Tape strips in dermatology research.

Tape strips have been used widely in dermatology research as a minimally invasive method to sample the epidermis, avoiding the need for skin biopsies. Most research has focused on epidermal pathology, such as atopic eczema, but there is increasing research into the use of tape strips in other dermatoses, such as skin cancer, and the microbiome. This review summarizes the technique of tape stripping, and discusses which dermatoses have been studied by tape stripping and alternative minimally invasive sampling methods. We review the number of tape strips needed from each patient and the components of the epidermis that can be obtained by tape stripping. With a focus on protein and RNA extraction, we address the techniques used to process tape strips. There is no optimal protocol to extract protein, as this depends on the abundance of the protein studied, its level of expression in the epidermis and its solubility. Many variables can alter the amount of protein obtained from tape strips, which must be standardized to ensure consistency between samples. No study has compared different RNA extraction techniques, but our own experience is that RNA yield is optimized by using 20 tape strips and the use of a cell scraper.

Effects of educational efforts in tribal homes and schools to reduce asthma triggers, symptoms and missed school days.

Native American asthma prevalence has been estimated higher than for the U.S. population average, and uncontrolled asthma results in absence from schools. This study analyzed effects of targeted education campaigns in both homes and schools on reducing asthma triggers and symptoms among 119 children with asthma, who were recruited from Cherokee and Nez Perce communities. The education campaigns were developed in collaboration with the researchers and the tribes, tailoring strategies to reduce asthma triggers adapted to tribal lifestyles. There was a special emphasis in identification of Traditional Ecological Knowledge (TEK) aspects, which were woven into the study plan and implementation. Some 62 study homes were investigated and the parents received targeted education at the beginning of the study, whereas the remaining control homes received the same education at the end of the study. In Cherokee homes, allergen levels were significantly increased in both control and study groups throughout the study. In Nez Perce homes, dog and cockroach allergen levels decreased significantly in the study homes. The parents reported asthma control test (ACT) scores of the children were significantly improved in both study and control groups, whereas respiratory illness days were reduced in the study group. In schools, allergen levels, particularly cat, dog and cockroach allergens, increased throughout the year in both study and control schools. However, high contact surface cleaning effectiveness based on adenosine tri phosphate (ATP) readings was improved in the study schools. It was also found that high contact surface cleaning effectiveness may have effects on students' absence rates tracked by schools. In conclusion, the results indicate partial improvements in parent reported health symptoms, although the improvements could not be definitively attributed to reduction of any specific exposure in the home environment. In the school environment, effective cleaning throughout the flu season could result in decreased absence rates.

Hierarchical Black Hole Mergers in Active Galactic Nuclei.

The origins of the stellar-mass black hole mergers discovered by LIGO/Virgo are still unknown. Here we show that if migration traps develop in the accretion disks of active galactic nuclei (AGNs) and promote the mergers of their captive black holes, the majority of black holes within disks will undergo hierarchical mergers-with one of the black holes being the remnant of a previous merger. 40% of AGN-assisted mergers detected by LIGO/Virgo will include a black hole with mass ≳50M_{⊙}, the mass limit from stellar core collapse. Hierarchical mergers at traps in AGNs will exhibit black hole spins (anti)aligned with the binary's orbital axis, a distinct property from other hierarchical channels. Our results suggest, although not definitively (with odds ratio of ∼1), that LIGO's heaviest merger so far, GW170729, could have originated from this channel.

Differential expression of secreted factors SOSTDC1 and ADAMTS8 cause profibrotic changes in linear morphoea fibroblasts.

BACKGROUND: Linear morphoea (LM) is a rare connective tissue disorder characterized by a line of thickened skin and subcutaneous tissue and can also affect the underlying muscle and bone. Little is known about the disease aetiology, with treatment currently limited to immune suppression, and disease recurrence post-treatment is common. OBJECTIVES: In order to uncover new therapeutic avenues, the cell-intrinsic changes in LM fibroblasts compared with site-matched controls were characterized. METHODS: We grew fibroblasts from site-matched affected and unaffected regions from five patients with LM, we subjected them to gene expression analysis and investigation of SMAD signalling. RESULTS: Fibroblasts from LM lesions showed increased migration, proliferation, altered collagen processing, and abnormally high basal levels of phosphorylated SMAD2, thereby rendering them less responsive to transforming growth factor (TGF)-ß1 and reducing the degree of myofibroblast differentiation, which is a key component of the wound-healing and scarring process in normal skin. Conditioned media from normal fibroblasts could reverse LM-affected fibroblast migration and proliferation, suggesting that the LM phenotype is driven by an altered secretome. Gene array analysis and RNA-Seq indicated upregulation of ADAMTS8 and downregulation of FRAS1 and SOSTDC1. SOSTDC1 knock-down recapitulated the reduced TGF-ß1 responsiveness and LM fibroblast migration, while overexpression of ADAMTS8 induced myofibroblast markers. CONCLUSIONS: We demonstrate that cell-intrinsic changes in the LM fibroblast secretome lead to changes observed in the disease, and that secretome modulation could be a viable therapeutic approach in the treatment of LM.

The reestablishment of microbial communities after surface cleaning in schools.

AIMS: The goal of this study was to quantify the indoor microbiome dynamics of bacterial and fungal communities on school desk surfaces during a cleaning intervention. METHODS AND RESULTS: Quantitative PCR and DNA sequenced-based approaches were employed to describe microbial community dynamics on ten desk surfaces, spread across three schools, located in the Northeast region of the United States. Six samples were taken from each desk, one precleaning, and five postcleaning at 30 min, 1, 3, 7 and 21 days. Cleaning of the desks physically removed c. 50% of bacteria, fungi, and human cells and a full recovery of the surface microbial concentrations occurred within 2-5 days. This recovery period is much shorter than the schools' once per semester cleaning schedule. The dominant source of bacteria and fungi on desks at all time points came from the human microbiome (skin, oral cavity, and gut). More than 50% fungi on desks were members of genera that contain known allergens. CONCLUSIONS: Microbial communities on these school desks are primarily generated and maintained from the deposition of human-associated bacteria and fungi. Current school surface cleaning protocols and cycles may be ineffective at reducing student exposure to fungal allergens and microbes of human origin. SIGNIFICANCE AND IMPACT OF STUDY: Multiple students often share desks in schools. Results on the removal and reestablishment of microbial communities on these surfaces are critical for setting cleaning schedules and practices that effectively interrupt exposure to surface-associated pathogens and allergens.

Improved ethical guidance for the return of results from psychiatric genomics research.

There is an emerging consensus that genomic researchers should, at a minimum, offer to return to individual participants clinically valid, medically important and medically actionable genomic findings (for example, pathogenic variants in BRCA1) identified in the course of research. However, this is not a common practice in psychiatric genetics research. Furthermore, psychiatry researchers often generate findings that do not meet all of these criteria, yet there may be ethically compelling arguments to offer selected results. Here, we review the return of results debate in genomics research and propose that, as for genomic studies of other medical conditions, psychiatric genomics researchers should offer findings that meet the minimum criteria stated above. Additionally, if resources allow, psychiatry researchers could consider offering to return pre-specified 'clinically valuable' findings even if not medically actionable-for instance, findings that help corroborate a psychiatric diagnosis, and findings that indicate important health risks. Similarly, we propose offering 'likely clinically valuable' findings, specifically, variants of uncertain significance potentially related to a participant's symptoms. The goal of this Perspective is to initiate a discussion that can help identify optimal ways of managing the return of results from psychiatric genomics research.

AKT1-mediated Lamin A/C degradation is required for nuclear degradation and normal epidermal terminal differentiation.

Nuclear degradation is a key stage in keratinocyte terminal differentiation and the formation of the cornified envelope that comprises the majority of epidermal barrier function. Parakeratosis, the retention of nuclear material in the cornified layer of the epidermis, is a common histological observation in many skin diseases, notably in atopic dermatitis and psoriasis. Keratinocyte nuclear degradation is not well characterised, and it is unclear whether the retained nuclei contribute to the altered epidermal differentiation seen in eczema and psoriasis. Loss of AKT1 function strongly correlated with parakeratosis both in eczema samples and in organotypic culture models. Although levels of DNAses, including DNase1L2, were unchanged, proteomic analysis revealed an increase in Lamin A/C. AKT phosphorylates Lamin A/C, targeting it for degradation. Consistent with this, Lamin A/C degradation was inhibited and Lamin A/C was observed in the cornified layer of AKT1 knockdown organotypic cultures, surrounding retained nuclear material. Using AKT-phosphorylation-dead Lamin A constructs we show that the retention of nuclear material is sufficient to cause profound changes in epidermal terminal differentiation, specifically a reduction in Loricrin, Keratin 1, Keratin 10, and filaggrin expression. We show that preventing nuclear degradation upregulates BMP2 expression and SMAD1 signalling. Consistent with these data, we observe both parakeratosis and evidence of increased SMAD1 signalling in atopic dermatitis. We therefore present a model that, in the absence of AKT1-mediated Lamin A/C degradation, DNA degradation processes, such as those mediated by DNAse 1L2, are prevented, leading to parakeratosis and changes in epidermal differentiation.

Rab3Gap1 mediates exocytosis of Claudin-1 and tight junction formation during epidermal barrier acquisition.

Epidermal barrier acquisition during late murine gestation is accompanied by an increase in Akt kinase activity and cJun dephosphorlyation. The latter is directed by the Ppp2r2a regulatory subunit of the Pp2a phosphatase. This was accompanied by a change of Claudin-1 localisation to the cell surface and interaction between Occludin and Claudin-1 which are thought to be required for tight junction formation. The aim of this study was to determine the nature of the barrier defect caused by the loss of AKT/Ppp2r2a function. There was a paracellular barrier defect in rat epidermal keratinocytes expressing a Ppp2r2a siRNA. In Ppp2r2a knockdown cells, Claudin-1 was located to the cytoplasm and its expression was increased. Inhibiting cJun phosphorylation restored barrier function and plasma membrane localisation of Claudin-1. Expression of the Rab3 GTPase activating protein, Rab3Gap1, was restored in Ppp2r2a siRNA cells when cJun phosphorylation was inhibited. During normal mouse epidermal development, Claudin-1 plasma membrane localisation and Rab3Gap1 cell surface expression were co-incident with Akt activation in mouse epidermis, strongly suggesting a role of Rab3Gap1 in epidermal barrier acquisition. Supporting this hypothesis, siRNA knockdown of Rab3Gap1 prevented plasma membrane Claudin-1 expression and the formation of a barrier competent epithelium. Replacing Rab3Gap1 in Ppp2r2a knockdown cells was sufficient to rescue Claudin-1 transport to the cell surface. Therefore these data suggest Rab3Gap1 mediated exocytosis of Claudin-1 is an important component of epidermal barrier acquisition during epidermal development.

The mTOR inhibitor rapamycin opposes carcinogenic changes to epidermal Akt1/PKBα isoform signaling.

Epidermal squamous cell carcinoma (SCC) is the most aggressive non-melanoma skin cancer and is dramatically increased in patients undergoing immunosuppression following solid organ transplantation, contributing substantially to morbidity and mortality. Recent clinical studies show that use of the mammalian target of rapamycin (mTOR) inhibitor rapamycin as a post-transplantation immunosuppressive significantly reduces SCC occurrence compared with other immunosuppressives, though the mechanism is not fully understood. We show that rapamycin selectively upregulates epidermal Akt1, while failing to upregulate epidermal Akt2. Rapamycin increases epidermal Akt1 phosphorylation via inhibition of the mTOR complex 1-dependent regulation of insulin receptor substrate-1. Epidermal Akt1 is commonly downregulated in SCC while Akt2 is upregulated. We now demonstrate similar Akt1 downregulation and Akt2 upregulation by ultraviolet (UV) radiation, the most important skin carcinogen. Hence, rapamycin's upregulation of Akt1 signaling could potentially oppose the effects of UV radiation and/or tumor-associated changes on Akt1 signaling. We show in skin culture that rapamycin does enhance restoration of Akt1 phosphorylation in skin recovering from UV radiation, suggesting a mechanism for rapamycin's antitumor activity in epidermis in spite of its efficient immunosuppressive properties.

Circulating cell-free fetal DNA for the detection of RHD status and sex using reflex fetal identifiers.

OBJECTIVE: To determine the sensitivity and specificity of circulating cell-free fetal DNA in determining the fetal RHD status and fetal sex. METHODS: Maternal blood was collected in each trimester of pregnancy from RhD negative nonalloimmunized women. Whole blood was centrifuged, separated into plasma and buffy coat, and frozen at -80°C. DNA analysis was conducted via allele-specific primer extensions for exons 4, 5, and 7 of the RHD gene and for a 37-base pair insertion in exon 4 (RHD pseudogene; psi) three Y-chromosome sequences (SRY, DBY, and TTY2), and an extraction control (TGIFL-like X/Y). RhD serotyping on cord blood and gender assessment of the newborns were entered into a Web-based database. RESULTS: One hundred twenty women were enrolled. The median gestational age at the first venipuncture was 12.4 (range: 10.6-13.9) weeks with 120 samples drawn; 118 samples were drawn at 17.6 (16-20.9) weeks; and 113 samples at 28.7 (27.9-33.9) weeks. Overall accuracy for RHD was 99.1%, 99.1%, and 98.1% for each trimester and was 99.1%, 99.1%, and 100% for fetal sex determination. CONCLUSIONS: Fetal RHD genotyping and sex can be very accurately determined in all three trimesters using circulating cell-free fetal DNA in the maternal circulation.

Occurrence of moisture problems in schools in three countries from different climatic regions of Europe based on questionnaires and building inspections - the HITEA study.

UNLABELLED: The aim of this study was to assess occurrence of dampness and mold in school buildings in three European countries (the Netherlands, Spain, and Finland), representing different climatic regions. An assessment was performed utilizing both questionnaires and on-site building investigations, and the agreement between these two methods was evaluated for validation purposes. On the basis of questionnaire data from a representative sample of schools, different types of moisture problems were reported in 24-47% of all school buildings at the time of the study. Most commonly reported was dampness in the Netherlands, moisture/water damage in Spain, and mold odor in Finland. Subsequently, 20-24 schools per country were selected for on-site inspections by trained staff. The overall agreement between the questionnaire and inspection data was good (kappa-value 0.62), however, with large differences (0.39-0.91) between countries. Extrapolating from the inspection data, the minimum estimates for prevalence of moisture problems in school buildings are 20% in the Netherlands, 41% in Spain, and 24% in Finland. In conclusion, moisture problems (such as moisture damage, dampness, and mold) are relatively common in schools. The occurrence and severity may vary across geographical areas, which can be partly explained by building characteristics. PRACTICAL IMPLICATIONS: On the basis of this study, the prevalence of verified moisture problems in school buildings was highest in Spain, but lower and similar in Finland and the Netherlands. Questionnaire-based surveys can be used to assess moisture problems in school buildings, but because of large variation in agreement with inspection data, the questionnaire needs to be validated by on-site inspections in a subsample of the surveyed buildings.

Association between substandard classroom ventilation rates and students' academic achievement.

UNLABELLED: This study focuses on the relationship between classroom ventilation rates and academic achievement. One hundred elementary schools of two school districts in the southwest United States were included in the study. Ventilation rates were estimated from fifth-grade classrooms (one per school) using CO(2) concentrations measured during occupied school days. In addition, standardized test scores and background data related to students in the classrooms studied were obtained from the districts. Of 100 classrooms, 87 had ventilation rates below recommended guidelines based on ASHRAE Standard 62 as of 2004. There is a linear association between classroom ventilation rates and students' academic achievement within the range of 0.9-7.1 l/s per person. For every unit (1 l/s per person) increase in the ventilation rate within that range, the proportion of students passing standardized test (i.e., scoring satisfactory or above) is expected to increase by 2.9% (95%CI 0.9-4.8%) for math and 2.7% (0.5-4.9%) for reading. The linear relationship observed may level off or change direction with higher ventilation rates, but given the limited number of observations, we were unable to test this hypothesis. A larger sample size is needed for estimating the effect of classroom ventilation rates higher than 7.1 l/s per person on academic achievement. PRACTICAL IMPLICATIONS: The results of this study suggest that increasing the ventilation rates toward recommended guideline ventilation rates in classrooms should translate into improved academic achievement of students. More studies are needed to fully understand the relationships between ventilation rate, other indoor environmental quality parameters, and their effects on students' health and achievement. Achieving the recommended guidelines and pursuing better understanding of the underlying relationships would ultimately support both sustainable and productive school environments for students and personnel.

Fetal blood sampling in baboons (Papio spp.): important procedural aspects and literature review.

BACKGROUND: The baboons (Papio cynocephalus) have similarities with human placentation and fetal development. Fetal blood sampling allows investigators to assess fetal condition at a specific point in gestation as well as transplacental transfer of medications. Unfortunately, assessing fetal status during gestation has been difficult and fetal instrumentation associated with high rate of pregnancy loss. Our objectives are to describe the technique of ultrasound guided cordocentesis (UGC) in baboons, report post-procedural outcomes, and review existing publications. METHODS: This is a procedural paper describing the technique of UGC in baboons. After confirming pregnancy and gestational age via ultrasound, animals participating in approved research protocols that required fetal assessment underwent UGC. RESULTS: We successfully performed UGC in four animals (five samples) using this technique. Animals were sampled in the second and third trimesters with fetal blood sampling achieved by sampling a free cord loop, placental cord insertion site or the intrahepatic umbilical vein. All procedures were without complication and these animals delivered at term. CONCLUSIONS: Ultrasound guided fetal umbilical cord venipuncture is a useful and safe technique to sample the fetal circulation with minimal risk to the fetus or mother. We believe this technique could be used for repeated fetal venous blood sampling in the baboons.