Inhibition of T Cell Receptor-mediated Signal Transduction by Erbstatin

Erbstatin, a tyrosine kinase inhibitor, inhibited p59^fyn- and p56^lck-kinases in vitro with IC₅₀s of 0.21 and 0.18 μg/ml, respectively. Inositol phosphates formation, enhanced by anti-CD3, was also inhibited by erbstatin. Moreover, erbstatin treatment prevented anti-CD3-induced interleukin 2 (IL-2) production, but phorbol ester-induced IL-2 production was not affected by erbstatin.     

Synergistic Inhibition of Growth by Aluminum and Iron of Tobacco (Nicotiana tabacum L.) Cells in Suspension Culture

Growth of tobacco cells at the logarithmic phase was inhibitedafter treatment with both AlCl₃ and FeSO₄ in modified Murashige-Skoogmedium prepared without P_i and EDTA at pH 5.0 for up to 20 h,whereas cells treated with either AlCl₃ or FeSO₄ (at concentrationsup to 100 µM, respectively) grew normally. These resultssuggest the synergistic inhibition of growth by Al³⁺ and Fe²⁺ions. During the exposure to both salts, there was a lag timeof about 10 h before growth inhibition became apparent in culturesof cells upon treatment with the two salts together. Then theextent of inhibition increased and reached a maximum value afterexposure for 18 h. After the lag period, cells treated withboth salts, but not cells treated with either salt alone, exhibitedsignificant increases in the amounts of both aluminum and ironin the cells and could be stained with hematoxylin, with thenuclei staining especially strongly. Cells treated with bothsalts also exhibited a decrease in the number of cells thatcould be plasmolyzed in 1 M mannitol, a decrease in the numberof cells that could be stained with fluorescein diacetate, adecrease in the amount of potassium in cells, and an increasein the extent of lipid peroxidation. These results suggest thataluminum causes the peroxidation of lipids in the presence ofiron and that lipid peroxidation alters the permeability ofthe plasma membrane and leads to cell death. (Received April 18, 1994; Accepted November 11, 1994)     

Studies on the Culture Conditions of Higher Plant Cells in Suspension Culture

To study the influence of cultural conditions on higher plant cells in suspension culture, the effects of nutritional conditions on the growth of suspended cells were investigated. Calluses were induced from 39 species of Nicotiana plants and 6 species of Populus plants on agar slant media, then these were transferred to suspension cultures. Concentrations of 2,4-D and kinetin suitable for incubation of callus from each plant were investigated and species having high growth rates in the appropriate medium were selected.

The effects of concentrations of auxins and kinetin, a variety of carbon and nitrogen sources, thiamin and myo-inositol on growth of the selected calluses were also studied. Of these calluses studied, N. glutinosa, N. tabacum var. Xanthi ova and P. hybrids were selected as calluses having high growth rates. Myo-inositol had no effect on any callus growth, and thiamin gave a distinct effect on Populus callus only. Nitrate as a nitrogen and sucrose as a carbon sources, and 2,4-D as an auxin were most effective in all calluses studied. Kinetin was essential for N. glutinosa among the calluses studied. Although high sugar concentrations tended to lengthen the lag period in the growth curve, there was no difference in the growth rates of the logarithmic phase among the concentrations.     

Effects of Nutritional Factors on the Formation of Ubiquinone by Tobacco Plant Cells in Suspension Culture

The ubiquinone (UQ) content of BY–2 cells on surface culture was examined to compare with that in suspension culture. The UQ content on surface culture was a little lower than that in suspension culture, but the pattern of the time-course of the UQ formation on surface culture was similar.

The changes of UQ content in BY–2 cells during autolysis were also examined. UQ in the cells subjected to autolysis was not rapidly metabolized nor excreted into the medium.

In order to obtain basic information for UQ formation by BY–2 cells in suspension culture, the cultural conditions, especially nutritional ones were investigated. Addition of 2,4-D was remarkably effective on UQ formation and a higher UQ content was observed with a higher 2,4-D concentration. Sucrose and glucose concentrations in the original medium were also influential factors. The UQ content tends to increase with the decrease of the sugar content. Precursors of UQ, amino acids, vitamins and organic acids were not effective on the UQ formation.     

Effects of Inorganic Nitrogen Sources and Physical Factors on the Formation of Ubiquinone by Tobacco Plant Cells in Suspension Culture

In order to obtain basic information on ubiquinone (UQ) formation by BY-2 cells in suspension culture, effects of inorganic phosphate and nitrogen sources and such physical factors as initial pH, light irradiation, shaking condition and temperature were investigated. The concentration of phosphate and nitrogen sources had no marked effect, but the ratio of ammonium nitrogen to nitrate nitrogen was significantly effective on UQ formation. The UQ content in BY-2 cells tends to increase at higher ratios of ammonium nitrogen. The increase in the UQ content was recognized at higher concentrations of 2, 4-d, especially with lower concentrations of sucrose. Physical factors had no marked effect on UQ formation except temperature. The UQ content was considerably raised at higher temperatures.     

Effects of Physical Factors and Antibiotics on the Growth of Higher Plant Cells in Suspension Culture

To study the influence of culture conditions on higher plant cells in suspension culture, the effects of antibiotics and physical factors, i.e. pH, temperature, shaking conditions and light, on the growth of suspended cells were investigated using three different callus cells; Populus hybrids (P. maximowiczii × P. nigra), N. glutinosa and N. tabacum var. xanthi ova. Of the conditions tested, red light as the light source, 32°C as the temperature, 7.5 as the initial pH value of the medium and 90 reci/min as the reciprocations of the shaker were the most favourable conditions for cell growth. In addition, the influence of various antibiotics was investigated. Of the antibiotics tested, penicillin, oleandomycin, cephaloridin and oxytetracycline did not inhibit cell growth and blasticidin-S was most toxic to all cultures.     

几种营养物质对烟草细胞生长和CoQ_(10)含量的影响

研究了蔗糖、KH2PO4、NH 4/NO3比对烟草细胞生长和CoQ10含量的影响,结果表明,当蔗糖浓度为30g/L时,CoQ10总量最高,此时细胞产量、CoQ10含量和总量分别为198g/L、4147μg/g(dwt)、8212μg/L。在上述蔗糖浓度下,当KH2PO4起始浓度为340mg/L、NH 4/NO-3为12时,细胞产量、CoQ10含量和总量分别最高,高于此比例时有利于CoQ10形成,但不利于细胞生长。     

不同培养条件对悬浮培养茶叶细胞生长及茶氨酸合成的影响

婺源绿茶嫩叶用MS培养基（加IBA 2mg／L,6-BA 4mg／L）进行茶叶愈伤组织悬浮培养,研究了不同培养条件对茶叶细胞悬浮培养过程中细胞生长与茶氨酸合成的影响。结果显示,NH4^＋／NO3^- 1．0／60．0mmol／L、K^＋ 100．0mmol／L、Mg^2＋ 3．0mmol／L、H2PO4^- 3．0mmol／L、蔗糖30．0g／L、水解酪蛋白2．0g/L条件下,茶叶细胞生长量和茶氨酸积累量均达到最高值;提高培养基中蔗糖和水解酪蛋白浓度可使细胞对数生长期和稳定期得到延长,从而有利于茶氮酸积累;H2PO4^-浓度主要影响细胞生长速率和茶氨酸积累速率的同步性,低H2PO4^-浓度环境中茶氨酸积累速率峰值滞后于细胞增长速率峰值,高H2PO4^-浓度环境中早于细胞生长速率峰值出现时间;K^＋和Mg^2＋对细胞生长的影响不明显,但影响细胞茶氨酸合成酶活性,维持适量的K^＋和Mg^2＋有利于茶氨酸积累。添加盐酸乙胺可大幅度提高茶氨酸积累量,并且先加入一定量盐酸乙胺再每天进行少量补充,茶氨酸合成量比一次性加入的效果要好。茶叶细胞生长和茶氨酸积累高峰期在整个培养过程的第19～22天出现,从生产效率考虑,培养周期以19～22天为宜。     

机械刺激诱导的烟草悬浮细胞一氧化氮产生途径及其与C矿和钙调素的关系

通过提高摇床转速对烟草细胞施加机械刺激（Ms）可诱导其胞内一氧化氮（No）的快速产生和一氧化氮合酶（Nos）活性的提高,这种MS诱导的NO产生可被N0清除剂cPTIO和NOS抑制剂L-NMMA显著抑制。此外,Ca2＋螯合剂EGTA、质膜Ca＋通道阻断剂La3＋、胞内Ca2＋通道拮抗剂钌红,以及钙调素抑制剂CPZ和TFP预处理均不同程度地抑制了机械刺激诱导的烟草细胞NO的产生,而机械刺激过程中钙调素活性显著上升并与NOS活性和NO含量的变化相一致。这些结果暗示着（类）Nos酶催化的精氨酸依赖途径可能是机械刺激诱发烟草细胞NO产生的主要途径,Ca2＋／CAM可能通过调节（类）NOS活性来调控No的产生。