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Three-dimensional human epidermal equivalents (HEEs) are a state-of-the-art organotypic culture model in preclinical investigative dermatology and regulatory toxicology. In this study, we investigated the utility of electrical impedance spectroscopy (EIS) for noninvasive measurement of HEE epidermal barrier function. Our setup comprised a custom-made lid fit with 12 electrode pairs aligned on the standard 24-transwell cell culture system. Serial EIS measurements for 7 consecutive days did not impact epidermal morphology, and readouts showed comparable trends with HEEs measured only once. We determined 2 frequency ranges in the resulting impedance spectra: a lower frequency range termed EISdiffcorrelated with keratinocyte terminal differentiation independent of epidermal thickness and a higher frequency range termed EISSCcorrelated with stratum corneumthickness. HEEs generated from CRISPR/Cas9-engineered keratinocytes that lack key differentiation genes FLG, TFAP2A, AHR, or CLDN1confirmed that keratinocyte terminal differentiation is the major parameter defining EISdiff. Exposure to proinflammatory psoriasis- or atopic dermatitis-associated cytokine cocktails lowered the expression of keratinocyte differentiation markers and reduced EISdiff. This cytokine-associated decrease in EISdiffwas normalized after stimulation with therapeutic molecules. In conclusion, EIS provides a noninvasive system to consecutively and quantitatively assess HEE barrier function and to sensitively and objectively measure barrier development, defects, and repair.

Low Earth Orbit satellites have been used since the 1960s for the most reliable position for reconnaissance in space. CORONA was the first recon satellite put into orbit, in 1960. The new form of war today is spread over valleys, hills and deserts. Historical and current use of satellites is explored.

ABSTRACTObjective: To evaluate the source of employment of athletic trainers (ATs), access to team physicians, and usage of up-to-date (implementation of the most current international sports medicine societies’ position statements and evidence-based literature policy and procedure manuals in secondary schools.Methods: We conducted a cross-sectional study among National Athletic Trainers’ Association (NATA) members. NATA ‘Research Survey Request’ was contacted to obtain 1,000 e-mail addresses of ATs who actively worked in the secondary school setting. We evaluated AT employment, access to team physicians, and usage of up-to-date policy and procedure manuals within different residential areas (urban, suburban, and rural) and school distinctions (public vs. private).Results: Two-hundred ninety-six responses were received (30% response rate). The majority (72%) of ATs reported having an assigned team physician for their school. Approximately one-third (36%) of ATs reported being employed by a hospital, clinic, or outreach facility. Fifty-one percent of ATs reported having a policy and procedure manual that was developed and reviewed with their team physician. There was a significantly greater amount of suburban ATs who reported having an up-to-date policy and procedure manual (urban 45% vs. suburban 60% vs. rural 39%; p = 0.007). Significantly more ATs who were employed in a public school setting reported conducting annual policy and procedure manual reviews than those employed in private school settings (public 63% vs. private 49%; p = 0.045).Conclusions: The majority of ATs have assigned team physicians and a third is employed by a medical group. A high number of ATs reported not having a policy and procedure manual that was developed and reviewed with their team physician. Sports medicine teams within secondary schools, particularly in private schools, should strive to develop and maintain an up-to-date policy and procedure manual that is frequently reviewed with their team physician.

Various culture media are used to propagate keratinocytes (KCs) in vitro. The COVID-19 pandemic resulted in supply chain shortages necessitating substitutions to standard laboratory protocols, which resulted in many laboratories having to use culture media different from those they typically use. We screened available media on the KC line N/TERT2G and found that biological responses varied considerably across three culture media: KC serum-free media, KC growth medium 2, and defined media. We observed qualitative and quantitative differences in proliferation; KCs cultured in defined media had significantly lower proliferative capacity. KC differentiation was assessed by western blot for CLDN1, occludin, cytokeratin-10, and loricrin. Elevated expression of differentiation markers was observed in cells cultured in either KC growth medium 2 or defined media compared with those in cells cultured in KC serum-free media. KC barrier function was measured by transepithelial electrical resistance. KCs cultured in KC growth medium 2 and defined media developed significantly higher transepithelial electrical resistance than those cultured in KC serum-free media, and when treated with IL-4 and IL-13 or IL-17A, we observed variable responses. H&E staining on day 5 -post-differentiation showed greater epithelial thickness in KCs cultured in defined media and KC growth medium 2 than in those cultured in KC serum-free media. These findings show that the choice of culture media impacts the biological response of KCs in a manner that persists through differentiation in the same media.

ABSTRACTEntamoeba histolyticais the causative agent of amoebic dysentery, a worldwide protozoal disease that results in approximately 100,000 deaths annually. The virulence of E. histolyticamay be due to interactions with the host bacterial flora, whereby trophozoites engulf colonic bacteria as a nutrient source. The engulfment process depends on trophozoite recognition of bacterial epitopes that activate phagocytosis pathways. E. histolyticaGPCR-1 (EhGPCR-1) was previously recognized as a putative G-protein-coupled receptor (GPCR) used by Entamoeba histolyticaduring phagocytosis. In the present study, we attempted to characterize EhGPCR-1 by using heterologous GPCR expression in Saccharomyces cerevisiae. We discovered that bacterial lipopolysaccharide (LPS) is an activator of EhGPCR-1 and that LPS stimulates EhGPCR-1 in a concentration-dependent manner. Additionally, we demonstrated that Entamoeba histolyticaprefers to engulf bacteria with intact LPS and that this engulfment process is sensitive to suramin, which prevents the interactions of GPCRs and G-proteins. Thus, EhGPCR-1 is an LPS-recognizing GPCR that is a potential drug target for treatment of amoebiasis, especially considering the well-established drug targeting to GPCRs.

Discusses the plans for lunar explorations. Reasons for returning to the moon; How to get the requirements for the explorations to the moon; How to implement the space exploration.

CRISPR/Cas9 technology is a powerful tool used to alter the genetic landscape of various hosts. This has been exemplified by its success in the transgenic animal world where it has been utilized to develop novel mouse lines modeling numerous disease states. The technology has helped to develop both in vitro and in vivo systems that simulate diseases within the fields of epithelial biology, skin cancer biology, dermatology, and beyond. Importantly, the delivery of the single-guide RNA/Cas9 editing complex to the host cell is key for its success. In this paper, we discuss the various methods that have been utilized as delivery techniques for CRISPR/Cas9 components, the benefits and pitfalls of each, and how successful they have been at genetically modifying epidermal cells. In addition, we acknowledge recent advances in the field of dermatology that have harnessed these methods to better understand epidermal biology, identify potential therapeutic targets, or serve as novel methods to treat disease states.

Small-scale zooplankton swimming behaviors can affect aquatic predator-prey interactions. Difficulties in controlling prey swimming behavior however, have restricted the ability to test hypotheses relating differences in small-scale swimming behavior to frequency of predation by fish. We report here a Virtual Plankton (VP) system that circumvents this problem by allowing the observation of fish “preying"on computer-generated prey images whose size, shape, color and swimming behavior can be precisely controlled. Two experiments were performed in which bluegill sunfish (Lepomis macrochirus) were given a choice of either two VP images, one of which moved twice as fast as the other, or six VP, one of which moved either faster (1.25 x, 1.5 x or 2 x ) or slower (0.5 x) than the other five. Current predator-prey models based on encounter probabilities and prey visibility predict that moving faster increases predation risk and conversely, moving slower decreases predation risk. In agreement with existing predator-prey models, in both experiments, fish chose faster moving VP significantly more often than their slower moving neighbors. Contrary to the predictions of existing models, in the second experiment with six VP, the rate at which fish chose a prey image moving half as fast as the five surrounding images did not differ significantly from the rate predicted by chance(l/6). These results suggest that current fish-zooplankton predation models would benefit by the incorporation of small-scale swimming behavior and assessments of its influence on overall prey visibility.

The pre–and post–contact mating behaviours of Daphnia pulicariaare investigated by direct observations of vertical distributions, swimming behaviours and male–female interactions. Analysis of vertical distributions in a 1 m deep, thermally stratified migration chamber reveals that females were always located in the upper layer of the water column but males exhibited a bimodal depth distribution, in which an individual's depth was a function of body length and water temperature. The observed distributions of males may be the result of several interacting pressures; predation avoidance, life–history optimization, and avoidance of assortative mating. Male swimming behaviour was faster and orthogonal to that of females, which is in agreement with the predictions of encounter–rate maximization models. Video recordings of males and females interacting in a 1–litre vessel showed that males both pursued and contacted other males more often than females. Thus, there was no evidence that Daphniaare able to use water–borne chemical signals to locate and identify potential mates. However, the average duration of male–female contacts (13.8 s) was much longer than those between males (1.6 s), suggesting that males can determine the sex of contacted individuals.Daphniamating behaviour is significantly more complex than previously acknowledged. In contrast to the conventional view of Daphniamales swimming more–or–less randomly and mating with any individual encountered, they exhibit behaviours which increase the potential of mating with females while reducing the risk of predation. Several male behaviours, such as ‘scanning’ and the performance of area–restricted spirals upon encounter, are similar to those reported for some copepods and may be common to zooplankton that lack sophisticated chemosensory abilities. The possibility that Daphniamay also be able to assess such important female attributes as species and reproductive status is discussed.

Epidermal cell models are critical in studying skin biology and diseases. The gold standard used by the scientific community has historically been primary cell cultures from discarded tissue, typically from neonates (foreskin). Although directly applicable to humans, this system suffers from multiple issues, including substantial donor-to-donor variability and a finite number of divisions in culture. As such, we have identified a faithful alternative called N/TERT2G cells. These cells show many of the characteristics of primary cells, including barrier formation, differentiation kinetics and/or protein expression, and pathogenesis. From our observations, this model is highly beneficial to the scientific community and can serve as a reproducible and continuously manipulatable platform in studying skin biology.

Keratinocytes express many pattern recognition receptors that enhance the skin’s adaptive immune response to epicutaneous antigens. We have shown that these pattern recognition receptors are expressed below tight junctions (TJ), strongly implicating TJ disruption as a critical step in antigen responsiveness. To disrupt TJs, we designed peptides inspired by the first extracellular loop of the TJ transmembrane protein CLDN1. These peptides transiently disrupted TJs in the human lung epithelial cell line 16HBE and delayed TJ formation in primary human keratinocytes. Building on these observations, we tested whether vaccinating mice with an epicutaneous influenza patch containing TJ-disrupting peptides was an effective strategy to elicit an immunogenic response. Application of a TJ-disrupting peptide patch resulted in barrier disruption as measured by increased transepithelial water loss. We observed a significant increase in antigen-specific antibodies when we applied patches with TJ-disrupting peptide plus antigen (influenza hemagglutinin) in either a patch-prime or a patch-boost model. Collectively, these observations demonstrate that our designed peptides perturb TJs in human lung as well as human and murine skin epithelium, enabling epicutaneous vaccine delivery. We anticipate that this approach could obviate currently used needle-based vaccination methods that require administration by health care workers and biohazard waste removal.