Integrate CRISPR/Cas9 for protein expression of HLA-B*38:68Q via precise gene editing.

The determination of null- or low-expressed HLA alleles is clinically relevant in both hematopoietic stem cell transplantation and solid organ transplantation. We studied the expression level of a questionable (Q) HLA-B*38:68Q allele, which carries a 9-nucleotide (nt) deletion at codon 230-232 in exon 4 of HLA-B*38:01:01:01 using CRISPR/Cas9 gene editing technology. CRISPR/Cas9 gene editing of HLA-B*38:01:01:01 homozygous EBV B cell line resulted in one HLA-B*38:68Q/B*38:01:01:01 heterozygous and one HLA-B*38:68Q homozygous clone. Flow cytometric analysis of monoclonal anti-Bw4 antibody showed the protein expression of HLA-B*38:01:01:01 in homozygous cells was 2.2 fold higher than HLA-B*38:68Q/B*38:01:01:01 heterozygous cells, and the expression of HLA-B*38:68Q/B*38:01:01:01 heterozygous cells was over 2.0 fold higher than HLA-B*38:68Q homozygous cells. The HLA-B*38:68Q expression was further confirmed using anti-B38 polyclonal antibody. Similarly, the expression of the HLA-B*38:01:01:01 homozygous cells was 1.5 fold higher than that of HLA-B*38:68Q/B*38:01:01:01 heterozygous cells, and the HLA-B*38:68Q/B*38:01:01:01 heterozygous cells was over 1.6 fold higher than that of HLA-B*38:68Q homozygous cells. The treatment of HLA-B*38:68Q homozygous cells with IFN-γ significantly increased its expression. In conclusion, we demonstrate that HLA-B*38:68Q is a low-expressing HLA allele. The CRISPR/Cas9 technology is a useful tool to induce precise gene editing in HLA genes to enable the characterization of HLA gene variants on expression and function

Not All Antibodies Are Created Equal: Factors That Influence Antibody Mediated Rejection.

Consistent with Dr. Paul Terasaki's "humoral theory of rejection" numerous studies have shown that HLA antibodies can cause acute and chronic antibody mediated rejection (AMR) and decreased graft survival. New evidence also supports a role for antibodies to non-HLA antigens in AMR and allograft injury. Despite the remarkable efforts by leaders in the field who pioneered single antigen bead technology for detection of donor specific antibodies, a considerable amount of work is still needed to better define the antibody attributes that are associated with AMR pathology. This review highlights what is currently known about the clinical context of pre and posttransplant antibodies, antibody characteristics that influence AMR, and the paths after donor specific antibody production (no rejection, subclinical rejection, and clinical dysfunction with AMR)

Spatial Ecology of and Public Attitudes toward Monk Parakeets Nesting on Electric Utility Structures in Dallas and Tarrant Counties, Texas

Monk parakeets (Myiopsitta monachus) were introduced to the United States (US) where they established naturalized populations. They often build their bulky twig nests on electric utility structures, causing economic damage. From May 2010–February 2013, we examined the spatial ecology of and public attitudes toward monk parakeets nesting on electric utility structures in Dallas and Tarrant counties, Texas, US. As nest sites, monk parakeets selected electric switchyards and substations constructed with multiple flat surfaces and acute-angles, within small fenced areas, with large canopy trees and taller anthropogenic structures within 100 m. Multi-scale analysis of urban land use-land cover (LULC) suggested the surrounding landscape had little impact on nest-site selection. Monk parakeets used canopy LULC more often than pavement, grass, buildings, or water. They traveled farthest from their nests during winter. Flock sizes were highly variable, yet largest during nonbreeding season. They foraged on a broad range of native vegetation and exhibited a diverse diet of flowers, fruits, acorns, grass blades, wild dry seeds, leaf buds, insect larvae, and commercial bird seed. We evaluated sociological variables as predictors of opposition to managing monk parakeets nesting on electric utility structures. Most survey participants were affluent, well-educated, older Caucasians who were unknowledgeable about, inexperienced with, and unsure about the impacts of monk parakeets. They indicated least opposition to nest removal and structural modification. When opposing, they would most likely do so socially and through petitions. Participants were influenced by their desire for monk parakeets to feed at their bird feeders or nest in their yard, people and groups important to them, and their perceived ease of opposing. Our results suggest LULC manipulation and food-based strategies are not reasonable for controlling urban monk parakeets. We recommend an outreach program explaining monk parakeet biology and the impacts of their nesting habits. We suggest targeting affluent areas adjacent to electric structures with nests and the predictors driving participants’ behaviors. To provoke least opposition, we advise nest removal and structural modification. We recommend electric companies conduct cost-benefit analyses exploring feasibility of modifying construction elements preferred by monk parakeets and redesigning new construction to reduce risk of future nesting on electric utility structures

ESTUDO SOBRE CORANTES ARTIFICIAIS EM ALIMENTOS: QUAIS OS RISCOS MAIS COMUNS PELO CONSUMO EXCESSIVO

Pretende-se realizar um estudo no qual serão verificados alimentos que contêm corantes artificiais e os danos que estes podem causar à saúde. Estudos sobre os efeitos à saúde causados pelos corantes artificiais além de insuficientes são contraditórios, havendo diferentes opiniões quanto à sua inocuidade. Muitos estudos mostram que esses aditivos podem causar uma série de males à saúde quando consumidos de forma incorreta, seja por excessos por parte da indústria ou exagero no consumo. Todavia, desde que se obedeçam aos percentuais máximos estabelecidos pela ANVISA (Decreto nº 50040, de 24 de janeiro de 1961), o consumo destes aditivos é considerado inofensivo à saúde (SHIMADZU, 2007). Espera-se que o estudo possa esclarecer e ajudar as pessoas a terem uma alimentação mais saudável, visando prevenir doenças e contribuir para melhor qualidade de vida

Subtle deregulation of the Wnt-signalling pathway through loss of Apc2 reduces the fitness of intestinal stem cells

The importance of the Wnt-signaling pathway on the regulation and maintenance of the intesti- nal stem cell (ISC) population is well recognized. However, our current knowledge base is founded on models using systems of gross deregulation of the Wnt-signaling pathway. Given the importance of this signaling pathway on intestinal homeostasis, there is a need to explore the role of more subtle alterations in Wnt-signaling levels within this tissue. Herein, we have used a model of Apc2 loss to meet this aim. Apc2 is a homolog of Apc which can also form a destruction complex capable of binding b-catenin, albeit less efficiently than Apc. We show that systemic loss of Apc2 results in an increase in the number of cells displaying nuclear b-catenin at the base of the intestinal crypt. This subsequently impacts the expression levels of several ISC markers and the fitness of ISCs as assessed by organoid formation efficiency. This work pro- vides the first evidence that the function and fitness of ISCs can be altered by even minor mis- regulation of the Wnt-signaling pathway. Our data highlights the importance of correct maintenance of this crucial signaling pathway in the maintenance and function of the ISC popu- lation

The response to influenza vaccination is associated with DNA methylation-driven regulation of T cell innate antiviral pathways.

BACKGROUND: The effect of vaccination on the epigenome remains poorly characterized. In previous research, we identified an association between seroprotection against influenza and DNA methylation at sites associated with the RIG-1 signaling pathway, which recognizes viral double-stranded RNA and leads to a type I interferon response. However, these studies did not fully account for confounding factors including age, gender, and BMI, along with changes in cell-type composition. RESULTS: Here, we studied the influenza vaccine response in a longitudinal cohort vaccinated over two consecutive years (2019-2020 and 2020-2021), using peripheral blood mononuclear cells and a targeted DNA methylation approach. To address the effects of multiple factors on the epigenome, we designed a multivariate multiple regression model that included seroprotection levels as quantified by the hemagglutination-inhibition (HAI) assay test. CONCLUSIONS: Our findings indicate that 179 methylation sites can be combined as potential signatures to predict seroprotection. These sites were not only enriched for genes involved in the regulation of the RIG-I signaling pathway, as found previously, but also enriched for other genes associated with innate immunity to viruses and the transcription factor binding sites of BRD4, which is known to impact T cell memory. We propose a model to suggest that the RIG-I pathway and BRD4 could potentially be modulated to improve immunization strategies