New Insights on the Role of ß-Cyanoalanine Synthase CAS-C1 in Root Hair Elongation through Single-Cell Proteomics.

(1) Background: Root hairs are specialized structures involved in water and plant nutrient uptake. They elongate from epidermal cells following a complex developmental program. ß-cyanoalanine synthase (CAS), which is mainly involved in hydrogen cyanide (HCN) detoxification in Arabidopsis thaliana, plays a role in root hair elongation, as evidenced by the fact that cas-c1 mutants show a severe defect in root hair shape. In addition to root hairs, CAS C1 is expressed in the quiescent center and meristem. (2) Methods: To identify its role in root hair formation, we conducted single-cell proteomics analysis by isolating root hair cells using Fluorescence-activated Cell Sorting (FACS) from wild-type and cas-c1 mutants. We also analyzed the presence of S-cyanylation, a protein post-translational modification (PTM) mediated by HCN and affecting cysteine residues and protein activity in proteins of wild type and cas-c1 mutants. (3) Results and Conclusions: We have found that the cas-c1 mutation has no visible effect on quiescent center or meristem root tissue, in both control and nutrient-deprivation conditions. We have identified more than 3900 proteins in root hairs and we have found that several proteins involved in root hair development, related to the receptor kinase FERONIA signaling and DNA methylation, are modified by S-cyanylation.

Luminescence imaging of water irradiated by protons under FLASH radiation therapy conditions.

Objective.FLASH radiation therapy with ultrahigh dose rates (UHDR) has the potential to reduce damage to normal tissue while maintaining anti-tumor efficacy. However, rapid and precise dose distribution measurements remain difficult for FLASH radiation therapy with proton beams. To solve this problem, we performed luminescence imaging of water following irradiation by a UHDR proton beam captured using a charge-coupled device camera.Approach. We used 60 MeV proton beams with dose rates of 0.03-837 Gy s-1from a cyclotron. Therapeutic 139.3 MeV proton beams with dose rates of 0.45-4320 Gy s-1delivered by a synchrotron-based proton therapy system were also tested. The luminescent light intensity induced by the UHDR beams was compared with that produced by conventional beams to compare the dose rate dependency of the light intensity and its profile.Main results. Luminescence images of water were clearly visualized under UHDR conditions, with significantly shorter exposure times than those with conventional beams. The light intensity was linearly proportional to the delivered dose, which is similar to that of conventional beams. No significant dose-rate dependency was observed for 0.03-837 Gy s-1. The light-intensity profiles of the UHDR beams agreed with those of conventional beams. The results did not differ between accelerators (synchrotron or cyclotron) and beam energies.Significance. Luminescence imaging of water is achievable with UHDR proton beams as well as with conventional beams. The proposed method should be suitable for rapid and easy quality assurance investigations for proton FLASH therapy, because it facilitates real-time, filmless measurements of dose distributions, and is useful for rapid feedback.

Root Canal Treatment of Oehlers Type III Dens Invaginatus in Maxillary Lateral Incisor and Remote Sinus Tract Using Dental Surgical Microscope and Cone-Beam Computed Tomography.

Dens invaginatus is a morphological abnormality of the tooth that results from a developmental anomaly during tooth formation, in which part of the enamel and dentin of the crown invaginates into the pulp cavity. This report describes a case of a maxillary lateral incisor with apical periodontitis apparently caused by Oehlers Type III dens invaginatus. The patient was a 69-year-old man who visited our clinic complaining of discomfort in the maxillary right lateral incisor. Cone-beam computed tomography (CBCT) revealed dens invaginatus of the maxillary lateral incisor and a sinus tract in the maxillary central incisor region, which was derived from apical periodontitis of the maxillary lateral incisor. The dens invaginatus was accompanied by a complex root canal morphology. Treatment, which was performed using a dental surgical microscope, had a favorable outcome. The patient remains in good condition at 1 year postoperatively.

NAC103 mutation alleviates DNA damage in an Arabidopsis thaliana mutant sensitive to excess boron.

Excess boron (B) is toxic to plants and thereby causes DNA damage and cell death in root meristems. However, the underlying mechanisms which link boron and DNA damage remain unclear. It has been reported that the rpt5a-6 mutant of the 26S proteasome is sensitive to excess boron, resulting in more frequent cell death in root meristem and reduced root elongation. In this study, we showed that a reduction in root growth in the rpt5a mutant in the presence of high boron levels is repressed by a mutation in NAC domain containing transcription factor NAC103, a substrate of the proteasome, which functions in the unfolded protein response pathway. The mutation in NAC103 alleviated excess-B-induced DNA damage and cell death in root meristems of the rpt5a mutant. Superoxide ( O 2 - ) staining with nitroblue tetrazolium revealed that boron stress causes O 2 - accumulation in root tips, which was higher in the rpt5a-6 mutant, whereas the accumulation was lower in the rpt5a-6 nac103-3 double mutant. Our work demonstrates the overall involvement of NAC103 in maintaining healthy root meristem under excess boron conditions in the absence of RPT5A proteasome subunit.

PECT1, a rate-limiting enzyme in phosphatidylethanolamine biosynthesis, is involved in the regulation of stomatal movement in Arabidopsis.

An Arabidopsis mutant displaying impaired stomatal responses to CO2 , cdi4, was isolated by a leaf thermal imaging screening. The mutated gene PECT1 encodes CTP:phosphorylethanolamine cytidylyltransferase. The cdi4 exhibited a decrease in phosphatidylethanolamine levels and a defect in light-induced stomatal opening as well as low-CO2 -induced stomatal opening. We created RNAi lines in which PECT1 was specifically repressed in guard cells. These lines are impaired in their stomatal responses to low-CO2 concentrations or light. Fungal toxin fusicoccin (FC) promotes stomatal opening by activating plasma membrane H+ -ATPases in guard cells via phosphorylation. Arabidopsis H+ -ATPase1 (AHA1) has been reported to be highly expressed in guard cells, and its activation by FC induces stomatal opening. The cdi4 and PECT1 RNAi lines displayed a reduced stomatal opening response to FC. However, similar to in the wild-type, cdi4 maintained normal levels of phosphorylation and activation of the stomatal H+ -ATPases after FC treatment. Furthermore, the cdi4 displayed normal localization of GFP-AHA1 fusion protein and normal levels of AHA1 transcripts. Based on these results, we discuss how PECT1 could regulate CO2 - and light-induced stomatal movements in guard cells in a manner that is independent and downstream of the activation of H+ -ATPases. [Correction added on 15 May 2023, after first online publication: The third sentence is revised in this version.].

Effectiveness and Cost-effectiveness of Online Brief Mindfulness-based Cognitive Therapy for the Improvement of Productivity in the Workplace: Study Protocol for a Randomized Controlled Trial.

BACKGROUND: Numerous studies have demonstrated the effectiveness of mindfulness-based programs (MBPs) among both clinical and nonclinical populations. These data document positive impacts in the workplace, including reducing perceived stress and burnout and increasing well-being. However, the effectiveness for productivity, which is of most interest to managers and administrators, is still unclear. In addition, MBPs in the workplace tend to be modified by reducing the number of the program sessions or delivering content online to improve accessibility. To date, however, the impact of MBPs that feature these modifications on productivity in the workplace has not been investigated. OBJECTIVE: The study aims to investigate the effectiveness and cost-effectiveness of online-delivered brief mindfulness-based cognitive therapy (bMBCT) for improving productivity and other work-related outcomes among healthy workers compared to the waitlist control. METHODS: We will conduct a 4-week randomized controlled trial (RCT) with a 6-month follow-up. Employees are included in the study if they (1) are between the ages of 20 and 65 years and (2) work longer than 30 hours weekly. Employees are randomly allocated to either the bMBCT group or the waitlist control group. The primary outcome of the study is the mean difference of productivity measured by the World Health Organization Health and Work Performance Questionnaire (WHO-HPQ) between the groups at 4, 16, and 28 weeks. Secondary outcomes include several clinical outcomes and health economics evaluation. RESULTS: We started recruiting participants in August 2021, and the intervention began in October 2021. A total of 104 participants have been enrolled in the study as of October 2021. The intervention is scheduled to be completed in December 2023. Data collection will be completed by the end of January 2024. CONCLUSIONS: The novelty of the study is that (1) it will investigate bMBCT's effectiveness on productivity, which is still unclear, and (2) samples are recruited from 3 companies in different industries. The limitations of the study are that (1) all measures assessed are in self-report format and (2) we lack an active control group. This study has the potential to provide new data on the relationship between MBPs and occupational health and productivity. TRIAL REGISTRATION: University Hospital Medical Information Network Clinical Trials Registry UMIN000044721; https://tinyurl.com/4e2fh873. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/36012.

Effectiveness of Mindfulness-Based Cognitive Therapy With Follow-Up Sessions for Pharmacotherapy-Refractory Anxiety Disorders: Protocol for a Feasibility Randomized Controlled Trial.

BACKGROUND: Augmented mindfulness-based cognitive therapy (MBCT) with treatment as usual (mainly pharmacotherapy) is reported to be effective after treatment for anxiety disorders. However, whether its effectiveness persists in the long term is unclear. OBJECTIVE: This study aims to examine the feasibility, acceptability, and effectiveness of a follow-up program by conducting a feasibility randomized controlled trial (RCT) that compares augmented MBCT with follow-up sessions and that without follow-up sessions in preparation for a definitive RCT. METHODS: The study involves an 8-week MBCT with a 10-month follow-up. Patients aged 20 to 65 years who meet the Diagnostic and Statistical Manual of Mental Disorders, 4th edition (DSM-IV) criteria for panic disorder, agoraphobia, or social anxiety disorder, which is not remitted with usual treatment for at least 4 weeks, will be included in the study and randomly allocated to receive augmented MBCT with follow-up sessions or augmented MBCT without follow-up sessions. For this feasibility RCT, the primary outcomes are (1) study inclusion rate, (2) dropout rate, (3) attendance rate, and (4) mean and standard deviation of several clinical measures at 8 weeks and 5, 8, and 12 months. RESULTS: We started recruiting participants in January 2020, and 43 participants have been enrolled up to January 2021. The study is ongoing, and data collection will be completed by May 2022. CONCLUSIONS: This study is novel in terms of its design, which compares augmented MBCT with and without follow-up sessions. The limitations of the trial are as follows: (1) mixed participants in terms of the delivery mode of the intervention, and (2) lack of a pharmacotherapy-alone arm. Owing to its novelty and significance, this study will provide fruitful knowledge for future definitive RCTs. TRIAL REGISTRATION: UMIN Clinical Trials Registry UMIN000038626; https://tinyurl.com/2p9dtxzh. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/33776.

The Roles of Peptide Hormones and Their Receptors during Plant Root Development.

Peptide hormones play pivotal roles in many physiological processes through coordinating developmental and environmental cues among different cells. Peptide hormones are recognized by their receptors that convey signals to downstream targets and interact with multiple pathways to fine-tune plant growth. Extensive research has illustrated the mechanisms of peptides in shoots but functional studies of peptides in roots are scarce. Reactive oxygen species (ROS) are known to be involved in stress-related events. However, recent studies have shown that they are also associated with many processes that regulate plant development. Here, we focus on recent advances in understanding the relationships between peptide hormones and their receptors during root growth including outlines of how ROS are integrated with these networks.

Intentional Replantation to Treat Apical Periodontitis of Maxillary First Molar with Foreign Body Located Outside Apical Foramen Using CBCT: A Case Report.

The recent use of cone beam computed tomography (CBCT) in the field of dentistry to obtain 3-dimensional (3D) images has enabled more effective examination and diagnosis in endodontic treatment. Such information has also been reported to be of benefit in surgical endodontic treatment such as intentional replantation. Here, we report a case of intentional replantation with the assistance of CBCT in which a good therapeutic outcome was achieved. The patient was a 30-year-old woman who visited our hospital with the chief complaint of spontaneous pain in the right maxillary first molar. Dental radiographs revealed a radiolucent area in the apical portion of the distal root, and a radiopaque area thought to be a broken shard from a small surgical instrument measuring approximately 1.5 mm in length. The shard was located outside the distal apical foramen. Based on these findings, the diagnosis was acute suppurative apical periodontitis of the right maxillary first molar. Infected root canal therapy was subsequently commenced. The patient's symptoms showed no improvement, however, and the pain persisted. Therefore, dental CBCT was performed to obtain 3D images, which confirmed a radiopaque area thought to be a broken shard from a small surgical instrument located outside the apical foramen of the distal root and facing in a direction that made it impossible to remove from the root canal. Because the patient's symptoms had shown no improvement and a foreign body was observed outside the apical foramen, intentional replantation combined with root resection was performed with informed consent. At a 1-year follow-up visit, progress was good, and there were no reported symptoms or signs. When reaching a diagnosis is difficult based on clinical findings and dental radiographs alone, the 3D images provided by CBCT offer a means of securing a more reliable diagnosis, allowing planning of treatment to be more effective.

Comparison of Characteristics of Dental Malpractice Trials between Medical Malpractice and Ordinary Divisions in District Courts.

The purpose of this study was to elucidate the characteristics of dental malpractice trials undertaken by medical malpractice divisions and ordinary divisions in district courts. Dentistry disputes in a total of 84 trials held between 1977 and 2014 were investigated. A total of 45 were conducted by medical malpractice divisions, resulting in 18 approvals and 27 dismissals, while 39 were undertaken by ordinary divisions, leading to 24 approvals and 15 dismissals. The parameters analyzed comprised category of dental treatment, judgment, amount claimed, and amount accepted. The results revealed that the mean amount claimed in trials held by medical malpractice divisions (¥12,563,324) was lower than that sought in trials conducted by ordinary divisions. The amount accepted was also found to exceed 50% of the amount claimed in 6 trials held by ordinary divisions (maximum 75.2%), but in only 2 trials conducted by medical malpractice divisions (maximum 54.8%); the mean amount accepted in trials held by medical malpractice divisions was 24.2%. These results indicate that judgments in trials conducted by medical malpractice divisions place a stronger emphasis on resolution of dental malpractice dispute than on pursuing truth or carrying out proper legal procedures.

Longitudinal and cross-sectional detection of four bovine enteric viruses by multiplex- reverse transcription polymerase chain reaction: Identification of possible indicator viruses to assess biosecurity level at bovine farms.

It can be judged that if the detection frequency of prevalent pathogenic viruses decreases, biosecurity has been enhanced. To monitor bovine farm biosecurity levels, one-step multiplex reverse transcription polymerase chain reaction (RT-PCR) for the simultaneous detection of group A rotavirus (RVA), bovine torovirus (BToV), bovine enterovirus (BEV), and bovine coronavirus (BCV) was designed, with the aim of configuring candidates for "viral pathogen indicators". A total of 322 bovine fecal samples were collected from calves aged less than three months at 48 bovine farms in Ibaraki and Chiba prefectures. At farm A, 20 calves were selected and sampled weekly for 12 weeks (184 samples); at farm B, 10 calves were selected and sampled for five weeks (50 samples); and at the rest of the 46 farms, 88 calves were sampled once. The screening on the 358 field samples proved positive for 27 RVA, 4 BToV, 55 BEV, and 52 BCV. In the successive sampling, RVA was detected once but not continuously, whereas BEV and BCV were detected in succession for up to five weeks. The results revealed that RVA was the primary agent among the positive samples obtained from calves aged three weeks or less, while BEV was the primary among those from the older than three weeks old. They can be employed as useful viral pathogen indicators for soundly evaluating biosecurity at bovine farms.

RGF1 controls root meristem size through ROS signalling.

The stem cell niche and the size of the root meristem in plants are maintained by intercellular interactions and signalling networks involving a peptide hormone, root meristem growth factor 1 (RGF1)1. Understanding how RGF1 regulates the development of the root meristem is essential for understanding stem cell function. Although five receptors for RGF1 have been identified2-4, the downstream signalling mechanism remains unknown. Here we report a series of signalling events that follow RGF1 activity. We find that the RGF1-receptor pathway controls the distribution of reactive oxygen species (ROS) along the developmental zones of the Arabidopsis root. We identify a previously uncharacterized transcription factor, RGF1-INDUCIBLE TRANSCRIPTION FACTOR 1 (RITF1), that has a central role in mediating RGF1 signalling. Manipulating RITF1 expression leads to the redistribution of ROS along the root developmental zones. Changes in ROS distribution in turn enhance the stability of the PLETHORA2 protein, a master regulator of root stem cells. Our results thus clearly depict a signalling cascade that is initiated by RGF1, linking this peptide to mechanisms that regulate ROS.

Respective optimal calcium concentrations for proliferation on type I collagen fibrils in two keratinocyte line cells, HaCaT and FEPE1L-8.

Keratinocyte line cells HaCaT and FEPE1L-8 are used for skin model with type I collagen fibrils (gels). For this purpose, not only differentiation but also regulation of proliferation on type I collagen gels by exogenous calcium concentration is important. When exogenous calcium concentration is low, primary keratinocyte proliferation is repressed and eventually cells are induced to apoptosis on type I collagen gels. The apoptosis induced on type I collagen gels is suppressed by increasing calcium concentration in the medium. That is, higher exogenous calcium concentration is necessary for primary keratinocyte survival on type I collagen gels than for that on dish surface culture. Meanwhile much higher exogenous calcium causes cell differentiation and inhibition of proliferation. The optimal calcium concentrations for proliferation on type I collagen gels have not been clarified in keratinocyte line cells. HaCaT cells have a unique calcium sensitivity in comparison with primary keratinocytes, whereas FEPE1L-8 cells have a similar sensitivity to primary keratinocytes. In this study, we compared the effect of calcium concentrations on proliferation of HaCaT and FEPE1L-8 cells on type I collagen gels. On type I collagen gels, both line cells required higher calcium concentrations for proliferation than on dish surface. HaCaT cells proliferated better in a wider range of calcium concentrations than FEPE1L-8 cells.

Virucidal efficacy of food additive grade calcium hydroxide against surrogate of human norovirus.

An alkaline agent, namely, food additive grade calcium hydroxide (FdCa(OH)2) in the solution, powder and suspension forms was evaluated as a virucidal agent, using a murine norovirus (MNV) as the surrogate for human norovirus. The main constituent of FdCa(OH)2 is Ca(OH)2, which has pH 13 in 0.17% solution. The results showed that 0.17% FdCa(OH)2 solution could inactivate MNV within 30s even in the presence of organic materials (5% fetal bovine serum (FBS)). In a contaminated surface experiment, MNV with 5% FBS was inoculated on rayon sheets, and the result showed FdCa(OH)2 solution could markedly reduce virus titer within 1min. When mouse feces were spiked with MNV and FdCa(OH)2 powder as 10% and 20% w/w was added to the feces, these concentrations could inactivate the virus within 30min and 15min, respectively. Whereas, FdCa(OH)2 suspension at 2.5% and 5% could inactivate the virus within 30min and at 1% within 45min. These and additional results obtained here indicate that FdCa(OH)2 is an effective virucidal agent against MNV, and can serve as a useful alternative disinfectant for inactivation and prevention of human norovirus in house and hospital.

Enhancement of bactericidal effects of sodium hypochlorite in chiller water with food additive grade calcium hydroxide.

An alkaline agent, namely food additive grade calcium hydroxide (FdCa(OH)2) in solution at 0.17%, was evaluated for its bactericidal efficacies in chiller water with sodium hypochlorite (NaOCl) at a concentration of 200 ppm total residual chlorine. Without organic material presence, NaOCl could inactivate Salmonella Infantis and Escherichia coli within 5 sec, but in the presence of fetal bovine serum (FBS) at 0.5%, the bactericidal effects of NaOCl were diminished completely. FdCa(OH)2 solution required 3 min to inactivate bacteria with or without 5% FBS. When NaOCl and FdCa(OH)2 were mixed at the final concentration of 200 ppm and 0.17%, respectively, the mixed solution could inactivate bacteria at acceptable level (103 reduction of bacterial titer) within 30 sec in the presence of 0.5% FBS. The mixed solution also inhibited cross-contamination with S. Infantis or E. coli on chicken meats. It was confirmed and elucidated that FdCa(OH)2 has a synergistic effect together with NaOCl for inactivating microorganisms.

Durability of alkaline agents' bactericidal efficacies in litter under field conditions.

Alkaline agents are well-known for their disinfection capacities against pathogens even at the presence of organic materials, but the durability of their bactericidal efficacies under field conditions is unknown. Therefore, within the present study, two alkaline agents, namely bioceramic (BCX) derived from chicken feces and food additive grade Ca(OH)2 (FdCa(OH)2) derived from natural lime stone, were evaluated for the persistence of their bactericidal efficacies in litter, under simulated field conditions. BCX powder mixed at 50% concentration in litter or FdCa(OH)2 powder at 20% concentration in litter lost their bactericidal efficacies at 3 days post exposure of chicks, and thereafter, both mentioned alkaline agents could not inactivate bacteria down to the acceptable level (≥3 log10 CFU/ml reduction).

Functions of long intergenic non-coding (linc) RNAs in plants.

Whole transcriptome analyses in many organisms have revealed that most transcribed RNAs do not encode proteins. These non-coding RNAs likely contribute to the regulation of gene expression during the development of multicellular organisms. In eukaryotes, the roles of small RNAs, one class of non-coding RNAs, in transcriptional and post-transcriptional regulation have been well characterized. However, the functions of a second class of non-coding RNAs, long intergenic noncoding (linc) RNAs, are relatively unknown, especially in plants. Recent advances in RNA-seq and tiling microarray technologies have revealed the presence of many lincRNAs across plant species. This review focuses on the functions of lincRNAs that have been recently reported in plants. One of the most well characterized functions of lincRNAs is to epigenetically regulate gene expression by recruiting proteins for chromosome modification to specific loci. Second, lincRNAs are known to inhibit the physical interaction between microRNAs (miRNAs) and their target mRNAs thus controling protein levels of the target mRNAs. Lastly, lincRNAs control alternative splicing by binding and sequestering the proteins required for alternative splicing.

Accuracy of the evaluation method for alkaline agents' bactericidal efficacies in solid, and the required time of bacterial inactivation.

An alkaline agent, namely food additive grade calcium hydroxide (FdCa (OH)2) in the powder form, was evaluated for its bactericidal efficacies in chicken feces at pH 13. The point for this evaluation was neutralization of the alkaline agent's pH at the time of bacterial recovery, since otherwise the results are substantially misleading. Without neutralization of the FdCa (OH)2 pH, the spiked bacteria were killed within min at the time of recovery in aqueous phase, but not in the solid form in feces, hence, it has been demonstrated that when bacteria were in solid, it took longer time than in liquid for the alkaline agent to inactivate them down to the acceptable level (≥3 log10 CFU/ml).

High-Resolution Expression Map of the Arabidopsis Root Reveals Alternative Splicing and lincRNA Regulation.

The extent to which alternative splicing and long intergenic noncoding RNAs (lincRNAs) contribute to the specialized functions of cells within an organ is poorly understood. We generated a comprehensive dataset of gene expression from individual cell types of the Arabidopsis root. Comparisons across cell types revealed that alternative splicing tends to remove parts of coding regions from a longer, major isoform, providing evidence for a progressive mechanism of splicing. Cell-type-specific intron retention suggested a possible origin for this common form of alternative splicing. Coordinated alternative splicing across developmental stages pointed to a role in regulating differentiation. Consistent with this hypothesis, distinct isoforms of a transcription factor were shown to control developmental transitions. lincRNAs were generally lowly expressed at the level of individual cell types, but co-expression clusters provided clues as to their function. Our results highlight insights gained from analysis of expression at the level of individual cell types.

Virucidal Properties of Bioceramic Derived from Chicken Feces pH 13 and its Stability in Harsh Environments.

Bioceramic derived from chicken feces (BCX) is a material produced by a sintering process for the purpose of use in animal farms to control livestock infectious diseases. In the present study, BCX at pH 13 was evaluated for the durability of its virucidal activity in simulated field conditions. First it was shown that BCX had activity toward Newcastle disease virus, infectious bursal disease virus, and goose parvovirus within 3 min and toward avian influenza virus (AIV) within 1 hr. BCX was further tested by keeping it under simulated harsh environmental conditions with sunlight for several weeks as well as by repeatedly soaking it with water and drying under sunlight many times. After sampling every 2 consecutive weeks and every 2 (of 9) consecutive resuspensions, BCX was evaluated for its efficacy against AIV. Evaluation under the harsh conditions illustrated that BCX could retain its satisfactory efficacy toward AIV throughout 7 wk and through 9 resuspensions. It is hence concluded that BCX is an excellent material for applying in livestock farming as a trapping disinfectant, due to its efficacy to inactivate various viruses, and that this efficacy is prolonged even under harsh environmental conditions.