Clonal dynamics in early human embryogenesis inferred from somatic mutation.

Cellular dynamics and fate decision in early human embryogenesis remain largely unknown owing to the challenges of performing studies in human embryos1. Here, we explored whole-genomes of 334 single-cell colonies and targeted deep sequences of 379 bulk tissues obtained from various anatomical locations of seven recently deceased adult human donors. Using somatic mutations as an intrinsic barcode, we reconstructed early cellular phylogenies that demonstrate (1) an endogenous mutational rate that is higher in the first cell division but decreases to approximately one per cell per cell division later in life; (2) universal unequal contribution of early cells to embryo proper, resulting from early cellular bottlenecks that stochastically set aside epiblast cells within the embryo; (3) examples of varying degrees of early clonal imbalances between tissues on the left and right sides of the body, different germ layers and specific anatomical parts and organs; (4) emergence of a few ancestral cells that will substantially contribute to adult cell pools in blood and liver; and (5) presence of mitochondrial DNA heteroplasmy in the fertilized egg. Our approach also provides insights into the age-related mutational processes and loss of sex chromosomes in normal somatic cells. In sum, this study provides a foundation for future studies to complete cellular phylogenies in human embryogenesis.

Multiscale landscaping of droplet wettability on fibrous layers of facial masks.

Liquid mobility is ubiquitous in nature, with droplets emerging at all size scales, and artificial surfaces have been designed to mimic such mobility over the past few decades. Meanwhile, millimeter-sized droplets are frequently used for wettability characterization, even with facial mask applications, although these applications have a droplet-size target range that spans from millimeters to aerosols measuring less than a few micrometers. Unlike large droplets, microdroplets can interact sensitively with the fibers they contact with and are prone to evaporation. However, wetting behaviors at the single-microfiber level remain poorly understood. Herein, we characterized the wettability of fibrous layers, which revealed that a multiscale landscape of droplets ranged from the millimeter to the micrometer scale. The contact angle (CA) values of small droplets on pristine fibrous media showed sudden decrements, especially on a single microfiber, owing to the lack of air cushions for the tiny droplets. Moreover, droplets easily adhered to the pristine layer during droplet impact tests and then yielding widespread areas of contamination on the microfibers. To resolve this, we carved nanowalls on the pristine fibers by plasma etching, which effectively suppressed such wetting phenomena. Significantly, the resulting topographies of the microfibers managed the dynamic wettability of droplets at the multiscale, which reduced the probability of contamination with impact droplets and suppressed the wetting transition upon evaporation. These findings for the dynamic wettability of fibrous media will be useful in the fight against infectious droplets.

Correction: A vesicular stomatitis virus-based prime-boost vaccination strategy induces potent and protective neutralizing antibodies against SARS-CoV-2.

[This corrects the article DOI: 10.1371/journal.ppat.1010092.].

Identification of novel Nrf2-activating neuroprotective agents: Elucidation of structural congeners of (-)-galiellalactone and congener-based novel Nrf2 activators.

Herein, (-)-galiellalactone 1 congeners responsible for the nuclear factor erythroid 2-related factor 2 (Nrf2)-activating neuroprotective effects were elucidated. Additionally, novel congener-based Nrf2 activators were identified using a drug repositioning strategy. (-)-Galiellalactone, which comprises a tricyclic lactone skeleton, significantly activates antioxidant response element (ARE)-mediated transcription in neuroblastoma SH-SY5Y cells. Interestingly, two cyclohexene-truncated [3.3] bicyclic lactone analogs, which possess an exocyclic α-methylene-γ-butyrolactone moiety, exhibited higher Nrf2/ARE transcriptional activities than the parent (-)-galiellalactone. We confirmed that the cyclohexene moiety embedding the [3.3] bicyclic lactone congener does not play the essential role of (-)-galiellalactone for Nrf2/ARE activation. Nrf2/ARE activation by novel analogs resulted in the upregulation of downstream antioxidative and phase II detoxifying enzymes, heme oxygenase-1, and NAD(P)H quinone oxidoreductase 1, which are closely related to the cytoprotective effects on neurodegenerative diseases. (-)-Galiellalactone and its [3.3] bicyclic variants 3l and 3p increased the expression of antioxidant genes and exhibited neuroprotective effects against 6-hydroxydopamine-mediated neurotoxicity in the neuroblastoma SH-SY5Y cell line.

Optimized trans-cranial direct current stimulation for prolonged consciousness disorder in a patient with titanium mesh cranioplasty.

BACKGROUND: Transcranial direct current stimulation (tDCS) has been used for the restoration of awareness in patients with a minimal consciousness state (MCS). Most brains of patients in MCS may structurally and electrophysiologically differ from un-damaged brains. Moreover, tDCS is currently contraindicated for patients with craniotomy or skull with metallic implants. CASE PRESENTATION: We present a case with prolonged MCS over 1 year, who had severe brain damage, ventriculoperitoneal shunt, and cranioplasty with a titanium mesh, which was treated with tDCS which optimized with the simulation of the electric field based on the patient's brain MRI. The patient was resulting in emergence from MCS. Six months later, she ate meals orally and started walking with assistance. DISCUSSION AND PERSPECTIVE: This personalized simulation based on MRI would make the treatment available even to patients with severe brain structural changes and metallic instrumentation.

Personalized iPSC-Derived Dopamine Progenitor Cells for Parkinson's Disease.

We report the implantation of patient-derived midbrain dopaminergic progenitor cells, differentiated in vitro from autologous induced pluripotent stem cells (iPSCs), in a patient with idiopathic Parkinson's disease. The patient-specific progenitor cells were produced under Good Manufacturing Practice conditions and characterized as having the phenotypic properties of substantia nigra pars compacta neurons; testing in a humanized mouse model (involving peripheral-blood mononuclear cells) indicated an absence of immunogenicity to these cells. The cells were implanted into the putamen (left hemisphere followed by right hemisphere, 6 months apart) of a patient with Parkinson's disease, without the need for immunosuppression. Positron-emission tomography with the use of fluorine-18-L-dihydroxyphenylalanine suggested graft survival. Clinical measures of symptoms of Parkinson's disease after surgery stabilized or improved at 18 to 24 months after implantation. (Funded by the National Institutes of Health and others.).

A vesicular stomatitis virus-based prime-boost vaccination strategy induces potent and protective neutralizing antibodies against SARS-CoV-2.

The development of safe and effective vaccines to prevent SARS-CoV-2 infections remains an urgent priority worldwide. We have used a recombinant vesicular stomatitis virus (rVSV)-based prime-boost immunization strategy to develop an effective COVID-19 vaccine candidate. We have constructed VSV genomes carrying exogenous genes resulting in the production of avirulent rVSV carrying the full-length spike protein (SF), the S1 subunit, or the receptor-binding domain (RBD) plus envelope (E) protein of SARS-CoV-2. Adding the honeybee melittin signal peptide (msp) to the N-terminus enhanced the protein expression, and adding the VSV G protein transmembrane domain and the cytoplasmic tail (Gtc) enhanced protein incorporation into pseudotype VSV. All rVSVs expressed three different forms of SARS-CoV-2 spike proteins, but chimeras with VSV-Gtc demonstrated the highest rVSV-associated expression. In immunized mice, rVSV with chimeric S protein-Gtc derivatives induced the highest level of potent neutralizing antibodies and T cell responses, and rVSV harboring the full-length msp-SF-Gtc proved to be the superior immunogen. More importantly, rVSV-msp-SF-Gtc vaccinated animals were completely protected from a subsequent SARS-CoV-2 challenge. Overall, we have developed an efficient strategy to induce a protective response in SARS-CoV-2 challenged immunized mice. Vaccination with our rVSV-based vector may be an effective solution in the global fight against COVID-19.

The Prognostic Utilities of DNA Mismatch Repair Status and KRAS and BRAF Mutation in Korean Colorectal Cancer Patients: The KASID Multicenter Study.

INTRODUCTION: KRAS, BRAF, and DNA mismatch repair (MMR) mutations aid clinical decision-making for colorectal cancer (CRC) patients. To ensure accurate predictions, the prognostic utilities of these biomarkers and their combinations must be individualized for patients with various TNM stages. METHODS: Here, we retrospectively analyzed the clinicopathological features of 904 Korean CRC patients who underwent CRC surgery in three teaching hospitals from 2011 to 2013; we also assessed the prognostic utilities of KRAS, BRAF, and MMR mutations in these patients. RESULTS: The overall frequencies of KRAS and BRAF mutations were 35.8% and 3.2%, respectively. Sixty-nine patients (7.6%) lacking expression of ≥1 MMR protein were considered MMR protein deficient (MMR-D); the remaining patients were considered MMR protein intact. KRAS mutations constituted an independent risk factor for shorter overall survival (OS) in TNM stage I-IV and stage III patients. BRAF mutations were associated with shorter OS in TNM stage I-IV patients. MMR-D status was strongly positive prognostic in TNM stage I-II patients. DISCUSSION/CONCLUSION: To our knowledge, this is the first multicenter study to explore the prognostic utilities of KRAS, BRAF, and MMR statuses in Korean CRC patients. Various combinations of KRAS, BRAF, and DNA MMR mutations serve as genetic signatures that affect tumor behavior; they are prognostic in CRC patients.

Visual prognosis and surgical timing of Ahmed glaucoma valve implantation for neovascular glaucoma secondary to diabetic vitrectomy.

BACKGROUND: Evaluate the visual outcomes of Ahmed glaucoma valve implantation (AGVI) in patients with neovascular glaucoma (NVG) who underwent diabetic vitrectomy and suggest appropriate AGVI timing. METHODS: Medical records of patients who underwent AGVI due to NVG after diabetic vitrectomy were reviewed. Successful intraocular pressure (IOP) control was defined as an IOP between 6 and 21 mmHg. Visual outcome was compared before NVG diagnosis and after AGVI, and the "favorable" visual outcome was defined as a postoperative deterioration in BCVA of less than 0.3 logMAR units compared to those before the development of NVG. Various factors including surgical timing were evaluated to identify the risk factors associated with unfavorable visual outcome. RESULTS: A total of 35 eyes were enrolled and divided into group 1(medically uncontrolled NVG group, IOP more than 30mmHg, 16 eyes) and group 2(NVG group responded well to the initial non-surgical treatment but eventually required AGVI, 19 eyes). Despite the favorable rate of normalization of post-AGVI IOP (85.7%), 43.8% in Group 1 and 26.3% in Group 2 showed unfavorable visual outcomes. In group 1, delayed surgical timing more than 1 week from the NVG diagnosis showed a significant association with unfavorable visual outcomes (P = 0.041). In group 2, poor patient compliance (follow up loss, refuse surgery) was the main factor of unfavorable visual outcomes. CONCLUSION: When NVG occurs in patients with proliferative diabetic retinopathy after vitrectomy, physicians should be cautious not to delay the surgical intervention, especially in patients with IOP of 30 or more despite non-surgical treatment. Early AGVI within six days might be necessary to preserve useful vision in these patients.

Structural Congeners of Izenamides Responsible for Cathepsin D Inhibition: Insights from Synthesis-Derived Elucidation.

This study aimed to elucidate the structural congeners of natural izenamides A, B, and C (1-3) responsible for cathepsin D (CTSD) inhibition. Structurally modified izenamides were synthesized and biologically evaluated, and their biologically important core structures were identified. We confirmed that the natural statine (Sta) unit (3S,4S)-γ-amino-ß-hydroxy acid is a requisite core structure of izenamides for inhibition of CTSD, which is closely related to the pathophysiological roles in numerous human diseases. Interestingly, the statine-incorporated izenamide C variant (7) and 18-epi-izenamide B variant (8) exhibited more potent CTSD-inhibitory activities than natural izenamides.

Ferroelectric Modulation of Surface Electronic States in BaTiO3 for Enhanced Hydrogen Evolution Activity.

Ferroelectric nanomaterials offer the promise of switchable electronic properties at the surface, with implications for photo- and electrocatalysis. Studies to date on the effect of ferroelectric surfaces in electrocatalysis have been primarily limited to nanoparticle systems where complex interfaces arise. Here, we use MBE-grown epitaxial BaTiO3 thin films with atomically sharp interfaces as model surfaces to demonstrate the effect of ferroelectric polarization on the electronic structure, intermediate binding energy, and electrochemical activity toward the hydrogen evolution reaction (HER). Surface spectroscopy and ab initio DFT+U calculations of the well-defined (001) surfaces indicate that an upward polarized surface reduces the work function relative to downward polarization and leads to a smaller HER barrier, in agreement with the higher activity observed experimentally. Employing ferroelectric polarization to create multiple adsorbate interactions over a single electrocatalytic surface, as demonstrated in this work, may offer new opportunities for nanoscale catalysis design beyond traditional descriptors.

Single Nucleotide Polymorphisms May Increase the Risk of Aspiration Pneumonia in Post-Stroke Patients with Dysphagia.

This study aimed to evaluate whether genetic polymorphism is associated with an increased risk of infection, specifically post-stroke aspiration pneumonia. Blood samples were obtained from a total of 206 post-stroke participants (males, n = 136; mean age, 63.8 years). Genotyping was performed for catechol-O-methyltransferase (rs4680, rs165599), dopamine receptors (DRD1; rs4532, DRD2; rs1800497, DRD3; rs6280), brain-derived neurotrophic factor (rs6265), apolipoprotein E (rs429358, rs7412), and the interleukin-1 receptor antagonist gene (rs4251961). The subjects were stratified into two groups, aged < 65 (young) and ≥ 65 (elderly). Functional parameters and swallowing outcomes were measured at enrollment and at 3 months post-onset. The primary outcome was the incidence of aspiration pneumonia. Analysis of the association between genetic polymorphisms and aspiration pneumonia history showed that a minor C rs429358 allele was associated with the occurrence of aspiration pneumonia in the young group, both in the additive and the dominant models (odds ratio: 4.53; 95% CI: 1.60−12.84, p = 0.004). In the multivariable analysis, the minor C rs429358 allele increased the risk of post-stroke aspiration pneumonia in young stroke patients by 5.35 (95% CI: 1.64−20.88). In contrast, no such association was observed in the elderly group. Apolipoprotein E polymorphism may affect the risk of post-stroke aspiration pneumonia.

Discovery of new ERRγ agonists regulating dopaminergic neuronal phenotype in SH-SY5Y cells.

The discovery of small molecules that regulate specific neuronal phenotypes is important for the development of new therapeutic candidates for neurological diseases. Estrogen-related receptor γ (ERRγ), an orphan nuclear receptor widely expressed in the central nervous system (CNS), is closely related to the regulation of neuronal metabolism and differentiation. We previously reported that upregulation of ERRγ could enhance dopaminergic neuronal phenotypes in the neuroblastoma cell line, SH-SY5Y. In this study, we designed and synthesized a series of new ERRγ agonists using the X-ray crystal structure of the GSK4716-bound ERRγ complex and known synthetic ligands. Our new ERRγ agonists exhibited increased transcriptional activities of ERRγ. In addition, our molecular docking results supported the experimental findings for ERRγ agonistic activity of the potent analogue, 5d. Importantly, 5d not only enhanced the expression of dopaminergic neuronal-specific molecules, TH and DAT but also activated the relevant signaling events, such as the CREB-mediated signaling pathway. The results of the present study may provide useful clues for the development of novel ERRγ agonists for neurological diseases related to the dopaminergic nervous system.

Nanocrack-regulated self-humidifying membranes.

The regulation of water content in polymeric membranes is important in a number of applications, such as reverse electrodialysis and proton-exchange fuel-cell membranes. External thermal and water management systems add both mass and size to systems, and so intrinsic mechanisms of retaining water and maintaining ionic transport in such membranes are particularly important for applications where small system size is important. For example, in proton-exchange membrane fuel cells, where water retention in the membrane is crucial for efficient transport of hydrated ions, by operating the cells at higher temperatures without external humidification, the membrane is self-humidified with water generated by electrochemical reactions. Here we report an alternative solution that does not rely on external regulation of water supply or high temperatures. Water content in hydrocarbon polymer membranes is regulated through nanometre-scale cracks ('nanocracks') in a hydrophobic surface coating. These cracks work as nanoscale valves to retard water desorption and to maintain ion conductivity in the membrane on dehumidification. Hydrocarbon fuel-cell membranes with surface nanocrack coatings operated at intermediate temperatures show improved electrochemical performance, and coated reverse-electrodialysis membranes show enhanced ionic selectivity with low bulk resistance.

Synthesis of 7,2'-Dihydroxy-4',5'-Dimethoxyisoflavanone, a Phytoestrogen with Derma Papilla Cell Proliferative Activity.

This paper reports a concise and scalable method for the synthesis of the phytoestrogen 7,2'-dihydroxy-4',5'-dimethoxyisoflavanone 1 via an optimized synthetic route. Compound 1 was readily obtained in 11 steps and 11% overall yield on a gram scale from commercially available 3,4-dimethoxyphenol. The key features of the synthesis include the construction of the deoxybenzoin unit through a sequence of Claisen rearrangement, oxidative cleavage, and aryllithium addition and the efficient synthesis of the isoflavanone architecture from highly functionalized 2-hydroxyketone.

Application of deep learning artificial intelligence technique to the classification of clinical orthodontic photos.

BACKGROUND: Taking facial and intraoral clinical photos is one of the essential parts of orthodontic diagnosis and treatment planning. Among the diagnostic procedures, classification of the shuffled clinical photos with their orientations will be the initial step while it was not easy for a machine to classify photos with a variety of facial and dental situations. This article presents a convolutional neural networks (CNNs) deep learning technique to classify orthodontic clinical photos according to their orientations. METHODS: To build an automated classification system, CNNs models of facial and intraoral categories were constructed, and the clinical photos that are routinely taken for orthodontic diagnosis were used to train the models with data augmentation. Prediction procedures were evaluated with separate photos whose purpose was only for prediction. RESULTS: Overall, a 98.0% valid prediction rate resulted for both facial and intraoral photo classification. The highest prediction rate was 100% for facial lateral profile, intraoral upper, and lower photos. CONCLUSION: An artificial intelligence system that utilizes deep learning with proper training models can successfully classify orthodontic facial and intraoral photos automatically. This technique can be used for the first step of a fully automated orthodontic diagnostic system in the future.

OST1 Activation by the Brassinosteroid-Regulated Kinase CDG1-LIKE1 in Stomatal Closure.

Crosstalk between signaling pathways is an important feature of complex regulatory networks. How signal crosstalk circuits are tailored to suit different needs of various cell types remains a mystery in biology. Brassinosteroid (BR) and abscisic acid (ABA) antagonistically regulate many aspects of plant growth and development through direct interactions between components of the two signaling pathways. Here, we show that BR and ABA synergistically regulate stomatal closure through crosstalk between the BR-activated kinase CDG1-LIKE1 (CDL1) and the OPEN STOMATA1 (OST1) of the ABA signaling pathway in Arabidopsis thaliana We demonstrate that the cdl1 mutant displayed reduced sensitivity to ABA in a stomatal closure assay, similar to the ost1 mutant. CDL1 and the BR receptor BR-INSENSITIVE1, but not other downstream components of the BR signaling pathway, were required for BR regulation of stomatal movement. Genetic and biochemical experiments demonstrated that CDL1 activates OST1 by phosphorylating it on residue Ser-7. BR increased phosphorylation of OST1, and the BR-induced OST1 activation was abolished in cdl1 mutants. Moreover, we found that ABA activates CDL1 in an OST1-dependent manner. Taken together, our findings illustrate a cell-type-specific BR signaling branch through which BR acts synergistically with ABA in regulating stomatal closure.

Concise syntheses and anti-inflammatory effects of isocorniculatolide B and corniculatolide B and C.

The first total syntheses of isocorniculatolide B, corniculatolide B, and corniculatolide C, consisting of isomeric corniculatolide skeletons, have been accomplished in a divergent manner. The key features of the synthesis involve the construction of diaryl ether linkages by nucleophilic aromatic substitution, installation of a C14-substituted alkyl side chain via a sequence of Baeyer-Villiger reaction and Claisen rearrangement, and efficient construction of corniculatolide and isocorniculatolide frameworks, including 17-membered (exterior) macrolactone skeletons from a versatile diaryl ether intermediate by Mitsunobu macrolactonization. Moreover, we prepared the structural congeners of isomeric corniculatolides via diverted total synthesis approach including desmethyl analogues and related dimeric macrolides. The anti-inflammatory activities of the synthesized natural products, analogues and synthetic intermediates were also investigated. In particular, corniculatolide B significantly inhibited the protein expression of COX-2 and the mRNA expressions of TNF-α, IL-1ß and IL-6 by inhibiting of NF-κB signaling in intestinal epithelial cells induced by lipopolysaccharide treatment. It also significantly inhibited the promoter activity and the phosphorylation of subunits p50 and p65 of NF-κB to the same extent as Bay 11-7082, a potent IκB kinase inhibitor. These results suggest that corniculatolide B might have therapeutic potential in inflammatory bowel disease via NF-κB signaling pathway.

Prediction of hand-wrist maturation stages based on cervical vertebrae images using artificial intelligence.

OBJECTIVE: To predict the hand-wrist maturation stages based on the cervical vertebrae (CV) images, and to analyse the accuracy of the proposed algorithms. SETTINGS AND POPULATION: A total of 499 pairs of hand-wrist radiographs and lateral cephalograms of 455 orthodontic patients aged 6-18 years were used for developing the prediction model for hand-wrist skeletal maturation stages. MATERIALS AND METHODS: The hand-wrist radiographs and the lateral cephalograms were collected from two university hospitals and a paediatric dental clinic. After identifying the 13 anatomic landmarks of the CV, the width-height ratio, width-perpendicular height ratio and concavity ratio of the CV were used as the morphometric features of the CV. Patients' chronological age and sex were also included as input data. The ground truth data were the Fishman SMI based on the hand-wrist radiographs. Three specialists determined the ground truth SMI. An ensemble machine learning methods were used to predict the Fishman SMI. Five-fold cross-validation was performed. The mean absolute error (MAE), round MAE and root mean square error (RMSE) values were used to assess the performance of the final ensemble model. RESULTS: The final ensemble model consisted of eight machine learning models. The MAE, round MAE and RMSE were 0.90, 0.87 and 1.20, respectively. CONCLUSION: Prediction of hand-wrist SMI based on CV images is possible using machine learning methods. Chronological age and sex increased the prediction accuracy. An automated diagnosis of the skeletal maturation may aid as a decision-supporting tool for evaluating the optimal treatment timing for growing patients.

JI017, a Complex Herbal Medication, Induces Apoptosis via the Nox4-PERK-CHOP Axis in Ovarian Cancer Cells.

Many anti-cancer drugs, including paclitaxel and etoposide, have originated and been developed from natural products, and traditional herbal medicines have fewer adverse effects and lesser toxicity than anti-tumor reagents. Therefore, we developed a novel complex herbal medicine, JI017, which mediates endoplasmic reticulum (ER) stress and apoptosis through the Nox4-PERK-CHOP signaling pathway in ovarian cancer cells. JI017 treatment increases the expression of GRP78, ATF4, and CHOP and the phosphorylation of PERK and eIF2α via the upregulation of Nox4. Furthermore, it increases the release of intracellular reactive oxygen species (ROS), the production of intracellular Ca2+, and the activation of exosomal GRP78 and cell lysate GRP78. Combination treatment using the sarco/endoplasmic reticulum Ca2+-ATPase inhibitor thapsigargin (TG) and JI017 reportedly induces increased ER stress and cell death in comparison to the control; however, knockdown experiments of PERK and CHOP indicated suppressed apoptosis and ER stress in JI017-treated ovarian cancer cells. Furthermore, targeting Nox4 using specific siRNA and pharmacological ROS inhibitors, including N-acetylcystein and diphenylene iodonium, blocked apoptosis and ER stress in JI017-treated ovarian cancer cells. In the radioresistant ovarian cancer model, when compared to JI017 alone, JI017 co-treatment with radiation induced greater cell death and resulted in overcoming radioresistance by inhibiting epithelial-mesenchymal-transition-related phenomena such as the reduction of E-cadherin and the increase of N-cadherin, vimentin, Slug, and Snail. These findings suggest that JI017 is a powerful anti-cancer drug for ovarian cancer treatment and that its combination treatment with radiation may be a novel therapeutic strategy for radioresistant ovarian cancer.