Identification of the cloned gene for the murine transplantation antigen H-2Kb by hybridization with synthetic oligonucleotides.

The H-2K(b) gene is a member of the large major histocompatibility complex class I gene family. Since many members of this family cross-hybridize with class I cDNA probes, the cloned H-2K(b) gene was identified by hybridization with specific oligonucleotide probes. This clone was definitively shown to encode the H-2K(b) polypeptide by partial DNA sequencing and by serological and tryptic peptide analyses of the expressed product.

Membrane elasticity of mouse dorsal root ganglion neurons decreases with aging.

Elastic area compressibility modulus (EACM) method was applied to measure the membrane elasticity of cultured dorsal root ganglion neurons from fetal and 3-month-old mice. The values of the EACM were 21.3 dyn/cm in 3-month-old neurons and 2.8 dyn/cm in fetal neurons. These results indicate that neural cell membrane elasticity decreases with aging.

Purification and characterization of choline oxidase from Arthrobacter globiformis.

Choline oxidase was purified from the cells of Arthrobacter globiformis by fractionations with acetone and ammonium sulfate, and column chromatographies on DEAE-cellulose and on Sephadex G-200. The purified enzyme preparation appeared homogeneous on disc gel electrophoresis. The enzyme was a flavoprotein having a molecular weight of approx. 83,000 (gel filtration) or approx. 71,000 (sodium dodecyl sulfate--polyacrylamide disc gel electrophoresis) and an isoelectric point (pI) around pH 4.5. Identification of the reaction products showed that the enzyme catalyzed the following reactions: choline + O2 leads to betaine aldehyde + H2O2, betaine aldehyde + O2 + H2O leads to betaine + H2O2. The enzyme was highly specific for choline and betaine aldehyde (relative reaction velocities: choline, 100%; betaine aldehyde, 46%; N,N-dimethylaminoethanol, 5.2%; triethanolamine, 2.6%; diethanolamine, 0.8%; monoethanolamine, N-methylaminoethanol, methanol, ethanol, propanol, formaldehyde, acetaldehyde, and propionaldehyde, 0%). Its Km values were 1.2 mM for choline and 8.7 mM for betaine aldehyde. The optimum pH for the enzymic reaction was around pH 7.5.

Oxidative pathway of choline to betaine in the soluble fraction prepared from Arthrobacter globiformis.

One strain of bacteria which showed high H2O2-generating activity was isolated from soil and characterized as Arthrobacter globiformis based on its morphological, nutritional, and physiological characteristics. The activities of H2O2 generation, NAD reduction and oxygen consumption in the bacterial cells were examined using choline, betaine aldehyde or betaine as substrate. Choline was oxidized to betaine aldehyde under aerobic conditions in a reaction coupled with H2O2 generation and oxygen consumption. On the other hand, betaine aldehyde seemed to be oxidized to betaine through two distinct oxidative reactions, H2O2 generation (oxygen consumption) under aerobic conditions and NAD reduction under either aerobic or anaerobic conditions. These enzyme activities were found in the supernatant fraction of the sonicated cell preparation.

Adaptation of cultured mammalian neurons to a hypotonic environment with age-related response.

The response of neurons to osmolal concentration changes has not been well-documented compared to erythrocytes, urinary bladder and epithelial cells. The effects of a hypotonic solution on morphology and electrophysiological functions in cultured dissociated neurons can be precisely studied. From the analysis of video pictures from Nomarski optics, diameters of the cells were seen to increase and then recover to the initial values after the application of a hypotonic solution. The rate of increase of cell size in fetal neurons was 4-5 times faster than in mature neurons. This age-related transient response was accompanied by a change of resting potential and membrane resistance. This transient depolarization and decrease of the resistance corresponded to morphological changes. However, the amplitude of an action potential scarcely changed during the cell membrane expansion. It is plausible that increased membrane tension caused by the membrane expansion might facilitate the opening of the channel. The cell membrane expansion might also increase ionic permeability through the membrane. This may reduce a swollen cell volume to the initial one by diluting their intracellular solute concentration. After this adaptation the cultured neurons were able to survive for a long time and extend processes in the hypotonic environment.

Expression of cytoskeletal-associated protein tyrosine phosphatase PTPH1 mRNA in human hepatocellular carcinoma.

We investigated the mRNA expression of cytosolic protein tyrosine phosphatase (PTPH1), which has a homologous domain to cytoskeletal-associated proteins, in human hepatocellular carcinomas (HCCs) by using reverse transcriptase-polymerase chain reaction (RT-PCR). PTPH1 mRNA was detected in all HCC cell lines (n = 6), and HCC and adjacent noncancerous tissues (n = 8) examined, indicating that PTPH1 was expressed in HCCs and hepatocytes. There was no remarkable difference in the level expression of PTPH1 mRNA between HCC and adjacent noncancerous tissues. We also performed RT-PCR single-strand conformation polymorphism (SSCP) analysis in HCC cell lines and tissues in the C-terminal region of the catalytic domain of PTPH1. In the cHc4 cell line and a HCC tissue specimen, a shifted band was detected, although it was not found in the non-cancerous tissue of the HCC specimen. Nucleotide sequence analysis showed a common mutation from T to C at the third letter of codon 919 which did not lead to amino acid substitution. These results suggest that another mutation leading to the development of HCC could occur in some region of PTPH1 other than that investigated in this study.

Hepatic infarction with portal thrombosis.

A case of hepatic infarction with portal thrombosis is reported. A 63-year-old woman with liver cirrhosis and esophageal varices was admitted for treatment of the esophageal varices. Endoscopic variceal ligation (EVL) and endoscopic injection sclerotherapy (EIS) were performed. Two months later, she experienced right hypochondralgia and right flank pain. Serum transaminase levels were suddenly elevated, and computed tomography scans of the liver showed multiple small nodular lesions. Her condition worsened, and she died of hepatic failure. Autopsy revealed splenic and portal vein thrombosis, multiple hepatic infarction, and evidence of chronic pancreatitis. We believe that liver cirrhosis and chronic pancreatitis were the main risk factors for the portal thrombosis, and the treatment for esophageal varices appeared to have triggered the thrombosis. The hepatic infarction was caused by the portal thrombosis.

Interactions of water-soluble porphyrins with hexadeoxyribonucleotides: resonance raman, UV-visible and 1H NMR studies.

The interactions of the water-soluble porphyrins M(TMpy-P4) [M = H2, Cu(II), Ni(II), and Co(III); TMpy-P4 = tetrakis(4-N-methylpyridyl)porphyrinato ion], with the hexadeoxyribonucleotides d(CGTACG)2, d(TACGTA)2, d(GCATGC)2, d(TGTGCA)2, and d(CTATAG)2 have been investigated by resonance Raman and/or UV-visible spectroscopy. The results indicate that all hexamers containing the 5'CG3' as well as the 5'GC3' site, and also the mismatched hexamer d(TGTGCA)2, are capable of intercalating the H2, Cu(II) and Ni(II) porphyrins. 1H nuclear magnetic resonance spectra of d(CGTACG)2 mixed with Cu(TMpy-P4) have provided further evidence for the intercalation. For the other cases, outside binding by localized electrostatic interaction is suggested. There is no evidence of groove binding to any of the hexamers. Possible reasons for different binding properties of long and short helices are discussed.

An autopsy case of intramedullary venous malformation of the spinal cord with spreading hematomyelia.

A 23-year-old Japanese woman experienced sudden severe shoulder pain, and subsequently died of cardiorespiratory arrest after an 8-day course of illness. Autopsy revealed a venous malformation in the spinal cord at the C6 level with fresh massive hemorrhage showing widespread rostrocaudal extension. The upper limit of the hemorrhage was in the left gracile nucleus of the medulla oblongata, and the lower limit was in the left posterior horn of the spinal cord at L4. The hemorrhage extended through the spinal cord, involving the left posterior horn and intermediate zone of the gray matter and the ventral part of the posterior funiculus. The mechanism of the extension of the hemorrhage and the pathogenesis of the spinal venous malformation are discussed.

Integrator for use in bombardment experiments.

A convenient integrator was developed for use in bombardment experiments with a charged particle accelerator such as a linac. In this device an irradiation dose is digitally displayed in the number of electrons per cm(2). Furthermore, a linac is controlled to deliver a preset dose by this integrator.

Prevalence of HBV and HCV infection in Japanese patients with hepatocellular carcinoma.

The prevalence of hepatitis B virus (HBV) and hepatitis C virus (HCV) infection was investigated in 63 Japanese patients with hepatocellular carcinoma (HCC). HBV infection was confirmed by measuring hepatitis B surface antigen and HBV-DNA in the serum, and HCV infection was confirmed by measuring antibody to HCV using a 2nd generation test and HCV-RNA in serum. Some 54.0% of the patients had HCV infection only, 27.0% had HBV infection only, and 9.5% had both HCV and HBV infection. Only 9.5% of HCC patients had neither HCV nor HBV markers. These results indicate that, in Japan, HCV and HBV infection is an important factor associated with HCC, and that the hepatitis virus may have a role in the carcinogenesis of HCC.

Synthesis and assignment of novel [125I]-labeled 1 alpha,25-dihydroxyvitamin D3 derivatives.

This paper describes the synthesis and structural assignment of noval 125I-1 alpha,25-dihydroxyvitamin D3 derivatives (1a) and (1b) labeled with 125I-Bolton-Hunter reagent [N-succinimidyl 3-(4-hydroxy-3-iodo[125I]phenyl)propionate] (1), which is known as a protein-labeling reagent, as tracers for radioimmunoassay (RIA). The radiospecific activities of these tracers (1a) and (1b) were calculated as 2,200 Ci/mmol (81.4 TBq/mmol).

Conformation and dynamics of Z-DNA oligomer duplex of d[(CG)3TATA(CG)3] in solution.

The conformation and dynamics of a DNA oligomer, d[(CG)3TATA(CG)3], in 4M NaClO4 (Z-TATA 16 mer) have been studied by 1H NMR. The principal results of our investigation are: (i) at low temperature d[(CG)3TATA(CG)3] exists as duplexes in both low (0.1M NaCl) and high (4M NaClO4) ionic strength solutions; (ii) CGCGCG segments undergo the B-to-Z transition in 4M NaClO4; (iii) even in 4M NaClO4 the TATA box exhibits non-Z-structures and possesses multiple conformations which are slowly exchanging on the NMR chemical shift difference time scale; and, (iv) the Z-type structure of the CGCGCG segments induced in 4M NaClO4 is more conformationally mobile than its B-type counterpart in 0.1M NaCl on the nanosecond time scale.

Solid phase synthesis of polynucleotides. VI. Further studies on polystyrene copolymers for the solid support.

A simple solid phase method for the synthesis of oligodeoxyribonucleotides has been developed using the phosphotriester approach. Mononucleotide coupling units are sequentially added to the polystyrene copolymer with 1% divinylbenzene and two kinds of oligonucleotides, d(CACGACCCCTCCACGT) and d(AACTGGTATTACTGGGCG), are synthesized in a relatively high yield. One cycle of the mononucleotide addition is about 70 minutes, and this method is particularly suitable for the automation of the synthesis upon availability of an automatic synthesizer.