Highly Efficient Fluorescent Probe to Visualize Protein Interactions at the Superresolution.

Superresolution microscopy (SR microscopy) of protein-protein interactions (PPIs) occurring in subcellular structures is essential for understanding cellular functions. However, a powerful and useful technology for SR microscopy of PPIs remains elusive. Here, we develop a highly efficient photoconvertible fluorescent probe, named split-Dendra2, for SR microscopy of PPIs in the cell. We found that split-Dendra2 enables a highly efficient detection of PPIs, making it possible to perform SR microscopy of PPIs with high spatial resolution and high image reconstruction fidelity. We demonstrate the utility of split-Dendra2 by visualizing PPIs occurring in small subcellular structures at the superresolution, such as clathrin-coated pits and focal adhesions, which cannot be visualized by the existing tools. Split-Dendra2 offers a powerful and useful tool that greatly expands the possibility of SR microscopy and can contribute to revealing the function of PPIs at the nanoscale resolution.

Effects of consecutive bed bathing with weak versus ordinary pressure on skin barrier recovery of hospitalised older adults: A within-person randomised controlled trial.

AIM: Wiping pressure (WP [mmHg]) during bed baths is essential to maintain skin integrity and care quality for older adults. However, effects of different wiping pressures on skin barrier recovery over multiple days remain unclear. This study evaluated and compared the effects of consecutive bed bathing with weak pressure and that with ordinary pressure on skin barrier recovery of hospitalised older adults. METHODS: This within-person, randomised, controlled trial involved 254 forearms (127 patients) and was conducted at a general hospital. Forearms were blinded and randomly assigned a site and sequence of two bed bathing sessions: wiping three times with weak (10≤WP<20) and ordinary pressure (20≤WP<30) once per day for 2 consecutive days. The skin barrier was assessed daily based on transepidermal water loss (TEWL) and stratum corneum hydration (SCH) before and 15 min after the interventions. Dry skin was assessed using the overall dry skin score. RESULTS: A linear mixed model showed that the time courses of TEWL and SCH differed significantly between groups. Impaired skin barrier function caused by ordinary pressure on the first day did not recover to baseline values the next day, whereas weak pressure did not cause significant changes. During subgroup analyses, TEWL of patients with dry skin was more likely to increase with ordinary pressure. CONCLUSIONS: Despite decreased skin barrier recovery experienced by older adults, our findings suggest the safety of weak pressure and highlight the importance of WP during bed baths. Weak pressure is particularly desirable for patients with dry skin. TRIAL REGISTRATION: UMIN000048838.

Improved hourly estimate of aerosol optical thickness over Asian land by fusing geostationary satellites Fengyun-4B and Himawari-9.

Asian over-land aerosols are complexities due to a mixture of anthropogenic air pollutants and natural dust. The accuracy of the aerosol optical thickness (AOT) retrieved from the satellite is crucial to their application in the aerosol data assimilation system. Fusion of AOTs with high spatiotemporal resolution from next-generation geostationary satellites such as Fengyun-4B (FY-4B) and Himawari-9, provides a new high-quality dataset capturing the aerosol spatiotemporal variability for data assimilation. This study develops a complete fusion algorithm to estimate the optimal AOT over-land in Asia from September 2022 to August 2023 at 10 km × 10 km resolution with high efficiency. The data fusion involves four steps: (1) investigating the spatiotemporal variability of FY-4B AOT within the past 1 h and 12 km radius calculation domain; (2) utilizing the aerosol spatiotemporal variability characteristics to estimate FY-4B pure and hourly merged AOTs; (3) performing bias corrections for FY-4B and Himwari-9 hourly merged AOT for different observation times and seasons considering pixel-level errors for each satellite; (4) fusing the bias-corrected FY-4B and Himawari-9 hourly merged AOT based on maximum-likelihood estimation (MLE) method. Compared to the original FY-4B AOT, validation with AERONET observation confirms that the root mean square error (RMSE) of hourly merged FY-4B AOT decreases by around 40.6 % and the correlation coefficient (CORR) increases by about 27.8 %. Compared to FY-4B and Himawari-9 merged AOT, the fused AOT significantly decreases (increases) RMSE (CORR) by around 24.7 % (7.3 %) and 20.2 % (5.6 %). In addition, fused AOT is double the number of single-sensor merged AOT. Fusion aerosol map accurately describes the spatial and temporal variations in Asian regions controlled by air pollution and dust storms. Further studies are required for other landscapes with different satellite combinations to promote the application in the data assimilation system.

Evaluation and uncertainty analysis of Himawari-8 hourly aerosol product version 3.1 and its influence on surface solar radiation before and during the COVID-19 outbreak.

The hourly Himawari-8 version 3.1 (V31) aerosol product has been released and incorporates an updated Level 2 algorithm that uses forecast data as an a priori estimate. However, there has not been a thorough evaluation of V31 data across a full-disk scan, and V31 has yet to be applied in the analysis of its influence on surface solar radiation (SSR). This study firstly investigates the accuracy of V31 aerosol products, which includes three categories of aerosol optical depth (AOD) (AODMean, AODPure, and AODMerged) as well as the corresponding Ångström exponent (AE), using ground-based measurements from the AERONET and SKYNET. Results indicate that V31 AOD products are more consistent with ground-based measurements compared to previous products (V30). The highest correlation and lowest error were seen in the AODMerged, with a correlation coefficient of 0.8335 and minimal root mean square error of 0.1919. In contrast, the AEMerged shows a larger discrepancy with measurements unlike the AEMean and AEPure. Error analysis reveals that V31 AODMerged has generally stable accuracy across various ground types and geometrical observation angles, however, there are higher uncertainties in areas with high aerosol loading, particularly for fine aerosols. The temporal analysis shows that V31 AODMerged performs better compared to V30, particularly in the afternoon. Finally, the impacts of aerosols on SSR based on the V31 AODMerged are investigated through the development of a sophisticated SSR estimation algorithm in the clear sky. Results demonstrate that the estimated SSR is significant consistency with those of well-known CERES products, with preservation of 20 times higher spatial resolution. The spatial analysis reveals a significant reduction of AOD in the North China Plain before and during the COVID-19 outbreak, resulting in an average 24.57 W m-2 variation of the surface shortwave radiative forcing in clear sky daytime.

Development of a predictive model for vitamin D deficiency based on the vitamin D status in young Japanese women: A study protocol.

BACKGROUND: Vitamin D deficiency (VDD) is associated with an increased risk for lifestyle-related diseases. In Japan, VDD is quite prevalent in all age groups, with its high risk in young women. Furthermore, its association during pregnancy with gestational hypertension and low birth weight has also been reported. VDD can be diagnosed by serum 25-hydroxyvitamin D [25(OH)D] levels, which, however, is not suited for screening. Therefore, we will create a predictive model for serum 25(OH)D concentration and prevalence of VDD based on such data as region, sun exposure habit, and vitamin D intake in young women. METHODS: From 2020 to 2022, we conduct a cross-sectional study of 600 young women in four regions of Japan, identify the indices associated with serum 25(OH)D concentrations such as sun exposure habits, habitual vitamin D intake, ultraviolet-B irradiation, seasons (summer and winter) and latitude, and construct prediction models for serum 25(OH)D concentrations and VDD risk. This study has been registered with UMIN-CTR (ID: UMIN000041527). RESULTS: One hundred and fifteen subjects have been collected from 6 institutions in winter as of May 2021. When data from more than 200 subjects have become available, we will conduct the interim analysis, summarize the data by region and facility, review the inclusion criteria for analysis, and check for missing values and outliers. Prediction models for serum 25(OH)D concentration and VDD will be determined in the final analysis when all cases have been collected. CONCLUSIONS: A screening tool for VDD risk to be developed in our study based on the predictive model would help the public and medical professionals prevent lifestyle-related diseases through improving VDD. Additionally, the results may serve as the scientific basis for determining the appropriate vitamin D intake and sun exposure standards.

A Survey on Health Care and Health Concerning Workers for Considering Appropriate Personal Health Record Service.

To prevent worsening of diabetes mellitus, we conducted a developmental research of personal health record (PHR), whereby affected individuals, medical staff and an insurer cooperate and manage effective treatment. Medical expenses can be suppressed by preventing the onset of lifestyle-related diseases. Companies can benefit from improved employee productivity by promoting healthy and efficient working styles. We conducted a health and medical consciousness survey to examine PHR models that are attractive for both employers and employees.

Blood asymmetric dimethylarginine and nitrite/nitrate concentrations in short-stature children born small for gestational age with and without growth hormone therapy.

Objective To investigate the basal amino acid metabolism and impact of growth hormone (GH) therapy in short-stature children born small for gestational age (short SGA children). Methods In this age-matched case-control study, the basal blood levels of amino acids, asymmetric dimethylarginine (ADMA), and nitrite/nitrate (NOx) were compared between 24 short SGA children and 25 age-matched normal children. Changes in these parameters were assessed for 12 months in 12 short SGA children initiating GH therapy (Group A) and 12 age-matched short SGA children without GH therapy (Group B). Results The arginine levels were significantly lower in the short SGA than in normal children. The ADMA levels were significantly higher and NOx levels were significantly lower in the short SGA than normal children. In Group A, the ADMA level was significantly lower and NOx level was significantly higher at 6 months than at baseline. At 12 months, the ADMA level in Group A began to increase, but the NOx level remained the same. Group B showed no significant changes. Conclusions This study is the first to show that ADMA is promoted and nitric oxide is suppressed in short SGA children and that GH therapy affects the production of ADMA and nitric oxide.

RBM24 promotes U1 snRNP recognition of the mutated 5' splice site in the IKBKAP gene of familial dysautonomia.

The 5' splice site mutation (IVS20+6T>C) of the inhibitor of κ light polypeptide gene enhancer in B cells, kinase complex-associated protein (IKBKAP) gene in familial dysautonomia (FD) is at the sixth intronic nucleotide of the 5' splice site. It is known to weaken U1 snRNP recognition and result in an aberrantly spliced mRNA product in neuronal tissue, but normally spliced mRNA in other tissues. Aberrantly spliced IKBKAP mRNA abrogates IKK complex-associated protein (IKAP)/elongator protein 1 (ELP1) expression and results in a defect of neuronal cell development in FD. To elucidate the tissue-dependent regulatory mechanism, we screened an expression library of major RNA-binding proteins (RBPs) with our mammalian dual-color splicing reporter system and identified RBM24 as a regulator. RBM24 functioned as a cryptic intronic splicing enhancer binding to an element (IVS20+13-29) downstream from the intronic 5' splice site mutation in the IKBKAP gene and promoted U1 snRNP recognition only to the mutated 5' splice site (and not the wild-type 5' splice site). Our results show that tissue-specific expression of RBM24 can explain the neuron-specific aberrant splicing of IKBKAP exon 20 in familial dysautonomia, and that ectopic expression of RBM24 in neuronal tissue could be a novel therapeutic target of the disease.

Behavior of cyphenothrin in aquatic environment.

The behavior of cyphenothrin (1) [(RS)-α-cyano-3-phenoxybenzyl (1RS)-cis-trans-2,2-dimethyl-3-(2-methylprop-1-enyl)cyclopropanecarboxylate] in an aquatic environment was investigated by using the 14C-labeled trans and cis isomers. In parallel with the rapid partition from water phase to bottom sediment, 1 was degraded with the first-order half-lives of 2.0 (trans-1) and 7.3 days (cis-1) in the water-sediment system under dark conditions. 1 underwent extensive microbial degradation via ester cleavage to form 3-phenoxybenzoic acid, finally forming bound residues and mineralizing to CO2. Aqueous photolysis significantly accelerated the degradation of 1 with a half-life of <1 day, mainly via photo-induced oxidation at the 2-methylprop-1-enyl group and ester cleavage without cis-trans isomerization. These results strongly suggest that 1 is unlikely to persist in the actual aquatic environment due to its rapid photolysis and extensive microbial degradation.

In vitro screening for inhibitor of cloned Drosophila melanogaster tyramine-ß-hydroxylase and docking studies.

Biogenic amines are common biologically active substances extended within the whole animal kingdom where they play vital roles as signal transducer as well as regulator of cell functions. One of these biogenic amines called octopamine (OA) is synthesized from tyramine (TA) by the catalysis of tyramine-ß-hydroxylase (TßH) originated in the insect nervous system. Both TA and OA act as neurotransmitters, neurohormones and neuromodulators in the arthropod nervous system. Herein, the inhibitory activity of 1-arylimidazole-2(3H)-thiones (AITs) was tested on cloned Drosophila tyramine-ß-hydroxylase (DmTßH) expressed in Bombyx mori strain. Radiolabelled 3H-TA was used to analyze the activity of AITs exhibited inhibitory effects on DmTßH, whose ID50 values range from 0.02 to 2511nM where DmTßH was inhibited in a dose-dependent manner at pH 7.6 and 25°C during a 30min of incubation. To understand the catalytic role of the TßH, a three dimensional structure of the TßH from Drosophila melanogaster was constructed by homology modeling using the Phyre2 web server with 100% confidence. The modeled three-dimensional structure of TßH was used to perform the docking study with AITs. This may give more insights to precise design of inhibitors for TßH to control insect's population.

Rectifier of aberrant mRNA splicing recovers tRNA modification in familial dysautonomia.

Familial dysautonomia (FD), a hereditary sensory and autonomic neuropathy, is caused by missplicing of exon 20, resulting from an intronic mutation in the inhibitor of kappa light polypeptide gene enhancer in B cells, kinase complex-associated protein (IKBKAP) gene encoding IKK complex-associated protein (IKAP)/elongator protein 1 (ELP1). A newly established splicing reporter assay allowed us to visualize pathogenic splicing in cells and to screen small chemicals for the ability to correct the aberrant splicing of IKBKAP. Using this splicing reporter, we screened our chemical libraries and identified a compound, rectifier of aberrant splicing (RECTAS), that rectifies the aberrant IKBKAP splicing in cells from patients with FD. Here, we found that the levels of modified uridine at the wobble position in cytoplasmic tRNAs are reduced in cells from patients with FD and that treatment with RECTAS increases the expression of IKAP and recovers the tRNA modifications. These findings suggest that the missplicing of IKBKAP results in reduced tRNA modifications in patients with FD and that RECTAS is a promising therapeutic drug candidate for FD.

An extract of the medicinal plant Artemisia annua modulates production of inflammatory markers in activated neutrophils.

PURPOSE: To investigate the ability of a commercial extract from the medicinal plant Artemisia annua to modulate production of the cytokine, tumor necrosis factor-alpha (TNF-α), and the cyclooxygenase (COX) inflammatory marker, prostaglandin E2 (PGE2) in activated neutrophils. METHODS: Neutrophils were harvested from rat whole blood and cultured in the presence of plant extract or control samples. Neutrophils, except unactivated control cells, were activated with 10 µg/mL lipopolysaccharide (LPS). The cells were cultured with a range of different concentrations of the A. annua extracts (400-1 µg/mL) and artemisinin (200 and 100 µg/mL) and the supernatants were then tested by enzyme-linked immunosorbent assay (ELISA) for the concentrations of TNF-α and PGE2. Each sample was assayed in triplicate. Positive controls with an inhibitor were assayed in triplicate: chloroquine 2.58 and 5.16 µg/mL for TNF-α, and ibuprofen 400 µg/mL for PGE2. An unsupplemented group was also assessed in triplicate as a baseline control. RESULTS: Neutrophils were stimulated to an inflammatory state by the addition of LPS. A. annua extract significantly inhibited TNF-α production by activated neutrophils in a dose-dependent manner. There was complete inhibition by the A. annua extract at 200, 100, and 50 µg/mL (all P≤0.0003). At A. annua extract concentrations of 25, 10, and 5 µg/mL, TNF-α production was inhibited by 89% (P<0.0001), 54% (P=0.0002), and 38% (P=0.0014), respectively. A. annua 1 µg/mL did not significantly inhibit TNF-α production (8.8%; P>0.05). Concentrations of 400, 200, and 100 µg/mL A. annua extract significantly inhibited PGE2 production by 87% (P=0.0128), 91% (P=0.0017), and 93% (P=0.0114), respectively. CONCLUSION: An extract of A. annua was shown to be a potent inhibitor of TNF-α and a strong inhibitor of PGE2 production in activated neutrophils at the concentrations tested. Further studies are warranted with this promising plant extract.

Successful treatment of skin and soft tissue infection due to carbapenem-resistant Acinetobacter baumannii by ampicillin-sulbactam and meropenem combination therapy.

In recent years, carbapenem-resistant Acinetobacter baumannii infections have been responsible for outbreaks in medical facilities. A 35-year-old Japanese woman developed a skin and soft tissue infection due to carbapenem-resistant A. baumannii. The isolate was resistant to antibiotics other than ampicillin-sulbactam and colistin, suggesting drug resistance due to carbapenemase production by OXA-23. We selected a combination therapy consisting of intravenous ampicillin-sulbactam and meropenem. No changes were observed in aspartate aminotransferase, alanine aminotransferase, blood urea nitrogen, or serum creatinine during therapy, and carbapenem-resistant A. baumannii was not detected in wound exudates 3 days after therapy initiation. In our patient's case, combination therapy with ampicillin-sulbactam and meropenem was successful. Thus, combination therapy with ampicillin-sulbactam and meropenem is effective against skin and soft tissue infection due to carbapenem-resistant A. baumannii. Combination therapy with intravenous ampicillin-sulbactam and meropenem may be an option for skin and soft tissue infections due to carbapenem-resistant A. baumannii.

Concise synthesis of the anti-HIV nucleoside EFdA.

EFdA (4'-ethynyl-2-fluoro-2'-deoxyadenosine), a nucleoside reverse transcriptase inhibitor with extremely potent anti-HIV activity, was concisely synthesized from (R)-glyceraldehyde acetonide in an 18% overall yield by a 12-step sequence involving highly diastereoselective ethynylation of an α-alkoxy ketone intermediate. The present synthesis is superior, both in overall yield and in the number of steps, to the previous one which required 18 steps from an expensive starting material and resulted in a modest overall yield of 2.5%.

Firefly luciferase genes from the subfamilies Psilocladinae and Ototretinae (Lampyridae, Coleoptera).

Firefly luciferase genes have been isolated from approximately 20 species of Lampyrinae, Luciolinae, and Photurinae. These are mostly nocturnal luminescent species that use light signals for sexual communication. In this study, we isolated three cDNAs for firefly luciferase from Psilocladinae (Cyphonocerus ruficollis) and Ototretinae (Drilaster axillaris and Stenocladius azumai), which are diurnal non-luminescent or weakly luminescent species that may use pheromones for communication. The amino acid sequences deduced from the three cDNAs showed 81-89% identities to each other and 60-81% identities with known firefly luciferases. The three purified recombinant proteins showed luminescence and fatty acyl-CoA synthetic activities, as observed in other firefly luciferases. The emission maxima by the three firefly luciferases (λmax, 545-546nm) were shorter than those by known luciferases from the nocturnal fireflies (λmax, 550-568nm). These results suggest that the primary structures and enzymatic properties of luciferases are conserved in Lampyridae, but the luminescence colors were red-shifted in nocturnal species compared to diurnal species.

Control of dry weight and tube feeding improved the general condition of a hemodialysis patient: report of a case.

Patients on long-term hemodialysis are at risk of developing malnutrition,and poor nutrient intake is an important factor in this. In the present case, we encountered a 55-year-old Japanese woman with end-stage renal failure and a past history of schizophrenia. Severe systemic edema was observed. Hemodialysis was started, but after one year she suddenly became unable to consume food orally, despite provision of a dietary plan by the nutrition support team (NST). Tube feeding was eventually implemented. Because the systemic edema did not improve, we decided to remove body fluid by intense hemodialysis. Hypotension was often observed during this hemodialysis, requiring dopamine. Over approximately 2 months, the patient's dry weight fell from 73 kg to 62 kg, the patient's activity improved and she became able to eat orally again, allowing tube feeding to be stopped. Although the reason for the sudden anorexia has not been clarified, tube feeding and dry weight control was successful in the treatment of this malnourished hemodialysis patient.

Enantioselective total synthesis of the potent anti-HIV nucleoside EFdA.

A concise enantioselective total synthesis of 4'-ethynyl-2-fluoro-2'-deoxyadenosine (EFdA), an extremely potent anti-HIV agent, has been accomplished from (R)-glyceraldehyde acetonide in 18% overall yield by a 12-step sequence involving a highly diastereoselective ethynylation of an α-alkoxy ketone intermediate.

Effects of granule-bound starch synthase I-defective mutation on the morphology and structure of pyrenoidal starch in Chlamydomonas.

Lowering of the CO2 concentration in the environment induces development of a pyrenoidal starch sheath, as well as that of pyrenoid and CO2-concentrating mechanisms, in many microalgae. In the green algae Chlamydomonas and Chlorella, activity of granule-bound starch synthase (GBSS) concomitantly increases under these conditions. In this study, effects of the GBSS-defective mutation (sta2) on the development of pyrenoidal starch were investigated in Chlamydomonas. Stroma starch- and pyrenoid starch-enriched samples were obtained from log-phase cells grown with air containing 5% CO2 (high-CO2 conditions favouring stromal starch synthesis) and from those transferred to low-CO2 conditions (air level, 0.04% CO2, favouring pyrenoidal starch synthesis) for 6h, respectively. In the wild type, total starch content per culture volume did not increase during the low-CO2 conditions, in spite of the development of pyrenoidal starch, suggesting that degradation of some part of stroma starch and synthesis of pyrenoid starch simultaneously occur under these conditions. Even in the GBSS-deficient mutants, pyrenoid and pyrenoid starch enlarged after lowering of the CO2 concentration. However, the morphology of the pyrenoid starch was thinner and more fragile than the wild type, suggesting that GBSS does affect the morphology of pyrenoidal starch. Surprisingly normal GBSS activity is shown to be required to obtain the high A-type crystallinity levels that we now report for pyrenoidal starch. A model is presented explaining how GBSS-induced starch granule fusion may facilitate the formation of the pyrenoidal starch sheath.

Specific Y14 domains mediate its nucleo-cytoplasmic shuttling and association with spliced mRNA.

Pre-mRNA splicing deposits multi-protein complexes, termed exon junction complexes (EJCs), on mRNAs near exon-exon junctions. The core of EJC consists of four proteins, eIF4AIII, MLN51, Y14 and Magoh. Y14 is a nuclear protein that can shuttle between the nucleus and the cytoplasm, and binds specifically to Magoh. Here we delineate a Y14 nuclear localization signal that also confers its nuclear export, which we name YNS. We further identified a 12-amino-acid peptide near Y14's carboxyl terminus that is required for its association with spliced mRNAs, as well as for Magoh binding. Furthermore, the Y14 mutants, which are deficient in binding to Magoh, could still be localized to the nucleus, suggesting the existence of both the nuclear import pathway and function for Y14 unaccompanied by Magoh.