Kei Inai

Outcomes of childhood pulmonary arterial hypertension in BMPR2 and ALK1 mutation carriers

Mutations in the bone morphogenetic protein receptor type 2 (BMPR2) gene and the activin receptor-like kinase 1 (ALK1) gene have been reported in heritable pulmonary arterial hypertension (HPAH) and idiopathic pulmonary arterial hypertension (IPAH). However, the relation between clinical characteristics and each gene mutation in IPAH and HPAH is still unclear, especially in childhood. The aim of this study was to determine, in a retrospective study, the influence and clinical outcomes of gene mutations in childhood IPAH and HPAH. Fifty-four patients with IPAH or HPAH whose onset of disease was at <16 years of age were included. Functional characteristics, hemodynamic parameters, and clinical outcomes were compared in BMPR2 and ALK1 mutation carriers and noncarriers. Overall 5-year survival for all patients was 76%. Eighteen BMPR2 mutation carriers and 7 ALK1 mutation carriers were detected in the 54 patients with childhood IPAH or HPAH. Five-year survival was lower in BMPR2 mutation carriers than mutation noncarriers (55% vs 90%, hazard ratio 12.54, p = 0.0003). ALK1 mutation carriers also had a tendency to have worse outcome than mutation noncarriers (5-year survival rate 64%, hazard ratio 5.14, p = 0.1205). In conclusion, patients with childhood IPAH or HPAH with BMPR2 mutation have the poorest clinical outcomes. ALK1 mutation carriers tended to have worse outcomes than mutation noncarriers. It is important to consider aggressive treatment for BMPR2 or ALK1 mutation carriers.

Mutations of NOTCH3 in childhood pulmonary arterial hypertension

Mutations of BMPR2 and other TGF‐β superfamily genes have been reported in pulmonary arterial hypertension (PAH). However, 60–90% of idiopathic PAH cases have no mutations in these genes. Recently, the expression of NOTCH3 was shown to be increased in the pulmonary artery smooth muscle cells of PAH patients. We sought to investigate NOTCH3 and its target genes in PAH patients and clarify the role of NOTCH3 signaling. We screened for mutations in NOTCH3, HES1, and HES5 in 41 PAH patients who had no mutations in BMPR2, ALK1, endoglin, SMAD1/4/8, BMPR1B, or Caveolin‐1. Two novel missense mutations (c.2519 G>A p.G840E, c.2698 A>C p.T900P) in NOTCH3 were identified in two PAH patients. We performed functional analysis using stable cell lines expressing either wild‐type or mutant NOTCH3. The protein‐folding chaperone GRP78/BiP was colocalized with wild‐type NOTCH3 in the endoplasmic reticulum, whereas the majority of GRP78/BiP was translocated into the nuclei of cells expressing mutant NOTCH3. Cell proliferation and viability were higher for cells expressing mutant NOTCH3 than for those expressing wild‐type NOTCH3. We identified novel NOTCH3 mutations in PAH patients and revealed that these mutations were involved in cell proliferation and viability. NOTCH3 mutants induced an impairment in NOTCH3‐HES5 signaling. The results may contribute to the elucidation of PAH pathogenesis.

Characterization of a novel KRAS mutation identified in Noonan syndrome

Noonan syndrome (NS) is the most common non‐chromosomal syndrome seen in children and is characterized by short stature, dysmorphic facial features, chest deformity, a wide range of congenital heart defects and developmental delay of variable degree. Mutations in the Ras/mitogen‐activated protein kinase (MAPK) signaling pathways cause about 70% of NS cases with a KRAS mutation present in about 2%. In a cohort of 65 clinically confirmed NS patients of Japanese origin, we screened for mutations in the RAS genes by direct sequencing. We found a novel mutation in KRAS with an amino acid substitution of asparagine to serine at codon 116 (N116S). We analyzed the biological activity of this mutant by ectopic expression of wild‐type or mutant KRAS. NS‐associated KRAS mutation resulted in Erk activation and active Ras–GTP levels, and exhibited mild cell proliferation. In addition, kras‐targeted morpholino knocked‐down zebrafish embryos caused heart and craniofacial malformations, while the expression of mutated kras resulted in maldevelopment of the heart. Our findings implicate that N116S change in KRAS is a hyperactive mutation which is a causative agent of NS through maldevelopment of the heart.

Relationships among red cell distribution width, anemia, and interleukin-6 in adult congenital heart disease.

BACKGROUND Red cell distribution width (RDW) is known to be associated with anemia and mortality in cardiovascular diseases, while anemia itself is related to increased mortality. RDW may also be related to cytokine activation. We investigated the potential of RDW to predict anemia-adjusted mortality in patients with adult congenital heart disease (ACHD) and we evaluated the relationships among RDW, anemia, and interleukin-6 (IL-6). METHODS AND RESULTS This was a single-center, retrospective cohort study. Blood RDW and IL-6 levels were measured in 144 patients with ACHD (median age [interquartile range (IQR)], 28 [22-36] years), 84% in New York Heart Association class I/II. During a mean 4.8-year follow-up, 21 (15%) patients died of cardiovascular causes. Elevated RDW (>15.0%) correlated significantly with mortality risk in a univariate analysis (RDW hazard ratio [HR]: 1.570; 95% confidence interval [CI]: 1.208-2.040 per 1 standard deviation increase; P=0.001). Elevated RDW levels correlated significantly with increased anemia-adjusted mortality (adjusted RDW HR: 1.912; 95% CI: 1.369-2.670; P<0.001). The high RDW group had significantly elevated serum IL-6 levels (RDW >15%, median [IQR], 3.7 [0.9-13.9] pg/ml vs. RDW ≤15%, 1.4 [0.8-2.5 pg/ml]; P=0.001), as did patients with anemia (anemia, 1.9 [0.9-5.2] pg/ml vs. no anemia, 1.4 [0.8-2.5 pg/ml]; P=0.021). CONCLUSIONS Elevation of RDW may be related with increased IL-6 and anemia-adjusted cardiovascular mortality in patients with ACHD.

Incidence, Clinical Course, and Risk Factors of Amiodarone-Induced Thyroid Dysfunction in Japanese Adults With Congenital Heart Disease

BACKGROUND Although amiodarone (AMD)-induced thyroid dysfunction (AITD) is an important complication of AMD therapy, little is known about AITD in adult Japanese patients with congenital heart disease (CHD). METHODS AND RESULTS We retrospectively studied 131 adult patients with CHD who were on low-dose AMD (median, 150 mg/day). The median patient age was 28 years, and the median follow-up was 44 months. The incidence, clinical course, and risk factors of AITD, including AMD-induced thyrotoxicosis (AIT) and AMD-induced hypothyroidism (AIH), were evaluated. The total incidence of AITD was 30% (AIT: 18%, n=24; AIH: 12%, n=16). Approximately 67% of patients with AIT displayed deterioration of tachyarrhythmia, and 38% patients underwent steroid therapy. Although thyroid function and symptoms associated with AIT improved within 6 months after diagnosis in most patients with AIT (92%), 1 patient died suddenly during an acute phase of AIT. No patient with AIH exhibited deterioration of tachyarrhythmia, and 9 patients underwent thyroid hormone replacement therapy. Cox multivariate analysis identified no independent risk factor for AIT, whereas liver dysfunction (hazard ratio 2.573; 95% confidence interval 1.102-5.795) was an independent risk factor for AIH. CONCLUSIONS AITD commonly occurred in adult Japanese patients with CHD even though they were on a low-dose AMD regimen. Risk factors for AITD may vary according to ethnicity and diet.

Monitoring of cerebral oxygenation during hypoxic gas management in congenital heart disease with increased pulmonary blood flow.

In the preoperative management of congenital heart disease (CHD) with increased pulmonary blood flow, hypoxic gas management to control pulmonary blood flow is useful. However, the cerebral oxygenation state has rarely been studied, and there is concern about neurologic development. In eight infants with CHD accompanied by increased pulmonary blood flow, hypoxia was induced after a 1-h baseline period in room air (FiO2, 0.21). The infants were simultaneously monitored in both the front-temporal region and the right-brachial region for 90 min using near-infrared spectroscopy (NIRS). The minimum SaO2 (pulse oximetry) after hypoxic gas administration was 80.8 ± 2.9% when the minimum FiO2 was 16.2 ± 1.1%. With a decrease in SaO2, oxy-Hb (O2Hb) decreased and total Hb [cHb: O2Hb + deoxy-Hb (HHb)] increased in both regions in the majority of infants. HHb increased in both regions with a decrease in SaO2. The maximum change in the tissue oxygenation index (TOI: O2Hb/cHb × 100) was –8.3 ± 2.6% in the front-temporal region and –3.6 ± 2.3% in the right-brachial region. Cerebral oxygenation decreased despite an increase in cerebral blood flow during hypoxic gas management. The change in TOI was ≤10% when the SaO 2 was ≥80%. Safer control of SaO2 should be maintained over 80% for hypoxia management in CHD based on the results of the present study.

Blood coagulation abnormalities and the usefulness of D-dimer level for detecting intracardiac thrombosis in adult Fontan patients

BACKGROUND Coagulation abnormality is associated with a high incidence of intracardiac thrombus (ICT) and systemic thromboembolism in Fontan patients. The biomarker for detecting ICT is currently unknown. METHODS We retrospectively investigated the underlying coagulation abnormality and useful biomarkers to screen for ICT in adult Fontan patients. We measured various biomarkers of blood coagulation, fibrinolysis, and platelet activity in 122 Fontan patients (Fontan group: median age [P25-P75]: 27 [20-34] years) and compared them to those in 50 patients with atrial septal defect (ASD group: 31 [24-40] years). RESULTS Regardless of whether the patient had ICT, the Fontan group showed significantly lower levels of antithrombin III, thrombomodulin, and α2-antiplasmin; lower protein C and protein S activities; and significantly higher levels of thrombin-antithrombin complex and α2-plasmin inhibitor complex than the ASD group. Among various biomarkers, D-dimer level measured by using latex immunoassay was significantly higher in the patients with ICT (thrombus group: n=21) than in the patients without ICT (non-thrombus group: n=101). Fifteen (26%) of 57 patients on warfarin achieved prothrombin time international normalized ratios (PT-INRs) of >2. The proportion of patients with PT-INRs of >2 tended to be lower in the thrombus group than in the non-thrombus group (13% vs 31%). Persistent atrial arrhythmia and D-dimer level were significant risk factors associated with ICT formation in the multivariate analysis (persistent atrial arrhythmia: hazard ratio [HR], 6.89; 95% confidence interval [CI], 1.44-34.5; D-dimer: HR, 0.29; 95% CI, 0.13-0.50). Receiver-operating characteristic curve analysis revealed that the appropriate cutoff D-dimer level for screening for ICT was 1.8μg/mL (area under the curve, 0.94), with a negative predictive value of 95%. CONCLUSIONS In the adult Fontan patients, blood coagulation abnormalities existed regardless of the absence of ICT. D-dimer level may be a useful biomarker for screening for ICT in adult Fontan patients.

Prognostic predictive value of gene mutations in Japanese patients with hypertrophic cardiomyopathy

Although some studies have attempted to find useful prognostic factors in hypertrophic cardiomyopathy (HCM), those results are not fully helpful for use in actual clinical practice. Furthermore, several genetic abnormalities associated with HCM have been identified. However, the genotype–phenotype correlation in HCM remains to be elucidated. Here, we attempted to assess patients with different types of gene mutations causing HCM and investigate the prognosis. A total of 140 patients with HCM underwent a screening test for myofilament gene mutations by direct sequencing of eight sarcomeric genes. Patients with a single mutation in cardiac troponin T, cardiac troponin I, α-tropomyosin, and regulatory and essential light chains were excluded from the study because the number of cases was too small. The clinical presentations and outcomes of the remaining 127 patients with HCM, 31 β-myosin heavy chain (MYH7) mutation carriers, 19 cardiac myosin-binding protein C (MYBPC3) mutation carriers, and 77 mutation non-carriers were analyzed retrospectively. MYBPC3 mutation carriers had a high frequency of ventricular arrhythmia and syncope. Kaplan–Meier curves revealed no significant difference in prognosis among the three groups, but a lack of family history of sudden death (SD) and a past history of syncope were significantly related to poor prognosis. An absence of family history of SD and past history of syncope are useful prognostic factors in patients with HCM. MYH7 and MYBPC3 mutations did not significantly influence prognosis compared to non-carriers. However, patients with the MYBPC3 mutation should be closely followed for the possibility of SD.

Vitamin D Kinetics and Parathyroid Gland Function in Patients with Congenital Heart Disease.

OBJECTIVE It has been recently reported that vitamin D deficiency may contribute to systemic illnesses that accompany chronic heart failure. These reports also suggest the serum levels of parathormone, which activates vitamin D in the liver, can be a useful marker of heart failure. This study was designed to evaluate the clinical implications of vitamin D and parathormone levels in patients with congenital heart diseases and chronic heart failure. DESIGN We measured 25-hydroxyvitamin D and parathormone serum levels in 103 adult patients with congenital heart diseases (age range 20-89 years). Of 103 patients, 54 were in New York Heart Association functional classes II or III. Their clinical data regarding cardiothoracic ratio, fractional shortening of the systemic ventricle, brain natriuretic peptide plasma levels, and pulse oximetry were also evaluated. RESULTS Of 54 patients with chronic heart failure, 50 (93%) exhibited vitamin D deficiency (25-hydroxyvitamin D serum levels <50 nmol/L) or elevation of parathormone (serum levels >65 pg/mL). These two parameters were inversely correlated. In multivariate analyses including age, gender, 25-hydroxyvitamin D, parathormone, pulse oximetry, cardiothoracic ratio, calcium, phosphorus, glomerular filtration rate, albumin, creatine kinase, end-diastolic diameter and fractional shortening of the systemic ventricle, and ratio of early diastolic transmitral flow velocity to mitral annular velosity, only the parathormone serum levels (P < .01) remained independently associated with brain natriuretic peptide plasma levels. Moreover, in multivariate analyses including the same variables minus parathormone serum levels, both pulse oximetry (P < .01) and glomerular filtration rate (P < .01) remained independently associated with parathormone levels. CONCLUSIONS Vitamin D deficiency and secondary hyperparathyroidism are common in patients with congenital heart diseases and heart failure. Serum parathormone and 25-hydroxyvitamin D levels correlated with several clinical heart failure markers, suggesting that vitamin D deficiency may deteriorate heart function in congenital heart disease patients.

The use of pethidine as pre-medication to prevent anoxic spells in patients with tetralogy of Fallot

Objective : The aim of this study was to determine whether morphine‐HCl or pethidine‐HCl differ as pre‐medication to prevent anoxic spell in patients with tetralogy of Fallot (TOF) at catheterization.