Effect of Brønsted Acids on the Activation of Mixed Anhydride/Mixed Carbonic Anhydride and C-Terminal-Free N-Methylated Peptide Synthesis in a Micro-Flow Reactor.

Amidations employing mixed (carbonic) anhydrides have long been favoured in peptide synthesis because of their cost-effectiveness and less waste generation. Despite their long history, no study has compared the effects of additives on the activation of mixed anhydrides and carbonic anhydrides. In this study, we investigated the amidation of mixed (carbonic) anhydride in the presence of a base and/or Brønsted acids. The use of NMI·HCl significantly improved the conversion of the mixed carbonic anhydride, while expediting nucleophilic attacks on the desired carbonyl group. In contrast, in the case of mixed carbonic anhydrides, neither the conversion nor the desired nucleophilic attack improved significantly. We developed a C-terminus-free N-methylated peptide synthesis method using mixed carbonic anhydrides in a micro-flow reactor. Fourteen N-alkylated peptides were synthesized in moderate to high yields (55-99%) without severe racemization (<1%). Additionally, a significant enhancement in the amidation between mixed carbonic anhydrides and bis-TMS-protected N-methyl amino acids with the inclusion of NMI·HCl was observed for the first time. In addition, we observed unexpected C-terminal epimerization of the C-terminus-free N-methyl peptides.

Pulmonary function and chest CT abnormalities 3 months after discharge from COVID-19, 2020-2021: A nation-wide multicenter prospective cohort study from the Japanese respiratory society.

BACKGROUND: No comprehensive analysis of the pulmonary sequelae of coronavirus disease 2019 (COVID-19) in Japan based on respiratory function tests and chest computed tomography (CT) has been reported. We evaluated post-COVID-19 conditions, especially focusing on pulmonary sequelae assessed by pulmonary function tests and chest CT. METHODS: For this prospective cohort study, we enrolled 1069 patients who presented pneumonia at the time of admission in 55 hospitals from February 2020 to September 2021. Disease severity was classified as moderateⅠ, moderate II, and severe, defined primarily according to the degree of respiratory failure. The data on post-COVID-19 conditions over 12 months, pulmonary function, and chest CT findings at 3 months were evaluated in this study. Additionally, the impact of COVID-19 severity on pulmonary sequelae, such as impaired diffusion capacity, restrictive pattern, and CT abnormalities, was also evaluated. RESULTS: The most frequently reported post-COVID-19 conditions at 3 months after COVID-19 were muscle weakness, dyspnea, and fatigue (48.4%, 29.0%, and 24.7%, respectively). The frequency of symptoms gradually decreased over subsequent months. In pulmonary function tests at 3 months, the incidence of impaired diffusion capacity and restrictive pattern increased depending on disease severity. There also were differences in the presence of chest CT abnormalities at the 3 months, which was markedly correlated with the severity. CONCLUSION: We reported a comprehensive analysis of post-COVID-19 condition, pulmonary function, and chest CT abnormalities in Japanese patients with COVID-19. The findings of this study will serve as valuable reference data for future post-COVID-19 condition research in Japan.

Rapid and column-chromatography-free peptide chain elongation via a one-flow, three-component coupling approach.

Short peptides are extremely important as drugs and building blocks for the syntheses of longer peptides. Both solid- and liquid-phase peptide syntheses suffer from a large number of synthetic steps, high cost, and/or tedious purification. Here, we developed a rapid, mild, inexpensive, and column-chromatography-free peptide chain elongation via a one-flow, three-component coupling (3CC) approach that is the first to use α-amino acid N-carboxy anhydrides (α-NCAs) both as electrophiles and nucleophiles. We demonstrated the high-yielding and column-chromatography-free syntheses of 17 tripeptides, as well as a gram-scale synthesis of a tripeptide. The total synthesis of beefy meaty peptide was achieved by repeating the 3CC approach with the addition of only one column chromatographic purification. We also demonstrated a one-flow tripeptide synthesis via in situ preparation of α-NCA starting from three readily available protected amino acids. With this study, we achieved dramatic reductions in both time and cost compared with typical solid-phase synthesis.

Impact of respiratory bacterial infections on mortality in Japanese patients with COVID-19: a retrospective cohort study.

BACKGROUND: Although cases of respiratory bacterial infections associated with coronavirus disease 2019 (COVID-19) have often been reported, their impact on the clinical course remains unclear. Herein, we evaluated and analyzed the complication rates of bacterial infections, causative organisms, patient backgrounds, and clinical outcome in Japanese patients with COVID-19. METHODS: We performed a retrospective cohort study that included inpatients with COVID-19 from multiple centers participating in the Japan COVID-19 Taskforce (April 2020 to May 2021) and obtained demographic, epidemiological, and microbiological results and the clinical course and analyzed the cases of COVID-19 complicated by respiratory bacterial infections. RESULTS: Of the 1,863 patients with COVID-19 included in the analysis, 140 (7.5%) had respiratory bacterial infections. Community-acquired co-infection at COVID-19 diagnosis was uncommon (55/1,863, 3.0%) and was mainly caused by Staphylococcus aureus, Klebsiella pneumoniae and Streptococcus pneumoniae. Hospital-acquired bacterial secondary infections, mostly caused by Staphylococcus aureus, Pseudomonas aeruginosa, and Stenotrophomonas maltophilia, were diagnosed in 86 patients (4.6%). Severity-associated comorbidities were frequently observed in hospital-acquired secondary infection cases, including hypertension, diabetes, and chronic kidney disease. The study results suggest that the neutrophil-lymphocyte ratio (> 5.28) may be useful in diagnosing complications of respiratory bacterial infections. COVID-19 patients with community-acquired or hospital-acquired secondary infections had significantly increased mortality. CONCLUSIONS: Respiratory bacterial co-infections and secondary infections are uncommon in patients with COVID-19 but may worsen outcomes. Assessment of bacterial complications is important in hospitalized patients with COVID-19, and the study findings are meaningful for the appropriate use of antimicrobial agents and management strategies.

Diagnostic significance of secondary bacteremia in patients with COVID-19.

OBJECTIVES: We investigated the occurrence of non-respiratory bacterial and fungal secondary infections, causative organisms, impact on clinical outcomes, and association between the secondary pathogens and mortality in hospitalized patients with coronavirus disease 2019 (COVID-19). METHODS: This was a retrospective cohort study that included data from inpatients with COVID-19 from multiple centers participating in the Japan COVID-19 Taskforce (April 2020 to May 2021). We obtained demographic, epidemiological, and microbiological data throughout the course of hospitalization and analyzed the cases of COVID-19 complicated by non-respiratory bacterial infections. RESULTS: Of the 1914 patients included, non-respiratory bacterial infections with COVID-19 were diagnosed in 81 patients (4.2%). Of these, 59 (3.1%) were secondary infections. Bacteremia was the most frequent bacterial infection, occurring in 33 cases (55.9%), followed by urinary tract infections in 16 cases (27.1%). Staphylococcus epidermidis was the most common causative organism of bacteremia. Patients with COVID-19 with non-respiratory secondary bacterial infections had significantly higher mortality, and a multivariate logistic regression analysis demonstrated that those with bacteremia (aOdds Ratio = 15.3 [5.97-39.1]) were at higher risk of death. Multivariate logistic regression analysis showed that age, male sex, use of steroids to treat COVID-19, and intensive care unit admission increased the risk for nosocomial bacteremia. CONCLUSIONS: Secondary bacteremia is an important complication that may lead to poor prognosis in cases with COVID-19. An appropriate medical management strategy must be established, especially for patients with concomitant predisposing factors.

BMP3b Is a Novel Antifibrotic Molecule Regulated by Meflin in Lung Fibroblasts.

Fibroblasts play a central role in the lung fibrotic process. Our recent study identified a novel subpopulation of lung fibroblasts expressing meflin (mesenchymal stromal cell- and fibroblast-expressing Linx paralogue), antifibrotic properties of which were confirmed by murine lung fibrosis model. Meflin-expressing fibroblasts were resistant to fibrogenesis induced by TGF-ß (transforming growth factor-ß), but its underlying mechanisms remain unknown. In this study, evaluation of a silica-nanoparticle-induced lung fibrosis model confirmed the antifibrotic effect of meflin via the regulation of TGF-ß signaling. We conducted comparative gene expression profiling in lung fibroblasts, which identified growth differentiation factor 10 (Gdf10) encoding bone morphogenic protein 3b (BMP3b) as the most downregulated gene in meflin-deficient cells under the profibrotic condition with TGF-ß. We hypothesized that BMP3b can be an effector molecule playing an antifibrotic role downstream of meflin. As suggested by single-cell transcriptomic data, restricted expressions of Gdf10 (Bmp3b) in stromal cells including fibroblasts were confirmed. We examined possible antifibrotic properties of BMP3b in lung fibroblasts and demonstrated that Bmp3b-null fibroblasts were more susceptible to TGF-ß-induced fibrogenic changes. Furthermore, Bmp3b-null mice exhibited exaggerated lung fibrosis induced by silica-nanoparticles in vivo. We also demonstrated that treatment with recombinant BMP3B was effective against TGF-ß-induced fibrogenesis in fibroblasts, especially in the suppression of excessive extracellular matrix production. These lines of evidence suggested that BMP3b is a novel humoral effector molecule regulated by meflin which exerts antifibrotic properties in lung fibroblasts. Supplementation of BMP3B could be a novel therapeutic strategy for fibrotic lung diseases.

COVID-19-Triggered Acute Exacerbation of IPF, an Underdiagnosed Clinical Entity With Two-Peaked Respiratory Failure: A Case Report and Literature Review.

Because severe coronavirus disease 2019 (COVID-19) affects the respiratory system and develops into respiratory failure, patients with pre-existing chronic lung disorders, such as idiopathic pulmonary fibrosis (IPF), are thought to be at high risk of death. Patients with IPF often suffer from a lethal complication, acute exacerbation (AE), a significant part of which is assumed to be triggered by respiratory viral infection. However, whether mild to moderate COVID-19 can trigger AE in patients with IPF remains unknown. This is the case report of a 60-year-old man with a 4-year history of IPF who successfully recovered from moderate COVID-19 but subsequently developed more severe respiratory failure, which was considered to be a COVID-19-triggered acute exacerbation of idiopathic pulmonary fibrosis (AE-IPF). It is important to be aware of the risk of AE-IPF after COVID-19 and to properly manage this deadly complication of IPF. Recent literature reporting cases with chronic interstitial lung diseases which developed respiratory failure by complications with COVID-19 is also reviewed and discussed.

Prognostic value of modified coronary flow capacity by 13N-ammonia myocardial perfusion positron emission tomography in patients without obstructive coronary arteries.

BACKGROUND: Vasodilator capacity of coronary circulation is an important diagnostic and prognostic tool in patients with coronary artery disease (CAD). We aimed to clarify the incidence of coronary microvascular dysfunction (CMD), defined as impaired modified coronary flow capacity (mCFC) proposed by Johnson and Gould and measured by 13N-ammonia myocardial perfusion positron emission tomography (PET), in patients without obstructive CAD and to evaluate the risk of future cardiovascular events. METHODS: This retrospective study recruited 407 consecutive CAD-suspected patients who underwent both pharmacological stress/rest 13N-ammonia PET and coronary angiography. Of the 407 patients, 137 patients (median age, 70 years; 63 women) were eligible and followed up (median, 19.8 months). Endpoints were defined as cardiovascular death or major adverse cardiovascular events (MACEs), such as cardiovascular death, nonfatal myocardial infarction, unplanned hospitalization for any cardiac reasons, and unplanned coronary revascularization. The impaired mCFC group included patients with mildly to severely reduced regional CFC in, at least, one vascular territory (n=34), while the remaining patients (n=103) were categorized as having preserved mCFC. RESULTS: Overall, cardiovascular death and MACEs occurred in five (4%) patients. The Kaplan-Meier curve showed a significant reduction in event-free survival for cardiovascular death (p=0.004) and MACEs (p<0.0001) in the impaired mCFC group, compared to the preserved mCFC group. Impaired mCFC was independently associated with the incidence of both cardiovascular death and MACEs after propensity-score adjustments [hazard ratio (HR), 10.7; 95% confidence interval (CI), 1.0-106.0; p=0.04 and HR, 9.5; 95% CI, 2.5-36.2; p<0.001, respectively]. CONCLUSIONS: In CAD-suspected patients without obstructive coronary arteries, impaired mCFC was observed in approximately 25% and was associated with a higher risk of cardiovascular death and MACEs. The mCFC concept can help identify patients who would benefit from specific therapies or lifestyle modifications to prevent future MACEs and can clarify potential mechanisms of CMD.

Successful Treatment with High-dose Steroids for Acute Exacerbation of Idiopathic Pulmonary Fibrosis Triggered by COVID-19.

We herein report a case of acute exacerbation of idiopathic pulmonary fibrosis (AE-IPF) triggered by COVID-19. An 87-year-old woman tested positive for COVID-19 on a polymerase chain reaction test, and computed tomography revealed ground-glass opacity (GGO) superimposed on a background pattern consistent with usual interstitial pneumonia. Considering these data, we diagnosed her with AE-IPF. She experienced worsening of dyspnea and expansion of the GGO. Therefore, we introduced high-dose steroids (methylprednisolone 250 mg/day for 3 days). After the treatment, the pulmonary infiltrates improved. She was discharged from our hospital without severe disability. High-dose steroids can be a viable treatment option for AE-IPF triggered by COVID-19.

Contracted joint capsule affects labral and chondral lesions in patients with frozen shoulder.

PURPOSE: Intra-articular pathologies, such as labral and chondral lesions, are common in patients with frozen shoulder. This study evaluated the correlations between the range of motion and labral and chondral lesions in patients with frozen shoulder and investigated their pathophysiologies. METHODS: In total, 125 individuals (53 men and 72 women) who underwent arthroscopic pan-capsular release between 2014 and 2020 were included in the study. The range of motion was measured using scapular fixation and true glenohumeral motion under general anaesthesia. The American Shoulder and Elbow Surgeons Shoulder score and the Shoulder Rating Scale score of the University of California, Los Angeles were used to compare intra-articular pathologies. RESULTS: More than 80% of patients with frozen shoulder had labral pathologies, and nearly half of them had chondral pathologies. Labral lesions extending to the anterior rim of the glenoid had a greater range of motion and the greatest total American Shoulder and Elbow Surgeons Shoulder score. More severe chondral lesions had a lesser range of motion, but presented the greatest function scores and the lowest strength scores according to the Shoulder Rating Scale of the University of California, Los Angeles. The pain scores of the American Shoulder and Elbow Surgeons Shoulder score and the Shoulder Rating Scale of the University of California, Los Angeles were not correlated with the degree of these pathologies. The traction force affected the labrum during true range of motion, and the compression force occurred on the articular cartilage during internal rotation at 90° of forward flexion during diagnostic arthroscopy. CONCLUSION: Labral and chondral lesions are common in patients with frozen shoulder. Adherence to the capsulolabral complex induced a limited range of motion and labral and chondral pathologies. Diagnostic arthroscopy with motion is an important method of reproducing the pathogenesis of intra-articular structures for patients with frozen shoulder. LEVEL OF EVIDENCE: Level III.

Size and surface modification of silica nanoparticles affect the severity of lung toxicity by modulating endosomal ROS generation in macrophages.

BACKGROUND: As the application of silica nanomaterials continues to expand, increasing chances of its exposure to the human body and potential harm are anticipated. Although the toxicity of silica nanomaterials is assumed to be affected by their physio-chemical properties, including size and surface functionalization, its molecular mechanisms remain unclear. We hypothesized that analysis of intracellular localization of the particles and subsequent intracellular signaling could reveal a novel determinant of inflammatory response against silica particles with different physico-chemical properties. RESULTS: We employed a murine intratracheal instillation model of amorphous silica nanoparticles (NPs) exposure to compare their in vivo toxicities in the respiratory system. Pristine silica-NPs of 50 nm diameters (50 nm-plain) induced airway-centered lung injury with marked neutrophilic infiltration. By contrast, instillation of pristine silica particles of a larger diameter (3 µm; 3 µm-plain) significantly reduced the severity of lung injury and neutrophilic infiltration, possibly through attenuated induction of neutrophil chemotactic chemokines including MIP2. Ex vivo analysis of alveolar macrophages as well as in vitro assessment using RAW264.7 cells revealed a remarkably lower cellular uptake of 3 µm-plain particles compared with 50 nm-plain, which is assumed to be the underlying mechanism of attenuated immune response. The severity of lung injury and neutrophilic infiltration was also significantly reduced after intratracheal instillation of silica NPs with an amine surface modification (50 nm-NH2) when compared with 50 nm-plain. Despite unchanged efficacy in cellular uptake, treatment with 50 nm-NH2 induced a significantly attenuated immune response in RAW264.7 cells. Assessment of intracellular redox signaling revealed increased reactive oxygen species (ROS) in endosomal compartments of RAW264.7 cells treated with 50 nm-plain when compared with vehicle-treated control. In contrast, augmentation of endosomal ROS signals in cells treated with 50 nm-NH2 was significantly lower. Moreover, selective inhibition of NADPH oxidase 2 (NOX2) was sufficient to inhibit endosomal ROS bursts and induction of chemokine expressions in cells treated with silica NPs, suggesting the central role of endosomal ROS generated by NOX2 in the regulation of the inflammatory response in macrophages that endocytosed silica NPs. CONCLUSIONS: Our murine model suggested that the pulmonary toxicity of silica NPs depended on their physico-chemical properties through distinct mechanisms. Cellular uptake of larger particles by macrophages decreased, while surface amine modification modulated endosomal ROS signaling via NOX2, both of which are assumed to be involved in mitigating immune response in macrophages and resulting lung injury.

Fibroblasts positive for meflin have anti-fibrotic properties in pulmonary fibrosis.

The prognosis of elderly individuals with idiopathic pulmonary fibrosis (IPF) remains poor. Fibroblastic foci, in which aggregates of proliferating fibroblasts and myofibroblasts are involved, are the pathological hallmark lesions in IPF to represent focal areas of active fibrogenesis. Fibroblast heterogeneity in fibrotic lesions hampers the discovery of the pathogenesis of pulmonary fibrosis. Therefore, to determine the pathogenesis of IPF, identification of functional fibroblasts is warranted. The aim of this study was to determine the role of fibroblasts positive for meflin, identified as a potential marker for mesenchymal stromal cells, during the development of pulmonary fibrosis.We characterised meflin-positive cells in a single-cell atlas established by single-cell RNA sequencing (scRNA-seq)-based profiling of 243 472 cells from 32 IPF lungs and 29 normal lung samples. We determined the role of fibroblasts positive for meflin using bleomycin (BLM)-induced pulmonary fibrosis.scRNA-seq combined with in situ RNA hybridisation identified proliferating fibroblasts positive for meflin in fibroblastic foci, not dense fibrosis, of fibrotic lungs in IPF patients. A BLM-induced lung fibrosis model for meflin-deficient mice showed that fibroblasts positive for meflin had anti-fibrotic properties to prevent pulmonary fibrosis. Although transforming growth factor-ß-induced fibrogenesis and cell senescence with the senescence-associated secretory phenotype were exacerbated in fibroblasts via the repression or lack of meflin, these were inhibited in meflin-deficient fibroblasts with meflin reconstitution.These findings provide evidence to show the biological importance of meflin expression on fibroblasts and myofibroblasts in the active fibrotic region of pulmonary fibrosis.

Effect of a single bout of morning or afternoon exercise on glucose fluctuation in young healthy men.

The timing of exercise plays an important role in the effect of the exercise on physiological functions, such as substrate oxidation and circadian rhythm. Exercise exerts different effects on the glycemic response to exercise and meal intake depending on when the exercise performed. Here, we comprehensively investigated the effects of the timing (morning or afternoon) of exercise on glucose fluctuation on the basis of several indices: glycemic variability over 24 h (24-h SD), J-index, mean amplitude of glucose excursions (MAGE), continuous overall net glycemic action (CONGA), and detrended fluctuation analysis (DFA). Eleven young men participated in 3 trials in a repeated measures design in which they performed a single bout of exercise at 60% of their maximal oxygen uptake for 1 h beginning either at 7:00 (morning exercise), 16:00 (afternoon exercise), or no exercise (control). Glucose levels were measured using a continuous glucose monitoring system (CGMs). Glucose fluctuation was slightly less stable when exercise was performed in the afternoon than in the morning, indicated by higher CONGA at 2 h and α2 in DFA in the afternoon exercise trial than in the control trial. Additionally, decreased stability in glucose fluctuation in the afternoon exercise trial was supported by the descending values of the other glucose fluctuation indices in order from the afternoon exercise, morning exercise, and control trials. Meal tolerance following exercise was decreased after both exercise trials. Glucose levels during exercise were decreased only in the afternoon exercise trial, resulting in less stable glucose fluctuations over 24 h.

Strain-induced modulation of temperature characteristics in ferrimagnetic Tb-Fe films.

This study investigates the effect of strain on the compensation temperature of ferrimagnetic Tb-Fe films formed on a flexible substrate. The compensation temperature is determined by the anomalous Hall measurement, and an application of 1.2% tensile strain reduces the compensation temperature by 12 K. X-ray magnetic circular dichroism reveals that approximately 5% of Fe magnetic moment and approximately 1% of Tb magnetic moment are reduced by an application of 0.9% tensile strain at the room temperature. To understand the greater reduction in Fe magnetization compared with that in Tb and the compensation temperature reduction simultaneously, a model applying molecular field theory is analyzed. Changes in three types of exchange coupling between Fe and Tb atoms are speculated to be caused by the strain.

Validation and reliability of a Japanese version of the Simple Shoulder Test: a cross-sectional study.

BACKGROUND: The Simple Shoulder Test (SST) is a widely used patient-reported outcome assessment. The purpose of this study was to develop and validate a Japanese version of the SST (SST-Jp). METHODS: A two-stage observational study was conducted to validate the cross-cultural adaptation of the SST. A total of 100 patients with shoulder disorders completed the SST-Jp; the Disability of Arm, Shoulder, and Hand assessment; and the Medical Outcomes Short-Form 36 (SF-36) at an initial visit. Thirty-four of the patients repeated the SST-Jp one week after the first examination. The test-retest reliability was quantified using the interclass correlation coefficient, and Cronbach's alpha (α) was calculated to assess internal consistency. Construct validity was assessed using Spearman's rank correlation coefficient. RESULTS: The internal consistency of the SST-Jp was very high (α = 0.826). The interclass correlation coefficient of the SST-Jp was also high (0.859). There was a strong, positive correlation between the Disability of Arm, Shoulder, and Hand and the SST-Jp (r = 0.717, P < .001). The SST-Jp was significantly correlated with most of the SF-36 subscales. The correlations of the SST-Jp with physical subscales of the SF-36 were stronger than those with the other subscales. CONCLUSIONS: The SST-Jp was found to be a valid and reliable measurement for shoulder joint pain and function assessment among the Japanese population.

Development of small high-voltage AC power supply for a dielectric barrier discharge plasma actuator.

A dielectric-barrier discharge plasma actuator (DBDPA) is a promising flow control device that can prevent flow separation around an airfoil using electrical discharges. Miniaturizing the DBDPA power supply remains a crucial technological challenge because its size and weight determine the performance of fluid devices equipped with this type of actuator. In this study, we propose a compact high-voltage AC power supply for a DBDPA intended for installation on small airplanes, including unmanned aerial vehicles. The power supply, which consists of a power supply board, a main control board, and a DC/AC converter board, is ∼110 g in weight. It can drive a 300-mm long DBDPA without any substantial voltage drop. The power consumption in standby remains below 1 W, and the maximum consumption during discharge in burst mode at a burst ratio of 5% is 24 W. The power supply uses a lithium-ion battery with a capacity of 1800 mA h, which allows continuous DBDPA operation for ∼1.5 h. An experiment was conducted in a wind tunnel using an airfoil model whose cross-section corresponds to that of an airfoil from a commercial glider airplane. Experimental results reveal that the surface pressure around the airfoil is modified by DBDPA operation, clearly demonstrating the effectiveness of the developed power supply for operating a DBDPA as a flow control device. The size and weight of the proposed power supply can be established as a benchmark to further miniaturize and optimize DBDPA power supplies.