Equol, a soybean metabolite with estrogen-like functions, decreases lipopolysaccharide-induced human neutrophil extracellular traps in vitro.

OBJECTIVE: Neutrophil extracellular traps (NETs) defend against acute infections. However, their overexpression causes organ failure during sepsis. Control of NET formation may improve the outcomes of patients with sepsis. Equol, a soybean isoflavone, is a female hormone analog, which prevents inflammation. We evaluated the effects of equol on NET formation in human neutrophils during inflammatory stimulation in vitro. METHODS: Healthy volunteers provided blood samples. An enzyme-linked immunosorbent assay (ELISA) assessed serum equol concentrations. NET formation in neutrophils was induced by lipopolysaccharide (LPS) treatment. ELISA quantified DNA-binding elastase, and immunostaining assessed NET formation. Reverse-transcription quantitative PCR and western blotting detected G-protein-coupled receptor 30 (GPR30) or peptidyl arginine deiminase 4 (PAD4) expression. Flow cytometry assessed neutrophil phagocytic ability with inactivated Escherichia coli. RESULTS: In neutrophils derived from males with low-serum equol levels (low-serum equol group), equol significantly decreased DNA-binding elastase levels and NET formation. Equol did not decrease NETs in neutrophils from males with high-serum equol levels. GPR30 expression of neutrophils was higher in the low-serum than in the high-serum equol group. PAD4 mRNA levels and nuclear PAD4 protein expression also decreased than the vehicle control in the low-serum equol group. Equol did not alter the phagocytic ability of neutrophils. In neutrophils from young females, equol had no inhibitory effect on NET formation. CONCLUSIONS: Equol decreases LPS-induced NET formation in neutrophils from males via inhibition of PAD4 expression. Our findings provide a rationale for investigating a new therapeutic approach using equol to control neutrophil activity during sepsis.

The role of death receptor signaling pathways in mouse Sertoli cell avoidance of apoptosis during LPS- and IL-18-induced inflammatory conditions.

Lipopolysaccharide (LPS) triggers infectious acute inflammation, and interleukin (IL)-18 is an inflammasome-mediated cytokine. We previously demonstrated that endogenous IL-18 induces testicular germ cell apoptosis during acute inflammation when plasma IL-18 levels are high. Additionally, high-dose recombinant IL-18 (rIL-18) induced Leydig cell apoptosis. The blood-testis barrier formed by Sertoli cells protects testicular germ cells from both exogenous and endogenous harmful substances. However, the impact of LPS and IL-18 on Sertoli cells remained unclear. We stimulated TM4 cells, a mouse Sertoli cell line, with LPS (200 or 1000 ng/mL) or rIL-18 (0.1-100 ng/mL) at levels that induced Leydig cell apoptosis in our previous study and assessed caspase 3 cleavage and the mRNA expression of inflammatory cytokines and markers of apoptotic pathways (Tnfr1, Fasl, Fas, Fadd) after stimulation. Il6 mRNA was increased by LPS stimulation. Tnfα mRNA was increased by 200 ng/mL LPS but not 1000 ng/mL LPS. Fas was increased, but Fasl was decreased, by LPS. LPS had little influence on Tnfr1 or Fadd mRNA expression and did not induce apoptosis. Il18 mRNA was not increased, and Il18r1 was significantly decreased following LPS treatment. Treatment with rIL-18 increased Il18r1 mRNA and induced inflammation, but decreased Tnfr1 and had little influence on apoptosis, as indicated by Tnfα, Fasl, Fas, Fadd and cleaved caspase 3. These results suggested that Sertoli cells do not easily undergo apoptosis despite strong inflammatory stimuli. Additionally, Sertoli cells may resist inflammation and play a larger role in protecting testicular homeostasis than other component cells of the testis.

High Plasma tPAPAI-1C Levels May Be Related to a Poor Prognosis in Patients with Severe or Critical COVID-19: A Single-Center Retrospective Study.

Severe novel coronavirus disease 2019 (COVID-19) patients have a high incidence of thrombotic complications and mortality. The pathophysiology of coagulopathy involves fibrinolytic system impairment and vascular endothelial damage. This study examined coagulation and fibrinolytic markers as outcome predictors. In an observational study of 164 COVID-19 patients admitted to our emergency intensive care unit, hematological parameters on days 1, 3, 5, and 7 were retrospectively compared between survivors and nonsurvivors. Nonsurvivors had a higher APACHE II score, SOFA score, and age than survivors. Nonsurvivors also had a significantly lower platelet count and significantly higher plasmin/α2plasmin inhibitor complex (PIC), tissue plasminogen activator/plasminogen activator inhibitor-1 complex (tPAPAI-1C), D-dimer, and fibrin/fibrinogen degradation product (FDP) levels than survivors throughout the measurement period. The 7-day maximum or minimum values of the tPAPAI-1C, FDP, and D-dimer levels were significantly higher in nonsurvivors. A multivariate logistic regression analysis showed that the maximum tPAPAI-1C (OR = 1.034; 95% CI,1.014-1.061; p = 0.0041) was an independent factor affecting mortality, with an area under the curve (AUC) of 0.713 (optimum cut-off of 51 ng/mL; sensitivity, 69.2%; and specificity, 68.4%). COVID-19 patients with poor outcomes exhibit exacerbated coagulopathy with fibrinolysis inhibition and endothelial damage. Consequently, plasma tPAPAI-1C might be a useful predictor of the prognosis in patients with severe or critical COVID-19.

Use of Slack for COVID-19 disaster response in Hyogo Prefecture.

INTRODUCTION: The coronavirus disease 2019 (COVID-19) pandemic has caused great disruptions in education and healthcare systems. However, before the COVID-19 pandemic, Hyogo Prefecture did not have a centralized system for sharing information among hospitals, public healthcare centers, the government, fire departments, and medical associations. In March 2021, we used Slack,® an enterprise social network, to invite people from hospitals, healthcare centers, the government, fire departments, and medical associations to join our workspace. This study was undertaken to verify whether the use of Slack can improve information sharing during a disaster. METHODS: A questionnaire on whether information-sharing tools such as Slack are useful for disaster response in Hyogo Prefecture, including for COVID-19, was administered using a Google Form. RESULTS: Of the 19 people who responded to the questionnaire, 15 (78.9 percent) were first-time users of Slack and 18 (94.7 percent) found it easy to use. All 19 participants stated that Slack proved useful in providing pertinent information regarding preventive measures for the control of COVID-19, and they believed that it would be a helpful tool if such disasters were to occur in the future. Regarding the launch time of Slack, 11 respondents (57.9 percent) answered that it was launched at the appropriate time, while five respondents (26.3 percent) answered that it was launched too late. CONCLUSIONS: The participants thought that Slack was effective for accelerating their communication. The use of Slack could also improve future disaster response communications.

Improving residential care amid COVID-19: The link between risk factors and hospitalization.

AIM: This study was conducted to investigate the relationship between the hospitalizations and backgrounds of patients with coronavirus disease 2019 to identify specific risk factors. METHODS: This retrospective study used health observation records to analyze the relationship between certain risk factors and the subsequent hospitalization of 321 patients who were discharged from a residential care facility between January 16 and February 8, 2021. The usefulness of a hospitalization prediction score, created based on the presence of comorbidities and sex, was examined. RESULTS: Being older, male, and having a history of high blood pressure or vascular disease were all risk factors. A multivariate analysis with age and hospitalization predictive score as independent variables and hospitalization as the dependent variable showed that age (odds ratio: 1.07, 95 percent confidence interval: 1.03-1.11, p < 0.01) significantly increased hospitalization risk by 7 percent for every 1-year age increase. The median time from illness onset to hospitalization for all patients was 9 days (interquartile range: 8-10). Hypoxia was the most common cause of hospitalization. However, hypoxia and other symptoms, such as cough and dyspnea, were not correlated. CONCLUSION: Given the pandemic, there may come another time when hospitals are not able to accommodate all patients who require care. In such instances, age, sex, the presence of comorbidities, and checking oxygen saturation regularly using a pulse oximeter around 9 days after the onset of the disease should all be considered important, as it may lead to improved and safer operation of overnight care facilities.

Dysfunctional Sars-CoV-2-M protein-specific cytotoxic T lymphocytes in patients recovering from severe COVID-19.

Although the importance of virus-specific cytotoxic T lymphocytes (CTL) in virus clearance is evident in COVID-19, the characteristics of virus-specific CTLs related to disease severity have not been fully explored. Here we show that the phenotype of virus-specific CTLs against immunoprevalent epitopes in COVID-19 convalescents might differ according to the course of the disease. We establish a cellular screening method that uses artificial antigen presenting cells, expressing HLA-A*24:02, the costimulatory molecule 4-1BBL, SARS-CoV-2 structural proteins S, M, and N and non-structural proteins ORF3a and nsp6/ORF1a. The screen implicates SARS-CoV-2 M protein as a frequent target of IFNγ secreting CD8+ T cells, and identifies M198-206 as an immunoprevalent epitope in our cohort of HLA-A*24:02 positive convalescent COVID-19 patients recovering from mild, moderate and severe disease. Further exploration of M198-206-specific CD8+ T cells with single cell RNA sequencing reveals public TCRs in virus-specific CD8+ T cells, and shows an exhausted phenotype with less differentiated status in cells from the severe group compared to cells from the moderate group. In summary, this study describes a method to identify T cell epitopes, indicate that dysfunction of virus-specific CTLs might be an important determinant of clinical outcomes.

Prognostic Factors Associated With Mortality of Patients With COVID-19 Requiring Ventilator Management: A Retrospective Cohort Study.

Aim There are few reports on the prognostic factors associated with mortality in coronavirus disease (COVID-19) patients with critical disease. This study assessed prognostic factors associated with mortality of patients with critical COVID-19 who required ventilator management. Methods This single-center, retrospective cohort study used medical record data of COVID-19 patients admitted to an emergency ICU at a hospital in Japan between March 1, 2020 and September 30, 2021, and provided with ventilator management. Multivariable logistic regression was used to identify factors associated with mortality. Results Seventy patients were included, of whom 29 (41.4%) died. The patients who died were significantly older (median: 69 years) (interquartile range [IQR]: 47-82 years) than the patients who survived (62 years [38-84 years], p<0.007). In addition, patients who died were significantly less likely to have received steroid therapy than patients who survived (25 [86.2%] vs. 41 [100%], p=0.026). In the multivariable analysis, age was identified as a significant prognostic factor for mortality and the risk of death increased by 6% for every one-year increase in age (OR: 1.06; 95% CI: 1.00-1.13; p=0.048). Medical history was not a risk factor for death. Conclusion Age was a predictor of mortality in critically ill patients with COVID-19. Therefore, the indications for critical care in older patients with COVID-19 should be carefully considered.

N95 masks worn to protect against COVID-19 prevented tuberculosis exposure in healthcare workers.

Since the start of the COVID-19 pandemic, the healthcare workers in our institution have been equipped with N95 masks when performing aerosol-generating procedures, as these are associated with an increased risk of infection. We present a case in which using an N95 mask prevented tuberculosis (TB) exposure among healthcare workers administering prehospital care in rapid response vehicles. Even after the resolution of the COVID-19 pandemic in the future, wearing N95 masks among healthcare workers is recommended to protect against pathogens, including TB, when performing aerosol-producing procedures or prehospital activities for patients suspected of having respiratory diseases.

Stenotrophomonas maltophilia Bacteremia Associated With Severe COVID-19: Successful Treatment With Appropriate Antimicrobial Therapy.

Stenotrophomonas maltophilia, an opportunistic pathogen, can cause bacteremia in immunocompromised and debilitated patients. A 50-year-old man with severe coronavirus disease 2019 (COVID-19) was admitted to our hospital's intensive care unit where he underwent extracorporeal membrane oxygenation and ventilatory support. On day 25, he developed S. maltophilia bacteremia originating from an indwelling central venous catheter. After confirming susceptibility, trimethoprim-sulfamethoxazole (80 mg/400 mg) was administered thrice daily. Following improvement, he was weaned from ventilation, recovered sufficiently, and was discharged on day 53. To the best of our knowledge, this is the first report of a patient recovering after antimicrobial treatment for S. maltophilia bacteremia associated with severe COVID-19.

Effect of Tocilizumab Treatment on Patients with Coronavirus Disease 2019 and Bacteremia: A Retrospective Cohort Study.

INTRODUCTION: This study aimed to determine if tocilizumab treatment for coronavirus disease 2019 (COVID-19) increases bacteremia and suppresses fever and inflammatory reactants. METHODS: In this single-center, retrospective, observational study, all patients with COVID-19 admitted to our emergency intensive care unit from March 2020 to August 2021 were categorized into tocilizumab-treated and tocilizumab-naïve groups, and the incidence of bacteremia and other factors between the two groups were compared. Patients with bacteremia were further classified into tocilizumab-treated and tocilizumab-naïve groups to determine if fever and inflammatory reactants were suppressed. RESULTS: Overall, 144 patients were included in the study, 51 of whom received tocilizumab, which was administered on the day of admission. Further, of the 24 (16.7%) patients with bacteremia, 13 were in the tocilizumab-treated group. Results revealed a significant difference in the C-reactive protein level (p < 0.001) at the onset of bacteremia between the tocilizumab-treated group [median 0.42 mg/dL (0.27-0.44 mg/dL)] and the tocilizumab-naïve group [7.48 mg/dL (4.56-13.9 mg/dL)]. The median number of days from admission to onset of bacteremia was not significantly different between the tocilizumab-treated group [10 days (9-12 days)] and the tocilizumab-naïve group [9 days (7.5-11 days)] (p = 0.48). There was no significant difference in fever between the groups. Multivariate logistic analysis showed that tocilizumab treatment did not affect the probability of bacteremia. CONCLUSION: Treatment of patients with COVID-19 with tocilizumab does not increase the risk of bacteremia. Tocilizumab suppresses C-reactive protein levels but not fever. Therefore, careful monitoring of fever can reduce the risk of missed bacteremia.

Effect of Tocilizumab Treatment on Patients with Coronavirus Disease 2019 and Bacteremia: A Retrospective Cohort Study.

INTRODUCTION: This study aimed to determine if tocilizumab treatment for coronavirus disease 2019 (COVID-19) increases bacteremia and suppresses fever and inflammatory reactants. METHODS: In this single-center, retrospective, observational study, all patients with COVID-19 admitted to our emergency intensive care unit from March 2020 to August 2021 were categorized into tocilizumab-treated and tocilizumab-naïve groups, and the incidence of bacteremia and other factors between the two groups were compared. Patients with bacteremia were further classified into tocilizumab-treated and tocilizumab-naïve groups to determine if fever and inflammatory reactants were suppressed. RESULTS: Overall, 144 patients were included in the study, 51 of whom received tocilizumab, which was administered on the day of admission. Further, of the 24 (16.7%) patients with bacteremia, 13 were in the tocilizumab-treated group. Results revealed a significant difference in the C-reactive protein level (p < 0.001) at the onset of bacteremia between the tocilizumab-treated group [median 0.42 mg/dL (0.27-0.44 mg/dL)] and the tocilizumab-naïve group [7.48 mg/dL (4.56-13.9 mg/dL)]. The median number of days from admission to onset of bacteremia was not significantly different between the tocilizumab-treated group [10 days (9-12 days)] and the tocilizumab-naïve group [9 days (7.5-11 days)] (p = 0.48). There was no significant difference in fever between the groups. Multivariate logistic analysis showed that tocilizumab treatment did not affect the probability of bacteremia. CONCLUSION: Treatment of patients with COVID-19 with tocilizumab does not increase the risk of bacteremia. Tocilizumab suppresses C-reactive protein levels but not fever. Therefore, careful monitoring of fever can reduce the risk of missed bacteremia.

Determination of Ultra-trace Tellurium in Steel by ID-ICP-MS/MS with Liquid-Liquid Extraction.

Inductively coupled plasma mass spectrometry using isotope dilution (ID-ICP-MS) with liquid-liquid extraction was used for determining ultra-trace tellurium (Te) in steels to improve the sensitivity, accuracy, and precision of the analysis. Single quadrupole-type ICP-MS cannot be used to determine the trace amount of Te because of the contribution of mass spectrometric interference by Xe. To overcome this, tandem mass-filter-type ICP-MS (ICP-MS/MS) was used to determine Te. A pretreatment by liquid-liquid extraction was also employed to compensate for the decrease in the signal intensity due to the use of ICP-MS/MS. The relative standard deviations of the resulting abundances, defined by repeated analysis from separated three steel samples, were <1.3%. Furthermore, the determined values of Te in standard reference materials were close to the reference values. Thus, the developed determination method is useful for the analysis of ultra-trace Te in steels.

Interleukin-18 levels and mouse Leydig cell apoptosis during lipopolysaccharide-induced acute inflammatory conditions.

Interleukin (IL)-18 is an inflammasome-mediated cytokine produced by germ cells, Leydig cells, and resident macrophages that is indispensable in the maintenance of homeostasis in the testis. We previously demonstrated that endogenous IL-18 induces testicular germ cell apoptosis during acute inflammation when plasma IL-18 levels are very high. However, the impact of acute inflammation and IL-18 on Leydig cells remained unclear. TM3 cells, a mouse Leydig cell line, and RAW264.7 cells, a mouse macrophage cell line, were stimulated with lipopolysaccharide (LPS) or recombinant IL-18 (rIL-18). We assessed the expression of inflammatory cytokines, caspase cleavage, and markers of apoptotic pathways. In Leydig cells, caspase 3 cleavage was increased and death-receptor-mediated apoptotic pathways were activated after LPS stimulation. However, LPS stimulation did not increase IL-18 expression in the Leydig cell line. When high-dose rIL-18 was administered to the Leydig cell line to mimic levels seem after inflammation, rIL-18 upregulated Tnf-α mRNA, Fadd mRNA, and Fas protein, promoted cleavage of caspase-8 and caspase-3, and induced apoptosis. Low-dose rIL-18 did not stimulate apoptosis. To determine if the high level of IL-18 seen in the testes after inflammation was derived from immune cells, we examined IL-18 protein expression in a macrophage cell line, RAW264.7. In contrast to the TM3 cells, IL-18 was significantly increased in RAW264.7 cells after LPS stimulation. These results suggest that high-dose IL-18 derived from macrophages is harmful to Leydig cells. Reducing the overexpression of IL-18 could be a new therapeutic approach to prevent Leydig cell apoptosis as a result of acute inflammation.

A Japanese case of cerebellar ataxia, spastic paraparesis and deep sensory impairment associated with a novel homozygous TTC19 mutation.

Mitochondrial complex III (CIII) deficiency comprises a group of complex and heterogeneous genetic disorders. TTC19 mutations constitute a rare cause of CIII deficiency and are associated with neurological disorders in childhood and adulthood. Herein, we describe a 27-year-old Japanese man with cerebellar ataxia, spastic paraparesis, loss of deep sensation, mild frontal lobe dysfunction and transient psychiatric symptoms. Brain magnetic resonance imaging showed cerebellar atrophy and bilateral high-intensity signals in the inferior olives and regions adjacent to periaqueductal gray matter, on T2-weighted images. On whole-exome sequencing, we detected a novel homozygous frameshift mutation c.157_158dup [p.Pro54Alafs*48] in TTC19. Mitochondrial enzyme assays confirmed mild impairment of CIII enzymatic activity in lymphoblasts, which was consistent with TTC19-related CIII deficiency. His symptoms and radiological findings demonstrated an early stage or mild form of this disease, and further clarify the characteristics of patients with rare TTC19 mutations.

Diagnosis and long-term management of hydrolyzed wheat protein wheat-dependent exercise-induced anaphylaxis.

Case: A 37-year-old woman was admitted to the emergency room with bilateral periorbital edema, hypotension, and expiratory stridor that developed 30 min after starting to hang out washing following consumption of a jam bun. Despite no food allergies or similar episodes, she had recently developed facial wheals after bathing. Outcome: She was immediately and repeatedly administered adrenalin and succinic acid hydrocortisone sodium for wheat-dependent exercise-induced anaphylaxis. On the third hospital day, radioallergosorbent testing reactions to wheat, gluten, and omega-5 gliadin were mildly positive; skin-prick tests for hydrolyzed wheat protein and a face-wash challenge were positive. Therefore, we diagnosed hydrolyzed wheat protein wheat-dependent exercise-induced anaphylaxis. Despite advising her about hyposensitization, the episode recurred and an adrenalin auto-injector was prescribed. Conclusion: Differentiating hydrolyzed wheat protein wheat-dependent exercise-induced anaphylaxis from conventional wheat-dependent exercise-induced anaphylaxis is important owing to their severity and similarities. Each requires long-term management of patients' etiological conditions by advising them about hyposensitization and prescribing adrenalin auto-injectors.

Determination method of multi elements in ferromanganese samples by LA-ICP-MS.

A dilution method using MnO(2) for the manganese nodule standard, JMn1, has been newly adopted to obtain accurate calibration lines. Moreover, an internal standard correction method was applied to correct changes in the sample introduction efficiency for the prime purpose of laser ablation-inductively coupled plasma-mass spectrometry (LA-ICP-MS) analysis. Using the dilution method and the correction method, good calibration lines were successfully obtained for 13 elements. The methods were also applied to the analysis of ferromanganese nodules. The results were in accordance with those obtained by ICP-MS, which requires chemical treatments of samples. The results are encouraging, and imply a high potential of LA-ICP-MS for the analysis of trace elements in ferromanganese nodules.

A role for IL-18 in human neutrophil apoptosis.

Decreased neutrophil apoptosis is associated with persistent inflammation, the severity of which correlates with serum IL-18 levels. IL-18 receptors as well as Toll-like receptors, including Toll-like receptor 4, a receptor for LPS, possess a highly conserved intracellular domain called "Toll-IL-1R domain" and activate overlapping signaling pathways. Here, we show that IL-18 modulates neutrophil apoptosis and compare its mechanism of action with LPS. We found that both IL-18 and LPS decreased neutrophil apoptosis in a similar dose- and time-dependent fashion. However, pretreatment with the phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002 increased apoptosis more effectively in IL-18- than in LPS-stimulated cells, whereas the ERK inhibitor PD98059 had the same effect in both. In contrast, the p38 mitogen-activated protein kinase (MAPK) inhibitor SB203580 had no influence on apoptosis at all. Neutrophils constitutively expressed mRNA for IL-18 receptor beta, but little or no receptor alpha, both of which increased during coculture with either IL-18 or LPS in a time- and dose-dependent manner. Of the Bcl-2 family, antiapoptotic A1/Bfl-1 tended to increase on IL-18 and LPS stimulation, but was further increased despite increased apoptosis in the presence of MAPK inhibitors. Thus, human neutrophils can express mRNA for IL-18 receptors alpha and beta, and IL-18, like LPS, inhibits neutrophil apoptosis by activating PI3K and ERK pathways but not p38MAPK. However, PI3K may play more important role(s) in IL-18- than in LPS-induced inhibition of apoptosis. Mitogen-activated protein kinases seem to mediate antiapoptotic signals through factors other than Bcl-2 gene family expression.

Effect of high pressure carbon dioxide on the clumping of the bacterial spores.

The formation of spore clumps of Bacillus coagulans and Bacillus licheniformis during high-pressure carbon dioxide treatment (HCT) was investigated. As the treatment time increased, the number of spore clumps increased. After 120 min, single spore decreased to 20-35% of the population. Addition of a surfactant decreased the hydrophobicity of spore surface and increased both the number of single spores and the rate of inactivation ratio of B. coagulans and B. licheniformis spores.