also explain the earlier documented decreased sweating in patients with vitiligo (6).
The neurotrophin network in human skin
Authors: Marconi A. , Dumas M. , Fila C. , Truzzi F. , Atzei P. , Pignatti M. , Bonté F. , Giannetti A. , Pincelli C.
Published: August 2010

The neurotrophin (NT) family includes nerve growth factor (NGF), brain‐derived neurotrophic factor (BDNF), neurotrophin‐3 (NT‐3) and NT‐4/5. Keratinocytes synthesize and release all NTs. Keratinocytes express the low‐affinity p75 receptor which binds all NTs, the high‐affinity NGF receptor trkA and the NT‐3 receptor trkC. By contrast, keratinocytes express only a truncated form of trkB, the BDNF high‐affinity receptor. NT‐3 stimulates keratinocyte proliferation. NTs other than NGF fail to protect keratinocytes from UVB‐induced apoptosis. While NGF decreases upon UVB irradiation, NT‐3 and NT‐4 are upregulated. UVA dose dependently increases NT‐3 levels. NT‐3 and NGF stimulate each other release. Both fibroblasts and myofibroblasts synthesize and release all NTs, as well as their trk receptors except for trkC. p75 is more expressed in myofibroblast than in fibroblasts. NGF reduces secretion of collagen I in myofibroblasts. Myofibroblasts secrete more collagen I than fibroblasts. NGF reduces production of metalloproteinase 2 (MMP2) in myofibroblasts. Melanocytes synthesize and release all NTs. UVB irradiation upregulates the release of NT‐3, while it downregulates the release of NT‐4. Melanocytes express p75, which is downregulated by UVB. Melanocytes also express trkA and the extracellular domain of trkB. Taken together, these data indicate that a complex NT network exists in human skin with potential functions, partly to be determined.
Neurotrophins exert immunomodulatory functions on peripheral blood eosinophils in atopic dermatitis
Authors: Raap U. , Deneka N. , Goltz C. , Bruder M. , Renz H. , Kapp A. , Wedi B.
Published: August 2010

Recently, the functional role of neurotrophins on eosinophils in allergic asthma has been described. The aim of this study was to investigate the possible role of BDNF, NT‐3, NT‐4 and NGF on peripheral blood eosinophils in atopic dermatitis (AD). AD patients were defined according to the criteria of Hanifin and Rajika. Peripheral blood eosinophils were purified by CD16‐negative selection (purity >98%) and stimulated with BDNF, NT‐3, NT‐4 (10, 50 and 100 ng/ml), or NGF (100, 500 and 1000 ng/ml) for 24 up to 120 h. Apoptotic eosinophils were investigated by determining their hypodiploid DNA peak and Annexin V method, respectively. Chemotactic index was assessed in a modified Boyden chamber assay and respiratory burst by lucigenin‐dependent chemiluminescence. Stimulation with BDNF, NT‐3 and NGF significantly inhibited the programmed cell death of AD eosinophils (P < 0.05–0.01) at each time point (24 h up to 120 h in culture). Chemotactic index was significantly increased in AD eosinophils after stimulation with BDNF, NT‐3 and NT‐4 (P < 0.05–0.01). Respiratory burst of AD eosinophils was not modified after stimulation with BDNF, NT‐3, NT‐4 or NGF. To summarize, BDNF, NT‐3 and NGF significantly inhibited the programmed cell death of AD eosinophils. However, BDNF, NT‐3, NT‐4 and NGF did not modulate respiratory burst of AD peripheral blood eosinophils. On the other hand, BDNF, NT3 and NT‐4 significantly induced AD eosinophil chemotaxis. Taken together, neurotrophins have a functional role on peripheral blood eosinophils revealing neuro–immunological interactions in AD.

Supported by HILF grant.
IgE‐independent atopic dermatitis is associated with a β‐2 adrenergic receptor gene polymorphism
Authors: Roguedas A. M. , Audrezet M. P. , Scotet V. , Dupré D. , Ferec C. , Misery L.
Published: August 2010

IgE levels are not elevated in about 20% of patients with atopic dermatitis (AD). In this intrinsic AD (IAD), allergic mechanisms are not very important and pathogeny could be mainly neurogenic. β2‐adrenergic receptors are localized on cells involved in AD: Langerhans' cells, keratinocytes and lymphocytes. We wondered whether IAD could be associated with gene polymorphisms 16 and 27 of this receptor. We studied 98 healthy subjects and 83 subjects suffering from DA (UKWP criteria). IgE levels were normal in 12 of them and elevated in 71 (EAD). After DNA extraction, the genotyping was done by PCR and Direct Sequencing of candidate gene. Statistical analysis was performed with EPI‐INFO 6.04 for χ2 test. We found a significant association of Gln27Glu polymorphism with IAD (P = 0.00071 and χ2 = 14.51). There was no difference between healthy subjects and EAD patients. Adrenergic receptor agonists are known to attenuate the proliferative response of human lymphocytes after activation, through the inhibition of interleukin‐2 release. It is known that catecholamines inhibit the antigen‐presenting capability of epidermal Langerhans' cells. Long‐term agonist‐promoted downregulation of receptor number is absent when glu is at position 27. We suggest that the suppression of inhibiting effects of catecholamines could be involved in IAD pathogeny. Dichotomic nature of AD (EAD and IAD) is also associated with polymorphisms (SNP) of the interleukin‐4/interleukin‐13 receptor gene and the differences of cutaneous variables (transepidermal water loss, capacitance and pH). Altogether, these findings indicate that IAD patients exhibit phenotypic and also genotypic features which differ from those patients with EAD. Otherwise, the presence of this polymorphism could provide an explication of rarity of hypertension with AD, because Glu27Gln has been identified as a susceptibility polymorphism for HTA.
Stress modulates peptidergic innervation and alters the cutaneous immune response: exacerbating pathomechanisms in atopic dermatitis?
Authors: Peters E. M. J. , Kuhlmey A. , Knackstedt M. , Paus R. , Klapp B. F. , Arck P. C.
Published: August 2010

Stress is said to induce itchiness of the skin and exacerbate inflammatory skin diseases such as atopic dermatitis. In this context, stress mediators such as the neuropeptide substance P play a role as immunmodulators and in a wider sense growth factors. For example, we were recently able to show that stress or treatment of mice with substance P is associated with mast cell degranulation, increased cutaneous inflammation and increased apoptosis in the hair follicle. However, local interactions between the nervous and immune systems, especially under perceived stress, have rarely been reported. Here, we show for the first time, that 24 and 48 h after sonic stress exposure, the number of SP‐immunoreactive nerve fibres in the back skin of C57BL/6 mice with all there hair follicles in the resting phase of the hair cycle (telogen, low numbers of cutaneous nerve fibres) increased significantly over non‐stressed mice with the strongest increase after 24 h. Such substance P immunoreactive nerve fibres contacted mast cells more frequently, which became significant after 48 h. At the same time, the percentage of degranulated mast cells increased significantly after 24 and 48 h with the strongest increase after 48 h when apoptotic cells also became significantly upregulated. The same stressor increased dermal infiltration, e.g. by eosinophils in C57BL/6 mice with experimentally induced allergic dermatitis over mice that were either stressed or had allergic dermatitis as well as over untreated controls. Increased infiltration was associated with increased epidermal thickness in stressed mice with allergic dermatitis and with an increased number of VCAM‐immunoreactive blood vessels. At the same time, the percentage of degranulated mast cells increased significantly, and the number of substance P‐immunoreactive peptidergic sensory nerve fibres decreased in the acute allergic dermatitis lesions. By semiquantitative RT‐PCR, allergic dermatitis increased cutaneous IL‐4 and to a lesser degree IFN‐γ production, but this was not affected by stress. Ultrastructural investigation showed unmyelinated peptidergic nerve fibres in a state of deterioration close to degranulating mast cells and eosinophils in the skin of stressed mice with allergic dermatitis, suggesting a decreased number of substance P‐immunoreactive nerve fibres due to active release of SP. This may lead to an upregulation of endothelial adhesion molecules and increased infiltration by immunocytes to the skin but at mRNA level does not alter the production of classical atopy‐related cytokines in skin. These data provide first evidence for stress‐induced exacerbation of cutaneous allergic diseases such as atopic dermatitis by local interaction of the peripheral nervous system with substance P.
Proteinase‐activated receptor‐2 mediates itch: a novel pathway for pruritus in human skin
Authors: Steinhoff M. , Neisius U. , Ikoma A. , Fartasch M. , Heyer G. , Skov P. S. , Luger T. A. , Schmelz M.
Published: August 2010

Proteinase‐activated receptors are G‐protein‐coupled receptors with seven‐transmembrane domains activated by serine proteinases. PAR‐2 is a receptor for mast cell tryptase, house dust mite allergens, bacterial antigens and trypsin, for example, indicating a role of PAR‐2 during inflammation and immune responses. In the skin, PAR‐2 is expressed by keratinocytes, endothelial cells, certain immune cells and nerves, suggesting a broad regulatory role of proteases in the skin. Recently, PAR‐2 has been demonstrated to be involved in neurogenic inflammation. Therefore, we examined whether neuronal PAR‐2 may be involved in pruritus of human skin. The endogenous PAR‐2 agonist tryptase was increased up to fourfold in atopic dermatitis (AD) patients. PAR‐2 was markedly enhanced on primary afferent nerve fibres in skin biopsies of AD patients. Intracutaneous injection of endogenous PAR‐2 agonists provoked enhanced and prolonged itch when applied intralesionally. Interestingly, itch upon mast cell degranulation prevailed despite local antihistamines in AD patients only. Thus, we identified enhanced PAR‐2 signalling as a new link between inflammatory and sensory phenomena in AD patients. PAR‐2 antagonists, thus, represent a promising therapeutic target for the treatment of cutaneous neurogenic inflammation and pruritus.
β‐endorphin modulates lipogenesis in human sebocytes
Authors: Böhm M. , Li Z. , Ottaviani M. , Picardo M. , Zouboulis Ch. C. , Ständer S. , Luger T. A.
Published: August 2010

Previous research in our laboratories has demonstrated that the human sebaceous gland is a target organ for several stress hormones including α‐melanocyte‐stimulating hormone (α‐MSH) and corticotropin‐releasing hormone (CRH). To further investigate the role of stress hormones in acne, we examined the expression of opioid receptors (ORs) and the biological actions of β‐endorphin (β‐ED), a natural high‐affinity ligand for the μ‐OR (MOR) and low‐affinity ligand for the Δ‐OR (DOR), in human sebocytes. RT‐PCR, Western immunoblotting and immunofluorescence studies identified the MOR but not the DOR in the human sebocyte cell line SZ95. Expression of the MOR was also confirmed in the sebaceous gland in normal human adult skin and was detectable primarily in peripherally located sebocytes. SZ95 sebocytes did not express the β‐ED precursor pro‐opiomelanocortin (POMC) as shown by RT‐PCR analysis and Western immunoblotting and stimulation of cells with prototypical POMC inducers such as tumour necrosis factor‐α, α‐MSH, dbcAMP, phorbol ester and CRH failed to induce POMC. On the functional level, β‐ED significantly suppressed the growth of SZ95 sebocytes induced by epidermal growth factor in chemically defined calcium‐rich medium. On the other hand, β‐ED enhanced lipogenesis as shown by Nile red staining and gas chromatography. Accordingly, β‐ED dramatically increased the amount of C16 : 0, C16 : 1, C18 : 0, C18 : 1 and C18 : 2 fatty acids in an extent similar to linoleic acid used as a positive stimulus. Our data demonstrate that human sebocytes express the MOR and respond to β‐ED with reduced in vitro proliferation and increased lipogenesis. These data establish a further link between stress and acne, the latter in which excessive sebum production is an invariable feature.
Red hair, fair skin and melanoma – melanocortin 1 receptor
Authors: Rees J. L. , Waterston K. , Naysmith L. , Oh C. , Hennessy A. , Bisset Y. , Diffey B. , Ito S. , Wakamatsu K.
Published: August 2010

We have previously shown that the MC1R is a key determinant of pigmentary phenotype in man. A range of common and uncommon alleles show diminished function leading to a change in the relative amounts of eumelanin and pheomelanin. As expected, these particular allelic variants are associated with both non‐melanoma and melanoma skin cancer and other pigmentary phenotypic characteristics such as freckling. We have recently shown that even against very different genetic backgrounds, the MC1R variants show a phenotypic effect [J Invest Dermatol 2003: 121 (1): 207]. We will present data to explain how the human pigmentary phenotypes can be quantified more appropriately, in terms of both hair melanins and cutaneous response to ultraviolet radiation (submitted and in press). Our results, we would argue, are relevant to those interested in melanocortin signalling in skin and to studies of the genetics of human skin colour and evolution of skin colour.
Solar‐simulated, ultraviolet radiation‐induced, upregulation of the melanocortin‐1 receptor, pro‐opiomelanocortin, and α‐melanocyte‐ stimulating hormone in human epidermis in vivo
Authors: Schiller M. , Brzoska T. , Böhm M. , Metze D. , Scholzen T. , Rougier A. , Luger T. A.
Published: August 2010

Ultraviolet (UV) light is one of the most crucial environmental factors with regard to its capacity to induce skin cancer, premature aging of the skin, and immunosuppression. Although UV directly affects the function of epidermal cells, many of these effects are mediated by the induction of cytokines, growth factors, and neuropeptides such as α‐melanocyte‐stimulating hormone (α‐MSH). Recently, in addition to its well‐known pigmentation inducing activity, a strong anti‐inflammatory as well as an immunomodulatory potential of α‐MSH has been recognized. The aim of this study was to determine whether UV irradiation affects the expression of both α‐MSH and the melanocortin‐1 receptor (MC‐1R) in human epidermis in vivo. The volar aspects of the forearms were exposed to twice the minimal erythema dose of solar simulating radiation (SSR). Three, 6, and 24 h after irradiation, the pro‐opiomelanocortin (POMC) and interleukin‐10 (IL‐10) mRNA levels in suction blister‐induced epidermal sheets were considerably upregulated as detected by semiquantitative RT‐PCR. Furthermore, α‐MSH and IL‐10 protein levels in blister fluids were significantly increased 24 h after UV irradiation, an effect which could be abolished by the application of the broadspectrum sunscreen AnthéliosXL® prior to UV (SSR) exposure. In addition, enhanced MC‐1R mRNA and receptor protein expression upon SSR was ascertained by RT‐PCR and immunohistochemistry of the epidermal sheets, respectively. POMC‐derived neuropeptides such as α‐MSH may therefore play an important role in modulating UV‐induced inflammation.
Neurogenic skin inflammation in stress‐triggered inhibition of hair growth in mice is promoted via nerve growth factor‐dependent pathways
Authors: Arck P. C. , Peters E. M. J. , Handjiski B. , Kuhlmei A. , Paus R.
Published: August 2010

Recently, we have pointed to the existence of a brain‐hair follicle axis (BFA), with neuropeptide substance P (SP) as one candidate mediator, to which stress‐triggered hair loss is imputable. Based on findings indicating that levels of nerve growth factor (NGF) increase upon exposure to stressful events, which is particularly striking within the context of the BFA, because NGF is known to increase the release of SP, we then aimed at dissecting the role of NGF in stress‐triggered hair loss. We observed increased expression of NGF, analyzed by real time PCR and immunohistochemistry, in stress‐exposed mice with a depilation‐induced hair cycle. Expression of NGF receptor p75 was also upregulated with stress, and TrkA receptor was moderately downregulated. Upon neutralization of NGF by antibody injection, stress‐triggered premature onset of catagen, which was accompanied by apoptosis and increased number/activation of perifollicular mast cells and macrophages, was significantly inhibited. Interestingly, subcutaneous injection of recombinant NGF to mimick stress effects resulted in an increased percentage of SP‐positive neurons in dorsal root ganglia. Taken together, our data indicate that an interactive communication network between sensory nerves and immune cells in the skin is promoted by stress‐triggered release of NGF and results in mast cell activation and migration of macrophages, the release of proinflammatory neuropeptides, i.e. SP. Such disequilibrium, which may be referred to as neurogenic inflammation, constitutes the prerequisite of increased hair loss.
CRH peptides modulate proliferation, melanogenesis and dendri‐city in human follicular melanocytes
Authors: Kauser S. , Slominski A. , Wei E. T. , Tobi D. J.
Published: August 2010

Corticotropin‐releasing hormone (CRH) is the most proximal element of the hypothalamic‐pituitary‐adrenal axis (HPA) and is the chief regulator of pituitary POMC gene expression and the subsequent production and secretion of POMC peptides. Previously, our laboratories documented cutaneous expression of CRH, urocortin and functional CRH receptors (CRH‐Rs), suggesting their role in skin physiology and pathology. Human skin predominately expressed CRH‐R1 with CRH‐R2 being expressed primarily in the adnexal structures. While CRH‐R activity has been implicated in the regulation of epidermal cell function, a role for these receptors in human hair biology has not yet been demonstrated. This study was designed to investigate the effects of modified CRH peptides (D‐Glu20)‐CRH, (D‐Pro5)‐CRH and (D‐Pro4)‐urocortin with respective selectivity for CRH‐R1 and CRH‐R2 on behaviour of cultured hair follicle melanocytes (HFMs) derived from scalp of seven normal individuals. HFMs were stimulated with these peptides (10−7−10−10 m) for 72 h. (D‐Glu20)‐CRH (10−8 m) and (D‐Pro5)‐CRH (10−9 and 10−10 m) markedly increased cell dendricity, melanogenesis and proliferation (P < 0.01) compared with unstimulated levels. While (D‐Pro4)‐urocortin failed to stimulate cell dendricity, this peptide did stimulate melanogenesis (10−8 m) (P < 0.01) and exhibited a biphasic proliferative response; stimulating pigment cell division at 10−7 and 10−8 m (P < 0.01) but inhibiting proliferation at 10−9 and 10−10 m (P < 0.01). Here, we demonstrate the existence of functionally active CRH‐Rs in cultured human scalp HFM and show that signalling via these receptors modulates follicular melanocyte dendricity, melanogenesis and proliferation. Thus, activation of CRH‐Rs may have a pivotal role in the regulation of follicular melanocyte homeostasis.
The murine hair follicle is a melatonin target
Authors: Kobayashi H. , Dunlop T. W. , Tychsen B. , Conrad F. , Ito T. , Ito N. , Aiba S. , Carlberg C. , Paus R.
Published: August 2010

The pineal hormone, melatonin exerts many functional effects on mammalian skin (e.g. melanogenesis inhibition, melanocyte growth inhibition, and regulation of seasonal pelage hair growth). However, its cutaneous expression, regulation, and functional role are still obscure. The aim of this study was to check whether murine hair follicles are indeed direct, peripheral melatonin targets which express melatonin membrane receptors (MT1 and MT2) and orphan nuclear receptor α (RORα) which interact with melatonin. Immunohistochemistry revealed that murine hair follicle keratinocytes show both MT1‐like immunoreactivity (IR) and ROR‐like IR, both of which changed substantially in a hair cycle‐dependent manner. Both semiquantitive RT‐PCR for MT1 and MT2, and quantitive real‐time PCR for MT1, MT2, and ROR on murine skin cDNA revealed that all three genes are transcribed in normal mouse skin in hair cycle‐dependent manner. Functionally, melatonin significantly inhibited the constitutional level of epidermal and hair follicle keratinocyte apoptosis in short‐term mouse skin organ culture. In conclusion, we here provide evidence that normal murine hair follicles are prominent direct target for melatonin bioregulation which express MT1, MT2, and ROR, at least some of which are functionally active in situ. These receptors are regulated in a hair cycle‐dependent manner, suggesting a role of melatonin in hair cycle control.
The potential role of acetylcholine receptors in acne inversa (HS) pathogenesis
Authors: Kurzen Hjalmar , Hana Anisa , Frongia Giovanni , Goerdt Sergij , Gratchev Alexei
Published: August 2010

Acne inversa is a chronic inflammatory disorder that has been shown to be influenced by tobacco smoking, which may represent a ‘natural’ model of exogenous nicotine‐mediated activation of the nicotinic acetylcholine receptors (AChR). In previous works, we have provided a concise mapping of AChR present in normal skin and have demonstrated a crucial role of AChR in terminal differentiation and barrier formation. In addition, several studies suggest a role for the non‐neuronal cholinergic system in immunomodulation. To date, the AChR composition of the cells involved in the pathogenesis of acne inversa has not yet been characterized.

Using immunohistochemistry and RT – PCR, we could show that nicotinic α3, α5, α7 and α9 AChR as well as muscarinic M1–M5 AChR are produced in lesional epidermis in a pattern comparable with normal epidermis. In vitro and in lesional skin, we could demonstrate the presence of α3, α5 and α7 nAChR as well as M1–M5 mAChR in variable amounts on macrophages. In lesional epidermis, the choline‐acetyltransferase (ChAT)‐reactivity was particularly pronounced in the epidermal basal layer, while in the sinus tracts, ChAT reactivity was extended to all epithelial layers. High levels of ChAT especially in the hair follicle infundibulum indicate that endogenously produced ACh may act synergistically with tobacco‐delivered nicotine in aggravating infundibular hyperkeratosis. ChAT could not be detected in macrophages in vitro or in vivo indicating that macrophages do not actively contribute to ACh signalling in acne inversa but may rather be targets of ACh, and hence nicotine induced immunomodulation.
The role of stem cells in suppurative environments
Authors: Herreros Dolores , García‐Arranz Mariano , Pascual Isabel , García‐Olmo Damián
Published: August 2010

Purpose: The management of suppurative perianal lesions presents an extremely challenging problem. Stem cells (SC) extracted from certain tissues, such as adipose tissue, can differentiate into various cell types. Therefore, we have tried to use such cells to stimulate healing in a purulent environment.

Methods: In the beginning, we designed a phase I clinical trial, involving five patients with Crohn's disease. We inoculated nine fistulas in four patients with autologous adipose‐derived stem cells (ADSC) and were followed at least 8 weeks. Seventy‐five percent became healed, and 25% showed a decrease in output flow. No adverse effects were observed in any patient. This study evidenced that such cells are safe. Then, we started a research line using SC in different suppurative environments. During the course of these studies, we had the opportunity to treat a patient with perianal hidradenitis suppurativa using our current protocol of ADSC transplantation. Eight weeks after injection, patient had no perianal suppuration, and a year later remains well.

Discussion: The biological mechanism that underlies the therapeutic success of ADSC transplantation is unknown. Cell differentiation, secretion of growth factors or immunomodulatory effects have been suggested. No ethical conflicts were identified by our Ethics Committee, because the cells were autologous.

Conclusions: Our study shows that ADSC are safe for the treatment of suppurative processes. The actual number of patients included and the uncontrolled nature of these pilot studies do not allow demonstration of the effectiveness of the treatment. However, the results encourage the performance of further studies.
Indices of condition in tropical ruminants*
Authors: SINCLAIR A. R. E. , DUNCAN P.
Published: August 2010

Bone marrow dry weight expressed as a percentage of its fresh weight was found to be a good indication of its fat content in wildebeest, kongoni, and buffalo. The calculated regression equations were similar enough to suggest a more general formula for the larger ruminants.

Bone marrow fat is mobilized after the kidney fat and hence is a better measure of condition when an animal is under more extreme stresses. Fluctuations in kidney fat reflect physiological events related to reproduction.

Visual estimates of bone marrow were found to reflect true differences in fat content, and the method is useful when marrow samples cannot be collected readily. It is pointed out that estimates of condition of naturally dying wild animals must be obtained before primary and secondary causes can be identified.
Clinical trial: the combination of rifaximin with partially hydrolysed guar gum is more effective than rifaximin alone in eradicating small intestinal bacterial overgrowth
Authors: Furnari M. , Parodi A. , Gemignani L. , Giannini E. G. , Marenco S. , Savarino E. , Assandri L. , Fazio V. , Bonfanti D. , Inferrera S. , Savarino V.
Published: August 2010

Aliment Pharmacol Ther2010;32:1000–1006

Background Abnormal intestinal clearance is involved in the pathogenesis of small intestinal bacterial overgrowth (SIBO). It is known that partially hydrolysed guar gum affects intestinal motility. Eradication therapy of SIBO is based on antibiotic treatment: no data are available on the role of fibre supplementation in eradicating SIBO.

Aim To assess whether the combination of partially hydrolysed guar gum and rifaximin is more effective than rifaximin alone in the treatment of SIBO.

Methods A 50 g‐glucose breath test was given to 500 consecutive patients. Patients with a positive glucose breath test and predisposing conditions to SIBO entered into the study, and were randomized to receive rifaximin 1200 mg/day or rifaximin 1200 mg/day plus partially hydrolysed guar gum 5 g/day for 10 days. Patients completed a symptom questionnaire and glucose breath test both in basal condition and 1 month after withdrawal of therapy.

Results Seventy‐seven patients had SIBO. Eradication rate of SIBO was 62.1% in the rifaximin group (both on per‐protocol and intention‐to‐treat analyses), and 87.1% (per‐protocol,P = 0.017) and 85.0% (intention‐to‐treat,P = 0.036) in the rifaximin‐plus‐partially hydrolysed guar gum group. Clinical improvement was observed in 86.9% and 91.1% of eradicated cases in rifaximin and rifaximin‐plus‐partially hydrolysed guar gum groups respectively (P = 0.677).

Conclusion The combination of rifaximin with partially hydrolysed guar gum seems to be more useful in eradicating SIBO compared with rifaximin alone.
Cortisol response to individualised graded insulin infusions: a reproducible biomarker for CNS compounds inhibiting HPA activation
Authors: Chen Christopher L. H. , Willis Brian A. , Mooney Louise , Ong Guan Koon , Lim Chay Ngee , Lowe Stephen L. , Tauscher‐Wisniewski Sitra , Cutler Jr Gordon B. , Wiss Stephen D.
Published: August 2010

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT

• The insulin tolerance test (ITT) is commonly used to induce cortisol release in response to physiological stress. This methodology can be employed in studies evaluating the hypothalamic‐pituitary‐adrenocortical axis.

WHAT THIS STUDY ADDS

• An exploration of the threshold for cortisol release in response to blood glucose lowering, indicating that the induction of hypoglycaemia with significant symptoms is not required for a highly reproducible maximal cortisol response.

AIM

To determine the potential of cortisol secretion, in response to a physiological stressor, as a biomarker for centrally active compounds targeting the hypothalamic‐pituitary‐adrenocortical (HPA) axis.

METHODS

Cortisol response to hypoglycaemia was measured in 26 healthy males in two stages: firstly to derive an algorithm for individualized, graded insulin infusion rates to achieve defined hypoglycaemic targets over 3 h and secondly to determine the inter‐ and intra‐subject variability of cortisol response to hypoglycaemia over two identical periods by measuring the maximum (tmax), time to maximum (Cmax) response and cortisol area under the response curve (AUC).

RESULTS

Hypoglycaemia induced a consistent cortisol response starting at approximately 1 h, corresponding to blood glucose concentrations of approximately 3.3 mmol l−1, and peaking approximately 3 h after the start of infusion. The inter‐ and intra‐subject coefficients of variation (CVs) of cortisol response were approximately 19 and 19% (AUC), 15 and 19 % (Cmax) and 10 and 14% (tmax), respectively. The intra‐subject CVs for the ratio of maximum cortisol response to baseline concentration and rate of initial cortisol response between study days were more variable (32.8% and 59.0%, respectively). The blood glucose‐cortisol response model derived from the study was predictive of the individual observed cortisol responses, and estimated a blood glucose EC50 associated with onset of the cortisol response of 3.3 mmol l−1.

CONCLUSIONS

Gradual hypoglycaemia is an effective, reproducible and well‐tolerated method of stimulating a cortisol response and may therefore be useful in assessing the neuroendocrine response to HPA axis inhibitors, such as corticotropin‐releasing hormone‐1 (CRH‐1) antagonists.
Iron availability from intermediate moisture beef
Authors: OBANU Z. A. , BIGGIN RUTH J. , NEALE R. J. , LEDWARD D. A. , LAWRIE R. A.
Published: August 2010

The availability of the iron present in intermediate moisture beef before and during storage at 38°C was evaluated in both normal and anaemic rats. Irrespective of the marked chemical changes that occur in the nature of the haemoproteins during storage the availability, in all samples, was as good as that in freshly cooked beef.
Extraction and composition of pectins and hemicelluloses of cell walls of sugar beet roots grown in Morocco
Authors: Fares Khalid , Renard C. M. G. C. , R’Zina Qamar , Thibault Jean‐François
Published: August 2010

Summary Cell‐wall polysaccharides were isolated as alcohol‐insoluble residues (AIR) from Moroccan sugar beet slices at two stages of the harvest season (early and late samples). Pectins and hemicelluloses were extracted by a sequential treatment with water, EDTA, hot dilute hydrochloric acid, cold dilute sodium hydroxide, cold 1 M and 4 M sodium hydroxide and finally water at pH 7. The highest extraction yields were obtained for the dilute acid and alkali treatments, which extracted pectins rich in arabinose. The extracts from the late sample were rich in glucose which, was not retained on DEAE Sepharose CL 6 B, and which may come from microbial growth on the fields after harvesting. Yields for hemicelluloses after extraction with 1 M NaOH were 12% and 13% of AIR, respectively, for early and late beet slices, and after extraction with 4 M NaOH 5% and 6%, respectively. The extracts were rich in arabinose, xylose and glucose, and contained fucose and mannose. Fractionation on DEAE Sepharose gave one major non‐retained fraction rich in neutral sugars, corresponding to xyloglucans and a minor retained fraction corresponding to heteroxylans, and residual pectins. Washing with water at pH 7 extracted pectins rich in rhamnose and galactose.
Consumer preferences and fungal and mycotoxin contamination of dried cassava products from Ghana
Authors: Wareing Peter W. , Westby Andrew , Gibbs John A. , Allotey Liquenda T. , Halm Mary
Published: August 2010

Summary Members of one hundred and twenty five households from 19 villages producing dried cassava products were interviewed in Ghana. Kokonte was the most important cassava product in 19% of the households processing it. Most kokonte was produced between January and March. Mould growth during processing or storage was a problem during June and July, which is part of the rainy season. Most producers and market traders preferred non‐mouldy kokonte, although many (59%) would consume a mouldy product. There was a price premium for non‐mouldy kokonte. The most commonly isolated fungi were yeasts and Cladosporium spp. (44 out of 49 samples). Other fungi isolated included Aspergillus spp. (20 samples); Penicillium spp. (15 samples) and Fusarium spp. (30 samples). Sterigmatocystin was detected in 10 samples at 0.17–1.67 mg kg–1; patulin in 4 samples at 0.55–0.85 mg kg–1; cyclopiazonic acid in 4 samples at 0.08–0.72 mg kg–1; penicillic acid in 5 samples at 0.06–0.23 mg kg–1 and tenuazonic acid in 3 samples at 0.02–0.34 mg kg–1. Mycotoxin contamination of mouldy kokonte was a potential problem; there is therefore the need to improve kokonte processing to avoid mould growth.
Effect of fish to starch ratio on viscoelastic properties and microstructure of fish cracker (‘keropok’) dough
Authors: Kyaw Z. Y. , Yu S. Y. , Cheow C. S. , Dzulkifly M. H. , Howell N. K.
Published: August 2010

Starch granules found at the centre of fish crackers (‘keropok’) which contained from 0 to 10% fish did not gelatinize after 2 h steaming. This was because of water movement from the centre to the periphery during steaming. A fish protein network started to develop in dough with a 15% fish content and this entrapped water molecules. These water molecules helped to complete gelatinization. The viscoelastic properties of ‘keropok’ dough increased with an increasing fish to starch ratio and also because of the formation of a fish‐protein network in the dough.
Use of UV‐C irradiation to prevent decay and maintain postharvest quality of ripe ‘Tommy Atkins’ mangoes
Authors: González‐Aguilar G. A. , Wang C. Y. , Buta J. G. , Krizek D. T.
Published: August 2010

Ripe mangoes ‘Tommy Atkins’ were exposed to UV‐C irradiation for 10 and 20 min, prior to storage for 14 days at 5 or 20 °C and a shelf‐life period of 7 days at 20 °C. UV‐C‐treated fruit maintained better visual appearance than unirradiated controls. UV‐C irradiation for 10 min was the most effective regime in suppressing decay symptoms and maintaining firmness during storage at 5 or 20 °C. Such fruit (treated with UV‐C for 10 min) showed greater levels of putrescine and spermidine after cold storage than controls and those treated with UV‐C for 20 min. Higher levels of sugars and lower levels of organic acids were observed in mangoes treated with UV‐C for 20 min. However, the most effective UV‐C treatment (10 min) for reducing decay maintained sugar and organic acid levels similar to the controls. No UV‐damage was observed on treated fruits after storage. These results indicate that UV‐C irradiation could be used as an effective and rapid method to preserve the postharvest life of ripe mangoes without adversely affecting certain quality attributes.