Sensitivity assessment of diphacinone by pharmacokinetic analysis in invasive black rats in the Bonin (Ogasawara) Archipelago, Japan.

The Bonin Archipelago is a United Nations Educational, Scientific and Cultural Organization's World Natural Heritage Site in Japan with a unique ecosystem; however, the invasive rodents preying on endemic species have been a significant concern. The anticoagulant rodenticide, diphacinone, sprayed by the Ministry of the Environment, has succeeded; however, its repeated use leads to rodenticide resistance. This study evaluated the sensitivity by in vivo pharmacokinetics/pharmacodynamics (PK/PD) analysis and physiologically-based pharmacokinetic modeling to diphacinone in black rats (Rattus rattus) captured on the Bonin Archipelago in February 2022. The Bonin rats exhibited prolonged coagulation time after diphacinone administration. They recovered earlier than susceptible black rats, indicating that Bonin rats were less susceptible, though there were no genetic mutations in Vkorc1, the target enzyme of diphacinone. After the administration of diphacinone, hepatic expression levels of Fsp1, identified as the vitamin K reductase, was decreased, however, the Bonin rats exhibited the most minor suppression. The PK analysis showed that the excretion capacity of the Bonin rats was lower than that of the resistant black rats. In the PBPK modeling, the resistant black rats showed higher clearance than the Bonin and susceptible black rats due to high hepatic metabolic capacity. The Bonin rats demonstrated slow absorption and relatively low clearance. This study highlighted the reduced rodenticide-sensitive tendency of wild black rats in the Bonin Archipelago at an in vivo phenotype level. At the same time, they do not have known rodenticide resistance mechanisms, such as hepatic metabolic enhancement or Vkorc1 mutations. It is crucial to monitor the biological levels to evaluate rodenticide sensitivity accurately.

Verification of image quality improvement of low-count bone scintigraphy using deep learning.

To improve image quality for low-count bone scintigraphy using deep learning and evaluate their clinical applicability. Six hundred patients (training, 500; validation, 50; evaluation, 50) were included in this study. Low-count original images (75%, 50%, 25%, 10%, and 5% counts) were generated from reference images (100% counts) using Poisson resampling. Output (DL-filtered) images were obtained after training with U-Net using reference images as teacher data. Gaussian-filtered images were generated for comparison. Peak signal-to-noise ratio (PSNR) and structural similarity (SSIM) to the reference image were calculated to determine image quality. Artificial neural network (ANN) value, bone scan index (BSI), and number of hotspots (Hs) were computed using BONENAVI analysis to assess diagnostic performance. Accuracy of bone metastasis detection and area under the curve (AUC) were calculated. PSNR and SSIM for DL-filtered images were highest in all count percentages. BONENAVI analysis values for DL-filtered images did not differ significantly, regardless of the presence or absence of bone metastases. BONENAVI analysis values for original and Gaussian-filtered images differed significantly at ≦25% counts in patients without bone metastases. In patients with bone metastases, BSI and Hs for original and Gaussian-filtered images differed significantly at ≦10% counts, whereas ANN values did not. The accuracy of bone metastasis detection was highest for DL-filtered images in all count percentages; the AUC did not differ significantly. The deep learning method improved image quality and bone metastasis detection accuracy for low-count bone scintigraphy, suggesting its clinical applicability.

AMPK/FOXO3a Pathway Increases Activity and/or Expression of ATM, DNA-PKcs, Src, EGFR, PDK1, and SOD2 and Induces Radioresistance under Nutrient Starvation.

Most solid tumors contain hypoxic and nutrient-deprived microenvironments. The cancer cells in these microenvironments have been reported to exhibit radioresistance. We have previously reported that nutrient starvation increases the expression and/or activity of ATM and DNA-PKcs, which are involved in the repair of DNA double-strand breaks induced by ionizing radiation. In the present study, to elucidate the molecular mechanisms underlying these phenomena, we investigated the roles of AMPK and FOXO3a, which play key roles in the cellular response to nutrient starvation. Nutrient starvation increased clonogenic cell survival after irradiation and increased the activity and/or expression of AMPKα, FOXO3a, ATM, DNA-PKcs, Src, EGFR, PDK1, and SOD2 in MDA-MB-231 cells. Knockdown of AMPKα using siRNA suppressed the activity and/or expression of FOXO3a, ATM, DNA-PKcs, Src, EGFR, PDK1, and SOD2 under nutrient starvation. Knockdown of FOXO3a using siRNA suppressed the activity and/or expression of AMPKα, ATM, DNA-PKcs, FOXO3a, Src, EGFR, PDK1, and SOD2 under nutrient starvation. Nutrient starvation decreased the incidence of apoptosis after 8 Gy irradiation. Knockdown of FOXO3a increased the incidence of apoptosis after irradiation under nutrient starvation. AMPK and FOXO3a appear to be key molecules that induce radioresistance under nutrient starvation and may serve as targets for radiosensitization.

Emphysematous pyelonephritis with ST elevation accompanied by reciprocal changes mimicking acute coronary syndrome.

Patients with infectious diseases including sepsis can develop ST segment changes on an electrocardiogram (ECG) in the absence of coronary artery disease. However, ST elevation with "reciprocal ST segment depression (RSTD)", which is recognized as a specific finding for ST-elevated myocardial infarction, is rare in such patients. Although a small number of cases have reported ST-segment elevation in gastritis, cholecystitis, and sepsis, regardless of coronary artery disease, none presented with reciprocal changes. Here, we describe a rare case of a patient with emphysematous pyelonephritis complicating septic shock who developed ST elevation accompanied by reciprocal changes with no coronary occlusion. Emergency physicians should consider the possibility of acute coronary syndrome mimicking, and choose non-invasive diagnostic procedures when investigating the causes of ECG abnormalities associated with critically ill patients.

Delivery accuracy of VMAT on two beam-matched linacs provided by accelerated go live service.

INTRODUCTION: Dosimetric accuracy is critical when a patient treated with volumetric modulated arc therapy (VMAT) is transferred to another beam-matched linac. To evaluate the performance of Accelerated Go Live (AGL) service, the measured beam characteristics and patient specific quality assurance (QA) results between two AGL-matched linacs were compared. MATERIALS AND METHODS: Two VersaHD linacs were installed using the AGL service. After the installation, the beam data such as percentage depth dose (PDD), lateral profiles and output factors for all photon beams were measured. Relative doses were also measured as a function of the multi-leaf collimator (MLC) leaf gap width. Subsequently, VMAT plans were created for prostate, pelvis, head and neck, liver, lung cancers and multiple brain metastases. Dose distributions and point doses were measured by multi-dimensional detectors and ionization chambers for patient specific quality assurance, and comparisons were made between the two linacs. RESULTS: Dose differences in PDDs were all within ± 1% except the entrance region, and the averaged gamma indices of the lateral profiles were within 0.3. The differences in doses as a function of the MLC leaf gap width between the two linacs were within ±0.5%. For all the plans, gamma passing rates were all higher than 95% with criteria of 2%/2 mm. The average and the SD of dose differences on the multi-dimensional detector between both measurements was 0.06 ± 2.12%, and the average of point dose differences was -0.03 ± 0.33%. CONCLUSION: We have evaluated the AGL performance in the context of beam characteristics and patient specific QA. It was demonstrated that the AGL service provides an accurate VMAT treatment reproducibility for many tumor sites with gamma pass rates greater than 95% under criteria of 2%/2 mm.

Anesthetic management of pheochromocytoma and paraganglioma for patients with Fontan circulation: a case series.

BACKGROUND: Anesthetic management of pheochromocytoma and paraganglioma with Fontan circulation is challenging for physicians, with attention to cardiovascular physiology. CASE PRESENTATION: We performed anesthetic management for pheochromocytoma and paraganglioma in three patients with Fontan circulation. We maintained intraoperative central venous pressure at preoperative level under fluid infusion and administrating nitric oxide to decrease pulmonary arterial resistance. We administered noradrenaline or vasopressin if low blood pressure was present despite adequate central venous pressure. Although noradrenaline is prevalent for the case of noradrenaline-secreting tumor especially after resection, we could maintain blood pressure to administrate vasopressin without increasing central venous pressure. Retroperitoneal laparoscopic approach which could avoid intra-abdominal adhesions might be selectable as case 3. CONCLUSIONS: Sophisticated management is required for pheochromocytoma and paraganglioma with Fontan circulation.

Non-volatile optical memory based on cooperative orientation switching: improvement of recording speed and contrast by utilizing out-of-plane orientation mode.

Herein, we report a novel strategy toward non-volatile optical memory with high-contrast, high-speed recording, and non-destructive readout capability based on the cooperative out-of-plane orientation of a fluorescent dye doped into azobenzene liquid crystalline polymer film. By employing the out-of-plane orientation switching upon irradiation with UV light and thermal heating, high-contrast turn-on fluorescence switching was successfully achieved and the optical recording was demonstrated with non-destructive fluorescence readout capability. Furthermore, the recording speed and the fluorescence on/off contrast in the present system were dramatically improved compared to the previous in-plane orientation mode.

Sequence divergence and retrotransposon insertion underlie interspecific epigenetic differences in primates.

Changes in the epigenome can affect the phenotype without the presence of changes in the genomic sequence. Given the high identity of the human and chimpanzee genome sequences, a substantial portion of their phenotypic divergence likely arises from epigenomic differences between the two species. In this study, the transcriptome and epigenome were determined for induced pluripotent stem cells (iPSCs) generated from human and chimpanzee individuals. The transcriptome and epigenomes for trimethylated histone H3 at lysine-4 (H3K4me3) and lysine-27 (H3K27me3) showed high levels of similarity between the two species. However, there were some differences in histone modifications. Although such regions, in general, did not show significant enrichment of interspecies nucleotide variations, gains in binding motifs for pluripotency-related transcription factors, especially POU5F1 and SOX2, were frequently found in species-specific H3K4me3 regions. We also revealed that species-specific insertions of retrotransposons, including the LTR5_Hs subfamily in human and a newly identified LTR5_Pt subfamily in chimpanzee, created species-specific H3K4me3 regions associated with increased expression of nearby genes. Human iPSCs have more species-specific H3K27me3 regions, resulting in more abundant bivalent domains. Only a limited number of these species-specific H3K4me3 and H3K27me3 regions overlap with species-biased enhancers in cranial neural crest cells, suggesting that differences in the epigenetic state of developmental enhancers appear late in development. Therefore, iPSCs serve as a suitable starting material for studying evolutionary changes in epigenome dynamics during development.

Supplemental Clostridium butyricum MIYAIRI 588 Affects Intestinal Bacterial Composition of Finishing Pigs.

Animal gastrointestinal tracts are populated by highly diverse and complex microbiotas. The gut microbiota influences the bioavailability of dietary components and is closely associated with physiological processes in the host. Clostridium butyricum reportedly improves growth performance and affects the gut microbiota and immune functions in post-weaning piglets. However, the effects of C. butyricum on finishing pigs remain unclear. Therefore, we herein investigated the effects of C. butyricum MIYAIRI 588 (CBM588) on the gut microbiota of finishing pigs. 16S rRNA gene sequencing was performed using fecal samples and ileal, cecal, and colonic contents collected after slaughtering. The α-diversity of the small intestinal microbiota was lower than that of the large intestinal microbiota, whereas ß-diversity showed different patterns depending on sample collection sites. The administration of CBM588 did not significantly affect the α- or ß-diversity of the microbiotas of fecal and intestinal content samples regardless of the collection site. However, a linear discriminant ana-lysis Effect Size revealed that the relative abundance of Lactobacillaceae at the family level, Bifidobacterium at the order level, and Lactobacillus ruminis and Bifidobacterium pseudolongum at the species level were higher in the fecal samples and cecal and colonic contents of the treatment group than in those of the control group. Therefore, the administration of CBM588 to finishing pigs affected the composition of the gut microbiota and increased the abundance of bacteria that are beneficial to the host. These results provide important insights into the effects of probiotic administration on relatively stable gut microbial ecosystems.

Toxicokinetic analysis of the anticoagulant rodenticides warfarin & diphacinone in Egyptian fruit bats (Rousettus aegyptiacus) as a comparative sensitivity assessment for Bonin fruit bats (Pteropus pselaphon).

Anticoagulant rodenticides have been widely used to eliminate wild rodents, which as invasive species on remote islands can disturb ecosystems. Since rodenticides can cause wildlife poisoning, it is necessary to evaluate the sensitivity of local mammals and birds to the poisons to ensure the rodenticides are used effectively. The Bonin Islands are an archipelago located 1000 km southeast of the Japanese mainland and are famous for the unique ecosystems. Here the first-generation anticoagulant rodenticide diphacinone has been used against introduced black rats (Rattus rattus). The only land mammal native to the archipelago is the Bonin fruit bat (Pteropus pselaphon), but little is known regarding its sensitivity to rodenticides. In this study, the Egyptian fruit bats (Rousettus aegyptiacus) was used as a model animal for in vivo pharmacokinetics and pharmacodynamics analysis and in vitro enzyme kinetics using their hepatic microsomal fractions. The structure of vitamin K epoxide reductase (VKORC1), the target protein of the rodenticide in the Bonin fruit bat, was predicted from its genome and its binding affinity to rodenticides was evaluated. The Egyptian fruit bats excreted diphacinone slowly and showed similar sensitivity to rats. In contrast, they excreted warfarin, another first-generation rodenticide, faster than rats and recovered from the toxic effect faster. An in silico binding study also indicated that the VKORC1 of fruit bats is relatively tolerant to warfarin, but binds strongly to diphacinone. These results suggest that even chemicals with the same mode of action display different sensitivities in different species: fruit bat species are relatively resistant to warfarin, but vulnerable to diphacinone.

Complete atrioventricular block under epidural ropivacaine infusion in a patient with first-degree atrioventricular block and myasthenia gravis: a case report.

BACKGROUND: First-degree atrioventricular block (AVB) may lead to complete AVB. Herein, we present a case of a complete AVB under thoracic epidural catheter infusion of ropivacaine with fentanyl in a patient with first-degree AVB and myasthenia gravis. CASE PRESENTATION: A 74-year-old woman with first-degree AVB underwent thymectomy for myasthenia gravis. Continuous thoracic epidural catheter infusion of 0.2% ropivacaine with fentanyl was initiated at 15 min before the end of the surgery. At 9 h postoperatively, the electrocardiogram showed a 10-s-long pause due to complete AVB. Thus, a temporary pacemaker was implanted, and at 19 h postoperatively on postoperative day 1, cardiac pacing was initiated and lasted approximately 30 s. After catheter removal, she had no further episodes of complete AVB. CONCLUSION: First-degree AVB may lead to complete AVB under the influence of thoracic epidural infusion of ropivacaine in patients with myasthenia gravis.

AMPK increases expression of ATM through transcriptional factor Sp1 and induces radioresistance under severe hypoxia in glioblastoma cell lines.

We have previously reported that severe hypoxia increases expression and activity of the DNA damage sensor ATM by activation of the key energy sensor AMPK. Here, to elucidate molecular mechanisms underlying increased expression and activity of ATM by AMPK under severe hypoxia, we investigated roles of transcriptional factors Sp1 and FoxO3a using human glioblastoma cell lines T98G and A172. Severe hypoxia increased expression of ATM, AMPKα and Sp1 but not that of FoxO3a. Knockdown of AMPKα suppressed expression of ATM and Sp1 and suppressed cellular radioresistance under severe hypoxia without affecting cell cycle distribution. Knockdown of Sp1 suppressed expression of ATM. These results suggest that increased expression and activity of AMPK under severe hypoxia induce cellular radioresistance through AMPK/Sp1/ATM pathway.

Development of attenuation correction methods using deep learning in brain-perfusion single-photon emission computed tomography.

PURPOSE: Computed tomography (CT)-based attenuation correction (CTAC) in single-photon emission computed tomography (SPECT) is highly accurate, but it requires hybrid SPECT/CT instruments and additional radiation exposure. To obtain attenuation correction (AC) without the need for additional CT images, a deep learning method was used to generate pseudo-CT images has previously been reported, but it is limited because of cross-modality transformation, resulting in misalignment and modality-specific artifacts. This study aimed to develop a deep learning-based approach using non-attenuation-corrected (NAC) images and CTAC-based images for training to yield AC images in brain-perfusion SPECT. This study also investigated whether the proposed approach is superior to conventional Chang's AC (ChangAC). METHODS: In total, 236 patients who underwent brain-perfusion SPECT were randomly divided into two groups: the training group (189 patients; 80%) and the test group (47 patients; 20%). Two models were constructed using Autoencoder (AutoencoderAC) and U-Net (U-NetAC), respectively. ChangAC, AutoencoderAC, and U-NetAC approaches were compared with CTAC using qualitative analysis (visual evaluation) and quantitative analysis (normalized mean squared error [NMSE] and the percentage error in each brain region). Statistical analyses were performed using the Wilcoxon signed-rank sum test and Bland-Altman analysis. RESULTS: U-NetAC had the highest visual evaluation score. The NMSE results for the U-NetAC were the lowest, followed by AutoencoderAC and ChangAC (P < 0.001). Bland-Altman analysis showed a fixed bias for ChangAC and AutoencoderAC and a proportional bias for ChangAC. ChangAC underestimated counts by 30-40% in all brain regions. AutoencoderAC and U-NetAC produced mean errors of <1% and maximum errors of 3%, respectively. CONCLUSION: New deep learning-based AC methods for AutoencoderAC and U-NetAC were developed. Their accuracy was higher than that obtained by ChangAC. U-NetAC exhibited higher qualitative and quantitative accuracy than AutoencoderAC. We generated highly accurate AC images directly from NAC images without the need for intermediate pseudo-CT images. To verify our models' generalizability, external validation is required.

Surgical treatment for breast cancer in a patient with erythropoietic protoporphyria and photosensitivity: a case report.

BACKGROUND: Erythropoietic protoporphyria (EPP) is a rare disorder of heme synthesis. Patients with EPP mainly show symptoms of photosensitivity, but approximately 20% of EPPs are associated with the liver-related complications. We report a case of breast cancer in a 48-year-old female patient with EPP in whom meticulous perioperative management was required in order to avoid complications resulting from this disease. CASE PRESENTATION: The patient was diagnosed with EPP at the age of 33 and had a rich family history of the disease. For right breast cancer initially considered as TisN0M0 (Stage 0), the right mastectomy and sentinel lymph node biopsy were performed, while the final stage was pT1bN0M0, pStage I. In the perioperative period, we limited the drug use and monitored light wavelength measurements. Besides, we covered surgical lights, headlights, and laryngoscope's light with a special polyimide film that filtered the wavelength of light causing dermal photosensitivity. After the surgery, any emerging complications were closely monitored. CONCLUSIONS: The surgery, internal medicine, anesthesiology, and operation departments undertook all possible measures through close cooperation to ensure a safe surgery for the patient with a rare condition.

Application of peptides with an affinity for phospholipid membranes during the automated purification of extracellular vesicles.

Extracellular vesicles (EVs), such as exosomes, have garnered increasing interest because of their potential clinical applications that range from diagnostics to therapeutics. The development of an automated and reproducible EV purification platform would therefore aid the introduction of EV biomarkers and therapies into the clinic. Here, we demonstrate that K8- as well as K-16 peptides (containing 8 and 16 lysine residues with dissociation constants of 102 nM and 11.6 nM for phosphatidylserine, respectively) immobilized on magnetic beads can capture small EVs (< 0.2 µm) from culture supernatants of MCF7 human breast cancer cells. Importantly, the bound EVs could be dissociated from the beads under mild conditions (e.g. 0.5 M NaCl), and the isolated EVs had the typical shapes of EVs under SEM and TEM with a mean particle size of 99 nm. Using the peptide-immobilized beads, we adapted a pre-existing bench top instrument for magnetic separation to perform automated EV purification with higher purity and yield than that obtained using the standard ultracentrifugation method.

DNA-PKcs is activated under nutrient starvation and activates Akt, MST1, FoxO3a, and NDR1.

BACKGROUND: Presence of unperfused regions containing cells under hypoxia and nutrient starvation; contributes to radioresistance in solid human tumors. We have previously reported that cultured cells; under nutrient starvation show resistance to ionizing radiation compare with cells under normal; condition, and that nutrient starvation increases ATM activity, which causes cellular resistance to; ionizing radiation (Murata et al., BBRC2018). For further investigation of molecular mechanisms; underlying radioresistance of cells under nutrient starvation, effects of nutrient starvation on activity; of DNA-PKcs have been investigated because both DNA-PKcs and ATM belong to the PIKK family; and are required for DNA DSBs repair. In addition to DNA-PKcs, effects of nutrient starvation on; activities of FoxO3a and its regulators Akt, MST1 and AMPK have been investigated because FoxO3a; mediates cellular responses to stress and is activated under nutrient starvation. METHODS: A human glioblastoma cell line, T98G was used to examine the effects of nutrient starvation on activities and expression of DNA-PKcs, Akt, MST1, FoxO3a, NDR1, and AMPK. To elucidate; signal transduction pathways for FoxO3a activation under nutrient starvation, we examined effects of; specific inhibitors or siRNA for DNA-PKcs or Akt on activities and expression of MST1, FoxO3, NDR1, andAMPK. RESULTS: Under nutrient starvation, phosphorylations of DNA-PKcs at Ser2056, Akt at Ser473, MST at Thr183, FoxO3a at Ser413, NDR1 at Ser281 and Thr282, and AMPK at Thr172 were increased, which suggests their activation. Nutrient starvation did not affect expression of DNA-PKcs, Akt, MST1, or NDR1, with decreased expression of FoxO3a and increased expression of AMPK. Inhibition; of DNA-PK suppressed phosphorylation of Akt under nutrient starvation. Inhibition of DNA-PK or; Akt suppressed phosphorylations of MST1, FoxO3a, and NDR1 under nutrient starvation, which; suggests DNA-PKcs and Akt activate MST1, FoxO3a, and NDR1. Inhibition of DNA-PK did not; suppress phosphorylation ofAMPK under nutrient starvation. CONCLUSION: Our data suggest that DN-PKcs is activated under nutrient starvation and activates AktMST1, FoxO3a, and NDR1.

Severe hypoxia increases expression of ATM and DNA-PKcs and it increases their activities through Src and AMPK signaling pathways.

BACKGROUND: Solid tumors often contain hypoxic regions because an abnormal and inefficient tumor vasculature is unable to supply sufficient oxygen. Tissue hypoxia is generally defined as a low oxygen concentration of less than 2%. It is well known that tumor cells under severe hypoxia, where oxygen concentration is less than 0.1%, show radioresistance. It has been reported that cells under severe hypoxia show different responses from those under mild hypoxia, where oxygen concentration is 0.5-2.0%. In the present study, we investigated the effects of severe hypoxia on expression and activities of ATM and DNA-dependent protein kinase catalytic subunit (DNA-PKcs), both of which regulate DNA double-strand breaks (DSBs) repair and radiation sensitivity. Signaling pathways for increasing expression and activities of ATM and DNA-PKcs under severe hypoxia were also investigated. METHODS: SV40-transformed human fibroblast cell lines, LM217 and LM205, and normal human dermal fibroblasts (NHDF) were used. Cells were cultured at an oxygen concentration of less than 0.05% for 12 or 24 h. Activities and/or expression of ATM, DNA-PKcs, Src, Caveolin-1, EGFR, HIF-1α, PDK1, Akt, AMPKα, and mTOR were estimated by Western blot analyses. RESULTS: Severe hypoxia increased expression and activities of ATM, DNA-PKcs, Src, Caveolin-1, EGFR, PDK1, Akt, and AMPKα, and decreased expression and activity of mTOR. A specific Src inhibitor, PP2 suppressed activation of ATM, DNA-PKcs, Caveolin-1, EGFR, and Akt under severe hypoxia. Treatment with siRNA for AMPKα suppressed activation of ATM and DNA-PKcs and increase of ATM expression under severe hypoxia. CONCLUSION: Our data show that severe hypoxia increases activities of ATM and DNA-PKcs through Src and AMPK signaling pathways, and that activation of AMPK under hypoxia causes increase of ATM expression. Since ATM and DNA-PKcs play important roles in DSBs repair induced by ionizing radiation, those data provide novel insights on the molecular mechanism of the cellular radioresistance under severe hypoxia.

Knockdown of AMPKα decreases ATM expression and increases radiosensitivity under hypoxia and nutrient starvation in an SV40-transformed human fibroblast cell line, LM217.

BACKGROUND: Presence of unperfused regions containing cells under hypoxia and nutrient starvation contributes to radioresistance in solid human tumors. It is well known that hypoxia causes cellular radioresistance, but little is known about the effects of nutrient starvation on radiosensitivity. We have reported that nutrient starvation induced decrease of mTORC1 activity and decrease of radiosensitivity in an SV40-transformed human fibroblast cell line, LM217, and that nutrient starvation induced increase of mTORC1 activity and increase of radiosensitivity in human liver cancer cell lines, HepG2 and HuH6 (Murata et al., BBRC 2015). Knockdown of mTOR using small interfering RNA (siRNA) for mTOR suppressed radiosensitivity under nutrient starvation alone in HepG2 cells, which suggests that mTORC1 pathway regulates radiosensitivity under nutrient starvation alone. In the present study, effects of hypoxia and nutrient starvation on radiosensitivity were investigated using the same cell lines. METHODS: LM217 and HepG2 cells were used to examine the effects of hypoxia and nutrient starvation on cellular radiosensitivity, mTORC1 pathway including AMPK, ATM, and HIF-1α, which are known as regulators of mTORC1 activity, and glycogen storage, which is induced by HIF-1 and HIF-2 under hypoxia and promotes cell survival. RESULTS: Under hypoxia and nutrient starvation, AMPK activity and ATM expression were increased in LM217 cells and decreased in HepG2 cells compared with AMPK activity under nutrient starvation alone or ATM expression under hypoxia alone. Under hypoxia and nutrient starvation, radiosensitivity was decreased in LM217 cells and increased in HepG2 cells compared with radiosensitivity under hypoxia alone. Under hypoxia and nutrient starvation, knockdown of AMPK decreased ATM activity and increased radiation sensitivity in LM217 cells. In both cell lines, mTORC1 activity was decreased under hypoxia and nutrient starvation. Under hypoxia alone, knockdown of mTOR slightly increased ATM expression but did not affect radiosensitivity in LM217. Under hypoxia and nutrient starvation, HIF-1α expression was suppressed and glycogen storage was reduced. CONCLUSION: Our data suggest that AMPK regulates ATM expression and partially regulates radiosensitivity under hypoxia and nutrient starvation. The molecular mechanism underlying the induction of ATM expression by AMPK remains to be elucidated.

High-resolution structure discloses the potential for allosteric regulation of mitogen-activated protein kinase kinase 7.

Mitogen-activated protein kinase kinase 7 (MAP2K7) regulates stress and inflammatory responses, and is an attractive drug discovery target for several diseases including arthritis and cardiac hypertrophy. Intracellular proteins such as MAP2K7 are prone to aggregation due to cysteine-driven oxidation in in vitro experiments. MAP2K7 instability due to the four free cysteine residues on the molecular surface abrogated the crystal growth and led to a low-resolution structure with large residual errors. To acquire a higher resolution structure for promoting rational drug discovery, we explored stable mutants of MAP2K7 by replacing the surface cysteine residues, Cys147, Cys218, Cys276 and Cys296. Single-site mutations, except for Cys147, maintained the specific activity and increased the protein yield, while all the multi-site mutations massively reduced the activity. The C218S mutation drastically augmented the protein production and crystallographic resolution. Furthermore, the C218S crystals grown under microgravity in a space environment yielded a 1.3 Å resolution structure, providing novel insights for drug discovery: the precisely assigned water molecules in the active site, the double conformations in the flexible region and the C-terminal extension bound to the N-terminal region of the adjacent molecules. The latter insight is likely to promote the production of allosteric MAP2K7 inhibitors.

Inclusion of gaming disorder criteria in ICD-11: A clinical perspective in favor.

Data from a specialist treatment facility for Internet addiction (IA) in Japan showed that (a) the vast majority of treatment seekers are addicted to online games, (b) their symptoms are often quite severe, and (c) there is a significant demand for IA treatment. In addition, systemic obstacles to the delivery of medical services in Japan exist due to the exclusion of IA criteria from ICD-10. Consequently, the inclusion of GD criteria in ICD-11 will almost certainly increase the capacity and quality of treatment through advances in research and possible changes in national medical systems to meet treatment demand.