The Protection Effects Of Ginaton On The Vascular Endothelial Cell Injury In Vitro Due To Iodine Excess

[Objective]To research the excess iodine effects on cultured vascular endothelial cells and the protection effects of ginaton on the injury by various kinds of methods: cell culture technology, flow cytometry and other interrelated biochemical techniques. It will support on the study of the mechanism of excess iodine and enrich the study of ginaton.[Materials and Methods]1. Cell cultivation: The vascular endothelial cell lines ECV304 were routinely grown in RPMI 1640 cultivation medium containing 10% fetal calf serum, penicillin and streptomycin in a humidified incubator at 37℃ and 5% CO₂.2. Cell counting and morphologic observing: Prepare a single cell suspension with density at 0.8～1.0× 10⁵ cells per milliliter. Plant cell in the tissue culture plate with 24 wells, and add different concentration of KI and ginaton (Shuxuening) after the cells adherence. To observe the condition of cells after 4-hour,8-hour, 12-hour and 24-hour.3. MTT experimentation: To evaluate the proliferation of endothelial cell lines ECV304. The ECV304s were exposed to iodine and ginaton at different concentrations, and finish cultivation after 4-hour, 8-hour,12-hour and 24-hour exposure. Pipette out RPMI 1640 cultivation medium and discard carefully. Add 150μ 1 DMSO to each well until the cells begin to detach, concuss it for ten minutes. Remove plate cover and measure the absorbance in each well, including the blanks, at 490 nm in a microtiter plate reader.4. Flow Cytometry: Samples were washed with phosphate-buffered saline (PBS) (pH 7.2), fixed, suspended in PBS and then analyzed using flow cytometer. Data analysis was performed with ModFit software. We use SPR represent the activity of cell proliferation, SPR=S/ (G₀/G₁+S+G₂/M) .5. Test of cell function: Test superoxide dismutase(SOD) in medium.[Results]1. Counting and proliferation activation: High iodine concentration in a certain area can promote cell proliferation, such as the group above 3 000μg/L at 4h and 700-3000μg /L at 12h. It could inhibit cell proliferation as prolong of time or extension of concentration, such as at the group above 4000μg /L at 12h and above 3000μg /L at 24h. Cell proliferation activity changes insensitively, the iodine concentration which promote cell proliferation is higher, it could still promote cell proliferation at the group above 4000μg /L at 12h and 3000-5000μg /L at 24h.2. SPR: The SPR of the free control (9.19±2.13) % was lower than 3000μg /L group (10.75±1.96) % and higher than 5000μg /L group (5.03±1.21) %, P<0.05. The rate of G₀/G₁,SPR and G₂/M was no significant difference, but the rate of apoptosis decreased as the concentration of ginaton (Shuxuening) increased at 3000μg/L group; The rate of G₀/G₁ and apoptosis was decreased as the concentration of ginaton (Shuxuening) increased, and the rate of SPR and G₂/M increased as the concentration of ginaton (Shuxuening) increased at 5000μg/L group.3. After a certain time in the high iodine concentration, SOD increased with increasing iodine concentration, and SOD decreased as the iodine concentration increased after reaching the highest values. The iodine concentration of the highest values of SOD increased with increasing concentration ginaton (Shuxuening).[Conclusions]1. Excess iodine has the injury effects to the cultured endothelial cells and different concentration of ginaton (Shuxuening) could antagonize these effects.2. The effect of excess iodine could concerned with free radicals and ginaton could antagonize this effect because it could enhance the reducibility. But how the iodine and ginaton exert influence still needs lucubrate.