Elimination of physiological senescent cutaneous cells in a novel p16-dependent senolytic mouse model impacts lipid metabolism in skin aging.

The evidence of the correlation between cellular senescence and aging has increased in research with animal models. These models have been intentionally generated to target and regulate cellular senescent cells with the promoter activity of p16Ink4a or p19Arf, genes that are highly expressed in aging cells. However, the senolytic efficiency in various organs and cells from these models represents unexpected variation and diversity in some cases. We have generated a novel knock-in model, p16tdT-hDTR mice, which possess tdTomato and human diphtheria toxin receptor (hDTR) downstream of Cdkn2a, an endogenous p16Ink4a gene. We successfully demonstrated that p16-derived tdTomato and hDTR expressions are observed in these mouse embryo fibroblasts and following treatment with diphtheria toxin (DT) eliminates those cells. Furthermore, we demonstrated the efficacy of eliminating p16-positive cells in vivo, and also observed a tendency to decrease their cutaneous SA-ß-gal activity after subcutaneous DT injection into p16tdT-hDTR mice. In particular, comprehensive gene expression analysis in skin revealed that upregulated genes related to lipid metabolisms with aging exhibited remarkable expressions under the senolysis. These results clearly unveiled p16-positive senescent cells contribute to age-related changes in skin.

AnNoBrainer, An Automated Annotation of Mouse Brain Images using Deep Learning.

Annotation of multiple regions of interest across the whole mouse brain is an indispensable process for quantitative evaluation of a multitude of study endpoints in neuroscience digital pathology. Prior experience and domain expert knowledge are the key aspects for image annotation quality and consistency. At present, image annotation is often achieved manually by certified pathologists or trained technicians, limiting the total throughput of studies performed at neuroscience digital pathology labs. It may also mean that simpler and quicker methods of examining tissue samples are used by non-pathologists, especially in the early stages of research and preclinical studies. To address these limitations and to meet the growing demand for image analysis in a pharmaceutical setting, we developed AnNoBrainer, an open-source software tool that leverages deep learning, image registration, and standard cortical brain templates to automatically annotate individual brain regions on 2D pathology slides. Application of AnNoBrainer to a published set of pathology slides from transgenic mice models of synucleinopathy revealed comparable accuracy, increased reproducibility, and a significant reduction (~ 50%) in time spent on brain annotation, quality control and labelling compared to trained scientists in pathology. Taken together, AnNoBrainer offers a rapid, accurate, and reproducible automated annotation of mouse brain images that largely meets the experts' histopathological assessment standards (> 85% of cases) and enables high-throughput image analysis workflows in digital pathology labs.

Decreased Memory and Learning Ability Mediated by Bmal1/M1 Macrophages/Angptl2/Inflammatory Cytokine Pathway in Mice Exposed to Long-Term Blue Light Irradiation.

Humans are persistently exposed to massive amounts of blue light via sunlight, computers, smartphones, and similar devices. Although the positive and negative effects of blue light on living organisms have been reported, its impact on learning and memory remains unknown. Herein, we examined the effects of widespread blue light exposure on the learning and memory abilities of blue light-exposed mice. Ten-week-old male ICR mice were divided into five groups (five mice/group) and irradiated with blue light from a light-emitting diode daily for 6 months. After 6 months of blue light irradiation, mice exhibited a decline in memory and learning abilities, assessed using the Morris water maze and step-through passive avoidance paradigms. Blue light-irradiated mice exhibited a decreased expression of the clock gene brain and muscle arnt-like 1 (Bmal1). The number of microglia and levels of M1 macrophage CC-chemokine receptor 7 and inducible nitric oxide synthase were increased, accompanied by a decrease in M2 macrophage arginase-1 levels. Levels of angiopoietin-like protein 2 and inflammatory cytokines interleukin-6, tumor necrosis factor-α, and interleukin-1ß were elevated. Our findings suggest that long-term blue light exposure could reduce Bmal1 expression, activate the M1 macrophage/Angptl2/inflammatory cytokine pathway, induce neurodegeneration, and lead to a decline in memory.

The Effect of Bacillus coagulans Induced Interactions among Intestinal Bacteria, Metabolites, and Inflammatory Molecules in Improving Natural Skin Aging.

BACKGROUND: Lactic acid bacteria consumption serves several health benefits to humans. However, their effect on natural skin aging is still unclear. METHODS: This study examined the effects of skin naturalization (particularly skin drying) by administering a spore-bearing lactic acid bacteria (Bacillus coagulans) in mice for 2 years. RESULTS: B. coagulans administration improved the natural skin of mice and significantly increased proportions of the genera Bacteroides and Muribaculum, among other intestinal bacteria. As metabolites, increases in nicotinic acid, putrescin, and pantothenic acid levels and a decrease in choline levels were observed. Increased hyaluronic acid, interleukin-10, and M2 macrophage levels indicate aging-related molecules in the skin. Intestinal permeability was also suppressed. Thus, these changes together improved natural skin aging. CONCLUSIONS: This study revealed that B. coagulans administration improved the natural skin aging in mice. This enhancement might be induced by the interaction of alterations in intestinal flora, metabolites, or inflammatory substances.

Induction of Skin Cancer by Long-Term Blue Light Irradiation.

Presently, people are not only exposed to sunlight but also to a large amount of blue light from personal computers and smartphones. This blue light has various effects on the living body. However, its effect on the induction of skin cancer is unknown. In this study, we investigated the induction of skin cancer by long-term blue light irradiation. Hairless mice were irradiated with blue light (LED; peak emission 479 nm) every day for one year, and a control was irradiated with white light (LED), green light (LED; peak emission 538 nm), and red light (LED; peak emission 629 nm) for one year, respectively. Skin cancer was induced only in the mice exposed to blue light. Long-term blue light irradiation also increased the migration of neutrophils and macrophages involved in carcinogenesis in the skin. In neutrophils, an increased expression of citH3 and PAD4 was observed, suggesting the possibility of NETosis. Conversely, in macrophages, inflammatory macrophages (type 1 macrophages) increased and anti-inflammatory macrophages (type 2 macrophages) decreased due to continuous blue light irradiation. These findings suggest that long-term continuous irradiation with blue light induces neutrophil NETosis and an increase in type 1 macrophages, resulting in skin cancer.

Periprocedural and 30-day outcomes of robotic-assisted percutaneous coronary intervention used in the intravascular imaging guidance.

In recent years, there have been several reports on robotic-assisted percutaneous coronary intervention (R-PCI), but few studies have been conducted on R-PCI performed under intravascular imaging guidance. To elucidate the periprocedural and postoperative 30-day outcomes of intravascular imaging-guided R-PCI, we performed a retrospective observational study on all patients in 102 consecutive cases who underwent R-PCI under intravascular imaging guidance at a single center in Japan from June 12, 2019 to February 18, 2021. The primary end point was 30-day survival, and the secondary end point was the incidence of complications. Intravascular imaging-guided R-PCI was performed 110 times in total on 125 lesions. The medians of procedural time, fluoroscopy time, contrast volume, patient entrance skin dose, and radiation exposure to the main operator were 49 min, 16 min, 67 mL, 0.62 Gy, and 0 µSv, respectively. Furthermore, 60.0% of target lesion branches were American College of Cardiology Foundation/American Heart Association classification type B2 or type C. However, in all cases, lesion dilatation was successful, and the final Thrombolysis in Myocardial Infarction flow grade was 3. The combination of manual operation was required in 12.7% of all cases, but 30-day survival was confirmed in all cases. There were two problems at the puncture site. One small distal branch artery dissection occurred due to manual operation, but no cardiovascular events (myocardial infarction, stroke) occurred and no target lesion restenosis was observed within 30 days of R-PCI. Hence, R-PCI using intravascular imaging demonstrated highly satisfactory treatment outcomes, and no complication caused by robotic operation was observed.

Incidence and natural history of coronary evagination after implanted biodegradable polymer sirolimus-eluting stent.

Aims: The incidence and temporal change in coronary evagination (CE) after first-generation drug-eluting stent implantation is well established, whereas that after biodegradable polymer sirolimus-eluting stent (BP-SES) implantation has not yet been evaluated. The aim of this study is to assess the incidence and natural history of CE after BP-SES implantation. Methods and results: In this multicenter registry, stable coronary lesions treated by Ultimaster BP-SES were evaluated by serial optical frequency domain imaging (OFDI) (at 0-1-12 or 0-3-12 months) and the incidence of CE was assessed. Coronary evagination was defined as the presence of an outward bulge in luminal vessel contour between apposed struts according to the following criteria: (i) evagination depth ≥10% of nominal stent diameter and (ii) evagination length ≥3.0 mm. Optical frequency domain imaging was obtained in 98, 47, 49, and 87 lesions at 0, 1, 3, and 12 months, respectively. Coronary evagination was observed in 20 (42.6%) and 12 (24.5%) lesions at 1 and 3 months, respectively, and all but one CE had resolved at 12 months. At 12 months, the mean CE area was almost zero and the mean malapposed stent area was also decreased. Comparison of the serial OFDI images indicated that CEs originated mostly from acute stent malapposition or coronary dissection behind the implanted stent. Conclusions: In stable lesions, CE was occasionally observed with Ultimaster BP-SES at 1-3 months but mostly resolved within 12 months, without late-acquired stent malapposition. These findings suggest the safety and feasibility of biodegradable polymer coating on DES.

Prevalence, clinical characteristics, and impact of active cancer in patients with acute myocardial infarction: data from an all-comer registry.

BACKGROUND: Although a history of cancer is a poor prognostic factor in patients with acute myocardial infarction (AMI), the clinical importance of coexisting active cancer remains unclear. METHODS: In this single-center retrospective study, we reviewed an AMI registry and assessed the prevalence and predictors of active cancer, 1-year incidence of cardiac death or major bleeding events (defined as a Bleeding Academy Research Consortium type 3 or 5), and the impact of coexisting active cancer on clinical outcomes. Active cancer was defined as either an already-diagnosed or undiagnosed occult cancer. RESULTS: Between January 2012 and December 2017, 1140 AMI patients (median age, 69 years; male, 76.0%) were enrolled. Active and historical cancers were diagnosed in 63 patients (5.5%) and 50 patients (4.4%), respectively. The most common location was the urinary tract (n=21). In the Kaplan-Meier analysis, the active cancer group had a higher incidence of 1-year cardiac death (17.5% vs. 5.3%, p < 0.001) and major bleeding events (19.0% vs. 5.6%, p < 0.001) than the non-cancer group. In the multivariate Cox proportional hazards regression models, active cancer was an independent predictor of both cardiac death and major bleeding at 1 year. Specifically, gastrointestinal tract and advanced-stage cancers had the poorest outcomes. Compared to the non-cancer group, the 1-year major bleeding rate was higher for all cancer types and stages. In contrast, early-stage cancers had a weaker impact on the 1-year cardiac mortality compared to advanced-stage cancers. Similarly, cardiac death during 1-year also occurred less frequently in occult cancers than in already-known cancers. CONCLUSIONS: In patients with AMI, coexisting active cancer was rare, but it significantly impacted cardiac death and major bleeding events.

[The Strength of the Sense of Shame of the Aged in Care Scenes: Comparison Between Whether or not There is Anyone Else Around Them].

Medical staff in a hospital or nursing facility should take care of aged individuals with dignity and respect. We conducted a survey on aged individuals to derive under what care circumstances they had a sense of shame, using 12 illustrations, drawn by ourselves, which were common daily care scenes where nurses and patients meet. This survey was conducted at 4 care facilities in A prefecture, Japan. The number of surveyed persons was 43, with the following exclusion criteria: over 60 years old, more than third level of care needed, and non suspected of having dementia. We got the following results from the answers of 41 persons: 1. When elder persons are surrounded by people other than the care staff, they feel more of a sense of shame than when alone; 2. They feel more sense of shame when they use a wheelchair than when they use crutches; 3. They do not feel much shame when they get a bed-bath, even if other persons are there; and 4. Male patients feel more shame than females when they meet their family. These results suggest that elderly patients feel a stronger sense of shame when they are seen by others than when they are seen by care staff. The result 2 suggests that the use of a wheelchair exposes their physical weakness to others. Males feel a stronger sense of shame when they show a weakness in their gender role. We conclude that the sense of shame of aged individuals in daily life scenes in a care facility depends on their gender and whether or not they are surrounded by other persons.

[Nursing Care Students' Image of Shame -Comparison Between First and Second Year Nursing Students].

It is not easy for nurses to estimate a patient's degree of shame, as the sense of shame depends on each person's personality, but nurses are requested to evaluate it as correctly as possible and to reduce the patient's mental load. We presume that most of the sense of shame is generated by body defects or disadvantages recognized by the patient. In this study, we tried to measure the degree of shame and to improve the basic nursing curriculum, depending on students' school year, under the assumption of what cases the nurses would frequently meet in a hospital. We prepared 13 figures that show common cases in hospitals. In these figures: 1) 6 figures show cases in which a nurse touched a patient's body; 2) 3 figures show common daily life; and 3) 4 figures show cases in which there are other people around the patient. A questionnaire was given to the first and second year students in A Nursing University, and we allocated scores of 1 to 10: 1 is "no-shame", and 10 is "very much shame". The students answered that patients must feel shame when: 1) they take off their clothes, 2) they show their disability to another person even without taking their clothes off, and 3) having people other than medical staff around them. In the results, as 2) appeared more strongly in the second year students than in the first, we thought that the second year students could surmise a patient's position in a hospital through the effect of the nursing education.

A trafficking-deficient KCNQ1 mutation, T587M, causes a severe phenotype of long QT syndrome by interfering with intracellular hERG transport.

BACKGROUND: KCNQ1-T587M is a C-terminal mutation correlated with severe phenotypes of long QT syndrome (LQTS). However, functional analysis of KCNQ1 channels with the T587M mutation showed a mild genotype in the form of haploinsufficiency in a heterologous expression system. This study sought to explore the molecular mechanism underlying the phenotype-genotype dissociation of LQTS patients carrying the KCNQ1-T587M mutation. METHODS: cDNAs for wild-type (WT) and KCNQ1 mutations (R259C and T587M) were transiently transfected into HEK293 cells stably expressing hERG (hERG-HEK), and whole-cell patch-clamp technique was performed to examine the effect of KCNQ1 mutations on IKr-like currents. In addition, fluorescence resonance energy transfer (FRET) was conducted to demonstrate the molecular interaction between KCNQ1 and hERG when co-expressed in HEK293 cells. RESULTS: KCNQ1-T587M mutation produced a significant (p<0.01) decrease in IKr-like tail current densities without affecting the gating kinetics, while KCNQ1-R259C mutation had no significant effect on the IKr-like tail current densities. Consistent with this result, FRET experiments demonstrated that both KCNQ1-WT and -R259C interacted with hERG in the cytosol and on the plasma membrane; however, the interaction between KCNQ1-T587M and hERG was observed only in the cytosol, and hERG proteins were seldom transported to the cell membrane, suggesting that the KCNQ1-T587M mutation impaired the trafficking of hERG to the cell membrane. CONCLUSIONS: The disruption of hERG trafficking caused by the KCNQ1-T587M mutation is likely the reason why some patients exhibit severe LQTS phenotypes.

Nursing Education Trial Using a Virtual Nightingale Ward.

Nursing department students are expected to correctly grasp the entire concept of nursing through their education. The authors created a movie of a Nightingale ward (virtual ward, hereafter) with an architectural computer design software for education. The students' reaction to the virtual ward was categorized into three viewpoints: that of nurses, of patients, and of nurses and patients in common. Most of the reactions in each viewpoint were: "easy to observe patients" in the nurses' viewpoint; "no privacy" in the patients' viewpoint; and "wide room" in the common viewpoint, respectively. These reactions show the effectiveness of using a virtual ward in nursing education. Because these reactions are characteristics of a Nightingale ward, and even students, who have generally less experiences, recognized these characteristics from the both viewpoints of nurses and patients.

Development of Teaching Materials in Basic Nursing Using Virtual Reality －Ceiling Image Teaching Materials of a Hospital Room－.

It is difficult to understand nursing practically for students who have just started studying nursing, because they cannot imagine the actual medical scene. The authors expect that they can improve nursing learning through experiences in virtual medical environments supported by ICT (information and commnication technology). We developed teaching materials in nursing education, and in a previous study we reported that the use of a virtual hospital was useful for students who had not had medical experience. The aim of the present study was to have students consider the meaning of a hospital room ceiling. First, we showed them a few ceiling patterns, and they selected the best one among the patterns. After that, we showed the same one in a virtual hospital room. They had negative opinions about what they had thought was the best one at this time. This demonstrates that the virtual hospital room is effective when students evaluate the ceiling as a part of the hospital room.

[Evaluation of Educational Effect of Problem-Posing System in Nursing Processing Study].

The nursing processing study is generally difficult, because it is important for nursing college students to understand knowledge and utilize it. We have developed an integrated system to understand, utilize, and share knowledge. We added a problem-posing function to this system, and expected that students would deeply understand the nursing processing study through the new system. This system consisted of four steps: create a problem, create an answer input section, create a hint, and verification. Nursing students created problems related to nursing processing by this system. When we gave a lecture on the nursing processing for second year students of A university, we tried to use the creating problem function of this system. We evaluated the effect by the number of problems and the contents of the created problem, that is, whether the contents consisted of a lecture stage or not. We also evaluated the correlation between those and regular examination and report scores. We derived the following: 1. weak correlation between the number of created problems and report score (r=0.27), 2. significant differences between regular examination and report scores of students who created problems corresponding to the learning stage, and those of students who created problems not corresponding to it (P<0.05). From these results, problem-posing is suggested to be effective to fix and utilize knowledge in the lecture of nursing processing theory.

[Nursing Education Utilizing Experiences in a Virtual Hospital Ward].

Environmental design should be required at medical facilities for conducting medical practice safely and for making hospitalization comfortable. Many medical nursing students cannot imagine medical facilities, especially hospital wards, when they study medical environments in a basic nursing lecture. As a result, they cannot connect well with patient assistance. We employed a computer assisted designing software, "3D My Home Designer" (Mega Soft Company) that runs on Windows 8, and considered the usefulness of it for lectures on environmental design showing how to design a hospital ward for patients' optimal hospital stay. We drew a medical facility in 2-D first, transformed it into 3D images, and then created movies of a virtual hospital ward in which a patient walked around. These movies consisted of 3 kinds: a) hospital room with changeable wall color, b) different allocations of hospital room and nurse station, and c) a blurred ward which corresponded to how a patient with poor eyesight (cataract) would see a ward. We prepared as controls: a') still images of a hospital room, b') still images of ward, and c') a documentation on how a ward is seen by a patient with a cataract. We gave a questionnaire to students and nurses about these movies and still images (controls). In a) and b), there were no differences between the movies and still images in both students and nurses. In c), both students and nurses had a viewpoint from the patient with poor eyesight. From these results, we consider that the students, who have fewer experiences in a hospital, may understand the environments well by movies and the application of a virtual movie ward to nursing education may be useful in a lecture, depending on the readiness of the students.

A novel KCNJ2 nonsense mutation, S369X, impedes trafficking and causes a limited form of Andersen-Tawil syndrome.

BACKGROUND: Mutations in KCNJ2, a gene encoding the inward rectifier K(+) channel Kir2.1, are associated with Andersen-Tawil syndrome (ATS), which is characterized by (1) ventricular tachyarrhythmias associated with QT (QU)-interval prolongation, (2) periodic paralysis, and (3) dysmorphic features. METHODS AND RESULTS: We identified a novel KCNJ2 mutation, S369X, in a 13-year-old boy with prominent QU-interval prolongation and mild periodic paralysis. The mutation results in the truncation at the middle of the cytoplasmic C-terminal domain that eliminates the endoplasmic reticulum (ER)-to-Golgi export signal. Current recordings from Chinese hamster ovary cells transfected with KCNJ2-S369X exhibited significantly smaller K(+) currents compared with KCNJ2 wild type (WT) (1 µg each) (-84 ± 14 versus -542 ± 46 picoamperes per picofarad [pA/pF]; -140 mV; P<0.0001). Coexpression of the WT and S369X subunits did not show a dominant-negative suppression effect but yielded larger currents than those of WT+S369X (-724 ± 98 pA/pF>-[84+542] pA/pF; 1 µg each; -140 mV). Confocal microscopy analysis showed that the fluorescent protein-tagged S369X subunits were predominantly retained in the ER when expressed alone; however, the expression of S369X subunits to the plasma membrane was partially restored when coexpressed with WT. Fluorescence resonance energy transfer analysis demonstrated direct protein-protein interactions between WT and S369X subunits in the intracellular compartment. CONCLUSIONS: The S369X mutation causes a loss of the ER export motif. However, the trafficking deficiency can be partially rescued by directly assembling with the WT protein, resulting in a limited restoration of plasma membrane localization and channel function. This alleviation may explain why our patient presented with a relatively mild ATS phenotype.

Adrenergic regulation of the rapid component of delayed rectifier K+ current: implications for arrhythmogenesis in LQT2 patients.

BACKGROUND: KCNH2 gene mutations disrupting rapid component of I(K) (I(Kr)) underlie type 2 congenital long QT syndrome (LQT2). Startled auditory stimuli are specific symptomatic triggers in LQT2, thus suggesting fast arrhythmogenic mechanism. OBJECTIVE: We investigated acute alpha(1A)- and cyclic adenosine monophosphate (cAMP)-related beta-adrenergic modulation of I(Kr) in HL-1 cardiomyocytes, wild type (WT)- and 2 LQT2-associated mutant Kv11.1 channels (Y43D- and K595E-Kv11.1) reconstituted in Chinese hamster ovary (CHO) cells. METHODS: I(Kr) and Kv11.1 currents were recorded using the whole-cell patch-clamp technique and confocal microscopy of HL-1 cardiomyocytes transfected with green fluorescent protein (GFP)-tagged pleckstrin homology domain of phospholipase C-delta(1) visualized fluctuations of membrane phosphatidylinositol 4,5-bisphosphate (PIP(2)) content. RESULTS: In HL-1 cardiomyocytes expressing human alpha(1A)-adrenoceptor, superfusion with phenylephrine significantly reduced I(Kr) amplitude, shifted current activation to more positive potentials, and accelerated kinetics of deactivation. Confocal images showed a decline of membrane PIP(2) content during phenylephrine exposure. Simultaneous application of adenylyl cyclase activator forskolin and phosphodiesterase inhibitor 3-isobutyl-1-methylxantine (IBMX) shifted I(Kr) activation to more negative potentials and decreased tail current amplitudes after depolarizations between +10 and +50 mV. In CHO cells, alpha(1A)-adrenoceptor activation downregulated WT-Kv11.1 channels and forskolin/IBMX produced a dual effect. Expressed alone, the Y43D-Kv11.1 or K595E-Kv11.1 channel had no measurable function. However, co-expression of WT-Kv11.1 and each mutant protein evoked currents with loss-of-function alterations but identical to WT-Kv11.1 alpha(1A)- and forskolin/IBMX-induced regulation. CONCLUSION: Acute adrenergic regulation of at least 2 Kv11.1 mutant channels is preserved as in WT-Kv11.1 and native I(Kr). Suppression of alpha(1A)-adrenoceptor-related transduction might have therapeutic implications in some cases of LQT2.