Functional analysis of titin/connectin N2-B mutations found in cardiomyopathy

Hypertrophic cardiomyopathy and dilated cardiomyopathy are two major clinical phenotypes of “idiopathic” cardiomyopathy. Recent molecular genetic analyses have now revealed that “idiopathic” cardiomyopathy is caused by mutations in genes for sarcomere components. We have recently reported several mutations in titin/connectin gene found in patients with hypertrophic cardiomyopathy or dilated cardiomyopathy. A hypertrophic cardiomyopathy-associated titin/connectin mutation (Arg740Leu) was found to increase the binding to actinin, while other dilated cardiomyopathy-associated titin/connectin mutations (Ala743Val and Val54Met) decreased the binding to actinin and Tcap/telethonin, respectively. We also reported several other mutations in the N2-B region of titin/connectin found in hypertrophic cardiomyopathy and dilated cardiomyopathy. Since the N2-B region expresses only in the heart, it was speculated that functional alterations due to the mutations cause cardiomyopathies. In this study, we investigated the functional changes caused by the N2-B region mutations by using yeast-two-hybrid assays. It was revealed that a hypertrophic cardiomyopathy-associated mutation (Ser3799Tyr) increased the binding to FHL2 protein, whereas a dilated cardiomyopathy-associated mutation (Gln4053ter) decreased the binding. In addition, another TTN mutation (Arg25618Gln) at the is2 region was found in familial DCM. Because FHL2 protein is known to tether metabolic enzymes to N2-B and is2 regions of titin/connectin, these observations suggest that altered recruitment of metabolic enzymes to the sarcomere may play a role in the pathogenesis of cardiomyopathies.     

Intraspinal Metastasis of Neuroblastoma -- Report of a Case Detected at Autopsy --

Metastasis to the brain or spinal cord parenchyma is extremelyrare in cases of neuroblastoma. We present a 3-year-7-month-old boy with neuroblastoma, stageIV, with intraspinal metastasis. He had no neurologic manifestationexcept incontinentia urinae and ataxia at the terminal stage.His cranial computed tomography scan showed high density areasin both cerebellar hemispheres which seemed to be distant metastaticmasses. At autopsy, the metastases in the cerebellum were foundto be due to dural infiltration but in the spinal cord parenchymaof the lumbar spine metastases were detected macroscopically.There were multiple distant metastatic areas near the roots,anterior and posterior. The neuroblastoma seemed to have spreadalong the roots by direct invasion from the meninges. In the future, the number of patients with metastasis into thebrain or spinal cord parenchyma will increase because patientswith progressive disease could live for a long time as a resultof intensive chemotherapy. Observation of these cases will behelpful to clarify the routes of metastasis to these areas.     

Thoracoscopic Radiofrequency Ablation of the Myocardium

Radiofrequency (RF) catheter ablation has been used for the treatment of ventricular tachycardia (VT), however, in some patients VT might result from subepicardiai macroreentry that could be successfully terminated by epicardial approach. This study examined the feasibility of thoracoscopic RF ablation of myocardium from epicardium using a custom made electrode. In five mongrel dogs, the thoracoscope was introduced through the 7th intercostal space. A 500-kHz continuous wave RF energy was connected to a custom made multiple electrode probe. Under thoracoscopic guidance, the heart was exposed and the RF probe was introduced. RF ablation was performed on the nonvascular ventricular wall of the beating heart. The left ventricular free wall and right ventricular outflow tract were satisfactorily visualized and ablated. The total dose of RF energy ranged from 50 to 500 J. and the estimated volume of ablated lesions ranged from 41.0–799 mm³. There were significant correlations between the RF discharge output and the irradiated lesion volume (P < 0.01), and the depth of the lesions (P < 0.01). Grossly, after RF ablation the ventricular myocardium demonstrated a circular, well-demarcated area of thermal injury. Volume and depth of the lesion depended upon the total dose of delivered RF energy. Thoracoscopic RF ablation appears to be a minimally invasive and useful method for creating irradiated myocardial lesions from epicardial surface. This method could he technically feasible for the treatment of Vts for which endocardial RF ablation is ineffective.     

Retrospective study on stage B prostate cancer in the Hokuriku District, Japan

BACKGROUND: The present study was conducted to investigate how patients with clinically localized prostate cancer were treated in the Hokuriku District, Japan. METHODS: Medical records of 536 patients with stage B prostate cancer were retrospectively reviewed. The patients were diagnosed and treated at four university hospitals and 32 collaborating hospitals in the Hokuriku District. RESULTS: Because their medical records were incomplete and/or they not available for follow up, 79 cases were excluded from this study. Conservative treatment with hormone therapy was used for 248 cases. Radical prostatectomy was performed in 199 cases, only 27 of whom underwent surgical monotherapy. There was no significant difference in disease-specific survival rates between the hormone (69.0%) and surgery group (83.2%) after 110 months. Results of the analysis of disease-specific survival rates according to histologic grade showed that patients with poorly differentiated cancers treated with hormone therapy were the only subset with significant differences when compared against the other patients. CONCLUSION: The value of prostatectomy alone or added was marginal in terms of survival. Only patients with poorly differentiated cancer might benefit from prostatectomy.     

Alcohol effect on sleep electroencephalography by fast Fourier transformation

Abstract The influence of alcohol (ethanol) on sleep was investigated in 10 men. Polysomnography (PS) was recorded on a baseline night (BL-N) and an ethanol (0.8 g/kg) night (Et-N). On visual score rapid eye movement (REM) sleep was reduced, REM latency was prolonged on Et-N as compared to BL-N. Using the fast Fourier transformation method, electroencephalographic power density of REM sleep in δ frequencies band and in the 10–12 Hz range of non-REM sleep were enhanced. REM sleep and non-RJEM sleep changes were prominent in the second-half and first-half of the night, respectively.     

Interaction of lipophilic peptides derived from mastoparan with phospholipid vesicles

Various kinds of lipophilic peptides were prepared by acylation of an ct-helical peptide, mastoparan, to investigate the effects of acyl groups on the interaction of peptides with phospholipid membranes. α-Helicity of the peptides was increased by introduction of long acyl groups. Acyl peptides showed different membrane-perturbation activities for neutral and acidic phospholipid vesicles, whereas a peptide with a dialkycarbamoyl group always exhibited a strong activity. High hemolytic activities were observed for the peptides with long acyls (single or double chain). These results indicate that lipophilic groups introduced to mastoparan contribute greatly to the interaction of the peptide with phospholipid membranes with lengthening of the acyl chain and that the structural character of the lipophilic group also influences the conformation of the peptide.     

STIMULATION OF CYCLIC AMP PRODUCTION IN FRTL-5 THYROID CELLS BY CRUDE IMMUNOGLOBULIN FRACTIONS OF SERUM FROM PREGNANT WOMEN

In an assay for thyroid-stimulating antibodies, in which FRTL-5 thyroid cells were incubated with crude immunoglobulin (Ig) fractions precipitated from serum with 15% polyethylene glycol, significant increase in cAMP production was elicited by the samples from 25 (35.7%) out of 70 pregnant women. The highest value was 529.5%. There was a close correlation between thyroid stimulating activities and serum hCG concentrations (r = 0.708, P less than 0.001). When 125I-hCG was added to serum from pregnant women, about 20% of the radioactivity was incorporated into the Ig fractions. hCG preparations within a range of concentrations of 30-300 IU/m elicited 2.3-16.5 times increase in cAMP in a dose-dependent manner. Nine pregnant women with serum TSH concentrations less than the lower limit of the normal range (less than 0.25 mU/l) displayed significantly higher values for both thyroid stimulating activities and serum hCG concentrations (P less than 0.001, respectively) compared with those who had normal TSH levels in serum. These data suggest that hCG or its variant may stimulate the thyroid sufficiently to suppress secretion of TSH from the pituitary in some pregnant women.     

Interaction of α-helical peptides with phospholipid membrane: effects of chain length and hydrophobicity of peptides

To investigate the interaction of amphiphilic α-helical peptides with phospholipid membranes, we synthesized Ac-(Leu-Ala-Arg-Leu)₃-NHcH₃ (4₃) and three derivatives, in which the chain length and the size of the hydrophobic region of the peptides were different from each other. These peptides formed an α-helical structure in the presence of vesicles. In the membrane-perturbation measurement, only 4₃ showed a strong membrane-perturbation activity below phase-transition temperature (25°C), but above phase-transition temperature (50°C), most peptides showed similar strong activities. On the other hand, in membrane-fusion measurement the long peptides, e.g., Ac-(Leu-Ala-Arg-Leu)₃-(Leu-Arg-Ala-Leu)₃-NHCH₃, had strong activities at low peptide concentrations at 25°C. The present study indicated a parallel relationship did not always exist between membrane fusion and perturbation caused by peptides.     

Marked increase of peripheral blood myeloblasts following G-CSF therapy in a patient with acute lymphoblastic leukemia

A 9 year old boy with acute lymphoblastic leukemia (ALL) received recombinant human granulocyte colony-stimulating factor (rhG-CSF) and showed a marked increment of myeloblasts in the peripheral blood. He was administered repeated courses of intermediate-dose cytosine arabinoside (Ara-C) therapy (1500 mg/m², days 1–5) for frequent central nervous system (CNS) relapse of ALL. The peripheral white blood cell nadir was less than 1000/μL, so he was treated with rhG-CSF. A marked increment of peripheral blood blasts was noted 3–5 days after rhG-CSF treatment. These cells decreased with the appearance of mature myeloid cells and disappeared about 2 weeks after the start of treatment. These findings suggested that the blasts might have the ability to differentiate into mature myeloid cells. A control patient with repeated CNS relapse of ALL showed no increment of peripheral blood blasts after similar repeated courses of Ara-C without rhG-CSF treatment. Cultured peripheral blood blasts obtained from the present patient showed differentiation into mature myeloid cells by morphological studies and surface marker analysis. These findings indicate that the peripheral blood blasts drawn by G-CSF were not leukemic blasts but normal myeloblasts.