4-Cl-edaravone and (E)-2-chloro-3-[(E)-phenyldiazenyl]-2-butenoic acid are the specific reaction products of edaravone with hypochlorite.

3-Methyl-1-phenyl-2-pyrazolin-5-one (edaravone) is a synthetic one-electron antioxidant used as a drug for treatment against acute phase cerebral infarction in Japan. This drug also reacts with two-electron oxidants like peroxynitrite to give predominantly 4-nitrosoedaravone but no one-electron oxidation products. It is believed that this plays a significant role in amelioration of amyotrophic lateral sclerosis. The drug was approved for treatment of amyotrophic lateral sclerosis in Japan and USA in 2015 and 2017, respectively. In this study, we examined the reaction of edaravone with another two-electron oxidant, hypochlorite anion (ClO-). Edaravone reacted with ClO- in 50% methanolic phosphate buffer (pH 7.4) solution containing typical two-electron reductants, such as glutathione, cysteine, methionine, and uric acid, as internal references. The concentration of edaravone decreased at a similar rate as each co-existing reference, indicating that it showed comparable reactivity toward ClO- as those references. Furthermore, 4-Cl-edaravone and (E)-2-chloro-3-[(E)-phenyldiazenyl]-2-butenoic acid (CPB) were identified as primary and end products, respectively, and no one-electron oxidation products were detected. These results suggest that edaravone treatment can bring greater benefit against ClO--related injury such as inflammation, and 4-Cl-edaravone and CPB can be good biomarkers for ClO--induced oxidative stress.

New Marine Antifouling Compounds from the Red Alga Laurencia sp.

Six new compounds, omaezol, intricatriol, hachijojimallenes A and B, debromoaplysinal, and 11,12-dihydro-3-hydroxyretinol have been isolated from four collections of Laurencia sp. These structures were determined by MS and NMR analyses. Their antifouling activities were evaluated together with eight previously known compounds isolated from the same samples. In particular, omaezol and hachijojimallene A showed potent activities (EC50 = 0.15-0.23 µg/mL) against larvae of the barnacle Amphibalanus amphitrite.

Single-Facility Analysis of COVID-19 Status of Healthcare Employees during the Eighth and Ninth Pandemic Waves in Japan after Introducing Regular Rapid Antigen Testing.

BACKGROUND: Community infections of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have increased rapidly since the emergence of the Omicron strain. During the eighth and ninth pandemic waves-when movement restrictions in the community were eased-the all-case registration system was changed, and the actual status of infection became uncertain. METHODS: We conducted regular rapid antigen tests (R-RATs) once or twice a week as self-testing to examine the actual state of coronavirus disease (COVID-19) diagnosis among healthcare employees. RESULTS: Overall, 320 (1.42/day) and 299 (1.76/day) employees were infected in the eighth and ninth pandemic waves. During both periods, 59/263 doctors (22.4%), 335/806 nurses (41.6%), 92/194 administrative employees (47.4%), and 129/218 clinical laboratory technicians (59.2%) were infected. In the eighth wave, 56 of 195 employees were infected through close contact; in the ninth wave, 26 of 62 employees were infected. No significant difference was observed in the number of vaccinations between infected and non-infected employees. The positivity rate of R-RATs was 0.41% and 0.45% in the eighth and ninth waves. R-RATs detected infection in 212 and 229 employees during the eighth and ninth waves, respectively; the ratio of R-RAT-detected positive employees to those who reported infection was significantly higher during the ninth wave (odds ratio: 1.67, 95% confidence interval: 1.17-2.37, p < 0.001). CONCLUSIONS: The number of infected healthcare employees remained high during the eighth and ninth pandemic waves in Japan. The R-RAT is considered effective for detecting mild or asymptomatic COVID-19 at an early stage and at a high rate in healthcare employees.

Using Video Calls to Reduce Risky Behaviors and Enhance Instruction Understanding of Patients in Acute Care Hospitals During the COVID-19 Pandemic.

Backgrounds During the COVID-19 pandemic, visitor restrictions in healthcare settings adversely affected patients. Video calls have emerged as an essential digital alternative that can decrease patients' anxiety and improve satisfaction. This study investigated whether family-initiated video calls could mitigate delirium symptoms and risky behaviors and enhance patients' comprehension of instructions. Methods This observational study used medical chart data and the Diem Payment System from a single acute care hospital in Fukuoka, Japan. The study involved patients hospitalized between May 2020 and August 2021 who used video chat systems. Patients or their relatives used video calls through Skype. The frequency of video chat use served as the primary exposure. Changes in the patients' risky behaviors and instruction comprehension upon discharge were the primary outcomes. Results A total of 532 patients were included in the study, with an average age of over 70 years. After implementing the inverse probability of treatment weighting adjustment, an improved balance across age, sex, BMI categories, and other variables was observed. The effects of video calls on risky behaviors and instruction comprehension varied. Patients with three or more video calls showed distinct effects compared with those with fewer calls. When hospitalization was limited to three weeks, video calls noticeably influenced risky behaviors (p=0.022, 95% CI:1.08-2.63), but not instruction comprehension (p=0.226, 95% CI:0.43-1.22). Conclusions The use of video calls as a visitation method in acute care hospitals during a pandemic suggests that video calls reduce risky behaviors in patients with a three-week stay. This alternative to physical visitations contributes positively to patient safety and supports ongoing efforts to prevent the spread of COVID-19.

Integration of methylation quantitative trait loci (mQTL) on dietary intake on DNA methylation levels: an example of n-3 PUFA and ABCA1 gene.

BACKGROUND: Epigenetic studies have reported relationships between dietary nutrient intake and methylation levels. However, genetic variants that may affect DNA methylation (DNAm) pattern, called methylation quantitative loci (mQTL), are usually overlooked in these analyses. We investigated whether mQTL change the relationship between dietary nutrient intake and leukocyte DNAm levels with an example of estimated fatty acid intake and ATP-binding cassette transporter A1 (ABCA1). METHODS: A cross-sectional study on 231 participants (108 men, mean age: 62.7 y) without clinical history of cancer and no prescriptions for dyslipidemia. We measured leukocyte DNAm levels of 8 CpG sites within ABCA1 gene by pyrosequencing method and used mean methylation levels for statistical analysis. TaqMan assay was used for genotyping a genetic variant of ABCA1 (rs1800976). Dietary fatty acid intake was estimated with a validated food frequency questionnaire and adjusted for total energy intake by using residual methods. RESULTS: Mean ABCA1 DNAm levels were 5% lower with the number of minor alleles in rs1800976 (CC, 40.6%; CG, 35.9%; GG, 30.6%). Higher dietary n-3 PUFA intake was associated with lower ABCA1 DNAm levels (1st (ref) vs. 4th, ß [95% CI]: -2.52 [-4.77, -0.28]). After controlling for rs180076, the association between dietary n-3 PUFA intake and ABCA1 DNAm levels was attenuated, but still showed an independent association (1st (ref) vs. 4th, ß [95% CI]: -2.00 [-3.84, -0.18]). The interaction of mQTL and dietary n-3 PUFA intake on DNAm levels was not significant. CONCLUSIONS: This result suggested that dietary n-3 PUFA intake would be an independent predictor of DNAm levels in ABCA1 gene after adjusting for individual genetic background. Considering mQTL need to broaden into other genes and nutrients for deeper understanding of DNA methylation, which can contribute to personalized nutritional intervention.

Identification of an active site of EMMPRIN for the augmentation of matrix metalloproteinase-1 and -3 expression in a co-culture of human uterine cervical carcinoma cells and fibroblasts.

OBJECTIVE: Extracellular matrix metalloproteinase inducer (EMMPRIN) is highly expressed on malignant tumor cell surface and accelerates tumor invasion. We previously reported that human uterine cervical carcinoma SKG-II cells exhibit the progression of in-vitro invasiveness by utilizing the enhanced production of matrix metalloproteinase (MMP) in human uterine cervical fibroblasts (HUCF) under an in-vitro co-culture model (Sato T et al., Gynecol Oncol 2004; 92:47-56). The aim of this study was to clarify the active site of EMMPRIN in the augmentation of MMP production in the co-culture of SKG-II cells and HUCF. METHODS: Western and Northern blot analyses were used to examine EMMPRIN and MMP expression in a co-culture of SKG-II cells or EMMPRIN-transfected COS-7 cells and HUCF. A systematic peptide screening method using nine synthetic EMMPRIN peptides was used to identify active site(s) of EMMPRIN for MMP induction. RESULTS: SKG-II cells constitutively expressed 53-kDa EMMPRIN on the cell surface and EMMPRIN production was enhanced under the co-culture. The concomitant augmentation of proMMP-3 production was diminished by adding an EMMPRIN antibody. EMMPRIN-transfected COS-7 cells stimulated HUCF to predominantly augment proMMP-1 and -3 expressions. A systematic peptide screening method revealed that (42)SLNDSATEVTGHRWLK(57) in the first loop domain of EMMPRIN participated in the augmentation of proMMP-1 production. CONCLUSIONS: These results provide a novel mechanism of malignancy of uterine cervical carcinoma, in that the augmentation of EMMPRIN expression by tumor-stromal cell interaction progresses tumor invasion along with the increase of MMP expression via an active site of EMMPRIN, (42)SLNDSATEVTGHRWLK(57).

Identification of a gene cluster of polyether antibiotic lasalocid from Streptomyces lasaliensis.

Elucidation of enzymatic polyether formation is a long-standing controversial issue in organic chemistry. To address this intriguing issue, identifying the actual substrate for epoxidation and sequential cyclization is essential. We selected the representative polyether ionophore, lasalocid, which has been proposed to undergo no modification at the late stage of biosynthesis. Cloning and a sequence analysis revealed seven polyketide synthase (PKS) genes, epoxidase and epoxide hydrolase genes for sequential ether formation, and several putative genes for supplying ethylmalonyl-CoA. Based on bioinformatic data, we propose the lasalocid biosynthetic pathway which involves characteristic aromatic ring formation and sequential cyclic ether formation. The finding of a thioesterase domain at the C-terminal of the seventh PKS indicates that intriguing oxidative cascade cyclization would occur after cleavage of the polyketide intermediate from PKS. Based on this observation, we have recently reported the enzymatic transformation of a bisepoxide intermediate to lasalocid with the recombinant epoxide hydrolase, Lsd19.

Epoxide hydrolase Lsd19 for polyether formation in the biosynthesis of lasalocid A: direct experimental evidence on polyene-polyepoxide hypothesis in polyether biosynthesis.

Polyether metabolites are an important class of natural products. Although their biosynthesis, especially construction of polyether skeletons, attracted organic chemists for many years, no experimental data on the enzymatic polyether formation has been obtained. In this study, a putative epoxide hydrolase gene lsd19 found on the biosynthetic gene cluster of an ionophore polyether lasalocid was cloned and successfully overexpressed in Escherichia coli. Using the purified Lsd19, a proposed substrate, bisepoxyprelasalocid, and its synthesized analogue were successfully converted into lasalocid A and its derivative via a 6-endo-tet cyclization mode. On the other hand, treatment of the bisepoxide with trichloroacetic acid gave isolasalocid A via a 5-exo-tet cyclization mode. Therefore, the enzymatic conversion observed in this study unambiguously showed that the bisepoxyprelasalocid is an intermediate of the lasalocid biosynthesis and that Lsd19 catalyzes the sequential cyclic ether formations involving an energetically disfavored 6-endo-tet cyclization. This is the first example of the enzymatic epoxide-opening reactions leading to a polyether natural product.

Individualized antiplatelet therapy after drug-eluting stent deployment: Implication of clinical trials of different durations of dual antiplatelet therapy.

At present, there is consensus that prolonged dual antiplatelet therapy (DAPT) is effective to reduce cardiovascular events at the expense of bleeding complication events. A causal relationship of prolonged DAPT with an increase in mortality remains debatable, however, it appears to be obvious that bleeding complications are associated with an increase in cardiac events. Thus, individualized optimal DAPT duration balancing the risk and benefit of DAPT should be applied. In addition, strategy to minimize bleeding complications is highly recommended. Several risk scores have been reported to discriminate the risk and benefits of DAPT. However, in general, bleeding risk and event risk are correlated with each other, thus predictability of these scores is limited to moderate. Therefore, interpretation of previous trials is important to overcome the shortcome in outcomes. In this review, we provide an overview of DAPT trials and clarify the shortfalls to consider in Japan. Finally, possible future trends with reference to the results of recent clinical trials will be presented.

In vivo treatment with the K+ channel blocker 4-aminopyridine protects against kainate-induced neuronal cell death through activation of NMDA receptors in murine hippocampus.

Activation of NMDA receptors has been shown to induce either neuronal cell death or neuroprotection against excitotoxicity in cultured neurons in vitro. To elucidate in vivo neuroprotective role of NMDA receptors, we investigated the effects of activation of NMDA receptors by endogenous glutamate on kainate-induced neuronal damage to the mouse hippocampus in vivo. The systemic administration of the K+ channel blocker 4-aminopyridine (4-AP, 5 mg/kg, i.p.) induced expression of c-Fos in the hippocampal neuronal cell layer, which expression was completely abolished by the noncompetitive NMDA receptor antagonist MK-801, thus indicating that the administration of 4-AP would activate NMDA receptors in the hippocampal neurons. The prior administration of 4-AP at 1 h to 1 day before significantly prevented kainate-induced pyramidal cell death in the hippocampus and expression of pyramidal cells immunoreactive with an antibody against single-stranded DNA. Further immunohistochemical study on deoxyribonuclease II revealed that the pretreatment with 4-AP led to complete abolition of deoxyribonuclease II expression induced by kainate in the CA1 and CA3 pyramidal cells. The neuroprotection mediated by 4-AP was blocked by MK-801 and by the adenosine A1 antagonist 8-cyclopenthyltheophylline. Taken together, in vivo activation of NMDA receptors is capable of protecting against kainate-induced neuronal damage through blockade of DNA fragmentation induced by deoxyribonuclease II in the murine hippocampus.

Enhanced binding activity of nuclear antioxidant-response element through possible formation of Nrf2/Fos-B complex after in vivo treatment with kainate in murine hippocampus.

To evaluate whether in vivo glutamate signals modulate signaling processes mediated by antioxidant-response element (ARE), we examined ARE binding in nuclear extracts from the hippocampus after in vivo treatment of mice with kainate. Enhancement of ARE binding was found at 2 h to 3 days after kainate treatment. Supershift analysis indicated possible involvement of Nrf2, Fos-B, and c-Fos in ARE binding in hippocampal nuclear extracts obtained from kainate-treated animals. On super-supershift analysis by combination of these antibodies, ARE probe/protein complex was shifted by the anti-Fos-B antibody alone, but not by the anti-c-Fos antibody alone, and further addition of the anti-Nrf2 antibody dramatically eliminated binding of the complex shifted by the anti-Fos-B antibody in hippocampal nuclear extracts from kainate-treated animals. Kainate treatment induced a profound increase in levels of c-Fos and Fos-B, without markedly affecting that of Nrf2 in nuclear extracts from the hippocampus. Co-localization of Nrf2 with both Fos-B and c-Fos was found in neuronal cell layers of the hippocampus in kainate-treated animals. RT-PCR analysis revealed that kainate treatment increases glutathione-S-transferase mRNA level in the hippocampus. Taken together, kainate signals may enhance nuclear ARE binding through an interaction between constitutive Nrf2 with inducible Fos-B expressed in murine hippocampus.

In vivo activation of c-Jun N-terminal kinase signaling cascade prior to granule cell death induced by trimethyltin in the dentate gyrus of mice.

The systemic administration of trimethyltin (TMT, 2.8 mg/kg, i.p.) induced granule cell death in the mouse dentate gyrus selectively 2 days later. The administration of TMT not only enhanced activator protein-1 DNA binding, along with an increase in expression of c-Jun and Fra-2, in the hippocampus 1 day later, but also facilitated phosphorylation of c-Jun N-terminal kinase (JNK) within the cytosol and nucleus. There was also a concomitant increase in the level of phosphorylated JNK kinase (MKK4/SEK1) in the cytosol 16-24 h after the administration. Moreover, TMT markedly elevated endogenous levels of both phosphorylated c-Jun and phosphorylated activating transcription factor-2 (ATF-2), in addition to activating JNK activity in the nuclear extracts obtained 16-24 h post-administration. Immunohistochemical analysis revealed that whereas Fra-2 and phosphorylated ATF-2 were expressed in the CA1 pyramidal cell layer predominantly, phosphorylated c-Jun was observed in both the CA1 pyramidal and dentate granule cell layers after TMT administration. Taken together, our data indicate that TMT activates the JNK pathway in the hippocampus prior to neuronal cell death. The prior activation of this pathway could be at least in part involved in the TMT-induced neural damage seen in the dentate granule cells of mice.

Diurnal and circadian rhythm in compound eye of cricket (Gryllus bimaculatus): changes in structure and photon capture efficiency.

Day-night changes in rhabdom size of compound eyes were investigated in three groups of crickets (Gryllus bimaculatus): nymphs and adult males and females. In both adults and nymphs, the rhabdoms were larger at night than during a day. In adults, the mean rhabdom occupation ratios (RORs) of ommatidial retinulae at midnight were about two times greater than the values at midday. This change contributes to control of the photon capture efficiency (PCE) of the eye according to photic environment. The RORs of adult males at midnight were higher than those of both adult females and nymphs. This suggests that the PCE of the compound eye of adult males is the greatest of all groups. Under constant darkness, day-night changes in ROR were detected only in adult males, but neither in adult females nor in nymphs. On the other hand, no day-night changes were detected in any experimental group under constant light. These results suggest that the change in rhabdom size between day and night is an adaptation to the photic environment that is controlled mainly by the light-dark (day-night) cycle. However, the change in male adults is induced by an endogenous circadian clock.

A novel functional site of extracellular matrix metalloproteinase inducer (EMMPRIN) that limits the migration of human uterine cervical carcinoma cells.

EMMPRIN (extracellular matrix metalloproteinase inducer)/CD147, a membrane-bound glycoprotein with two extracellular loop domains (termed loops I and II), progresses tumor invasion and metastasis by increasing the production of matrix metalloproteinase (MMP) in peritumoral stoma cells. EMMPRIN has also been associated with the control of migration activity in some tumor cells, but little is known about how EMMPRIN regulates tumor cell migration. In the present study, EMMPRIN siRNA suppressed the gene expression and production of EMMPRIN in human uterine cervical carcinoma SKG-II cells. An in vitro scratch wound assay showed enhancement of migration of EMMPRIN-knockdown SKG-II cells. In addition, the SKG-II cell migration was augmented by adding an E. coli-expressed human EMMPRIN mutant with two extracellular loop domains (eEMP-I/II), which bound to the cell surface of SKG-II cells. However, eEMP-I/II suppressed the native EMMPRIN-mediated augmentation of proMMP-1/procollagenase-1 production in a co-culture of the SKG-II cells and human uterine cervical fibroblasts, indicating that the augmentation of SKG-II cell migration resulted from the interference of native EMMPRIN functions by eEMP-I/II on the cell surface. Furthermore, a systematic peptide screening method using nine synthetic EMMPRIN peptides coding the loop I and II domains (termed EM1-9) revealed that EM9 (170HIENLNMEADPGQYR184) facilitated SKG-II cell migration. Moreover, SKG-II cell migration was enhanced by administration of an antibody against EM9, but not EM1 which is a crucial site for the MMP inducible activity of EMMPRIN. Therefore, these results provide novel evidence that EMMPRIN on the cell surface limits the cell migration of human uterine cervical carcinoma cells through 170HIENLNMEADPGQYR184 in the loop II domain. Finally, these results should provide an increased understanding of the functions of EMMPRIN in malignant cervical carcinoma cells, and could contribute to the development of clinical strategies for cervical cancer therapy.

Intriguing substrate tolerance of epoxide hydrolase Lsd19 involved in biosynthesis of the ionophore antibiotic lasalocid A.

Recently, we reported that the epoxide hydrolase Lsd19, the first enzyme shown to catalyze epoxide-opening cascades, can catalyze the conversion of a putative bisepoxide intermediate to polyether antibiotic lasalocid, which involves energetically disfavored 6-endo-tet cyclization of the epoxy alcohol. Here, we examined the substrate tolerance of Lsd19. Lsd19 accepts various substrate analogues differing in the left segment of lasalocid and epoxide stereochemistry to afford either THF-THP or THF-THF products with excellent regioselectivity.

Mother-to-daughter occurrence of biliary atresia: a case report.

We herein report a case of mother-to-daughter occurrence of biliary atresia (BA). The mother underwent a Kasai hepatic portoenterostomy at the age of 64 days with a diagnosis of type III BA. Her postoperative course was uneventful, with a good bile discharge, and she became pregnant at the age of 20 years. At 21 weeks of gestational age, fetal ultrasonographic examination of the baby revealed a cystic mass at the hepatic hilum. The baby was born via vaginal delivery at 41 weeks of gestation with a birth weight of 3269 g. Because of an increasing direct bilirubin level and the absence of bile excretion, a laparotomy was performed at the age of 29 days. Intraoperative cholangiography revealed the diagnosis of BA I-cyst, and a hepaticojejunostomy was performed with a good bile discharge. To our knowledge, these patients are the first occurrence of BA in a transmission pattern of mother to daughter.

Gastrectomy performed with noninvasive positive pressure ventilation for a patient with severe chronic obstructive pulmonary disease: report of a case.

We report the case of a patient with severe chronic obstructive pulmonary disease (COPD) for whom gastrectomy was successfully performed with the use of noninvasive positive pressure ventilation (NPPV). A 63-year-old man who had been suffering from chronic pulmonary emphysema for 12 years and receiving home oxygen therapy (HOT) for 9 years was diagnosed with gastric carcinoma. The patient required supplemental oxygen via nasal cannulae even at rest, and had labored breathing through pursed lips after a short conversation. The forced expiratory volume in 1 s was 400 ml. He underwent conventional gastrectomy under general anesthesia, and was extubated 90 min after surgery and given NPPV support. He was successfully weaned from NPPV on postoperative day (POD) 10 and discharged from our hospital on POD 28. Noninvasive positive pressure ventilation is useful for the perioperative management of patients with severe COPD and for extending the possibilities of surgery for patients on HOT.

Sequences and evolutionary analyses of eukaryotic-type protein kinases from Streptomyces coelicolor A3(2).

Four eukaryotic-type protein serine/threonine kinases from Streptomyces coelicolor A3(2) were cloned and sequenced. To explore evolutionary relationships between these and other protein kinases, the distribution of protein serine/threonine kinase genes in prokaryotes was examined with the TFASTA program. Genes of this type were detected in only a few species of prokaryotes and their distribution was uneven; Streptomyces, Mycobacterium, Synechocystis and Myxococcus each contained more than three such genes. Homology analyses by GAP and Rdf2 programs suggested that some kinases from one species were closely related, whilst others were only remotely related. This was confirmed by examining phylogenetic trees constructed by the neighbour-joining and other methods. For each species, analysis of the coding regions indicated that the G+C content of protein kinase genes was similar to that of other genes. Considered with the fact that in phylogenetic trees the amino acid sequences of STPK from Aquifex aeolicus and some other eukaryotic-type protein kinases in prokaryotes form a cluster with protein kinases from eukaryotes, this suggests that the eukaryotic-type protein kinases were present originally in both prokaryotes and eukaryotes, but that most of these genes have been lost during the evolutionary process in prokaryotes because they are not needed. This conclusion is supported by the observation that the prokaryotes retaining several of these kinases undergo complicated morphological and/or biochemical differentiation.