Histochemical staining of exogenous chromium and iron with the catalytic oxidation of phenylamines

Summary A highly sensitive and specific method for staining exogenous chromium and iron in tissues is described. This method is superior to conventional complex-forming methods with regard to its sensitivity and specificity for these metals. The staining reaction is based on the metalcatalysed oxidation of phenylamines. Tissue sections were incubated in a medium containing hydrogen peroxide and phenylamines, p-phenylenediamine or phenylhydrazine. Results obtained from test-tube experiments concerning the catalytic activities of metals indicated that the staining reactions depends on the activities of metals in tissues.     

Nucleosidediphosphate kinase from Ehrlich ascites tumor cells

A phosphate-incorporating protein has been highly purified from the cytosol of Ehrlich ascites tumor cells (EAT cells). The nitrocellulose membrane method was used to follow the progress of the purification by quantitation of the [32P]phosphorylated form of the protein. The purified protein was identified as an NDP-kinase since it exhibited NDP-kinase activity and had enzyme characteristics in common with other NDP-kinases from various mammalian cells. The purified NDP-kinase was found to have a molecular weight of approximately 76,000 daltons. Moreover, the enzyme appears to consist of two distinct polypeptides (18,000 and 20,000 daltons). This enzyme contained 19 amino acids, with high levels of glycine (9.8%) and lysine (9.0%). The enzyme rapidly formed a [32P]phosphoenzyme when incubated with [gamma-32P]ATP in the presence of Mg2+ (1 mM) at the optimum pH of 7.5 even at low temperature (below 4 degrees C). This phosphoenzyme is an enzyme-bound, high-energy-phosphate intermediate, because ATP was formed from it on incubation with ADP in the presence of Mg2+ (1 mM). This finding suggests that the phosphoenzyme functions as an intermediate in NDP-kinase action.     

Isolation of functional cDNA clones for human thymidylate synthase

Thymidine auxotrophic mutants of mouse FM3A cells due to thymidylate synthase deficiency can be transformed into prototrophs by DNA-mediated gene transfer using total human DNA (Ayusawa, D., Shimizu, K., Koyama, H., Takeishi, K., and Seno, T. (1983) J. Biol. Chem. 258, 48-53). From one such transformed cell clone, cloned recombinant lambda phages containing DNA fragments were obtained recently that were concluded by circumstantial genetic evidence to have been derived from the human thymidylate synthase gene (Takeishi, K., Ayusawa, D., Kaneda, S., Shimizu, K., and Seno, T. (1984) J. Biochem. (Tokyo) 95, 1477-1483). Using a DNA segment derived from the cloned genomic DNA fragment and free of repetitive sequences as a probe, functional cDNA corresponding to thymidylate synthase mRNA could be cloned from a cDNA library of SV40 transformed human fibroblasts constructed by Okayama and Berg (Okayama, H. and Berg, P. (1983) Mol. Cell. Biol. 3, 280-289). The cloned cDNA plasmid containing an insert of approximately 1.7-kilobase transformed mouse thymidine auxotrophic mutant cells to thymidine prototrophic cells at a frequency of 2-3 transformants/micrograms of DNA/10(5) cells, a value almost comparable to the highest so far reported. The resultant transformants retained the introduced cDNA and expressed human thymidylate synthase protein sufficient for supporting normal growth of otherwise auxotrophic mouse cells.     

Folic acid as a dietary factor affecting the wing morph of the planthopper, Laodelphax striatellus (hemiptera, delphacidae)

A factor effecting the production of brachypterous adults in Laodelphax striatellus was studied by comparing a diet on which brachypterous adults sometimes developed and a diet on which they never developed. By substituting components distinctive of the latter diet into the former, or by varying the concentration of certain components of the latter diet, the production of brachypterous adults was found to vary with the concentration of a vitamin, folic acid. Brachypterous adults were produced only when the diet contained folic acid at concentrations between 0.5 mg/l and 7.5 mg/l.

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Résumé Un facteur efficace pour la production d'adultes brachyptères chez Laodelphax striatellus a été recherché en comparant une nourriture sur laquelle des adultes brachyptères se développent parfois et une nourriture sur laquelle ils ne se développent jamais. En substituant les constituants spécifiques de cette dernière par ceux de la première ou en variant les concentrations de certains des constituants de la dernière on a montré que la production d'adultes brachyptères varie avec la concentration d'une vitamine, l'acide folique. Ceux-ci sont produits seulement lorsque l'alimentation contient de l'acide folique à des concentrations comprises entre 0,5 mg/l et 7,5 mg/l.

     

Necessity of IgE antibodies and mast cells for manifestation of resistance against larval Haemaphysalis longicornis ticks in mice

Genetically mast cell-deficient WBB6F1-W/Wv mice showed an apparent defect in manifestation of the resistance against larval Haemaphysalis longicornis ticks, but their serum IgE levels increased more than 100-fold after the second tick infestation. Immune sera obtained from the WBB6F1-W/Wv mice were adoptively transferred to the other WBB6F1-W/Wv mice which had received intracutaneous injections of WBB6F1-+/+ mouse-derived cultured mast cells. Because the resistance against ticks was detectable only when both mast cells and IgE antibodies were available, immediate hypersensitivity reaction appeared to have a physiologic role in the manifestation of the resistance against H. longicornis ticks.     

Utilization of Anhydrous Aluminum Phosphate as a Sole Source of Phosphorous by a Selected Carrot Cell Line

Two variants of carrot (Daucus carota L. cv. MS Yonsun) celllines, which had been selected with Al-phosphate as a sole sourceof phosphorous, were characterized on their mechanisms of phosphate-utilizationfrom Al-phosphate. Both cell lines excreted citrate into themedium. The amount of citrate excretion was highly correlatedwith cell growth in the presence of Al-phosphate. There wasabout a 1 to 1 correlation between solublized-Al and excreted-citratein the medium during cell growth. These results suggest that1) the citrate could chelate with Al, at a 1 to 1 ratio, inAl-phosphate, 2) the citrate-chelated Al remains outside thecells, and 3) solublized phosphate from Al-phosphate is utilizedfor the growth of carrot cells. The characteristics of the selectedcells were very stable, since the rate of citrate-excretionshowed no change after subculturing 25 passages without Al-phosphate. (Received August 7, 1989; Accepted October 19, 1989)     

Effects of the Cortex on Light- and Kinetin-Induced Accumulation of Betacyanin in the Epidermal Tissue of Phytolacca americana

Accumulation of betacyanin in the peeled green epidermis fromthe stem of P. americana was induced by incubating the epidermisin Murashige and Skoog's medium, under light, and was promotedby the presence of kinetin. However, in the epidermal tissuewith cortex attached, the accumulation of betacyanin was inhibited. (Received March 27, 1989; Accepted January 24, 1990)     

Demonstration of Antigenic and Genotypic Variation in Orientia tsutsugamushi Which Were Isolated in Japan,and Their Classification into Type and Subtype

A total of 40 strains of Orientia tsutsugamushi (34 isolates from patients and trombiculid mites in Japan, and 6 prototype strains of antigenic variants) were examined for classification based on the reactivities with type-specific monoclonal antibodies in indirect immunofluorescence tests, and on the restriction fragment length polymorphism of a polymerase chain reaction (PCR)-amplified 56-kilodalton type-specific antigenic protein gene. By these methods, several antigenic and genotypic variants were found among the strains, and these variants were classified into types and further into subtypes. These results suggest that there are many variants in O. tsutsugamushi, and the methods used here seem to be useful for the systematic classification of the numerous variants. A strain which may be a new type distinguishable from those identified previously was also found in this study. Furthermore, variety in the degree of pathogenicity in mice related to type and/or subtype classification were observed.     

Age-dependent aluminum accumulation in the human aorta and cerebral artery

The purpose of this study was to determine the extent of aluminum (Al) accumulation in the human aorta and cerebral arteries. The Al contents in the aortae and in the cerebral arteries from 23 human subjects was determined by inductively coupled plasma atomic emission spectrophotometry (ICP-AES). The subjects' age range was 45–99-yr-old; 15 of the subjects were males and 8 were females. Al was detected in twelve aortae and in six cerebral arteries, when the entire specimen was analyzed. Two specimens where Al was found in the cerebral arteries contained no Al in the aorta. No relationship to the subject's sex was found. When related to age, two groups were established. Group L (45–75 yr old) and group H (>75 yr old), which exhibited aortal Al concentrations of 33.3 and 72.7%, respectively. When the aortic wall was dissected into the tunica intima, media, and adventitia, Al was found mainly in the tunica media. In the aorta, significant relationships were found between Al and phosphorus (P) levels (r=0.801,p<0.01) and between Al and calcium (Ca) (r=0.661,p<0.05). We have concluded that Al accumulation is age-dependent and that it occurs both in the aorta and in the cerebral artery. In the aorta, accumulation occurs mainly in the tunica media. Both P and Ca appear to enhance aortal Al accumulation.