Emberger syndrome (ES), an autosomal dominant disorder characterized by congenital deafness, primary lymphedema, and predisposition to myeloid malignancies, is caused by mutations in the GATA2 gene. Although primary lymphedema is an important hallmark of ES, the pathophysiology remains unclear due to the lack of a suitable experimental model. In this study, we isolated induced pluripotent stem cells (iPSCs) from two patients with ES (i.e., ES-iPSCs) and analyzed their in vitro lymphatic differentiation potential via the mesodermal progenitor stage. KDR+ CD34+ early mesodermal progenitors generated from either ES-iPSCs or wild-type iPSCs during a 6-days serum- and feeder-free culture supplemented with bone morphogenetic protein 4 and vascular endothelial growth factor (VEGF) had almost equivalent developmental potential. However, upon co-culture with OP9 stromal cells, KDR+ CD34+ cells derived from ES-iPSCs developed into CD31+ lymphatic vessel endothelial hyaluronan receptor-1+ VEGF receptor 3+ lymphatic endothelial cells less efficiently than KDR+ CD34+ cells derived from wild-type iPSCs. Thus, patient-derived iPSCs recapitulate impairments at an early stage of lymphangiogenesis, making them a useful experimental tool for dissecting the pathophysiology of primary lymphedema in ES and developing potential therapeutic approaches.

Ruptured aneurysms or pseudoaneurysms associated with the vasa corona, in the absence of cerebral arteriovenous malformation or dural arteriovenous fistula, are extremely rare but should be recognized as a possible cause of subarachnoid hemorrhage (SAH). We report the case of a 51-year-old female who presented with SAH due to a ruptured vasa corona pseudoaneurysm, with no associated anatomical abnormalities involving the vertebral artery or the posterior inferior cerebellar artery. She recovered after direct surgery. In previous cases, anterior spinal artery dilatation due to posterior inferior cerebellar artery anomalies prompted endovascular treatment. A thorough assessment remains crucial, and direct surgery can be effective.

BACKGROUND:Infantile-onset Pompe disease (IOPD) is a lethal disorder, but enzyme replacement therapy (ERT) has dramatically improved the prognosis of these patients. CASE SUMMARY:An 18-year-old male patient, who had been diagnosed with IOPD based on decreased α-glucosidase activity in his infancy and received ERT, visited an emergency department at Kyoto University Hospital with poor complexion. An electrocardiogram showed extreme bradycardia and complete atrioventricular block (cAVB), and permanent pacemaker was implanted for cAVB. DISCUSSION:ERT has reduced the risk of death in patients with IOPD; on the other hand, severe cAVB in those patients needs to be recognized as a rare complication. TAKE-HOME MESSAGE:cAVB is a rare but critical complication in long-term survivors of IOPD and deserves attention.

Pulmonary arteriovenous malformations (PAVM) are a major surgical complication of univentricular heart therapy, significantly limiting patient survival. Interruption of hepatic venous drainage into the lungs has been long hypothesized to play a critical role in PAVM development by angiogenesis modulation. To understand human plasma role on PAVM development, we conducted a prospective study analyzing paired plasma samples from superior vena cava (SVC) and hepatic vein (HV) origin from 10 infants with congenital heart disease. Two and three dimensional in vitro arteriovenous models were implemented to compare the angiogenic potential of SVC and HV plasma, alongside untargeted shotgun proteomic profiling. Compared to HV, SVC plasma exhibited a pro-angiogenic profile in vitro. SVC proteome was enriched with pro-angiogenic S100 calcium-binding family proteins (S100A7/A8/A9/P). In contrast, HV plasma had unique expression of actin binding proteins (profilin 1/filamin A/actinin alfa 1), and upregulation of pathways associated with vascular stability and cell adhesion maintenance, suggesting a plausible counteracting role to SVC. Our results emphasize the importance of considering not only angiogenesis, but also extracellular mechano-transduction and vascular tone regulation in PAVM pathogenesis; and highlights the value of in vitro arteriovenous modeling to bridge the gap towards PAVM understanding and advancement of future therapeutical alternatives.

BACKGROUND:DNM1L encodes dynamin-related protein 1, which plays an important role in mitochondrial and peroxisomal division. The DNM1L mutation leads to cardiac dysfunction in patients and animal models. However, the mechanism of cardiac dysfunction caused by DNM1L mutation has not been elucidated clearly at least in the studies of human cardiomyocytes. METHODS:We established human induced pluripotent stem cells (hiPSCs) from two pediatric patients with DNM1L mutation. The hiPSCs were differentiated into hiPSC-derived cardiomyocytes (hiPS-CMs). Mitochondrial morphology and function, cardiomyocyte Ca2+ dynamics, and contractile and diastolic function of hiPS-CMs were analyzed. RESULTS:The morphology of the mitochondria was abnormally elongated in patient-derived hiPS-CMs. The mitochondrial membrane potential and oxygen consumption rate were significantly decreased, resulting in reduced ATP production. In the analysis of Ca2+ dynamics, the 50% time to decay was significantly longer in patient-derived hiPS-CMs than in healthy control. High-precision live-imaging system analysis revealed that contractile and diastolic function was significantly impaired under isoproterenol stimulation. CONCLUSION:DNM1L mutations cause mitochondrial impairment with less production of ATP in cardiomyocytes. This leads to abnormal intracellular Ca2+ dynamics, resulting in contractile and diastolic dysfunction. IMPACT:DNM1L mutations was identified in two pediatric patients who developed cardiac dysfunction and human induced pluripotent stem cells (hiPSCs) were established from these two patients and differentiated into hiPSC-derived cardiomyocytes (hiPS-CMs). DNM1L mutations induced abnormal mitochondrial morphology, mitochondrial dysfunction, and insufficient ATP production in hiPS-CMs. In addition, hiPS-CMs with DNM1L mutation showed abnormal Ca2+ kinetics and impaired contractile and diastolic function. This is the first study that elucidate the mechanism of cardiac dysfunction caused by DNM1L mutations by using hiPSCs.

Background: Idiopathic/heritable pulmonary arterial hypertension (I/H-PAH), a disease with poor prognosis, often faces diagnostic delays due to nonspecific initial symptoms. In Japan, school ECG screening, performed in every 3 years for all students, identifies 41% of school-aged I/H-PAH patients; these typically show elevated pulmonary artery pressure (PAP) but have minimal symptoms (WHO-FC I/II) and preserved RV function. In this disease, the duration from the appearance of pulmonary vascular lesions to PH manifestation and the subsequent onset of RV failure are unknown. Objective: To address the precise pathological trajectory leading to PH manifestation, PH-related ECG findings at diagnosis (Dx) and pre-Dx in pediatric PAH patients were investigated utilizing Japan's school ECG screening system. Methods: Observational study of I/H-PAH patients (6-18 years), newly diagnosed 2005-2020, in Japanese pediatric cardiology teaching hospitals. School ECGs at/pre Dx were evaluated for PH-related findings (R≥1.5mV in V1, RV strain, Q in V1, IRBBB, inverted T waves) according to the criteria from the Japanese Society of Pediatric Cardiology and Cardiac Surgery. Results: In the survey, 118/143 institutions (82.5%) responded; 45 (38%) of those reported cases. 88 patients (51M, 37F; mean age 10.7y) were analyzed. Mode of detection of PAH were school ECG screening (n=36), symptoms (44), family history (5), and incidental findings (3). Compared to nonscreening group, the school screening group had higher proportion of WHO-FC I/II (91% vs 57%, p<.01) and lower plasma BNP (20 vs 102 pg/mL, p<.01), with comparable mean PAP (48 vs 55 mmHg, p=.65) and PVRI (12.1 vs 15.8 WU×m 2 , p=.38). PH-related ECG findings at Dx (n=87), which were present in 93% of all patients, were largely comparable between the groups (95% vs 91, p=.68), with the exception of higher prevalence of Q waves in V1 in the nonscreening (31% vs 15, p<.05) and R≥1.5mV in V1 in the screening group (59% vs 30, p<.05). Crucially, among 35 pre-Dx ECGs, PH-related abnormalities were observed in 77% (10/13) of those taken within 2 years of Dx, versus only 14% (3/22, p<.01) of those taken >2 years prior. Conclusion: Pre-Dx ECG analysis indicates PH-related findings can appear up to 2 years before symptom manifestation, suggesting a 1-2 year window of established PH with minimal symptoms in many pediatric patients. This highlights the potential for earlier diagnosis by performing ECG screening at even shorter intervals.

BACKGROUND/OBJECTIVES:Primary central nervous system lymphoma (PCNSL) has a markedly high proliferation rate, making early diagnosis and prompt therapeutic intervention essential. To accelerate diagnosis, our institution adopted flow cytometry (FCM) in conjunction with conventional histopathology, and this study therefore evaluated the diagnostic performance of FCM for PCNSL. METHODS:We retrospectively analyzed 67 consecutive patients with suspected PCNSL who underwent intraoperative FCM between 2010 and 2023 based on preoperative imaging. B-cell clonality was defined as ≥20% CD19/CD20-positive cells with a κ/λ ratio of >3.0 or <0.5. RESULTS:Using histopathology, we confirmed the presence of PCNSL in 42 patients, all diagnosed as having diffuse large B-cell lymphoma. Six cases (14.3%) were discordant (FCM-D). The sensitivity, specificity, and positive predictive value of FCM were 85.7%, 100%, and 100%, respectively. T-cell markers were significantly elevated in FCM-D cases (p < 0.01), although these were pathologically diagnosed as diffuse large B-cell lymphoma based on histology and immunohistochemistry. CONCLUSIONS:FCM yields reliable diagnostic information within hours of tissue collection and supports early therapeutic decisions in PCNSL. Discordant results may reflect reactive T-cell infiltration. This is the first study to present detailed subset analyses in PCNSL using FCM in correlation with pathology, underscoring its utility as a rapid diagnostic tool.

Abstract Nucleic acid therapeutics hold promise for cancer treatment but have yet to be effectively applied to malignant glioma due to challenges in stability and delivery. We explored the intracerebral delivery of lipid nanoparticles (LNPs) encapsulating siRNA as a drug delivery carrier. Our goal was to develop an siRNA therapy selectively targeting malignant glioma cells by combining LNPs with Fc-domain-binding peptide lipids (FcBPs), a modification system that allows antibody orientation control. Intracerebral Delivery of mRNA-Encapsulated Lipid Nanoparticles: We aimed to deliver mRNA-encapsulated LNPs to the brain using focused ultrasound (FUS) and microbubbles (MB). Following the administration of microbubbles and mRNA-encapsulated LNPs in normal mice, FUS irradiation was performed on the brain. The introduction of firefly luciferase mRNA resulted in gene expression that was 20 times higher in the FUS-irradiated brain compared to the non-irradiated side. Gene expression was observed in vascular endothelial cells throughout the brain and in parenchymal cells outside the brain blood vessels at the FUS-irradiated site. Development of siRNA Medicines: We prepared FcBP-modified siRNA-LNPs targeting integrin αvβ3, highly expressed in malignant gliomas, using siRNA-Luc2. The modifications with FcBP or anti-integrin αvβ3 antibody did not alter the physicochemical properties or polydispersity index (PDI). In vitro, anti-integrin αvβ3-FcBP-modified siRNA-LNPs inhibited the binding of integrin to U87-MG-Luc2 cells, demonstrating high binding affinity and knockdown effects via αvβ3.Discussion:Our findings suggest that the delivery of mRNA-encapsulated LNPs into the brain is feasible using FUS and MB. Additionally, we successfully developed siRNA-LNPs modified with anti-integrin αvβ3 antibody FcBP. Future work will evaluate the cell binding and antitumor effects of siRNA-VEGF encapsulated in these LNPs, aiming to develop new treatments for malignant glioma by enhancing blood-brain barrier penetration and targeting malignant glioma.

Abstract INTRODUCTION Primary central nervous system lymphoma (PCNSL) is characterized by an extremely high proliferation rate, making early diagnosis and therapeutic intervention crucial. To facilitate rapid diagnosis, our hospital has implemented flow cytometry (FCM) in addition to conventional histopathological methods. We evaluated the effectiveness of FCM for the rapid diagnosis of PCNSL. METHODS From 2010 to 2023, we retrospectively analyzed 67 consecutive patients with suspected PCNSL based on preoperative imaging findings, in whom FCM was performed using intraoperative specimens. B cell clonality was determined when CD19 or CD20 was expressed in 20% or more and the κ/λratio was either >3.0 or <0.5. RESULTS Pathological diagnosis confirmed PCNSL in 42 cases, all of which were diagnosed as diffuse large B-cell lymphoma (DLBCL). Six cases (14.3%) of PCNSL patients were discordant with FCM (FCM-D). In contrast, all cases of other tumor types were FCM-D. The sensitivity and specificity of FCM were 85.7% and 100%, respectively. Additionally, T cell surface markers such as CD3, CD4, CD5, and CD8 were significantly elevated in the FCM-D group of PCNSL patients (p<0.01). DISCUSSION FCM results can be obtained within a few hours after tissue collection. With a specificity of 100%, FCM provides a reliable basis for early therapeutic intervention and is anticipated to improve postoperative Karnofsky Performance Status (KPS) and overall survival (OS) in the future. The FCM-D cases among PCNSL patients may have indicated the infiltration of T lymphocytes. Sensitivity could potentially be improved by analyzing multiple samples with FCM. CONCLUSION FCM can serve as a reliable basis for the rapid diagnosis and therapeutic intervention of PCNSL patients.

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.

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.

Background: Dilated cardiomyopathy (DCM) is a progressive myocardial disorder characterized by impaired cardiac contraction and ventricular dilation. However, some patients with DCM improve when experiencing left ventricular reverse remodeling (LVRR). Currently, the detailed association between genotypes and clinical outcomes, including LVRR, particularly among children, remains uncertain. Methods: Pediatric patients with DCM from multiple Japanese institutions recorded between 2014 and 2023 were enrolled. We identified their DCM-related genes and explored the association between gene variants and clinical outcomes, including LVRR. Results: We included 123 pediatric patients (62 males; median age: 8 [1–51] months) and found 50 pathogenic variants in 45 (35.0%) of them. The most identified gene was MYH7(14.0%), followed by RYR2 (12.0%) and TPM1 (8.0%). LVRR was achieved in 47.5% of these patients. The left ventricular ejection fraction remained unchanged (31.4% to 39.8%, P = 0.1913) in patients with sarcomere gene variants but significantly increased in those with nonsarcomere gene variants (33.4% to 47.8%, P = 0.0466) and those without gene variants (33.6% to 54.1%, P = 0.003). Conclusions: LVRR was not uniform across functional gene groups. Hence, an individualized gene-guided prediction approach may be adopted for children with DCM.

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.

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 +/- 7.42mm/m(2), 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 +/- 2.84mm/m(2)) compared to the native unilateral PAs (16.11 +/- 5.46mm/m(2), P < 0.0001) and non-rbMAPCA (10.13 +/- 4.44mm/m(2), 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.

BACKGROUND: Living-donor lobar lung transplantation (LDLLT) remains a life-saving option for pedi-atric patients with respiratory failure. However, the long-term survival and post-transplant quality of adult lobar grafts transplanted into children are unknown. Therefore, this study aimed to evaluate the outcomes of pediatric LDLLT and post-transplant graft growth. METHODS: We retrospectively reviewed the prospectively collected clinical data of 25 living-donor lung transplantations performed in 24 pediatric recipients aged <= 17 years. The annual pulmonary function test data and computed tomography scans of 12 recipients, followed up for >5 years without significant complications, were used to evaluate growth in height, graft function, and radiological changes. The Kaplan-Meier method and simple linear regression were performed for analysis. RESULTS: Bilateral lower lobe transplantation was performed in 12 patients, unilateral lower lobe trans-plantation in 12, and bilateral middle lobe transplantation in 1. The median volumetric size matching at transplantation was 142% (range, 54%-457%). The 5-and 10-year overall survival rates were 87.7% and 75.1(sic), respectively. Chronic lung allograft dysfunction occurred in 2 patients. During a median follow-up of 6 years, the median increases in height and vital capacity were 14.4% (range, 0.80%-43.5%) and 58.5% (range, 6.7%-322%), respectively. Graft weight was positively correlated with graft volume (r(2)=0.622, p<0.001) after the graft volume exceeded the original lobar volume in the donor. CONCLUSIONS: This study shows that pediatric LDLLT offers satisfactory long-term survival, with the growth of mature adult lobes transplanted into growing children. J Heart Lung Transplant 2023;42:660-668 (c) 2022 International Society for Heart and Lung Transplantation. All rights reserved.

The prognosis of primary central nervous system lymphoma (PCNSL) in the elderly remains poor. We aimed to evaluate the outcome of rituximab, methotrexate, procarbazine, and vincristine (RMPV) chemotherapy in elderly patients with new-onset PCNSL. Twenty-eight patients aged ≥ 70 years treated for PCNSL between 2010 and 2020 were examined retrospectively. Nineteen patients received RMPV and nine did not qualify. Patients received five to seven cycles of RMPV plus response-adapted whole-brain radiotherapy (WBRT) and cytarabine. Ten of the 19 patients who received RMPV (52.6

Objective: The preset study evaluated the outcome of living-donor segmental lung transplantation for pediatric patients. Methods: Between August 2009 and May 2021, we performed living-donor segmental lung transplantation in 6 critically ill pediatric patients, including 1 patient on a ventilator alone and another patient on a ventilator and extracorporeal membrane oxygenation (ECMO). There were 4 male and 2 female patients, with a median age of 7 years (range, 4-15 years) and a median height of 112.7 cm (range, 95-125.2 cm). The diagnoses included complications of allogeneic hematopoietic stem cell transplantation (n = 4) and pulmonary fibrosis (n = 2). All patients received bilateral lung transplantation under cardiopulmonary bypass. A basal segment and a lower lobe were implanted in 3 patients, and a basal segment and an S6 segment were implanted in the other 3 patients. In 2 patients, the right S6 segmental graft was horizontally rotated 180 degrees and implanted as the left lung. Results: Among the 9 segmental grafts implanted, 7 functioned well after reperfusion. Two rotated S6 segmental grafts became congestive, with 1 requiring graft extraction and the other venous repair, which was successful. There was 1 hospital death (14 days) due to sepsis and 1 late death (9 years) due to leukoencephalopathy. The remaining 4 patients are currently alive at 9 months, 10 months, 1.3 years, and 1.9 years. Conclusions: Living-donor segmental lung transplantation was a technically difficult but feasible procedure with acceptable outcomes for small pediatric patients with chest cavities that were too small for adult lower lobe implantation.