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[
Genetics,
1997]
To identify genomic regions required for establishment and patterning of the epidermis, we screened 58 deficiencies that collectively delete at least approximately 67% of the Caenorhabditis elegans genome. The epidermal pattern of deficiency homozygous embryos was analyzed by examining expression of a marker specific for one of the three major epidermal cell types, the seam cells. The organization of the epidermis and internal organs was also analyzed using a monoclonal antibody specific for epithelial adherens junctions. While seven deficiencies had no apparent effect on seam cell production, 21 were found to result in subnormal, and five in excess numbers of these cells. An additional 23 deficiencies blocked expression of the seam cell marker, in some cases without preventing cell proliferation. Two deficiencies result in multinucleate seam cells. Deficiencies were also identified that result in subnormal numbers of epidermal cells, hyperfusion of epidermal cells into a large syncytium, or aberrant epidermal differentiation. Finally, analysis of internal epithelia revealed deficiencies that cause defects in formation of internal organs, including circularization of the intestine and bifurcation of the pharynx lumen. This study reveals that many regions of the C. elegans genome are required zygotically for patterning of the epidermis and other epithelia.
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[
International C. elegans Meeting,
1995]
We are investigating the mechanisms by which the epidermis is specified and patterned during embryogenesis. The epidermis consists of 3 major subtypes of cells approximately arranged in bilaterally symmetric rows. To identify loci that are required zygotically for normal epidermal development, we have screened ~60 deficiencies representative of most of the genome. Homozygous deficiency embryos were analyzed for expression of SCM, a seam cell-specific b-galactosidase construct (1994 East Coast C. elegans Meeting, p. 111) and the MH27 antigen which is localized in epithelial adherens junctions. The quantity and pattern of seam cells and the structure of the epidermis were evaluated for each deficiency at various developmental stages. Four classes of seam cell phenotypes, 4 classes of other epidermal phenotypes, and the minimum number of relevant loci for each are listed below.
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[
International C. elegans Meeting,
1993]
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[
Molecules,
2015]
Roemerine (RM) is an aporphine alkaloid isolated from the fresh rattan stem of Fibraurea recisa, and it has been demonstrated to have certain antifungal activity. This study aimed to investigate the antifungal activity of RM and the underlying mechanisms in Candida albicans (C. albicans). The in vitro antifungal activity of RM was evaluated by a series of experiments, including the XTT reduction assay, confocal laser scanning microscopy assay, scanning electron microscope assay. Results showed that 1 g/mL RM inhibited biofilm formation significantly (p < 0.01) both in Spider medium and Lee's medium. In addition, RM could inhibit yeast-to-hyphae transition of C. albicans in a dose-dependent manner. The biofilm-specific and hypha-specific genes such as YWP1, SAP5, SAP6, HWP1, ECE1 were up-regulated and EFG1 was down-regulated after 8 g/mL RM treatment. Furthermore, the toxicity of RM was investigated using C. elegans worms, three cancer cells and one normal cell. The date showed that RM had no significant toxicity. In conclusion, RM could inhibited the formation of C. albicans biofilm in vitro, but it had no fungicidal effect on planktonic C. albicans cells, and the anti-biofilm mechanism may be related to the cAMP pathway.
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[
Anal Chem,
2021]
The use of quality control samples in metabolomics ensures data quality, reproducibility, and comparability between studies, analytical platforms, and laboratories. Long-term, stable, and sustainable reference materials (RMs) are a critical component of the quality assurance/quality control (QA/QC) system; however, the limited selection of currently available matrix-matched RMs reduces their applicability for widespread use. To produce an RM in any context, for any matrix that is robust to changes over the course of time, we developed iterative batch averaging method (IBAT). To illustrate this method, we generated 11 independently grown <i>Escherichia coli</i> batches and made an RM over the course of 10 IBAT iterations. We measured the variance of these materials by nuclear magnetic resonance (NMR) and showed that IBAT produces a stable and sustainable RM over time. This <i>E. coli</i> RM was then used as a food source to produce a <i>Caenorhabditis elegans</i> RM for a metabolomics experiment. The metabolite extraction of this material, alongside 41 independently grown individual <i>C. elegans</i> samples of the same genotype, allowed us to estimate the proportion of sample variation in preanalytical steps. From the NMR data, we found that 40% of the metabolite variance is due to the metabolite extraction process and analysis and 60% is due to sample-to-sample variance. The availability of RMs in untargeted metabolomics is one of the predominant needs of the metabolomics community that reach beyond quality control practices. IBAT addresses this need by facilitating the production of biologically relevant RMs and increasing their widespread use.
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[
Orphanet J Rare Dis,
2020]
BACKGROUND: Pathogenic variations in the gene encoding the skeletal muscle ryanodine receptor (RyR1) are associated with malignant hyperthermia (MH) susceptibility, a life-threatening hypermetabolic condition and RYR1-related myopathies (RYR1-RM), a spectrum of rare neuromuscular disorders. In RYR1-RM, intracellular calcium dysregulation, post-translational modifications, and decreased protein expression lead to a heterogenous clinical presentation including proximal muscle weakness, contractures, scoliosis, respiratory insufficiency, and ophthalmoplegia. Preclinical model systems of RYR1-RM and MH have been developed to better understand underlying pathomechanisms and test potential therapeutics. METHODS: We conducted a comprehensive scoping review of scientific literature pertaining to RYR1-RM and MH preclinical model systems in accordance with the PRISMA Scoping Reviews Checklist and the framework proposed by Arksey and O'Malley. Two major electronic databases (PubMed and EMBASE) were searched without language restriction for articles and abstracts published between January 1, 1990 and July 3, 2019. RESULTS: Our search yielded 5049 publications from which 262 were included in this review. A majority of variants tested in RYR1 preclinical models were localized to established MH/central core disease (MH/CCD) hot spots. A total of 250 unique RYR1 variations were reported in human/rodent/porcine models with 95% being missense substitutions. The most frequently reported RYR1 variant was R614C/R615C (human/porcine total n=39), followed by Y523S/Y524S (rabbit/mouse total n=30), I4898T/I4897T/I4895T (human/rabbit/mouse total n=20), and R163C/R165C (human/mouse total n=18). The dyspedic mouse was utilized by 47% of publications in the rodent category and its RyR1-null (1B5) myotubes were transfected in 23% of publications in the cellular model category. In studies of transfected HEK-293 cells, 57% of RYR1 variations affected the RyR1 channel and activation core domain. A total of 15 RYR1 mutant mouse strains were identified of which ten were heterozygous, three were compound heterozygous, and a further two were knockout. Porcine, avian, zebrafish, C. elegans, canine, equine, and drosophila model systems were also reported. CONCLUSIONS: Over the past 30years, there were 262 publications on MH and RYR1-RM preclinical model systems featuring more than 200 unique RYR1 variations tested in a broad range of species. Findings from these studies have set the foundation for therapeutic development for MH and RYR1-RM.
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[
Dev Biol,
1994]
In the early Caenorhabditis elegans embryo most of the ectoderm arises from the AB blastomere, one of the six founder cells. We report that nonequivalent blastomeres are generated at the third division round in the AB lineage. Each AB granddaughter divides to produce one cell that has the potential to make abundant epidermis and one that instead produces primarily nervous system. This unequal distribution of the potential to make epidermis occurs in an AB granddaughter that is isolated by laser-ablation of all other cells or during the development of an isolated AB blastomere in culture. The fidelity of this event is normally masked by a signal from the MS founder cell, which induces mesoderm in particular AB descendants. When MS induction is prevented by laser cell-ablation or by a mutation in the
glp-1 gene, the epidermal fate map of the AB great granddaughters becomes left-right symmetrical. Cell lineage analyses demonstrate that, in fact, the AB lineage becomes entirely left-right symmetrical in the absence of MS induction. This accounts for the extra epidermal cells previously observed in a
glp-1 mutant. Our results suggest that epidermal differentiation in the nematode may be controlled by a cell-autonomous mechanism that differentially allocates epidermal potential during AB development and that MS induction generates the left-right asymmetry in the fates of AB descendants in part by overriding this potential.
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[
International C. elegans Meeting,
1995]
The AB founder cell is the primary progenitor of ectoderm in C. elegans, producing most of the epidermis and nervous system. Previous studies showed that, in the absence of inductive influences from P1 descendants, each of the four granddaughters of AB divides asymmetrically (1-4). Each gives rise to one AB8 cell that makes abundant epidermis and one that appears to make exclusively nervous system, marking the first event that subdivides the AB-derived ectoderm. We are attempting to identify genes required for this asymmetric division by performing a combined screen for zygotic- and maternal-effect lethal mutants in which this subdivision of the ectoderm is defective. Our strategy is to identify mutants in which the number of AB-derived epidermal cells is altered; perturbations in the asymmetric AB4 -> AB8 division would be expected to result in excess or too few AB-derived epidermal cells. As a marker of the AB epidermal precursor fate we are using a fusion construct that results in nuclear-localized GFP expression in the 20 AB-derived epidermal seam cells. Thus far, from a screen of 10,000 haploid genomes, three classes of mutants have been identified based on the expression pattern of the GFP marker. In class 1 mutants, a large excess of cells (>= 30 total) express the seam-specific marker; in class 2 mutants, a modest excess of cells (< 30 total) express the marker; in class 3 mutants, expression of GFP is eliminated entirely. These mutants are being further characterized to assess whether they are defective in the subdivision of the AB ectodermal lineages. 1. Mello et al., Cell 77, 95 (1994). 2. Hutter and Schnabel, Devel. 120, 2051 (1994). 3. Moskowitz et al., Devel. 120, 3325 (1994). 4. Gendreau et al., Dev. Biol. 166, 770 (1994).
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[
Elife,
2020]
Ryanodine receptor type I-related myopathies (RYR1-RMs) are a common group of childhood muscle diseases associated with severe disabilities and early mortality for which there are no available treatments. The goal of this study is to identify new therapeutic targets for RYR1-RMs. To accomplish this, we developed a discovery pipeline using nematode, zebrafish, and mammalian cell models. We first performed large-scale drug screens in <i>C. elegans</i> which uncovered 74 hits. Targeted testing in zebrafish yielded positive results for two
p38 inhibitors. Using mouse myotubes, we found that either pharmacological inhibition or siRNA silencing of
p38 impaired caffeine-induced Ca<sup>2+</sup> release from wild type cells while promoting intracellular Ca<sup>2+</sup> release in <i>Ryr1</i> knockout cells. Lastly, we demonstrated that
p38 inhibition blunts the aberrant temperature-dependent increase in resting Ca<sup>2+</sup> in myotubes from an RYR1-RM mouse model. This unique platform for RYR1-RM therapy development is potentially applicable to a broad range of neuromuscular disorders.
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[
J Environ Sci (China),
2011]
Sulfamethoxazole (SMX) is one of the most common detected antibiotics in the environment. In order to study whether SMX can affect behavior and growth and whether these effects could be transferred to the progeny, Caenorhabditis elegans was exposed at environmentally relevant concentrations for 24, 48, 72 and 96 hr, respectively. After exposure, the exposed parent generation (P0) was measured for behavior and growth indicators, which were presented as percentage of controls (POC). Then their corresponding unexposed progeny (F1) was separated and measured for the same indicators. The lowest POC for P0 after 96 hr-exposure at 100 mg/L were 37.8%, 12.7%, 45.8% and 70.1% for body bending frequency (BBF), reversal movement (RM), Omega turns (OT) and body length (BL), respectively. And F1 suffered defects with the lowest POC as 55.8%, 24.1%, 48.5% and 60.7% for BBF, RM, OT and BL, respectively. Defects in both P0 and F1 showed a time- and concentration-dependent fashion and behavior indicators showed better sensitivity than growth indicator. The observed effects on F1 demonstrated the transferable properties of SMX. Defects of SMX at environmental concentrations suggested that it is necessary to perform further systematical studies on its ecological risk in actual conditions.