Identification of α-Tocopherol succinate as an RFFL-substrate interaction inhibitor inducing peripheral CFTR stabilization and apoptosis.

The E3 ubiquitin ligase RFFL is an apoptotic inhibitor highly expressed in cancers and its knockdown suppresses cancer cell growth and sensitizes to chemotherapy. RFFL also participates in peripheral protein quality control which removes the functional cell surface ΔF508-CFTR channel and reduces the efficacy of pharmaceutical therapy for cystic fibrosis (CF). Although RFFL inhibitors have therapeutic potential for both cancer and CF, they remain undiscovered. Here, a chemical array screening has identified α-tocopherol succinate (αTOS) as an RFFL ligand. NMR analysis revealed that αTOS directly binds to RFFL's substrate-binding region without affecting the E3 enzymatic activity. Consequently, αTOS inhibits the RFFL-substrate interaction, ΔF508-CFTR ubiquitination and elimination from the plasma membrane of epithelial cells, resulting in the increased functional CFTR channel. Among the α-tocopherol (αTOL) analogs we tested, only αTOS inhibited the RFFL-substrate interaction and increased the cell surface ΔF508-CFTR, depending on RFFL expression. Similarly, the unique proapoptotic effect of αTOS was dependent on RFFL expression. Thus, unlike other αTOL analogs, αTOS acts as an RFFL protein-protein interaction inhibitor which may explain its unique biological properties among αTOL analogs. Moreover, αTOS may act as a CFTR stabilizer, a novel class of drugs that extend cell surface ΔF508-CFTR lifetime.

Lung transplantation for cystic fibrosis complicated by cirrhosis: A case report.

A 16-year-old girl with a genetic diagnosis of cystic fibrosis was referred to us for consideration of lung transplantation. She had been hospitalized repeatedly for pneumonia and pneumothoraxes and her respiratory function had worsened progressively. Although she also had liver cirrhosis, she was considered a candidate for lung transplantation because her liver disease was compensated and only slowly progressive. After bilateral lung transplantation from a brain-dead donor, she developed ascites that was well controlled with diuretics. Otherwise, her post-operative course was uneventful and she was transferred to another hospital for rehabilitation 39 days after lung transplantation.

NiFe Alloy Integrated with Amorphous/Crystalline NiFe Oxide as an Electrocatalyst for Alkaline Hydrogen and Oxygen Evolution Reactions.

The rational design of efficient and low-cost electrocatalysts based on earth-abundant materials is imperative for large-scale production of hydrogen by water electrolysis. Here we present a strategy to prepare highly active catalyst materials through modifying the crystallinity of the surface/interface of strongly coupled transition metal-metal oxides. We have thermally activated the catalysts to construct amorphous/crystalline Ni-Fe oxide interfaced with a conductive Ni-Fe alloy and systematically investigated their electrocatalytic performance toward the hydrogen evolution and oxygen evolution reactions (HER and OER) in alkaline solution. It was found that the Ni-Fe/oxide material with a crystalline surface oxide phase showed remarkably superior HER activity in comparison with its amorphous or poorly crystalline counterpart. In contrast, interestingly, the amorphous/poorly crystalline oxide significantly facilitated the OER activity in comparison with the more crystalline counterpart. On one hand, the higher HER activity can be ascribed to a favorable platform for water dissociation and H-H bond formation, enabled by the unique crystalline metal/oxide structure. On the other hand, the enhanced OER catalysis on the amorphous Ni-Fe oxide surfaces can be attributed to the facile activation to form the active oxyhydroxides under OER conditions. Both are explained based on density functional theory calculations. These results thus shed light onto the role of crystallinity in the HER and OER catalysis on heterostructured Ni-Fe/oxide catalysts and provide guidance for the design of new catalysts for efficient water electrolysis.

Investigation on the Usefulness of Sulfamethoxazole Trimethoprim Combination Small Tablets in Pediatric Pharmacotherapy: A Single Center Observational Study Using a Questionnaire.

Sulfamethoxazole trimethoprim (ST) combinations are used to prevent infection in immunocompromised patients. In pediatric patients, conventional ST combination tablets (cTab) are large and granules are not preferred due to their rough and bitter taste in the mouth. Since a new formulation of smaller tablets (sTab, 1 cTab = 1-gram granules = 4 sTab) was approved, a study regarding the usability of sTab in pediatric patients was conducted. Children who started taking sTab of the ST combination at our hospital between August 2021 and August 2022 were included. Using an anonymous questionnaire, the dosage of ST combinations, the child's response (3-point visual scale: positive, neutral, or negative), preparation and administration time, and method of taking the drug were asked. Twenty-two patients (median age: 11.0 years) receiving cTab. Median (range) number of tablets per dose was 1 (0.5-1.5) tablet, and was 4 tablets (1.0-4.0) after switching to sTab. Twenty patients (median age: 5.0 years) receiving granules. Median (range) single dose was 0.75 (0.2-2.0) gram, and was 2.0 (1.0-4.0) tablets after switching to sTab. Post-dose reactions were positive in 5, neutral in 7, and negative in 10 cases for cTab, and positive in 1, neutral in 7, and negative in 12 cases for granules. After switching to sTab, 9, 13 and 0 cases, and 10, 9 and 1 cases were positive, neutral, and negative, respectively. Median preparation and administration times were decreased after switching to sTab in both cTab and granules groups. The frequency of dosage manipulations was also decreased. The switch to sTab improved acceptability, and decreased burden of administration, suggesting that sTab is a user-friendly formulation in pediatric medications.

Detection of Anti-SARS-CoV-2 Nucleocapsid and Spike Antibodies in Patients with Coronavirus Disease 2019 in Japan.

OBJECTIVES: Coronavirus Disease 2019 (COVID-19) is caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Serological testing for anti-SARS-CoV-2 nucleocapsid (N) antibodies (Abs) and anti-SARS-CoV-2 spike (S) Abs is performed to detect prior COVID-19 infection. It is still controversial which antibodies are the most sensitive and specific, and which can be detected earliest after infection. Here, we evaluated the results of serological tests of anti-SARS-CoV-2 N and S Abs in Japan. METHODS: Symptomatic COVID-19 patients (n = 84) and control patients with rheumatoid arthritis (n = 93) were recruited at Tokyo National Hospital. Anti-SARS-CoV-2 N and S Abs were measured by commercial electrochemiluminescence immunoassays. RESULTS: The fraction of patients positive for anti-SARS-CoV-2 N and S Abs was highest >14 days after symptom onset. The frequency of anti-SARS-CoV-2 S Ab positivity at this time (80.4%) tended to be slightly but not significantly lower than anti-SARS-CoV-2 N Ab positivity (84.8%). Optimized cut-off levels for anti-SARS-CoV-2 N and S Ab positivity were lower than the manufacturer's recommended cut-off levels. Using multiple linear regression analyzes with anti-SARS-CoV-2 N and S Abs, we created an Ab-index with high sensitivity. CONCLUSION: To increase the sensitivity of serological diagnostic tests for COVID-19, it is suggested that both anti-SARS-CoV-2 N and S Abs should be measured and cut-off levels decreased.

Cardiac Power Output Is Independently and Incrementally Associated With Adverse Outcomes in Heart Failure With Preserved Ejection Fraction.

BACKGROUND: Cardiac power output is a measure of cardiac performance, and its prognostic significance has been shown in heart failure (HF) with reduced ejection fraction. Patients with HF with preserved ejection fraction may have altered cardiac performance, but the prognostic relevance of cardiac power output is unknown. This study sought to determine the association between cardiac power output and clinical outcomes in HF with preserved ejection fraction and to compare its prognostic effect to other measures of cardiac performance including ventricular-arterial coupling and mechanical efficiency. METHODS: Cardiac power output normalized to left ventricular mass was assessed by echocardiography in 408 patients with HF with preserved ejection fraction. Load-independent contractility (end-systolic elastance), arterial elastance, its coupling (arterial elastance/end-systolic elastance), left ventricular global longitudinal strain, and mechanical efficiency (stroke work/pressure-volume area) were also estimated noninvasively. The primary end point was a composite of cardiovascular mortality or HF hospitalization. RESULTS: The primary composite outcome occurred in 84 patients during a median follow-up of 19.4 months. There was a dose-dependent association between cardiac power output and the composite outcomes, in which patients with the lowest tertile of cardiac power output had >3-fold risk than those with the highest tertile (hazard ratio, 3.04 [95% CI, 1.66-5.57]; P=0.0003). In a multivariable model, lower cardiac power output was independently associated with adverse outcomes (hazard ratio, 0.70 per 1 SD [95% CI, 0.49-0.97]; P=0.03). In contrast, left ventricular size, end-systolic elastance, arterial elastance, arterial elastance/end-systolic elastance ratio, and left ventricular mechanical efficiency were not associated with outcomes. Cardiac power output provided an incremental prognostic effect over the model based on clinical (age, gender, diastolic blood pressure, and atrial fibrillation) and echocardiographic markers (left atrial size, pulmonary pressures, global longitudinal strain, and the ratio of early diastolic mitral inflow velocity to early diastolic mitral annular tissue velocity; P=0.03). CONCLUSIONS: In patients with HF with preserved ejection fraction, cardiac power output was independently and incrementally associated with adverse outcomes whereas other markers of cardiac performance were not.

Diaphragmatic hernia during treatment of lung cancer harboring an EGFR mutation.

Epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) are a first-line treatment for patients with nonsmall-cell lung cancer harboring EGFR mutations. We report a 65-year-old Japanese woman with nonsmall-cell lung cancer taking an EGFR-TKI who visited the emergency department with acute nausea and vomiting. Imaging studies demonstrated an incarcerated diaphragmatic hernia. Urgent diagnostic surgery revealed a gap in the diaphragm acting as a hernial orifice, where a metastatic tumor was detected. We consider that regression of the diaphragmatic metastasis by EGFR-TKI therapy resulted in perforation of the diaphragm, causing the diaphragmatic hernia. Gastrointestinal adverse events, e.g. nausea, vomiting and diarrhea, are common during EGFR-TKI treatment. However, this case suggests that in patients with diaphragmatic metastasis, we should consider the rare possibility of diaphragmatic perforation and a subsequent hernia.

Structural modification of a novel inhibitor for mycobacterium enoyl-acyl carrier protein reductase assisted by In silico structure-based drug screening.

Background: Mycobacterium tuberculosis enoyl-acyl carrier protein reductase (mtInhA) is involved in the biosynthesis of mycolic acids, a major component of mycobacterial cell walls, and has been targeted in the development of anti-tuberculosis (TB) drugs. In our previous in silico structure-based drug screening study, we identified KES4, a novel class of mtInhA inhibitor. KES4 is composed of four ring structures (A-D-rings) and molecular dynamic simulation predicted that the D-ring is essential for the interaction with mtInhA. Methods: The structure-activity relationship study of the D-ring was attempted and aided by in silico docking simulations to improve the mtInhA inhibitory activity of KES4. A virtual chemical library of the D-ring-modified KES4 was then constructed and subjected to in silico docking simulation against mtInhA using the GOLD program. The candidate compound showing the highest GOLD score, referred to as KEN1, was synthesized, and its biological properties were compared with those of the lead compound KES4. Results: We achieved the synthesis of KEN1 and evaluated its effects on InhA activity, mycobacterial growth, and cytotoxicity. The antimycobacterial activity of KEN1 was comparable to that of the lead compound (KES4), although it exhibited superior activity in mtInhA inhibition. \. Conclusions: We obtained a KES4 derivative with high mtInhA inhibitory activity by in silico docking simulation with a chemical library consisting of a series of D-ring-modified KES4.

Efficient Large-Scale 2D Culture System for Human Induced Pluripotent Stem Cells and Differentiated Cardiomyocytes.

Cardiac regenerative therapies utilizing human induced pluripotent stem cells (hiPSCs) are hampered by ineffective large-scale culture. hiPSCs were cultured in multilayer culture plates (CPs) with active gas ventilation (AGV), resulting in stable proliferation and pluripotency. Seeding of 1 × 106 hiPSCs per layer yielded 7.2 × 108 hiPSCs in 4-layer CPs and 1.7 × 109 hiPSCs in 10-layer CPs with pluripotency. hiPSCs were sequentially differentiated into cardiomyocytes (CMs) in a two-dimensional (2D) differentiation protocol. The efficiency of cardiac differentiation using 10-layer CPs with AGV was 66%-87%. Approximately 6.2-7.0 × 108 cells (4-layer) and 1.5-2.8 × 109 cells (10-layer) were obtained with AGV. After metabolic purification with glucose- and glutamine-depleted and lactate-supplemented media, a massive amount of purified CMs was prepared. Here, we present a scalable 2D culture system using multilayer CPs with AGV for hiPSC-derived CMs, which will facilitate clinical applications for severe heart failure in the near future.

The inhibition/excitation ratio related to task-induced oscillatory modulations during a working memory task: A multtimodal-imaging study using MEG and MRS.

Detailed studies on the association between neural oscillations and the neurotransmitters gamma-aminobutyric acid (GABA) and glutamate have been performed in vitro. In addition, recent functional magnetic resonance imaging studies have characterized these neurotransmitters in task-induced deactivation processes during a working memory (WM) task. However, few studies have investigated the relationship between these neurotransmitters and task-induced oscillatory changes in the human brain. Here, using combined magnetoencephalography (MEG) and magnetic resonance spectroscopy (MRS), we investigated the modulation of GABA and glutamate + glutamine (Glx) concentrations related to task-induced oscillations in neural activity during a WM task. We first acquired resting-state MRS and MEG data from 20 healthy male volunteers using the n-back task. Time-frequency analysis was employed to determine the power induced during the encoding and retention phases in perigenual anterior cingulate cortex (pg-ACC), mid-ACC, and occipital cortex (OC). Statistical analysis showed that increased WM load was associated with task-induced oscillatory modulations (TIOMs) of the theta-gamma band relative to the zero-back condition (TIOM0B) in each volume of interest during the encoding phase of the n-back task. The task-induced oscillatory modulations in the two-back condition relative to the zero-back condition (TIOM2B-0B) were negatively correlated with the percent rate change of the correct hit rate for 2B-0B, but positively correlated with GABA/Glx. The positive correlation between TIOM2B-0B and GABA/Glx during the WM task indicates the importance of the inhibition/excitation ratio. In particular, a low inhibition/excitation ratio is essential for the efficient inhibition of irrelevant neural activity, thus producing precise task performance.

Near-infrared spectroscopic study of frontopolar activation during face-to-face conversation in major depressive disorder and bipolar disorder.

Major depressive disorder (MDD) and bipolar disorder (BD) patients show speech characteristics that vary greatly according to mood state. In a previous study, we found impaired temporal and right inferior frontal gyrus (IFG) activation in schizophrenia during face-to-face conversation; no study had, however, previously investigated mood disorders during face-to-face conversation. Here, we investigated frontal and temporal lobe activation during conversation in patients with MDD and BD. Frontal and temporal lobe activation was measured using near-infrared spectroscopy (NIRS) in 29 patients with MDD, 31 patients with BD, and 31 normal controls (NC). We compared continuous activation and rapid change of activation with talk/listen phase changes during the conversation and analyzed the correlation between these indices and clinical variables. Both the MDD and BD groups showed decreased continuous activation in the left dorsolateral prefrontal (DLPFC) and left frontopolar cortices (FPCs); they also showed decreased rapid change in bilateral FPC activation. In the MDD group, the rapid change of activation was positively correlated with Global Assessment of Functioning (GAF) scores. In the BD group, continuous activation was negatively correlated with age of onset. These results indicate that frontal activation during conversation decreases in both MDD and BD. However, both continuous activation and rapid change may reflect the pathophysiological character of MDD and BD; in particular, the reduced amount of rapid change in the right FPC may be related to impaired adaptive ability in MDD.

Protein phosphatase complex PP5/PPP2R3C dephosphorylates P-glycoprotein/ABCB1 and down-regulates the expression and function.

P-glycoprotein (P-gp)/ABCB1 is a key molecule of multidrug resistance in cancer. Protein phosphatase (PP) 2A, regulatory subunit B, gamma (PPP2R3C), which is a regulatory subunit of PP2A and PP5, was identified as a binding candidate to P-gp. Immunoprecipitation-western blotting revealed that PP5 and PPP2R3C were coprecipitated with P-gp, while PP2A was not. PP5/PPP2R3C dephosphorylated protein kinase A/protein kinase C-phosphorylation of P-gp. Knockdown of PP5 and/or PPP2R3C increased P-gp expression and lowered the sensitivity to vincristine and doxorubicin. Consequently, our results indicate that PP5/PPP2R3C negatively regulates P-gp expression and function.

Temporal lobe and inferior frontal gyrus dysfunction in patients with schizophrenia during face-to-face conversation: a near-infrared spectroscopy study.

Schizophrenia (SC) is marked by poor social-role performance and social-skill deficits that are well reflected in daily conversation. Although the mechanism underlying these impairments has been investigated by functional neuroimaging, technical limitations have prevented the investigation of brain activation during conversation in typical clinical situations. To fill this research gap, this study investigated and compared frontal and temporal lobe activation in patients with SC during face-to-face conversation. Frontal and temporal lobe activation in 29 patients and 31 normal controls (NC) (n = 60) were measured during 180-s conversation periods by using near-infrared spectroscopy (NIRS). The grand average values of oxyhemoglobin concentration ([oxy-Hb]) changes during task performance were analyzed to determine their correlation with clinical variables and Positive and Negative Syndrome Scale (PANSS) subscores. Compared to NCs, patients with SC exhibited decreased performance in the conversation task and decreased activation in both the temporal lobes and the right inferior frontal gyrus (IFG) during task performance, as indicated by the grand average of [oxy-Hb] changes. The decreased activation in the left temporal lobe was negatively correlated with the PANSS disorganization and negative symptoms subscores and that in the right IFG was negatively correlated with illness duration, PANSS disorganization, and negative symptom subscores. These findings indicate that brain dysfunction in SC during conversation is related to functional deficits in both the temporal lobes and the right IFG and manifests primarily in the form of disorganized thinking and negative symptomatology.

The role of G proteins in thromospondin-1-induced vascular smooth muscle cell migration.

PURPOSE: Thrombospondin-1 (TSP-1), which is a matricellular glycoprotein associated with chemotaxis of vascular smooth muscle cells (VSMCs), is relevant to the development of arterial lesions. Evidence suggests that TSP-1 receptors are linked to guanosine triphosphate-binding proteins (G proteins). The purpose of this study was to determine the role of G proteins in TSP-1-induced VSMC chemotaxis and whether this pathway was associated with extracellular signal-regulated kinase 1/2 (ERK) or p38 kinase activation (downstream pathways associated with VSMC chemotaxis). METHODS: In all studies, quiescent VSMCs were preincubated either with serum-free medium, cholera toxin, pertussis toxin, forskolin, or 3-isobutyl-1-methylxanthine. Using a microchemotaxis chamber, preincubated VSMCs were exposed to TSP-1 or serum-free medium. Migrated VSMCs were recorded as cells/5 fields (400x) and analyzed by paired t-test. To evaluate the effect of G proteins on TSP-1-induced ERK or p38 activation, preincubated VSMCs were exposed to serum-free medium or TSP-1 and analyzed by Western immunoblotting. For measurement of intracellular cyclic adenosine monophosphate (cAMP) levels, enzyme-linked immunosorbant assay was performed on preincubated VSMCs exposed to serum-free medium or TSP-1. RESULTS: Although pertussis toxin attenuated TSP-1-induced chemotaxis, cholera toxin abolished TSP-1-induced chemotaxis. Cholera toxin, but not pertussis toxin, inhibited both ERK and p38 activation. The cAMP stimulators forskolin and IBMX abolished TSP-1-induced chemotaxis and ERK and p38 activation. Although no changes were observed in cAMP levels in VSMCs treated with serum-free medium, TSP-1, or pertussis toxin, cholera toxin alone significantly increased cAMP levels. CONCLUSION: G(s) protein signaling inhibits TSP-1-induced VSMC chemotaxis by increasing the levels of cAMP. G(i) signaling is involved in the mechanism of TSP-1 stimulated chemotaxis and warrants additional study. Agents that increase cAMP levels may be beneficial in reducing TSP-1-induced chemotaxis in response to vascular injury.

The use of multi-detector row computed tomography (MDCT) as an alternative to specimen preparation for anatomical instruction.

The purpose of the study reported here was to establish a method of teaching veterinary anatomy, including radiologic anatomy, for clinical practice using computer-aided diagnosis (CAD). Two clinically healthy dogs and three cats were scanned using multi-detector row computed tomography (MDCT). Images were made by means of imaging processing software. At the workstation, by observing the transverse, dorsal-plane, or sagittal sections and three-dimensional (3D) images simultaneously, it is much easier to understand the 3D anatomical structure. With this educational support system, anatomical figures can be explained using living animals instead of specimens. In addition, clinical representative examples can be used to show anatomical disorders to students. Veterinary students (N = 62) who filled out a questionnaire evaluating how the method aided their understanding of both experimental study and clinical examples gave it a score of 88.2 +/- 20.6 (Mean +/- SD) on a visual analog scale. This system can enhance veterinary students' understanding and interest in anatomy and can enable us to offer them a quality veterinary medical education. We concluded that CAD is a useful new option not only for clinical service but also for veterinary education.

Extra-abdominal chondrolipoma in a dog.

A 10-year-old male maltise dog was presented for evaluation of a gradual abdominal enlargement, which had developed over 2 years. (On the radiograph of the abdomen, a significant distention of the abdomen with a large accumulation of fat could be seen.) The mass effect was noticeable because of the rightward displacement of the viscera. There was multifocal amorphous mineralization, mostly in the right abdomen. Computed tomography (CT) identified a 13 x 13 x 10 cm mass, most likely an intra-abdominal lipoma, by which the digestive tract had been displaced in the abdomen. It was only during surgery, however, that the mass was found to be located between the peritoneum and transversus abdominis muscles and the rectus abdominus muscle. It was completely resected. Histopathological examination revealed that the mass was chondrolipoma. The amorphous radiopaque lesion on the images corresponded to cartilaginous tissues with calcification. There has been no recurrence for 10 months. This is the first report of an extra-abdominal chondrolipoma in dogs. Cartilaginous metaplasia of lipoma, located in the abdominal area, should be included in the differential diagnosis for multifocal amorphous mineralization.

Change in liver and plasma ceramides during D-galactosamine-induced acute hepatic injury by LC-MS/MS.

In fulminant hepatic failure, various toxins causing multi-organ failure increase in plasma. As a novel toxin, ceramide, a well-studied lipid mediator of apoptosis, levels were determined by LC-MS/MS in the liver and plasma of D-galactosamine-intoxicated rats. 18 and 24h after intraperitoneal administration of D-galactosamine (1g/kg body weight) to rats, fulminant hepatic failure occurred as evidenced by a severe elevation in plasma GOT and GPT. The liver concentration of minor ceramide components (C18:0, C20:0, C22:1, C22:0, and C24:2) increased significantly compared to that in the control group that was given saline. The plasma concentration of major ceramides (C24:0, C24:1, C16:0, C22:0, C22:1, and C18:0) increased 24h after administration of D-galactosamine and the total ceramide concentration was also increased to 3.6 times that in the control. In conclusion, the increased concentrations of ceramides in plasma during fulminant hepatic failure may be one of important toxins causing damage in other organs including the brain and kidney.

Cyclic strain stimulates early growth response gene product 1-mediated expression of membrane type 1 matrix metalloproteinase in endothelium.

SUMMARY: Matrix metalloproteinases (MMPs) are hypothesized to be involved in the processes of endothelial cell (EC) migration and matrix remodeling during angiogenesis. Although hemodynamic forces (such as blood pressure, wall tension, and shear stress) are considered to be strong stimuli for angiogenesis, the role of hemodynamic forces on the regulation of MMPs including membrane type 1 matrix metalloproteinase (MT1-MMP) has not been fully elucidated. To study this, rat microvascular EC were exposed to 60 cycles/minute of 24% maximum strain for up to 24 hours. MT1-MMP mRNA and protein increased in a time-dependent manner through 24 hours of exposure to cyclic strain. Cyclic strain induced early growth response gene product (Egr-1) mRNA and protein within 1 hour. A specific nucleoprotein complex was formed when an oligonucleotide containing binding sites for Sp1 and Egr-1 was incubated with nuclear extracts from EC exposed to 1 hour of cyclic strain. Antibodies to Egr-1 completely supershifted this complex. Increased binding of Egr-1 by cyclic strain to the MT1-MMP promoter correlated with enhanced transcriptional activity. These results suggest that cyclic strain up-regulates the Egr-1-mediated expression of MT1-MMP in rat microvascular EC, emphasizing the importance of hemodynamic forces in the regulation of MT1-MMP in vivo.