Halvorsenbruhn9369

© 2020 The Author(s).We previously reported dysregulated expression of liver-derived messenger RNA (mRNA) and long noncoding RNA (lncRNA) in patients with advanced fibrosis resulting from nonalcoholic fatty liver disease (NAFLD). Here we sought to identify changes in mRNA and lncRNA levels associated with activation of hepatic stellate cells (HSCs), the predominant source of extracellular matrix production in the liver and key to NAFLD-related fibrogenesis. We performed expression profiling of mRNA and lncRNA from LX-2 cells, an immortalized human HSC cell line, treated to induce phenotypes resembling quiescent and myofibroblastic states. We identified 1964 mRNAs (1377 upregulated and 587 downregulated) and 1460 lncRNAs (665 upregulated and 795 downregulated) showing statistically significant evidence (FDR ≤0.05) for differential expression (fold change ≥|2|) between quiescent and activated states. Pathway analysis of differentially expressed genes showed enrichment for hepatic fibrosis (FDR = 1.35E-16), osteoarthritis (FDR = 1.47E-14), and axonal guidance signaling (FDR = 1.09E-09). selleck products We observed 127 lncRNAs/nearby mRNA pairs showing differential expression, the majority of which were dysregulated in the same direction. A comparison of differentially expressed transcripts in LX-2 cells with RNA-sequencing results from NAFLD patients with or without liver fibrosis revealed 1047 mRNAs and 91 lncRNAs shared between the two datasets, suggesting that some of the expression changes occurring during HSC activation can be observed in biopsied human tissue. These results identify lncRNA and mRNA expression patterns associated with activated human HSCs that appear to recapitulate human NAFLD fibrosis. © 2020 The Author(s).The Streptococcus pyogenes CRISPR/Cas9 (SpCas9) system is now widely utilized to generate genome engineered mice; however, some studies raised issues related to off-target mutations with this system. Herein, we utilized the Campylobacter jejuni Cas9 (CjCas9) system to generate knockout mice. We designed sgRNAs targeting mouse Tyr or Foxn1 and microinjected into zygotes along with CjCas9 mRNA. We obtained newborn mice from the microinjected embryos and confirmed that 50% (Tyr) and 38.5% (Foxn1) of the newborn mice have biallelic mutation on the intended target sequences, indicating efficient genome targeting by CjCas9. In addition, we analyzed off-target mutations in founder mutant mice by targeted deep sequencing and whole genome sequencing. Both analyses revealed no off-target mutations at potential off-target sites predicted in silico and no unexpected random mutations in analyzed founder animals. In conclusion, the CjCas9 system can be utilized to generate genome edited mice in a precise manner. © 2020 The Authors.Visualizing mitochondria in living Dictyostelium discoideum cells using fluorescent dyes is often problematic due to variability in staining, metabolism of the dyes, and unknown potential effects of the dyes on mitochondrial function. We show that fluorescent labelling of mitochondria, using an N-terminal mitochondrial localization sequence derived from the D. discoideum protein GcvH1 (glycine cleavage system H1) attached to a red fluorescent protein enables clear mitochondrial imaging. We also show that this labelling has no effect upon mitochondria load or respiratory function. © 2020 The Author(s).Background and aims Handling of PTB and EPTB patients with adequate standard detection of MTBC and anti-TB drug sensitivity using accurate and rapid methods could provide good TB management and clinical treatment outcomes. The Xpert MTB/RIF assay is an automated, cartridge-based NAAT that can simultaneously detect MTBC and RIF resistance within 2 h. The aim of this study was to evaluate the implementation of Xpert for determining diagnosis of PTB and EPTB in adults and children. Methods A descriptive study was performed using e-TB Manager data from the MDR-TB Clinic at Dr. Soetomo Academic Hospital. Suspected TB cases were from the areas of East Java Province from January 2016 to December 2018. Xpert assay was conducted using standardized criteria for clinically suspected TB, and MTBC-positive results with RR were examined by the culture method using MGIT 960 BACTEC System. Results A total of 1181 (1181/3009, 39.25%) sputum samples from suspected new MDR-PTB cases tested positive for MTBC with 3.02% RR. Among 3893 sputum samples from previously treated probable MDR-PTB cases tested using Xpert, 1936 (49.73%) were MTBC positive with 13.20% RR. Among 59 new suspected MDR-PTB cases tested using MGIT 960 BACTEC System, 55 tested positive for MTBC, although all RR strains were highly sensitive to amikacin (100%), kanamycin (95%), and ofloxacin (89%). A total of 49 children with suspected PTB were tested using Xpert, revealing low positivity (12%) for MTBC, with all RR strains being rifampicin sensitive (RS). Of the 86 suspected EPTB cases tested using Xpert, very few were MTBC-positive (26%), with 91% RS. Conclusions This study revealed that in adults and children with PTB and EPTB, the Xpert assay achieved a low positivity detection rate for MTBC in samples from new or previously treated cases, and this could be the result of many factors. © 2020 The Authors. Published by Elsevier Ltd.Introduction One-third of tuberculosis (TB) cases in Ethiopia are missing from care for reasons that are not well studied. The aim of this study was to assess TB burden and identify challenges related to TB screening and diagnosis in Ethiopia. Methods A facility-based cross-sectional study was conducted in seven health facilities selected from two regions and 2 city administrations of Ethiopia using stratified random sampling procedures. The data of 1,059,065 patients were included from outpatient department, HIV clinic, diabetic, and maternal-child health clinics. Data were collected from October to December 2018 using a retrospective review of three years' facility data (2015 to 2017) supplemented by a semi-structured interview with purposively selected health care workers and heads of the health facilities. Results A total of 1,059,065 patients visited the health facilities in three years, of these, 978,480 (92.4%) were outpatients. Of the total, 20,284 (2%) were presumptive TB cases (with 14 days or more cough), 12.