HOIL-1L deficiency induces cell cycle alteration which causes immaturity of skeletal muscle and cardiomyocytes.

HOIL-1L deficiency was recently reported to be one of the causes of myopathy and dilated cardiomyopathy (DCM). However, the mechanisms by which myopathy and DCM develop have not been clearly elucidated. Here, we sought to elucidate these mechanisms using the murine myoblast cell line C2C12 and disease-specific human induced pluripotent stem cells (hiPSCs). Myotubes differentiated from HOIL-1L-KO C2C12 cells exhibited deteriorated differentiation and mitotic cell accumulation. CMs differentiated from patient-derived hiPSCs had an abnormal morphology with a larger size and were excessively multinucleated compared with CMs differentiated from control hiPSCs. Further analysis of hiPSC-derived CMs showed that HOIL-1L deficiency caused cell cycle alteration and mitotic cell accumulation. These results demonstrate that abnormal cell maturation possibly contribute to the development of myopathy and DCM. In conclusion, HOIL-1L is an important intrinsic regulator of cell cycle-related myotube and CM maturation and cell proliferation.

Surgical treatment for a child with traumatic ventricular septal rupture and ventricular aneurysm.

A five-year-old boy was diagnosed with the ventricular septal rupture and ventricular aneurysm after blunt chest trauma in child abuse. Because of the intractable heart failure, he underwent operation in subacute period. Postoperative course was uneventful. The blunt cardiac injury in children can be caused by mild trauma and can be lethal. Surgical intervention should be considered when the clinical condition is unstable.

Chloroquine decreases cardiac fibrosis and improves cardiac function in a mouse model of Duchenne muscular dystrophy.

BACKGROUND: Duchenne muscular dystrophy (DMD), a severe degenerative skeletal and cardiac muscle disease, has a poor prognosis, and no curative treatments are available. Because decreased autophagy has been reported to contribute to skeletal muscle degeneration, therapies targeting autophagy are expected to improve skeletal muscle hypofunction. However, the role of this regulatory mechanism has not been evaluated clearly in DMD cardiomyocytes. METHODS: In this present study, we evaluated myocardial fibrosis and its mechanism in mdx mice, a model of DMD, and also evaluated changes in cardiac function. RESULTS: As assessed by LC3 immunohistochemistry, a small number of autophagosomes were detected in cardiomyocytes of both mdx mice and control wild-type (WT) mice. The number of autophagosomes was significantly enhanced by 4 weeks of isoproterenol-induced cardiac stress in cardiomyocytes of mdx but not WT mice. Simultaneously, isoproterenol increased cardiomyocyte fibrosis in mdx but not WT mice. Administration of chloroquine significantly decreased cardiomyocyte fibrosis in mdx mice, even after isoproterenol treatment. Left ventricle size and function were evaluated by echocardiography. Left ventricular contraction was decreased in mdx mice after isoproterenol treatment compared with control mice, which was alleviated by chloroquine administration. CONCLUSIONS: Heart failure in DMD patients is possibly treated with chloroquine, and the mechanism probably involves chloroquine's anti-inflammatory effects.

Influence of the bronchus on the vascular growth of major aortopulmonary collateral arteries.

OBJECTIVES: Patients with major aortopulmonary collateral arteries (MAPCAs) often require additional surgical or catheter intervention after unifocalization (UF) due to stenosis and poor growth. We hypothesized that the UF design influences vascular growth; assessment was based on the passing route related to the bronchus. METHODS: We enrolled 5 patients with pulmonary atresia (PA), ventricular septal defect and MAPCA who underwent UF and subsequent definitive repair at our institute from 2008 to 2020. Angiography and computed tomography scans were routinely performed before surgical intervention to clarify pulmonary circulation and the relationships between MAPCAs and the bronchus, which revealed peculiar MAPCAs directed to the pulmonary hilum passing behind the bronchus (defined as retro-bronchial MAPCAs; rbMAPCAs). Vascular growth of rbMAPCAs, non-rbMAPCAs and the native pulmonary artery were assessed using the angiograms before and after repair. RESULTS: The angiogram before UF [age 42 (24-76) days, body weight 3.2 (2.7-4.2) kg] showed that the diameter of the original unilateral PA, rbMAPCA and non-rbMAPCA was 19.95 ± 6.65, 20.72 ± 5.36 and 20.29 ± 7.42 mm/m2, respectively (P = 0.917). UF was completed in a single-stage with the placement of modified Blalock-Taussig shunt through median sternotomy at the age of 1.6 (1.0-2.5) months. Angiograms performed 3.0 (1.0-10.0) years after UF completion demonstrated a smaller rbMAPCA diameter at peri-bronchial region (3.84 ± 2.84 mm/m2) compared to the native unilateral PAs (16.11 ± 5.46 mm/m2, P < 0.0001) and non-rbMAPCA (10.13 ± 4.44 mm/m2, P = 0.0103). CONCLUSIONS: RbMAPCAs tend to be stenosed at the point where they cross the bronchus and emerge in the middle mediastinum after in situ UF.

Hemodynamic and Clinical Response to Liver Transplantation in Children and Young Adults POPH Patients.

Portopulmonary hypertension is an intractable form of pulmonary hypertension. Although liver transplantation is recommended for patients who respond poorly to treatments, the mechanisms by which liver transplantation improves pulmonary hypertension remain unclear. The present study investigated these mechanisms by retrospectively evaluating patients' data. This study retrospectively evaluated echocardiography and catheterization data before and after liver transplantation in 12 patients who underwent liver transplantation from 2001 to 2019. The 12 patients included one male and 11 females, of median age at liver transplantation of 10 years, 2 months. Nine patients underwent liver transplantation for congenital biliary atresia and three for portal vein aplasia or hypoplasia. Mean pulmonary arterial pressure was 44.1 ± 8.1 mmHg at the first cardiac catheter examination, 35.3 ± 7.8 mmHg before liver transplantation, and 29.5 ± 9.3 mmHg 6 months after liver transplantation. Pulmonary artery pressure was reduced by treatments of pulmonary hypertension and by liver transplantation. Pulmonary vascular resistance did not differ before and after liver transplantation, whereas the cardiac index decreased significantly, indicating that the significant reduction in mean pulmonary artery pressure was due to a decrease in cardiac index. Decreased cardiac index was thought to result from improvements in hyperdynamic conditions due to increased (normalized) systemic vascular resistance. Liver transplantation likely suppresses shear stress on pulmonary arteries, preventing further damage by hyper-circulation. A longer-term evaluation is required to determine the effect of improving pulmonary artery remodeling.

Atrial septal defect occlusion by a catheter during reperfusion of homograft in a case of living donor liver transplantation.

Liver transplantation for patients with atrial septal defect and pulmonary artery stenosis, causing high right atrium pressure, raises concerns about embolism in systemic vessels during reperfusion of the donor liver graft. Temporal atrial septal defect occlusion by a catheter is a simple and easy method of preventing the complication.

Phenotype-Based High-Throughput Classification of Long QT Syndrome Subtypes Using Human Induced Pluripotent Stem Cells.

For long QT syndrome (LQTS), recent progress in genome-sequencing technologies enabled the identification of rare genomic variants with diagnostic, prognostic, and therapeutic implications. However, pathogenic stratification of the identified variants remains challenging, especially in variants of uncertain significance. This study aimed to propose a phenotypic cell-based diagnostic assay for identifying LQTS to recognize pathogenic variants in a high-throughput manner suitable for screening. We investigated the response of LQT2-induced pluripotent stem cell (iPSC)-derived cardiomyocytes (iPSC-CMs) following IKr blockade using a multi-electrode array, finding that the response to IKr blockade was significantly smaller than in Control-iPSC-CMs. Furthermore, we found that LQT1-iPSC-CMs and LQT3-iPSC-CMs could be distinguished from Control-iPSC-CMs by IKs blockade and INa blockade, respectively. This strategy might be helpful in compensating for the shortcomings of genetic testing of LQTS patients.

The intracellular Ca2+ concentration is elevated in cardiomyocytes differentiated from hiPSCs derived from a Duchenne muscular dystrophy patient.

Duchenne muscular dystrophy (DMD) is the most common and severe form of muscular dystrophy. The major symptoms of this condition are walking difficulties, dyspnea caused by progressive skeletal muscle weakness, and cardiomyopathy. Recent advances in ventilator support devices have dramatically decreased mortality caused by respiratory distress. Consequently, cardiomyopathy resulting in heart failure is currently the major cause of death among DMD patients. One mechanism by which skeletal muscle is damaged in DMD patients involves elevation of the intracellular Ca2+ concentration. By contrast, the mechanisms underlying the development of cardiomyopathy are unclear. To investigate this, we examined the intracellular Ca2+ concentration and calcium transients in cardiomyocytes differentiated from human induced pluripotent stem cells (hiPSCs). hiPSCs were derived from a DMD patient (DMD-hiPSCs), in whom exon 44 of the gene encoding dystrophin was deleted, and from his parents (control-hiPSCs), who did not carry this mutation. The intracellular Ca2+ concentration was measured using the fluorescent indicator indo-1. The fluorescence ratio (410/490 nm) of indo-1 at rest (R0), the peak of this ratio (Rmax), and the amplitude (Rmax-R0) were significantly higher in cardiomyocytes differentiated from DMD-hiPSCs than in those differentiated from control-hiPSCs. Moreover, mechanical stretching significantly increased the intracellular Ca2+ concentration in cardiomyocytes differentiated from DMD-hiPSCs, but not in those differentiated from control-hiPSCs. These findings indicate that elevation of the intracellular Ca2+ concentration can cause cardiac damage leading to cardiomyopathy in DMD patients.

Urethral function and histopathology in aged female rats as a stress urinary incontinence model.

OBJECTIVE: Stress urinary incontinence (SUI) is a common disease condition in elderly women, suggesting that its etiology may be linked to aging. To investigate the hypothesis that urethral dysfunction and histopathological changes are possible contributors to SUI in elderly women, several parameters of urethral function, as well as histological parameters, were compared between young and aged rats. METHODS: Virgin female rats were examined at 3 different ages, namely 3, 12, and 24 months, corresponding to young, middle-aged, and aged rats, respectively. Urethral function was assessed by measuring the leak point pressure (LPP), pudendal nerve stimulation (PNS)-induced elevation in urethral pressure, and phenylephrine-induced increase in urethral perfusion pressure (UPP). Histopathological assessments were performed following hematoxylin and eosin (HE), Masson's trichrome, and immunofluorescence staining of urethral tissue. RESULTS: LPP of aged rats was significantly reduced compared to that of both young and middle-aged rats. PNS-induced elevation in urethral pressure in aged rats was also significantly lower than that in young rats. In contrast, there were no significant differences in the phenylephrine-induced increase in UPP between young and aged rats. Connective tissue area in the external urethral sphincter (EUS) layer was increased in aged rats, whereas the smooth muscle layer was histologically similar to that in young rats. The number of EUS fibers was significantly reduced in aged rats, whereas the cross-sectional area of EUS fibers increased from differed compared with young rats. CONCLUSION: We have demonstrated age-related changes in EUS function and morphology in the rat urethra, which are considered to be etiological risk factors for SUI in humans.

Living-donor lung transplantation after surgical repair of transposition of the great arteries.

Pediatric pulmonary hypertension after surgery for congenital heart disease is a significant complication. We present a case of living-donor lung transplantation for a 12-year-old girl with pulmonary hypertension after surgical repair of transposition of great arteries. Despite repairing the transposition of great arteries, her growth was severely restricted because of progressive pulmonary hypertension; thus, lung transplantation was discussed. Standard bilateral lobar transplantation seemed unfeasible due to oversized grafts, so we performed a single lobar transplantation. Unexpectedly, she developed complications and died 3 months postoperatively despite another emergent lobar transplantation. We discussed the challenges and potential solutions regarding lobar size mismatching.

Nitrogen oxide cycle regulates nitric oxide levels and bacterial cell signaling.

Nitric oxide (NO) signaling controls various metabolic pathways in bacteria and higher eukaryotes. Cellular enzymes synthesize and detoxify NO; however, a mechanism that controls its cellular homeostasis has not been identified. Here, we found a nitrogen oxide cycle involving nitrate reductase (Nar) and the NO dioxygenase flavohemoglobin (Fhb), that facilitate inter-conversion of nitrate, nitrite, and NO in the actinobacterium Streptomyces coelicolor. This cycle regulates cellular NO levels, bacterial antibiotic production, and morphological differentiation. NO down-regulates Nar and up-regulates Fhb gene expression via the NO-dependent transcriptional factors DevSR and NsrR, respectively, which are involved in the auto-regulation mechanism of intracellular NO levels. Nitrite generated by the NO cycles induces gene expression in neighboring cells, indicating an additional role of the cycle as a producer of a transmittable inter-cellular communication molecule.

Portable Micropatterns of Neuronal Cells Supported by Thin Hydrogel Films.

A grid micropattern of neuronal cells was formed on a free-standing collagen film (35 µm thickness) by directing migration and extension of neurons along a Matrigel pattern previously prepared on the film by the microcontact printing method. The neurons migrated to reach the nodes on the grid pattern and extended neurites to bridge cell bodies at the nodes. The resulting neuronal micropattern on the collagen film containing culture medium can be handled and deformed with tweezers with maintenance of physiological activity of the neurons, as examined by response of cytosolic Ca2+ concentration to a dose of bradykinin. This portability is the unique advantage of the present system that will open novel possibility of cellular engineering including the on-demand combination with analytical devices. The repetitive lamination of the film on a microelectrode chip was demonstrated for local electrical stimulation of a specific part of the grid micropattern of neurons, showing Ca2+ wave propagation along the neurites. The molecular permeability is the further advantage of the free-standing hydrogel substrate, which allows external supply of nutrients and dosing with chemical stimulants through the film even under rolled and laminated conditions.

Inhibitory effect of the selective serotonin 5-HT3 receptor antagonist ramosetron on duodenal acidification-induced gastric hypersensitivity in rats.

Irritable bowel syndrome (IBS) and functional dyspepsia (FD) are both functional gastrointestinal disorders and frequently co-occur in patients. While one cause of FD appears to be gastric hypersensitivity, whether the hypersensitivity is affected by IBS treatments remains unclear, given the lack of appropriate animal models for testing. Here, we established an experimental model of duodenal acidification-induced gastric hypersensitivity in conscious rats. The model involved duodenal acidification induced by the infusion of hydrochloric acid into the proximal duodenum, with the nociceptive response being determined as the change in mean arterial pressure (MAP) during gastric distension via an indwelling latex balloon. Using our model we evaluated the effects of duodenal acidification, increased distension pressure, and orally administered therapeutic agents for IBS with diarrhea (IBS-D). Duodenal acidification enhanced the pressor response during gastric distension, and pretreatment with the opioid κ-receptor agonist fedotozine (10mg/kg, intra-arterial) inhibited the pressor response. Pressure levels of 15-60 mm Hg increased MAP in response to gastric distension. The serotonin 5-HT3 receptor antagonist ramosetron (30 µg/kg) inhibited MAP increase induced by duodenal acidification, with no other IBS-D therapeutic agents showing any effect. In contrast, the serotonin 5-HT3 receptor agonist m-chlorophenylbiguanide (1mg/kg) significantly enhanced the pressor response during gastric distension. These findings indicate that the serotonin 5-HT3 receptor plays a key role in duodenal acidification-induced gastric hypersensitivity in rats, suggesting that ramosetron may reduce FD symptoms by ameliorating sensitized gastric perception.

Hydration structure of CO2-absorbed 2-aminoethanol studied by neutron diffraction with the 14N/15N isotopic substitution method.

Neutron diffraction measurements were carried out for CO2-absorbed aqueous 11 mol % 2-aminoethanol (MEA) D2O solutions (corresponding to 30 wt % MEA solution) in order to obtain information on both the intramolecular structure and intermolecular hydration structure of the MEA carbamate molecule in the aqueous solution. Neutron scattering cross sections observed for (MEA)0.11(D2O)0.89, (MEA)0.11(D2O)0.89(CO2)0.06, and (MEA)0.11(D2O)0.89(DCl)0.11 solutions with different (14)N/(15)N ratios were used to derive the first-order difference function, ΔN(Q), which involves environmental structural information around the nitrogen atom of the MEA molecule. Intramolecular geometry and intermolecular hydration structure of MEA, protonated MEA (MEAD(+)), and MEA carbamate (MEA-CO2) molecules were obtained through the least-squares fitting of the observed Δ(N)(Q) in the high-Q region and the intermolecular difference function, Δ(N)(inter)(Q), respectively. In the aqueous solution, the MEA molecule takes the gauche conformation (dihedral angle, ∠NCCO = 45 ± 3°), suggesting that an intramolecular hydrogen bond is formed. On the other hand, values of the dihedral angle ∠NCCO determined for MEAD(+) and MEA-CO2 molecules were 193 ± 4° and 214 ± 8°, respectively. These results imply that the intermolecular hydrogen bonds are dominated for MEAD(+) and MEA-CO2 molecules. The intermolecular nearest neighbor N···O(D2O) distance for the MEA molecule was determined to be 3.13 ± 0.01 Å, which suggests weak intermolecular interaction between the amino-nitrogen atom of MEA and water molecules in the first hydration shell. The nearest-neighbor N···O(D2O) distances for MEAD(+) and MEA-CO2 molecules, 2.79 ± 0.03 and 2.87 ± 0.04 Å, clearly indicate strong hydrogen bonds are formed among the amino group of these molecules and neighboring water molecules.

Ventricular septal defect complicated by infantile lobar emphysema.

An infant with ventricular septal defect, atrial septal defect, and Down syndrome (trisomy 21) was referred to our institution. The patient gradually developed infantile lobar emphysema of the right upper lobe and became dependent on mechanical ventilator support. When the patient was five months of age, we successfully performed a total correction of cardiac anomalies combined with a right upper lobectomy. The resection of the right upper lobe was performed under cardiopulmonary bypass through a median sternotomy. There was no major airway bleeding or mediastinitis. The patient was extubated on postoperative day 12 and discharged on postoperative day 29.

Effect of interfacial serum proteins on melanoma cell adhesion to biodegradable poly(l-lactic acid) microspheres coated with hydroxyapatite.

We have measured the interaction forces between a murine melanoma cell and a poly(l-lactic acid) (PLLA) microsphere coated with/without hydroxyapatite (HAp) nanoparticles (i.e., an HAp/PLLA or a bare PLLA microsphere) in a serum-free culture medium, using atomic force microscopy (AFM) with colloid probe technique, in order to investigate how the HAp-nanoparticle coating as well as interfacial serum proteins influence the cell-microsphere adhesion. The cell adhesion force of the HAp/PLLA microspheres was 1.4-fold stronger than that of the bare PLLA microspheres. When the microspheres were pretreated with a culture medium supplemented with 10% fetal bovine serum, the cell adhesion force of the HAp/PLLA microspheres was increased by a factor of 2.1; in contrast, no change was observed in the cell adhesion force of the bare PLLA microspheres before/after the pretreatment. Indeed, the cell adhesion force of the HAp/PLLA was 2.8-fold larger than that of the bare PLLA after the pretreatment. Additionally, we have investigated the effect of interfacial serum proteins on the zeta potentials of these microspheres. On the basis of the obtained results, possible mechanism of cell adhesion to the HAp/PLLA and bare PLLA microspheres in the presence/absence of the interfacial serum proteins is discussed.

Left pulmonary artery banding to repair ipsilateral diffuse pulmonary arteriovenous fistula.

Congenital pulmonary arteriovenous fistula (PAVF) is a rare disease which causes hypoxemia by shunting deoxygenated blood from the pulmonary artery into pulmonary venous return. Lung transplantation is the most effective therapy to treat severe, diffuse PAVF. However, the availability of lungs for transplantation is limited in most parts in the world. For patients with diffuse PAVF affecting only one side of the lungs, ipsilateral pulmonary artery banding (PAB) is an effective treatment, but not yet standard of care. We report successful treatment of a patient with diffuse left-sided PAVF with PAB. We believe that PAB is an effective therapy for severe unilateral PAVF and may serve as a bridge to lung transplantation.

Inhibitory effect of ramosetron on corticotropin releasing factor- and soybean oil-induced delays in gastric emptying in rats.

BACKGROUND AND AIM: Symptoms of functional dyspepsia (FD) are highly prevalent in patients with irritable bowel syndrome (IBS). However, the effects of therapeutic agents for IBS on the pathophysiology of FD are unclear. In this study, therefore, we examined the effects of ramosetron, a serotonin 5-HT(3) receptor antagonist, on corticotropin releasing factor (CRF)- and soybean oil-induced delays in gastric emptying of rats, in comparison with anti-diarrheal agent and spasmolytics. The involvement of 5-HT and the 5-HT(3) receptor in delayed gastric emptying was also evaluated. METHODS: Corticotropin releasing factor was administered intravenously to rats 10min before oral administration of 0.05% phenol red solution, and the amount remaining in the stomach was measured after 30min. Soybean oil was administered orally with glass beads, and the number of residual beads in the stomach was counted 1h later. RESULTS: Both CRF and soybean oil inhibited gastric emptying dose-dependently. Ramosetron and itopride, a gastro-prokinetic agent, significantly reduced both CRF- and soybean oil-induced delays in gastric emptying, while an anti-diarrheal agent and spasmolytics aggravated them. Pretreatment with p-chlorophenylalanine for 2days to reduced the synthesis of endogenous 5-HT diminished the effects of both CRF and soybean oil on gastric emptying. A 5-HT(3) receptor agonist m-chlorophenylbiguanide suppressed gastric emptying of both phenol red and glass beads, and those effects were reversed by ramosetron. CONCLUSIONS: These results suggest that CRF and soybean oil suppress gastric emptying in rats by activating 5-HT(3) receptors, and that by antagonizing these receptors, ramosetron may ameliorate symptoms of FD in clinical settings.