Cell proliferation and abnormal nuclei induced by radiation in renal tubule epithelium as an early manifestation of late damage

The time of appearance and the dose response of radiation effects in the mouse kidney were assessed from the determination of increases in labeling index, the appearance of proximal tubule cells with abnormally large nuclei, and kidney weight loss. Increased labeling indices and abnormally large nuclei were observed in the irradiated proximal tubule cells before any other histological changes were seen. The labeling index increased with dose (from 3 to 15 Gy) but not with time (from 1 to 12 months after irradiation). Increased labeling was evident as soon as 1 month after irradiation. Cell depletion as measured by a decrease in kidney weights compared to those of age-matched controls was not significant until 6 or more months after 11-, 13-, or 15-Gy irradiation. The frequency of cells with large nuclei increased steadily during the first 9 months after 15 Gy and tended to decline between 9 and 12 months, coincident with accelerating renal weight loss. These findings are consistent with the hypothesis that the production of these cells is a result of an abortive mitotic division and their loss is an eventual result of such an aberration. The increased proliferation induced by irradiation increases the chance for an abortive mitosis and death, presumably at a subsequent mitosis, of radiation-damaged proximal tubule cells, which is a major factor in the appearance of late radiation damage in the kidney.     

Increased plasma level of endothelin with vasoconstriction after dextran infusion.

Our previous study demonstrated that volume expansion with dextran produced blood pressure elevation due to vasoconstriction 3 hours after the cessation of infusion. To examine whether endogenous endothelin contributes to this vasoconstriction, we measured plasma level of endothelin before, immediately after, and 3 hours after the administration of dextran. Plasma level of endothelin decreased immediately after the administration (from 1.5 +/- 0.3 pg/ml to 1.1 +/- 0.2 pg/ml, P less than 0.05), and increased 3 hours after the administration (2.1 +/- 0.3 pg/ml, P less than 0.05). However, the changes in the plasma level of endothelin did not significantly correlated with those in blood pressure or total peripheral resistance. Thus, vasoconstriction after dextran infusion was accompanied by an increase in the plasma level of endothelin, but further evaluation is needed for the direct role of this peptide in the vasoconstrictive blood pressure elevation.     

Divergence pattern and selective mode in protein evolution: The example of vertebrate myoglobins and hemoglobin chains

Summary The evolutionary relation of vertebrate myoglobin and the hemoglobin chains including the agnathan hemoglobin chain is investigated on the basis of a new view of amino acid changes that is developed by canonical discriminant analysis of amino acid residues at individual sites. In contrast to the clear discrimination of amino acid residues between myoglobin, hemoglobin a chain, and hemoglobin chain in warm-blood vertebrates, the three types of globins in the lower class of vertebrates show so much variation that they are not well discriminated. This is seen particularly at the sites that are ascertained in mammals to carry the amino acid residues participating in stabilizing the monomeric structure in myoglobin and the residues forming the subunit contacts in hemoglobin. At these sites, agnathan hemoglobin chains are evaluated to be intermediate between the myoglobin and hemoglobin chains of gnathostomes. The variation in the phylogenetically lower class of globins is also seen in the internal region; there the amino acid residues of myoglobin and hemoglobin chains in the phylogenetically higher class exhibit an example of parallel evolution at the molecular level. New quantities, the distance of sequence property between discriminated groups and the variation within each group, are derived from the values of discriminant functions along the peptide chain, and this set of quantities simply describes an overall feature of globins such that the distinction between the three types of globins has been clearer as the vertebrates have evolved to become jawed, landed, and warm-blooded. This result strongly suggests that the functional constraint on the amino acid sequence of a protein is changed by living conditions and that severe conditions constitute a driving force that creates a distinctive protein from a less-constrained protein.Offprint requests to: J. Otsuka     

Karyotyping by PFGE of clinical isolates of Sporothrix schenckii

Abstract From October 1991 to December 1992 we had eight patients with sporotrichosis at Tsukuba University Hospital in Japan. With 8 strains isolated from these patients, PFGE (pulsed-field gel electrophoresis) analyses were carried out to examine whether the karyotype of S. schenckii is distinguished by our method and whether this molecular approach is a useful means of biotyping of S. schenckii strains. Chromosomes were separated by contour-clamped homogeneous electric field (CHEF) gel electrophoresis. The strains had six to eight chromosomes and a total genome size was approx. 28 Mbp. Although these karyotypes of all the isolates looked closely similar to each other, they were grouped into three types.     

Predominance of beta-structure in solubilized zona pellucida from porcine ova

The isolated zona pellucida from porcine ova was effectively solubilized in water at 60 degrees C within one hour. The circular dichroic spectra of zona in water with and without dithiothreitol showed the beta-form. Although sodium dodecyl sulfate partially induced helical structure, the beta-form was considerably retained. These results indicate that the zona glycoproteins have a structure-forming potential for the beta-structure and the hydrogen bonds of the beta-structure stabilize the supramolecular complex of the zona pellucida. The beta-form was also detected in zona solubilized by tryptic digestion. Porcine acrosin, however, did not solubilize the zona.     

Stimulative effect of prostaglandins on production of hexosamine-containing substances by cultured fibroblasts (2) early effect of various prostaglandins at various doses

Early effects of various prostaglandins on the production of hexosamine-containing substances by cultured fibroblasts, which were derived from a rat carrageenin granuloma, were studied. At the stationary phase, the cells were exposed for 6 h to one of the prostaglandin A₁ (PGA₁), A₂, B₁, B₂, D₂, F_1α, F_2α, E₁, E₂ or arachidonic acid in various concentrations ranging from 0.01 to 10 μg/ml for all the stimuli and from 10 pg to 10 μg/ml for PGF_2α. The activity of the cells in incorporating ³H-glucosamine into hexosamine-containing substances (acidic) glycosaminoglycans and glycoproteins) during this period was compared with that of control cells. All the stimuli tested showed more or less stimulative effect on the synthesis of hexosamine-containing substances at their specific concentrations. PGF_2α was found to be the most potent stimulant and its stimulative effect was found significant even at the low concentration of 100 pg/ml. PGD₂, F_1α and E₂ were the next potent stimuli. Their optimum dose were around 1 μg/ml but they still had significant stimulation at the concentration of 0.01 μg/ml. Effect of PGE₂ was rather mild. Stimulation by PGA₁, A₂, B₁ and B₂ or arachidonic acid was seen at high dose, and its seemed to be non-specific. The results suggested that these prostaglandins such as PGF_2α, D₂, F_1α and E₂ play some important role on regulating the production of intercellular ground substances.     

Biochemical and immuno- and lectin-histochemical studies of solubility and retention of bone matrix proteins during EDTA demineralization

Summary The present study utilized biochemical and immuno-and lectin-histochemical methods to demonstrate solubility and retention of mineral-binding non-collagenous proteins in rat midshaft subperiosteal bone during EDTA demineralization. A monoclonal antibody (9-A-2) specific for chondroitin 4-sulphate and dermatan sulphate and wheat germ agglutinin (WGA) specific forN-acetyl-d-glucosamine,N-acetylneuraminic acid, andN-acetyl-d-galactosamine were used. Bone proteins were extracted from fresh unfixed or aldehyde-fixed specimens with a three step extraction procedure, 4 M guanidine HCl (GdnCl), aqueous EDTA without GdnCl, followed by GdnCl. For comparison with the second extraction step, ethanolic trimethylammonium EDTA (ethanolic EDTA) was substituted for aqueous EDTA. Based on protein staining and Western blot analysis of SDS-polyacrylamide gel electrophoresis of each extract using 9-A-2 and WGA, retention of mineral-binding proteins extractable from fresh specimens with aqueous EDTA was greatly increased in tissue when ethanolic EDTA was used. Their retention was even greater with prior aldehyde fixation. Maximum retention with no detectable solubility of 9-A-2 and WGA reactive proteins was obtained after ethanolic EDTA extraction of aldehyde-fixed specimens, which concomitantly provided the strongest immuno- and lectin staining. These results indicate that this combined method dramatically improves retention of PGs and glycoproteins during demineralization of bone tissues and provides the best method for localizing these glycoconjugates.     

Nucleotide sequence of Physarum polycephalum 5.8S rRNA gene and its flanking regions.

The nucleotide sequence of Physarum polycephalum 5.8S rRNA gene and its flanking regions has been determined. The homologies of the 5.8S rRNA sequence with those of Saccharomyces, Chlamydomonas and Xenopus were 56%, 50% and 52%, respectively. In spite of these relatively low homologies, its possible secondary structure was very similar to those of other species.