An unusual root tip formation in hairy root culture of Hyoscyamus muticus

Summary Hairy root cultures of Hyoscyamus muticus were established using Agrobacterium rhizogenes ATCC 15834. In one out of 8 clones established, an unusual root tip formation was observed after transfer of cultures from half-strength Murashige and Skoog (1962) to White's medium (1939). This phenomenon was associated with the production of a fine brownish cell suspension culture. Hairy root development resumed after transfer of the root tips from White to half-strength Murashige and Skoog medium. After plating the isolated brownish cells on hormone-free half-strength Murashige and Skoog or White solid medium, callus proliferation was observed, and then redifferentiation of hairy roots occurred. The polymerase chain reaction analysis of the H. muticus hairy root (clone Z2) revealed that only the tl region of the T-DNA was integrated. The growth and the production of five tropane alkaloids by this clone were examined.Abbreviations PCR Polymerase Chain Reaction - MS medium Murashige and Skoog Medium - 1/2 MS medium half-strength MS medium - WP medium Woody Plant medium - RC medium Root Culture medium - WH medium White medium - HPLC High Performance Liquid Chromatography - wt. weight     

Mechanisms of Fatal Cardiotoxicity following High-Dose Cyclophosphamide Therapy and a Method for Its Prevention

Observed only after administration of high doses, cardiotoxicity is the dose-limiting effect of cyclophosphamide (CY). We investigated the poorly understood cardiotoxic mechanisms of high-dose CY. A rat cardiac myocardial cell line, H9c2, was exposed to CY metabolized by S9 fraction of rat liver homogenate mixed with co-factors (CYS9). Cytotoxicity was then evaluated by 3-(4,5-dimethyl-2-thiazolyl)¬2,5-diphenyl¬2H-tetrazolium bromide (MTT) assay, lactate dehydrogenase release, production of reactive oxygen species (ROS), and incidence of apoptosis. We also investigated how the myocardial cellular effects of CYS9 were modified by acrolein scavenger N-acetylcysteine (NAC), antioxidant isorhamnetin (ISO), and CYP inhibitor β-ionone (BIO). Quantifying CY and CY metabolites by means of liquid chromatography coupled with electrospray tandem mass spectrometry, we assayed culture supernatants of CYS9 with and without candidate cardioprotectant agents. Assay results for MTT showed that treatment with CY (125–500 μM) did not induce cytotoxicity. CYS9, however, exhibited myocardial cytotoxicity when CY concentration was 250 μM or more. After 250 μM of CY was metabolized in S9 mix for 2 h, the concentration of CY was 73.6 ± 8.0 μM, 4-hydroxy-cyclophosphamide (HCY) 17.6 ± 4.3, o-carboxyethyl-phosphoramide (CEPM) 26.6 ± 5.3 μM, and acrolein 26.7 ± 2.5 μM. Inhibition of CYS9-induced cytotoxicity occurred with NAC, ISO, and BIO. When treated with ISO or BIO, metabolism of CY was significantly inhibited. Pre-treatment with NAC, however, did not inhibit the metabolism of CY: compared to control samples, we observed no difference in HCY, a significant increase of CEPM, and a significant decrease of acrolein. Furthermore, NAC pre-treatment did not affect intracellular amounts of ROS produced by CYS9. Since acrolein seems to be heavily implicated in the onset of cardiotoxicity, any competitive metabolic processing of CY that reduces its transformation to acrolein is likely to be an important mechanism for preventing cardiotoxicity.     

Electrical excitability of proteinoid microspheres composed of basic and acidic proteinoids

Material flow equilibration, an endogenous interaction adjustment process underlying fulfillment of material flow continuity during material self-assembly, also underlies electrical excitability observed in the proteinoid microspheres as simulated models of the protocell. The spikings of the membrane potentials are attributed to a singular character of the interaction rate coefficients measuring the strengths of the coupling between basic and acidic proteinoids, in which the rates change singularly with time due to material flow equilibration.     

Behaviour of mouse macrophage cell line and mouse fibroblast on copolymers containing cellulose triacetate.

Copolymers containing cellulose triacetate were prepared under various conditions. A bacteriological plastic dish was coated with the copolymer (HCTA-MDI copolymer) composed of hydrolysed cellulose triacetate (HCTA) and diphenylmethane diisocyanate (MDI). The deacetylated copolymer (DA-copolymer) was prepared by the deacetylation of HCTA-MDI copolymer. The behaviour of animal cells such as a mouse macrophage cell line (A640-BB-2 cells) and a mouse fibroblast (3T6 cells) on prepared dishes was investigated morphologically. A640-BB-2 cells showed good adhesion and smooth spreading, and 3T6 cells also showed good adhesion and sufficient cell growth on the dish coated with HCTA-MDI copolymer. These results suggest that these copolymers are useful for biomedical materials.     

Genetic study on mating type genes by a new type of tetrad analysis inClosterium ehrenbergii

In a heterothallic line ofClosterium ehrenbergii, a zygote germinates to produce two gones (meiotic products) as a result of the survival of two non-sister nuclei after meiosis. The map distance of any gene from the centromere can be determined by a new type of tetrad analysis, “representative tetrad analysis”. A mating type gene was demonstrated to be 5.0 map units apart from the centromere under chemically and physiologically controlled conditions. Concurrently, some bisexual and non-sexual gones were observed (2.3% and 5.3%, respectively), for the first time. Several mechanisms for producing these bisexual and non-sexual gones are discussed. Contribution from the Laboratory of Genetics, Faculty of Agriculture, Kyoto University, No. 437.     

A new species of genus <Emphasis Type="Italic">Stictochironomus</Emphasis> (Diptera: Chironomidae), collected in the Oze river basin,Hiroshima, Japan

A new species of the chironomid genus Stictochironomus, collected in the Oze river basin, Hiroshima, Japan, is described in terms of of adult male morphology.