Gene-based association study of rare variants in children of diverse ancestries implicates TNFRSF21 in the development of allergic asthma.

BACKGROUND: Most genetic studies of asthma and allergy have focused on common variation in individuals primarily of European ancestry. Studying the role of rare variation in quantitative phenotypes and in asthma phenotypes in populations of diverse ancestries can provide additional, important insights into the development of these traits. OBJECTIVE: We sought to examine the contribution of rare variants to different asthma- or allergy-associated quantitative traits in children with diverse ancestries and explore their role in asthma phenotypes. METHODS: We examined whole-genome sequencing data from children participants in longitudinal studies of asthma (n = 1035; parent-identified as 67% Black and 25% Hispanic) to identify rare variants (minor allele frequency < 0.01). We assigned variants to genes and tested for associations using an omnibus variant-set test between each of 24,902 genes and 8 asthma-associated quantitative traits. On combining our results with external data on predicted gene expression in humans and mouse knockout studies, we identified 3 candidate genes. A burden of rare variants in each gene and in a combined 3-gene score was tested for its associations with clinical phenotypes of asthma. Finally, published single-cell gene expression data in lower airway mucosal cells after allergen challenge were used to assess transcriptional responses to allergen. RESULTS: Rare variants in USF1 were significantly associated with blood neutrophil count (P = 2.18 × 10-7); rare variants in TNFRSF21 with total IgE (P = 6.47 × 10-6) and PIK3R6 with eosinophil count (P = 4.10 × 10-5) reached suggestive significance. These 3 findings were supported by independent data from human and mouse studies. A burden of rare variants in TNFRSF21 and in a 3-gene score was associated with allergy-related phenotypes in cohorts of children with mild and severe asthma. Furthermore, TNFRSF21 was significantly upregulated in bronchial basal epithelial cells from adults with allergic asthma but not in adults with allergies (but not asthma) after allergen challenge. CONCLUSIONS: We report novel associations between rare variants in genes and allergic and inflammatory phenotypes in children with diverse ancestries, highlighting TNFRSF21 as contributing to the development of allergic asthma.

Evaluation of protected benzoic acid on growth performance, nutrient digestibility, and gut health indices in starter pigs.

Benzoic acid is a common alternative for antibiotic and zinc oxide use in nursery diets. Free benzoic acid (BZA) is often supplied, but this form is absorbed before it can exert any effect on distal segments of the gut. The study aimed to evaluate the effects of protected benzoic acid on growth performance, nutrient digestibility, plasma metabolites, and gut health indices in starter pigs. A total of 192 pigs were weaned at 28 ± 1 d age (initial body weight, 8.72 ± 1.13 kg). Pens were assigned to one of four treatment diets (n = 8 pens per treatment): (1) no additive (NC), (2) free benzoic acid (BZA; 0.6%), (3) protected benzoic acid (BC50; 0.2%, supplied at a ratio of one to three equivalents of BZA), and (4) antibiotic growth promoter (AGP; Carbadox, 50 ppm). Diets were fed for three weeks over two periods (period 1, 7 d; period 2, 14 d). Body weight and feed intake were measured for each period. Feces were collected at the end of each period to determine apparent total tract digestibility (ATTD) of organic matter (OM), gross energy (GE), and crude protein (CP). One pig per pen was euthanized per period to determine plasma metabolites; jejunum and ileum morphology; jejunum, ileum, and colon cytokine abundance; and jejunum, ileum, and colon tight junction protein expression. The AGP group had increased average daily gain (ADG) and average daily feed intake (ADFI) compared to other groups in period 1 and overall (P < 0.05); however, ADG and ADFI of the BC50 group was intermediate between the NC and BZA groups and the AGP group in period 2. The ATTD of OM, GE, and CP were greater in the AGP group compared to the NC and BC50 groups (P < 0.05), whereas the BZA group was intermediate. Jejunum and ileum villus height and crypt depth increased from period 1 to period 2 (P < 0.01) but were similar across groups. Ileum and colon tumor necrosis factor-α (TNF-α) abundances were greater, whereas colon interleukin (IL)-1ß and colon and ileum IL-8 abundances were less, in the AGP group compared to the BZA group (P < 0.05); the NC and BC50 groups exhibited intermediate TNF-α, IL-1ß, and IL-8 abundance in the ileum and colon. Jejunum cytokine abundance did not vary among groups but declined from period 1 to period 2 (P < 0.05). Tight junction protein expression also did not vary among groups. In summary, protected BZA supported a slight increase in growth performance in starter pigs, suggesting its potential as an alternative feed additive in nursery diets.

Role of Chondroitin Sulfate Proteoglycan 5 in Steroid-Induced Cataract.

Steroid-induced cataracts (SIC) are defined as cataracts associated with the administration of corticosteroids. Long-term glucocorticoid treatment for inflammatory diseases reportedly increases the risk of SIC, and steroids can induce cataracts by disrupting ocular growth factor balance or homeostasis. In this study, we verified the effect of chondroitin sulfate proteoglycan 5 (CSPG5) using dexamethasone (dexa)-treated human lens epithelial (HLE-B3) cells and the lens epithelium from the anterior capsule of SIC patients obtained during cataract surgery. CSPG5 expression increased in the lens epithelium of SIC patients. The downregulation of CSPG5 suppressed the dexa-induced epithelial-mesenchymal transition (EMT)-related protein expression and motility in HLE-B3 cells. The disruption of the transcription factors EZH2 and B-Myb downregulated CSPG5, dexa-induced fibronectin expression, and cell migration in HLE-B3 cells, reaffirming that CSPG5 expression regulates EMT in lens epithelial cells. Taken together, these results indicate that the steroid-induced effects on lens epithelial cells are mediated via alterations in CSPG5 expression. Therefore, our study emphasizes the potential of CSPG5 as a therapeutic target for the prevention and treatment of SIC.

A closed, autologous bioprocess optimized for TCR-T cell therapies.

Autologous cell therapy has proven to be an effective treatment for hematological malignancies. Cell therapies for solid tumors are on the horizon, however the high cost and complexity of manufacturing these therapies remain a challenge. Routinely used open steps to transfer cells and reagents through unit operations further burden the workflow reducing efficiency and increasing the chance for human error. Here we describe a fully closed, autologous bioprocess generating engineered TCR-T cells. This bioprocess yielded 5-12 × 10e9 TCR-expressing T cells, transduced at low multiplicity of infections, within 7-10 days, and cells exhibited an enriched memory T-cell phenotype and enhanced metabolic fitness. It was demonstrated that activating, transducing, and expanding leukapheresed cells in a bioreactor without any T-cell or peripheral blood mononuclear cell enrichment steps had a high level of T-cell purity (~97%). Several critical process parameters of the bioreactor, including culturing at a high cell density (7e6 cells/mL), adjusting rocking agitations during phases of scale-up, lowering glycolysis through the addition of 2-deoxy- d-glucose, and modulating interleukin-2 levels, were investigated on their roles in regulating transduction efficiency, cell growth, and T-cell fitness such as T-cell memory phenotype and resistance to activation-induced cell death. The bioprocess described herein supports scale-out feasibility by enabling the processing of multiple patients' batches in parallel within a Grade C cleanroom.

Multi-omic association study identifies DNA methylation-mediated genotype and smoking exposure effects on lung function in children living in urban settings.

Impaired lung function in early life is associated with the subsequent development of chronic respiratory disease. Most genetic associations with lung function have been identified in adults of European descent and therefore may not represent those most relevant to pediatric populations and populations of different ancestries. In this study, we performed genome-wide association analyses of lung function in a multiethnic cohort of children (n = 1,035) living in low-income urban neighborhoods. We identified one novel locus at the TDRD9 gene in chromosome 14q32.33 associated with percent predicted forced expiratory volume in one second (FEV1) (p = 2.4x10-9; ßz = -0.31, 95% CI = -0.41- -0.21). Mendelian randomization and mediation analyses revealed that this genetic effect on FEV1 was partially mediated by DNA methylation levels at this locus in airway epithelial cells, which were also associated with environmental tobacco smoke exposure (p = 0.015). Promoter-enhancer interactions in airway epithelial cells revealed chromatin interaction loops between FEV1-associated variants in TDRD9 and the promoter region of the PPP1R13B gene, a stimulator of p53-mediated apoptosis. Expression of PPP1R13B in airway epithelial cells was significantly associated the FEV1 risk alleles (p = 1.3x10-5; ß = 0.12, 95% CI = 0.06-0.17). These combined results highlight a potential novel mechanism for reduced lung function in urban youth resulting from both genetics and smoking exposure.

Multi-residue analytical method for detecting pesticides, veterinary drugs, and mycotoxins in feed using liquid- and gas chromatography coupled with mass spectrometry.

Modified QuEChERS and triple quadrupole mass spectrometry (LC and GC-MS/MS) technology were used to sequentially analyze pesticides, veterinary drugs, and mycotoxins in feed. In order to analyze the harmful substances that may remain or occur in the feed, we performed optimization experiments for sample preparation and LC-MS/MS and GC-MS/MS conditions. Optimized sample preparation involves extracting 5 g of sample with 15 mL of 0.25 M EDTA and 10 mL of acetonitrile. And some extracts were diluted 10-fold with 100 mM ammonium formate aqueous solution and analyzed by LC-MS/MS, and some extracts were purified through 25 mg PSA and analyzed by GC-MS/MS by adding an analyte protectant. We confirmed the matrix effect of feed ingredients and compound feeds, and added a dilution process after extraction to increase on-site efficiency. Matrix-matched calibration was applied for quantification. Method validation was performed for 197 pesticides, 56 components for veterinary drugs, and 5 components for toxins. All the components showed good linearity (r2 ≥ 0.98) in the developed analytical method. For most compounds, the limit of quantitation was 0.05 mg/kg. The recovery rate experiment was repeated three times at three concentrations including LOQ in feed ingredient, compound feed for livestock, and compound feed for pets. The recovery rate was 70.09-119.76% and relative standard deviations were ≤ 18.91%. And the accuracy and precision were further verified through cross-validation between laboratories. The developed analytical method was used to monitor 414 domestically distributed and imported feeds.

Method to Minimize the Errors of AI: Quantifying and Exploiting Uncertainty of Deep Learning in Brain Tumor Segmentation.

Despite the unprecedented success of deep learning in various fields, it has been recognized that clinical diagnosis requires extra caution when applying recent deep learning techniques because false prediction can result in severe consequences. In this study, we proposed a reliable deep learning framework that could minimize incorrect segmentation by quantifying and exploiting uncertainty measures. The proposed framework demonstrated the effectiveness of a public dataset: Multimodal Brain Tumor Segmentation Challenge 2018. By using this framework, segmentation performances, particularly for small lesions, were improved. Since the segmentation of small lesions is difficult but also clinically significant, this framework could be effectively applied to the medical imaging field.

Rational design and synthesis of 2-(1H-indazol-6-yl)-1H-benzo[d]imidazole derivatives as inhibitors targeting FMS-like tyrosine kinase 3 (FLT3) and its mutants.

Fms-like tyrosine kinase 3 (FLT3) has been verified as a therapeutic target for acute myeloid leukaemia (AML). In this study, we report a series of 2-(1H-indazol-6-yl)-1H-benzo[d]imidazol-5-yl benzamide and phenyl urea derivatives as potent FLT3 inhibitors based on the structural optimisation of previous FLT3 inhibitors. Derivatives were synthesised as benzamide 8a-k, 8n-z, and phenyl urea 8l-m, with various substituents. The most potent inhibitor, 8r, demonstrated strong inhibitory activity against FLT3 and FLT3 mutants with a nanomolar IC50 and high selectivity profiles over 42 protein kinases. In addition, these type II FLT3 inhibitors were more potent against FLT3 mutants correlated with drug resistance. Overall, we provide a theoretical basis for the structural optimisation of novel benzimidazole analogues to develop strong inhibitors against FLT3 mutants for AML therapeutics.

A Decision Tree Model for Breast Reconstruction of Women with Breast Cancer: A Mixed Method Approach.

The number of breast reconstructions following mastectomy has increased significantly during the last decades, but women are experiencing a number of conflicts with breast reconstruction decisions. The aim of this study was to develop a decision tree model of breast reconstruction and to examine its predictability. Mixed method design using ethnographic decision tree modeling was used. In the qualitative stage, data were collected using individual and focus group interviews and analyzed to construct a decision tree model. In the quantitative stage, the questionnaire was developed questions based on the criteria identified in the qualitative stage. A total of 61 women with breast cancer participated in 2017. Five major criteria: recovery of body image; impact on recurrence; recommendations from others; financial resources; and confirmation by physicians. The model also included nine predictive pathways. It turns out that the model predicted 90% of decisions concerning whether or not to have breast reconstruction. The findings indicate that the five criteria play a key role in decision-making about whether or not to have breast reconstruction. Thus, more comprehensive issues, including these five criteria, need to be integrated into an intervention for women with breast cancer to make their best decision on breast reconstruction.

Breast Milk Transforming Growth Factor ß Is Associated With Neonatal Gut Microbial Composition.

BACKGROUND AND OBJECTIVES: Breast milk is a complex bioactive fluid that varies across numerous maternal and environmental conditions. Although breast-feeding is known to affect neonatal gut microbiome, the milk components responsible for this effect are not well-characterized. Given the wide range of immunological activity breast milk cytokines engage in, we investigated 3 essential breast milk cytokines and their association with early life gut microbiota. METHODS: A total of 52 maternal-child pairs were drawn from a racially diverse birth cohort based in Detroit, Michigan. Breast milk and neonatal stool specimens were collected at 1-month postpartum. Breast milk transforming growth factor (TGF)ß1, TGFß2, and IL-10 were assayed using enzyme-linked immunosorbent assays, whereas neonatal gut microbiome was profiled using 16S rRNA sequencing. RESULTS: Individually, immunomodulators TGFß1 and TGFß2 were significantly associated with neonatal gut microbial composition (R = 0.024, P = 0.041; R = 0.026, P = 0.012, respectively) and increased richness, evenness, and diversity, but IL-10 was not. The effects of TGFß1 and TGFß2, however, were not independent of one another, and the effect of TGFß2 was stronger than that of TGFß1. Higher levels of TGFß2 were associated with the increased relative abundance of several bacteria, including members of Streptococcaceae and Ruminococcaceae, and lower relative abundance of distinct Staphylococcaceae taxa. CONCLUSIONS: Breast milk TGFß concentration explains a portion of variability in gut bacterial microbiota composition among breast-fed neonates. Whether TGFß acts in isolation or jointly with other bioactive components to alter bacterial composition requires further investigation. These findings contribute to an increased understanding of how breast-feeding affects the gut microbiome-and potentially immune development-in early life.

Joint effects of pregnancy, sociocultural, and environmental factors on early life gut microbiome structure and diversity.

The joint impact of pregnancy, environmental, and sociocultural exposures on early life gut microbiome is not yet well-characterized, especially in racially and socioeconomically diverse populations. Gut microbiota of 298 children from a Detroit-based birth cohort were profiled using 16S rRNA sequencing: 130 neonates (median age = 1.2 months) and 168 infants (median age = 6.6 months). Multiple factors were associated with neonatal gut microbiome composition in both single- and multi-factor models, with independent contributions of maternal race-ethnicity, breastfeeding, mode of delivery, marital status, exposure to environmental tobacco smoke, and indoor pets. These findings were consistent in the infants, and networks demonstrating the shared impact of factors on gut microbial composition also showed notable topological similarity between neonates and infants. Further, latent groups defined by these factors explained additional variation, highlighting the importance of combinatorial effects. Our findings also have implications for studies investigating the impact of the early life gut microbiota on disease.

[Decision Making Experience on Breast Reconstruction for Women with Breast Cancer].

PURPOSE: The purpose of this study was to explore decision making experiences of Korean women with breast cancer who underwent breast reconstruction with/after a mastectomy. METHODS: Data were collected during 2015-2016 through individual in-depth interviews with 10 women who had both mastectomy and breast reconstruction, and analyzed using phenomenological method to identify essential themes on experiences of making a decision to have breast reconstruction. RESULTS: Five theme clusters emerged. First, "expected loss of sexuality and discovery of autonomy" illustrates various aims of breast reconstruction. Second, "holding tight to the reputation of doctors amid uncertainty" specifies the importance of a trust relationship with their physician despite a lack of information. Third, "family members to step back in position" describes support or opposition from family members in the decision making process. Fourth, "bewilderment due to the paradox of appearance-oriented views" illustrates paradoxical environment, resulting in confusion and anger. Lastly, "decision to be made quickly with limited time to oneself" describes the crazy whirling process of decision making. CONCLUSION: Findings highlight aims, worries, barriers, and facilitators that women with breast cancer experience when making a decision about breast reconstruction. Deciding on breast reconstruction was not only a burden for women in a state of shock with a diagnosis of breast cancer, but also an opportunity to decide to integrate their body, femininity, and self which might be wounded from a mastectomy. These findings will help oncology professionals provide effective educational counselling before the operation to promote higher satisfaction after the operation.

Mobile-Based Video Learning Outcomes in Clinical Nursing Skill Education: A Randomized Controlled Trial.

Mobile devices are a regular part of daily life among the younger generations. Thus, now is the time to apply mobile device use to nursing education. The purpose of this study was to identify the effects of a mobile-based video clip on learning motivation, competence, and class satisfaction in nursing students using a randomized controlled trial with a pretest and posttest design. A total of 71 nursing students participated in this study: 36 in the intervention group and 35 in the control group. A video clip of how to perform a urinary catheterization was developed, and the intervention group was able to download it to their own mobile devices for unlimited viewing throughout 1 week. All of the students participated in a practice laboratory to learn urinary catheterization and were blindly tested for their performance skills after participation in the laboratory. The intervention group showed significantly higher levels of learning motivation and class satisfaction than did the control. Of the fundamental nursing competencies, the intervention group was more confident in practicing catheterization than their counterparts. Our findings suggest that video clips using mobile devices are useful tools that educate student nurses on relevant clinical skills and improve learning outcomes.

Effects of Mesenchymal Stem Cell Treatment on the Expression of Matrix Metalloproteinases and Angiogenesis during Ischemic Stroke Recovery.

BACKGROUND: The efficacy of mesenchymal stem cell (MSC) transplantation in ischemic stroke might depend on the timing of administration. We investigated the optimal time point of MSC transplantation. After MSC treatment, we also investigated the expression of matrix metalloproteinases (MMPs), which play a role in vascular and tissue remodeling. METHODS: Human bone marrow-derived MSCs (2 × 10(6), passage 5) were administrated intravenously after permanent middle cerebral artery occlusion (MCAO) was induced in male Sprague-Dawley rats. First, we determined the time point of MSC transplantation that led to maximal neurological recovery at 1 h, 1 day, and 3 days after MCAO. Next, we measured activity of MMP-2 and MMP-9, neurological recovery, infarction volume, and vascular density after transplanting MSCs at the time that led to maximal neurological recovery. RESULTS: Among the MSC-transplanted rats, those of the MSC 1-hour group showed maximal recovery in the rotarod test (P = 0.023) and the Longa score (P = 0.018). MMP-2 activity at 1 day after MCAO in the MSC 1-hour group was significantly higher than that in the control group (P = 0.002), but MMP-9 activity was not distinct. The MSC 1-hour group also showed smaller infarction volume and higher vascular density than did the control group. CONCLUSIONS: In a permanent model of rodent MCAO, very early transplantation of human MSCs (1 h after MCAO) produced greater neurological recovery and decreased infraction volume. The elevation of MMP-2 activity and the increase in vascular density after MSC treatment suggest that MSCs might help promote angiogenesis and lead to neurological improvement during the recovery phase after ischemic stroke.

Water Lubrication of Stainless Steel using Reduced Graphene Oxide Coating.

Lubrication of mechanical systems using water instead of conventional oil lubricants is extremely attractive from the view of resource conservation and environmental protection. However, insufficient film thickness of water due to low viscosity and chemical reaction of water with metallic materials have been a great obstacle in utilization of water as an effective lubricant. Herein, the friction between a 440 C stainless steel (SS) ball and a 440 C stainless steel (SS) plate in water lubrication could be reduced by as much as 6-times by coating the ball with reduced graphene oxide (rGO). The friction coefficient with rGO coated ball in water lubrication was comparable to the value obtained with the uncoated ball in oil lubrication. Moreover, the wear rate of the SS plate slid against the rGO coated ball in water lubrication was 3-times lower than that of the SS plate slid against the uncoated ball in oil lubrication. These results clearly demonstrated that water can be effectively utilized as a lubricant instead of oil to lower the friction and wear of SS components by coating one side with rGO. Implementation of this technology in mechanical systems is expected to aid in significant reduction of environmental pollution caused by the extensive use of oil lubricants.

5,3'-Dihydroxy-6,7,4'-trimethoxyflavanone exerts its anticancer and antiangiogenesis effects through regulation of the Akt/mTOR signaling pathway in human lung cancer cells.

5,3'-Dihydroxy-6,7,4'-trimethoxyflavanone (DHTMF) is one of the constituents of Vitex rotundifolia, a medicinal herb that is used for the treatment of various disorders in China and Korea. In this study we evaluated the antitumor and antiangiogeneic activities of DHTMF. DHTMF significantly suppressed growth and induced apoptosis in lung carcinoma cells in a dose-dependent manner, as indicated by a decrease in Bcl-2 levels and increases in Bax and cleaved caspase-3 levels. In addition, DHTMF treatment significantly reduced the phosphorylation of Akt and mammalian target of rapamycin (mTOR), accompanied by reductions in the protein level of hypoxia-inducible factor (HIF-1α) and vascular endothelial growth factor (VEGF), which are key angiogenic molecules in H522 lung cancer cells. Furthermore DHTMF inhibited VEGF-induced angiogenesis, as indicated by reduced expression of CD34, tube formation and migration in human umbilical vein endothelial cells (HUVECs), as well as reduced neovascularization in an in vivo mouse Matrigel plug assay. DHTMF also inhibited phosphorylation of Akt, mTOR, and p70S6K in HUVECs and lung cancer cells. Taken together, our finding indicated that DHTMF inhibits Akt/mTOR signaling and reduces the expression of HIF-1 α and VEGF in tumor cells, which in turns inhibits endothelial cell-mediated angiogenesis. These results suggest that DHTMF inhibits angiogenesis as well as induces apoptosis via the Akt/mTOR pathway and might elicit pharmacological effects that are useful for treatment of lung cancer.

Cytochalasin H, an active anti-angiogenic constituent of the ethanol extract of Gleditsia sinensis thorns.

Angiogenesis, the process of new vessel formation from the pre-existing blood vasculature, is critical for continuous tumor growth and is considered to be a validated antitumor target. The results of our previous study demonstrate the anti-angiogenic potential of an extract of Gleditsia sinensis thorns, which has been traditionally used in Korean medicine to remedy diverse diseases, including tumors. In the present study, we attempted to identify the active anti-angiogenic constituents of the ethanol extract of G. sinensis thorns (EEGS). By virtue of in vitro activity-guided fractionation using human umbilical vein endothelial cells (HUVEC) primary endothelial cells, chromatographic separation, and NMR spectral analyses, we isolated and identified the potent active constituent, cytochalasin H, a biologically active secondary metabolite of fungi. This unexpected active constituent may have originated from the endophytic fungi, Chaetomium globosum, which naturally populate G. sinensis, the identity of which was determined by analysis of fungal community. Cytochalasin H isolated from the EEGS showed in vitro anti-angiogenic activities such as suppressed cell growth and mobility in HUVEC, and inhibited the pro-angiogenic protein-induced formation of new blood vessels in vivo. The anti-angiogenic effect of cytochalasin H was in part due to reduced expression of pro-angiogenic factor, such as endothelin-1. This is the first report regarding the isolation and identification of cytochalasin H, as an active anti-angiogenic constituent of G. sinensis thorns.

Percutaneous vertebroplasty at C7 for the treatment of painful metastases -A case report-.

The cervical spine is a less common site for metastatic disease than is the thoracolumbar spine. Percutaneous vertebroplasty (PVP) in the cervical spine can be performed using an anteromedial or lateral approach. A 51-year-old woman with breast cancer had been experiencing severe weight-bearing neck pain for 2 months, even after undergoing radiation therapy. Imaging studies revealed an osteolytic compression fracture in the C7 vertebra. For performing PVP at C7 using the anteromedial approach, a needle was inserted from the left side of the patient's neck. The needle was advanced to the anterior 1/3 anterior 1/3 or 1/2 of the vertebral body by hammering, and approximately 2 ml of cement was injected. Immediately after the operation, the patient could move her neck without pain. In conclusion, PVP using an anterolateral approach may be an option for treating metastatic osteolytic vertebral lesions in the cervical spine for alleviating intractable axial neck pain.