Properties of lead deposits in cell walls of radish (Raphanus sativus) roots.

Various mechanisms are involved in detoxification of heavy metals such as lead (Pb) in plant cells. Most of the Pb taken up by plants accumulates in their roots. However, the detailed properties of Pb complexes in roots remain unclear. We have investigated the properties of Pb deposits in root cell walls of radish (Raphanus sativus L.) seedlings grown on glass beads bed containing Pb pellets, which are the source of Pb-contamination in shooting range soils. Pb deposits were tightly bound to cell walls. Cell wall fragments containing about 50,000 ppm Pb were prepared from the roots. After extracting Pb from the cell wall fragments using HCl, Pb ions were recombined with the Pb-extracted cell wall fragments in a solution containing Pb acetate. When the cell wall fragments were treated with pectinase (E.C. 3.2.1.15) and were chemically modified with 1-ethyl-3-dimethylamino-propylcarboimide, the Pb-rebinding ability of the treated cell wall fragments decreased. When acid-treated cell wall fragments were incubated in a solution containing Pb(2+) and excess amounts of a chelating agent, Pb recombined with the cell wall fragments were measured to estimate the affinity between Pb(2+) and the cell wall fragments. Our data show that Pb(2+) binds to carboxyl groups of cell walls. The source of the carboxyl groups is suggested to be pectic compounds. A stability constant of the Pb-cell wall complex was estimated to be about 10(8). The role of root cell walls in the mechanism underlying heavy metal tolerance was discussed.

Cytochemical and electron probe X-ray microanalysis studies on the distribution change of intracellular calcium in columella cells of soybean roots under simulated microgravity.

The columella cells of soybean roots grown under gravity and simulated microgravity induced by a clinostat were examined using potassium pyroantimonate (PA) and quantitative X-ray microanalysis of cryosections to determine the role of Ca in the regulation of the gravitropic response. Amyloplasts in the columella cells were localized exclusively at the bottom under gravity, but diffusely distributed in the cytoplasmic matrix under simulated microgravity, thus supporting the statolith theory. In the columella cells, PA precipitates containing Ca were diffusely distributed in the cytoplasmic matrix under gravity. Under simulated microgravity, however, they decreased in number and size in the cytoplasmic matrix, whereas increased only in number in the vacuole, indicating that Ca moved from the cytoplasmic matrix into the vacuole. The vacuole of columella cells contained mostly electron-dense granular structures localized along the inner surface of tonoplasts, which closely resembled the tannin vacuole reported in Mimosa pulvinar motor cells. Under simulated microgravity, their configuration changed dramatically from a granular shape to a flat plate. The quantitative X-ray microanalysis of cryosections showed that the vacuolar electron-dense structures contained a large amount of Ca. Under simulated microgravity, the concentration of Ca increased conspicuously in these vacuolar electron-dense structures, concomitantly with a marked decrease of K in the vacuoles and an increase of K in the cell walls. These results suggest that the release of Ca(2+) from, and uptake by, the vacuolar electron-dense structures is closely related to the signal transmission in the gravitropic response and that Ca movement occurs opposite to that of K.

Quest for anti-inflammatory substances using IL-1ß-stimulated gingival fibroblasts.

BACKGROUND: We have previously reported that azulene-related compounds, and alkaline extract of Sasa senanensis Rehder potently inhibited nitric oxide (NO) production by lipopolysaccharide (LPS)-stimulated mouse macrophages. We investigated here whether they can inhibit pro-inflammatory cytokine production, by activated human gingival fibroblast (HGF). MATERIALS AND METHODS: HGF was established from the periodontal tissues of extracted tooth. Viable cell number was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) method. Production of Prostaglandin E(2) (PGE(2)) and cytokines was determined by enzyme immunoassay, and enzyme-linked immunosorbent assay, respectively. RESULTS: Interleukin (IL)-1ß did not inhibit, but rather slightly stimulated the growth of HGF cells. IL-1ß stimulated the production of PGE(2), IL-6, IL-8 and monocyte chemotactic protein-1 very potently, but not that of nitric oxide and tumor necrosis factor-α. Native LPS and synthetic lipid A from E. coli and P. gingivalis was much less stimulatory. Dexamethasone, not indomethacin, was an efficient inhibitor of IL-8 production. Among five azulene-related compounds, benzo[b]cyclohepta[e][1,4]thiazine most potently inhibited the IL-8 production by HGF cells, as well as NO production by activated RAW264.7 cells. The alkaline extract of Sasa senanensis Rehder significantly inhibited IL-8 production, without affecting the cell viability. CONCLUSION: The present system may be applicable for use in the search for anti-gingivitis substances.

Hormetic and anti-radiation effects of tropolone-related compounds.

We have previously investigated a total of 173 azulene-, tropolone- and azulenequinone-related compounds for their tumor-specificity and anti-inflammatory activity. In this study, we selected six compounds that showed tumor-specific cytotoxicity (referred to as group I compounds) and five compounds that inhibited nitric oxide production by activated macrophages (referred to as group II compounds) to investigate their possible hormetic and anti-radiation effects. We have established three oral normal cell type, human gingival fibroblast HGF-1, pulp cell HPC-1 and periodontal ligament fibroblast HPLF-1, from extracted teeth and periodontal tissue. These normal cells expressed p53 protein, regardless of the growth stage (either at growing or near confluent phase), more than oral squamous cell carcinoma cell line (HSC-2). Group I compounds slightly stimulated the growth of HPL-1 cells only at restricted durations and concentrations, but did not affect that of HGF-1 and HPC-1 cells, suggesting the minor hormetic effects displayed by these compounds. We established a new evaluation system for UV-induced cellular damage using an intact HSC-2 cell system in which sodium ascorbate (vitamin C) and gallic acid, but not N-acetyl-l-cysteine nor catalase, exerted protective effects. Three group I compounds and two group II compounds significantly protected the cells from UV-induced injury, suggesting their possible anti-UV effect.

Elucidation of the mode of interaction in the UP1-telomerase RNA-telomeric DNA ternary complex which serves to recruit telomerase to telomeric DNA and to enhance the telomerase activity.

We found that UP1, a proteolytic product of heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1), both enhances and represses the telomerase activity. The formation of the UP1-telomerase RNA-telomeric DNA ternary complex was revealed by a gel retardation experiment. The interactions in the ternary and binary complexes were elucidated by NMR. UP1 has two nucleic acid-binding domains, BD1 and BD2. In the UP1-telomerase RNA binary complex, both BD1 and BD2 interact with telomerase RNA. Interestingly, when telomeric DNA was added to the binary complex, telomeric DNA bound to BD1 in place of telomerase RNA. Thus, BD1 basically binds to telomeric DNA, while BD2 mainly binds to telomerase RNA, which resulted in the formation of the ternary complex. Here, UP1 bridges telomerase and telomeric DNA. It is supposed that UP1/hnRNP A1 serves to recruit telomerase to telomeric DNA through the formation of the ternary complex. A model has been proposed for how hnRNP A1/UP1 contributes to enhancement of the telomerase activity through recruitment and unfolding of the quadruplex of telomeric DNA.