Dynamic movement of the Golgi unit and its glycosylation enzyme zones.

Knowledge on the distribution and dynamics of glycosylation enzymes in the Golgi is essential for better understanding this modification. Here, using a combination of CRISPR/Cas9 knockin technology and super-resolution microscopy, we show that the Golgi complex is assembled by a number of small 'Golgi units' that have 1-3 µm in diameter. Each Golgi unit contains small domains of glycosylation enzymes which we call 'zones'. The zones of N- and O-glycosylation enzymes are colocalised. However, they are less colocalised with the zones of a glycosaminoglycan synthesizing enzyme. Golgi units change shapes dynamically and the zones of glycosylation enzymes rapidly move near the rim of the unit. Photobleaching analysis indicates that a glycosaminoglycan synthesizing enzyme moves between units. Depletion of giantin dissociates units and prevents the movement of glycosaminoglycan synthesizing enzymes, which leads to insufficient glycosaminoglycan synthesis. Thus, we show the structure-function relationship of the Golgi and its implications in human pathogenesis.

A mouse model of autoimmune inner ear disease without endolymphatic hydrops.

Autoimmune inner ear disease (AIED) is an organ-specific disease characterized by irreversible, prolonged, and progressive hearing and equilibrium dysfunctions. The primary symptoms of AIED include asymmetric sensorineural hearing loss accompanied by vertigo, aural fullness, and tinnitus. AIED is divided into primary and secondary types. Research has been conducted using animal models of rheumatoid arthritis (RA), a cause of secondary AIED. However, current models are insufficient to accurately analyze vestibular function, and the mechanism underlying the onset of AIED has not yet been fully elucidated. Elucidation of the mechanism of AIED onset is urgently needed to develop effective treatments. In the present study, we analyzed the pathogenesis of vertigo in autoimmune diseases using a mouse model of type II collagen-induced RA. Auditory brain stem response analysis demonstrated that the RA mouse models exhibited hearing loss, which is the primary symptom of AIED. In addition, our vestibulo-oculomotor reflex analysis, which is an excellent vestibular function test, accurately captured vertigo symptoms in the RA mouse models. Moreover, our results revealed that the cause of hearing loss and vestibular dysfunction was not endolymphatic hydrops, but rather structural destruction of the organ of Corti and the lateral semicircular canal ampulla due to an autoimmune reaction against type II collagen. Overall, we were able to establish a mouse model of AIED without endolymphatic hydrops. Our findings will help elucidate the mechanisms of hearing loss and vertigo associated with AIED and facilitate the development of new therapeutic methods.

Prenatal methamphetamine exposure causes dysfunction in glucose metabolism and low birthweight.

Methamphetamine (METH) is a psychostimulant drug that induces addiction. Previous epidemiological studies have demonstrated that maternal METH abuse during pregnancy causes low birthweight (LBW) in the offspring. As a source of essential nutrients, in particular glucose, the placenta plays a key role in fetal development. LBW leads to health problems such as obesity, diabetes, and neurodevelopmental disorders (NDDs). However, the detailed mechanism underlying offspring's LBW and health hazards caused by METH are not fully understood. Therefore, we investigated the effects of prenatal METH exposure on LBW and fetal-placental relationship by focusing on metabolism. We found dysfunction of insulin production in the pancreas of fetuses exposed to METH. We also found a reduction of the glycogen cells (GCs) storing glycogens in the junctional zone of placenta, all of which suggest abnormal glucose metabolism affects the fetal development. These results suggest that dysfunction in fetal glucose metabolism may cause LBW and future health hazards. Our findings provide novel insights into the cause of LBW via the fetal-placental crosstalk.

SR 57227A, a serotonin type-3 receptor agonist, as a candidate analgesic agent targeting nociplastic pain.

Pain is influenced by various factors, such as fear, anxiety, and memory. We previously reported that pain-like behaviors in mice can be induced by environmental cues in which a pain stimulus was previously presented, and that pain was reduced using fentanyl (an opioid). Although opioid analgesics are currently used to treat persistent pain, their inappropriate use causes a significant number of deaths in the United States. Thus, alternative medicines to opioids are needed. Here, we reported that SR 57227A, a serotonin type-3 receptor agonist, significantly reduced pain-like behaviors. The number of c-Fos positive cells increased by environmental cues in PFC was decreased by SR 57227A. Moreover, SR 57227A reduced pain-like behaviors of the formalin test, and restored reductions in paw withdrawal thresholds by acidic saline intramuscular injection and sciatic nerve ligation. Unlike opioids, SR 57227A induced no preference behaviors as measured by the conditioned place preference test. These data suggested that SR 57227A is an effective alternative pain reliever to opioids that targets chronic pain.

A new therapy against ulcerative colitis via the intestine and brain using the Si-based agent.

Ulcerative colitis (UC) is a non-specific inflammatory bowel disease that causes ulcers and erosions in the colonic mucosa and becomes chronic with cycles of amelioration and exacerbation. Because its exact etiology remains largely unclear, and the primary therapy is limited to symptomatic treatment, the development of new therapeutic agent for UC is highly desired. Because one of the disease pathogenesis is involvement of oxidative stress, it is likely that an appropriate antioxidant will be an effective therapeutic agent for UC. Our silicon (Si)-based agent, when ingested, allowed for stable and persistent generation of massive amounts of hydrogen in the gastrointestinal tract. We demonstrated the Si-based agent alleviated the mental symptom as well as the gastrointestinal symptoms, inflammation, and oxidation associated with dextran sodium sulfate-induced UC model through Hydrogen and antioxidant sulfur compounds. As the Si-based agent was effective in treating UC in the brain and large intestine of mice, it was considered to be capable of suppressing exacerbations and sustaining remission of UC.

Combined prognostic value of malnutrition using GLIM criteria and renal insufficiency in elderly heart failure.

AIMS: We aimed to investigate the prognostic impact of malnutrition, defined by the Global Leadership Initiative on Malnutrition (GLIM) criteria, stratified by renal function in hospitalized patients with acute decompensated heart failure (HF). METHODS AND RESULTS: In this retrospective study, 314 patients who were hospitalized for acute decompensated HF from August 2019 to October 2020 were enrolled. We evaluated malnutrition using the GLIM criteria during the time of admission. The primary outcome was 90-day all-cause mortality. The median patient age was 82 years, and 90-day mortality was 14.0%. In total, 76 (24.2%) patients were malnourished according to the GLIM criteria. Malnutrition defined by the GLIM criteria [adjusted hazard ratio (HR) 1.41, 95% confidence interval (CI) 1.02-1.91, P = 0.036] and renal insufficiency [adjusted HR 2.59, 95% CI 1.07-6.28, P = 0.035 for estimated glomerular filtration rate (eGFR) < 30 mL/min/1.73 m2 vs. ≥60 mL/min/1.73 m2 ] were identified as independent predictors of 90-day mortality after adjustment for age, systolic blood pressure, and serum sodium level. In the combined setting of both variables, patients with malnutrition and eGFR < 30 mL/min/1.73 m2 had a markedly higher risk of 90-day mortality compared with those without malnutrition and eGFR ≥ 60 mL/min/1.73 m2 (adjusted HR 3.92, 95% CI 1.10-13.9, P = 0.035). Adding both eGFR and malnutrition, defined by the GLIM criteria, to the baseline model with established risk factors improved both net reclassification and integrated discrimination greater than that of the baseline model (0.606, P < 0.001 and 0.050, P = 0.002, respectively), even when compared with the model with malnutrition by the GLIM alone (0.463, P = 0.002 and 0.034, P < 0.001, respectively). CONCLUSIONS: Nutrition screening using the GLIM criteria stratified by renal function could clearly predict 90-day mortality in hospitalized patients with acute decompensated HF.

Pain-like behavior in mice can be induced by the environmental context in which the pain stimulus was previously given.

It has been known for decades that classical conditioning influences pain perception. However, the precise mechanism of pain modified by conditioning remains unclear, partly because of the lack of dedicated behavioral tests. In the present study, we aimed to develop a new method to detect conditioned pain using mice that were injected with formalin as an unconditioned nociceptive stimulus into the hind paw repetitively under a neutral environment. On the test day, the mice exhibited a pain-like behavior without the application of a pain stimulus in the environment. These results demonstrate that a conditioned nociceptive response can be induced by exposure alone to the environmental context in which the pain was previously experienced. The conditioned nociceptive response was sustained for at least 2 weeks. Furthermore, the conditioned nociceptive response was reduced by fentanyl but not by ibuprofen, pregabalin or fluvoxamine. This method may be useful for studying the mechanisms of irritable chronic pain and for the development of novel therapeutic strategies.

＜Editors' Choice＞ Prognostic utility of multipoint nutritional screening for hospitalized patients with acute decompensated heart failure.

This study aimed to evaluate the impact of serial changes in nutritional status on 1-year events including all-cause mortality or rehospitalization owing to heart failure (HF) among hospitalized patients with acute decompensated HF (ADHF). The study subjects comprised 253 hospitalized patients with ADHF. The controlling nutritional status (CONUT) score was assessed both at hospital admission and discharge. The subjects were divided into three groups according to nutritional status using CONUT score: normal (0 and 1), mild risk (2-4), and moderate to severe risk defined as malnutrition (5-12). We observed nutritional status was improved or not. The incidence of malnutrition was 30.4% at hospital admission and 23.7% at discharge, respectively. Malnutrition was independently associated with 1-year events among hospitalized patients with ADHF. Presence or absence of improvement in nutritional status was significantly associated with 1-year events (P < 0.05), that was independent of percentage change in plasma volume in multivariate Cox regression analyses. We determined a reference model, including gender and estimated glomerular filtration rate, using multivariate logistic regression analysis (P < 0.05). Adding the absence of improvement in nutritional status during hospitalization to the reference model significantly improved both NRI and IDI (0.563, P < 0.001 and 0.039, P = 0.001). Furthermore, malnutrition at hospital discharge significantly improved NRI (0.256, P = 0.036) In conclusion, serial changes in the nutritional status evaluated on the basis of multiple measurements may provide more useful information to predict 1-year events than single measurement at hospital admission or discharge in hospitalized patients with ADHF.

Si-Based Hydrogen-Producing Nanoagent Protects Fetuses From Miscarriage Caused by Mother-to-Child Transmission.

Mother-to-child transmission of viruses and bacteria increases the risk of miscarriage and various diseases in children. Such transmissions can result in infections and diseases in infants or the induction of an inflammatory immune response through the placenta. Recently, we developed a silicon (Si)-based hydrogen-producing nanoagent (Si-based agent) that continuously and effectively produces hydrogen in the body. Since medical hydrogen has antioxidative, anti-inflammatory, antiallergic, and antiapoptotic effects, we investigated the effects of our Si-based agent on mother-to-child transmission, with a focus on the rate of miscarriage. In pregnant mice fed a diet containing the Si-based agent, lipopolysaccharide (LPS)-induced miscarriage due to mother-to-child transmission was reduced and inflammation and neutrophil infiltration in the placenta were suppressed. We also found that the Si-based agent suppressed IL-6 expression in the placenta and induced the expression of antioxidant and antiapoptotic genes, such as Hmox1 and Ptgs2. The observed anti-inflammatory effects of the Si-based agent suggest that it may be an effective preventative or therapeutic drug for miscarriage or threatened miscarriage during pregnancy by suppressing maternal inflammation caused by bacterial and viral infections.

Clinical Impact of Worsening Renal Function in Elderly Patients with Acute Decompensated Heart Failure.

Background and Objectives: The clinical significance of worsening renal function (WRF) in elderly patients with acute decompensated heart failure (ADHF) is not completely understood. We compared the clinical conditions between younger and elderly patients with ADHF after the appearance of WRF to establish its prognostic influence. Methods: We included 654 consecutive patients (37% women) admitted for ADHF. We divided the patients into four groups according to their age (<80 years, under-80, n=331; ≥80 years, over-80, n=323) and to their WRF statuses (either WRF or non-WRF group). We defined WRF as an increase in serum creatinine level ≥0.3 mg/dL or ≥150% within 48 hours after hospital arrival (under-80, n=62; over-80, n=75). The primary endpoint was a composite of cardiac events within 1 year. Results: The survival analyses revealed that the WRF group had significantly more cardiac events than the non-WRF group in patients in the over-80 group (log-rank p=0.025), but not in those of the under-80 group (log-rank p=0.50). The patients in the over-80, WRF group presented more significant mean blood pressure (MBP) drops than those in the over-80 non-WRF group (p=0.003). Logistic regression analyses revealed that higher MBP at admission was a significant predictor of WRF. Conclusions: WRF is a predictor of poor outcomes in elderly patients with ADHF.

Impact of predictive value of Fibrosis-4 index in patients hospitalized for acute heart failure.

BACKGROUND: Abnormalities in liver function tests commonly occur in patients with acute heart failure (AHF). The Fibrosis-4 (FIB4) index, a non-invasive and easily calculated marker, has been used for hepatic diseases and reflects adverse prognosis. It is not clearly established whether the FIB4 index at admission can predict adverse outcomes in patients with AHF. METHODS AND RESULTS: From a multicenter AHF registry, we retrospectively evaluated 1162 consecutive patients admitted due to AHF (median age 78 [69-85] years and 702 patients [60.4%] were male). The FIB4 index at admission was calculated as: age (yrs) × aspartate aminotransferase [U/L]/(platelets count [103/µL] × âˆšalanine aminotransferase [U/L]. The median value of the FIB4 index at admission was 2.79. All-cause mortality and rehospitalization due to HF at 12 months were investigated as a composite endpoint and occurred in 142 (12.2%) patients and 232 (20%) patients, respectively. Kaplan-Meyer analysis shows a significant increase in the composite endpoint from the first to fourth quartile group of the FIB4 index values (log-rank, p < 0.001). Multivariate Cox regression model revealed the FIB4 index was an independent risk predictor for composite endpoint in patients with AHF (3 months: HR ratio 1.013 [95% Confidence interval (CI):1.001-1.025]; p = 0.03, 12 months: HR 1.015 [95% CI:1.005-1.025]; p = 0.003, respectively). However, neither aspartate aminotransferase, alanine aminotransferase, nor platelet count was found to be a significant predictor. CONCLUSIONS: Hepatic dysfunction evaluated with the FIB4 index at admission is a predictor of the composite endpoint of all-cause mortality and rehospitalization in AHF patients.

Efficacy of Rapid Decongestion Strategy in Patients Hospitalized for Acute Heart Failure.

BACKGROUND: Clinical congestion is the most dominant feature in patients with acute decompensated heart failure (HF). However, uncertainty exists due to the permutations and combinations of congestion status and decongestion strategies. This study investigated the effect of congestion status and its improvement on 1-year mortality.Methods and Results:In all, 453 consecutive patients hospitalized for acute decompensated HF between July 2015 and March 2017 were prospectively included in the study. Congestion was evaluated using the congestion score. The 1-year mortality rate was 22.7%. The mean (±SD) congestion scores at admission, on Day 3, and at discharge were 10.7±3.9, 3.4±3.5, and 0.3±0.8, respectively. The improvement rate in congestion scores during the first 3 days was 78%; 46.6% of patients had residual congestion. The Day 3 congestion score and the improvement rate during the first 3 days were related to 1-year all-cause mortality and cardiovascular mortality. Combined predictive values were examined by calculating multivariable-adjusted hazard ratios for associations of residual congestion and improvement rate during the first 3 days, and prognostic variables identified by the Cox regression model. Residual congestion and lesser improvement (<64%) were associated with higher relative risk of 1-year all-cause mortality and cardiovascular mortality than residual congestion and higher improvement (≥64%) or resolved congestion. CONCLUSIONS: Rapid decongestion could be a prerequisite regardless of residual congestion in hospitalized acute decompensated HF patients.

Prognostic Importance of Multiple Nutrition Screening Indexes for 1-Year Mortality in Hospitalized Acute Decompensated Heart Failure Patients.

Background: The purpose of the study was to evaluate the impact of nutritional status on 1-year mortality in hospitalized patients with acute decompensated heart failure (ADHF). Methods and Results: We enrolled 457 hospitalized ADHF patients. Previously established objective nutritional indexes (controlling nutritional status [CONUT], prognostic nutritional index [PNI], geriatric nutritional risk index [GNRI], and subjective global assessment [SGA]) were evaluated at hospital admission. Malnutrition was defined as CONUT score ≥5, PNI score <38, GNRI score <92, and SGA scores B and C. The frequencies of malnutrition based on CONUT, PNI, GNRI, and SGA were 31.5%, 21.4%, 44.9%, and 27.8%, respectively. All indexes were related to the occurrence of 1-year mortality on univariate Cox regression analysis (P<0.05). We constructed a reference model using age, body mass index, systolic blood pressure, sodium concentration, and renal function on multivariable Cox regression analysis. Adding SGA to the reference model significantly improved both net reclassification improvement (NRI) and integrated discrimination improvement (0.344, P=0.002; 0.012, P=0.049; respectively). Other indexes (CONUT, PNI, and GNRI scores) significantly improved NRI (0.254, P=0.019; 0.273, P=0.013; 0.306, P=0.006; respectively). Conclusions: Nutritional screening assessed at hospital admission was appropriate for the prediction of 1-year mortality in hospitalized patients with ADHF.

Structural insight into microtubule stabilization and kinesin inhibition by Tau family MAPs.

The Tau family microtubule-associated proteins (MAPs) promote microtubule stabilization and regulate microtubule-based motility. They share the C-terminal microtubule-binding domain, which includes three to five tubulin-binding repeats. Different numbers of repeats formed by alternative splicing have distinct effects on the activities of these proteins, and the distribution of these variants regulates fundamental physiological phenomena in cells. In this study, using cryo-EM, we visualized the MAP4 microtubule complex with the molecular motor kinesin-1. MAP4 bound to the C-terminal domains of tubulins along the protofilaments stabilizes the longitudinal contacts of the microtubule. The strongest bond of MAP4 was found around the intertubulin-dimer interface such that MAP4 coexists on the microtubule with kinesin-1 bound to the intratubulin-dimer interface as well. MAP4, consisting of five repeats, further folds and accumulates above the intertubulin-dimer interface, interfering with kinesin-1 movement. Therefore, these cryo-EM studies reveal new insight into the structural basis of microtubule stabilization and inhibition of kinesin motility by the Tau family MAPs.

Structural Insights into the Altering Function of CRMP2 by Phosphorylation.

Collapsin response mediator protein 2 (CRMP2) regulates neuronal polarity by controlling microtubule dynamics. CRMP2 activity is regulated by semaphorin-induced phosphorylation at the C-terminal tail domain. Unphosphorylated CRMP2 induces effective axonal microtubule formation to give the axonal characteristics to a neurite, whereas phosphorylated CRMP2 leads to the apparently opposite effect, growth cone collapse. We have recently characterized the structural detail of CRMP2-induced axonal microtubule formation (Niwa et al. (2017) Sci. Rep., 7: 10681). CRMP2 forms the hetero-trimer with GTP-tubulin to induce effective axonal microtubule formation in the future axon. Phosphorylation of CRMP2 has been reported to decrease the affinity between CRMP2 and the microtubule, albeit the molecular mechanisms of how the phosphorylation of CRMP2 changes the structure to achieve distinct effects from unphosphorylated CRMP2 is not well understood. Here we performed a series of biochemical and structural analyses of phospho-mimic CRMP2. Phosphorylation of CRMP2 undergoes small conformational changes at the C-terminal tail with shifting the surface charge, which not only alters the interactions within the CRMP2 tetramer but also alters the interactions with GTP-tubulin. Consequently, phospho-mimic CRMP2 fails to form a hetero-trimer with GTP-tubulin, thus losing the ability to establish and maintain the axonal microtubules.Key words: CRMP2, phosphorylation, microtubule, axon, crystal structure.

Impact of high-density lipoprotein 3 cholesterol subfraction on periprocedural myocardial injury in patients who underwent elective percutaneous coronary intervention.

BACKGROUND: Periprocedural myocardial injury (PMI) is a major complication of percutaneous coronary intervention (PCI) and is associated with atherosclerotic coronary plaque and worse clinical outcomes. High-density lipoprotein cholesterol (HDL-C) is a protective factor for cardiovascular disease. However, the role of HDL-C subfractions, such as HDL2 cholesterol (HDL2-C) or HDL3 cholesterol (HDL3-C), in cardiovascular disease remains unclear. The purpose of the study was to investigate the relationship between HDL2-C and HDL3-C subfractions and the incidence of PMI in patients who underwent elective PCI. METHODS: We enrolled 129 patients who underwent elective PCI for stable angina pectoris. PMI was defined as an increase in high-sensitivity troponin T levels > 5 times the upper normal limit (> 0.070 ng/mL) at 24 h after PCI. Serum HDL-C subfractions (HDL2-C and HDL3-C) were assessed using ultracentrifugation in patients with and those without PMI. RESULTS: HDL3-C levels were significantly lower in patients with PMI than in those without (15.1 ± 3.0 mg/dL vs. 16.4 ± 2.9 mg/dL, p = 0.016) and had an independent and inverse association with PMI (odds ratio, 0.86; 95% confidence interval, 0.74-0.99; p = 0.038). When divided by the cut-off value of HDL3-C for PMI (14.3 mg/dL), the incidence of PMI was significantly higher in low HDL3-C patients than in high HDL3-C patients (51.2% vs. 30.2%, p = 0.020). CONCLUSIONS: HDL3-C was an independent inverse predictor of PMI in patients who underwent elective PCI.

Clinical Impact of Circulating Irisin on Classified Coronary Plaque Characteristics.

BACKGROUND: Myokines are hormones secreted by skeletal muscles during physical activity. Low myokine levels may contribute to metabolic dysfunction and cardiovascular disorders. Irisin, a newly identified myokine, has been the focus of recent research. The aim of the present study was to analyze the association between circulating irisin levels and tissue characteristics of nonculprit left main coronary artery (LMCA) plaques with the use of integrated backscatter (IB) intravascular ultrasound (IVUS). METHODS: This observational study enrolled 55 Japanese patients following successful percutaneous coronary intervention for lesions in the left anterior descending arteries or left circumflex arteries. Circulating myokine levels, including myostatin, brain-derived neurotrophic factor, and irisin, were measured by an enzyme-linked immunosorbent assay. Tissue characteristics of LMCA plaque were evaluated by IB-IVUS. RESULTS: Circulating irisin levels were negatively associated with percent lipid volume (%LV) [r = -0.31 (95% CI, -2.52 to -0.21), P = 0.02] and positively associated with percent fibrous volume (%FV) [r = 0.32 (95% CI, 0.22-2.20), P = 0.02]. The optimal cutoff value of circulating irisin for the prediction of lipid-rich LMCA plaques was 6.02 µg/mL [area under the curve = 0.713, P < 0.01 (95% CI, 0.58-0.85)]. Multivariate linear regression analysis identified circulating irisin levels as independent predictors for %LV and %FV of the LMCA [ß = -0.29 (95% CI, -2.53 to -0.07), P = 0.04 and ß = 0.30 (95% CI, 0.10-2.23), P = 0.03, respectively]. CONCLUSIONS: Circulating irisin levels are significantly associated with tissue characteristics of nonculprit LMCA plaques.

Impact of post-dilatation on longitudinal stent elongation: An in vitro study.

OBJECTIVES: To evaluate whether balloon inflation for post-dilatation causes longitudinal stent deformation (LSD). METHODS AND RESULTS: Two stents, sized 2.5mm×28mm and 3.5mm×28mm (Nobori®, biodegradable polymer biolimus-eluting stent; Ultimaster®, biodegradable polymer sirolimus-eluting stent; Terumo Co., Tokyo, Japan), were deployed at nominal pressure in straight and tapered silicon vessel models. Then, post-dilatation was performed in two ways: dilatation from the distal (D-P group) or proximal (P-D group) side of the stent. Microscopic findings showed that the stents were elongated during every step of the procedure regardless of the post-dilatation method and type of vessel model. The D-P group showed linear elongation during each step of post-dilatation (straight model: 28.7±0.3mm vs. 29.9±0.3mm, p=0.002; tapered model: 28.0±0.1mm vs. 29.9±0.1mm, p<0.001). In contrast, in the P-D group, the most significant change was observed in the first step of post-dilatation and only slight changes were observed thereafter (straight model: 28.6±0.1mm vs. 29.5±0.1mm, p<0.001; tapered model: 28.2±0.1mm vs. 29.5±0.1mm, p<0.001). Optical frequency domain imaging analysis showed that the frequency of stent strut malapposition was positively correlated with the percentage change in stent length (r=0.74, p<0.0001). CONCLUSION: LSD was observed during every step of post-dilatation in both the straight and tapered vessel models. However, some differences were observed between the D-P and P-D groups. Minimizing stent strut malapposition may reduce the risk of LSD.