Functional linkage of the cardiac ATP-sensitive K+ channel to the actin cytoskeleton

The role of the cytoskeleton in the rundown and reactivation of adenosine triphosphate (ATP) sensitive K⁺ channels (KATP channels) was examined by perturbing selectively the intracellular surface of inside-out membrane patches excised from guinea-pig ventricular myocytes. Actin filament-depolymerizing agents (cytochalasins and desoxyribonuclease I) accelerated channel rundown, while actin filament stabilizer (phalloidin) or phosphatidylinositol biphosphate (PIP₂; inhibitor of F-actin-severing proteins) inhibited spontaneous and/or Ca²⁺-induced rundown. When rundown was induced by cytochalasin D or by long exposure to high Ca²⁺, channel activity could not be restored by exposure to MgATP, but application of F-actin with MgATP could reinstitute channel activity. The processes of rundown and reactivation of cardiac K_ATP channels may thus be influenced by the assembly and disassembly of the actin cytoskeletal network, which provides a novel regulatory mechanism of this channel.     

Small round and spindle cell sarcoma with neuronal differentiation and oncocyte-like features of the thoracic wall: A case report with histological,immunohistochemical and ultrastructural examinations

A case of small round and spindle cell sarcoma with neuronal differentiation and oncocyte-like features is presented. The tumor was encountered in a 32 year old Japanese woman with an initial presentation of palpable tumor in the left lateral region of the thorax. The resected tumor was a partially well encapsulated whitish medullary one and consisted of small round and spindle tumor cells, together with so-called rhabdoid cells in the small round cell area. Although pseudorosettes were often observed, true rosette formation could not be detected anywhere. Ultrastructurally, despite a histologic variety of tumor cells, most tumor cells possessed numerous mitochondria, some of which occasionally contained abnormal filamentous or crystalloid structures. Various amounts of microfilaments were present in most tumor cells and microtubules were present in a few. A minority of small round cells possessed a small number of neurosecretory granules, especially in short cytoplasmic processes. A positive immunoreaction for neuron specific enolase was found by immunohistochemical examination in several small round tumor cells and for neurofilaments in lesser numbers. Despite the lack of S-100 protein, MB2 was detected in both small round and spindle cells. On the basis of these findings, the tumor of the present case corresponds to malignant peripheral nerve sheath tumor with neuronal differentiation and oncocytic features.     

Reduced-intensity unrelated cord blood transplantation for patients with advanced malignant lymphoma.

We report the results of reduced-intensity unrelated cord blood transplantation (RI-UCBT) in patients with advanced malignant lymphoma. Twenty patients (median age, 46.5 years; range, 27-66 years) underwent RI-UCBT with a preparative regimen consisting of fludarabine 125 mg/m2 , melphalan 80 mg/m 2 , and 4 Gy of total body irradiation. The median infused total cell dose was 2.75 x 10(7)/kg (range, 2.3-3.4 x 10(7)/kg). Graft-versus-host disease (GVHD) prophylaxis was composed of cyclosporine or tacrolimus alone. Fifteen patients achieved primary neutrophil engraftment after a median of 20 days. Eight patients developed grade II to IV acute GVHD, and 2 developed chronic GVHD. Of the 16 patients with evaluable disease, 10 achieved a complete response. Primary disease recurred in 1 patient, and transplant-related mortality within 100 days occurred in 8 of 20 patients. The estimated 1-year probability of progression-free survival was 50%. These data suggest that RI-UCBT is a feasible option for patients with refractory lymphoma who lack an HLA-matched donor.     

Mostly separate distributions of CLAC- versus Abeta40- or thioflavin S-reactivities in senile plaques reveal two distinct subpopulations of beta-amyloid deposits

Collagenous Alzheimer amyloid plaque component (CLAC) is a unique non-Abeta amyloid component of senile plaques (SP) derived from a transmembrane collagen termed CLAC-precursor. Here we characterize the chronological and spatial relationship of CLAC with other features of SP amyloid in the brains of patients with Alzheimer's disease (AD), Down syndrome (DS), and of PSAPP transgenic mice. In AD and DS cerebral cortex, CLAC invariably colocalized with Abeta42 but often lacked Abeta40- or thioflavin S (thioS)-reactivities. Immunoelectron microscopy of CLAC-positive SP showed labeling of fibrils that are more loosely dispersed compared to typical amyloid fibrils in CLAC-negative SP. In DS cerebral cortex, diffuse plaques in young patients were negative for CLAC, whereas a subset of SP became CLAC-positive in patients aged 35 to 50 years, before the appearance of Abeta40. In DS cases over 50 years of age, Abeta40-positive SP dramatically increased, whereas CLAC burden remained at a constant level. In PSAPP transgenic mice, CLAC was positive in the diffuse Abeta deposits surrounding huge-cored plaques. Thus, CLAC and Abeta40 or thioS exhibit mostly separate distribution patterns in SP, suggesting that CLAC is a relatively early component of SP in human brains that may have inhibitory effects against the maturation of SP into beta-sheet-rich amyloid deposits.     

Establishment and Characterization of Cell Lines from Human Adenovirus Type 12-induced Murine Tumors Producing Endogenous Virus Particles

Two cell lines designated IC KMS and D KMS were established from human adenovirus type 12 induced tumors of C3Hf/OK mouse. The cell lines retained the characteristics of the original tumor i.e., production of numerous C type and intracisternal A-type particles, integration of Adl2 El region DNA and amplification of the myc gene family. Chromosomal analysis revealed chromosome aberrations in both IC KMS and D KMS cells. The modal chromosome number of IC KMS cells was 54 and that of D-KMS cells was 48. Metacentric chromosomes and mini-chromosomes were found. Trisomy of chromosome 3, 7 and 12 was seen frequently in D KMS cells. Although DNA aneuploidy was revealed by flow cytometry, the DNA indices of these cells showed no relation to the copy number of integrated Adl2 DNA. These cells have been propagated by serial culture during the past 17 months. Production of endogenous virus particles is a unique characteristic of IC KMS and D KMS cells. These cell lines would be useful materials for examining the contribution of Adl2 carcinogenesis to activation of endogenous virus particles, and also the correlation between Adl2 carcinogenesis and cancer related genes. Acta Pathol Jpn 42: 242-248, 1992.     

Human ORFeome Version 1.1: A Platform for Reverse Proteomics

The advent of systems biology necessitates the cloning of nearly entire sets of protein-encoding open reading frames (ORFs), or ORFeomes, to allow functional studies of the corresponding proteomes. Here, we describe the generation of a first version of the human ORFeome using a newly improved Gateway recombinational cloning approach. Using the Mammalian Gene Collection (MGC) resource as a starting point, we report the successful cloning of 8076 human ORFs, representing at least 7263 human genes, as mini-pools of PCR-amplified products. These were assembled into the human ORFeome version 1.1 (hORFeome v1.1) collection. After assessing the overall quality of this version, we describe the use of hORFeome v1.1 for heterologous protein expression in two different expression systems at proteome scale. The hORFeome v1.1 represents a central resource for the cloning of large sets of human ORFs in various settings for functional proteomics of many types, and will serve as the foundation for subsequent improved versions of the human ORFeome.     

Identification of a large novel imprinted gene cluster on mouse proximal chromosome 6

Mice with maternal duplication of proximal chromosome 6 die in utero at an early embryonic stage. Recently, two imprinted genes, paternally expressed Sgce and maternally expressed Asb4, were identified in this region. This report analyzes the imprinting status of genes within a 1-Mb region containing these two genes. Peg10, which is next to Sgce, shows complete paternal expression, like Sgce. Conversely, Neurabin, Pon2, and Pon3 show preferential maternal expression at embryonic stages, although they all show biallelic expression in neonatal tissues. These results demonstrate that there is a large novel imprinted gene cluster in this region. 5'-RACE (Rapid Amplification of cDNA Ends) analysis of Peg10 revealed the existence of a novel first exon separate from the second exon, which encoded two putative ORFs similar to the viral Gag and Pol proteins. A differentially methylated region established in sperm and eggs is located just within the region containing the two first exons of Peg10 and Sgce, and may play an important role in regulating the two paternally expressed genes: Peg10 and Sgce.     

Nondestructive evaluation of blood flow in a dialyzer using X-ray computed tomography

It is very important to observe the concentrations and flow patterns of blood through a dialyzer to evaluate its function and to obtain the most appropriate design. We established a visualization method for the blood flow pattern in a dialyzer using X-ray computed tomography, and investigated the so-called internal filtration phenomenon. The results obtained were as follows: (1) The influence of 5% BaSO₄, which was added to the blood as a contrast medium, on the filtration rate of the dialyzer was minimal. (2) The relationship between the concentration of BaSO₄ and the Hounsfield unit value was expressed by linear regression. (3) Hounsfield unit values increased massively just after blood entered the dialyzer and peak values increased with dialysate perfusion under the following conditions: the dialyzer (BS-1.6UL, polysulfone hollow fibers) was used, and bovine blood with 5% BaSO₄ added was used at a blood flow rate of 200ml/min. The dialysate flow rate was 500ml/min and the slice thickness of X-ray computed tomography was 1–10mm. (4) It was observed that blood flowed slightly faster in the center than the peripheral portion of the dialyzer, when the flow pattern was followed after pulse injection of blood containing 20% BaSO₄ into the dialyzer. It was concluded that this method could possibly be utilized not only qualitatively but also quantitatively for observation of the real state of blood flow and in designing dialyzers.     

The ultrasound examination of the deep vein thrombosis

Ultrasonography is very useful for detection of deep vein thrombosis. The purpose of this paper is to show a method for detecting them efficiently by high resolution transducer and color Doppler system. We examined patients in the supine and prone positions. To detect the venous flow easily and differentiate thrombi from simple venous dilatation, some maneuvers are useful; one is pushing the vein area using the transducer on examination, the second is breathing overload, and the last is so-called milking. We can find throombi in the external iliac or femoral veins of patients who have symptoms of lower leg swelling, however, we need to better detect venous thrombi in the lower leg in patients with a history of pulmonary embolism. Because deep venous thrombi are increasing, the role of ultrasound will expand in the future.