1,410 research outputs found
The Genetics of CFTR: Genotype - Phenotype Relationship, Diagnostic Challenge and Therapeutic Implications
A Genotypic-oriented View of CFTR Genetics Highlights Specific Mutational Patterns Underlying Clinical Macro-categories of Cystic Fibrosis.
Cystic Fibrosis (CF) is a monogenic disease caused by mutations of the Cystic Fibrosis Transmembrane conductance Regulator (CFTR) gene. The genotype-phenotype relationship in this disease is still unclear, and diagnostic, prognostic and therapeutic challenges persist. We enrolled 610 patients with different forms of CF and studied them from a clinical, biochemical, microbiological and genetic point of view. Overall, 125 different mutated alleles (11 of which with novel mutations and 10 of which complex) and 225 genotypes were found. A strong correlation between mutational patterns at the genotypic level and phenotypic macro-categories emerged. This specificity appears to be largely dependent on rare and individual mutations, as well as on the varying prevalence of common alleles in different clinical macro-categories. However, 19 genotypes appeared to underlie different clinical forms of the disease. The dissection of the pathway from the CFTR mutated genotype to the clinical phenotype allowed to identify at least two components of the variability usually found in the genotype - phenotype relationship. One component seems to depend on the genetic variation of CFTR, the other component on the cumulative effect of variations in other genes and cellular pathways independent from CFTR. The experimental dissection of the overall biological CFTR pathway appears to be a powerful approach for a better comprehension of the genotype - phenotype relationship. However, a change from an allele-oriented to a genotypic-oriented view of CFTR genetics is mandatory, as well as a better assessment of sources of variability within the CFTR pathway
On Cas A, Cassini, comets, and King Charles
We re-examine the long-standing problem of the date of the Cassiopeia A supernova (SN), in view of recent claims that it might be the 1630 ânoon-starâ seen at the birth of King Charles II. We do not support this identification, based on the expected brightness of a Type-IIb SN (too faint to be seen in daylight), the extrapolated motion of the ejecta (inconsistent with a date earlier than 1650), the lack of any scientific follow-up observations, the lack of any mention of it in Asian archives. The origin of the 1630 noon-star event (if real) remains a mystery; there was a bright comet in 1630 June but no evidence to determine whether or not it was visible in daylight. Instead, we present French reports about a fourth-magnitude star discovered by Cassini in Cassiopeia in or shortly before 1671, which was not seen before or since. The brightness is consistent with what we expect for the Cas A SN; the date is consistent with the extrapolated motion of the ejecta. We argue that this source could be the long-sought SN
Correction of a urea cycle defect after ex vivo gene editing of human hepatocytes
Ornithine transcarbamylase deficiency (OTCD) is a monogenic disease of ammonia metabolism in hepatocytes. Severe disease is frequently treated by orthotopic liver transplantation. An attractive approach is the correction of a patient's own cells to regenerate the liver with gene-repaired hepatocytes. This study investigates the efficacy and safety of ex vivo correction of primary human hepatocytes. Hepatocytes isolated from an OTCD patient were genetically corrected ex vivo, through the deletion of a mutant intronic splicing site achieving editing efficiencies >60% and the restoration of the urea cycle in vitro. The corrected hepatocytes were transplanted into the liver of FRGN mice and repopulated to high levels (>80%). Animals transplanted and liver repopulated with genetically edited patient hepatocytes displayed normal ammonia, enhanced clearance of an ammonia challenge and OTC enzyme activity, as well as lower urinary orotic acid when compared to mice repopulated with unedited patient hepatocytes. Gene expression was shown to be similar between mice transplanted with unedited or edited patient hepatocytes. Finally, a genome-wide screening by performing CIRCLE-seq and deep sequencing of >70 potential off-targets revealed no unspecific editing. Overall analysis of disease phenotype, gene expression, and possible off-target editing indicated that the gene editing of a severe genetic liver disease was safe and effective.
Keywords: CRISPR; FRGN; ex vivo; genome editing; hepatocyte transplantation; liver-humanized mouse; primary hepatocytes; urea cycle disorder
Resistance to mesenchymal reprogramming sustains clonal propagation in metastatic breast cancer
The acquisition of mesenchymal traits is considered a hallmark of breast cancer progression. However, the functional relevance of epithelial-to-mesenchymal transition (EMT) remains controversial and context dependent. Here, we isolate epithelial and mesenchymal populations from human breast cancer metastatic biopsies and assess their functional potential in vivo. Strikingly, progressively decreasing epithelial cell adhesion molecule (EPCAM) levels correlate with declining disease propagation. Mechanistically, we find that persistent EPCAM expression marks epithelial clones that resist EMT induction and propagate competitively. In contrast, loss of EPCAM defines clones arrested in a mesenchymal state, with concomitant suppression of tumorigenicity and metastatic potential. This dichotomy results from distinct clonal trajectories impacting global epigenetic programs that are determined by the interplay between human ZEB1 and its target GRHL2. Collectively, our results indicate that susceptibility to irreversible EMT restrains clonal propagation, whereas resistance to mesenchymal reprogramming sustains disease spread in multiple models of human metastatic breast cancer, including patient-derived cells in vivo
Role of Chromatin Structural Changes in Regulating Human CYP3A Ontogeny
Non-standard abbreviations: bp(s), base pair(s); C/EBP, CCAAT/enhancer binding protein; ChIP, chromatin immunoprecipitation; CLEM4, constitutive liver enhancer module 4; Cq, quantification cycle; DME, drug metabolizing enzyme; HNF4α, hepatocyte nuclear factor 4 alpha; PXR, pregnane X receptor; qPCR, quantitative polymerase chain reaction; TBP, TATAbox binding protein; TFIID, transcription factor II D; TSS, transcription start site; USF1, upstream stimulatory factor 1; XREM, xenobiotic-response enhancer module DMD #69344 3 Abstract Variability in drug metabolizing enzyme developmental trajectories contributes to interindividual differences in susceptibility to chemical toxicity and adverse drug reactions, particularly in the first years of life. Factors linked to these interindividual differences are largely unknown, but molecular mechanisms regulating ontogeny are likely involved. To evaluate chromatin structure dynamics as a likely contributing mechanism, age-dependent changes in modified and variant histone occupancy were evaluated within known CYP3A4 and 3A7 regulatory domains. Chromatin immunoprecipitation using fetal or postnatal human hepatocyte chromatin pools followed by quantitative polymerase chain reaction DNA amplification was used to determine relative chromatin occupancy by modified and variant histones. Chromatin structure representing a poised transcriptional state (bivalent chromatin), indicated by the occupancy by modified histones associated with both active and repressed transcription, was observed for CYP3A4 and most 3A7 regulatory regions in both postnatal and fetal livers. However, the CYP3A4 regulatory regions had significantly greater occupancy by modified histones associated with repressed transcription in the fetal liver. Conversely, some modified histones associated with active transcription exhibited greater occupancy in the postnatal liver. CYP3A7 regulatory regions also had significantly greater occupancy by modified histones associated with repressed transcription in the fetus. The occupancy by modified histones observed is consistent with chromatin structural dynamics contributing to CYP3A4 ontogeny, although the data is less conclusive regarding CYP3A7. Interpretation of the latter data may be confounded by cell-type heterogeneity in the fetal liver. DMD #69344
Measurement of Branching Fractions and Rate Asymmetries in the Rare Decays B -> K(*) l+ l-
In a sample of 471 million BB events collected with the BABAR detector at the
PEP-II e+e- collider we study the rare decays B -> K(*) l+ l-, where l+ l- is
either e+e- or mu+mu-. We report results on partial branching fractions and
isospin asymmetries in seven bins of di-lepton mass-squared. We further present
CP and lepton-flavor asymmetries for di-lepton masses below and above the J/psi
resonance. We find no evidence for CP or lepton-flavor violation. The partial
branching fractions and isospin asymmetries are consistent with the Standard
Model predictions and with results from other experiments.Comment: 16 pages, 14 figures, accepted by Phys. Rev.
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