Regulation of epidermal tight junctions by calcium ATPases and p38

The epidermis is the upper layer of the skin and keratinocytes are its most abundant cells. Tight junctions are cell junctions located in the granular layer of the epidermis. They maintain the polarity of the cells and regulate the movement of water-soluble molecules. Epidermal tight junctions may lose their integrity when there are defects in intercellular calcium regulation. Hailey-Hailey and Darier´s disease are dominantly inherited, blistering skin diseases. Hailey-Hailey disease is caused by mutations in the ATP2C1 gene encoding a calcium/manganese ATPase SPCA1 of the Golgi apparatus. Darier´s disease is caused by mutations in the ATP2A2 gene encoding a calcium ATPase SERCA2 of the endoplasmic reticulum. p38 regulates the differentiation of keratinocytes. The overall regulation of epidermal tight junctions is not well understood. The present study examined the regulation of tight junctions in the human epidermis with a focus on calcium ATPases and p38. Skin from Hailey-Hailey and Darier´s disease patients was studied by using immunofluorescence labeling which targeted intercellular junction proteins. Transepidermal water loss was also measured. ATP2C1 gene expression was silenced in cultured keratinocytes, by siRNA, which modeled Hailey-Hailey disease. Expression of intercellular junction proteins was studied at the mRNA and protein levels. Squamous cell carcinoma and normal human keratinocytes were used as a model for impaired and normal keratinocyte differentiation, and the role of p38 isoforms alpha and delta in the regulation of intercellular junction proteins was studied. Both p38 isoforms were silenced by adenovirus cell transduction, chemical inhibitors or siRNA and keratinocyte differentiation was assessed. The results of this thesis revealed that: i.) intercellular junction proteins are expressed normally in acantholytic skin areas of patients with Hailey-Hailey or Darier´s disease but the localization of ZO-1 expanded to the stratum spinosum; ii.) tight junction proteins, claudin-1 and -4, are regulated by ATP2C1 in non-differentiating keratinocytes; and iii.) p38 delta regulates the expression of tight junction protein ZO-1 in proliferating keratinocytes and in squamous cell carcinoma derived cells. ZO-1 silencing, however, did not affect the expression of other tight junction proteins, suggesting that they are differently regulated. This thesis introduces new mechanisms involved in the regulation of tight junctions revealing new interactions. It provides novel evidence linking intracellular calcium regulation and tight junctions.

Leaf anatomy of Araucaria angustifolia (Bertol.) Kuntze (Araucariaceae)

Immature and mature leaves of juvenile and adult plants of Araucaria angustifolia (Araucariaceae) were observed with the objective of updating the morphoanatomical data of the leaves of this species, which were restricted to basic descriptions in previous studies. The observations, made in optical allowed to establish anatomical differences among mature leaves of juvenile and adult plants in relation to the number of palisade parenchimal layers, the number of compartmented cells and the transfusion tissue development. Epidermis, the albuminous cells, the phloem, and the transfusion tissue descriptions are in disagreement with the data obtained data by different authors. The epidermal tissue and the hypodermis differ entirely when the plant is still juvenile, being inferred that these tissues would soon perform the protection function against mechanical damages and water loss, the vital characteristics during the first development months of young offspring.

Anatomical and developmental aspects of leaf galls induced by Schizomyia macrocapillata Maia (Diptera: Cecidomyiidae) on Bauhinia brevipes Vogel (Fabaceae)

Samples of healthy leaves and galls induced by Schizomyia macrocapillata Maia on Bauhinia brevipes Vogel were submitted to routine techniques to investigate gall anatomy and development. Pouch galls are induced on the abaxial surface of unfolded immature leaves, and become spheroid with long reddish hairs covering their external surface. Galls occur isolated or coalesce when in larger numbers. Gall development was divided into six phases: 1) initiation; 2) tissue re-arrangement; 3) tissue differentiation; 4) maturation; 5) growth phase; and 6) dehiscence. This last phase corresponds to gall senescence, which takes place just after the larva exits the chamber to pupate. An important developmental phase of tissue reorientation was recorded after the initiation phase. The presence of hyphae close to the covering layer characterizes this gall as an ambrosia gall and the feeding mode of the gall migde is discussed. Few hyphae were found during the first developmental phases and fungi may play an important role during gall morphogenesis. Neoformed trichomes may provide not only photoprotection but also protection against natural enemies and water loss. The neoformation of phloematic bundles suggests host manipulation and indicates the establishment of a deviating sink.

Relationships between crown architecture and available irradiance in two cerrado species with different leaf phenologies

Structural differences between cerrado species with different leaf phenologies are linked to crown architecture, leaf production, and biomass allocation to shoots and leaves. The present study characterized crown structures and the patterns of biomass allocation to leaves and shoots in two woody cerrado species with contrasting leaf phenologies and quantified the irradiance reaching their leaves to determine the best period during the day for photosynthetic activity. The shoots and leaves of five individuals of both Annona coriacea (deciduous) and Hymenaea stigonocarpa (evergreen) were collected along a 50 m transect in a cerrado fragment within the urban perimeter of Catalão - GO, to determine their patterns of biomass allocation in their crowns. The evergreen H. stigonocarpa had significantly higher mean values of shoot inclination (SI), petiole length (PL), leaf area (LA), leaf display index (LDI), and individual leaf area per shoot (ILA), while the deciduous species A. coriacea had significantly higher leaf numbers (LN). The more complex crown of H. stigonocarpa had shoots in more erect positions (orthotropic), with intense self-shading within shoots; A. coriacea, on the other hand, had slanting (plagiotropic) shoots in the crown, allowing similar irradiance levels to all leaf surfaces. The production of plagiotropic shoots by the deciduous species (A. coriacea) is a strategy that enables its use of incident sunlight early in the morning and preventing excessive water loss or excessive irradiance. Hymenaea stigonocarpa (an evergreen), by contrast, had orthotropic shoots and uses intense self-shading as a strategy to avoid excessive irradiance, especially at midday. Differences in crown architectures between evergreen and deciduous species of cerrado sensu stricto can therefore be viewed as adaptations to the environmental light regime.

Time motion analysis of football (soccer) referees during official matches in relation to the type of fluid consumed

The aim of the present study was to determine the effect of volume and composition of fluid replacement on the physical performance of male football referees. Ten referees were evaluated during three official matches. In one match the participants were asked to consume mineral water ad libitum, and in the others they consumed a pre-determined volume of mineral water or a carbohydrate electrolyte solution (6.4% carbohydrate and 22 mM Na+) equivalent to 1% of their baseline body mass (half before the match and half during the interval). Total water loss, sweat rate and match physiological performance were measured. When rehydrated ad libitum (pre-match and at half time) participants lost 1.97 ± 0.18% of their pre-match body mass (2.14 ± 0.19 L). This parameter was significantly reduced when they consumed a pre-determined volume of fluid. Sweat rate was significantly reduced when the referees ingested a pre-determined volume of a carbohydrate electrolyte solution, 0.72 ± 0.12 vs 1.16 ± 0.11 L/h ad libitum. The high percentage (74.1%) of movements at low speed (walking, jogging) observed when they ingested fluid ad libitum was significantly reduced to 71% with mineral water and to 69.9% with carbohydrate solution. An increase in percent movement expended in backward running was observed when they consumed a pre-determined volume of carbohydrate solution, 7.7 ± 0.5 vs 5.5 ± 0.5% ad libitum. The improved hydration status achieved with the carbohydrate electrolyte solution reduced the length of time spent in activities at low-speed movements and increased the time spent in activities demanding high-energy expenditure.

Mathematical modelling of the osmotic dehydration of cherry tomato (Lycopersicon esculentum var. cerasiforme)

Osmotic dehydration of cherry tomato as influenced by osmotic agent (sodium chloride and a mixed sodium chloride and sucrose solutions) and solution concentration (10 and 25% w/w) at room temperature (25°C) was studied. Kinetics of water loss and solids uptake were determined by a two parameter model, based on Fick's second law and applied to spherical geometry. The water apparent diffusivity coefficients obtained ranged from 2.17x10-10 to 11.69x10-10 m²/s.

Osmotic dehydration of sweet potato (Ipomoea batatas) in ternary solutions

The aim of this work was to evaluate the osmotic dehydration of sweet potato (Ipomoea batatas) using hypertonic sucrose solutions, with or without NaCl, at three different concentrations, at 40 °C. Highest water losses were obtained when the mixture of sucrose and NaCl was used. The addition of NaCl to osmotic solutions increases the driving force of the process and it is verified that the osmotic dehydration process is mainly influenced by changes in NaCl concentration, but the positive effect of the salt-sucrose interaction on soluble solids also determined the decrease of solid gain when solutes were at maximum concentrations. Mass transfer kinetics were modeled according to Peleg, Fick and Page's equations, which presented good fittings of the experimental data. Peleg's equation and Page's model presented the best fitting and showed excellent predictive capacity for water loss and salt gain data. The effective diffusivity determined using Fick's Second Law applied to slice geometry was found to be in the range from 3.82 x 10-11 to 7.46 x 10-11 m²/s for water loss and from 1.18 x 10-10 to 3.38 x 10-11 m²/s for solid gain.

Process variables in the osmotic dehydration of sliced peaches

This paper evaluated the influence of temperature and concentration of the sucrose syrup on the pre-osmotic dehydration of peaches. Physical (colour and texture) and chemical variables (soluble solid content; total sugar, reducing and non-reducing sugar contents; and titratable acidity) were investigated, as well as the osmotic dehydration parameters (loss of weight and water; solids incorporation). An experimental central composite design was employed varying the temperature (from 30 to 50 ºC) and concentration (from 45 to 65 ºBrix) and maintaining the syrup to fruit ratio (4:1), process time (4 hours), and format (slices). The degree of acceptance was used in the sensory analysis evaluating the following characteristics: appearance, taste, texture, colour, and overall quality using a hedonic scale. The results were modelled using the Statistica program (v. 6.0) and the Response Surface Methodology. The mathematical models of the following dimensionless variations yielded significant (p < 0.05) and predictive results: soluble solids content, total and non-reducing sugar contents, titratable acidity, colour parameter L*, and water loss. The models of the attributes colour and appearance yielded significant (p < 0.10) but not predictive results. Temperature was the prevalent effect in the models. The process conditions in the range from 50 to 54.1 ºC and from 45 to 65 ºBrix led to greater water losses and better sensory performances.

Integration of cooking and vacuum cooling of carrots in a same vessel

Cooked vegetables are commonly used in the preparation of ready-to-eat foods. The integration of cooking and cooling of carrots and vacuum cooling in a single vessel is described in this paper. The combination of different methods of cooking and vacuum cooling was investigated. Integrated processes of cooking and vacuum cooling in a same vessel enabled obtaining cooked and cooled carrots at the final temperature of 10 ºC, which is adequate for preparing ready-to-eat foods safely. When cooking and cooling steps were performed with the samples immersed in boiling water, the effective weight loss was approximately 3.6%. When the cooking step was performed with the samples in boiling water or steamed, and the vacuum cooling was applied after draining the boiling water, water loss ranged between 15 and 20%, which caused changes in the product texture. This problem can be solved with rehydration using a small amount of sterile cold water. The instrumental textural properties of carrots samples rehydrated at both vacuum and atmospheric conditions were very similar. Therefore, the integrated process of cooking and vacuum cooling of carrots in a single vessel is a feasible alternative for processing such kind of foods.

Physicochemical changes in cubiu fruits (Solanum sessiliflorum Dunal) at different ripening stages

Cubiu shrubs (Solanum sessiliflorum Dunal) have drawn the attention of researchers for their biological versatility (preferential heliophilous or facultative ombrophilous shrubs), their capacity to grow in upland or lowland areas, and the good technological quality of their fruits for the food industry. The aim of this study was to verify physicochemical changes in cubiu fruits during maturation. The fruits were harvested from the experimental station of olericulture of the Instituto Nacional de Pesquisas da Amazônia (INPA), Brazil. The analyses were performed in whole cubiu fruits (peel, pulp, and placenta) at four traditional ripening stages (green, turning, ripe, and fully ripe) for the determination of weight, moisture, total solids, total carotenoids, proteins, lipids, and ash. Cubiu fruits showed large weight variation, with amodal distribution. The ripe stage was critical to maintain moisture, and from that stage on, water loss became evident. The lipids increased steadily over the four ripening stages, maintaining, however, insignificant calorie content. Total carotenoids, proteins, and ash reached the maximum level at the fully ripe stage. With the exception of weight and moisture, all physicochemical changes exhibited the same general behavior, i.e. they increased as the fruits ripened at the four investigated stages.

Effect of process variables on the osmotic dehydration of star-fruit slices

The objective of this work was to study the effect of blanching and the influence of temperature, solution concentration, and the initial fruit:solution ratio on the osmotic dehydration of star-fruit slices. For blanching, different concentrations of citric and ascorbic acids were studied. The samples immersed in 0.75% citric acid presented little variation in color in relation to the fresh star-fruit. Osmotic dehydration was carried out in an incubator with orbital shaking, controlled temperature, and constant shaking at 120 rpm. The influence of process variables was studied in trials defined by a complete 23 central composite design. In general, water loss and solids gain were positively influenced by temperature and by solution concentration. Nevertheless, lower temperatures reduced water loss throughout the osmotic dehydration process. An increase in the amount of dehydrating solution (initial fruit:solution ratio) slightly influenced the evaluated responses. The process carried out at 50 ºC with a solution concentration of 50% resulted in a product with lower solids gain and greater water loss. Under these conditions, blanching minimized the effect of the osmotic treatment on star-fruit browning, and therefore the blanched fruits showed little variation in color in relation to the fresh fruit.

Chemical, biochemical, and microbiological aspects of chitosan quaternary salt as active coating on sliced apples

The biocompatibility of chitosan and chitosan quaternary salt coatings was evaluated for use as edible coatings for sliced apple. Measurement of water loss, color change, and fungal growth appearance were monitored as a function of time. A significant brownish effect was observed on chitosan coated slices, varying greatly from L* = 76.5 and Hue angle = 95.9° (t = 0) to L* = 45.3 and Hue angle = 69.8° (t = 3 days), whilst for TMC coated samples the variation was considerable lower (L* = 74.1; Hue angle = 95.0°) to (L* = 67.0; Hue angle = 83.8°) within the same period. The hydrosoluble derivative N,N,N-trimethylchitosan demonstrated good antifungal activity against P. expansum although highly dependent on the polymer properties such as degree of quaternization. The most efficient formulation was that prepared from derivative having a degree of quaternization of 45%, high solubility, and high viscosity. This formulation restrained fungus spreading up to 30%, while for the control it reached almost 80% of the total assessed surfaces during 7 days of storage.

Analysis of applicability of Peleg model to the cooking-infusion of mackerel (Scomber japonicus) slices

Mass transfer kinetics in osmotic dehydration is usually modeled by Fick's law, empirical models and probabilistic models. The aim of this study was to determine the applicability of Peleg model to investigate the mass transfer during osmotic dehydration of mackerel (Scomber japonicus) slices at different temperatures. Osmotic dehydration was performed on mackerel slices by cooking-infusion in solutions with glycerol and salt (a w = 0.64) at different temperatures: 50, 70, and 90 ºC. Peleg rate constant (K1) (h(g/gdm)-1) varied with temperature variation from 0.761 to 0.396 for water loss, from 5.260 to 2.947 for salt gain, and from 0.854 to 0.566 for glycerol intake. In all cases, it followed the Arrhenius relationship (R²>0.86). The Ea (kJ / mol) values obtained were 16.14; 14.21, and 10.12 for water, salt, and glycerol, respectively. The statistical parameters that qualify the goodness of fit (R²>0.91 and RMSE<0.086) indicate promising applicability of Peleg model.

Biochemical and color changes of fresh-cut melon (Cucumis melo L. cv. Galia) treated with UV-C

The importance of minimally processed commodities in the retail groceries of most developed countries has been rising continuously during the last decades. Cantaloupe melon is used more than any other fruit in fresh-cut processing. Ultraviolet (UV) light has been extensively used to simulate biological stres in plants and for determining resistance mechanisms of plant tissues. In this study the effect of ultraviolet irradiation on some properties of fresh-cut cantalope melon was determined during storage. Freshly cut cantalope melons cubes treated with ultraviolet irradiation at the doses of 1, 2 or 3 min before storage, and then placed in a cold room at 5±1°C temperature and 85-90% RH. Hue angle values of control group is low compared to UV-C treated samples, whereas L values of is high. EL of UV treated samples higher than those of control group. Total soluble solids of fresh-cut melon samples in UC3 treatment increased during storage. The results indicate that UV-C treatments on fresh-cut cantaloupe melon cubes increased total soluble solids independently from water loss.

The role of Stl1p in glycerol accumulation in osmotically stressed icewine yeast Saccharomyces cerevisiae K1 V1116

The high sugar concentration in Icewine juice exerts hyperosmotic stress in the wine yeast causing water loss and cell shrinkage. To counteract the dehydration, yeast synthesize and accumulate glycerol as an internal osmolyte. In a laboratory strain of S. cerevisiae, STLl encodes for Stllp, an H+ /glycerol symporter that is glucose inactivated, but induced upon hyperosmotic stress. STLl, was found to be a highly upregulated gene in Icewine fermenting cells and its expression was 25-fold greater than in yeast cells fermenting diluted Icewine juice, making it one of the most differentially expressed genes between the two fermentation conditions. In addition, Icewine fermenting cells showed a two-fold higher glycerol production in the wine compared to yeast fermenting diluted Icewine juice. We proposed that Stllp is (1) active during Icewine fermentation and is not glucose inactivated and (2) its activity contributes to the limited cell growth observed during Icewine fermentation as a result of the dissipation of the plasma membrane proton gradient. To measure the contribution ofStl1p in active glycerol transport (energy dependent) during Icewine fermentation, we first developed an Stllp-dependent (14C]glycerol uptake assay using a laboratory strain of S. cerevisiae (BY 4742 and LiSTLl) that was dependent on the plasma membrane proton gradient and therefore energy-dependent. Wine yeast K1-Vll16 was also shown to have this energy dependent glycerol uptake induced under salt stress. The expression of STLl and Stllp activity were compared between yeast cells harvested from Icewine and diluted Icewine fermentations. Northern blot analysis revealed that STLl was expressed in cells fermenting Icewine juice but not expressed under the diluted juice conditions. Glycerol uptake by cells fermenting Icewine juice was not significantly different than cells fermenting diluted Icewine juice on day 4 and day 7 of Vidal and Riesling fermentations respectively, despite encountering greater hyperosmotic stress. Furthermore, energy- dependent glycerol uptake was not detected under either fermentation conditions. Because our findings show that active glycerol uptake was not detected in yeast cells harvested from Icewine fermentation, it is likely that Stllp was glucose inactivated despite the hyperosmotic stress induced by the Icewine juice and therefore did not play a role in active glycerol uptake during Icewine fermentation.