RPGRIP1L is required for stabilizing epidermal keratinocyte adhesion through regulating desmoglein endocytosis.

Cilia-related proteins are believed to be involved in a broad range of cellular processes. Retinitis pigmentosa GTPase regulator interacting protein 1-like (RPGRIP1L) is a ciliary protein required for ciliogenesis in many cell types, including epidermal keratinocytes. Here we report that RPGRIP1L is also involved in the maintenance of desmosomal junctions between keratinocytes. Genetically disrupting the Rpgrip1l gene in mice caused intraepidermal blistering, primarily between basal and suprabasal keratinocytes. This blistering phenotype was associated with aberrant expression patterns of desmosomal proteins, impaired desmosome ultrastructure, and compromised cell-cell adhesion in vivo and in vitro. We found that disrupting the RPGRIP1L gene in HaCaT cells, which do not form primary cilia, resulted in mislocalization of desmosomal proteins to the cytoplasm, suggesting a cilia-independent function of RPGRIP1L. Mechanistically, we found that RPGRIP1L regulates the endocytosis of desmogleins such that RPGRIP1L-knockdown not only induced spontaneous desmoglein endocytosis, as determined by AK23 labeling and biotinylation assays, but also exacerbated EGTA- or pemphigus vulgaris IgG-induced desmoglein endocytosis. Accordingly, inhibiting endocytosis with dynasore or sucrose rescued these desmosomal phenotypes. Biotinylation assays on cell surface proteins not only reinforced the role of RPGRIP1L in desmoglein endocytosis, but also suggested that RPGRIP1L may be more broadly involved in endocytosis. Thus, data obtained from this study advanced our understanding of the biological functions of RPGRIP1L by identifying its role in the cellular endocytic pathway.

Measurement of local magnetic fields in actinide tetrafluorides.

Fluorine-19 magnetic shielding tensors have been measured in a series of actinide tetrafluorides (AnF4) by solid state nuclear magnetic resonance spectroscopy. Tetravalent actinide centers with 0-8 valence electrons can form tetrafluorides with the same monoclinic structure type, making these compounds an attractive choice for a systematic study of the variation in the electronic structure across the 5f row of the Periodic Table. Pronounced deviations from predictions based on localized valence electron models have been detected by these experiments, which suggests that this approach may be used as a quantitative probe of electronic correlations.

A new non-diffusional gas bubble production route in used nuclear fuel: implications for fission gas release, cladding corrosion, and next generation fuel design.

A novel relationship between noble metal phase particles and fission gas bubble production in used nuclear fuel is described. The majority of Te atoms within noble metal phase undergo radioactive decay to form stable Xe within a few hours after particle formation. This results in the production of clusters of Xe atoms contained within the solid metal matrix exhibiting an equivalent gas bubble pressure approaching 1 GPa. These high pressure bubbles are stabilized by the UO2 within the bulk of the fuel. However, when these bubbles form near the fuel/cladding interface, in combination with local and temporal damage caused by fission recoil, they are capable of overcoming the fracture strength of the UO2 and rupturing catastrophically. The force of the resulting bubble rupture is sufficient to eject noble metal phase particles several microns into the cladding. This proposed mechanism explains the observance of noble metal phase in cladding and is consistent with a host of morphological features found near the fuel/cladding interface.

Chemical and Isotopic Characterization of Noble Metal Phase from Commercial UO2 Fuel.

We report elemental and isotopic analysis for the noble metal fission product phase found in irradiated nuclear fuel. The noble metal phase was isolated from three commercial irradiated UO2 fuels by chemically dissolving the UO2 fuel matrix, leaving the noble metal phase as the undissolved residue. Macro amounts of this residue were dissolved using a KOH + KNO3 fusion and then chemically separated into individual elements for analysis by mass spectrometry. Though the composition of this phase has been previously reported, this work is the most comprehensive chemical analysis of the isolated noble metal phase to date. We report both elemental and isotopic abundances of the five major components of the noble metal phase (Mo, Tc, Ru, Rh, Pd). In addition, we report a sixth element present in high quantities in this phase, tellurium. Tellurium appears to be an integral component of noble metal particles.

Fibronectin-derived Epiviosamines enhance PDGF-BB-stimulated human dermal fibroblast migration in vitro and granulation tissue formation in vivo.

Fibronectin (FN) is a multimodular glycoprotein that is a critical component of the extracellular matrix (ECM) anlage during embryogenesis, morphogenesis, and wound repair. Our laboratory has previously described a family of FN-derived peptides collectively called "epiviosamines" that enhance platelet-derived growth factor-BB (PDGF-BB)-driven tissue cell survival, speed burn healing, and reduce scarring. In this study, we used an agarose drop outmigration assay to report that epiviosamines can enhance PDGF-BB-stimulated adult human dermal fibroblast (AHDF) outmigration in a dose-dependent manner. Furthermore, these peptides can, when delivered topically, stimulate granulation tissue formation in vivo. A thiol-derivatized hyaluronan hydrogel cross-linked with polyethyleneglycol diacrylate (PEGDA) was used to topically deliver a cyclized epiviosamine: cP12 and a cyclized engineered variant of cP12 termed cNP8 to porcine, full-thickness, excisional wounds. Both cP12 and cNP8 exhibited dose-dependent increases in granulation tissue formation at day 4, with 600 µM cNP8 significantly enhancing new granulation tissue compared to vehicle alone. In contrast to previous studies, this study suggests that epiviosamines can be used to increase granulation tissue formation without an exogenous supply of PDGF-BB or any cell-binding peptides. Thus, epiviosamine may be useful topically to increase granulation tissue formation in acute wounds.

Blood vessel occlusion with erythrocyte aggregates causes burn injury progression-microvasculature dilation as a possible therapy.

Burns are dynamic injuries characterized by progressive tissue death and continuous severe pain over the course of several days. The extent of burn injury progression determines the ultimate patient outcome. Initial burns result in a central zone of necrosis surrounded by a potentially viable zone of ischemia. Several mechanisms have been proposed to explain injury progression, including oxidant and cytokine stress resulting from either ischemia/reperfusion and/or inflammation, but no proven therapy has emerged. To address the unmet need to limit burn injury progression, the root cause of this process must be delineated. For this reason, we have recently focused on post-burn blood vessel occlusion, currently ascribed to microthrombi. We have found that blood vessel occlusion is initially, mainly and persistently caused by erythrocyte aggregation. Although thermal-induced cell necrosis is the immediate cause of cell death, apoptotic cells from persistent ischemia/anoxia, admixed with inflammatory cells, form a band between viable and nonviable tissue 24 hours later. The delayed cell death by apoptosis appears to be the main attractant for inflammatory cells. Finally, we posit that fibrinogen elevation arising from inflammation provides stimulus for additional erythrocyte aggregation, further extending blood vessel occlusion. In our view this persistent occlusion with resultant prolonged tissue ischemia/anoxia, not ischemia/reperfusion, is the root cause of burn injury progression concomitant with associated severe and persistent pain. Epiviosamines, a new class of peptides, appear to selectively dilate microvasculature, and may provide therapy for burn injury progression.

Vasoactive effect of fibronectin-derived epiviosamine-1 and related peptides in quiescent and stress models.

OBJECTIVE: Following thermal burn injury, plasma fibronectin degrades within the interstitium; one possible product is EVA-1, PSHISKYILRWRPK found within the FNIII1 . EVA-1 ameliorates thermal burn injury progression, and binds to and enhances PDGF-BB in promoting cell metabolism, growth and survival; shorter related peptides lose these abilities. Here we study the effect of EVA-1 and shorter peptides for their vasoactivity under quiescent and stress conditions. METHODS: Using the hamster cheek pouch intravital microscopy model, five EVA-1 related peptides were applied to small arterioles via micropipette (10-16 -10-4 mol L-1 ) in quiescent tissue and after defined stress: nitric oxide or heat. RESULTS: Peak dilation occurred with nanomolar doses (longer peptides) or below (shorter peptides), blocked by propranolol (beta-adrenergic receptor antagonist). Micromolar doses of the same peptides induced only constriction, not antagonized by phentolamine (alpha-adrenergic receptor antagonist). Scrambled variants of two peptides yielded only constriction, suggesting constriction might be due peptide charge. Each stressor caused a left shift in dilation response, blocked by carazolol. CONCLUSIONS: Thus, this important region of FNIII1 contains sequences that have a gradation of biological functions dependent on the length of the peptide sequence, with increased efficacy for dilation following stressors.

Platelet-Derived Growth Factor in Heart Failure.

Defective vascular and cardiomyocyte function are implicated in the development and progression of both heart failure with reduced and preserved ejection fraction. Any treatment option that augments these myocardial processes may therefore be of significant value. The platelet-derived growth factor (PDGF) family is involved in a wide range of growth processes and plays a key role in both regulating angiogenesis and mesenchymal cell development. Thus, PDGF may serve as a potent therapy for heart failure. While numerous animal studies have demonstrated beneficial cardiovascular effects of growth factor therapy, promising laboratory data has not yet translated to effective therapies. In this review, we outline the biological role of PDGF and summarize previous studies that have focused on the cardiovascular effects of normal PDGF signaling, administration of PDGF, and the effects of PDGF on stem cell therapy.

Blood vessel occlusion in peri-burn tissue is secondary to erythrocyte aggregation and mitigated by a fibronectin-derived peptide that limits burn injury progression.

Although vascular occlusion has long been noted in peri-burn tissue, the literature is inconsistent regarding the nature of the occlusion, with articles in the 1940s claiming that erythrocytes were the culprit and in the 1980s-1990s that microthrombi were responsible. To better define the nature of vessel occlusion, we studied two porcine burn models, a hot comb horizontal injury model and a vertical injury progression model. In both cases, tissue from the first two days after burn were stained with hemotoxylin and eosin, or probed for platelets or for fibrinogen/fibrin. Erythrocytes, identified as nonstained, clumped, anuclear, 5 µm cells, occluded most blood vessels (BVs) in both burn models. In contrast, platelet or fibrinogen/fibrin antibodies stained BV occlusions minimally at early time points, and only up to 16% of deep dermal BVs at 48 hours in the hot comb model and up to 7% at 24 hours in the vertical injury progression model. Treatment of animals with a fibronectin-derived peptide (P12), which limits burn injury progression and can dilate peripheral microvasculature, reduced erythrocyte occlusion by at least 50%, speeded healing and reduced scarring. Early erythrocyte aggregation, rather than thrombosis, explains the ineffectiveness of anticoagulants to prevent burn injury progression.

Cutaneous wound healing in aging small mammals: a systematic review.

As the elderly population grows, so do the clinical and socioeconomic burdens of nonhealing cutaneous wounds, the majority of which are seen among persons over 60 years of age. Human studies on how aging effects wound healing will always be the gold standard, but studies have ethical and practical hurdles. Choosing an animal model is dictated by costs and animal lifespan that preclude large animal use. Here, we review the current literature on how aging effects cutaneous wound healing in small animal models and, when possible, compare healing across studies. Using a literature search of MEDLINE/PubMed databases, studies were limited to those that utilized full-thickness wounds and compared the wound-healing parameters of wound closure, reepithelialization, granulation tissue fill, and tensile strength between young and aged cohorts. Overall, wound closure, reepithelialization, and granulation tissue fill were delayed or decreased with aging across different strains of mice and rats. Aging in mice was associated with lower tensile strength early in the wound healing process, but greater tensile strength later in the wound healing process. Similarly, aging in rats was associated with lower tensile strength early in the wound healing process, but no significant tensile strength difference between young and old rats later in healing wounds. From studies in New Zealand White rabbits, we found that reepithelialization and granulation tissue fill were delayed or decreased overall with aging. While similarities and differences in key wound healing parameters were noted between different strains and species, the comparability across the studies was highly questionable, highlighted by wide variability in experimental design and reporting. In future studies, standardized experimental design and reporting would help to establish comparable study groups, and advance the overall knowledge base, facilitating the translatability of animal data to the human clinical condition.

Electron ionization mass spectrum of tellurium hexafluoride.

The electron ionization mass spectrum of tellurium hexafluoride (TeF6) is reported for the first time. The starting material was produced by direct fluorination of Te metal or TeO2 with nitrogen trifluoride. Formation of TeF6 was confirmed through cryogenic capture of the tellurium fluorination product and analysis through Raman spectroscopy. The eight natural abundance isotopes were observed for each of the set of fragment ions: TeF5(+), TeF4(+) TeF3(+), TeF2(+), TeF1(+), and Te(+), Te2(+). A trend in increasing abundance was observed for the odd fluoride bearing ions, TeF1(+) < TeF3(+) < TeF5(+), and a decreasing abundance was observed for the even fragment series, Te(F0)(+) > TeF2(+) > TeF4(+) > TeF6(+), with the molecular ion TeF6(+) not observed at all. Density functional theory based electronic structure calculations were used to calculate optimized ground state geometries of these gas phase species, and their relative stabilities explain the trends in the data and the lack of observed signal for TeF6(+).

Intravenous curcumin efficacy on healing and scar formation in rabbit ear wounds under nonischemic, ischemic, and ischemia-reperfusion conditions.

Curcumin, a spice found in turmeric, is widely used in alternative medicine for its purported anti-inflammatory and antioxidant activities. The goal of this study was to test the curcumin efficacy on rabbit ear wounds under nonischemic, ischemic, and ischemia-reperfusion conditions. Previously described models were utilized in 58 New Zealand White rabbits. Immediately before wounding, rabbits were given intravenous crude or pure curcumin (6 µg/kg, 30 µg/kg, or 60 µg/kg) dissolved in 1% ethanol. Specimens were collected at 7-8 days to evaluate the effects on wound healing and at 28 days to evaluate the effects on hypertrophic scarring. Student's t test was applied to screen difference between any treatment and control group, whereas analysis of variance was applied to further analyze for all treatment groups in aggregate in some specific experiments. Treatment with crude curcumin suggested accelerated wound healing that reached significance for reepithelialization in lower and medium doses and granulation tissue formation in lower dose. Purified curcumin became available and was used for all later experiments. Treatment with pure curcumin suggested accelerated wound healing that reached significance for reepithelialization in lower and medium doses and granulation tissue formation in lower dose. Treatment with pure curcumin significantly promoted nonischemic wound healing in a dose-response fashion compared with controls as judged by increased reepithelialization and granulation tissue formation. Improved wound healing was associated with significant decreases in pro-inflammatory cytokines interleukin (IL)-1 and IL-6 as well as the chemokine IL-8. Curcumin also significantly reduced hypertrophic scarring. The effects of curcumin were examined under conditions of impaired healing including ischemic and ischemia-reperfusion wound healing, and beneficial effects were also seen, although the dose response was less clear. Systemically administrated pure curcumin significantly promotes nonischemic wound healing and reduces hypertrophic scarring. Improvements in wound healing were associated with decreased inflammatory markers in wounds. Further study is needed to optimize dosing in ischemic and ischemia-reperfusion wound healing. In aggregate, the studies strongly support the systemic administration of curcumin to improve wound healing.

Endothelial necrosis at 1 hour postburn predicts progression of tissue injury.

Burn injury progression has not been well characterized at the cellular level. To define burn injury progression in terms of cell death, histopathologic spatiotemporal relationships of cellular necrosis and apoptosis were investigated in a validated porcine model of vertical burn injury progression. Cell necrosis was identified by high mobility group box 1 protein and apoptosis by Caspase 3a staining of tissue samples taken 1 hour, 24 hours, and 7 days postburn. Level of endothelial cell necrosis at 1 hour was predictive of level of apoptosis at 24 hours (Pearson's r = 0.87) and of level of tissue necrosis at 7 days (Pearson's r = 0.87). Furthermore, endothelial cell necrosis was deeper than interstitial cell necrosis at 1 hour (p < 0.001). Endothelial cell necrosis at 1 hour divided the zone of injury progression (Jackson's zone of stasis) into an upper subzone with necrotic endothelial cells and initially viable adnexal and interstitial cells at 1 hour that progressed to necrosis by 24 hours and a lower zone with initially viable endothelial cells at 1 hour but necrosis and apoptosis of all cell types by 24 hours. Importantly, this spatiotemporal series of events and rapid progression resembles myocardial infarction and stroke and implicates mechanisms of these injuries, ischemia, ischemia reperfusion, and programmed cell death in burn progression.

The uni-directional association of atopic dermatitis and rheumatoid arthritis: a systematic review and meta-analysis.

Atopic dermatitis (AD) is a highly pruritic, inflammatory skin disease with a strong immune component. Rheumatoid arthritis (RA) is a systemic autoimmune disease that causes synovitis and destruction of small joints. Researchers have attempted to quantify an association between both diseases with mixed conclusions. This systematic review and meta-analysis will study the association between AD and RA. Additionally, we conducted a systematic review between AD and other arthritic conditions including osteoarthritis (OA), psoriatic arthritis (PsA), and juvenile idiopathic arthritis (JIA). Medline, Web of Science, Cochrane, and EMBASE databases were searched for relevant studies from inception to March 2021. Observational studies examining relationships between AD and arthritic conditions were selected. 2539 studies were screened; nine were found suitable for quantitative analysis, all of which examined AD and RA. All studies had low risk of bias as determined by the Newcastle-Ottawa Scale. Patients with RA did not have significantly increased odds of comorbid AD. These findings were consistent across multiple study designs. However, patients with AD had significantly increased odds of comorbid RA. There were not enough studies identified to perform quantitative analysis between AD and other arthritic conditions. Two studies, one on JIA and one PsA, found no association with AD. Two studies on AD and OA had conflicting results. The present study provides definitive evidence of increased odds of comorbid RA in AD patients. There were no such increased odds of comorbid AD in RA patients. No such association was found between AD and PsA, OA or JIA.

Histopathologic staining of low temperature cutaneous burns: comparing biomarkers of epithelial and vascular injury reveals utility of HMGB1 and hematoxylin phloxine saffron.

Histopathology remains the gold standard for evaluation of burn depth, progression, and healing, but burn literature offers little guidance on the best stains for analysis of these complex and evolving injuries. A battery of histochemical and immunohistochemical stains was compared on adjacent sections to determine the best stains for histopathologic study and imaging of burns. Using a validated porcine model of vertical burn progression, full-thickness cutaneous biopsies were stained using hematoxylin and eosin, Hematoxylin phloxine saffron (HPS), Masson Trichrome, Elastin Von Gieson, Movatt's Pentachrome, vimentin, CD31, KI-67, caspase 3a, and high mobility group box 1. Depth of collagen degeneration, cellular necrosis, apoptosis, and vascular occlusion; and reparative processes of cellular hyperplasia, reepithelialization, and new collagen deposition were measured by ocular microscopy. High mobility group box 1 was superior for necrosis between 1 and 24 hours postburn. Vimentin underestimated necrosis until 48 hours postburn. For overall assessment, hematoxylin and eosin and HPS were comparable, except for analysis of thermally injured collagen, vessel occlusion, erythrocyte extravasation, and polariscopic study of collagen deposition, where HPS was superior. HPS stain offers specific advantages in histopathologic burn analysis. Inexpensive and rapid to produce, HPS allows users to analyze eosinophilic components more precisely than standard hematoxylin and eosin.

High cell density attenuates reactive oxygen species: implications for in vitro assays.

In vitro cell-based assays are an essential and universally used step in elucidation of biological processes as well as in drug development. However, results obtained depend on the validity of protocols used. This statement certainly pertains to in vitro assays of oxidative stress. The holy grail of in vitro models is reliability and predictability of outcomes that relate to a single variable like addition of hydrogen peroxide or xanthine oxidase. Without such validated outcomes, comparison of results among different laboratories is not possible. Achieving this goal requires a thorough understanding of the complex interplay between the cells, their environment, and the experimental assays. Furthermore, as this knowledge is attained, it must be disseminated and used to update and standardize existing protocols. Here, we confirm and extend the effect of pyruvate and cell density on in vitro oxidative stress assays. Cell viability was assessed using a colorimetric assay measuring the reduction of a tetrazolium salt (XTT) into a colored formazan dye. Extracellular hydrogen peroxide concentrations were measured using the foxp3 assay. We confirmed a previously reported finding that pyruvate, a common ingredient in cell culture media, acts as an extracellular scavenger of reactive oxygen species. We also demonstrated that cell density directly correlates with resistance to oxidative stress in tissue culture. It is theorized that the protective effect due to cell density predominantly relates to intracellular factors such as reduced glutathione and extracellular factors such as catalase.

Analysis of a nuclear accident: fission and activation product releases from the Fukushima Daiichi nuclear facility as remote indicators of source identification, extent of release, and state of damaged spent nuclear fuel.

Researchers evaluated radionuclide measurements of environmental samples taken from the Fukushima Daiichi nuclear facility and reported on the Tokyo Electric Power Co. Website following the 2011 tsunami-initiated catastrophe. This effort identified Units 1 and 3 as the major source of radioactive contamination to the surface soil near the facility. Radionuclide trends identified in the soils suggested that: (1) chemical volatility driven by temperature and reduction potential within the vented reactors' primary containment vessels dictated the extent of release of radiation; (2) all coolant had likely evaporated by the time of venting; and (3) physical migration through the fuel matrix and across the cladding wall were minimally effective at containing volatile species, suggesting damage to fuel bundles was extensive. Plutonium isotopic ratios and their distance from the source indicated that the damaged reactors were the major contributor of plutonium to surface soil at the source, decreasing rapidly with distance from the facility. Two independent evaluations estimated the fraction of the total plutonium inventory released to the environment relative to cesium from venting Units 1 and 3 to be ∼0.002-0.004%. This study suggests significant volatile radionuclides within the spent fuel at the time of venting, but not as yet observed and reported within environmental samples, as potential analytes of concern for future environmental surveys around the site. The majority of the reactor inventories of isotopes of less volatile elements like Pu, Nb, and Sr were likely contained within the damaged reactors during venting.

Advantages and Disadvantages of Using Small and Large Animals in Burn Research: Proceedings of the 2021 Research Special Interest Group.

Multiple animal species and approaches have been used for modeling different aspects of burn care, with some strategies considered more appropriate or translatable than others. On April 15, 2021, the Research Special Interest Group of the American Burn Association held a virtual session as part of the agenda for the annual meeting. The session was set up as a pro/con debate on the use of small versus large animals for application to four important aspects of burn pathophysiology: burn healing/conversion, scarring, inhalation injury, and sepsis. For each of these topics, two experienced investigators (one each for small and large animal models) described the advantages and disadvantages of using these preclinical models. The use of swine as a large animal model was a common theme due to anatomic similarities with human skin. The exception to this was a well-defined ovine model of inhalation injury; both of these species have larger airways which allow for incorporation of clinical tools such as bronchoscopes. However, these models are expensive and demanding from labor and resource standpoints. Various strategies have been implemented to make the more inexpensive rodent models appropriate for answering specific questions of interest in burns. Moreover, modeling burn-sepsis in large animals has proven difficult. It was agreed that the use of both small and large animal models has merit for answering basic questions about the responses to burn injury. Expert opinion and the ensuing lively conversations are summarized herein, which we hope will help inform experimental design of future research.