Transgenic Rescue of the LARGEmyd Mouse: A LARGE Therapeutic Window?

LARGE is a glycosyltransferase involved in glycosylation of α-dystroglycan (α-DG). Absence of this protein in the LARGEmyd mouse results in α-DG hypoglycosylation, and is associated with central nervous system abnormalities and progressive muscular dystrophy. Up-regulation of LARGE has previously been proposed as a therapy for the secondary dystroglycanopathies: overexpression in cells compensates for defects in multiple dystroglycanopathy genes. Counterintuitively, LARGE overexpression in an FKRP-deficient mouse exacerbates pathology, suggesting that modulation of α-DG glycosylation requires further investigation. Here we demonstrate that transgenic expression of human LARGE (LARGE-LV5) in the LARGEmyd mouse restores α-DG glycosylation (with marked hyperglycosylation in muscle) and that this corrects both the muscle pathology and brain architecture. By quantitative analyses of LARGE transcripts we also here show that levels of transgenic and endogenous LARGE in the brains of transgenic animals are comparable, but that the transgene is markedly overexpressed in heart and particularly skeletal muscle (20–100 fold over endogenous). Our data suggest LARGE overexpression may only be deleterious under a forced regenerative context, such as that resulting from a reduction in FKRP: in the absence of such a defect we show that systemic expression of LARGE can indeed act therapeutically, and that even dramatic LARGE overexpression is well-tolerated in heart and skeletal muscle. Moreover, correction of LARGEmyd brain pathology with only moderate, near-physiological LARGE expression suggests a generous therapeutic window

Prenatal muscle development in a mouse model for the secondary dystroglycanopathies

The defective glycosylation of α-dystroglycan is associated with a group of muscular dystrophies that are collectively referred to as the secondary dystroglycanopathies. Mutations in the gene encoding fukutin-related protein (FKRP) are one of the most common causes of secondary dystroglycanopathy in the UK and are associated with a wide spectrum of disease. Whilst central nervous system involvement has a prenatal onset, no studies have addressed prenatal muscle development in any of the mouse models for this group of diseases. In view of the pivotal role of α-dystroglycan in early basement membrane formation, we sought to determine if the muscle formation was altered in a mouse model of FKRP-related dystrophy

Chemical Evolution of Atmospheric Organic Carbon over Multiple Generations of Oxidation

The evolution of atmospheric organic carbon (OC) as it undergoes oxidation has a controlling influence on concentrations of key atmospheric species, including particulate matter, ozone, and oxidants. However, the full characterization of OC over hours to days of atmospheric processing has been stymied by its extreme chemical complexity. Here we study the multigenerational oxidation of -pinene in the laboratory, characterizing products with several state-of-the-art analytical techniques. While quantification of some early-generation products remains elusive, full carbon closure is achieved (within uncertainty) by the end of the experiments. This enables new insights into the effects of oxidation on OC properties (volatility, oxidation state, and reactivity), and the atmospheric lifecycle of OC. Following an initial period characterized by functionalization reactions and particle growth, fragmentation reactions dominate, forming smaller species. After approximately one day of atmospheric aging, most carbon is sequestered in two long-lived reservoirs, volatile oxidized gases and low-volatility particulate matter

Staphylococcus aureus persistence in osteocytes: weathering the storm of antibiotics and autophagy/xenophagy

Staphylococcus aureus is a major causative pathogen of osteomyelitis. Intracellular infections of resident bone cells including osteocytes can persist despite gold-standard clinical intervention. The mechanisms by which intracellular S. aureus evades antibiotic therapy are unknown. In this study, we utilised an in vitro S. aureus infection model of human osteocytes to investigate whether antibiotic-mediated dysregulation of autophagy contributes to this phenomenon. Infected or non-infected osteocyte-like cells were exposed to combinations of rifampicin, vancomycin, and modulators of autophagy. Intracellular bacterial growth characteristics were assessed using colony-forming unit (CFU) analysis, viable bacterial DNA abundance, and the rate of escape into antibiotic-free medium, together with measures of autophagic flux. Rifampicin, alone or in combination with vancomycin, caused a rapid decrease in the culturability of intracellular bacteria, concomitant with stable or increased absolute bacterial DNA levels. Both antibiotics significantly inhibited autophagic flux. However, modulation of autophagic flux did not affect viable bacterial DNA levels. In summary, autophagy was shown to be a factor in the host–pathogen relationship in this model, as its modulation affected the growth state of intracellular S. aureus with respect to both their culturability and propensity to escape the intracellular niche. While rifampicin and vancomycin treatments moderately suppressed autophagic flux acutely, this did not explain the paradoxical response of antibiotic treatment in decreasing S. aureus culturability whilst failing to clear bacterial DNA and hence intracellular bacterial load. Thus, off-target effects of rifampicin and vancomycin on autophagic flux in osteocyte-like cells could not explain the persistent S. aureus infection in these cells

Ethylene oxide monitor with part-per-trillion precision for in situ measurements

An Aerodyne tunable infrared laser direct absorption spectrometer with a multipass cell with a 413 m pathlength for the detection of ethylene oxide (EtO) is presented (TILDAS-FD-EtO). This monitor achieves precisions of &lt;75 ppt or &lt;0.075 ppb s−1 and &lt;20 ppt in 100 s (1σ). We demonstrate precisions averaging down to 4 ppt h−1 (1σ precision) when operated with frequent humidity-matched zeroes. A months-long record of 2022 ambient concentrations at a site in the eastern United States is presented. Average ambient EtO concentration is on the order of 18 ppt (22 ppt standard deviation, SD). Enhancement events of EtO lasting a few hours are observed, with peaks as high as 600 ppt. Back-trajectory simulations suggest an EtO source nearly 35 km away. This source along with another are confirmed as emitters through mobile near-source measurements, with downwind concentrations in the 0.5 to 700 ppb range depending on source identity and distance downwind.</p

Changes in ozone and precursors during two aged wildfire smoke events in the Colorado Front Range in summer 2015

The relative importance of wildfire smoke for air quality over the western US is expected to increase as the climate warms and anthropogenic emissions decline. We report on in situ measurements of ozone (O3), a suite of volatile organic compounds (VOCs), and reactive oxidized nitrogen species collected during summer 2015 at the Boulder Atmospheric Observatory (BAO) in Erie, CO. Aged wildfire smoke impacted BAO during two distinct time periods during summer 2015: 6–10 July and 16–30 August. The smoke was transported from the Pacific Northwest and Canada across much of the continental US. Carbon monoxide and particulate matter increased during the smoke-impacted periods, along with peroxyacyl nitrates and several VOCs that have atmospheric lifetimes longer than the transport timescale of the smoke. During the August smoke-impacted period, nitrogen dioxide was also elevated during the morning and evening compared to the smoke-free periods. There were nine empirically defined high-O3 days during our study period at BAO, and two of these days were smoke impacted. We examined the relationship between O3 and temperature at BAO and found that for a given temperature, O3 mixing ratios were greater (∼ 10 ppbv) during the smoke-impacted periods. Enhancements in O3 during the August smoke-impacted period were also observed at two long-term monitoring sites in Colorado: Rocky Mountain National Park and the Arapahoe National Wildlife Refuge near Walden, CO. Our data provide a new case study of how aged wildfire smoke can influence atmospheric composition at an urban site, and how smoke can contribute to increased O3 abundances across an urban–rural gradient

CDH1 mutation distribution and type suggests genetic differences between the etiology of orofacial clefting and gastric cancer

Pathogenic variants in CDH1, encoding epithelial cadherin (E-cadherin), have been implicated in hereditary diffuse gastric cancer (HDGC), lobular breast cancer, and both syndromic and non-syndromic cleft lip/palate (CL/P). Despite the large number of CDH1 mutations described, the nature of the phenotypic consequence of such mutations is currently not able to be predicted, creating significant challenges for genetic counselling. This study collates the phenotype and molecular data for available CDH1 variants that have been classified, using the American College of Medical Genetics and Genomics criteria, as at least ‘likely pathogenic’, and correlates their molecular and structural characteristics to phenotype. We demonstrate that CDH1 variant type and location differ between HDGC and CL/P, and that there is clustering of CL/P variants within linker regions between the extracellular domains of the cadherin protein. While these differences do not provide for exact prediction of the phenotype for a given mutation, they may contribute to more accurate assessments of risk for HDGC or CL/P for individuals with specific CDH1 variants

Characterization of ozone production in San Antonio, Texas, using measurements of total peroxy radicals

Observations of total peroxy radical concentrations ([XO2]&thinsp;≡&thinsp;[RO2]&thinsp;+&thinsp;[HO2]) made by the Ethane CHemical AMPlifier (ECHAMP) and concomitant observations of additional trace gases made on board the Aerodyne Mobile Laboratory (AML) during May 2017 were used to characterize ozone production at three sites in the San Antonio, Texas, region. Median daytime [O3] was 48&thinsp;ppbv at the site downwind of central San Antonio. Higher concentrations of NO and XO2 at the downwind site also led to median daytime ozone production rates (P(O3)) of 4.2&thinsp;ppbv&thinsp;h−1, a factor of 2 higher than at the two upwind sites. The 95th percentile of P(O3) at the upwind site was 15.1&thinsp;ppbv&thinsp;h−1, significantly lower than values observed in Houston. In situ observations, as well as satellite retrievals of HCHO and NO2, suggest that the region was predominantly NOx-limited. Only approximately 20&thinsp;% of observations were in the VOC-limited regime, predominantly before 11:00&thinsp;EST, when ozone production was low. Biogenic volatile organic compounds (VOCs) comprised 55&thinsp;% of total OH reactivity at the downwind site, with alkanes and non-biogenic alkenes responsible for less than 10&thinsp;% of total OH reactivity in the afternoon, when ozone production was highest. To control ozone formation rates at the three study sites effectively, policy efforts should be directed at reducing NOx emissions. Observations in the urban center of San Antonio are needed to determine whether this policy is true for the entire region.</p