Comprehensive analysis of non-selective and selective autophagy in yeast atg mutants and characterization of autophagic activity in the absence of the Atg8 conjugation system.

Most autophagy-related genes, or ATG genes, have been identified in studies using budding yeast. Although the functions of the ATG genes are well understood, the contributions of individual genes to non-selective and various types of selective autophagy remain to be fully elucidated. In this study, we quantified the activity of non-selective autophagy, the cytoplasm-to-vacuole targeting (Cvt) pathway, mitophagy, endoplasmic reticulum (ER)-phagy, and pexophagy in all Saccharomyces cerevisiae atg mutants. Among the mutants of the core autophagy genes considered essential for autophagy, the atg13 mutant and mutants of the genes involved in the two ubiquitin-like conjugation systems retained residual autophagic functionality. In particular, mutants of the Atg8 ubiquitin-like conjugation system (the Atg8 system) exhibited substantial levels of non-selective autophagy, the Cvt pathway, and pexophagy, although mitophagy and ER-phagy were undetectable. Atg8-system mutants also displayed intravacuolar vesicles resembling autophagic bodies, albeit at significantly reduced size and frequency. Thus, our data suggest that membranous sequestration and vacuolar delivery of autophagic cargo can occur in the absence of the Atg8 system. Alongside these findings, the comprehensive analysis conducted here provides valuable datasets for future autophagy research.

Marked gallbladder wall thickening caused by Epstein-Barr virus-induced infectious mononucleosis.

Key Clinical Message: In patients with symptoms of viral infection and marked thickening of the gallbladder wall, it is important to suspect acalculous cholecystitis due to Epstein-Barr virus-induced infectious mononucleosis. Abstract: A 35-year-old Japanese man presented with fever, abdominal right upper quadrant pain, and liver dysfunction. Positive immunoglobulin M and -G antibodies and negative nuclear antigen for Epstein-Barr virus were observed. Abdominal ultrasonography revealed a markedly thickened gallbladder wall. Acalculous cholecystitis due to Epstein-Barr virus-induced infectious mononucleosis was diagnosed.

How to Overcome the Barriers Behind Writing Case Reports for Beginners and Young General Physicians.

Background: Case reports are fundamental to clinical medicine that trace back to ancient Egypt preceding Hippocrates in the history of medicine. Case reports contribute to academic development and new clinical research. However, among cases presented at an annual academic conference for Japanese generalists, only a few cases were later published in peer-reviewed journals, highlighting potential barriers regarding the writing of case reports, such as mentorship absence. This paper aimed to offer guidance and strategies to novice and young general physicians in overcoming barriers and effectively composing case reports for international peer-reviewed journals. Methods: This paper focuses on case reports for general physicians with extensive experience in writing case reports for international peer-reviewed journals. We conducted a narrative review to help beginners and young general physicians in writing case reports and discussed strategies for overcoming these barriers. Results: We propose the following three tips as important processes for writing case reports: recognize the types of suitable cases for case reports; select a journal for submission using a list of candidate journals for general physicians; and organize the discussion section with one theme per paragraph. In addition, we provide a list of journals that specifically focus on case reports, along with important pointers for beginners and young general physicians that will assist authors in the field of general medicine in choosing appropriate journals for submission. Conclusion: We hope that understanding and applying these tips will aid beginners and young general physicians in writing case reports.

Atg44/Mdi1/mitofissin facilitates Dnm1-mediated mitochondrial fission.

Mitochondria undergo fission and fusion, and their coordinated balance is crucial for maintaining mitochondrial homeostasis. In yeast, the dynamin-related protein Dnm1 is a mitochondrial fission factor acting from outside the mitochondria. We recently reported the mitochondrial intermembrane space protein Atg44/mitofissin/Mdi1/Mco8 as a novel fission factor, but the relationship between Atg44 and Dnm1 remains elusive. Here, we show that Atg44 is required to complete Dnm1-mediated mitochondrial fission under homeostatic conditions. Atg44-deficient cells often exhibit enlarged mitochondria with accumulated Dnm1 and rosary-like mitochondria with Dnm1 foci at constriction sites. These mitochondrial constriction sites retain the continuity of both the outer and inner membranes within an extremely confined space, indicating that Dnm1 is unable to complete mitochondrial fission without Atg44. Moreover, accumulated Atg44 proteins are observed at mitochondrial constriction sites. These findings suggest that Atg44 and Dnm1 cooperatively execute mitochondrial fission from inside and outside the mitochondria, respectively.Abbreviation: ATG: autophagy related; CLEM: correlative light and electron microscopy; EM: electron microscopy; ER: endoplasmic reticulum; ERMES: endoplasmic reticulum-mitochondria encounter structure; GA: glutaraldehyde; GFP: green fluorescent protein; GTP: guanosine triphosphate: IMM: inner mitochondrial membrane; IMS: intermembrane space; OMM: outer mitochondrial membrane; PB: phosphate buffer; PBS: phosphate-buffered saline; PFA: paraformaldehyde; RFP: red fluorescent protein; WT: wild type.

Effects of a 60-Minute Lecture About Diagnostic Errors for Medical Students: A Single-Center Interventional Study.

INTRODUCTION: The danger of diagnostic errors exists in daily medical practice, and doctors are required to avoid such errors as much as possible. Although various factors, including cognitive, system-related, and patient-related factors, are involved in the occurrence of diagnostic errors, the percentage of doctors with insufficient medical knowledge among those factors is extremely low. Therefore, lectures on diagnostic errors might also be useful for medical students without experience working as doctors. This study investigated whether a 60-minute lecture on diagnostic errors would enable Japanese medical students to consider the factors involved in diagnostic errors and how their perceptions of diagnostic errors change. METHODS AND MATERIALS: This single-center interventional study was conducted in October 2022 among fourth-year medical students at the Faculty of Medicine, Saga University. A questionnaire survey was conducted before and immediately after the lecture to investigate changes in the perceptions of medical students regarding diagnostic errors. One mock case question was given on an exam the day after the lecture, and the number of responses to cognitive biases and system-related and patient-related factors involved in diagnostic errors were calculated. RESULTS: A total of 83 students were analyzed. After the lecture, medical students were significantly more aware of the existence of the concept of diagnostic error, the importance of learning about it, their willingness to continue learning about it, and their perception that learning about diagnostic errors improves their clinical skills. They were also significantly less likely to feel blame or shame over diagnostic errors. The mean numbers of responses per student for cognitive bias, system-related factors, and patient-related factors were 1.9, 3.4, and 0.9, respectively. The mean number of responses per student for all factors was 5.6. CONCLUSION: A 60-minute lecture on diagnostic errors among medical students is beneficial because it significantly changes their perception of diagnostic errors. The results of the present study also suggest that lectures may enable Japanese medical students to consider the factors involved in diagnostic errors.

Mitophagy mediated by BNIP3 and NIX protects against ferroptosis by downregulating mitochondrial reactive oxygen species.

Mitophagy plays an important role in the maintenance of mitochondrial homeostasis and can be categorized into two types: ubiquitin-mediated and receptor-mediated pathways. During receptor-mediated mitophagy, mitophagy receptors facilitate mitophagy by tethering the isolation membrane to mitochondria. Although at least five outer mitochondrial membrane proteins have been identified as mitophagy receptors, their individual contribution and interrelationship remain unclear. Here, we show that HeLa cells lacking BNIP3 and NIX, two of the five receptors, exhibit a complete loss of mitophagy in various conditions. Conversely, cells deficient in the other three receptors show normal mitophagy. Using BNIP3/NIX double knockout (DKO) cells as a model, we reveal that mitophagy deficiency elevates mitochondrial reactive oxygen species (mtROS), which leads to activation of the Nrf2 antioxidant pathway. Notably, BNIP3/NIX DKO cells are highly sensitive to ferroptosis when Nrf2-driven antioxidant enzymes are compromised. Moreover, the sensitivity of BNIP3/NIX DKO cells is fully rescued upon the introduction of wild-type BNIP3 and NIX, but not the mutant forms incapable of facilitating mitophagy. Consequently, our results demonstrate that BNIP3 and NIX-mediated mitophagy plays a role in regulating mtROS levels and protects cells from ferroptosis.

Writing Case Reports Can Improve Seven Components in Clinical Reasoning.

Case reports provide scientific knowledge and opportunities for new clinical research. However, it is estimated that less than 5% of cases presented by Japanese generalists at academic conferences are published due to various barriers such as the complex process of writing articles, conducting literature searches, the significant time required, the reluctance to write in English, and the challenge of selecting appropriate journals for publication. Therefore, the purpose of this opinion paper is to provide clinicians with practical tips for writing case reports that promote diagnostic excellence. In recent years, clinical practitioners have been striving for diagnostic excellence and optimal methods to accurately and comprehensively understand the patient's condition. To write a case report, it is essential to be mindful of the elements of diagnostic excellence and consider the quality of the diagnostic reasoning process. We (the authors) are seven academic generalists who are members of the Japanese Society of Hospital General Medicine (JSHGM) - Junior Doctors Association, with a median of 7 years after graduation and extensive experience publishing case reports in international peer-reviewed journals. We conducted a narrative review and discussed ways to write case reports to promote diagnostic excellence, leveraging our unique perspectives as academic generalists. Our review did not identify any reports addressing the critical points in writing case reports that embody diagnostic excellence. Therefore, this report proposes a methodology that describes the process involved in writing diagnostic excellence-promoting case reports and provides an overview of the lessons learned. Based on our review and discussion, we explain the essential points for promoting diagnostic excellence through case reports categorized into seven components of clinical reasoning. These strategies are useful in daily clinical practice and instrumental in promoting diagnostic excellence through case reports.

Imeglimin mitigates the accumulation of dysfunctional mitochondria to restore insulin secretion and suppress apoptosis of pancreatic ß-cells from db/db mice.

Mitochondrial dysfunction in pancreatic ß-cells leads to impaired glucose-stimulated insulin secretion (GSIS) and type 2 diabetes (T2D), highlighting the importance of autophagic elimination of dysfunctional mitochondria (mitophagy) in mitochondrial quality control (mQC). Imeglimin, a new oral anti-diabetic drug that improves hyperglycemia and GSIS, may enhance mitochondrial activity. However, chronic imeglimin treatment's effects on mQC in diabetic ß-cells are unknown. Here, we compared imeglimin, structurally similar anti-diabetic drug metformin, and insulin for their effects on clearance of dysfunctional mitochondria through mitophagy in pancreatic ß-cells from diabetic model db/db mice and mitophagy reporter (CMMR) mice. Pancreatic islets from db/db mice showed aberrant accumulation of dysfunctional mitochondria and excessive production of reactive oxygen species (ROS) along with markedly elevated mitophagy, suggesting that the generation of dysfunctional mitochondria overwhelmed the mitophagic capacity in db/db ß-cells. Treatment with imeglimin or insulin, but not metformin, reduced ROS production and the numbers of dysfunctional mitochondria, and normalized mitophagic activity in db/db ß-cells. Concomitantly, imeglimin and insulin, but not metformin, restored the secreted insulin level and reduced ß-cell apoptosis in db/db mice. In conclusion, imeglimin mitigated accumulation of dysfunctional mitochondria through mitophagy in diabetic mice, and may contribute to preserving ß-cell function and effective glycemic control in T2D.

Validation and Improvement of the Saga Fall Risk Model: A Multicenter Retrospective Observational Study.

Purpose: We conducted a pilot study in an acute care hospital and developed the Saga Fall Risk Model 2 (SFRM2), a fall prediction model comprising eight items: Bedriddenness rank, age, sex, emergency admission, admission to the neurosurgery department, history of falls, independence of eating, and use of hypnotics. The external validation results from the two hospitals showed that the area under the curve (AUC) of SFRM2 may be lower in other facilities. This study aimed to validate the accuracy of SFRM2 using data from eight hospitals, including chronic care hospitals, and adjust the coefficients to improve the accuracy of SFRM2 and validate it. Patients and Methods: This study included all patients aged ≥20 years admitted to eight hospitals, including chronic care, acute care, and tertiary hospitals, from April 1, 2018, to March 31, 2021. In-hospital falls were used as the outcome, and the AUC and shrinkage coefficient of SFRM2 were calculated. Additionally, SFRM2.1, which was modified from the coefficients of SFRM2 using logistic regression with the eight items comprising SFRM2, was developed using two-thirds of the data randomly selected from the entire population, and its accuracy was validated using the remaining one-third portion of the data. Results: Of the 124,521 inpatients analyzed, 2,986 (2.4%) experienced falls during hospitalization. The median age of all inpatients was 71 years, and 53.2% were men. The AUC of SFRM2 was 0.687 (95% confidence interval [CI]:0.678-0.697), and the shrinkage coefficient was 0.996. SFRM2.1 was created using 81,790 patients, and its accuracy was validated using the remaining 42,731 patients. The AUC of SFRM2.1 was 0.745 (95% CI: 0.731-0.758). Conclusion: SFRM2 showed good accuracy in predicting falls even on validating in diverse populations with significantly different backgrounds. Furthermore, the accuracy can be improved by adjusting the coefficients while keeping the model's parameters fixed.

How Do We Establish the Utility and Evidence of General Medicine in Japan?

Hospital Medicine in the United States has achieved significant progress in the accumulation of evidence. This development has influenced the increasing societal demand for General Medicine in Japan. Generalists in Japan actively engage in a wide range of interdisciplinary clinical practices, education, and management. Furthermore, Generalists have also contributed to advances in research. However, there is limited evidence regarding the benefits of General Medicine in Japan in all these areas, with most of the evidence derived from single-center studies. In Japan, the roles of Generalists are diverse, and the comprehensive definition of General Medicine makes it difficult to clearly delineate its scope. This results in an inadequate accumulation of evidence regarding the benefits of General Medicine, potentially making it less attractive to the public and younger physicians. Therefore, it is necessary to categorize General Medicine and collect clear evidence regarding its benefits.

Polybacterial Iliopsoas Muscle Abscess as an Indication for Early Diagnosis of Crohn's Disease.

BACKGROUND Crohn disease (CD) is a chronic, relapsing inflammatory bowel disease characterized by penetrations or fistulae in the gastrointestinal tract and abscesses in the surrounding tissues. Diagnosis of CD is difficult with an iliopsoas muscle abscess (IMA) as an initial presentation. CASE REPORT A 22-year-old Japanese man had right hip pain 17 days prior to admission. Because of worsening pain, he was admitted to our hospital. Physical examination revealed limitation of his right hip motion and a positive right psoas sign. Abdominal contrast-enhanced computed tomography (CT) revealed a large right IMA. Continuous drainage, which revealed polymicrobial pus, with intravenous administration of antibiotics dramatically decreased the size of the IMA. The drainage tube was removed on hospitalization day 9 because barium enema and contrast radiography of the abscess through the drainage tube showed no fistula. However, on day 19 of hospitalization, the IMA was redetected by abdominal CT. Continuous abscess drainage was resumed, and the third contrast radiograph of the abscess revealed contrast medium flow into the small intestine. Colonoscopy detected stenoses and circumferential ulceration of the terminal ileum. Histopathological examination of the ileum biopsy showed histocyte aggregation with lymphocyte or plasmacyte infiltration of the lamina propria, compatible with a CD diagnosis. Laparoscopic ileocecal resection was performed on day 64 of hospitalization. CONCLUSIONS Penetration of the intestinal tract caused by CD should be suspected in a patient with a polymicrobial IMA. It is essential to identify the fistula and subsequently perform surgical resection of the affected intestinal area.

Helicobacter cinaedi Infection Presenting with Myalgia and Cellulitis.

BACKGROUND Helicobacter cinaedi is a rare bacterium, accounting for only 0.2% of the positive isolates in blood cultures. Previous reports note that patients with H. cinaedi infection often have underlying diseases. H. cinaedi infection is diagnosed by blood culture. However, because of the slow growth of this bacterium in blood culture, the diagnosis can be missed. CASE REPORT A 78-year-old man gradually developed erythema and pain in his left arm, then left shoulder and both lower legs. The patient presented to our hospital on day 17. He was afebrile, but the examination was remarkable for tenderness in both gastrocnemius muscles and erythema from the distal left lower leg to the ankle. We suspected pyomyositis and cellulitis and started oral administration of amoxicillin-clavulanate. On day 22, H. cinaedi was detected in blood cultures. Based on these findings, we diagnosed pyogenic myositis and cellulitis caused by H. cinaedi bacteremia. On day 24, antibiotic therapy was changed to intravenous ampicillin, and symptoms improved. Additional examination did not reveal any underlying immunodeficiency disorder, such as malignancy or HIV infection. CONCLUSIONS H. cinaedi infection can occur in healthy patients. Myalgia can be caused by pyogenic myositis because of bacteremia. In cases of myalgia or cellulitis of unknown etiology, blood cultures can be useful when bacteremia is suspected; blood samples should be monitored over an extended period.

Acute abdominal pain due to atypical bilateral adrenal infarction in acute myeloid leukemia with alterations related to myelodysplasia: A case report.

Key Clinical Message: Acute myeloid leukemia (AML) can cause acute abdomen following adrenal insufficiency or adrenal infarction. Therefore, when diffusely enlarged adrenal glands and adrenal insufficiency of unknown cause are seen in a patient presenting with acute abdomen, adrenal infarction due to AML, or other hematologic diseases should be ruled out. Abstract: A 49-year-old man developed acute abdominal pain following adrenal insufficiency and was diagnosed with acute myeloid leukemia (AML) with myelodysplasia-related changes. Because AML can cause acute abdominal pain due to adrenal infarction following adrenal insufficiency, a patient with these conditions should be ruled out adrenal infarction due to AML or other hematologic diseases.

Listeria meningitis diagnosed by blood culture with fever, neurological symptoms, and no meningeal irritation signs.

Key Clinical Message: Listeria can cause neurological symptoms in immunocompromised and older patients. Additionally, it is impossible to rule out meningitis by the absence of typical meningeal irritation signs. Therefore, patients with fever and neurological impairments should be rapidly examined for blood and cerebrospinal fluid cultures to rule out Listeria meningitis. Abstract: A woman in her 90s developed fever, dysarthria, and transient disturbance of consciousness. Physical examination revealed no meningeal irritation signs. Listeria monocytogenes were detected in her blood culture the following day. Because of an increased number of cells in cerebrospinal fluid, she was diagnosed with Listeria meningitis.

Difficulty in diagnosing peritonitis caused by multidrug-resistant tuberculosis.

Key Clinical Message: The low sensitivity of ascites culture for acid-fast bacilli necessitates a peritoneal biopsy when tuberculous peritonitis is suspected. Findings in the peritoneum on computed tomography may prompt suspicion of tuberculous peritonitis. Abstract: A 47-year-old Nigerian man presented with fever, abdominal distention, and weight loss. Abdominal computed tomography revealed massive ascites and peritoneal thickening. Despite failing to culture acid-fast bacilli from ascites, histological examination and culture of peritoneum revealed multidrug-resistant tuberculosis peritonitis. Peritoneal biopsy is mandatory when tuberculosis peritonitis is suspected.

A Simple and Accurate Model for Predicting Fall Injuries in Hospitalized Patients: Insights from a Retrospective Observational Study in Japan.

BACKGROUND While several predictive models for falls have been reported such as we reported in 2020, those for fall "injury" have been unreported. This study was designed to develop a model to predict fall injuries in adult inpatients using simple predictors available immediately after hospitalization. MATERIAL AND METHODS This was a single-center, retrospective cohort study. We enrolled inpatients aged ≥20 years admitted to an acute care hospital from April 2012 to March 2018. The variables routinely obtained in clinical practice were compared between the patients with fall injury and the patients without fall itself or fall injury. Multivariable analysis was performed using covariables available on admission. A predictive model was constructed using only variables showing significant association in prior multivariable analysis. RESULTS During hospitalization of 17 062 patients, 646 (3.8%) had falls and 113 (0.7%) had fall injuries. Multivariable analysis showed 6 variables that were significantly associated with fall injuries during hospitalization: age (P=0.001), sex (P=0.001), emergency transport (P<0.001), medical referral letter (P=0.041), history of falls (P=0.012), and abnormal bedriddenness ranks (all P≤0.001). The area under the curve of this predictive model was 0.794 and the shrinkage coefficient was 0.955 using the same data set given above. CONCLUSIONS We developed a predictive model for fall injuries during hospitalization using 6 predictors, including bedriddenness ranks from official Activities of Daily Living indicators in Japan, which were all easily available on admission. The model showed good discrimination by internal validation and promises to be a useful tool to assess the risk of fall injuries.

The mitochondrial intermembrane space protein mitofissin drives mitochondrial fission required for mitophagy.

Mitophagy plays an important role in mitochondrial homeostasis by selective degradation of mitochondria. During mitophagy, mitochondria should be fragmented to allow engulfment within autophagosomes, whose capacity is exceeded by the typical mitochondria mass. However, the known mitochondrial fission factors, dynamin-related proteins Dnm1 in yeasts and DNM1L/Drp1 in mammals, are dispensable for mitophagy. Here, we identify Atg44 as a mitochondrial fission factor that is essential for mitophagy in yeasts, and we therefore term Atg44 and its orthologous proteins mitofissin. In mitofissin-deficient cells, a part of the mitochondria is recognized by the mitophagy machinery as cargo but cannot be enwrapped by the autophagosome precursor, the phagophore, due to a lack of mitochondrial fission. Furthermore, we show that mitofissin directly binds to lipid membranes and brings about lipid membrane fragility to facilitate membrane fission. Taken together, we propose that mitofissin acts directly on lipid membranes to drive mitochondrial fission required for mitophagy.

TIM23 facilitates PINK1 activation by safeguarding against OMA1-mediated degradation in damaged mitochondria.

PINK1 is activated by autophosphorylation and forms a high-molecular-weight complex, thereby initiating the selective removal of damaged mitochondria by autophagy. Other than translocase of the outer mitochondrial membrane complexes, members of PINK1-containing protein complexes remain obscure. By mass spectrometric analysis of PINK1 co-immunoprecipitates, we identify the inner membrane protein TIM23 as a component of the PINK1 complex. TIM23 downregulation decreases PINK1 levels and significantly delays autophosphorylation, indicating that TIM23 promotes PINK1 accumulation in response to depolarization. Moreover, inactivation of the mitochondrial protease OMA1 not only enhances PINK1 accumulation but also represses the reduction in PINK1 levels induced by TIM23 downregulation, suggesting that TIM23 facilitates PINK1 activation by safeguarding against degradation by OMA1. Indeed, deficiencies of pathogenic PINK1 mutants that fail to interact with TIM23 are partially restored by OMA1 inactivation. These findings indicate that TIM23 plays a distinct role in activating mitochondrial autophagy by protecting PINK1.

Hva22, a REEP family protein in fission yeast, promotes reticulophagy in collaboration with a receptor protein.

The endoplasmic reticulum (ER) undergoes selective autophagy called reticulophagy or ER-phagy. Multiple reticulon- and receptor expression enhancing protein (REEP)-like ER-shaping proteins, including budding yeast Atg40, serve as reticulophagy receptors that stabilize the phagophore on the ER by interacting with phagophore-conjugated Atg8. Additionally, they facilitate phagophore engulfment of the ER by remodeling ER morphology. We reveal that Hva22, a REEP family protein in fission yeast, promotes reticulophagy without Atg8-binding capacity. The role of Hva22 in reticulophagy can be replaced by expressing Atg40 independently of its Atg8-binding ability. Conversely, adding an Atg8-binding sequence to Hva22 enables it to substitute for Atg40 in budding yeast. Thus, the phagophore-stabilizing and ER-shaping activities, both of which Atg40 solely contains, are divided between two separate factors, receptors and Hva22, respectively, in fission yeast.Abbreviations: AIM: Atg8-family interacting motif; Atg: autophagy related; DTT: dithiothreitol; ER: endoplasmic reticulum GFP: green fluorescent protein; NAA: 1-naphthaleneacetic acid; REEP: receptor expression enhancing protein; RFP: red fluorescent protein; UPR: unfolded protein response.