of coronavirus disease (COVID-19) had been reported globally since December 2019 (1), severely burdening the healthcare system (2). The extremely fast transmission capability of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has aroused concern about its various transmission routes.The main transmission routes for SARS-CoV-2 are respiratory droplets and close contact (3). Knowing the extent of environmental contamination of SARS-CoV-2 in COVID-19 wards is critical for improving safety practices for medical staff and answering questions about SARS-CoV-2 transmission among the public. However, whether SARS-CoV-2 can be transmitted by aerosols remains controversial, and the exposure risk for close contacts has not been systematically evaluated. Researchers have detected SARS-CoV-2 on surfaces of objects in a symptomatic patient's room and toilet area (4). However, that study was performed in a small sample from regions with few confirmed cases, which might not reflect real conditions in outbreak regions where hospitals are operating at full capacity. In this study, we tested surface and air samples from an intensive care unit (ICU) and a general COVID-19 ward (GW) at Huoshenshan Hospital in Wuhan, China (Figure 1). The StudyFrom February 19 through March 2, 2020, we collected swab samples from potentially contaminated objects in the ICU and GW as described previously (5). The ICU housed 15 patients with severe disease and the GW housed 24 patients with milder disease. We also sampled indoor air and the air outlets to detect aerosol exposure. Air samples were collected by using a SASS 2300 Wetted Wall Cyclone Sampler (Research International, Inc., https://www.resrchintl.com) at 300 L/min for of 30 min. We used sterile premoistened swabs to sample the floors, computer mice, trash cans, sickbed handrails, patient masks, personal protective equipment, and air outlets. We tested air and surface samples for the open reading frame (ORF) 1ab and nucleoprotein (N) genes of SARS-CoV-2 by quantitative real-time PCR. (Appendix, https://wwwnc.cdc.gov/EID/ article/26/7/20-0885-App1.pdf).Almost all positive results were concentrated in the contaminated areas (ICU 54/57, 94.7%; GW 9/9, 100%); the rate of positivity was much higher for the ICU (54/124, 43.5%) than for the GW (9/114, 7.9%) (Tables 1, 2). The rate of positivity was

Twenty percent of the familial form of amyotrophic lateral sclerosis (ALS) is caused by mutations in the Cu, Zn-superoxide dismutase gene (SOD1) through the gain of a toxic function. The nature of this toxic function of mutant SOD1 has remained largely unknown. Here we show that WT SOD1 not only hastens onset of the ALS phenotype but can also convert an unaffected phenotype to an ALS phenotype in mutant SOD1 transgenic mouse models. Further analyses of the single-and double-transgenic mice revealed that conversion of mutant SOD1 from a soluble form to an aggregated and detergent-insoluble form was associated with development of the ALS phenotype in transgenic mice. Conversion of WT SOD1 from a soluble form to an aggregated and insoluble form also correlates with exacerbation of the disease or conversion to a disease phenotype in double-transgenic mice. This conversion, observed in the mitochondrial fraction of the spinal cord, involved formation of insoluble SOD1 dimers and multimers that are crosslinked through intermolecular disulfide bonds via oxidation of cysteine residues in SOD1. Our data thus show a molecular mechanism by which SOD1, an important protein in cellular defense against free radicals, is converted to aggregated and apparently ALS-associated toxic dimers and multimers by redox processes. These findings provide evidence of direct links among oxidation, protein aggregation, mitochondrial damage, and SOD1-mediated ALS, with possible applications to the aging process and other late-onset neurodegenerative disorders. Importantly, rational therapy based on these observations can now be developed and tested.crosslinked ͉ disulfide bonds ͉ oxidation ͉ protein aggregation ͉ neurodegeneration A myotrophic lateral sclerosis (ALS) is a progressive paralytic disorder caused by degeneration of the motor neurons in brain and spinal cord (1). Most of the ALS cases are sporadic, with Ϸ5-10% being familial. The progressive paralysis in ALS usually affects respiratory function, leading to ventilatory failure and death; 50% of patients die within 3 years of onset of symptoms, and 90% die within 5 years. The juvenile form of ALS usually has a prolonged course of two to four decades. There is no known effective treatment for this fatal disease, although marginal delay in mortality has been noted with the drug riluzole (2).Familial ALS can be transmitted as either a dominant or a recessive trait. We and our collaborators have previously shown that mutations in the Cu, Zn-superoxide dismutase gene (SOD1) are associated with Ϸ20% of familial ALS cases (3, 4). The pathogenic mechanisms underlying this disease are still largely unknown. Most, but not all, transgenic mice overexpressing ALS-associated SOD1 mutants develop ALS-like disease (5), and transgenic mice overexpressing human WT SOD1 (hwtSOD1) or SOD1-deficient mice do not develop ALS-like disease (5, 6), suggesting that mutant SOD1 requires a threshold of expression to cause the disease through the gain of a toxic property.Thus far, Ͼ100 mutations, widely distrib...

Venetoclax-based therapy can induce responses in approximately 70% of older previously untreated patients with acute myeloid leukemia (AML). However, upfront resistance as well as relapse following initial response demonstrates the need for a deeper understanding of resistance mechanisms. In the present study, we report that responses to venetoclax + azacitidine in patients with AML correlate closely with developmental stage, where phenotypically primitive AML is sensitive, but monocytic AML is more resistant. Mechanistically, resistant monocytic AML has a distinct transcriptomic profi le, loses expression of venetoclax target BCL2, and relies on MCL1 to mediate oxidative phosphorylation and survival. This differential sensitivity drives a selective process in patients which favors the outgrowth of monocytic subpopulations at relapse. Based on these fi ndings, we conclude that resistance to venetoclax + azacitidine can arise due to biological properties intrinsic to monocytic differentiation. We propose that optimal AML therapies should be designed so as to independently target AML subclones that may arise at differing stages of pathogenesis. SIGNIFICANCE: Identifying characteristics of patients who respond poorly to venetoclax-based therapy and devising alternative therapeutic strategies for such patients are important topics in AML. We show that venetoclax resistance can arise due to intrinsic molecular/metabolic properties of monocytic AML cells and that such properties can potentially be targeted with alternative strategies.

In vitro generation of functional gametes is a promising approach for treating infertility, although faithful replication of meiosis has proven to be a substantial obstacle to deriving haploid gamete cells in culture. Here we report complete in vitro meiosis from embryonic stem cell (ESC)-derived primordial germ cells (PGCLCs). Co-culture of PGCLCs with neonatal testicular somatic cells and sequential exposure to morphogens and sex hormones reproduced key hallmarks of meiosis, including erasure of genetic imprinting, chromosomal synapsis and recombination, and correct nuclear DNA and chromosomal content in the resulting haploid cells. Intracytoplasmic injection of the resulting spermatid-like cells into oocytes produced viable and fertile offspring, showing that this robust stepwise approach can functionally recapitulate male gametogenesis in vitro. These findings provide a platform for investigating meiotic mechanisms and the potential generation of human haploid spermatids in vitro.

Non-technical summary Our focus was on whether amyotrophic lateral sclerosis (ALS) might be precipitated by early developmental changes in large spinal motoneurons, which are vulnerable to early die-off in ALS. It has been shown that some electrical properties in motoneurons are profoundly altered soon after birth in mutant superoxide dismutase-1 (SOD1) mice, a standard animal model of ALS. These same properties undergo rapid developmental changes in normal mice during this time period. Our goal was to compare the development of motoneuron electrical properties in normal and SOD1 mice. Properties were measured from birth to 12 days of age, when the mouse is considered juvenile, but long before symptom onset. Most electrical properties in the SOD1 motoneurons showed an accelerated pace of maturation during this early developmental period compared with the normal motoneurons. If this trend persists, it could, along with other disease factors, hasten the onset of normal motoneuron degeneration due to ageing and result in the development of ALS.Abstract Spinal motoneurons are highly vulnerable in amyotrophic lateral sclerosis (ALS). Previous research using a standard animal model, the mutant superoxide dismutase-1 (SOD1) mouse, has revealed deficits in many cellular properties throughout its lifespan. The electrical properties underlying motoneuron excitability are some of the earliest to change; starting at 1 week postnatal, persistent inward currents (PICs) mediated by Na + are upregulated and electrical conductance, a measure of cell size, increases. However, during this period these properties and many others undergo large developmental changes which have not been fully analysed. Therefore, we undertook a systematic analysis of electrical properties in more than 100 normal and mutant SOD1 motoneurons from 0 to 12 days postnatal, the neonatal to juvenile period. We compared normal mice with the most severe SOD1 model, the G93A high-expressor line. We found that the Na + PIC and the conductance increased during development. However, mutant SOD1 motoneurons showed much greater increases than normal motoneurons; the mean Na + PIC in SOD1 motoneurons was double that of wild-type motoneurons. Additionally, in mutant SOD1 motoneurons the PIC mediated by Ca 2+ increased, spike width decreased and the time course of the after-spike after-hyperpolarization shortened. These changes were advances of the normal effects of maturation. Thus, our results show that the development of normal and mutant SOD1 motoneurons follows generally similar patterns, but that the rate of development is accelerated in the mutant SOD1 motoneurons. Statistical analysis of all measured properties indicates that approximately 55% of changes attributed to the G93A SOD1 mutation can be attributed to an increased rate of maturation.

Mutations in the enzyme superoxide dismutase 1 (SOD1) initiate a progressive motoneurone degeneration in amyotrophic lateral sclerosis (ALS). Transgenic mice overexpressing this mutation develop a similar progressive motoneurone degeneration. In spinal motoneurones cultured from presymptomatic mice expressing the glycine to alanine mutation at base pair 93 (G93A) SOD1 mutation, a marked increase in the persistent component of the Na + current was observed, without changes in passive properties. This increase only enhanced neuronal excitability in high input conductance cells, as low input conductance cells exhibited a compensatory outward shift in the current remaining after Na + blockade. High input conductance motoneurones tend to be large, so these results may explain the tendency of large motoneurones to degenerate first in ALS. Riluzole, at the therapeutic concentration used to treat ALS, decreased neuronal excitability and persistent Na + current in G93A motoneurones to levels observed in the control motoneurones. Aberrations in the intrinsic electrical properties may be among the first symptoms to emerge in SOD1-linked ALS.

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Point mutations in Cu, Zn-superoxide dismutase (SOD1) cause a familial form of the neurodegenerative disease amyotrophic lateral sclerosis (ALS). Aggregates of mutant SOD1 proteins are observed in histopathology and are invoked in several proposed mechanisms for motor neuronal death; however, the significant stability and activity of the mature mutant proteins are not readily explained in such models. Recent biochemical studies suggest that it is the immature disulfide-reduced forms of the familial ALS mutant SOD1 proteins that play a critical role; these forms tend to misfold, oligomerize, and readily undergo incorrect disulfide formation upon mild oxidative stress in vitro. Here we provide physiological support for this mechanism of aggregate formation and show that a significant fraction of the insoluble SOD1 aggregates in spinal cord of the ALS-model transgenic mice contain multimers cross-linked via intermolecular disulfide bonds. These insoluble disulfide-linked SOD1 multimers are found only in the spinal cord of symptomatic transgenic animals, are not observed in unafflicted tissue such as brain cortex and liver, and can incorporate WT SOD1 protein. The findings provide a biochemical basis for a pathological hallmark of this disease; namely, incorrect disulfide cross-linking of the immature, misfolded mutant proteins leads to insoluble aggregates. disulfide bond ͉ protein aggregation ͉ oxidative stress ͉ neurodegnerative disease A myotrophic lateral sclerosis (ALS) is one of the most common adult-onset neurodegenerative diseases, and, in 1993, several mutations in the Cu, Zn-superoxide dismutase (SOD1) gene were identified as a cause of a subset of familial ALS (fALS) (1, 2). Since that time, Ͼ100 types of SOD1 mutations have been linked with fALS; however, its molecular mechanism remains unresolved. SOD1, which is a homodimer with a copper and zinc ion in each subunit, functions as an antioxidant enzyme by converting superoxide radical to oxygen and hydrogen peroxide (3). Some of the SOD1 mutant proteins have comparable levels of dismutase activity, and the SOD1-knockout mouse does not develop ALS-like motor neuron symptoms (4), leading to the idea that SOD1-mediated toxicity in fALS is due to a new activity acquired by mutations in the SOD1 gene. One of the proposed toxic functions in the SOD1 mutant proteins is an aberrant copper chemistry. A subset of ALS mutations in SOD1 can lead to catalytic nitration of tyrosine residues and peroxide or superoxide production at the copper site (5); however, the copper-toxicity model is difficult to reconcile with the fact that ALS symptoms still are observed in the transgenic mouse expressing SOD1 proteins in which all four copper ligands are mutated (6). Although some copper chelators can reduce the toxicity of mutant proteins in transgenic ALS model mice (7), a role for aberrant copper chemistry in the disease has not been widely addressed.Another hypothesis for a gain of toxic function is that the mutations in SOD1 facilitate protein aggregation (8). An enzyma...