Cutaneous denervation of psoriasiform mouse skin improves acanthosis and inflammation in a sensory neuropeptide-dependent manner

Nervous system involvement in psoriasis pathogenesis is supported by increases in nerve fiber numbers and neuropeptides in psoriatic skin and by reports detailing spontaneous plaque remission following nerve injury. Using the KC-Tie2 psoriasiform mouse model, we investigated the mechanisms by which nerve injury leads to inflammatory skin disease remission. Cutaneous nerves innervating dorsal skin of KC-Tie2 animals were surgically axotomized and beginning 1 day after denervation, CD11c(+) cell numbers decreased by 40% followed by a 30% improvement in acanthosis at 7 days and a 30% decrease in CD4(+) T-cell numbers by 10 days. Restoration of substance P (SP) signaling in denervated KC-Tie2 skin prevented decreases in CD11c(+) and CD4(+) cells, but had no effect on acanthosis; restoration of calcitonin gene-related peptide (CGRP) signaling reversed the improvement in acanthosis and prevented denervated-mediated decreases in CD4(+) cells. Under innervated conditions, small-molecule inhibition of SP in KC-Tie2 animals resulted in similar decreases to those observed following surgical denervation for cutaneous CD11c(+) and CD4(+) cell numbers; whereas small-molecule inhibition of CGRP resulted in significant reductions in CD4(+) cell numbers and acanthosis. These data demonstrate that sensory nerve-derived peptides mediate psoriasiform dendritic cell and T-cell infiltration and acanthosis and introduce targeting nerve-immunocyte/KC interactions as potential psoriasis therapeutic treatment strategies.     

Defects in antioxidant defense and calcium transport in the epidermis of xeroderma pigmentosum patients

Summary A comparative study of the antioxidant enzymes superoxide dismutase, catalase, glutathione reductase and thioredoxin reductase was undertaken in two families with xeroderma pigmentosum (XP) and in healthy controls of corresponding skin phototypes. Epidermal blister roofs obtained from the XP patients revealed significant decreases in catalase, thioredoxin reductase, and superoxide dismutase, but glutathione reductase was unaffected. In addition, keratinocytes established from XP patients contained a significantly higher than normal intracellular calcium concentration compared with control cells from a corresponding skin type. Keratinocytes established from an XP obligate heterozygote revealed intermediate levels of calcium between XP homozygotes and controls. Previously high intracellular calcium has been shown to compromise the redox status of keratinocytes by allosteric inhibition of the thioredoxin reductase/thioredoxin electron transfer system. In XP homozygous keratinocytes from sun-exposed epidermis, the intracellular concentration of reduced thioredoxin was decreased to 50% compared with these cells from unexposed skin. Taken together, the results from this study indicate that the epidermis in XP patients lacks effective defense against free radicals and peroxides. In addition to the well-established defect in the normal rates of unscheduled DNA repair, these findings provide an even better explanation for the multiple cutaneous neoplasms in these patients.     

Effect of a new topically active antiandrogen (RU 38882) on the rat sebaceous gland: comparison with cyproterone acetate

RU 38882 is a new antiandrogen. When given by subcutaneous or oral route, RU 38882 is about 25 times less potent than cyproterone acetate. However, when applied topically to the intact rat skin, RU 38882 (0.25-25 mg/rat/day for 5 days or 3 weeks) decreases, in a dose-related manner, the volume density of the smooth endoplasmic reticulum vesicles of the differentiating cells of the sebaceous gland, a structure directly involved in sebum lipid synthesis. Under these conditions RU 38882 is about 100 times more potent than cyproterone acetate and unlike cyproterone acetate, does not modify the prostate weight. The lack of efficacy of cyproterone acetate on the sebaceous gland could be due to its partial androgenic activity while RU 38882, under these conditions, acts as a pure antiandrogen which inhibits the nuclear androgen-receptor translocation.     

A model system using tape stripping for characterization of Langerhans cell-precursors in vivo

Little is known about the immigration of bone marrow-derived progenitors of Langerhans cells (LC) into the epidermis. We developed an in vivo system based on the tape stripping method that allowed us to study the immigration of LC into the epidermis after intradermal injection of bone marrow-derived dendritic cells (DC). Tape stripping induced a mechanical disruption of the epidermal barrier that led to skin inflammation and subsequent emigration of LC and dermal DC from the skin. Emigrating LC and dermal DC were observed in lymphatic vessels, and the numbers of LC and dermal DC in the draining lymph node increased. Up to 500 times more injected precursors migrated into tape-stripped epidermis as compared with unstripped epidermis. Newly immigrated cells were slender with one or two dendrites and acquired a more dendritic morphology after 2-4 days. They were both MHC II-positive and negative and they did not express Langerin/CD207, nor macrophage-mannose receptor/CD206 and Fc-epsilon receptor I. In contrast, all cells that had entered the epidermis expressed CD11c and CCR6, suggesting that they were LC. We conclude that this experimental system may serve as a valuable tool for the further characterization of LC-precursors and the conditions necessary for LC-immigration into the epidermis.     

GADD45 regulates G2/M arrest,DNA repair,and cell death in keratinocytes following ultraviolet exposure

GADD45 is a multifunctional protein that is regulated by p53. p53 plays an important role in regulating DNA repair and in the response to ultraviolet light in keratinocytes. This study investigates the role of GADD45 in the response to ultraviolet B. Cell cycle analysis demonstrated that wild-type and Gadd45-deficient cells have transient G2/M arrest, but only in the wild-type cells was arrest sustained. Cdc2 kinase activity in immunoprecipitates from normal and Gadd45-deficient cells decreases after irradiation in normal cells but not in Gadd45-deficient cells. An immunofluorescent study with Cdc2 antibody demonstrated diffuse cellular fluorescence before ultraviolet irradiation in both Gadd45-deficient and wild-type cells, but upon ultraviolet irradiation only Gadd45-proficient cells showed Cdc2 sequestration in the cytoplasm. Gadd45-deficient cells also have a slower rate of nucleotide excision repair. The lack of G2/M arrest coupled with reduced DNA repair leads to a higher ultraviolet sensitivity of Gadd45-deficient cells. These results reveal that GADD45 promotes G2/M arrest via nuclear export and kinase activity of Cdc2, increases global genomic DNA repair, and inhibits cell death in keratinocytes. Thus, GADD45 plays an important role in maintaining genomic integrity in ultraviolet-exposed skin.     

Critical role of the interleukin-23/T-helper 17 cell axis in the pathogenesis of psoriasis

Psoriasis is an inflammatory disease with dynamic interactions between the immune system and the skin. Recent studies have demonstrated that the interleukin (IL)-23/T-helper (Th)17 cell axis plays an important role in the pathogenesis of psoriasis. Here, the biology and function of Th17 cells as well as the crucial role of IL-23 in the context of the Th17 cell-dependent chronic inflammation in psoriatic skins are reviewed. Recent study about the role of the IL-23/Th17 axis in the pathogenesis of psoriasis-like lesions in K5.Stat3C transgenic mice is also discussed. This model mouse for psoriasis not only verifies the therapeutic efficacies of biologics that specifically target the IL-23/Th17 axis, but also clarifies the pathogenesis of psoriasis.     

Melanin granula distribution and phagocytosis in psoriasis vulgaris after PUVA therapy

Summary Melanin-containing basal cells of the epidermis, melanin-containing macrophages, mast cells, eosinophilic granulocytes and plasma cells were quantitatively investigated with the purpose of gaining an understanding of the quantitative changes in these cell systems under PUVA therapy. This patients have been exposed to solar radiation some weeks ore months before the begin of the PUVA-treatment. Different dying-processes were used to investigate biopsy samples of psoriatically healthy and psoriatically affected skin, from 28 patients before, and 39 patients after PUVA therapy, using a 2 d m with a field of view of 0.1 mm². Altogether more than 9,000 fields of view have been analysed. The average radiation amount was 12 irradiations with an average total energy of 21.5 J/cm². It was found that the count of granula-containing basal layer cells decreases in the psoriatic healthy region due to pigment incontinence and increase in the psoriatically affected region. The subepidermal melanincontaining phagocytes increase in both regions to a similar extent. In the case of the mast cells there was no trend to degranulation. The count of eosinophilic granulocytes and plasma cells was unchanged.     

Radioautographic measurement of electron-induced epidermal kinetic effects in different aged rats

We have previously shown that the ability of rat epidermal cells to repair electron-induced DNA damage decreases as a function of age. The present investigation was performed to examine the relationship between this finding and sensitivity of epidermal cells to the cytotoxic effects of the radiation. Male CD rats at ages 2, 28, 100, 200, 420, and 728 days were injected with [3H]-thymidine [( 3H]Thd) at a dose of 2 mu Ci/g body weight. One hour later, the rats were anesthetized and the dorsal skin irradiated with various doses of 0.8 meV electrons at a dose rate of 660 rads/min. At 24 h after irradiation, radioautographs were made of a sheet of epidermis that was separated by trypsinization from the underlying dermis. Labeled cells were scored either as singlets or doublets (adjacent labeled cells). The percent labeled cells and percent labeled cells as doublets were determined. The estimated labeling index (the proportion of cells labeled by a single exposure to [3H]Thd) of the epidermal basal layer decreased as a function of age. The slope of the semilog plot of the percent labeled cells as doublets as a function of electron dose indicates that the Do value decreases with increasing age. The results show, however, that the greatest difference in sensitivity occurs between 2-day (neonatal) and 28-day (pubescent) animals and again between 420-day (adult) and 728-day (senescent) animals.     

Correlation between a Cutometer® and quantitative evaluation using Moire topography in age-related skin elasticity

BACKGROUND/PURPOSE: As aging occurs, our skin gets more wrinkles, becomes drier and loses its elasticity. Validating the evaluation of skin elasticity is especially important, because it is not as visible as other signs of aging such as wrinkles. So it is needed that the method for measuring skin elasticity is able to reflect perception about the change of the skin state. METHODS: Here, the correlation between age and the parameters given by a Cutometer is identified and the main parameters that reflect the decreases in skin elasticity in terms of ages are presented. Also, Moire's system, an evaluation method to quantify the sensory value of viewing, is developed. A five-grade standard of Moire topographic photo scale on the face is used to evaluate the state of skin elasticity and lifting 20- to 61-year-old women. Based on this photo standard, scoring is performed using a five-grade system by three specialists to obtain the consensus score. The score is compared with the result of a Cutometer. RESULTS: Significant negative correlations between age and results of a Cutometer (r=-0.687-0.725), Moire's topography scores (r=-938), were found. Some Cutometer parameters and the decreases in skin elasticity in terms of ages were highly correlated (r=-0.687-0.725). The results from Moire system and flexibility as sensory evaluation also had a very high correlation with age (r=-0.765-0.932). Finally, we have shown the significance of the correlation between the result of a Cutometer and the score of Moire topography (r=0.711). CONCLUSIONS: It is considered that Cutometer parameters R7 and R2 are used as main parameters to assess skin elasticity and aging. And our studies using Moire topography on the face have confirmed that instrumental measurements reflect the decrease in skin elasticity, which is perceived visually.     

Cell kinetics of the human hair follicle

The human hair follicle is a complex three-dimensional structure, and little is known about the control of growth or differentiation in this tissue. The base of the follicle is divided into two compartments: (1) the keratinocytes of the hair matrix and (2) the dermal papilla, which consists of fibroblasts and often a microvascular system. The dermal papilla exerts a strong influence on the behavior of the follicle keratinocytes. The keratinocytes in an anagen follicle cycle rapidly and migrate upward to form the hair fiber and inner root sheath. These cells thus express up to seven different differentiated phenotypes, yet we know nothing about how this is controlled or how the cells migrate into each layer.

An approach that has proved fruitful in other systems has been to define the rates of proliferation in different parts of the tissue. This can lead to identifying which cells are targets for growth regulation and provide information on how proliferation and differentiation are coordinated. One must understand the limitations of the techniques and know what can be derived from resultant data. Cell kinetics is an area where techniques are often misapplied or the results misinterpreted.     

Innate immunity in cutaneous melanoma

The skin immune system is composed of a vast network of immune cells, including lymphocytes, macrophages, neutrophils, dendritic cells and Langerhans cells, which not only are involved in inflammatory responses but also contribute to homeostatic function and may participate in the various steps of carcinogenesis. Many studies support the notion that innate immunity has a key role in the development, growth and prognosis of cutaneous malignant melanoma (MM), through the release of pro‐ and/or anti‐inflammatory cytokines and tumour growth factors. The tumour environment in a major subset of cutaneous MM shows evidence of a T cell‐infiltrated phenotype, but there is less known about the presence and the phenotype of other immune system cells. Response to immunotherapy is largely correlated with the presence of T cells in the tumour microenvironment, while the regulation exerted by stromal components such as macrophages and mast cells has been less investigated. In the current report, we review the recent literature, focusing our attention on the role of macrophages, dendritic cells, mast cells and natural killer cells in orchestrating MM progression, to better understand tumour immunobiology. The identification of new therapeutic targets and the application of approaches aimed at modulating crosstalk between immune and tumour cells, could have a crucial impact on immunotherapy and result in better clinical outcome. We hope this review will be helpful in cutaneous MM research.     

Sun protection factor persistence on human skin during a day without physical activity or ultraviolet exposure

Background/purpose: Recently, we showed that the sun protection factor (SPF) decreases by a constant factor to reach 55% during a day with activities. Organic sunscreens but not inorganic ones are absorbed through the skin. We wished to determine the SPF decrease caused by absorption by investigating the difference in SPF decreases between an organic and an inorganic sunscreen, assuming that the sunscreens are stable, and that the SPF decrease is time dependent if caused by absorption.
Methods: Sunscreens were used on the backs of 22 participants, who were physically inactive at 22 °C. SPF testing was performed 30 min, 4, and 8 h after application of 2 mg/cm² sunscreen. Whether cream evaporation changed the ultraviolet (UV) transmission was studied in vitro .
Results: The SPFs of the organic and inorganic sunscreens were reduced by about 25% after 8 h. Evaporation of the cream did not cause a change in UV transmission in vitro .
Conclusion: A similar decrease in SPF of the organic and inorganic sunscreen was seen during 8 h without activities, and is thus not likely to be caused by absorption or evaporation from the skin. The SPF decrease after 8 h is about 55% when the participants perform activities and 25% without activities.
Trial registration: Registered at http://www.clinicaltrials.gov . Register name: 'Sunscreen: Persistence of Sun Protection Factor and the Influence on Vitamin D'. Register number H-B-2007-120.     

Mild hyperbaric oxygen activates the proliferation of epidermal basal cells in aged mice

The proliferation of epidermal basal cells decreases with age. This study examined the effects of exposure to mild hyperbaric oxygen on the proliferative activity of epidermal basal cells in aged mouse skin. Hairless mice aged 5, 34 and 55 weeks were exposed to mild hyperbaric oxygen at 1266 hPa with 36% oxygen for 6 h/day for 1 or 2 weeks. Skin samples were then collected from the back area to evaluate epidermal thickness and the number and proliferative activity of epidermal basal cells. Exposure to mild hyperbaric oxygen had no effect on the epidermal thickness, irrespective of age, but accelerated the proliferative activity of epidermal basal cells in aged mouse skin.     

Migratory Langerhans cells in mouse lymph nodes in steady state and inflammation

Dendritic cells cells induce immunity or-in the steady state-maintain peripheral tolerance. Little is known in that regard about Langerhans cells. Therefore, we investigated migrating Langerhans cells in the steady-state versus inflammation. Increased numbers of Langerhans cells, as determined by immunostaining for Langerin/CD207, appeared in the lymph nodes in response to a contact allergen. Whereas a large proportion of Langerhans cells expressed CD86 in the steady state, CD40, and CD80 were found on a smaller percentage. During inflammation, more CD40(+), CD80(+), CD274/B7-H1/PD-L1(+), and CD273/B7-DC/PD-L2(+) Langerhans cells were found in the lymph nodes, and they expressed higher levels of these molecules. CD275/inducible T cell co-stimulator (ICOS) ligand was not detected. Langerhans cells in the nodes of contact allergen-treated mice produced more IL-12p40/70. This correlated with more interferon-gamma being produced by activated lymph node T cells. Epicutaneous immunization with ovalbumin under inflammatory conditions led to a more vigorous proliferation of antigen-specific CD4 T cells in vitro and in vivo as compared with immunization in the steady state. The latter modality, however did not induce strong CD4 T cell tolerance in this model. Thus, the overall phenotype of Langerhans cells is not an indicator for their immunogenic or tolerogenic potential.     

The significance of stress hormones (glucocorticoids, catecholamines) for eruptions and spontaneous remission phases in psoriasis

BACKGROUND: In an earlier paper, it was described how acute eruptions of psoriasis may be produced in phases of immune deficiency and in the presence of bacterial antigen-releasing inflammatory foci, whereas clinical spontaneous remissions are produced in phases of immunologic activity. Therefore, it was of interest to investigate whether the stress hormones cortisol/epinephrine are involved in triggering such deficiency and activity phases. METHODS: During a series of investigations lasting up to 3 years in 95 patients, the following were determined: cortisol/epinephrine levels, polyclonal serum immunoglobulins IgM, IgG, and IgA, total serum IgE, complement C3 and C4 proteins, T cells and subpopulations, as well as streptococcal titers ASO/ADNase B, severity index (PASI) RESULTS: Phases of clinical inactivity are associated with the mechanism, "immunologic regulation," where antibacterial titers are elevated, but all other parameters are unremarkable. Eruption phases (in 32 of 95 patients) showed absolute increases in serum cortisol levels and antibacterial titers, and decreases in serum epinephrine (adrenaline) levels. Phases of spontaneous remission (in 25 of 32 patients) showed, in contrast to the eruption phases, absolute increases in serum epinephrine levels, and significant falls in serum cortisol levels and bacterial titers. CONCLUSIONS: On the basis of these results, the participation of the immune system is confirmed in the pathogenesis of psoriasis, which is subject to control by higher neurohormonal systems. Cortisol may be involved in the clinical eruption phase, and epinephrine in the remission phase. Both hormones are true antagonists and have important effects on the human immune system if produced in excess via the pituitary-adrenal axis. Infection with Streptococcus pyogenes is an additional trigger for the dermatosis.     

Topically applied L-carnitine effectively reduces sebum secretion in human skin

Background Oily skin condition is caused by an excessive sebaceous gland activity, resulting in an overproduction of sebum, giving the skin an undesired shiny, oily appearance. Aims To identify an active substance that reduces sebum production in human sebaceous glands by regulating fat metabolism in a natural way. Patients/Methods The effects of l ‐carnitine on β‐oxidation and intracellular lipid content were investigated in vitro using the human sebaceous cell line SZ95. Penetration experiments utilizing pig skin as a model system were performed with a cosmetic formulation containing radioactively labeled l ‐carnitine. To determine the in vivo effects, a vehicle‐controlled, randomized study was carried out using a cosmetic formulation containing 2%l ‐carnitine for 3 weeks. Sebum production was investigated utilizing the lipid‐absorbent Sebutape^®. Results SZ95 cells treated with 0.5% or 1%l ‐carnitine demonstrated a significant concentration‐dependent increase in β‐oxidation compared to control cells. Following the treatment with l ‐carnitine, intracellular lipid concentrations decreased significantly in a dose‐dependent manner compared with untreated control cells. In skin penetration experiments, topically applied l ‐carnitine reached the dermis. In addition, topical in vivo application of a formulation containing 2%l ‐carnitine for 3 weeks significantly decreased the sebum secretion rate compared to the treatment with vehicle. Conclusions Our results show that the treatment of human sebocytes with l ‐carnitine significantly augments β‐oxidation and significantly decreases intracellular lipid content in human sebocytes. Topically applied l ‐carnitine is bioavailable and leads to a significant sebum reduction in vivo. In conclusion, l ‐carnitine represents a valuable compound, produced naturally within the body, for the topical treatment of oily skin in humans.     

Alopecia areata susceptibility in rodent models

With our current view of alopecia areata as an autoimmune disease, it is probable that disease development in an individual is dependent on multiple genetic and environmental factors interacting in a complex system. Rodent models afford the opportunity to investigate alopecia areata development and to define the significance of the different factors involved. Recently, rodent model characterization has been conducted using flow cytometry, microarray analysis, and functional studies. From these a pattern of events in alopecia areata development has emerged. Although the preliminary activation events for the onset of alopecia areata remain unknown, the response of the immune system is characterized by antigen presentation and costimulation of lymphocytes in the lymph nodes and skin, a deficiency of CD4+/CD25+ regulatory cells, and an action of activated lymphocytes on hair follicles via Fas/FasL signaling and cytokines. Thus, onset of disease may require appropriate (or inappropriate) expression of stimulatory antigens within the hair follicle, the breakdown of the putative hair follicle immune privilege, the presentation of antigens to the immune system, a failure of immune system regulation, and the ability of the activated immune system to disrupt anagen-stage hair follicles. Once the sequence of events is initiated, it may become a self-perpetuating cycle, with epitope spreading leading to a wider range of targets in chronic alopecia areata. Rodent model studies have provided significant insight into alopecia areata, but much more remains to be explained about the mechanisms of disease development.     

Analysis of quinolone resistance mechanisms in Neisseria gonorrhoeae isolates in vitro

BACKGROUND AND OBJECTIVES: Gonococcal fluoroquinolone resistance is now a significant problem in Japan. We generated gonococcal mutants resistant to norfloxacin in vitro from norfloxacin sensitive isolates and analysed the contribution of three known mechanisms of quinolone resistance in Neisseria gonorrhoeae. MATERIALS AND METHODS: Three clinical isolates of N gonorrhoeae susceptible to norfloxacin were exposed to increasing concentrations of norfloxacin. To identify mutations in the gyrA and parC genes of the gonococcal mutants, the quinolone resistance determining regions of the gyrA and parC genes were polymerase chain reaction (PCR) amplified and the PCR products were directly sequenced. Norfloxacin accumulation in the gonococcal cells was also measured. RESULTS: The MICs of norfloxacin for three variants containing a single GyrA mutation were 16-fold higher than that for their parent isolates. A variant showing reduced norfloxacin accumulation in the cells, without mutations in the GyrA or ParC proteins, was also less sensitive to norfloxacin, with a 16-fold increase in the MIC, compared with the parent strain. The MIC of norfloxacin for a variant which contained a single GyrA mutation with reduced norfloxacin accumulation in the cells was 128-fold higher than for the parent strain. A variant containing mutations in both GyrA and ParC proteins with reduced accumulation of norfloxacin in the cells showed a 256-fold increase in the norfloxacin MIC compared with the parent strain. There was no variant containing a ParC mutation without the simultaneous presence of a GyrA mutation. CONCLUSIONS: The results from this study suggest that not only a mutation in the gyrA gene but also reduced drug accumulation in cells contributes to the development of fluoroquinolone a mutation in the gyrA gene contributes to a high level of fluoroquinolone resistance in gonococci with decreases in accumulation in cells having an additional but lesser effect.


     

Qualitative and quantitative studies of Paget cells in the horny layer

One case of extramammary Paget's disease was studied by ultrastructural and morphometrical methods to clarify whether or not Paget cells change qualitatively or quantitatively in passing through the epidermis. Paget cells in living layers were observed as large cells with abundant clear cytoplasm, large nuclei, and prominent nucleoli. The cytoplasm was enriched with well developed rough-surfaced endoplasmic reticulum, mitochondria, free ribosomes, and glycogen particles. In contrast, filamentous structures were not developed. Paget cells are connected to each other and adjacent keratinocytes by small desmosomes. In contrast, in the horny layer, Paget cells contained shrunken, flattened nuclei. Cell organelles had degenerated and large lipid droplets were present in the cytoplasm. As disruption of the desmosomes took place, intercellular spaces between Paget cells and keratinocytes widened. Based on the morphometrical analysis by a sonic digitizer computer system, there were statistically significant decreases in areas and diameters of Paget cells between the horny layer and the living cell layers, including the basal, spinous and granular layers. In conclusion, it is suggested that Paget cells change qualitatively and quantitatively after they move into the horny layer from the living layers.