Glucose Tolerance and the Risk Factors for Transmission in Japanese SARS-CoV-2/WA-1/2020 Epicenter: A Retrospective Study.

Purpose: The severe pathogenic ancient-type COVID-19, SARS-CoV-2/WA-1/2020 was the predominant gene variant in early 2020 in Japan, however, its transmissibility was uncertain. The period before the public commenced using any personal protective equipment (PPE) was evaluating to describe the transmissibility of the SARS-CoV-2/WA-1/2020. We analyzed the secondary attack rate (SAR) among close contacts and the risk factor for SAR. Methods: This retrospective cohort study included a total of 539 patients who were anticipated for the SARS-CoV-2/WA-1/2020 infection at Toho University Medical Center Omori Hospital from February to May 2020. We selected 54 patients with 1) exclude other pathogens infection, 2) include "Three Cs" condition: crowded places between distance< 6 feet, closed spaces indoor and close contact settings involving contact >15min with a person tested positive for SARS-CoV-2/WA-1/2020 without PPE. We evaluated alternative infection risks: the body mass index (BMI) and diabetes (DM) status (non-DM, pre-DM, and DM) as demographic determinants of transmissibility and infectivity of SARS-CoV2/WA-1/2020 cases during the incubation period. Results: The calculated SAR was 79.3%. BMI was significantly associated with the PCR positivity rate, which was significant in the univariate (CI 95%, 1.02-1.51; P = 0.03) and multivariate (CI 95%, 1.02-1.60; P = 0.03) analyses. Comparing the different BMI groups, the highest BMI group (25.5-35.8 kg/m2) had an elevated risk of SAR compared to the lowest BMI group (14.0-22.8 kg/m2), with an odds ratio of 1.41 (95% CI, 1.02-1.59; P = 0.03). There were no significant differences in the risk of SAR among different DM statuses. Conclusion: The transmissibility of SARS-CoV2/WA-1/2020 was high (79.3%) among household members without PPE who had "Three Cs" exposure. Although pre-DM and established DM did not confer a risk for transmissibility, higher BMI was associated with an increased risk of SAR. Trial Registration: UMIN Clinical Trials Registry, UMIN0000 50905.

Role of albumin Cys34 redox state in the progression of differentiated thyroid carcinoma and induction of ferroptosis.

Differentiated thyroid cancer (DTC) is the most prevalent endocrine malignancy worldwide and requires effective prognostic markers and therapeutic targets to optimize patient outcomes. This study investigated the potential of human serum albumin (HSA) cysteine-34 (Cys34) redox state as a prognostic indicator and therapeutic avenue for DTC. A retrospective cohort study of 99 patients with DTC undergoing radioactive iodine therapy found that higher concentrations of HSA with the reduced form of Cys34 (i.e., human mercaptalbumin [HMA]) were associated with improved progression-free survival in metastatic DTC. In vitro experiments using a DTC cell line revealed that HMA induced cytotoxic effects by triggering ferroptosis, characterized by lipid peroxidation, intracellular ROS accumulation, and decreased cell viability. Ferroptosis inhibitors rescued cell viability, confirming their role in cytotoxicity. These results implicate the HSA-Cys34 redox state is a promising avenue for precision medicine in DTC, shedding light on the prognostic relevance and therapeutic potential of HMA-induced ferroptosis. They emphasize the opportunity for personalized treatment strategies to advance the management of patients with DTC.

Molecular Cloning, Expression Analyses, and Physiological Roles of Cathelicidins in the Bursa of Fabricius of the Japanese Quail, Coturnix japonica.

Antimicrobial peptides (AMPs) act directly on pathogens and maintain the anti-inflammatory effects and activation of immunocompetent cells. Therefore, the activation of the immune system in poultry via the elevation of endogenous AMPs has been attempted. In this study, we focused on the host defense mechanisms in the bursa of Fabricius (BF) of Japanese quail, cloned the cDNA of cathelicidin (CATH)-1 to -3, and analyzed their expression sites. In situ hybridization experiments revealed the mRNA expression of the CATHs in the interfollicular epithelium surrounding the lumen of the quail BF, which suggests that each CATH may exert its antimicrobial action directly in the BF. The intravenous injection of bacterial lipoteichoic acid and lipopolysaccharide endotoxins into the quail promoted the mRNA expression of CATH-1 and CATH-3 in the BF. The addition of CATH-1 or CATH-2 at the time of the antigen injection into mice resulted in antiserum with high antibody titers. Ad libitum administration of butyrate, a short-chain fatty acid, in the drinking water induced an increase in CATH-2 mRNA expression in the BF under certain conditions. These results may improve the defense mechanisms of quail by stimulating CATH expression in the BF through their diet.

Overcoming the Limitations of CRISPR-Cas9 Systems in Saccharomyces cerevisiae: Off-Target Effects, Epigenome, and Mitochondrial Editing.

Modification of the genome of the yeast Saccharomyces cerevisiae has great potential for application in biological research and biotechnological advancements, and the CRISPR-Cas9 system has been increasingly employed for these purposes. The CRISPR-Cas9 system enables the precise and simultaneous modification of any genomic region of the yeast to a desired sequence by altering only a 20-nucleotide sequence within the guide RNA expression constructs. However, the conventional CRISPR-Cas9 system has several limitations. In this review, we describe the methods that were developed to overcome these limitations using yeast cells. We focus on three types of developments: reducing the frequency of unintended editing to both non-target and target sequences in the genome, inducing desired changes in the epigenetic state of the target region, and challenging the expansion of the CRISPR-Cas9 system to edit genomes within intracellular organelles such as mitochondria. These developments using yeast cells to overcome the limitations of the CRISPR-Cas9 system are a key factor driving the advancement of the field of genome editing.

SGLT2 inhibitor versus carbohydrate-restricted isocaloric diet: reprogramming substrate oxidation in type 2 diabetes.

OBJECTIVE: Based on the whole-body energy metabolism and insulin action, the difference between increased excretion of carbohydrate in urine by SGLT2i and reduced same amount of oral carbohydrate intake are scarce. This study aimed to compare the effect of carbohydrate availability with reduced oral intake (carbohydrate-restricted isocaloric diet: CRIC diet) or lost in urine, as urinary glucosuria on sodium/glucose cotransporter-2 inhibitor (SGLT2i) treatment, focus on the insulin requirement and the macronutrient oxidation within insulin treated type 2 diabetes. METHODS: This is randomized 3-arm open-label prospective study. Subjects treated with titrated basal-bolus insulin regimen subsequent to three diet regimens, control diet (CON), administration of canagliflozin 100 mg/day to CON (SGLT2i), or CRIC diet, with a week admission to the endocrinology ward followed by 12 weeks outpatients' management. The main outcome measures including the total insulin dose (TID) required to achieve euglycemia, fasting and postprandial energy expenditure (EE) and respiratory quotient (RQ) at 1-week and 12-week. RESULTS: We enrolled 23 patients with type 2 diabetes (male/female: 14/9, age: 53.6 ± 14.2 years, body mass index: 26.9 ± 4.8 kg/m2, HbA1c: 12.5 ± 1.6%). The TID was similar with CON and SGLT2i at both 1 and 12-weeks. Although comparable net carbohydrate availability in SGLT2i and CRIC groups, the TID was significantly higher in the CRIC (p = 0.02) compare to the SGLT2i at both 1 and 12-weeks. Fasting EE was similar in all groups, postprandial EE was significantly elevated in the SGLT2i and CRIC groups compared to the CON group (p = 0.03 and 0.04). Compare to the CON, lower basal fasting RQ (p = 0.049) and decreased delta-RQ (postprandial RQ/fasting RQ) indicated continuous lipid substrate utilization in the SGLT2i (p = 0.04) and CRIC (p = 0.03) groups. CONCLUSION: The CRIC diet resulted in a similar fasting and postprandial EE and substrate oxidation compared to the SGLT2i. The increased insulin requirement in the CRIC diet indicates that a relatively highly lipid and protein consumption, compared to the SGLT2i and CON, may influence insulin requirement.

The mutual relationship between the host immune system and radiotherapy: stimulating the action of immune cells by irradiation.

The effects of irradiation on tumor tissue and the host immune system are interrelated. The antitumor effect of irradiation is attenuated in the immunocompromised hosts. In addition, radiation alone positively and negatively influences the host immune system. The positive effects of radiation are summarized by the ability to help induce and enhance tumor-antigen-specific immune responses. The cancer-immunity cycle is a multistep framework that illustrates how the tumor-antigen-specific immune responses are induced and how the induced antigen-specific immune cells exert their functions in tumor tissues. Irradiation affects each step of this cancer-immunity cycle, primarily in a positive manner. In contrast, radiation also has negative effects on the immune system. The first is that irradiation has the possibility to kill irradiated effector immune cells. The second is that irradiation upregulates immunosuppressive molecules in the tumor microenvironment, whereas the third is that irradiation to the tumor condenses immunosuppressor cells in the tumor microenvironment. When used in conjunction with radiotherapy, immune checkpoint inhibitors can further leverage the positive effects of radiation on the immune system and compensate for the negative effects of irradiation, which supports the rationale for the combination of radiotherapy and immune checkpoint inhibitors. In this review, we summarize the preclinical evidence for the reciprocal effects of radiation exposure and the immune system, and up-front topics of the combination therapy of immune checkpoint inhibitors and radiotherapy.

Development of Artificial System to Induce Chromatin Loosening in Saccharomyces cerevisiae.

In eukaryotic cells, loosening of chromatin causes changes in transcription and DNA replication. The artificial conversion of tightly packed chromatin (heterochromatin) to loosely packed chromatin (euchromatin) enables gene expression and regulates cell differentiation. Although some chemicals convert chromatin structures through histone modifications, they lack sequence specificity. This study attempted to establish a novel technology for inducing chromatin loosening in target regions of Saccharomyces cerevisiae. We focused on histone acetylation, which is one of the mechanisms of euchromatin induction. The sequence-recognizing ability of the dead Cas9 (dCas9) and guide RNA (gRNA) complex was used to promote histone acetylation at a targeted genomic locus. We constructed a plasmid to produce a fusion protein consisting of dCas9 and histone acetyltransferase Gcn5 and a plasmid to express gRNA recognizing the upstream region of heterochromatic URA3. Confocal microscopy revealed that the fusion proteins were localized in the nucleus. The yeast strain producing the fusion protein and gRNA grew well in the uracil-deficient medium, while the strain harboring empty plasmids or the strain containing the mutations that cause loss of nucleosomal histone acetylation activity of Gcn5 did not. This suggests that the heterochromatin was loosened as much as euchromatin through nucleosomal histone acetylation. The amount of euchromatic DNA at the target locus increased, indicating that chromatin loosening was induced by our system. Nucleosomal histone acetylation in heterochromatic loci by our developed system is a promising method for inducing euchromatic state in a target locus.

Quantitative evaluation of insulin-induced abdominal subcutaneous dystrophic tissue using shear wave elastography.

AIMS/INTRODUCTION: Subcutaneous dystrophic tissue (DT) produced by insulin injection causes dysglycemia owing to inadequate absorption of insulin. However, precise techniques for measuring DT have not been established. Shear wave elastography (SWE) is an imaging technology that can quantify tissue stiffness. In this study, insulin injection-induced DT was quantified using SWE to generate whole-abdominal wall subcutaneous tissue by three-dimensional (3D) imaging in patients with type 2 diabetes who were treated with multiple insulin injections. MATERIALS AND METHODS: Seven patients with type 2 diabetes were recruited who received long-standing multiple insulin injections. Using SWE, the shear wave velocity (SWV) of DT and control (normal subcutaneous tissue) was measured. Furthermore, two of seven patients underwent whole-abdominal SWE examination to calculate the proportion of DT. A subcutaneous insulin tolerance test was also performed in both the DT and control tissues. RESULTS: The SWV in DT was significantly higher than that in the control tissue (2.87 [2.66-2.98] vs 1.29 [1.23-1.44] m/s, P < 0.01). The proportion of the DT volume was 0.67% and 5.21% for two individuals from the entire abdominal subcutaneous tissue volume. The area under the curve for the subcutaneously injected insulin aspart concentration at the DT sites was lower than that of the control tissue (75.0 [52.1-111] vs 116 [86.9-152.5] h*mU/L, P = 0.1). CONCLUSIONS: SWE can be useful in quantifying abdominal subcutaneous insulin-induced DT, especially the 3D volume of insulin injection-induced DT from the entire abdominal subcutaneous tissue. This study is the first to examine the volume and distribution of abdominal subcutaneous DT using SWE.

Pre-Existing Diabetes Limits Survival Rate After Immune Checkpoint Inhibitor Treatment for Advanced Lung Cancer: A Retrospective Study in Japan.

BACKGROUND: Although immune checkpoint inhibitors (ICIs) are promising in the treatment of advanced cancer, their use is associated with immune-related adverse events (irAEs) that affect endocrine organ systems. Although development of irAEs was associated with improved cancer-specific survival, the risk of irAEs is unclear. We investigated the association of pre-ICI comorbidities-including diabetes-with irAEs, overall survival (OS), and progression-free survival (PFS) in advanced lung cancer. METHODS: Patients with lung cancer who were treated with ICIs during the period from September 1, 2015 through July 31, 2018 were retrospectively enrolled. All data were collected from the NEPTUNE database of university patients. Hazard ratios were estimated by using Cox regression weighted for propensity scores. Odds ratios were calculated by logistic regression and adjusted for unbalanced variables. The Kaplan-Meier method was used to compare OS, and the generalized Wilcoxon test was used to compare median survival. RESULTS: Among the 88 patients identified, 22 (25.0%) had diabetes (DM) before ICI treatment and 57 (75.0%) did not (non-DM); irAEs developed in 12.2% of patients with DM and in 9.1% of patients in non-DM (p=0.87). Diabetes status was not associated with irAE risk in relation to baseline characteristics (age, sex, TNM staging, thyroid and renal function) or in propensity score-matched analysis (age, TNM staging). During a mean follow-up of 30 months, OS and cancer-specific PFS were significantly higher in patients who developed irAEs (Kaplan-Meier estimates, p=0·04 and 0·03, respectively). In propensity score-matched analysis, diabetes was significantly associated with lower OS (multivariate hazard ratio, 0·36; 95% CI, 0·13-0·98) unrelated to irAEs. Irrespective of irAEs, PFS was also lower among patients with DM than among non-DM patients (Kaplan-Meier estimate, p=0·04). CONCLUSION: Pre-existing diabetes was associated with higher mortality in advanced lung cancer, regardless of irAE development during treatment with ICI.

Whole brain radiation therapy for primary central nervous system marginal zone lymphoma: a case report.

A standard radiation therapy protocol for primary central nervous system marginal zone lymphoma (CNS-MZL) has not been established. The International Lymphoma Radiation Oncology Group suggested a radiation therapy dose of 30-36 Gy for lesions of well-defined CNS-MZL. We report a case of relatively low-dose whole brain radiation therapy (WBRT) for ill-defined CNS-MZL. A 56-year-old man who presented with sudden left-sided convulsions and impaired consciousness was diagnosed with CNS-MZL. The tumor had an ill-defined lesion, without cerebrospinal fluid involvement. WBRT, consisting of 25.2 Gy in 14 fractions, was administered owing to the difficulty in target delineation for focal radiation therapy. No chemotherapy was administered during the treatment course. After the 36-month follow-up period, the patient maintained complete remission without neurological disorders. This report describes the usefulness of relatively low-dose WBRT for ill-defined CNS-MZL.

Long-term clinical outcomes of salvage pelvic radiation therapy for oligo-recurrent pelvic lymph nodes after definitive external-beam radiation therapy for non-metastatic prostate cancer.

Although salvage external-beam radiation therapy (EBRT) is an attractive treatment option for pelvic lymph nodal recurrence (PeNR) in patients with prostate cancer (PCa), limited data are available regarding its long-term efficacy. This study examined the long-term clinical outcomes of patients who underwent salvage pelvic radiation therapy (sPRT) for oligo-recurrent pelvic lymph nodes after definitive EBRT for non-metastatic PCa. Patients who developed PeNR after definitive EBRT and were subsequently treated with sPRT at our institution between November 2007 and December 2015 were retrospectively analyzed. The prescribed dose was 45-50.4 Gy (1.8-2 Gy per fraction) to the upper pelvis, with up to 54-66 Gy (1.8-2 Gy per fraction) for recurrent nodes. Long-term hormonal therapy was used as neoadjuvant and/or adjuvant therapy. The study population consisted of 12 consecutive patients with PeNR after definitive EBRT (median age: 73 years). The median follow-up period was 58.9 months. The 5-year overall survival, PCa-specific survival, biochemical failure-free, clinical failure-free, and castration-resistant PCa-free rates were 82.5, 100.0, 62.3, 81.8, and 81.8%, respectively. No grade 2 or higher sPRT-related late toxicities occurred. In conclusion, more than half of the study patients treated with sPRT had a long-term disease-free status with acceptable morbidities. Moreover, most of the patients maintained hormonal sensitivity. Therefore, this approach may be a promising treatment method for oligo-recurrent pelvic lymph nodes.

Taxonomic revision of Pseudolachnea and Pseudolachnella and establishment of Neopseudolachnella and Pseudodinemasporium gen. nov.

The taxonomy of Pseudolachnea and Pseudolachnella is controversial. Some authors have regarded them as congeneric, whereas others have considered them to be distinct genera differentiated merely on the number of conidial septa. A total of 26 isolates of Pseudolachnea-like fungi were subjected to morphological examination and phylogenetic analyses of nuc rDNA internal transcribed spacers 1 and 2 and partial 28S sequences and partial sequence of the translation elongation factor 1α gene. The results indicated that our materials should be classified in four genera: Pseudolachnea, Pseudolachnella, and two new genera, Neopseudolachnella and Pseudodinemasporium. Although the monophyly of both Pseudolachnea and Pseudolachnella was confirmed, it was concluded that differences observed in the conidiomatal structure, such as thickness of basal stroma and the excipulum, were more reliable for their circumscription, instead of conidial septation. Neopseudolachnella was similar to Pseudolachnea and Pseudolachnella in conidial morphology but was characterized by the conidiomata lacking an excipulum, unlike members of the latter two genera. Pseudodinemasporium bore conidia morphologically similar to those of Dinemasporium but was differentiated from the latter by the conidiomata, which was composed of a well developed peridial structure. A total of 12 new species, namely three in Neopseudolachnella (N. acutispora, N. magnispora, N. uniseptata), one in Pseudodinemasporium (P. fabiforme) and eight in Pseudolachnella (P. asymmetrica, P. botulispora, P. brevicoronata, P. campylospora, P. complanata, P. falcatispora, P. fusiformis and P. pachyderma) are described and illustrated.

An important role of endothelial hairy-related transcription factors in mouse vascular development.

The Hairy-related transcription factor family of Notch- and ALK1-downstream transcriptional repressors, called Hrt/Hey/Hesr/Chf/Herp/Gridlock, has complementary and indispensable functions for vascular development. While mouse embryos null for either Hrt1/Hey1 or Hrt2/Hey2 did not show early vascular phenotypes, Hrt1/Hey1; Hrt2/Hey2 double null mice (H1(ko) /H2(ko) ) showed embryonic lethality with severe impairment of vascular morphogenesis. It remained unclear, however, whether Hrt/Hey functions are required in endothelial cells or vascular smooth muscle cells. In this study, we demonstrate that mice with endothelial-specific deletion of Hrt2/Hey2 combined with global Hrt1/Hey1 deletion (H1(ko) /H2(eko) ) show abnormal vascular morphogenesis and embryonic lethality. Their defects were characterized by the failure of vascular network formation in the yolk sac, abnormalities of embryonic vascular structures and impaired smooth muscle cell recruitment, and were virtually identical to the H1(ko) /H2(ko) phenotypes. Among signaling molecules implicated in vascular development, Robo4 expression was significantly increased and activation of Src family kinases was suppressed in endothelial cells of H1(ko) /H2(eko) embryos. The present study indicates an important role of Hrt1/Hey1 and Hrt2/Hey2 in endothelial cells during early vascular development, and further suggests involvement of Robo4 and Src family kinases in the mechanisms of embryonic vascular defects caused by the Hrt/Hey deficiency.

Tmem100, an ALK1 receptor signaling-dependent gene essential for arterial endothelium differentiation and vascular morphogenesis.

Members of the transforming growth factor-ß superfamily play essential roles in various aspects of embryonic development and physiological organ function. Among them, bone morphogenetic protein (BMP) 9 and BMP10 regulate embryonic vascular development by activating their endothelial receptor ALK1 (activin receptor-like kinase 1, also called Acvrl1). ALK1-mediated intracellular signaling is implicated in the etiologies of human diseases, but their downstream functional proteins are largely unknown. In this study, we identified Tmem100, a gene encoding a previously uncharacterized intracellular transmembrane protein, to be an embryonic endothelium-enriched gene activated by BMP9 and BMP10 through the ALK1 receptor. Tmem100 null mice showed embryonic lethality due to impaired differentiation of arterial endothelium and defects of vascular morphogenesis, which phenocopied most of the vascular abnormalities observed with the Acvrl1/Alk1 deficiency. The activity of Notch- and Akt-mediated signaling, which is essential for vascular development, was down-regulated in Tmem100 null mice. Cre-mediated deletion of Tmem100 in endothelial cells was sufficient to recapitulate the null phenotypes. These data indicated that TMEM100 may play indispensable roles downstream of BMP9/BMP10-ALK1 signaling during endothelial differentiation and vascular morphogenesis.