Association of Placental Pathology with Physical and Neuronal Development of Infants: A Narrative Review and Reclassification of the Literature by the Consensus Statement of the Amsterdam Placental Workshop Group.

The placenta is the largest fetal organ, which connects the mother to the fetus and supports most aspects of organogenesis through the transport of nutrients and gases. However, further studies are needed to assess placental pathology as a reliable predictor of long-term physical growth or neural development in newborns. The Consensus Statement of the Amsterdam Placental Workshop Group (APWGCS) on the sampling and definition of placental lesions has resulted in diagnostic uniformity in describing the most common pathological lesions of the placenta and contributed to the international standardization of descriptions of placental pathology. In this narrative review, we reclassified descriptions of placental pathology from previously published papers according to the APWGCS criteria and comparatively assessed the relationship with infantile physical and/or neural development. After reclassification and reevaluation, placental pathology of maternal vascular malperfusion, one of the APWGCS criteria, emerged as a promising candidate as a universal predictor of negative infantile neurodevelopmental outcomes, not only in term and preterm deliveries but also in high-risk groups of very low birthweight newborns. However, there are few studies that examined placental pathology according to the full categories of APWGCS and also included low-risk general infants. It is necessary to incorporate the assessment of placental pathology utilizing APWGCS in the design of future birth cohort studies as well as in follow-up investigations of high-risk infants.

Interleukin-23 levels in umbilical cord blood are associated with neurodevelopmental trajectories in infancy.

Our previous study, which aimed to understand the early neurodevelopmental trajectories of children with and without neurodevelopmental disorders, identified five classes of early neurodevelopmental trajectories, categorized as high normal, normal, low normal, delayed, and markedly delayed. This investigation involved measurement using the Mullen Scale of Early Learning in a representative sample of Japanese infants followed up from the age of 0 to 2 years (Nishimura et al., 2016). In the present study, we investigated the potential association between cytokine concentrations in umbilical cord serum with any of the five classes of neurodevelopmental trajectories previously assigned, as follows: high normal (N = 85, 13.0%), normal (N = 322, 49.1%), low normal (N = 137, 20.9%), delayed (N = 87, 13.3%), and markedly delayed (N = 25, 3.8%) in infancy. Decreased interleukin (IL)-23 levels in the cord blood were associated with the markedly delayed class, independent of potential confounders (odds ratio, 0.44; 95%confidence interval: 0.26-0.73). Furthermore, IL-23 levels decreased as the developmental trajectory became more delayed, demonstrating that IL-23 plays an important role in development, and is useful for predicting the developmental trajectory at birth.

A Case of Consumptive Coagulopathy Before Cardiopulmonary Failure in Amniotic Fluid Embolism and Review of Literature: A Perspective of the Latent Onset and Progression of Coagulopathy.

Amniotic fluid embolism (AFE) induces cardiopulmonary insufficiency with consumptive coagulopathy. Previous studies reported that refractory coagulopathy has already advanced at the onset of maternal cardiovascular and/or respiratory symptoms. However, when the consumption of coagulation factors starts during the clinical course, AFE remains to be elucidated. We report an intrapartum AFE case of consumptive coagulopathy before dyspnea with hypotension developing during urgent cesarean delivery that was revealed by non-reassuring fetal heart rate tracing. The patient, a 42-year-old multiparous parturient, underwent induced labor after a premature rupture of membranes in week 39 of pregnancy. Coagulation screening was initially within the normal range. Fetal heart rate monitoring demonstrated bradycardia coincided with uterine tachysystole after three hours, which required urgent cesarean section with preoperative blood screening. The hemoglobin level was maintained at 129 g/L; however, the fibrinogen value reduced to 1.79 g/L with D-dimer elevation over 60 µg/mL. Ninety minutes later, she developed dyspnea with hypotension at suturing hysterotomy. At the end of surgery, her fibrinogen further decreased to below 0.3 g/L with prolonged prothrombin time. After vigorous intensive care, she was discharged without sequelae. Consumptive coagulopathy may initiate and progress before apparent cardiopulmonary symptoms in some AFE cases. Non-reassuring fetal heart rate tracing concomitant with abrupt uterine tachysystole and/or hypertonus may be an earlier time point for the detection and intervention of AFE-related coagulopathy.

Investigation of optimal weight gain during pregnancy: A retrospective analysis of the Japanese perinatal registry database.

AIM: This study aimed to determine the weight gain during pregnancy that minimizes the predicted probability of various perinatal adverse events according to the pre-pregnancy body mass index (BMI) and make recommendations for optimal weight gain in Japan. METHODS: The Japan Society of Obstetrics and Gynecology perinatal database for 2015-2017 was used. From the 719 723 deliveries included in this database, parturients with underlying diseases or missing data were excluded, and 419 114 deliveries were analyzed. A questionnaire survey was also conducted to weigh each perinatal adverse event. For each of the nine outcomes, a restricted cubic spline model was made to estimate the association between the "expected gestational weight gain at 40 weeks" and the outcome risk. RESULTS: Since the classes of medical facilities were generally the same, weights were assigned according to the mean of the questionnaires rather than by the class of the facility. For each pre-pregnancy BMI, the weight gains during pregnancy that minimized the predicted probability of various adverse perinatal events were 12-15, 10-13, 7-10, and upper limit of 5 kg for the underweight, normal-weight, obese 1, and obese ≥2 groups, respectively. CONCLUSIONS: The weight gain during pregnancy that minimizes the predicted probability of various perinatal adverse events according to the pre-pregnancy BMI was established.

Comparative analysis of hyperfibrinolysis with activated coagulation between amniotic fluid embolism and severe placental abruption.

Amniotic fluid embolism (AFE) and placental abruption (PA) are typical obstetric diseases associated with disseminated intravascular coagulation (DIC). AFE is more likely to be complicated with enhanced fibrinolysis than PA. AFE may have an additional mechanism activating fibrinolytic cascade. We aimed to compare the coagulation/fibrinolysis factors among AFE, PA, and peripartum controls. We assessed AFE cases registered in the Japanese AFE Registry, and PA cases complicated with DIC (severe PA) and peripartum controls recruited at our hospital. The following factors in plasma were compared: prothrombin fragment 1 + 2 (PF1 + 2), plasmin α2-plasmin inhibitor complex (PIC), tissue factor (TF), tissue plasminogen activator (tPA), annexin A2 (AnnA2), total thrombin activatable fibrinolysis inhibitor (TAFI) including its activated form (TAFIa), and plasminogen activator inhibitor-type 1 (PAI-1). PF1 + 2 and PIC were markedly increased in both AFE (n = 27) and severe PA (n = 12) compared to controls (n = 23), without significant difference between those disease groups; however, PIC in AFE showed a tendency to elevate relative to PF1 + 2, compared with severe PA. AFE had significantly increased tPA and decreased total TAFI levels compared with severe PA and controls, which might be associated with further plasmin production in AFE and underlie its specific fibrinolytic activation pathway.

Molecular Editing Enhances Oxidation Resistance of Menaquinone-Targeting Antibiotics Lysocin E and WAP-8294A2.

Lysocin E (1 a) and WAP-8294A2 (2 a) are peptidic natural products with 37- and 40-membered macrocycles, respectively. Compounds 1 a and 2 a have potent antibacterial activities against Gram-positive bacteria and share a unique mode of action. The electron-rich indole ring of d-Trp-10 of 1 a and 2 a interacts with the electron-deficient benzoquinone ring of menaquinone, which is a co-enzyme in the bacterial respiratory chain. Formation of the electron-donor-acceptor complex causes membrane disruption, leading to cell death. Despite the promising activities of 1 a and 2 a, the susceptibility of Trp-10 to oxidative degradation potentially deters the development of these compounds as antibacterial drugs. To address this issue, we replaced the indole ring with more oxidation-resistant aromatics having a similar shape and electron-rich character. Specifically, analogues with benzofuran (1 b/2 b), benzothiophene (1 c/2 c), and 1-naphthalene (1 d/2 d) rings were designed, and chemically prepared by full solid-phase total syntheses. Antibacterial assays of the six analogues revealed similar activities of 1 d/2 d and markedly reduced activities of 1 b/2 b and 1 c/2 c compared with 1 a/2 a. Equipotent 1 d and 2 d both showed high resistance to oxidation by peroxyl radicals. Hence, the present study demonstrates a new molecular editing strategy for conferring oxidation stability on natural products with pharmacologically useful functions.

Obstetric admission to intensive care units in Japan: a cohort study using the Japanese Intensive care PAtient Database.

PURPOSE: This study aimed to describe the epidemiology and annual trends of obstetric patients using a multicenter intensive care database. METHODS: This multicenter, retrospective cohort study used the Japanese Intensive care PAtient Database (JIPAD). We included obstetric patients registered in the JIPAD between 2015 and 2020. We investigated the proportion of obstetric patients among all patients in the intensive care unit (ICU). We also described the characteristics, procedures, and outcomes of obstetric patients. In addition, the annual trends were examined by nonparametric tests for trends. RESULTS: Of the 184,705 patients enrolled in the JIPAD, 750 (0.41%) were obstetric patients from 61 facilities. The median age was 34 years, the number of post-emergency surgeries was 450 (60.0%), and the median APACHE III score was 36. Mechanical ventilation was the most common procedure performed in 247 (32.9%) patients. There were five (0.7%) in-hospital deaths. The proportion of obstetric patients in the ICU did not change between 2015 and 2020 (P for trend = 0.32). However, there was a trend for a significant decrease in the severity of illness and length of hospital stay on an annual basis between 2015 and 2020. Most patients were admitted to the ICU because of a pregnancy-related disorder postoperatively. CONCLUSION: The proportion of obstetric patients was 0.41% of all ICU admissions. The proportion of obstetric patients admitted to the ICU did not change from 2015 to 2020, but the patients' severity of illness and length of hospital stay significantly decreased over time.

Construction of Five Tryptophan Isomers and Application of the Isomers to Solid-Phase Total Syntheses of Lysocin E Derivatives.

Tryptophan (Trp) plays a unique role in peptides and proteins as its indole ring possesses an electron-rich character and an N1-H hydrogen-bond donor. Because of its non-rotationally symmetric structure, synthetic alterations of the orientation of the indole ring would modulate the intrinsic structures and functions of peptides and proteins. Here we developed synthetic routes to the five Trp isomers in which the C3-substitution of the indole ring was changed to the C2/4/5/6/7-substitutions, and applied the five monomers to Fmoc-based solid-phase peptide synthesis. Specifically, the five monomers were prepared via Negishi cross-coupling reactions of C2/4/5/6/7-iodoindoles. To demonstrate the applicability of the monomers to the solid-phase synthesis, the five Trp isomers of macrocyclic antibiotic lysocin E were selected as target molecules and synthesized through peptide elongation, on-resin macrocyclization, and global deprotection. The Trp isomers displayed markedly weaker antibacterial activity than the parent natural product, revealing the biological importance of the precise three-dimensional shape of the original Trp residue of lysocin E. The present methods for the preparation and application of these five Trp isomers provide a new strategy for analyzing and modifying the specific functions of numerous Trp-containing peptides and proteins beyond this study.

Massive platelet-rich thrombus formation in small pulmonary vessels in amniotic fluid embolism: An autopsy study.

OBJECTIVE: To identify pulmonary/uterine thrombus formation in amniotic fluid embolism (AFE). DESIGN: Retrospective, observational. SETTING: Nationwide. POPULATION: Eleven autopsy cases of AFE and control cases. METHODS: We assessed pulmonary and uterine thrombus formation and thrombus area in AFE and pulmonary thromboembolism (PTE) as a control. The area of platelet glycoprotein IIb/IIIa, fibrin, neutrophil elastase, citrullinated histone H3 (a neutrophil extracellular trap marker) and mast cell chymase immunopositivity was measured in 90 pulmonary emboli, 15 uterine thrombi and 14 PTE. MAIN OUTCOME MEASURES: Pathological evidence of thrombus formation and its components in AFE. RESULTS: Amniotic fluid embolism lung showed massive thrombus formation, with or without amniotic emboli in small pulmonary arteries and capillaries. The median pulmonary thrombus size in AFE (median, 0.012 mm2 ; P < 0.0001) was significantly smaller than that of uterine thrombus in AFE (0.61 mm2 ) or PTE (29 mm2 ). The median area of glycoprotein IIb/IIIa immunopositivity in pulmonary thrombi in AFE (39%; P < 0.01) was significantly larger than that of uterine thrombi in AFE (23%) and PTE (15%). The median area of fibrin (0%; P < 0.001) and citrullinated histone H3 (0%; P < 0.01) immunopositivity in pulmonary thrombi in AFE was significantly smaller than in uterine thrombi (fibrin: 26%; citrullinated histone H3: 1.1%) and PTE (fibrin: 42%; citrullinated histone H3: 0.4%). No mast cells were identified in pulmonary thrombi. CONCLUSIONS: Amniotic fluid may induce distinct thrombus formation in the uterus and lung. Pulmonary and uterine thrombi formation may contribute to cardiorespiratory collapse and/or consumptive coagulopathy in AFE.

Interaction of genetic liability for attention deficit hyperactivity disorder (ADHD) and perinatal inflammation contributes to ADHD symptoms in children.

Objective: Genetic and environmental factors contribute to the development of Attention Deficit/Hyperactivity Disorder (ADHD). Perinatal inflammation is one of the promising environmental risk factors for ADHD, but the relationship between the genetic risk for ADHD and perinatal inflammation requires further examination. Methods: A possible gene-environmental interaction between perinatal inflammation and ADHD polygenic risk score (ADHD-PRS) on ADHD symptoms was investigated in children aged 8-9 from the Hamamatsu Birth Cohort for Mothers and Children (N = 531). Perinatal inflammation was evaluated by the level of concentration of three cytokines assayed in umbilical cord blood. The genetic risk for ADHD was assessed by calculating ADHD-PRS for each individual using a previously collected genome-wide association study of ADHD. Results: Perinatal inflammation (ß [SE], 0.263 [0.017]; P < 0.001), ADHD-PRS (ß [SE], 0.116[0.042]; P = 0.006), and an interaction between the two (ß [SE], 0.031[0.011]; P = 0.010) were associated with ADHD symptoms. The association between perinatal inflammation and ADHD symptoms measured by ADHD-PRS was evident only in the two higher genetic risk groups (ß [SE], 0.623[0.122]; P < 0.001 for the medium-high risk group; ß [SE], 0.664[0.152]; P < 0.001 for the high-risk group). Conclusion: Inflammation in the perinatal period both directly elevated ADHD symptoms and magnified the impact of genetic vulnerability on ADHD risk particularly among children aged 8-9 with genetically higher risk for ADHD.

Exposure to environmental chemicals and cancer risk: epidemiological evidence from Japanese studies.

Exposure to certain chemicals in the environment may contribute to the risk of developing cancer. Although cancer risk from environmental chemical exposure among general populations is considered low compared to that in occupational settings, many people may nevertheless be chronically exposed to relatively low levels of environmental chemicals which vary by such various factors as residential area, lifestyle, and dietary habits. It is therefore necessary to assess population-specific exposure levels and examine their association with cancer risk. Here, we reviewed epidemiological evidence on cancer risk and exposure to dichlorodiphenyltrichloroethane (DDT), hexachlorocyclohexane (HCH), polychlorinated biphenyls (PCBs), per- and polyfluoroalkyl substances (PFASs), cadmium, arsenic, and acrylamide. Japanese are widely exposed to these chemicals, mainly through the diet, and an association with increased cancer risk is suspected. Epidemiological evidence from Japanese studies to date does not support a positive association between blood concentrations of DDT, HCH, PCBs, and PFASs and risk of breast or prostate cancer. We established assessment methods for dietary intake of cadmium, arsenic, and acrylamide using a food frequency questionnaire. Overall, dietary intakes of cadmium, arsenic, and acrylamide were not significantly associated with increased risk of total cancer and major cancer sites in the Japan Public Health Center-based Prospective Study. However, statistically significant positive associations were observed between dietary cadmium intake and risk of estrogen receptor-positive breast cancer among postmenopausal women, and dietary arsenic intake and risk of lung cancer among male smokers. In addition, studies using biomarkers as exposure assessment revealed statistically significant positive associations between urinary cadmium concentration and risk of breast cancer, and between ratio of hemoglobin adducts of acrylamide and glycidamide and risk of breast cancer. Epidemiological studies of general populations in Japan are limited and further evidence is required. In particular, studies of the association of organochlorine and organofluorine compounds with risk of cancer sites other than breast and prostate cancer are warranted, as are large prospective studies of the association between biomarkers of exposure and risk of cancer.

C-Terminal modification of polytheonamide B uncouples its dual functions in MCF-7 cancer cells.

Polytheonamide B (1) is an exceptionally large peptide that forms a transmembrane ion channel. The potent cytotoxicity of 1 against MCF-7 cancer cells originates from its two ion transport functions. Compound 1 depolarizes the plasma membrane and neutralizes acidic lysosomes. Here, we describe how we uncoupled these functions by designing and synthesizing new analogues of 1.

Identifying Active Progeny Virus Particles in Formalin-Fixed, Paraffin-Embedded Sections Using Correlative Light and Scanning Electron Microscopy.

Immunohistochemical analysis of formalin-fixed paraffin-embedded (FFPE) tissue blocks is routinely used to identify virus-infected cells. However, detecting virus particles in FFPE sections using light microscopy is difficult because of the light diffraction resolution limitations of an optical microscope. In this study, light microscopy and field emission scanning electron microscopy were performed to observe 3-dimensional virus particles in FFPE sections in a nondestructive manner using NanoSuit or osmium conductive treatment methods. The virus particles in FFPE sections were immunostained with specific antibodies against the surface antigens of the viral particles and stained with 3,3'-diaminobenzidine. A metal solution (0.2% gold chloride or 2% osmium tetroxide) was applied to enhance the 3,3'-diaminobenzidine-stained area. This procedure is nondestructive for FFPE sections and is a simpler method than transmission electron microscopy. To validate the applicability of this technique, we performed 3-dimensional imaging of the virus particles of different sizes, such as human papillomavirus, cytomegalovirus, and varicella-zoster virus. Furthermore, ultrathin sections from the FFPE sections that were observed to harbor viral particles using field emission scanning electron microscopy were prepared and assessed using transmission electron microscopy. In the correlative areas, transmission electron microscopy confirmed the presence of large numbers of virus particles. These results indicated that the combination of marking viral particles with 3,3'-diaminobenzidine/metal staining and conductive treatment can identify active progeny virus particles in FFPE sections using scanning electron microscopy. This easy correlative imaging of field emission scanning electron microscopy of the identical area of FFPE in light microscopy may help elucidate new pathological mechanisms of virus-related diseases.

Outdoor Play as a Mitigating Factor in the Association Between Screen Time for Young Children and Neurodevelopmental Outcomes.

Importance: Whether the association between higher screen time in infancy and later suboptimal neurodevelopment can be mitigated by frequency of outdoor play is unknown. Objective: To investigate whether higher screen time at age 2 years is associated with neurodevelopmental outcomes at age 4 years and whether this association is mediated by frequency of outdoor play at age 2 years 8 months. Design, Setting, and Participants: Participants were a subsample of the Hamamatsu Birth Cohort Study for Mothers and Children (HBC Study, N = 1258). Children were born between December 2007 and March 2012 and followed up from 1 year 6 months to 4 years. The analysis was conducted from April 2021 to June 2022. Exposures: Screen time longer than 1 hour a day at age 2 years was coded as higher screen time. Main Outcomes and Measures: Standardized scores for communication, daily living skills, and socialization domains of the Vineland Adaptive Behavior Scale, second edition, at age 4 years were used (mean [SD], 100 [15]). The mediating factor was frequency of outdoor play at age 2 years 8 months, with 6 or 7 days per week coded as frequent outdoor play. Results: Of 885 participants, 445 children (50%) were female; mean (SD) screen time per day was 2.6 (2.0) hours. Causal mediation analyses revealed that higher screen time at age 2 years was associated with lower scores in communication at age 4 years (nonstandardized coefficient b = -2.32; 95% CI, -4.03 to -0.60), but the association was not mediated by frequency of outdoor play. Higher screen time was also associated with lower scores in daily living skills (b = -1.76; 95% CI, -3.21 to -0.31); 18% of this association was mediated by frequency of outdoor play. Frequency of outdoor play was associated with socialization (b = 2.73; 95% CI, 1.06 to 4.39), whereas higher screen time was not (b = -1.34; 95% CI, -3.05 to 0.36). Conclusions and Relevance: Higher screen time at age 2 years was directly associated with poorer communication at age 4 years. It was also associated with daily living skills, but frequency of outdoor play at age 2 years 8 months alleviated it, suggesting outdoor play mitigated the association between higher screen time and suboptimal neurodevelopment. Future research should specify the nature of the associations and intervention measures, enabling targeted interventions that reduce the potential risk in screen time.

Association between serum concentrations of perfluoroalkyl substances and global DNA methylation levels in peripheral blood leukocytes of Japanese women: A cross-sectional study.

Global DNA methylation levels in peripheral blood leukocytes can be a biomarker for cancer risk; however, levels can be changed by various factors such as environmental pollutants. We investigated the association between serum concentrations of perfluoroalkyl substances (PFASs) and global DNA methylation levels of leukocytes in a cross-sectional study using the control group of a Japanese breast cancer case-control study [397 women with a mean age of 54.1 (SD 10.1) years]. Importantly, our analysis distinguished branched PFAS isomers as different from linear isomers. The serum concentrations of 20 PFASs were measured by in-port arylation gas-chromatography negative chemical ionization mass spectrometry. Global DNA methylation levels in peripheral blood leukocytes were measured using a luminometric methylation assay. Associations between log10-transformed serum PFAS concentrations and global DNA methylation levels were evaluated by regression coefficients in multivariable robust linear regression analyses. Serum concentrations of 13 PFASs were significantly associated with increased global DNA methylation levels in leukocytes. Global DNA methylation was significantly increased by 1.45 %-3.96 % per log10-unit increase of serum PFAS concentration. Our results indicate that exposure to PFASs may increase global DNA methylation levels in peripheral blood leukocytes of Japanese women.

Predictors of Major Adverse Cardiovascular and Cerebrovascular Events After Acute Coronary Syndromes: A Retrospective Observational Study Using YoMDB Database.

Objectives: Despite the rapid aging of the population in Japan, clinical predictors for major adverse cerebrovascular and cardiovascular events (MACCE) in patients with new onset of acute coronary syndromes (ACS) have not been well studied. This study therefore aimed to identify the predictors of MACCE in the first onset of ACS patients requiring care. Materials and Methods: Using the Yokohama Original Medical Database, we identified 3,373 patients who experienced a first onset of ACS and had certified care information from April 2014 to March 2016. The incidence proportion of MACCE from June 2014 to March 2018 was retrospectively investigated. Each patient's independence of daily living (IDL) was classified as one of three categories (reference, mild and severe). Results: Predictors of MACCE were identified using multivariate logistic regression analysis. Impaired IDL was associated with increased MACCE, with adjusted odds ratios for reference, mild and severe of 1.00, 1.35 (95% confidence intervals 1.14-1.60) and 2.12 (95% confidence intervals 1.61-2.80; P for trend < 0.001), respectively. Conclusions: This study revealed that male sex, chronic kidney disease, atrial fibrillation, high-intensity statin use, low-intensity statin use, and lower IDL (representing less independence) were the predictors of MACCE requiring care for a first onset of ACS. Further research will be required to understand the results of interventions for the identified predictors of MACCE.

Development of the Japanese version of the Depression Literacy Scale.

BACKGROUND: Depression is a major social concern in Japan. It is therefore necessary to develop a scale in Japanese that can assess depression literacy. AIMS: The present study aimed to develop the Japanese version of the Depression Literacy Scale (D-Lit-J), and examined its validity and reliability. METHODS: Three groups were administered the D-Lit-J, including 117 first-year university English literature students, 112 first-year medical school students, and 53 psychiatrists. Among these, 112 (95.7%), 112 (100%), and 29 subjects (54.7%) returned completed questionnaires, respectively. The total D-Lit-J scores were compared between the three groups to assess known-group validity, and internal reliability was examined by calculating Cronbach's alpha coefficients. Medical students were asked to complete the questionnaire a second time, 3 weeks later (11 students did not respond), to assess the test-retest reliability using the intra-class correlation coefficient. RESULTS: The total D-Lit-J scores (mean ± SD) were 7.61 ± 4.18, 9.51 ± 4.37, and 17.7 ± 3.15, for English literature students, medical students, and psychiatrists, respectively, and there were significant differences between the three groups (p < .05). The Cronbach's alpha coefficients ranged from .800 to .834 in all students, and was .764 in psychiatrists, revealing a good internal consistency. The intra-class correlation coefficient of the scale was .769. CONCLUSIONS: The D-Lit-J showed a credible known-group validity, with good internal and test-retest reliabilities. Additional studies with a greater variety of subjects and that examine concurrent or discriminant validity will be necessary in the future.

Gramicidin A accumulates in mitochondria, reduces ATP levels, induces mitophagy, and inhibits cancer cell growth.

Gramicidin A (1) is a linear 15-mer peptidic natural product. Because of its sequence of alternating d- and l-chirality, 1 folds into a ß6.3-helix in a lipid bilayer and forms a head-to-head dimer to function as a transmembrane channel for monovalent cations (H+, Na+, and K+). The potent anticancer activity of 1 was believed to be mainly attributed to the free ion diffusion across the plasma membrane. In this study, we investigated the cytostatic action of 1 in nanomolar concentrations using the human breast cancer cell line MCF-7, and revealed the unprecedented spatiotemporal behavior of 1 for the first time. Compound 1 not only disrupted the ion concentration gradients of the plasma membrane, but also localized in the mitochondria and depolarized the inner mitochondrial membrane. The diminished H+ gradient in the mitochondria inhibited ATP synthesis. The resultant mitochondrial malfunction led to mitophagy, while the cellular energy depletion induced G1 phase accumulation. The multiple events occurred in a time-dependent fashion and ultimately caused potent inhibition of cell growth. The present study provides valuable information for the design and development of new cytostatic agents exploiting channel-forming natural products.