Japanese Planocerid Flatworms: Difference in Composition of Tetrodotoxin and Its Analogs and the Effects of Ingestion by Toxin-Bearing Fishes in the Ryukyu Islands, Japan.

Tetrodotoxin (TTX), known as pufferfish toxin, is a potent neurotoxin blocking sodium channels in muscle and nerve tissues. TTX has been detected in various taxa other than pufferfish, including marine polyclad flatworms, suggesting that pufferfish toxin accumulates in fish bodies via food webs. The composition of TTX and its analogs in the flatworm Planocera multitentaculata was identical to those in wild grass puffer Takifugu alboplumbeus. Previously, Planocera sp. from Okinawa Island, Japan, were reported to possess high level of TTX, but no information was available on TTX analogs in this species. Here we identified TTX and analogs in the planocerid flatworm using high-resolution liquid chromatography-mass spectrometry, and compared the composition of TTX and analogs with those of another toxic and non-toxic planocerid species. We show that the composition of TTX and several analogs, such as 5,6,11-trideoxyTTX, dideoxyTTXs, deoxyTTXs, and 11-norTTX-6(S)-ol, of Planocera sp. was identical to those of toxic species, but not to its non-toxic counterpart. The difference in the toxin composition was reflected in the phylogenetic relationship based on the mitochondrial genome sequence. A toxification experiment using predatory fish and egg plates of P. multitentaculata demonstrated that the composition of TTX and analogs in wild T. alboplumbeus juveniles was reproduced in artificially toxified pufferfish. Additionally, feeding on the flatworm egg plates enhanced the signal intensities of all TTX compounds in Chelonodon patoca and that of deoxyTTXs in Yongeichthys criniger.

Structural Development of Androgen Receptor Antagonists Using Phenylferrocene Framework as a Hydrophobic Pharmacophore.

We previously identified nitrophenylferrocenes and cyanophenylferrocenes as promising lead structures of novel androgen receptor (AR) antagonists, based on the structural similarity between ferrocene and the steroidal skeleton. In the present research, we explored the structure-activity relationship (SAR) of phenylferrocene derivatives. Introduction of a hydrophobic substituent such as a chlorine atom at the 2-position or 3-position of phenylferrocene derivatives significantly increased the antagonistic activity toward wild-type AR, and among the synthesized compounds, 3-chloro-4-cyanophenylferrocene (29) exhibited the most potent anti-proliferative activity toward the androgen-dependent growth of SC-3 cells expressing wild-type AR (IC50 14 nM). Like conventional antiandrogens such as hydroxyflutamide, the major active metabolite of flutamide, compound 29 exhibited agonistic activity toward T877A-AR, a mutant AR expressed in human prostate cancer cell line LNCaP. Notably, however, the 2-chloro isomer 27 showed potent antagonistic activity toward wild-type AR (IC50 49 nM) and also exhibited antagonistic activity toward T877A-AR. Our SAR data should prove helpful for the development of new-generation AR antagonists based on phenylferrocene as candidate agents to treat drug-resistant prostate cancer.

The highly developed symbiotic system between the solar-powered nudibranch Pteraeolidia semperi and Symbiodiniacean algae.

The intricate coexistence of Symbiodiniacean algae with a diverse range of marine invertebrates underpins the flourishing biodiversity observed within coral reef ecosystems. However, the breakdown of Symbiodiniaceae-host symbiosis endangers these ecosystems, necessitating urgent study of the symbiotic mechanisms. The symbiosis between nudibranchs and Symbiodiniaceae has been identified as an efficacious model for examining these mechanisms, yet a comprehensive understanding of their histological structures and cellular processes remains elusive. A meticulous histological exploration of the nudibranch Pteraeolidia semperi, employing optical, fluorescence, and electron microscopy, has revealed fine tubules extending to the body surface, with associated epithelial cells having been shown to adeptly encapsulate Symbiodiniaceae intracellularly. By tracing the stages of the "bleaching" in nudibranchs, it was inferred that algal cells, translocated via the digestive gland, are directly phagocytosed and expelled by these epithelial cells. Collectively, these insights contribute substantially to the scholarly discourse on critical marine symbiotic associations.

A Case of Leg Pain After the Initial Dose of the COVID-19 Vaccine, Followed by Deep Vein Thrombosis and Pulmonary Embolism After the Second Dose.

COVID-19 has been spreading worldwide. Therefore, the COVID-19 vaccine is recommended for prevention. However, adverse events after COVID-19 vaccination remain an issue, and we should monitor patients for adverse events and determine their association with COVID-19 vaccination. Here, we report a case involving a 48-year-old Japanese woman who experienced dull left leg pain that resolved spontaneously after the first vaccine dose, followed by deep vein thrombosis (DVT) and pulmonary embolism (PE) after the second dose. The findings from this case suggest that the COVID-19 vaccine could cause severe adverse events, such as DVT. Therefore, patients should understand their subjective symptoms and report any side effects experienced after the first dose before they take the second dose. Furthermore, medical providers should enquire about all possible symptoms experienced after the initial dose before they administer the second dose.

Complete genome sequence of Edwardsiella sp. NBRC12716 isolated in 1962 from the liver of diseased eel.

We report the complete genome sequence of Edwardsiella sp. NBRC12716 isolated from a diseased eel in 1962. The genome consists of a single, circular chromosome 3,771,060 bp in length with 59.74% GC content and encodes 25 rRNA, 96 tRNA, and 3,182 protein-coding genes.

The complete mitochondrial genome of Spurilla braziliana MacFarland 1909 (Nudibranchia, Aeolidiidae).

Spurilla braziliana MacFarland 1909 is a morphologically diverse nudibranch found in the Pacific and Western Atlantic. The complete mitochondrial genome of S. braziliana has been constructed using next-generation sequencing technology. The mitochondrial genome is 14,291 bp and contains 13 protein-coding genes, 2 rRNA genes, and 23 tRNA genes. Molecular phylogenetic analysis using the maximum likelihood method revealed that S. braziliana is included in the superfamily Aeolidioidea and forms a monophyletic group with Berghia stephanieae, a nudibranch of the family Aeolidiidae. This study reinforces existing taxonomic insights and provides a basis for further molecular phylogenetic analysis.

Case of nivolumab-induced delayed adrenal insufficiency during the treatment of renal cell carcinoma.

INTRODUCTION: Nivolumab is an immune checkpoint inhibitor used to treat advanced renal cell carcinoma (RCC). Adrenal insufficiency has been reported as an adverse event associated with nivolumab. We report a case of adrenal insufficiency that occurred more than 1 year after the initiation of nivolumab while patient was still receiving treatment. CASE REPORT: The patient was a 90-year-old Japanese woman. Fatigue and decreased cortisol levels were observed after 15 courses of nivolumab. MANAGEMENT & OUTCOME: The symptoms improved with the initiation of oral hydrocortisone 30 mg once a day. Nivolumab was not resumed, and the patient is still under outpatient observation. DISCUSSION: This is the first report of RCC with adrenal insufficiency occurring more than 1 year after the initiation of the nivolumab regimen. Symptoms of adrenal insufficiency are similar to those of cancer progression. When symptoms of fatigue occur in patients receiving nivolumab, adrenal insufficiency should be suspected, regardless of the duration from nivolumab initiation.

A case of blood triglyceride increased induced by ABVD therapy for classical Hodgkin lymphoma.

There are no reports of blood triglyceride (TG) levels increasing with the ABVD regimen. Herein, we present a case of Hodgkin's lymphoma that exhibited ABVD-induced blood TG increase. The patient was a 40-year-old Japanese man. Empiric therapy was initiated using the ABVD regimen for Hodgkin lymphoma. On day 58, the fasting blood TG concentration increased to 1,451 mg/dL. Since no adverse events were noted, 0.2 mg/day of pemafibrate was administered, and the ABVD regimen was continued. Blood TG levels should be periodically monitored during ABVD administration for the patients who are at high risk of increased blood TG levels.

Draft genome sequencing and assembly of Risso's dolphin, Grampus griseus.

The Risso's dolphin (Grampus griseus) is one of the migratory marine mammals and they have commonly dispersed in tropical and temperate seas. It is a least concerned species in the IUCN red list of threatened species. However, their population size and factors affecting their population structure are unknown. Due to the wide distribution of this species, their populations might be genetically stable and less structured. To support genetic studies on dolphins and other marine mammals, we assembled the draft genome of Risso's dolphin that was found in Japan. The tissue samples were used to extract high molecular DNA and subjected to sequencing by Illumina HiSeq X, Oxford Nanopore MinION, and Bionano Saphyr. The assembled hybrid genome was 75.9% of complete eukaryotic BUSCOs and the genome size was 2.256 Gb with 2.042 Mb of scaffold N50. De novo assembly of this genome by Bionano Saphyr recovered 2.036 Gb total genome map length and structural variations. The gene structures of this draft genome were identified by BRAKER2, and 9947 genes were recovered. The data will be useful for future studies of cetaceans.

Speech Recognition System Generates Highly Accurate Endoscopic Reports in Clinical Practice.

Objective Endoscopic reports are conventionally written at the end of each procedure, and the endoscopist must complete the report from memory. To make endoscopic reporting more efficient, we developed a new speech recognition (SR) system that generates highly accurate endoscopic reports based on structured data entry. We conducted a pilot study to examine the performance of this SR system in an actual endoscopy setting with various types of background noise. Methods In this prospective observational pilot study, participants who underwent upper endoscopy with our SR system were included. The primary outcome was the correct recognition rate of the system. We compared the findings generated by the SR system with the findings in the handwritten report prepared by the endoscopist. The initial correct recognition rate, number of revisions, finding registration time, and endoscopy time were also analyzed. Results Upper endoscopy was performed in 34 patients, generating 128 findings of 22 disease names. The correct recognition rate was 100%, and the median number of revisions was 0. The median finding registration time was 2.57 [interquartile range (IQR), 2.33-2.92] seconds, and the median endoscopy time was 234 (IQR, 194-227) seconds. Conclusion The SR system demonstrated high recognition accuracy in the clinical setting. The finding registration time was extremely short.

Construction of a chromosome-level Japanese stickleback species genome using ultra-dense linkage analysis with single-cell sperm sequencing.

It is still difficult to construct the genomes of higher organisms as their genome sequences must be extended to the length of the chromosome by linkage analysis. In this study, we attempted to provide an innovative alternative to conventional linkage analysis by devising a method to genotype sperm using 10× Genomics single-cell genome sequencing libraries to generate a linkage map without interbreeding individuals. A genome was assembled using sperm from the Japanese stickleback Gasterosteus nipponicus, with single-cell genotyping yielding 1 864 430 very dense hetero-SNPs and an average coverage per sperm cell of 0.13×. In total, 1665 sperm were used, which is an order of magnitude higher than the number of recombinations used for conventional linkage analysis. We then improved the linkage analysis tool scaffold extender with low depth linkage analysis (SELDLA) to analyze the data according to the characteristics of the single-cell genotyping data. Finally, we were able to determine the chromosomal location (97.1%) and orientation (64.4%) of the contigs in the 456 Mb genome of G. nipponicus, sequenced using nanopores. This method promises to be a useful tool for determining the genomes of non-model organisms for which breeding systems have not yet been established by linkage analysis.

Signal of safety due to adverse drug reactions induced by tacrolimus with or without azithromycin.

INTRODUCTION: We identified two reports of drug levels increased and acute kidney injury caused by the drug-drug interaction between azithromycin (AZM) and tacrolimus (TAC). However, it is unclear whether the combination of these two drugs causes additive or synergistic adverse drug reactions. Therefore, we evaluated the disproportionality in reporting drug level increased and acute kidney injury for these two drugs are used alone and in combination with each other. METHOD: Data from the US Food and Drug Administration's Adverse Event Reporting System from 1974 to Q3/2021 were used. Reports based on exposure to macrolide antibiotic alone, TAC alone, and each macrolide antibiotic + TAC were extracted. Proportional reporting ratios (PRRs) and 95% confidence intervals (CIs) were calculated, and a lower limit of the 95% CI (Lower95CI) value of 2.0 or higher was interpreted as a signal of safety. RESULTS: Lower95CIs for macrolide antibiotic alone and TAC showed no potential signals of safety, including drug level increase, acute kidney injury, and control event. The PRRs and 95% CI for drug levels increased were 3.27 (2.69-3.97) for AZM + TAC, and 10.81 (9.59-12.17) for clarithromycin (CAM) + TAC. For CAM + TAC, the PRR and 95% CI were 8.42 (7.51-9.44) in acute kidney injury. However, AZM + TAC was not associated with a signal of safety in acute kidney injury. CONCLUSIONS: This suggests that AZM + TAC has a low risk of causing acute kidney injury but may cause increased drug levels.

Selection of a reference gene for studies on lipid-related aquatic adaptations of toothed whales (Grampus griseus).

Toothed whales are one group of marine mammals that has developed special adaptations, such as echolocation for predation, to successfully live in a dynamic aquatic environment. Their fat metabolism may differ from that of other mammals because toothed whales have acoustic fats. Gene expression in the metabolic pathways of animals can change with respect to their evolution and environment. A real-time quantitative polymerase chain reaction (RT-qPCR) is a reliable technique for studying the relative expressions of genes. However, since the accuracy of RT-qPCR data is totally dependent on the reference gene, the selection of the reference gene is an essential step. In this study, 10 candidate reference genes (ZC3H10, FTL, LGALS1, RPL27, GAPDH, FTH1, DCN, TCTP, NDUS5, and UBIM) were initially tested for amplification efficiency using RT-qPCR. After excluding DCN, the remaining nine genes, which are nearly 100% efficient, were selected for the gene stability analysis. Stable reference genes across eight different fat tissue, liver, and muscle samples from Grampus griseus were identified by four algorithms, which were provided in Genorm, NormFinder, BestKeeper, and Delta CT. Finally, a RefFinder comprehensive ranking was performed based on the stability values, and the nine genes were ranked as follows: LGALS1 > FTL > GAPDH > ZC3H10 > FTH1 > NDUS5 > TCTP > RPL27 > UBIM. The LGALS1 and FTL genes were identified as the most stable novel reference genes. The third-ranked gene, GAPDH, is a well-known housekeeping gene for mammals. Ultimately, we suggest the use of LGALS1 as a reliable novel reference gene for genomics studies on the lipid-related aquatic adaptations of toothed whales.

Another polymorphic mitochondrial genome of Grampus griseus and phylogeny of family Delphinidae.

Risso's dolphin (Grampus griseus Cuvier, 1812) is the only species of genus Grampus and a cosmopolitan marine inhabitant. Here, we report a polymorphic complete mitochondrial genome of G. griseus. The size of the total mitochondrial genome was 16,386 bp in length and contains 13 protein-coding genes, 22 transfer RNA genes, two ribosomal RNA genes, and a control region. 37 single nucleotide polymorphic sites (SNPs) were identified compared to the references. Based on the available total mitochondrial dolphin genomes' phylogenetics, G. griseus has formed a clade with 0.1415 distance, sister to the following species of the subfamily Globicephalinae and the taxonomy of Orcinus orca still needs further investigations.

Simultaneous amplicon analysis of multiple soil samples using MinION sequencing.

The diversity and composition of soil microorganisms needs to be understood as they influence ecosystem processes. MinION is a relatively recent next-generation sequencer, which provides the advantage of sequencing long reads. In this study, two types of soil were prepared experimentally to investigate the possibility of simultaneously analyzing multiple environmental samples using MinION. The MinION sequencing of amplicons was adjusted by the different rounds of PCR performed. Soil fungi and bacteria were compared using ITS and 16S rRNA amplicons, respectively. For ITS, the number of reads available for MinION sequencing were simply increased by performing two PCRs and purification using Agencourt AMPure XP. However, the effect of performing PCR twice was not high for 16S rRNA. Therefore, performing PCR twice appears to be effective for analyzing ITS regions. Regarding the number of reads obtained using MinION sequencing, clustering the same sample was possible if a read of ∼2000 bases or more was obtained in 16S rRNA and ITS. Further, information on 80 samples was obtained by performing only one round of MinION sequencing. Thus, MinION sequencing can be used to analyze a large number of samples simultaneously, providing a strong tool for amplicon sequencing. • Soil microbial composition before and after treatment was compared between 16S rRNA and ITS amplicons using MinION sequencing • One PCR amplification and two PCR amplifications were also compared • Information on 80 samples was obtained by performing only one round of MinION sequencing.

Physical frailty predicts the development of social frailty: a prospective cohort study.

BACKGROUND: It has not been clarified whether physical frailty symptoms predict social. frailty. The purpose of this study was to elucidate the effect of physical frailty on social frailty, and to determine which domains of physical frailty predict the development of social frailty. METHODS: We employed a two-year prospective cohort study. A total of 342 socially robust community-dwelling older adults were recruited. We used a modified social frailty screening index consisting of four social domains including financial difficulties, living alone, social activity, and contact with neighbors. Physical frailty status was also assessed at baseline. At the two-year follow-up, we assessed the development of social frailty. Social status was assessed using four social subdomains for the primary analysis. Social status was assessed using the two social subdomains of social activity and contact with neighbors, which would be affected by the physical frailty component, for the secondary analysis. The risk ratios (RR) of physical frailty for the development of social frailty were estimated. RESULTS: Although physical frailty symptoms were not a significant risk factor for future development of social frailty as assessed by four social subdomains (adjusted RR 1.39, 95% CI 0.95-2.15), it became significant when development of social frailty was assessed by the two social subdomains (adjusted RR 1.78, 95% CI 1.10-2.88). An analysis using the physical frailty subdomain showed that slow gait speed (adjusted RR 3.41, 95% CI 1.10-10.53) and weakness (adjusted RR 1.06, 95% CI 1.01-1.12) were independent risk factors for development of social frailty as assessed by two social subdomains. CONCLUSIONS: Physical frailty symptoms predict the development of social frailty. Among physical frailty subdomains, gait speed and muscle strength are critical independent risk factors for future decline in the social aspect. The prevention of physical frailty, especially by maintaining gait ability and muscle strength, may be effective for avoiding social frailty.

Tyrphostin AG-related compounds attenuate H2O2-induced TRPM2-dependent and -independent cellular responses.

PURPOSE: TRPM2 is a Ca2+-permeable channel that is activated by H2O2. TRPM2-mediated Ca2+ signaling has been implicated in the aggravation of inflammatory diseases. Therefore, the development of TRPM2 inhibitors to prevent the aggravation of these diseases is expected. We recently reported that some Tyrphostin AG-related compounds inhibited the H2O2-induced activation of TRPM2 by scavenging the intracellular hydroxyl radical. In the present study, we examined the effects of AG-related compounds on H2O2-induced cellular responses in human monocytic U937 cells, which functionally express TRPM2. METHODS: The effects of AG-related compounds on H2O2-induced changes in intracellular Ca2+ concentrations, extracellular signal-regulated kinase (ERK) activation, and CXCL8 secretion were assessed using U937 cells. RESULTS: Ca2+ influxes via TRPM2 in response to H2O2 were blocked by AG-related compounds. AG-related compounds also inhibited the H2O2-induced activation of ERK, and subsequent secretion of CXCL8 mediated by TRPM2-dependent and -independent mechanisms. CONCLUSION: Our results show that AG-related compounds inhibit H2O2-induced CXCL8 secretion following ERK activation, which is mediated by TRPM2-dependent and -independent mechanisms in U937 cells. We previously reported that AG-related compounds blocked H2O2-induced TRPM2 activation by scavenging the hydroxyl radical. The inhibitory effects of AG-related compounds on TRPM2-independent responses may be due to scavenging of the hydroxyl radical.

Inhibitory effects of Tyrphostin AG-related compounds on oxidative stress-sensitive transient receptor potential channel activation.

Some transient receptor potential (TRP) proteins including TRPA1, TPRM2 and TRPV1 are oxidative stress-sensitive Ca(2+)-permeable channels. Ca(2+) signaling via these TRP channels activated by oxidative stress has been implicated in the aggravation of various inflammatory diseases and pain sensation. We recently reported that Tyrphostin AG490 exerted inhibitory effects on H2O2-induced TRPM2 activation by scavenging the hydroxyl radical. In order to identify stronger inhibitors of oxidative stress-sensitive TRP channels than AG490, we examined the inhibitory effects of Tyrphostin AG-related compounds on H2O2-induced TRP channel activation in human embryonic kidney 293 cells expressing TRP channels. AG555 and AG556 blocked the activation of TRPM2 by H2O2 more strongly than AG490. Regarding TRPV1 and TRPA1, none of the three compounds tested affected H2O2-induced TRPV1 activation; however, AG555 and AG556 reduced H2O2-induced TRPA1 activation more than AG490. Thus, we herein identified AG555 and AG556 as new compounds that exert stronger inhibitory effects on H2O2-induced TRPM2 and TRPA1 activation than AG490. Edaravone, a hydroxyl radical scavenger used in the treatment of cerebral hemorrhage and cerebral infarction, did not affect H2O2-induced TRPM2 or TRPA1 activation. AG555 and AG556 may be useful seed compounds as therapeutic agents for several TRP-related diseases associated with oxidative stress.

TRPM2 channels in alveolar epithelial cells mediate bleomycin-induced lung inflammation.

Lung inflammation is a major adverse effect of therapy with the antitumor drug bleomycin (BLM). Transient receptor potential melastatin 2 (TRPM2) is a Ca(2+)-permeable channel that is activated by oxidative stress through the production of ADP-ribose. We herein investigated whether TRPM2 channels contributed to BLM-induced lung inflammation. The intratracheal instillation of BLM into wild-type (WT) mice increased the number of polymorphonuclear leukocytes (PMNs) and inflammatory cytokine levels in the lung. Increases in inflammatory markers in WT mice were markedly reduced in trpm2 knockout (KO) mice, which demonstrated that the activation of TRPM2 channels was involved in BLM-induced lung inflammation. The expression of TRPM2 mRNA was observed in alveolar macrophages, alveolar epithelial cells, and lung fibroblasts. Actually, TRPM2 protein was expressed in lung tissues. Of these, TRPM2 channels in epithelial cells were activated by the addition of H2O2 following a BLM pretreatment, resulting in the secretion of macrophage inflammatory protein-2 (MIP-2). The H2O2-induced activation of TRPM2 by the BLM pretreatment was blocked by the poly(ADP-ribose) polymerase (PARP) inhibitors PJ34 and 3-aminobenzamide. The accumulation of poly(ADP-ribose) in the nucleus, a marker for ADP-ribose production, was strongly induced by H2O2 following the BLM pretreatment. Furthermore, administration of PRAP inhibitors into WT mice markedly reduced recruitment of inflammatory cells and MIP-2 secretion induced by BLM instillation. These results suggest that the induction of MIP-2 secretion through the activation of TRPM2 channels in alveolar epithelial cells is an important mechanism in BLM-induced lung inflammation, and the TRPM2 activation is likely to be mediated by ADP-ribose production via PARP pathway. TRPM2 channels may be new therapeutic target for BLM-induced lung inflammation.