A human skin equivalent burn model to study the effect of a nanocrystalline silver dressing on wound healing.

In this study, a deep burn wound model was established using a 3D human skin equivalent (HSE) model and this was compared to native skin. HSEs were constructed from dermis derived from abdominoplasty/breast surgery and this dermal template was seeded with primary keratinocytes and fibroblasts. The HSE model was structurally similar to native skin with a stratified and differentiated epidermis. A contact burn (60 °C, 80 °C, 90 °C) was applied with a modified soldering iron and wounds were observed at day 1 and 7 after burn. The HSEs demonstrated re-growth with keratinocyte proliferation and formation of a neo-epidermis after burn injury, whereas the ex vivo native skin did not. To assess the suitability of the 3D HSE model for penetration and toxicity studies, a nanocrystalline silver dressing was applied to the model for 7 days, with and without burn injury. The effect of silver on skin re-growth and its penetration and subcellular localization was assessed in HSEs histologically and with laser ablation-inductively coupled plasma mass spectrometry (LA-ICPMS). The silver treatment delayed or reduced skin re-growth, and silver particles were detected on the top of the epidermis, and within the papillary dermis. This novel in vitro 3D multicellular deep burn wound model is effective for studying the pathology and treatment of burn wound injury and is suitable for penetration and toxicity studies of wound healing treatments.

Deposits from giant floods in Gale crater and their implications for the climate of early Mars.

This study reports in-situ sedimentologic evidence of giant floods in Gale crater, Mars, during the Noachian Period. Features indicative of floods are a series of symmetrical, 10 m-high gravel ridges that occur in the Hummocky Plains Unit (HPU). Their regular spacing, internal sedimentary structures, and bedload transport of fragments as large as 20 cm suggest that these ridges are antidunes: a type of sedimentary structure that forms under very strong flows. Their 150 m wavelength indicates that the north-flowing water that deposited them was at least 24 m deep and had a minimum velocity of 10 m/s. Floods waned rapidly, eroding antidune crests, and re-deposited removed sediments as patches on the up-flow limbs and trough areas between these ridges forming the Striated Unit (SU). Each patch of the SU is 50-200 m wide and long and consists of 5-10 m of south-dipping layers. The strike and dip of the SU layers mimic the attitude of the flank of the antidune on which they were deposited. The most likely mechanism that generated flood waters of this magnitude on a planet whose present-day average temperature is - 60 °C was the sudden heat produced by a large impact. The event vaporized frozen reservoirs of water and injected large amounts of CO2 and CH4 from their solid phases into the atmosphere. It temporarily interrupted a cold and dry climate and generated a warm and wet period. Torrential rainfall occurred planetwide some of which entered Gale crater and combined with water roaring down from Mt. Sharp to cause gigantic flash floods that deposited the SU and the HPU on Aeolis Palus. The warm and wet climate persisted even after the flooding ended, but its duration cannot be determined by our study.

Characteristics and roles of extracellular vesicles released by epidermal keratinocytes.

Keratinocytes, which constitute 90% of the cells in the epidermis of the skin, have been demonstrated to communicate with other skin cells such as fibroblasts, melanocytes and immune cells through extracellular vesicles (EVs). This communication is facilitated by the enriched EV biomolecular cargo which regulates multiple biological processes within skin tissue, including cell proliferation, cell migration, anti-apoptosis, pigmentation transfer and extracellular matrix remodelling. This review will provide an overview of the current literature and advances in the field of keratinocyte-derived EV research with particular regard to the interactions and communication between keratinocytes and other skin cells, mediated by EVs and EV components. Importantly, this information may shed some light on the potential for keratinocyte-derived EVs in future biomedical studies.

Attenuated kallikrein-related peptidase activity disrupts desquamation and leads to stratum corneum thickening in human skin equivalent models.

BACKGROUND: Epidermal homeostasis is maintained through the balance between keratinocyte proliferation, differentiation and desquamation; however, human skin equivalent (HSE) models are known to differentiate excessively. In native tissue, proteases such as kallikrein-related peptidase (KLK) 5 and KLK7 cleave the extracellular components of corneodesmosomes; proteins corneodesmosin, desmocollin 1 and desmoglein 1, loosening the cellular connections and enabling desquamation. The actions of KLK7 are tightly controlled by protease inhibitors, skin-derived antileucoproteinase (SKALP) and lymphoepithelial Kazal-type-related inhibitor (LEKTI), which also inhibits KLK5, localizing protease activity to the stratum corneum. OBJECTIVES: To investigate the mechanisms that inhibit the desquamation cascade in HSE models. METHODS: Human skin tissue and HSE models were investigated using gene microarray, real-time polymerase chain reaction (PCR), immunohistochemistry and Western blot analysis to examine key components of the desquamation pathway. To elucidate proteolytic activity in HSEs and native skin, in situ and gel zymography was performed. RESULTS: Histological analysis indicated that HSE models form a well-organized epidermis, yet develop an excessively thick and compact stratum corneum. Gene microarray analysis revealed that the desquamation cascade was dysregulated in HSE models and this was confirmed using real-time PCR and immunohistochemistry. Immunohistochemistry and Western blot indicated overexpression of LEKTI and SKALP in HSEs. Although KLK7 was also highly expressed in HSEs, zymography indicated that protease activation and activity was lower than in native skin. CONCLUSIONS: These findings demonstrate that stratum corneum thickening is due to inhibited KLK5 and KLK7 activation and a subsequent lack of corneodesmosome degradation in the HSE model epidermis.

Deposition, exhumation, and paleoclimate of an ancient lake deposit, Gale crater, Mars.

The landforms of northern Gale crater on Mars expose thick sequences of sedimentary rocks. Based on images obtained by the Curiosity rover, we interpret these outcrops as evidence for past fluvial, deltaic, and lacustrine environments. Degradation of the crater wall and rim probably supplied these sediments, which advanced inward from the wall, infilling both the crater and an internal lake basin to a thickness of at least 75 meters. This intracrater lake system probably existed intermittently for thousands to millions of years, implying a relatively wet climate that supplied moisture to the crater rim and transported sediment via streams into the lake basin. The deposits in Gale crater were then exhumed, probably by wind-driven erosion, creating Aeolis Mons (Mount Sharp).

Ancient aqueous environments at Endeavour crater, Mars.

Opportunity has investigated in detail rocks on the rim of the Noachian age Endeavour crater, where orbital spectral reflectance signatures indicate the presence of Fe(+3)-rich smectites. The signatures are associated with fine-grained, layered rocks containing spherules of diagenetic or impact origin. The layered rocks are overlain by breccias, and both units are cut by calcium sulfate veins precipitated from fluids that circulated after the Endeavour impact. Compositional data for fractures in the layered rocks suggest formation of Al-rich smectites by aqueous leaching. Evidence is thus preserved for water-rock interactions before and after the impact, with aqueous environments of slightly acidic to circum-neutral pH that would have been more favorable for prebiotic chemistry and microorganisms than those recorded by younger sulfate-rich rocks at Meridiani Planum.

The highly abundant urinary metabolite urobilin interferes with the bicinchoninic acid assay.

Estimation of total protein concentration is an essential step in any protein- or peptide-centric analysis pipeline. This study demonstrates that urobilin, a breakdown product of heme and a major constituent of urine, interferes considerably with the bicinchoninic acid (BCA) assay. This interference is probably due to the propensity of urobilin to reduce cupric ions (Cu(2+)) to cuprous ions (Cu(1+)), thus mimicking the reduction of copper by proteins, which the assay was designed to do. In addition, it is demonstrated that the Bradford assay is more resistant to the influence of urobilin and other small molecules. As such, urobilin has a strong confounding effect on the estimate of total protein concentrations obtained by BCA assay and thus this assay should not be used for urinary protein quantification. It is recommended that the Bradford assay be used instead.

Stratum basale keratinocyte expression of the cell-surface glycoprotein CDCP1 during epidermogenesis and its role in keratinocyte migration.

BACKGROUND: Epidermogenesis and epidermal wound healing are tightly regulated processes during which keratinocytes must migrate, proliferate and differentiate. Cell-to-cell adhesion is crucial to the initiation and regulation of these processes. CUB-domain-containing protein (CDCP)1 is a transmembrane glycoprotein that is differentially tyrosine phosphorylated during changes in cell adhesion and survival signalling, and is expressed by keratinocytes in native human skin, as well as in primary cultures. OBJECTIVES: To investigate the expression of CDCP1 during epidermogenesis and its role in keratinocyte migration. METHODS: We examined both human skin tissue and an in vitro three-dimensional human skin equivalent model to examine the expression of CDCP1 during epidermogenesis. To examine the role of CDCP1 in keratinocyte migration we used a function-blocking anti-CDCP1 antibody and a real-time Transwell™ cell migration assay. RESULTS: Immunohistochemical analysis indicated that in native human skin CDCP1 is expressed in the stratum basale and stratum spinosum. In contrast, during epidermogenesis in a three-dimensional human skin equivalent model, CDCP1 was expressed only in the stratum basale, with localization restricted to the cell-cell membrane. No expression was detected in basal keratinocytes that were in contact with the basement membrane. Furthermore, an anti-CDCP1 function-blocking antibody was shown to disrupt keratinocyte chemotactic migration in vitro. CONCLUSIONS: These findings delineate the expression of CDCP1 in human epidermal keratinocytes during epidermogenesis and demonstrate that CDCP1 is involved in keratinocyte migration.

Ancient impact and aqueous processes at Endeavour Crater, Mars.

The rover Opportunity has investigated the rim of Endeavour Crater, a large ancient impact crater on Mars. Basaltic breccias produced by the impact form the rim deposits, with stratigraphy similar to that observed at similar-sized craters on Earth. Highly localized zinc enrichments in some breccia materials suggest hydrothermal alteration of rim deposits. Gypsum-rich veins cut sedimentary rocks adjacent to the crater rim. The gypsum was precipitated from low-temperature aqueous fluids flowing upward from the ancient materials of the rim, leading temporarily to potentially habitable conditions and providing some of the waters involved in formation of the ubiquitous sulfate-rich sandstones of the Meridiani region.

Pyroclastic activity at Home Plate in Gusev Crater, Mars.

Home Plate is a layered plateau in Gusev crater on Mars. It is composed of clastic rocks of moderately altered alkali basalt composition, enriched in some highly volatile elements. A coarsegrained lower unit lies under a finer-grained upper unit. Textural observations indicate that the lower strata were emplaced in an explosive event, and geochemical considerations favor an explosive volcanic origin over an impact origin. The lower unit likely represents accumulation of pyroclastic materials, whereas the upper unit may represent eolian reworking of the same pyroclastic materials.

Assessment of Mars Exploration Rover landing site predictions.

Comprehensive analyses of remote sensing data during the three-year effort to select the Mars Exploration Rover landing sites at Gusev crater and at Meridiani Planum correctly predicted the atmospheric density profile during entry and descent and the safe and trafficable surfaces explored by the two rovers. The Gusev crater site was correctly predicted to be a low-relief surface that was less rocky than the Viking landing sites but comparably dusty. A dark, low-albedo, flat plain composed of basaltic sand and haematite with very few rocks was expected and found at Meridiani Planum. These results argue that future efforts to select safe landing sites based on existing and acquired remote sensing data will be successful. In contrast, geological interpretations of the sites based on remote sensing data were less certain and less successful, which emphasizes the inherent ambiguities in understanding surface geology from remotely sensed data and the uncertainty in predicting exactly what materials will be available for study at a landing site.

Pancam multispectral imaging results from the Opportunity Rover at Meridiani Planum.

Panoramic Camera (Pancam) images from Meridiani Planum reveal a low-albedo, generally flat, and relatively rock-free surface. Within and around impact craters and fractures, laminated outcrop rocks with higher albedo are observed. Fine-grained materials include dark sand, bright ferric iron-rich dust, angular rock clasts, and millimeter-size spheroidal granules that are eroding out of the laminated rocks. Spectra of sand, clasts, and one dark plains rock are consistent with mafic silicates such as pyroxene and olivine. Spectra of both the spherules and the laminated outcrop materials indicate the presence of crystalline ferric oxides or oxyhydroxides. Atmospheric observations show a steady decline in dust opacity during the mission. Astronomical observations captured solar transits by Phobos and Deimos and time-lapse observations of sunsets.

Pancam multispectral imaging results from the Spirit Rover at Gusev Crater.

Panoramic Camera images at Gusev crater reveal a rock-strewn surface interspersed with high- to moderate-albedo fine-grained deposits occurring in part as drifts or in small circular swales or hollows. Optically thick coatings of fine-grained ferric iron-rich dust dominate most bright soil and rock surfaces. Spectra of some darker rock surfaces and rock regions exposed by brushing or grinding show near-infrared spectral signatures consistent with the presence of mafic silicates such as pyroxene or olivine. Atmospheric observations show a steady decline in dust opacity during the mission, and astronomical observations captured solar transits by the martian moons, Phobos and Deimos, as well as a view of Earth from the martian surface.

Effect of Lactobacillus plantarum 299v on colonic fermentation and symptoms of irritable bowel syndrome.

A number of recent clinical trials have promoted the use of probiotic bacteria as a treatment for irritable bowel syndrome (IBS). The recent demonstration of abnormal colonic fermentation in some patients with this condition provides an opportunity for the objective assessment of the therapeutic value of these bacteria. This study was designed to investigate the effects of Lactobacillus plantarum 299V on colonic fermentation. We conducted a double-blind, placebo-controlled, cross-over, four-week trial of Lactobacillus plantarum 299V in 12 previously untreated patients with IBS. Symptoms were assessed daily by a validated composite score and fermentation by 24-hr indirect calorimetry in a 1.4-m3 canopy followed by breath hydrogen determination for 3 hr after 20 ml of lactulose. On placebo, the median symptom score was 8.5 [6.25-11.25 interquartile range (IQR)], the median maximum rate of gas production was 0.55 ml/min (0.4-1.1 IQR), and the median hydrogen production was 189.7 ml/24 hr (118.3-291.1 IQR). On Lactobacillus plantarum 299V the median symptom score was 8 (6.75-13.5 IQR), the median maximum rate of gas production 0.92 ml/min (0.45-1.5 IQR), and the median hydrogen production 208.2 ml/24 hr (146-350.9 IQR). There was no significant difference. Breath hydrogen excretion after lactulose was reduced by the probiotic (median at 120 min, 6 ppm; placebo, 17 ppm; P = 0.019). In conclusion, Lactobacillus plantarum 299V in this study did not appear to alter colonic fermentation or improve symptoms in patients with the irritable bowel syndrome.

Irritable bowel syndrome: is the search for lactose intolerance justified?

OBJECTIVES: To determine if confirmation of hypolactasia offers any benefit to the dietary treatment of patients with irritable bowel syndrome (IBS). METHODS: One hundred and twenty-two consecutive IBS patients (37 male, 85 female) were given lactose hydrogen breath tests (LHBT). Those with positive LHBT followed a low lactose diet for 3 weeks. Those improving on the diet were given double-blind, placebo-controlled challenges (DBPCC) with 5 g, 10 g and 15 g of lactose and a placebo, to confirm lactose intolerance. Those who did not respond to the low lactose diet followed either an exclusion or low fibre diet. Symptoms scores were kept prior to the LHBT, 8 h post-LHBT and daily whilst following any dietary change. Patients with negative LHBT returned to clinic and subsequent dietary interventions were recorded. RESULTS: LHBT was positive in 33/122 (27%) IBS patients. Syrr otom scores prior to LHBT were not significantly different between the two groups, but after LHBT the symptoms in the positive group were significantly worse. Twenty-three patients followed a low-lactose diet of which only nine (39%) improved. Six who did not improve followed an exclusion diet, three improved and all were intolerant of milk. Three tried a low fibre diet with two improving. DBPCC were inconclusive. In the negative LHBT group 35 agreed to try a diet and 24 improved (69%). Eight were intolerant of cow's milk. CONCLUSIONS: Use of a low lactose diet was disappointing in IBS patients with lactose malabsorption. Food intolerance was demonstrated in IBS patients with positive or negative LHBT and milk was identified as a problem in both groups. DBPCC were inconclusive. There appears to be little advantage in trying to separate patients who malabsorb lactose from others with IBS.

Pharmacokinetic-pharmacodynamic modelling in the early development phase of anti-psychotics: a comparison of the effects of clozapine, S 16924 and S 18327 in the EEG model in rats.

1. The use of pharmacokinetic/pharmacodynamic (PK/PD) analysis in early compound development was investigated in the rat for two developmental anti-psychotic compounds with clozapine as a positive control. 2. Three plasma samples were collected from each of eight animals according to a pre-defined sampling matrix allowing a total of 12 time points for PK analysis. Quantitative electroencephalography (QEEG), particularly the theta and beta frequencies, was used as a measurement of pharmacological effect. 3. PK/KD modelling of the sparse PK data available relative to a rich set of PD data was achieved using a population approach in NONMEM (IV). Individual PK parameter estimates were incorporated into a PK/PD model. 4. Qualitative EEG changes in rat and human were similar for clozapine, but different for the two developmental compounds, suggesting that changes in these PD parameters may not be specifically related to the anti-psychotic activity. 5. Although no definitive data are available concerning the signal specificity of EEG frequency bands with respect to dopaminergic or serotonergic receptor activity, qualitative and quantitative differences seen in EEG parameters are likely to result from the multiple receptor occupancy for these compounds. 6. The results confirm the value of population PK/PD modelling in conjunction with sparse sampling to enable determination of concentration effect relationships in the pre-clinical development programme of CNS-active drugs.

Effects of nasal O2 on sleep-related disordered breathing in ambulatory patients with stable heart failure.

OBJECTIVE: The purpose of this study was 1) to determine the effects of nasal O2 on periodic breathing, arterial oxyhemoglobin desaturation and nocturnal ventricular arrhythmias in patients with heart failure and 2) determine the characteristics of patients whose periodic breathing will be reversed by O2 administration; our hypothesis was that patients with more severe periodic breathing and desaturation, will respond more favorably to oxygen. DESIGN: Prospective study. SETTING: Referral sleep laboratory of a Department of Veterans Affairs Medical Center. PARTICIPANTS: 36 ambulatory male patients with heart failure whose initial polysomnograms showed periodic breathing with fifteen or more episodes of apnea (A) and hypopnea (H) per hour (AH index, AHI) were treated with nasal O2 during the subsequent full night polysomnography. INTERVENTIONS: Oxygen. MEASUREMENTS AND RESULTS: Arterial blood gases and hydrogen ion concentrations were measured, and cardiac radionuclide ventriculography, Holter monitoring, and polysomnography were done. The studies were scored blindly. Treatment with O2 resulted in a significant reduction in AHI (49+/-19 vs 29+/-29, means+/-SD), central apnea index (28+/-23 vs 13+/-18 per hour), and the percent of total sleep time below an arterial oxyhemoglobin saturation of 90% (23+/-21% vs 0.8+/-2.3%). In spite of virtual normalization of saturation with O2 therapy, the number of ventricular arrhythmias during sleep did not change significantly. In 39% of the patients (14 out of 36), O2 therapy resulted in reversal of central sleep apnea (defined by a reduction in AHI to less than 15/hr). In this group, the AHI decreased by 78% which was significantly (p=0.0001) more than improved (22%) in AHI of the remaining patients (n=22). The main differences between baseline characteristics of the two groups was a significantly higher mean PaCO2 in patients who did respond fully to O2 (39.3+/-5.4 vs 36.1+/-4.2 mm Hg, p=0.03). In both groups, however, O2 administration resulted in significant and similar improvement in arterial oxyhemoglobin saturation (saturation <90%, percent total sleep time 0.1+/-0.3% vs 1+/-3%). CONCLUSION: In patients with stable heart failure, administration of nasal O2 significantly improves periodic breathing and virtually eliminates clinically significant arterial oxyhemoglobin desaturation. The beneficial effects of O2, however, may be modulated by the level of arterial PCO2. Acute O2 therapy has important benefits on sleep apnea and nocturnal arterial oxyhemoglobin desaturation in heart failure patients. Long term benefits of O2 therapy in heart failure and sleep apnea need to be determined.

Sleep apnea in 81 ambulatory male patients with stable heart failure. Types and their prevalences, consequences, and presentations.

BACKGROUND: Heart failure is a highly prevalent disorder that continues to be associated with repeated hospitalizations, high morbidity, and high mortality. Sleep-related breathing disorders with repetitive episodes of asphyxia may adversely affect heart function. The main aims of this study were to determine the prevalence, consequences, and differences in various sleep-related breathing disorders in ambulatory male patients with stable heart failure. METHODS AND RESULTS: This article reports the results of a prospective study of 81 of 92 eligible patients with heart failure and a left ventricular ejection fraction < 45%. There were 40 patients without (hourly rate of apnea/hypopnea, 4 +/- 4; group 1) and 41 patients with (51% of all patients; hourly rate of apnea/hypopnea, 44 +/- 19; group 2) sleep apnea. Sleep disruption and arterial oxyhemoglobin desaturation were significantly more severe and the prevalence of atrial fibrillation (22% versus 5%) and ventricular arrhythmias were greater in group 2 than in group 1. Forty percent of all patients had central sleep apnea, and 11% had obstructive sleep apnea. The latter patients had significantly greater mean body weight (112 +/- 30 versus 75 +/- 16 kg) and prevalence of habitual snoring (78% versus 28%). However, the hourly rate of episodes of apnea and hypopnea (36 +/- 10 versus 47 +/- 21), episodes of arousal (20 +/- 14 versus 23 +/- 11), and desaturation (lowest saturation, 72 +/- 11% versus 78 +/- 12%) were similar in patients with these different types of apnea. CONCLUSIONS: Fifty-one percent of male patients with stable heart failure suffer from sleep-related breathing disorders: 40% from central and 11% from obstructive sleep apnea. Both obstructive and central types of sleep apnea result in sleep disruption and arterial oxyhemoglobin desaturation. Patients with sleep apnea have a high prevalence of atrial fibrillation and ventricular arrhythmias.

Overview of the Mars Pathfinder mission and assessment of landing site predictions.

Chemical analyses returned by Mars Pathfinder indicate that some rocks may be high in silica, implying differentiated parent materials. Rounded pebbles and cobbles and a possible conglomerate suggest fluvial processes that imply liquid water in equilibrium with the atmosphere and thus a warmer and wetter past. The moment of inertia indicates a central metallic core of 1300 to 2000 kilometers in radius. Composite airborne dust particles appear magnetized by freeze-dried maghemite stain or cement that may have been leached from crustal materials by an active hydrologic cycle. Remote-sensing data at a scale of generally greater than approximately 1 kilometer and an Earth analog correctly predicted a rocky plain safe for landing and roving with a variety of rocks deposited by catastrophic floods that are relatively dust-free.

Results from the Mars Pathfinder camera.

Images of the martian surface returned by the Imager for Mars Pathfinder (IMP) show a complex surface of ridges and troughs covered by rocks that have been transported and modified by fluvial, aeolian, and impact processes. Analysis of the spectral signatures in the scene (at 440- to 1000-nanometer wavelength) reveal three types of rock and four classes of soil. Upward-looking IMP images of the predawn sky show thin, bluish clouds that probably represent water ice forming on local atmospheric haze (opacity approximately 0.5). Haze particles are about 1 micrometer in radius and the water vapor column abundance is about 10 precipitable micrometers.