Solution Radioactivated by Hadron Radiation Can Increase Sister Chromatid Exchanges

When energetic particles irradiate matter, it becomes activated by nuclear reactions. Radioactivation induced cellular effects are not clearly understood, but it could be a part of bystander effects. This investigation is aimed at understanding the biological effects from radioactivation in solution induced by hadron radiation. Water or phosphate buffered saline was activated by being exposed to hadron radiation including protons, carbon- and iron-ions. 1 mL of radioactivated solution was transferred to flasks with Chinese hamster ovary (CHO) cells cultured in 5 mL of complete media. The induction of sister chromatid exchanges (SCE) was used to observe any increase in DNA damage responses. The energy spectrum and the half-lives of the radioactivation were analyzed by NaI scintillation detector in order to identify generated radionuclides. In the radioactivated solution, 511 keV gamma-rays were observed, and their half-lives were approximately 2 min, 10 min, and 20 min. They respectively correspond to the beta+ decay of ¹⁵O, ¹³N, and ¹¹C. The SCE frequencies in CHO cells increased depending on the amount of radioactivation in the solution. These were suppressed with a 2-hour delayed solution transfer or pretreatment with dimethyl sulfoxide (DMSO). Our results suggest that the SCE induction by radioactivated solution was mediated by free radicals produced by the annihilated gamma-rays. Since the SCE induction and DMSO modulation are also reported in radiation-induced bystander effects, our results imply that radioactivation of the solution may have some contribution to the bystander effects from hadron radiation. Further investigations are required to assess if radioactivation effects would attribute an additional level of cancer risk of the hadron radiation therapy itself.     

Purification and characterization of a third glucosyltransferase from Streptococcus mutans serotype g

Streptococcus mutans strain AHT (serotype g) secretes at least two glucosyltransferases with different pI values. A novel glucosyltransferase with a pI of 5.8 was purified 244-fold from the ammonium sulphate fraction by DEAE-cellulose chromatography, FPLC (Mono Q column, Pharmacia) and hydrophobic chromatography. The enzyme preparation gave a single protein band on analysis by both PAGE and SDS-PAGE, and did not form multiple protein bands detectable by IEF. The Mr was estimated to be about 130,000 by SDS-PAGE and about 135,000 by ultracentrifugal analysis. The apparent Km value and pH optimum of the enzyme were 3.9 +/- 0.2 mM (mean +/- SD) and about 4.7, respectively. The enzyme synthesized water-soluble glucan from sucrose, and the glucan consisted of over 90 mol% 1,6-alpha-D-glucosidic linkages. The enzyme activity was not stimulated by primer dextran. Anti-enzyme serum produced a single precipitin band with the purified enzyme preparation, whereas it did not react with either of the other two known glucosyltransferases.     

Effects of proteinase inhibitors on preimplantation embryos in the rat

Proteinase inhibitors of microbial origin were injected into the uterine horns of mated rats at 14:00 h on Day 5 of pregnancy (spermatozoa in vaginal smear = Day 1), and 5 or 6 h later the embryos were flushed from the horns and examined. Chymostatin and alpha-MAPI, inhibitors of chymotrypsin-like serine proteinase and thiol proteinases, as well as thiolstatin, an inhibitor of thiol proteinases, significantly inhibited embryo growth. The inhibitory activity of alpha-MAPI on embryonic growth was distinctly greater than that of thiolstatin, although the ID50 values of the two inhibitors to papain are similar. Antipain and leupeptin which are inhibitors of trypsin-like and thiol proteinases, and talopeptin, an inhibitor of metal proteinases, significantly interrupted the removal of the zona pellucida from expanding blastocysts. These results suggest that (1) a chymotrypsin-like proteinase seems to be important to the growth of the embryo, (2) a thiol proteinase may participate in embryonic growth, and (3) a trypsin-like proteinase and a metal proteinase are likely to participate in zonalysis.     

A serum factor stimulating cathepsin D-release from erythrocytes or ghosts

A serum factor preparation extensively purified from bovine serum stimulated cathepsin D-release from the rat blood cells in a concentration-dependent fashion within a range of physiological concentrations of the factor. Among the blood cells only the erythrocytes (or ghosts) were responsive to the factor, and the leucocytes and lymphocytes were unresponsive. The effects of Ca2+-concentrations, SH- blocking reagents, protease-inhibitors, calmodulin-inhibitors, calmodulin or EGTA-pretreatment of the ghosts on cathepsin D-release from the erythrocytes of ghosts in the presence or the absence of serum factor were investigated. The results suggested that the serum factor may first activate the Ca2+-calmodulin system via the mobilization of "Ca2+-pool" and then the calmodulin-dependent SH-protease in the erythrocyte plasma membranes. The activated protease in turn may break the linkage between cathepsin D and the plasma membranes, liberating cathepsin D activity into the incubation medium. The name of "calciferin" was proposed for the serum factor.     

Properties of the strongly immobilized signal observed in spin-labeled erythrocytes

A strongly immobilized signal from fatty acid spin labels was observed in human erythrocytes treated with oxidizing agents such as glutaraldehyde, hydrogen peroxide, phenylhydrazine and copper-ortho-phenanthroline. This signal was also observed in freshly prepared ghosts treated with potassium superoxide and in old erythrocyte ghosts. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of these samples demonstrated the diffuse, nondiscrete bands of high molecular weight due to the cross-linking of membrane proteins. The temperature and pH dependences of the outer hyperfine splitting of this signal were very similar to those of bovine serum albumin. We propose that the strongly immobilized signal reflects the interaction of the lipids with the cross-linked products of membrane proteins.     

Dose-response relations for dicentric yields in G0 lymphocytes on man and crab-eating monkey following acute and chronic γ-irradiations

A comparison has been made of dicentric yields in G₀ lymphocytes between man and crab-eating monkey, Macaca fascicularis, after acute and chronic γ-irradiations. With acute irradiation (49.6 rad/min) there was no significant difference between them, but for the chronic irradiation (17.1 rad/h) a significant difference was observed between the species. When the dose-response relations were fitted to the linear-quadratic model (Y = αD + βD²), the species-difference observed for chronic irradiation was almost entirely due to change in the value of β. In addition, after chronic irradiation the β-value for monkey was almost negligible, but that for man was significant. Post-irradiation incubation experiment showed that cells with dicentrics were partly eliminated during the course of chronic irradiation, because there were appreciable reductions of dicentric yields (ca. 25% for both man and monkey at 400 rad) together with mitotic indices (ca. 30% and 60% for man and monkey, respectively, at 400 rad). Accordingly, it would be reasonable to postulate that G₀ repair for dicentrics other than selection mechanism must play a major role in the effects of low dose rate. It can be further suggested that G₀-repair capacity for chromosal damages leading to dicentrics may be different among different primate species.     

Continuous butanol/isopropanol fermentation in down-flow column reactor coupled with pervaporation using supported liquid membrane

Continuous butanol/isopropanol fermentation with immobilized Clostridium isopropylicum was performed in a downflow column reactor using molasses as the substrate. In order to prevent product inhibition and at the same time obtain high concentration of the products, the column reactor was coupled with a pervaporation module using a supported liquid membrane. The liquid membrane was prepared with oleyl alcohol nontoxic to the microorganism. In comparison with the continuous fermentation without product removal, the specific butanol production rate was 2 times higher. The butanol concentration in the permeate was 230 kg/m(3), which was about 50 times higher than that in the culture broth. A numerical investigation suggested a further increase in the productivity by improving the module construction.     

Deletion of kringle domains or the N-terminal hairpin structure in hepatocyte growth factor results in marked decreases in related biological activities.

To determine the essential domain for biological activity in the hepatocyte growth factor (HGF) molecule, we prepared various mutated recombinant HGFs using site-directed mutagenesis, and examined the effects on DNA synthesis in hepatocytes, scattering of MDCK cells and the antiproliferative activity on HepG2 hepatoma cells. Native HGF and mutant HGFs, in which Gln534 and/or Tyr673 were respectively substituted for His and Ser to coincide with the catalytic triad amino acids in plasmin, markedly stimulated DNA synthesis of hepatocytes and scattering of MDCK cells but inhibited DNA synthesis of HepG2 cells. The mutant HGF deleted with the third or fourth kringle domain resulted in marked decrease of all three biological activities, while deletion of the N-terminal hairpin structure or the first or second kringle domain almost completely inactivated biological activities. We propose that the N-terminal hairpin structure and the first and second kringle domains are essential for biological activities of HGF and possibly for binding to its receptor.     

Differential effect of UV irradiation on induction of intragenic and intergenic recombination during commitment to meiosis in Saccharomyces cerevisiae

A comparison was made between the induction of intragenic and intergenic recombinations during meiosis in a wild-type diploid of Saccharomyces cerevisiae. Under non-irradiated normal conditions, production of both intragenic and intergenic recombinants greatly increased in the cells with commitment to meiosis. The susceptibility of cells to the induction of both the spontaneous intra- and intergenic recombinations in meiotic cells was similar. However, under condition of UV irradiation, there were striking differences between intra- and intergenic recombinations. Susceptibility to induction of intragenic recombination by UV irradiation was not enhanced at meiosis compared with mitosis, and was not altered through commitment to meiotic processes. In contrast, however, susceptibility to the induction of intergenic recombination by UV irradiation was enhanced markedly during commitment to meiosis compared with mitosis. Genetic analysis suggested that the enhanced susceptibility to recombination during meiosis is specifically concerned with reciprocal-type recombination (crossing-over) but not non-reciprocal-type recombination (gene conversion). Hence it is concluded that the meiotic process appears to be intimately concerned with the mechanism(s) of induction of recombination, especially reciprocal-type recombination.     

Lignans of Larix leptolepis

Five lignans have been isolated from wood of Larix leptolepis. They are identified as 1-(4-hydroxy-3-methoxyphenyl)-2-4-[2-formyl-(E)-vinyl]-2-methoxyphenoxy-propane- 1,3-diol, 1-(4-hydroxy-3-methoxyphenyl)-2-2-methoxy-4-[1-(E)-propen-3-ol]-phenoxy- propane-1,3-diol, 1-(4-hydroxy-3-methoxyphenyl)-2-(4-formyl-2-methoxyphenoxy)-propane-1,3-diol, 1,2-bis-(4-hydroxy-3-methoxyphenyl)-propane-1,3-diol and a trilignol, leptolepisol C.