While prime editing enables precise sequence changes in DNA, cellular determinants of prime editing remain poorly understood. Using pooled CRISPRi screens, we discovered that DNA mismatch repair ...(MMR) impedes prime editing and promotes undesired indel byproducts. We developed PE4 and PE5 prime editing systems in which transient expression of an engineered MMR-inhibiting protein enhances the efficiency of substitution, small insertion, and small deletion prime edits by an average 7.7-fold and 2.0-fold compared to PE2 and PE3 systems, respectively, while improving edit/indel ratios by 3.4-fold in MMR-proficient cell types. Strategic installation of silent mutations near the intended edit can enhance prime editing outcomes by evading MMR. Prime editor protein optimization resulted in a PEmax architecture that enhances editing efficacy by 2.8-fold on average in HeLa cells. These findings enrich our understanding of prime editing and establish prime editing systems that show substantial improvement across 191 edits in seven mammalian cell types.
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•Pooled CRISPRi screens reveal that MMR inhibits prime editing efficiency and precision•PE4 and PE5 enhance editing outcomes through co-expression of dominant negative MLH1•Programming additional silent mutations can enhance prime editing by evading MMR•PEmax editor improves prime editing efficacy in synergy with PE4, PE5, and epegRNAs
PE4 and PE5 are efficient and precise prime editing systems developed by leveraging insights into the way DNA repair pathways impact genome editing outcomes
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
To evaluate the efficacy and safety of methylprednisolone in treating the coronavirus disease 2019 (COVID-19) patients.
A retrospective cohort study was conducted, and all COVID-19 patients were ...recruited who were admitted to the Yichang Third People's Hospital from February 1st to March 31st, 2020. One-to-one propensity score matching (PSM) was used for minimizing confounding effects. The primary outcome was hospital mortality, with the secondary outcomes being the time needed for a positive SARS-CoV-2 nucleic acid test to turn negative and the length of hospital stay.
Totaling 367 patients with COVID-19 hospitalized at the Yichang Third People's Hospital were identified, of whom 276 were mild or stable COVID-19, and 67 were serious or critically ill. Among them, 255 patients were treated using methylprednisolone, and 188 did not receive any corticosteroid-related treatment. After PSM, no statistically significant difference was found in the baseline characteristics between the two groups. Regarding the outcomes, there also were no statistically significant difference between the two groups. Patients without the use of methylprednisolone were more quickly to obtain negative results of their nasopharyngeal swab tests of SARS-CoV-2 nucleic acid after treatment, compared to those receiving methylprednisolone.
Methylprednisolone could not improve the prognosis of patients with COVID-19, and the efficacy and safety of the use of methylprednisolone in patients with COVID-19 still remain uncertain, thus the use of corticosteroids clinically in patients with COVID-19 should be with cautions.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
This study simulates and compares symmetric and asymmetric MoS 2 nanosheet transistor structures. The results show that the asymmetric MoS 2 nanosheet transistor achieves higher current density and ...better gate control, and thus, this structure is optimized to meet the current density requirement for low-power applications in the International Roadmap for Devices and Systems (IRDS). Dielectric materials of Al 2 O 3 and HfO 2 are investigated, and equivalent oxide thicknesses ranging from 0.8 to 2 nm are analyzed. Transistors with Al 2 O 3 as the dielectric exhibit better performance than those with HfO 2 due to the lower level of remote phonon scattering. The gate length and underlap region are scaled down from 14 to 5 nm and from 7 to 1 nm, respectively, in an attempt to further enhance the ON-current. The substoichiometric metal oxide AlO x is used to induce more electrons in the undoped MoS 2 channel. AlO x increases the ON-current with no gate control degradation. After optimization, the transistor demonstrates a subthreshold swing of 62.3 mV/dec, a drain-induced barrier lowering (DIBL) of 21.6 mV/dec, and a current density of <inline-formula> <tex-math notation="LaTeX">495 ~\mu \text{A}/\mu \text{m} </tex-math></inline-formula> with a supply voltage of 0.65 V. Compared with Si gate-all-around transistors, the MoS 2 nanosheet transistor exhibits excellent gate control, good electrostatics, and comparable current density.
Angelman syndrome (AS) and Prader-Willi syndrome (PWS) are neurodevelopmental disorders of genomic imprinting. AS results from loss of function of the ubiquitin protein ligase E3A (UBE3A) gene, ...whereas the genetic defect in PWS is unknown. Although induced pluripotent stem cells (iPSCs) provide invaluable models of human disease, nuclear reprogramming could limit the usefulness of iPSCs from patients who have AS and PWS should the genomic imprint marks be disturbed by the epigenetic reprogramming process. Our iPSCs derived from patients with AS and PWS show no evidence of DNA methylation imprint erasure at the cis-acting PSW imprinting center. Importantly, we find that, as in normal brain, imprinting of UBE3A is established during neuronal differentiation of AS iPSCs, with the paternal UBE3A allele repressed concomitant with up-regulation of the UBE3A antisense transcript. These iPSC models of genomic imprinting disorders will facilitate investigation of the AS and PWS disease processes and allow study of the developmental timing and mechanism of UBE3A repression in human neurons.
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BFBNIB, NMLJ, NUK, PNG, SAZU, UL, UM, UPUK
Over the last two decades, the Central Weather Bureau of Taiwan and the U.S. National Severe Storms Laboratory have been involved in a research and development collaboration to improve the monitoring ...and prediction of river flooding, flash floods, debris flows, and severe storms for Taiwan. The collaboration resulted in the Quantitative Precipitation Estimation and Segregation Using Multiple Sensors (QPESUMS) system. The QPESUMS system integrates observations from multiple mixed-band weather radars, rain gauges, and numerical weather prediction model fields to produce high-resolution (1 km) and rapid-update (10 min) rainfall and severe storm monitoring and prediction products. The rainfall products are widely used by government agencies and emergency managers in Taiwan for flood and mudslide warnings as well as for water resource management. The 3D reflectivity mosaic and QPE products are also used in high-resolution radar data assimilation and for the verification of numerical weather prediction model forecasts. The system facilitated collaborations with academic communities for research and development of radar applications, including quantitative precipitation estimation and nowcasting. This paper provides an overview of the operational QPE capabilities in the Taiwan QPESUMS system.
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BFBNIB, DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Succinic Semialdehyde Dehydrogenase Deficiency (SSADHD) is an ultra-rare autosomal recessive neurometabolic disorder caused by ALDH5A1 mutations presenting with autism and epilepsy. Here, we report ...the generation and characterization of human induced pluripotent stem cells (hiPSCs) derived from fibroblasts of three unrelated SSADHD patients – one female and two males with the CRISPR-corrected isogenic controls. These individuals are clinically diagnosed and are being followed in a longitudinal clinical study.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
Abstract
Deficiency of the adaptor protein complex 4 (AP-4) leads to childhood-onset hereditary spastic paraplegia (AP-4-HSP): SPG47 (AP4B1), SPG50 (AP4M1), SPG51 (AP4E1) and SPG52 (AP4S1). This ...study aims to evaluate the impact of loss-of-function variants in AP-4 subunits on intracellular protein trafficking using patient-derived cells. We investigated 15 patient-derived fibroblast lines and generated six lines of induced pluripotent stem cell (iPSC)-derived neurons covering a wide range of AP-4 variants. All patient-derived fibroblasts showed reduced levels of the AP4E1 subunit, a surrogate for levels of the AP-4 complex. The autophagy protein ATG9A accumulated in the trans-Golgi network and was depleted from peripheral compartments. Western blot analysis demonstrated a 3–5-fold increase in ATG9A expression in patient lines. ATG9A was redistributed upon re-expression of AP4B1 arguing that mistrafficking of ATG9A is AP-4-dependent. Examining the downstream effects of ATG9A mislocalization, we found that autophagic flux was intact in patient-derived fibroblasts both under nutrient-rich conditions and when autophagy is stimulated. Mitochondrial metabolism and intracellular iron content remained unchanged. In iPSC-derived cortical neurons from patients with AP4B1-associated SPG47, AP-4 subunit levels were reduced while ATG9A accumulated in the trans-Golgi network. Levels of the autophagy marker LC3-II were reduced, suggesting a neuron-specific alteration in autophagosome turnover. Neurite outgrowth and branching were reduced in AP-4-HSP neurons pointing to a role of AP-4-mediated protein trafficking in neuronal development. Collectively, our results establish ATG9A mislocalization as a key marker of AP-4 deficiency in patient-derived cells, including the first human neuron model of AP-4-HSP, which will aid diagnostic and therapeutic studies.
Tuberous sclerosis complex (TSC) is a genetic disorder caused by mutations in
or
Patients frequently have epilepsy, autism spectrum disorder, and/or intellectual disability, as well as other systemic ...manifestations. In this study, we differentiated human induced pluripotent stem cells (iPSCs) from a female patient with TSC with one or two mutations in
into neurons using induced expression of NGN2 to examine neuronal dysregulation associated with the neurological symptoms in TSC. Using this method, neuronal differentiation was comparable between the three genotypes of iPSCs. We observed that
neurons show mTOR complex 1 (mTORC1) hyperactivation and associated increased cell body size and process outgrowth, as well as exacerbation of the abnormalities by loss of the second allele of
in
neurons. Interestingly, iPSC-derived neurons with either a single or biallelic mutation in
demonstrated hypersynchrony and downregulation of FMRP targets. However, only neurons with biallelic mutations of
demonstrated hyperactivity and transcriptional dysregulation observed in cortical tubers. These data demonstrate that loss of one allele of
is sufficient to cause some morphological and physiological changes in human neurons but that biallelic mutations in
are necessary to induce gene expression dysregulation present in cortical tubers. Finally, we found that treatment of iPSC-derived neurons with rapamycin reduced neuronal activity and partially reversed gene expression abnormalities, demonstrating that mTOR dysregulation contributes to both phenotypes. Therefore, biallelic mutations in
and associated molecular dysfunction, including mTOR hyperactivation, may play a role in the development of cortical tubers.
In this study, we examined neurons derived from induced pluripotent stem cells with two, one, or no functional
(tuberous sclerosis complex 2) alleles and found that loss of one or both alleles of
results in mTORC1 hyperactivation and specific neuronal abnormalities. However, only biallelic mutations in
resulted in elevated neuronal activity and upregulation of cell adhesion genes that is also observed in cortical tubers. These data suggest that loss of heterozygosity of
or
may play an important role in the development of cortical tubers, and potentially epilepsy, in patients with TSC.
Abstract
The aim of this study is to survey the prevalence of sarcopenia and the factors that influence its development in Southern Taiwan's community-dwelling aged people.
This is an observational ...cross-sectional study using the 6-meter walking test, body composition, handgrip strength, body measurements, and basic personal information to identify sarcopenia in the participants. This study included 200 participants aged 65 or over living in Taiwan, but excluded the following:
The prevalence of sarcopenia in the elderly community is approximately 6.0%. It is less prevalent in females (1.5%) than in males (14.3%). The incidence of sarcopenia increases with age. Significantly related risk factors for sarcopenia are gender, age, smoking, and body mass index (BMI) (
P
< .05). Further analysis of the risk factors for sarcopenia reveal that the odds ratios (ORs) of having low muscle mass increase with every 1 year in age by a factor of 1.19 (
P
< .05); those who smoke show a higher incidence than those who do not smoke (OR = 2.69,
P
< .05). For every 1 kg / m
2
increase in BMI, the odds of sarcopenia decrease by a factor of 0.45.
For the elderly, the lower the BMI, the higher the risk of sarcopenia. Maintaining good exercise habits and keeping body weight in check might help to prevent sarcopenia by increasing functional ability and improving muscle strength.
Soluble microbial products (SMP) are one of the main causes of reverse osmosis (RO) membrane fouling in wastewater reclamation, and coagulation and adsorption are commonly used to prevent such ...fouling. However, the mechanism by which typical coagulants and adsorbents affect the RO membrane fouling caused by SMP remains unclear. In this study, two bacterial strains isolated from fouled RO membranes in a full-scale wastewater reclamation plant were used to produce SMP. Both bacterial strains were found to generate high SMP yield (0.23–0.25 g/g), which caused severe RO membrane fouling. Coagulation by polyaluminum chloride (PACl) and ferric chloride (FeCl3) and adsorption by granular activated carbon (GAC) were applied as pretreatment methods to alleviate the membrane fouling caused by SMP. Compared with PACl, FeCl3 performed better in removing SMP, decreasing fluorescence intensity, and reducing molecular weight (MW). GAC preferentially adsorbed protein and humic substances in SMP and unselectively and efficiently removed fluorescent compounds; however, it could only remove a limited amount of the high-MW compound. The RO membrane fouling potential of treated SMP was investigated, and FeCl3 and GAC were found to effectively alleviate the fouling. In addition to a decrease in the dissolved organic carbon content of SMP, a decrease in specific fluorescence intensity, MW, and polysaccharide content caused by coagulation or adsorption could result in lower RO membrane fouling.
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•Bacteria on RO membranes had high SMP yield and the SMP severely fouled RO membranes.•FeCl3 performed better in SMP removal than PAC.•Activated carbon prefers to remove fluorescent compounds and low-MW SMP.•FeCl3 and GAC could effectively alleviate RO membrane fouling caused by SMP.•Reducing polysaccharide content of SMP significantly alleviate RO membrane fouling.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
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