Analysis of Mouse Keratin 6a Regulatory Sequences in Transgenic Mice Reveals Constitutive, Tissue-Specific Expression by a Keratin 6a Minigene

The analysis of keratin 6 expression is complicated by the presence of multiple isoforms that are expressed constitutively in a number of internal stratified epithelia, in palmoplantar epidermis, and in the companion cell layer of the hair follicle. In addition, keratin 6 expression is inducible in interfollicular epidermis and the outer root sheath of the follicle, in response to wounding stimuli, phorbol esters, or retinoic acid. In order to establish the critical regions involved in the regulation of keratin 6a (the dominant isoform in mice), we generated transgenic mice with two different-sized mouse keratin 6a constructs containing either 1.3 kb or 0.12 kb of 5' flanking sequence linked to the lacZ reporter gene. Both constructs also contained the first intron and the 3' flanking sequence of mouse keratin 6a. Ectopic expression of either transgene was not observed. Double-label immunofluorescence analyses demonstrated expression of the reporter gene in keratin 6 expressing tissues, including the hair follicle, tongue, footpad, and nail bed, showing that both transgenes retained keratinocyte-specific expression. Quantitative analysis of beta -galactosidase activity verified that both the 1.3 and 0.12 kb keratin 6a promoter constructs produced similar levels of the reporter. Notably, both constructs were constitutively expressed in the outer root sheath and interfollicular epidermis in the absence of any activating stimulus, suggesting that they lack the regulatory elements that normally silence transcription in these cells. This study has revealed that a keratin 6a minigene contains critical cis elements that mediate tissue-specific expression and that the elements regulating keratin 6 induction lie distal to the 1.3 kb promoter region.

Acidity and photolability of ruthenium salen nitrosyl and aquo complexes in aqueous solutions

The photochemical behavior of nitrosyl complexes Ru(salen)(NO)(OH(2))(+) and Ru(salen)(NO) Cl (salen = N, N`-ethylenebis-(salicylideneiminato) dianion) in aqueous solution is described. Irradiation with light in the 350-450 nm range resulted in nitric oxide (NO) release from both. For Ru(salen)(NO) Cl secondary photoreactions also resulted in chloride aquation. Thus, in both cases the final photoproduct is the diaquo cation Ru(III) (salen) (OH(2))(2)(+), for which pK(a)`s of 5.9 and 9.1 were determined for the coordinated waters. The pK(a) of the Ru(salen)(NO)(OH(2))+ cation was also determined as 4.5 +/- 0.1, and the relative acidities of these ruthenium aquo units are discussed in the context of the bonding interactions between Ru(III) and NO. (C) 2007 Elsevier B.V. All rights reserved.

Decline in homicide rates in Sao Paulo, Brasil: a descriptive analysis

Objective. To describe homicide mortality in the municipality of Sao Paulo according to type of weapon, sex, race or skin color, age, and areas of socioeconomic inequalities, between 1996 and 2008. Method. For this ecological time-series study, data about deaths in the municipality of Sao Paulo were collected from the municipal program for improvement of mortality information, using International Classification of Diseases, 10th revision (ICD-10) codes. Homicide mortality rates (HMR) were calculated for the overall population and specifically for each sex, race or skin color, age range, type of weapon, and occurrence in social deprivation/affluence areas. HMR were adjusted for age using the direct method. The percentage age of variation in HMR was calculated for the study period. For areas of socioeconomic inequalities, the relative risk of death from homicide was calculated. Results. HMR fell 73.7% between 2001 and 2008. A reduction in HMR was observed in all groups, especially males (-74.5%), young men between 15 and 24 years of age (-78.0%), and residents in areas of extreme socioeconomic deprivation (-79.3%). The reduction occurred mostly in firearm homicide rates (-74.1%). The relative risk of death from homicide in areas of extreme socioeconomic deprivation, as compared to areas with some degree of socioeconomic deprivation, was 2.77 in 1996, 3.9 in 2001, and 2.13 in 2008. In areas of high socioeconomic deprivation, the relative risk was 2.07 in 1996 and 1.96 in 2008. Conclusions. To understand the reduction in homicide rates in the municipality of Sao Paulo, it is important to take into consideration macrodeterminants that affect the entire municipality and all population subgroups, as well as micro/local determinants that have special impact on homicides committed with firearms and on subgroups such as the young, males, and residents of areas of high socioeconomic deprivation.

Distribution of HDL-cholesterol and non-HDL-cholesterol in Brazilian children and adolescents - The Floripa study

Background and aims: HDL-cholesterol (HDL-C) and non-HDL-cholesterol (nHDL-C) are involved in atherosclerosis. The aim of this study was to determine the distribution of HDL-C and nHDL-C and its association with cardiovascular and socio-cultural variables in a pediatric Brazilian sample. Methods and results: Children and adolescents from Florianopolis were randomly selected and a structured questionnaire was administered, a physical examination was performed and a blood sample was collected. Enzymatic and Direct methods in vitro were used to determine the total cholesterol and HDL-cholesterol levels. The associations among HDL-C and nHDL-C and the described variables were tested by odds ratio and logistic regression. A total of 1009 individuals were examined. Based on the Brazilian criteria, 23% were classified with low levels of HDL-C and 25% with high levels of non-HDL-C. After multivariate analysis there were significant associations among low HDL-C and high C-reactive protein (OR, 3.3; 95% CI, 2.1-5.2), paternal tobacco use (OR, 1.5; 95% CI, 1.1-2.1), and high triceps-to-subscapular index (OR, 1.5; 95% CI, 1.1-2.2). There were also significant associations among high nHDL-C and high waist circumference (OR, 1.95; 95% CI, 1.16-3.29), black skin color (OR, 1.78; 95% CI, 1.06-3.06), and high income (OR, 1.48; 95% CI, 1.09-2.02). Conclusions: In this sample, low levels of HDL-C were associated with other clinical variables such as a centripetal fat pattern and C-reactive protein, and n-HDL-C was associated with abdominal obesity, skin color and economic class. (C) 2009 Elsevier B. V. All rights reserved.

Supersaturated solutions evaluated with an in vitro stratum corneum tape stripping technique

The flux of a compound across a membrane from any formulation, whether it contains penetration enhancers or not, is limited by its saturated solubility in the vehicle. Under such conditions the concentration of the permeant in the outer layers of the stratum corneum is also saturated. Consequently, when the permeation of a drug from a supersaturated solution leads to enhanced penetration, the concentration of the drug in the outer layers of the membrane is also supersaturated. Therefore, the stratum corneum may possess antinucleant properties which inhibit or retard the crystallisation process. In this study, the enhanced in vitro permeation of supersaturated solutions of piroxicam across human skin in diffusion cells was demonstrated. The amount of permeant in the stratum corneum was determined using a tape stripping technique. Supersaturated solutions up to four degrees of saturation were investigated which produced a linear relationship between the degree of saturation and the amount of piroxicam in the stratum corneum (R-2 = 0.970). Furthermore, the amount of piroxicam in the viable layers of the skin also increased with increasing degree of saturation. An analysis of the results suggested that enhanced penetration across human skin from supersaturated solutions of piroxicam may occur as a result of the antinucleating ability of the intercellular lipids of the stratum corneum. (C) 1997 Elsevier Science B.V.

Resting energy expenditure in white and non-white severely obese women

This study aimed to compare the resting energy expenditure (REE) of white and non-white severely obese Brazilian women. REE was examined in 83 severely obese Brazilian women (n = 58 white and 25 non-white) with mean (+/- SD) age 42.99 +/- 11.35 and body mass index 46.88 +/- 6.22 kg/m(2) who were candidates for gastric bypass surgery. Body composition was assessed by air displacement plethysmography (ADP) BOD PODO body composition system (Life Measurement Instruments, Concord, CA) and REE was measured, under established protocol, with an open-circuit calorimeter (Deltatrac II MBM-200, Datex-Ohmeda, Madison, WI, USA). There was no significant difference between the REE of white and non-white severely obese women (1,953 +/- 273 and 1,906 +/- 271 kcal/d, respectively; p = 0.48). However, when adjusted for fat free mass (MLG), REE was significantly higher in non-white severely obese women (difference between groups of 158.4 kcal, p < 0.01). REE in white women was positively and significantly correlated to C-reactive protein (PCR) (r = 0.41.8; P < 0.001) and MLG (r = 0.771; P < 0.001). In the non-white women, REE was only significantly correlated to MLG (r = 0.753; P < 0.001). The multiple linear regression analysis showed that skin color, MLG and PCR were the significant determinants of REE (R(2) = 0.55). This study showed that, after adjustment for MLG, non-white severely obese women have a higher REE than the white ones. The association of body composition inflammation factors and REE in severely obese Brazilian women remains to be further investigated.

Vitamin D status in a sunny country: Where has the sun gone?

Background & aims: Hypovitaminosis D [serum 25 vitamin D < 30 ng/ml] is related to the development of metabolic bone disease and greater risk of chronic illnesses. However, it is frequently under-diagnosed, mainly in countries where UV radiation is abundant. We prospectively determined the prevalence and the predictors of serum 25 vitamin D (s25(OH)D) in a healthy Brazilian population after the winter and after the summer. Methods: 603 (118M and 485F) healthy Brazilian volunteers aged 18-90 years from a universitary hospital were selected after the winter of 2006. From the initial sample, 209 volunteers (31M and 178F) accepted to participate in a second health check after the subsequent summer. Results: After the winter, median s25(OH)D was 21.4 ng/mL and 77.4% of the population presented hypovitaminosis D. s25(OH)D was significantly related to age, BMI, PTH and race. In multivariate linear regression analysis, s25(OH)D was significantly and independently dependent on age, glycemia and skin color. Significant increase in s25(OH)D was verified after summer [10.6 (3.7-19.3 ng/ml); p < 0.001] and this improvement was dependent on age. We also observed a significant decrease in hyperparathyroidism prevalence (20.8% vs. 4.9%; P < 0.0001). Conclusion: In Sao Paulo, at the end of winter, we observed a high prevalence of hypovitaminosis D and secondary hyperparathyroidism in healthy adults. s25(OH)D was dependent on age and skin color. After summer, we observed a decrease in the prevalence of hypovitaminosis D. This unexpected finding emphasizes the need for a strong recommendation to monitor s25(OH)D, even in a sunny country such as Brazil. (C) 2010 Elsevier Ltd and European Society for Clinical Nutrition and Metabolism. All rights reserved.

Photodynamic therapy for acne vulgaris: A critical review from basics to clinical practice Part II. Understanding parameters for acne treatment with photodynamic therapy

Photodynamic therapy requires a photosensitizer, oxygen, and activating light. For acne, pilosebaceous units are ""target"" structures. Porphyrins are synthesized in vivo from 5-aminolevulinic acid (ALA), particularly in pilosebaceous units. Different photosensitizers and drug delivery methods have been reported for acne treatment. There are a variety of porphyrin precursors with different pharmacokinetic properties. Among them, ALA and methyl-ester of ALA (MAT.) are available for possible off-label treatment of acne vulgaris. In addition, various light sources, light dosimetry, drug incubation time, and pre- and posttreatment care also change efficacy and side effects. None of these variables has been optimized for acne treatment, but a number of clinical trials provide helpful guidance. In this paper, we critically analyze clinical trials, case reports, and series of cases published through 2009. (J Am Acad Dermatol 2010;63:195-211.)

Thermoregulatory Efficiency is Increased after Heat Acclimation in Tropical Natives

To evaluate the effects of heat acclimation on sweat rate redistribution and thermodynamic parameters, 9 tropical native volunteers were submitted to 11 days of exercise-heat exposures (40 +/- 0 degrees C and 45.1 +/- 0.2% relative humidity). Sudomotor function was evaluated by measuring total and local (forehead, chest, arm, forearm, and thigh) sweat rates, local sweat sodium concentration, and mean skin and rectal temperatures. We also calculated heat production (H), heat storage (S), heat exchange by radiation (R) and by convection (C), evaporated sweat (E(sw)), sweating efficiency (eta(sw)), skin wettedness (w(sk)), and the ratio between the heat storage and the sum of heat production and heat gains by radiation and convection (S/(H+R+C)). The heat acclimation increased the whole-body sweat rate and reduced the mean skin temperature. There were changes in the local sweat rate patterns: on the arm, forearm, and thigh it increased significantly from day 1 to day 11 (all p<0.05) and the sweat rates from the forehead and the chest showed a small nonsignificant increase (p=0.34 and 0.17, respectively). The relative increase of local sweat rates on day 11 was not different among the sites; however, when comparing the limbs (arm, forearm, and thigh) with the trunk (forehead and chest), there was a significant higher increase in the limbs (32 +/- 5%) in comparison to the trunk (11 +/- 2%, p=0.001). After the heat acclimation period we observed higher w(sk) and E(sw) and reduced S/(H+R+C), meaning greater thermoregulatory efficiency. The increase in the limb sweat rate, but not the increase in the trunk sweat rate, correlated with the increased w(sk), E(sw), and reduced S/(H+R+C) (p<0.05 to all). Altogether, it can be concluded that heat acclimation increased the limbs` sweat rates in tropical natives and that this increase led to increased loss of heat through evaporation of sweat and this higher sweat evaporation was related to higher thermoregulatory efficiency. J Physiol Anthropol 29(1): 1-12, 2010 http://www.jstage.jst.go.jp/browse/jpa2 [DOI: 10.2114/jpa2.29.1]

In vitro susceptibility to antimycotic drug undecanoic acid, a medium-chain fatty acid, is nutrient-dependent in the dermatophyte Trichophyton rubrum

The antimycotic activity of fatty acids has long been known, and their presence in human skin and sweat appears to protect the host against superficial mycoses. Undecanoic acid is a medium-chain fatty acid that has been used in the treatment of dermatophytoses in humans. In this study, we selected one Trichophyton rubrum undecanoic acid-resistant strain that showed a marked reduction in its capacity to grow on human nail fragments, which correlated with the reduced activity of secreted keratinolytic proteases. Moreover, the susceptibility of T. rubrum to undecanoic acid is also dependent on the carbon source utilized by both control and resistant strains. The growth of the control strain was strongly inhibited by undecanoic acid in Sabouraud medium or in cultures supplemented with low-fat milk, whereas it was ineffective when the cultures were supplemented with Tween 20 or keratin as the carbon source, suggesting that nutrient conditions are crucial in establishing a susceptibility to antifungal drugs, which is helpful for the isolation and characterization of resistant strains, and in the screening for new antifungal drugs.

Birth weight and bone mass in young adults from Brazil

Background: Birth weight is positively associated with adult bone mass. However, it is not clear if its effect is already evident in early adulthood. Objective: To investigate the association between birth weight, adult body size, the interaction between them and bone mass in young adults. Methods: Bone densitometry by DXA was performed on 496 individuals (240 men) aged 23-24 years from the 1978/79 Ribeirao Preto (southern Brazil) birth cohort, who were born and still residing in the city in 2002. Birth weight and length as well as adult weight and height were directly measured and converted to z-scores. The influence of birth weight and length, and adult weight and height on bone area (BA), bone mineral content (BMC) and bone mineral density (BMD) at the lumbar spine, proximal femur and femoral neck were investigated through simple and multiple linear regression models. Adjustments were made for sex, skin color, gestational age, physical activity level, smoking status and dietary consumption of protein, calcium and alcohol. Interaction terms between birth weight and adult weight, and birth length and adult height were tested. Results: Men in the highest fertile of birth weight distribution had greater BA and BMC at all three bone sites when compared with their counterparts in the lowest tertiles (p<0.008). For BMD, this trend was observed only in the lumbar spine. Adult weight and height were positively associated with BA and BMC at all three bone sites (p<0.05). For BMD, these associations were seen for adult weight, but for adult height an association was observed only in the lumbar spine. Birth weight retained positive associations with proximal femur BA and BMC after adjustments for current weight and height. No interaction was observed between variables measuring prenatal growth and adult body size. Conclusion: Birth weight and postnatal growth are independent determinants of adult bone mass in a sample of Brazilian adults. This effect is already evident in early adulthood. (C) 2010 Elsevier Inc. All rights reserved.

Post-transcriptional regulation of the GLI1 oncogene by the expression of alternative 5 ' untranslated regions

The oncogene GLI1 is involved in the formation of basal cell carcinoma and other tumor types as a result of the aberrant signaling of the Sonic hedgehog-Patched pathway. In this study, we have identified alternative GLI1 transcripts that differ in their 5' untranslated regions (UTRs) and are generated by exon skipping. These are denoted (alpha -UTR, beta -UTR, and gamma -UTR according to the number of noncoding exons possessed (three, two, and one, respectively). The alpha- and beta -UTR forms represent the major Gli1 transcripts expressed in mouse tissues, whereas the gamma -UTR is present at relatively low levels but is markedly induced in mouse skin treated with 12-O-tetradecanoylphorbol 13-acetate, Transcripts corresponding to the murine beta and gamma forms were identified in human tissues, but significantly, only the gamma -UTR form was present in basal cell carcinomas and in proliferating cultures of a keratinocyte cell line. Flow cytometry analysis determined that the gamma -UTR variant expresses a heterologous reporter gene 14-23-fold higher than the alpha -UTR and 5-13-fold higher than the beta -UTR in a variety of cell types. Because expression of the gamma -UTR variant correlates with proliferation, consistent with a role for GLI1 in growth promotion, up-regulation of GLI1 expression through skipping of 5' noncoding exons may be an important tumorigenic mechanism.

The sequence of a 200 kb portion of a Plasmodium vivax chromosome reveals a high degree of conservation with Plasmodium falciparum chromosome 3

Within a 199 866 base pair (bp) portion of a Plasmodium vivax chromosome we identified a conserved linkage group consisting of at least 41 genes homologous to Plasmodium falciparum genes located on chromosome 3. There were no P. vivax homologues of the P. falciparum cytoadherence-linked asexual genes clag 3.2, clag 3.1 and a var C pseudogene found on the P. vivax chromosome. Within the conserved linkage group, the gene order and structure are identical to those of P. falciparum chromosome 3. This conserved linkage group may extend to as many as 190 genes. The subtelomeric regions are different in size and the P. vivax segment contains genes for which no P. falciparum homologues have been identified to date. The size difference of at least 900 kb between the homologous P. vivax chromosome and P. falciparum chromosome 3 is presumably due to a translocation. There is substantial sequence divergence with a much higher guanine + cytosine (G + C) content in the DNA and a preference for amino acids using GC-rich codons in the deduced proteins of P. vivax. This structural conservation of homologous genes and their products combined with sequence divergence at the nucleotide level makes the P. vivax genome a powerful tool for comparative analyses of Plasmodium genomes. (C) 2001 Elsevier Science B.V. All rights reserved.

Laminin 10/11: an alternative adhesive ligand for epidermal keratinocytes with a functional role in promoting proliferation and migration

We have investigated the expression and function of the isoforms of laminin bearing the alpha(5) chain, i.e. laminin-10/11 in neonatal and adult human skin. By immunostaining human skin derived from a variety of anatomic sites, we found that the laminin-alpha(5) chain is expressed abundantly in the basement membrane underlying the interfollicular epidermis and the blood vessels in the dermis. Interestingly, while the expression level of the well-studied laminin-5 isoform did not change significantly with age, laminin-10/11 (a5 chain) appeared to decrease in the basement membrane underlying the epidermis, in adult skin. In contrast, the levels of laminin-10/11 in the basement membrane underlying blood vessels remained unchanged in neonatal vs. adult skin. Importantly, in vitro cell adhesion assays demonstrated that laminin-10/11 is a potent adhesive substrate for both neonatal and adult keratinocytes and that this adhesion is mediated by the alpha(3)beta(1), and alpha(6)beta(4) integrins. Adhesion assays performed with fractionated basal keratinocytes showed that stem cells, transit amplifying cells and early differentiating cells all adhere to purified laminin-10/11 via these receptors. Further, laminin-10/11 provided a proliferative signal for neonatal foreskin keratinocytes, adult breast skin keratinocytes, and even a human papillomavirus type-18 transformed tumorigenic keratinocyte cell line in vitro. Finally, laminin-10/11 was shown to stimulate keratinocyte migration in an in vitro wound healing assay. These results provide strong evidence for a functional role for laminin-10/11 in epidermal proliferation during homeostasis, wound healing and neoplasia.

Pathways to Melanoma Development: Lessons from the Mouse

Because of subtle differences between mouse and human skin, mice have traditionally not been an ideal model to study melanoma development. Understanding of the molecular mechanisms of melanoma predisposition, however, has been greatly improved by modeling various pathway defects in the mouse. This review analyzes the latest developments in mouse models of melanoma, and summarizes what these may indicate about the development of this neoplasm in humans. Mutations of genes involved in human melanoma have been recapitulated with some unexpected results, particularly with respect to the role of the two transcripts (Ink4a and Arf) encoded by the Cdkn2a locus. Both the Ink4a/pRb and Arf/p53 pathways are involved in melanoma development in mice, and possible mechanisms of cross-talk between the two pathways are discussed. We also know from mouse models that Ras/mitogen-activated protein kinase pathway activation is very important in melanoma development, either through direct activation of Ras (e.g., Hras G12V), or via activation of Ras-effector pathways by other oncogenes (e.g., Ret, Hgf/Sf). Ras can cooperate with the Arf/p53 pathway, and probably the Ink4a/Rb pathway, to induce melanoma. These three growth regulation pathways (Ink4a/pRb, Arf/p53, and Ras/mitogen-activated protein kinase) seem to represent three major axes of melanoma development in mice. Finally, we summarize experiments using genetically modified mice that have given indications of the intensity and timing of ultraviolet radiation exposure that may be most responsible for melanoma development.