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Background: Idiopathic pulmonary arterial hypertension (IPAH) is a progressive disease caused by vascular remodeling of the pulmonary arteries with elevated pulmonary vascular resistance. Recently, various pulmonary vasodilator drugs have become available in the clinical field, and have dramatically ameliorated the prognosis of IPAH. However, little is known about how the mechanical properties of pulmonary arterial smooth muscle cells (PASMCs) are altered under drug supplementation. Methods: Atomic force microscopy (AFM) was used to investigate the mechanical properties of PASMCs derived from a patient with IPAH (PAH-PASMCs) and a healthy control (N-PASMCs) which received the supplementation of clinically used drugs for IPAH: sildenafil, macitentan, and riociguat. Results: PASMCs derived from PAH-PASMCs were stiffer than those derived from N-PASMCs. With sildenafil treatment, the apparent Young’s modulus ( E 0 ) of cells significantly decreased in PAH-PASMCs but remained unchanged in N-PASMCs. The decrease in E 0 of PAH-PASMCs was also observed in macitentan and riociguat treatment. The stress relaxation AFM revealed that the decrease in E 0 of PAH-PASMCs resulted from a decrease in the cell elastic modulus and/or increase in cell fluidity. The combination treatment of macitentan and riociguat showed an additive effect on cell mechanical properties, implying that this clinically accepted combination therapy for IPAH influences the intracellular mechanical components. Conclusions: Pulmonary vasodilator drugs affect the mechanical properties of PAH-PASMCs, and there exists a mechanical effect of combination treatment on PAH-PASMCs. Cardiol Res. 2021;12(4):231-237 doi: https://doi.org/10.14740/cr1282

BACKGROUND Bronchopulmonary dysplasia (BPD) is a chronic lung disease in premature neonates. Classical BPD is caused by hyperoxia and high-pressure mechanical ventilation, whereas BPD in recent era is caused by impaired pulmonary angiogenesis and alveolarization in extreme prematurity. Although sildenafil was reported to be effective in a hyperoxia-induced rat BPD model, several clinical trials could not demonstrate any significant improvement in the respiratory statuses of BPD infants. Riociguat is a soluble guanylate cyclase stimulator that increases cyclic guanosine monophosphate activity in a nitric oxide independent manner. However, a beneficial effect in BPD has not been established yet. METHODS AND RESULTS We established BPD model in rats by injection of SU5416 on day 1 followed by maintenance under normoxia, which resulted in oversimplified alveoli, sparse pulmonary capillary vessels, severe pulmonary hypertension, and growth retardation, which mimicked the features observed in recent clinical management of BPD. We administered riociguat from day 10, when BPD rats exhibited growth retardation. Histological analyses demonstrated that riociguat treatment significantly but partially ameliorated lung alveolarization, vascularization, and pulmonary hypertension. However, the survival rate was not significantly improved by riociguat treatment. CONCLUSIONS Riociguat could ameliorate pulmonary alveolarization, vascularization, and hypertension in the SU5416 induced BPD rat model, but could not improve the overall survival.

OBJECTIVES This study aimed to identify the histological characteristics associated with bridge to recovery using Berlin Heart EXCOR® (BHE) in paediatric patients METHODS Of the 10 consecutive patients RESULTS The degree of myocardial fibrosis was significantly lower, and the capillary vascular density was significantly higher in the recovery group than in the non-recovery group [16% (standard deviation 5.9%) vs 28% (5.9%), P = 0.021, and 65 (11) vs 43 (18) units/high-power field, P = 0.037, respectively]. The changes during 3 months after BHE implantation in LV diastolic dimension (z-score) and ejection fraction were significantly greater in the recovery group than in the non-recovery group [−9.6 (3.5) vs −3.6 (4.5), P = 0.045, and 36% (13%) vs 13% (13%), P = 0.032, respectively]. CONCLUSIONS In paediatric patients

Background Restrictive cardiomyopathy (RCM) is characterized by impaired ventricular relaxation. Although several mutations were reported in some patients, no mutations were identified in cardiomyocyte expressing genes of other patients, indicating that pathological mechanisms underlying RCM could not be determined by cardiomyocytes only. Cardiac fibroblasts (CFs) are a major cell population in the heart; however, the pathological roles of CFs in cardiomyopathy are not fully understood.Methods and Results:This study established 4 primary culture lines of CFs from RCM patients and analyzed their cellular physiology, the effects on the contraction and relaxation ability of healthy cardiomyocytes under co-culture with CFs, and RNA sequencing. Three of four patients hadTNNI3mutations. There were no significant alterations in cell proliferation, apoptosis, migration, activation, and attachment. However, when CFs from RCM patients were co-cultured with healthy cardiomyocytes, the relaxation velocity of cardiomyocytes was significantly impaired both under direct and indirect co-culture conditions. RNA sequencing revealed that gene expression profiles of CFs in RCM were clearly distinct from healthy CFs. The differential expression gene analysis identified that several extracellular matrix components and cytokine expressions were dysregulated in CFs from RCM patients. Conclusions The comprehensive gene expression patterns were altered in RCM-derived CFs, which deteriorated the relaxation ability of cardiomyocytes. The specific changes in extracellular matrix composition and cytokine secretion from CFs might affect pathological behavior of cardiomyocytes in RCM.

Patients with pulmonary atresia and ventricular septal defect (PA/VSD) demonstrate a wide variety of pulmonary and coronary arteries; additionally, coronary pulmonary artery fistula (CPAF) is a rare manifestation of PA/VSD and is seldom detected during pregnancy. In this report, we present a case of prenatal diagnosis of CPAF in PA/VSD with showing the present impactful images of CPAF in a neonate, which were obtained by using fetal echocardiography and postnatal electrocardiography-gated 320-row computed tomography. Prenatal diagnosis of CPAF can facilitate the provision of better therapeutic strategies after birth.

Pure restrictive cardiomyopathy is a strong risk factor for poor outcomes in children with cardiomyopathy on ventricular assist devices. Owing to concomitant right heart failure, children with end-staged restrictive cardiomyopathy who are supported with a ventricular assist device often require a biventricular assist device, which is another risk factor for waitlist mortality in heart transplantation candidates. Herein, we report the case of a 3-year-old boy with pure restrictive cardiomyopathy who successfully underwent heart transplantation after 12 months of support with staged biventricular assist devices. Owing to the progression of diastolic dysfunction, the left ventricular assist device could not provide adequate circulation support. Despite the provision of biventricular assist device support, the patient required a complex management strategy that involved balancing the left and right ventricular assist device supports. We were able to stabilize the patient by careful synchronization of the supports and proceeded to heart transplantation. TRIAL REGISTRATION: Clinical Registration No.: Institutional Review Board of Osaka University Hospital, approval no. 16105.

CIC-rearranged undifferentiated small round cell sarcoma (CIC-rearranged USRCS) is a recently established type of Ewing-like small round cell sarcomas, characterized by CIC gene rearrangement, most commonly CIC-DUX4 fusion. This report presents the second case of CIC-rearranged USRCS arising primarily in the cerebrum. A 64-year-old otherwise healthy woman presented with a 1 × 1 cm sized hemorrhagic subcortical tumor in the left temporo-parietal lobe. The tumor repeatedly recurred, and the patient underwent three surgeries, chemotherapy with doxorubicin and ifosfamide, and radiotherapy, as well as gamma knife surgery. Systemic examination revealed no other extracranial masses. Imprint cytology revealed small to moderate-sized round-to-ovoid tumor cells with mild pleomorphism and variations in size and shape. The nuclei contained finely granular chromatin, and some had easily-recognizable nucleoli. The tumor exhibited a mainly cytoplasmic pattern of CD99 immunostaining, rather than a diffuse membranous pattern. The tumor also exhibited diffuse positivity for calretinin and p16, as well as partial positivity for WT1 (nuclear and cytoplasmic staining pattern) and D2-40. FISH assessment showed CIC split signals. In conclusion, CIC-rearranged USRCSs can occur primarily in the cerebrum. It would be impossible to diagnose them through cytology alone, but cytology would be useful to rule out other small round cell brain tumors including gliomas, lymphomas, carcinomas, and germinoma. Immunohistochemical analysis including tests for CD99, calretinin, and WT1 would help to suggest CIC-rearranged USRCSs and distinguish them from Ewing sarcomas. Additionally, immunohistochemistry for p16 might be useful in the diagnosis. Diagn. Cytopathol. 2016;44:828-832. © 2016 Wiley Periodicals, Inc.

Purpose Epstein-Barr Virus (EBV) is well known to be associated with posttransplant lymphoproliferative disorders (PTLD). Due to moderate immunosuppression in heart transplant (HTx) recipients, many viruses other than EBV are also infected and activated and assessment of simultaneous viral loads of other viruses is also important. In the present study, the role of simultaneous assessment of plasma and peripheral mononuclear cells for multiple viral load quantification in peripheral blood of pediatric patients after heart transplantation was elucidated. Methods In a period of 9 months (from January to October 2009) we collected 28 samples of 3 adult and 6 pediatric cardiac transplant recipients. Quantitative polymerase chain reaction for EBV, Cytomegalovirus, Parvovirus 19 (B19), BK and JC virus, and HHV6 was simultaneously performed based on the TaqMan system using DNA which isolated from peripheral mononuclear cells (PBMCs) and plasma, as described elsewhere. Phenotyping of infected lymphocytes were also done. CD8, CD19, CD45RO, CD57, etc. Results Case 1: 2-year old girl who underwent HTx at 1 year of her age, developed B cell type PTLD and died of cytokine release syndrome, showed rapid increases in EBV load in both PMBC and plasma as well as HHV 6 and B19. Case 2: 22-year female with subclinical infection for 19 years after HTx, showed sudden decrease in both in EBV load in both PMBC and plasma after conversion of everolimus to mycophenolate mofetil and both EBV loads eliminated after cytotoxic T lymphocyte (CTL) formation against EBV. Three patients (clinical in 2 and subclinical in 1) showed an increase in B19 load. Although B19 load became negative in 2 patients, B19 load sustained positive for 9 months in one patients who experienced B cell type PLTD 6 years ago and had continuous B19 infection. In one 38-year old male with steroid treatment for nephrosis and one 11-year old boy with living renal transplantation 10 years after HTx showed increases in BK and JC virus loads and an rapid decrease in both viral loads after reduction of immunosuppression. Conclusion Simultaneous multiple viral load quantifications are useful to evaluate immunosuppressive status of HTx recipients as well as monitoring of viral infection itself.

The clarification of public concerns regarding heart transplantation is important for improving low organ donation rates in Japan. In the present study, we used the Twitter data of 4986 tweets (between August 2015 and January 2016) and 1429 tweets (between April 2016 and May 2016) to analyze public discourse on heart transplantation in Japan and identify the reasons for low organ donation rates. We manually categorized all tweets relevant to heart transplantation into nine categories and counted the number of tweets in each category per month. During the study period, the most popular category of tweets was related to the media, followed by money (tweets questioning or even criticizing the high price of fundraising goals to go overseas for heart transplantations), while some tweets were misconceptions. We also conducted a sentiment analysis, which revealed that the most popular negative tweets were related to money, while the most positive tweets were related to reports on the favorable outcomes of recipients. Our results suggest that listening to concerns, providing correct information (particularly for some misconceptions), and emphasizing the outcomes of recipients will facilitate an increase in the number of people contemplating heart transplantation and organ donation.

Objective To assess the safety and efficacy of the endothelin receptor antagonist ambrisentan in pediatric pulmonary arterial hypertension (PAH). Study design In this open-label, phase IIb study, patients with PAH aged 8 to Results Most of the 41 patients randomized (80%) experienced ≥1 TEAE; most were mild (22%) or moderate (49%) in severity (no difference between dose groups). Most common TEAEs were headache (24%), nausea (17%), abdominal pain (12%), and nasopharyngitis (12%). Eight patients had serious TEAEs; 2 were fatal (unrelated to study treatment). Improved 6-minute walking distance was observed from baseline to week 24: total mean (SD) change, +40.69 (84.58) meters; World Health Organization functional class was maintained or improved in 70% and 27% patients, respectively. Conclusions Ambrisentan was well tolerated; TEAEs were consistent with the adult safety profile. Efficacy was similar to previous findings in adult PAH; however, interpretation is limited by small sample size. Findings support a potentially similar benefit:risk profile in pediatric (8 to Trial registration ClinicalTrials.gov : NCT01332331

We recently found a constitutively active mutant of natriuretic peptide receptor 2 (caNPR2; V883M), which synthesizes larger amounts of cyclic guanosine monophosphate (cGMP) intracellularly without any ligand stimulation than existing drugs. The aim of this study was to investigate the therapeutic effects of gene transduction using caNPR2 for pulmonary arterial hypertension (PAH). In vitro gene transduction into human pulmonary arterial smooth muscle cells using Sendai virus (SeV) vectors carrying caNPR2 induced 10,000-fold increases in the synthesis of cGMP without ligand stimulation, and the proliferation of caNPR2-expressing cells was significantly attenuated. The PAH model rats generated by hypoxia and the administration of SU5416 were then treated with SeV vectors through a direct injection into the left pulmonary artery. Right ventricular systolic pressure was significantly decreased 2 weeks after the treatment, while systemic blood pressure remained unchanged. Histological analyses revealed that the medial wall thickness and occlusion rate of pulmonary arterioles were significantly improved in caNPR2-treated lungs. Neither the systemic integration of virus vectors nor side effects were observed. The massive stimulation of cGMP synthesis by gene therapy with caNPR2 was safe and effective in a PAH rat model and, thus, has potential as a novel therapy for patients with severe progressive PAH.

Background Elucidating public concerns regarding vaccinations is important for successful immunization programs. The objective of the present study was to categorize public concerns regarding influenza vaccinations in Japan by analyzing a massive web-based question dataset. Methods The Yahoo! Chiebukuro (Japanese Yahoo! Answers) Dataset, which includes more than 16 million questions collected between April 2004 and April 2009, was used in this study. We sequentially filtered data to obtain questions on influenza vaccinations. Any questions that met our exclusion criteria concerning veterinary vaccines or computer virus vaccines were removed from the analysis. Filtered questions and their answers were manually analyzed for their content by a team of board-certified pediatricians. Results After filtering data, we obtained 1950 questions regarding influenza vaccinations. The three most frequently asked questions were regarding the vaccination schedule, safety, and effectiveness. When we analyzed monthly trends in question contents, we noted the emergence of similar questions in the same period every year. Therefore, we classified the time periods of each year into three parts: (1) from April to the commencement of seasonal influenza vaccinations (September), (2) from October until the epidemic period, and (3) the epidemic period. Two interesting results were obtained: concerns regarding effectiveness abruptly increased during the epidemic period, and pregnant or breastfeeding women increasingly asked questions regarding feasibility between October and the epidemic period. Conclusions The questions and concerns collected and analyzed in this study illustrate that the public have questions about the influenza vaccine and also that questions changed with periodical consistency. These results highlight the possible usefulness of providing the public with the latest and correct information to their questions in a timely manner, for example via an official health website.

Background To investigate the possible role of sex hormones in the pathogenesis of pulmonary arterial hypertension (PAH), the effect of β-estradiol (E2) on bone morphogenetic protein (BMP) signaling, a key signaling pathway involved in PAH, was studied in human pulmonary arterial endothelial cells (HPAEC). Methods and results BMP signaling molecules, including BMP receptor, Smad1/5/8 and Id1, were studied in HPAEC under 1% O2 (hypoxia) and 21% O2 (normoxia) as well as the effect of hypoxia-inducible factor (HIF)-1α expression in the presence of E2 on BMP signaling. The effects of an estrogen receptor (ER) antagonist (ICI 182,780) and cycloheximide, and the interaction of ER with Smad or HIF-1α were also studied. In the presence of E2, BMP signaling was augmented under normoxia but suppressed under hypoxia. HIF-1α accumulation suppressed BMP signaling, whereas HIF-1α inhibition augmented signaling. These effects were cancelled by ICI 182,780. Moreover, binding between ER, HIF-1α and phosphorylated (p)-Smad1/5/8 proteins occurred only under hypoxia. On inhibition of de novo synthesis with cycloheximide, however, p-Smad1/5/8 expression was suppressed only under normoxia. Conclusions The effects of E2 on BMP signaling in HPAEC altered depending on O2 concentration and different mechanisms may be involved. BMP and sex hormones may play an important role in PAH development.

Endothelin-1 (ET-1), a potent vasoconstrictor, is considered to be implicated in failing Fontan circulation, however the expressions of ET-1 and endothelin receptor type A (ET(A)R) and type B (ET(B)R) in the pulmonary arteries of failed Fontan patients were not elucidated.Immunohistochemistry and quantitative real-time PCR were used to analyse the expression levels of ET-1 and its receptors in the pulmonary arteries of the autopsy lung tissues of the patients who died after the Fontan procedure (n=10). We divided these patients into 3 groups, failed Fontan (n=4), heart failure (n=3) and non-failed Fontan (n=3), and then compared those to the age-matched normal controls (n=4).The intra-acinar pulmonary arteries of failed Fontan patients showed significant medial hypertrophy. Computational optical density analyses of the immunostaining revealed that the expressions of ET-1, ET(A)R, and ET(B)R in the intra-acinar pulmonary arteries were significantly increased in the failed Fontan patients (P0.05 vs. normal controls), however no significant difference was observed between the non-failed Fontan patients and the normal controls. Quantitative real-time PCR analyses confirmed that the mRNA expressions of ET-1, ET(A)R, and ET(B)R were significantly increased in the failed Fontan patients (P0.05 vs. normal controls).The overexpression of ET-1 and its receptors in the pulmonary arteries can cause pulmonary vasoconstriction and vascular remodelling, leading to failed Fontan circulation. This study suggests a histopathological rationale for the potential benefits of endothelin receptor antagonists in patients with failing Fontan circulation.

Purpose: Post-transplant lymphoproliferative disorder (PTLD) caused by Epstein-Barr virus (EBV) has recently has been recognized as a serious complication of various organ transplantations. In our institute, we have followed-up 34 pediatric patients after heart transplantation. We retrospectively investigated clinical features of PTLD in heart transplant recipients. Methods: The evaluation period was from July 2004 to June 2015. Six patients with PTLD (4 boys, 2 girls) were diagnosed by blood examination, lymph node histology and EBV viral load in peripheral blood and invasive organ. Results: Median age at onset of PTLD was 4.5 years old (range, 3.2-16.2 years). Median time from transplantation to onset was 18 months (range, 7-45 months). Bone marrow and central nervous system invasion were not detected in all cases. Four patients were EBV-seronegative recipients for EBV-seropositive donors. We performed the lymphoma-based chemotherapy combined with the anti-CD20 monoclonal antibody rituximab and achieved remission. As a result, five patients remained alive in remission, and one patient died of therapy-related toxicity. Although one patient relapsed after chemotherapy, she achieved second remission after multimodal chemotherapy combined with rituximab. Abdominal lymph node involvement was detected in four cases, and one patient experienced severe abdominal symptoms including intestinal perforation. Conclusion: EBV serostatus and the period from transplantation to onset of PTLD appear to represent important prognostic factors. Early detection followed by pre-emptive reduction of immunosuppressive agents is essential to the management of PTLD, but may increase the risk of fatal rejection. We administered chemotherapy combined with rituximab, and this treatment strategy might be effective. However, severe fatal toxicity developed in one patient, and severe abdominal symptoms due to tumor lysis were noted in many cases. These findings suggested the importance of carefully selecting the induction treatment and perform various supportive therapies. We should establish appropriate treatment strategies according to prognostic factors because of the distinctive characteristics of PTLD by accumulating more cases. Disclosures No relevant conflicts of interest to declare.