Search

Summary: Background: Emergence of SARS-CoV-2 variants that escape neutralising antibodies hampers the development of vaccines and therapeutic antibodies against SARS-CoV-2. IGHV3-53/3-66-derived public antibodies, which are generally specific to the prototype virus and are frequently induced in infected or vaccinated individuals, show minimal affinity maturation and high potency against prototype SARS-CoV-2. Methods: Monoclonal antibodies isolated from a Delta breakthrough infection case were analysed for cross-neutralising activities against SARS-CoV-2 variants. The broadly neutralising antibody K4-66 was further analysed in a hamster model, and the effect of somatic hypermutations was assessed using the inferred germline precursor. Findings: Antibodies derived from IGHV3-53/3-66 showed broader neutralising activity than antibodies derived from IGHV1-69 and other IGHV genes. IGHV3-53/3-66 antibodies neutralised the Delta variant better than the IGHV1-69 antibodies, suggesting that the IGHV3-53/3-66 antibodies were further maturated by Delta breakthrough infection. One IGHV3-53/3-66 antibody, K4-66, neutralised all Omicron subvariants tested, including EG.5.1, BA.2.86, and JN.1, and decreased the viral load in the lungs of hamsters infected with Omicron subvariant XBB.1.5. The importance of somatic hypermutations was demonstrated by the loss of neutralising activity of the inferred germline precursor of K4-66 against Beta and Omicron variants. Interpretation: Broadly neutralising IGHV3-53/3-66 antibodies have potential as a target for the development of effective vaccines and therapeutic antibodies against newly emerging SARS-CoV-2 variants. Funding: This work was supported by grants from AMED (JP23ym0126048, JP22ym0126048, JP21ym0126048, JP23wm0125002, JP233fa627001, JP223fa627009, JP24jf0126002, and JP22fk0108572), and the JSPS (JP21H02970, JK23K20041, and JPJSCCA20240006).

IntroductionThe severe acute respiratory syndrome coronavirus (SARS-CoV-2) spike (S) protein is essential in mediating membrane fusion of the virus with the target cells. Several reports demonstrated that SARS-CoV-2 S protein fusogenicity is reportedly closely associated with the intrinsic pathogenicity of the virus determined using hamster models. However, the association between S protein fusogenicity and other virological parameters remains elusive.MethodsIn this study, we investigated the virological parameters (e.g., S1/S2 cleavage efficiency, plaque size, pseudoviral infectivity, pseudovirus entry efficiency, and viral replication kinetics) of eleven previous variants of concern (VOCs) and variants of interest (VOIs) correlating with S protein fusogenicity.Results and discussionS protein fusogenicity was found to be strongly correlated with S1/S2 cleavage efficiency and plaque size formed by clinical isolates. However, S protein fusogenicity was less associated with pseudoviral infectivity, pseudovirus entry efficiency, and viral replication kinetics. Taken together, our results suggest that S1/S2 cleavage efficiency and plaque size could be potential indicators to predict the intrinsic pathogenicity and S protein fusogenicity of newly emerged SARS-CoV-2 variants.

Abstract Background Despite physicians’ vital role in advance care planning, a limited number of physicians practice it. This study assessed factors associated with physicians’ knowledge, attitudes, and practices regarding advance care planning. Methods This cross-sectional study used data from an anonymous survey conducted by the Japanese Ministry of Health, Labour and Welfare. Questionnaires were mailed to 4500 physicians in November and December 2022. Data from 1260 respondents were analyzed. Results Of the respondents, 46.4%, 77.0%, and 82.0% reported good knowledge of advance care planning, agreed with promoting it, and with its provision by medical/care staff, respectively. Male physicians were significantly less likely to support advance care planning (odds ratio: 0.54, 95% confidence interval: 0.35–0.84) or agree to its provision by medical/care staff (odds ratio: 0.47, 95% confidence interval: 0.29–0.78) but significantly more likely to practice it (odds ratio: 1.58, 95% confidence interval: 1.05–2.36). Physicians specialized in surgery or internal/general/palliative medicine were more knowledgeable about advance care planning and more likely to practice it. Physicians working in clinics were significantly less knowledgeable (odds ratio: 0.33, 95% confidence interval: 0.25–0.44) about advance care planning and less likely to support it (odds ratio: 0.37, 95% confidence interval: 0.27–0.50), agree with its provision by medical/care staff (odds ratio: 0.54, 95% confidence interval: 0.39–0.75), or to practice it (odds ratio: 0.16, 95% confidence interval: 0.12–0.22). Conclusions Physicians working in clinics had less knowledge of advance care planning, less supportive attitudes, and less likely to practice it. Knowledge, attitudes and practice also varied by gender and specialty. Interventions should target physicians working in clinics.

The COVID-19 pandemic highlighted the need for advance care planning (ACP) as a way to help mitigate the various care concerns that accompanied the healthcare crisis. However, unique obstacles to typical ACP practice necessitated the need for guidance and innovation to help facilitate these vital conversations. The aim of this systematic review was to identify the various ACP barriers and facilitators that arose during the pandemic and determine how ACP practice was affected across different contexts and among different populations. This systematic review (PROSPERO registration number: CRD42022359092), which adheres to the PRISMA guidelines for reporting systematic reviews, examined studies on ACP in the context of the COVID-19 pandemic. The review involved searches of five databases, including MEDLINE and Embase. Of the 843 identified studies, 115 met the inclusion criteria. The extracted ACP barriers and facilitators were codified and quantified. The most frequently occurring ACP barrier codes were: Social distancing measures and visitation restrictions, Uncertainty surrounding the COVID-19 prognosis, and Technological/Telehealth barriers. The most frequently occurring ACP facilitator codes were the following: Telehealth/virtual ACP platforms, Training for clinicians, and Care team collaboration. Identifying the ACP barriers and facilitators is essential for developing effective, resilient ACP promotion strategies and improving its delivery, accessibility, and acceptability.

Objective The COVID-19 pandemic has impacted the capacity for advance care planning (ACP) among patients, families and healthcare teams. We sought to identify and review the barriers to and facilitators of ACP implementation for medical staff in different settings (eg, hospitals, outpatient palliative care, nursing and care homes) during the pandemic.Design This study employed an overview of reviews design. We searched the MEDLINE, CENTRAL, Web of Science and Embase databases for studies published between 8 December 2019 and 30 July 2023. We used AMSTAR 2 to assess the risk of bias.Results We included seven reviews. Common barriers to ACP implementation included visitation restrictions, limited resources and personnel and a lack of coordination among healthcare professionals. In care and nursing homes, barriers included a dearth of palliative care physicians and the psychological burden on facility staff. Using telemedicine for information sharing was a common facilitator across settings. In hospitals, facilitators included short-term training in palliative care and palliative care physicians joining the acute care team. In care and nursing homes, facilitators included ACP education and emotional support for staff.Conclusions Visitation restrictions and limited resources during the pandemic posed obstacles; however, the implementation of ACP was further hindered by insufficient staff education on ACP in hospitals and facilities, as well as a scarcity of information sharing at the community level. These pre-existing issues were magnified by the pandemic, drawing attention to their significance. Short-term staff training programmes and immediate information sharing could better enable ACP.PROSPERO registration number CRD42022351362.

Background Undertriaged patients have worse outcomes than appropriately triaged patients. Machine learning provides better triage prediction than conventional triage in emergency departments, but no machine learning-based undertriage prediction models have yet been developed for prehospital telephone triage. We developed and validated machine learning models for telephone triage.Materials and methods We conducted a retrospective cohort study with the largest after-hour house-call (AHHC) service dataset in Japan. Participants were ≥16 years and used the AHHC service between 1 November 2018 and 31 January 2021. We developed five prediction models based on support vector machine (SVM), lasso regression (LR), random forest (RF), gradient-boosted decision tree (XGB), and deep neural network (DNN). The primary outcome was undertriage, and predictors were telephone triage level and routinely available telephone-based data, including age, sex, 80 chief complaint categories and 10 comorbidities. We measured the area under the receiver operating characteristic curve (AUROC) for all the models.Results We identified 15,442 eligible patients (age: 38.4 ± 16.6, male: 57.2%), including 298 (1.9%; age: 58.2 ± 23.9, male: 55.0%) undertriaged patients. RF and XGB outperformed the other models, with the AUROC values (95% confidence interval; 95% CI) of the SVM, LR, RF, XGB and DNN for undertriage being 0.62 (0.55–0.69), 0.79 (0.74–0.83), 0.81 (0.76–0.86), 0.80 (0.75–0.84) and 0.77 (0.73–0.82), respectively.Conclusions We found that RF and XGB outperformed other models. Our findings suggest that machine learning models can facilitate the early detection of undertriage and early intervention to yield substantially improved patient outcomes.KEY MESSAGESUndertriaged patients experience worse outcomes than appropriately triaged patients; thus, we developed machine learning models for predicting undertriage in the prehospital setting. In addition, we identified the predictors of risk factors associated with undertriage.Random forest and gradient-boosted decision tree models demonstrated better prediction performance, and the models identified the risk factors associated with undertriage.Machine learning models aid in the early detection of undertriage, leading to significantly improved patient outcomes and identifying undertriage-associated risk factors, including chief complaint categories, could help prioritize conventional telephone triage protocol revision.

Bronchial organoids (BO) and BO-derived air-liquid interface models (BO-ALI) are used to model SARS-CoV-2 infection of human bronchial epithelial cells, reporting that ciliated cells were permissive to the virus, unlike basal cells.

It has been reported that many receptors and proteases are required for severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection. Although angiotensin-converting enzyme 2 (ACE2) is the most important of these receptors, little is known about the contribution of other genes. In this study, we examined the roles of neuropilin-1, basigin, transmembrane serine proteases (TMPRSSs), and cathepsins (CTSs) in SARS-CoV-2 infection using the CRISPR interference system and ACE2-expressing human induced pluripotent stem (iPS) cells. Double knockdown of TMPRSS2 and cathepsin B (CTSB) reduced the viral load to 0.036% ± 0.021%. Consistently, the combination of the CTPB inhibitor CA-074 methyl ester and the TMPRSS2 inhibitor camostat reduced the viral load to 0.0078% ± 0.0057%. This result was confirmed using four SARS-CoV-2 variants (B.1.3, B.1.1.7, B.1.351, and B.1.1.248). The simultaneous use of these two drugs reduced viral load to less than 0.01% in both female and male iPS cells. These findings suggest that compounds targeting TMPRSS2 and CTSB exhibit highly efficient antiviral effects independent of gender and SARS-CoV-2 variant.

Summary: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused an epidemic and spread rapidly all over the world. Because the analysis of host factors other than receptors and proteases has not been sufficiently performed, we attempted to identify and characterize host factors essential for SARS-CoV-2 infection using iPS cells and airway organoids (AO). Based on previous CRISPR screening and RNA-seq data, we found that exocyst complex component 2 (EXOC2) is one important host factor for SARS-CoV-2 infection. The intracellular SARS-CoV-2 nucleocapsid (N) expression level was decreased to 3.7% and the virus copy number in cell culture medium was decreased to 1.6% by EXOC2 knockdown. Consistently, immunostaining results showed that N protein-positive cells were significantly decreased by EXOC2 knockdown. We also found that EXOC2 knockdown downregulates SARS-CoV-2 infection by regulating IFNW1 expression. In conclusion, controlling the EXOC2 expression level may prevent SARS-CoV-2 infection and deserves further study.

Abstract The Japanese Clinical Practice Guidelines for Management of Sepsis and Septic Shock 2020 (J-SSCG 2020), a Japanese-specific set of clinical practice guidelines for sepsis and septic shock created as revised from J-SSCG 2016 jointly by the Japanese Society of Intensive Care Medicine and the Japanese Association for Acute Medicine, was first released in September 2020 and published in February 2021. An English-language version of these guidelines was created based on the contents of the original Japanese-language version. The purpose of this guideline is to assist medical staff in making appropriate decisions to improve the prognosis of patients undergoing treatment for sepsis and septic shock. We aimed to provide high-quality guidelines that are easy to use and understand for specialists, general clinicians, and multidisciplinary medical professionals. J-SSCG 2016 took up new subjects that were not present in SSCG 2016 (e.g., ICU-acquired weakness [ICU-AW], post-intensive care syndrome [PICS], and body temperature management). The J-SSCG 2020 covered a total of 22 areas with four additional new areas (patient- and family-centered care, sepsis treatment system, neuro-intensive treatment, and stress ulcers). A total of 118 important clinical issues (clinical questions, CQs) were extracted regardless of the presence or absence of evidence. These CQs also include those that have been given particular focus within Japan. This is a large-scale guideline covering multiple fields; thus, in addition to the 25 committee members, we had the participation and support of a total of 226 members who are professionals (physicians, nurses, physiotherapists, clinical engineers, and pharmacists) and medical workers with a history of sepsis or critical illness. The GRADE method was adopted for making recommendations, and the modified Delphi method was used to determine recommendations by voting from all committee members. As a result, 79 GRADE-based recommendations, 5 Good Practice Statements (GPS), 18 expert consensuses, 27 answers to background questions (BQs), and summaries of definitions and diagnosis of sepsis were created as responses to 118 CQs. We also incorporated visual information for each CQ according to the time course of treatment, and we will also distribute this as an app. The J-SSCG 2020 is expected to be widely used as a useful bedside guideline in the field of sepsis treatment both in Japan and overseas involving multiple disciplines.

Summary: Genetic differences are a primary reason for differences in the susceptibility and severity of COVID-19. As induced pluripotent stem (iPS) cells maintain the genetic information of the donor, they can be used to model individual differences in SARS-CoV-2 infection in vitro. We found that human iPS cells expressing the SARS-CoV-2 receptor angiotensin-converting enzyme 2 (ACE2) (ACE2-iPS cells) can be infected w SARS-CoV-2. In infected ACE2-iPS cells, the expression of SARS-CoV-2 nucleocapsid protein, budding of viral particles, and production of progeny virus, double membrane spherules, and double-membrane vesicles were confirmed. We performed SARS-CoV-2 infection experiments on ACE2-iPS/ embryonic stem (ES) cells from eight individuals. Male iPS/ES cells were more capable of producing the virus compared with female iPS/ES cells. These findings suggest that ACE2-iPS cells can not only reproduce individual differences in SARS-CoV-2 infection in vitro but also are a useful resource to clarify the causes of individual differences in COVID-19 due to genetic differences.

Human hepatocytes are essential materials in pharmaceutical researches. Not only primary human hepatocytes (PHH) but also human iPS cell-derived hepatocyte-like cells (human iPS-HLCs) are expected to be applied as materials for pharmaceutical researches. To date, several culture media have been developed for culturing human hepatocytes. However, there have been no reports comparing these media to determine which is most suitable for culturing human hepatocytes. In this study, we compared five commercial media (Hepatocyte Culture Medium (HCM), HepatoZYME-SFM, Cellartis Power Primary HEP Medium, DMEM/F12, and William's E Medium (WEM)) to determine which is most suitable for culturing PHH and human iPS-HLCs. In hepatic differentiation of human iPS cells (day 14-25 of differentiation), albumin (ALB) and urea secretion abilities and CYP2C9, CYP2C19, and CYP3A4 activities were the highest when using HCM or WEM. During maintenance of human iPS-HLCs, ALB and urea producing abilities and CYP2C9, CYP2C19, and CYP3A4 activities were the highest when using HCM. Importantly, we found that human iPS-HLCs cultured in HCM were maintained for 3 weeks or more without impairment of their hepatic functions. These results suggest that it is necessary to select an optimal medium for hepatic differentiation and maintenance of human iPS-HLCs. In the case of PHH culture, there was little difference in hepatic functions among the five media. However, the CYP2C9, CYP2C19, and CYP3A4 activities were the highest when using HCM and WEM. In conclusion, it is important to select the optimal medium for specific application when carrying out pharmaceutical researches using human hepatocytes.