Establishment of the human hepatocellular carcinoma cell line and its characteristics

c-Hc-4 has been established and maintained for more than seven years. The hepatocellular carcinoma originated in 45-year old man with liver cirrhosis. The cell grew in vitro forming a sheet of monolayered cells and firmly attaching to the inner surface of cultured flasks. Morphologically they showed epithelial-like pattern. The doubling time was about 20 hours. Their modal chromosome number was 58. Serial heterologous transplantation in nude mice was successful. The histological finding was almost the same patterns as those in the primary tumor. The cultured cells produced alpha-fetoprotein (AFP) and carcinoembryonic antigen (CEA).     

Lipoxygenase activities of human platelets and their subcellular fractions: Comparison between lipoxygenase-deficient platelets and normal platelets

Lipoxygenase activities were estimated in washed platelets (intact platelets) and their subcellular fractions obtained from 7 patients with deficient platelet lipoxygenase activities and 9 normal subjects. From sonicated platelet preparations, 12,000 g supernatant (F-I), cytosol (F-II) and microsomal fractions (F-III) were prepared by differential centrifugation. When 12-hydroxyeicosatetraenoic acid (12-HETE) produced by the incubation of arachidonic acid with intact platelets or each of their subcellular fractions from normal subjects was measured by reversed-phase high-performance liquid chromatography analysis, the lipoxygenase activities of F-I, F-II and F-III were 87%, 31% and 17%, respectively, of the enzyme activity of intact platelets. One of the patients showed no detectable lipoxygenase activity in any preparation, while the other patients showed reduced enzyme activities in all preparations. The addition of CaCl2 significantly increased 12-HETE synthesis solely by F-I from these patients. In most of these patients, contrary to normal subjects, it appeared that the lipoxygenase activity was not fully expressed in intact platelets, since the F-I produced more 12-HETE than the intact platelets.     

Acidic fibroblast growth factor prevents death of hippocampal CA1 pyramidal cells following ischemia.

Ischemic insult induces neuronal death in the CA1 subfields of the hippocampus which are designated generally as the most vulnerable brain region. Recent studies have shown that acidic and basic fibroblast growth factors are potent trophic factors that support the survival of neurons in many brain regions including the hippocampus. Here we demonstrate that continuous infusion of acidic fibroblast growth factor into the lateral cerebral ventricles beginning 2 days before ischemia prevents the death of the CA1 pyramidal cells in the hippocampus of gerbils. Furthermore, delayed continuous administration of acidic fibroblast growth factor starting 5 min after ischemia is equally protective. The results suggest a possible physiological function for acidic fibroblast growth factor in the normal support of hippocampal CA1 pyramidal cells and neurons in some other brain regions in considering the broad spectrum of responsive neurons.     

Studies on the association of NIDDM in Japanese patients with hyperthyroid Graves' disease.

We attempted to analyze the association of hyperthyroid Graves' disease with non-insulin-dependent diabetes mellitus (NIDDM). Forty-nine patients (23 males and 26 females; 7.6%) of a total of 647 patients with hyperthyroid Graves' disease had NIDDM, several years before or after Graves' disease was diagnosed. Only 1 patient had insulin-dependent diabetes mellitus. Compared with the general Japanese population (n = 9,133), the incidence of NIDDM (n = 348; 3.9%) in patients with Graves' disease was higher in all age groups. Only 4 patients (8.2%) of the 49 hyperthyroid patients with NIDDM had a history of being overweight (body mass index > 25). In contrast, 276 (79.9%) of the 348 diabetic patients were currently or previously overweight. Moreover, the incidence of a family history of diabetes (13 of the 49 hyperthyroid Graves' patients with NIDDM; 26.5%) was also lower in the patients with NIDDM in the general Japanese population (50% incidence). The male:female ration in patients with Graves' disease and NIDDM was 1:1.1; much different from that in the total Graves' disease population (1:4.1). Analysis of the HLA loci A, B, C, DR and DQ (35 determinations) in 35 hyperthyroid patients with NIDDM and in 386 subjects from the general population revealed a highly significant difference between them in the incidence of HLA-Cw4, -DR2, -DQw1, -DQw3 and -DQw4. This study suggests that there was an association of Graves' disease with NIDDM. A significant association of HLA-DR and -DQ loci was observed in hyperthyroid Graves' patients with NIDDM.     

Complete nucleotide sequence of the mitochondrial DNA from a liverwort,Marchantia polymorpha

Libraries of cosmid and plasmid clones covering the entire region of mtDNA from the liverwortMarchantia polymorpha were constructed. These clones were used for the determination of the complete nucleotide sequence of the liverwort mtDNA totally 186,608 bp (GenBank no. M68929) and including genes for 3 species of ribosomal RNAs, 29 genes for 27 species of transfer RNAs, and 30 genes for functionally known proteins (16 ribosomal proteins, 3 subunits of cytochromec oxidase, apocytochromeb protein, 3 subunits of H⁺-ATPase, and 7 subunits of NADH ubiquinone oxidoreductase). The genome also contains 32 unidentified open reading frames. Thus the complete nucleotide sequences from both chloroplast and mitochondrial genomes have been determined in the same organism. Plasmid clones are available upon the request. Gene names are represented according to Lonsdale and Leaver (1988) with modifications recommended by Lonsdale (personal communication).     

Enclosure experiment on the control mechanism of planktonic bacterial standing stock

In order to understand the control mechanisms of a large, stable bacterial standing stock, enclosure experiments were conducted in a eutrophic lake, where both bacterial productivity and grazing pressure were very high. Total bacterial number in the different enclosures ranged from 1.2 to 2.7×10⁷ cells mL⁻¹ throughout the experiment. The average bacterial cell production rate estimated from a grazer eliminating experiment was 6.3×10⁵ cells mL⁻¹ h⁻¹. Difference in the bacterial cell production rate between shaded and unshaded enclosures was not apparent. Bacteria showed a reduction in standing stock of only about 25–30% even after the supply of light was cut to 1%. Bacteria in the shaded enclosures then recovered their production rate in the first 12 days of perturbation. Grazing pressure in the shaded enclosures was not less than that for the control. Thus, it was considered a control mechanism of bacterial stable standing stock that the bacteria shifted their organic substrate from extracellular dissolved organic carbon freshly released from phytoplankton to that already stocked in the water column, though it is not known whether the dominant bacteria were the same.     

Mutations in a Saccharomyces cerevisme host showing increased holding stability of the heterologous plasmid pSRI

Summary We have isolated Saccharomyces cerevisiae mutants, smp, showing stable maintenance of plasmid pSRI, a Zygosaccharomyces rouxii plasmid. The smp mutants were recessive and were classified into at least three different complementation groups. The three mutants also showed increased stability of YRp plasmids and the mutations are additive for plasmid stability. One mutation, smp1, confers a respiration-deficient (rho ⁰) phenotype and several Rho^– mutants independently isolated by ethidium bromide treatment of the same yeast strain also showed increased stabilities of pSR1 and YRp plasmids. The wild-type S. cerevisiae cells showed a strongly biased distribution of pSR1 molecules as well as YRp plasmids to the mother cells at mitosis, while the smpf mutant did not show this bias. Another mutation, smp3, at a locus linked to ade2 on chromosome XV, confers temperature-sensitive growth. The SMP3 gene encodes a 59.9 kDa hydrophobic protein and disruption of the gene is lethal.     

Determination of the chromosomal site for the human radiosensitive ataxia telangiectasia gene by chromosome transfer

The chromosomal localization of the gene which complements radiation hypersensitivity of AT cells was studied by microcell-mediated chromosome transfer. A 6-thioguanine-resistant derivative of an immortalized AT cell line, AT2KYSVTG, was used as a recipient for microcell-mediated chromosome transfer from 4 strains of mouse A9 cells, 3 of which carried a human X/11 recombinant chromosome containing various regions of chromosome 11, while the other carried an intact X chromosome. HAT-resistant microcell hybrids were isolated and examined for their radiosensitivity and chromosome constitution. The microcell hybrid clones obtained from the transfer of an intact X chromosome or an X/11 chromosome bearing the pter → q13 region of chromosome 11 did not show a difference in radiosensitivity from parental AT cells, while those obtained from the transfer of X/11 chromosomes bearing either the p11 → qter or the pter → q23 region of chromosome 11 exhibited a marked radioresistance which was comparable to normal human fibroblasts. A HAT-resistant but radiosensitive variant was further obtained from the microcell fusion with an A9 cell strain carrying an X/11 chromosome bearing the 11p11 → qter region, in which a deletion at the 11q23 region was found. The results indicate that the gene which complements a radiosensitive phenotype of AT is located at the q23 region of chromosome 11.