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71.
Time course changes in the cell shape and in the patterns of microfilament distribution were analyzed quantitatively using cultured porcine aortic endothelial cell monolayers before and after a shear flow exposure. Geometrical parameters of the cell and of the microfilament were measured on fluorescent photomicrographs of the cells stained with rhodamine-phalloidin. After the shear flow exposure (20 dyn cm-2, 0-24 h), the endothelial cells on glass were elongated and oriented to the direction of the flow. Under the no-flow condition, F-actin filaments were mainly localized at the periphery of the cell, although some filaments were seen in the more central portion. The angles of the filaments were randomly distributed. After 3 h, the stress fiber-like structure of an F-actin bundle was formed in the central part of the cells, and these filaments were oriented to the direction of the flow. The degree of orientation increased as the time of exposure to shear stress became longer. This change in F-actin preceded cell elongation and orientation; these changes were statistically significant only after 6 h. After 24 h, peripheral filaments were again observed, and the fluorescence intensity of rhodamine-phalloidin-stained cells was enhanced. These findings suggest that the redistribution of F-actin filaments is one of the early cellular responses to the onset of shear stress and that it is one of the most important factors controlling cell elongation and orientation to the direction of the flow. 相似文献
72.
Bacillus circulans IAM1165 produces at least two extracellular beta-1,3-glucanases that lyse fungal cell walls. One of these extracellular enzymes was purified to homogeneity. The molecular mass was 87 kDa, and the pI was 4.3. The optimum temperature of the enzyme reaction was 70 degrees C when laminarin (a soluble beta-1,3-glucan) was used as the substrate. The pH range of the enzyme was broad (pH 4.5 to 9.0), and the optimum pH was 6.5. The enzyme is an endo beta-1,3-glucanase and has a random cleavage pattern. 相似文献
73.
ATP-dependent Sr2+ transport was examined in vitro using basolateral membrane (BLM) vesicles isolated from rat renal cortex to clarify the discrimination
mechanisms between strontium (Sr) and calcium (Ca) in renal tubules during reabsorption. ATP-dependent Sr2+ uptake and Ca2+ uptake were observed in renal BLM vesicles and were inhibited by vanadate. Hill plots indicate similar kinetic behavior for
Ca2+ and Sr2+ uptake. The apparentK
m andV
max of ATP-dependent Sr2+ uptake were both higher than those for Ca2+ uptake. ATP-dependent Sr2+ uptake by BLM vesicles diminished in the presence of 0.1 μM Ca2+ and was more markedly inhibited by 1 μM Ca2+. Hill plots of Sr2+ uptake data with and without 0.1 μM Ca2+ showed that the cooperative behavior of Sr2+ uptake was not changed by Ca2+. In the presence of 0.1 μM Ca2+, the affinity of the transport system for Sr2+ and the velocity of Sr2+ uptake in the BLM were both decreased. However, the rate of Ca2+ uptake was not diminished by Sr2+ concentrations of <1.6 μM. These results suggest that Ca2+ is preferentially transported in the renal cortex BLM when Ca2+ and Sr2+ are present at the same time. 相似文献
74.
75.
We assessed pulmonary mechanics in six open-chest rabbits (3 young and 3 adult) by the forced oscillation technique between 0.16 and 10.64 Hz. Under control conditions, pulmonary resistance (RL) decreased markedly between 0.16 and 4 Hz, after which it became reasonably constant. Measurements of alveolar pressure from two alveolar capsules in each rabbit showed that the large decrease of RL with increasing frequency below 4 Hz was due to lung tissue rheology and that tissue resistance was close to zero above 4 Hz. Estimates of resistance and elastance, also obtained by fitting tidal ventilation data at 1 Hz to the equation of the linear single-compartment model, gave values for RL motion that were slightly higher than those obtained by forced oscillations at the same frequency, presumably because of the flow dependence of airways resistance. After treatment with increasing doses of aerosolized methacholine, RL and pulmonary elastance between 0.16 and 1.34 Hz progressively increased, as did the point at which the pulmonary reactance crossed zero (the resonant frequency). The alveolar pressure measurements showed the lung to become increasingly inhomogeneously ventilated in all six animals, whereas in the three younger rabbits lobar atelectasis developed at high methacholine concentrations and the alveolar capsules ceased to communicate with the central airways. We conclude that the low-frequency pulmonary impedance of rabbits exhibits the same qualitative features observed in other species and that it is a sensitive indicator of the changes in pulmonary mechanics occurring during bronchoconstriction. 相似文献
76.
Hydroperoxide decomposition by the NADP-glutathione system in rat liver mitochondria was analyzed. Mitochondria were found to contain high concentrations of the reduced form of glutathione (GSH) (4.32 +/- 0.50 nmol/mg) and NADPH (4.74 +/- 0.64 nmol/mg), and high activities of glutathione peroxidase and reductase. In the initial phase of the reaction, the rate of hydroperoxide decomposition was proportional to both the GSH level and the activity of GSH peroxidase. However, in the later steady state, the step of NADP reduction was rate-limiting, and the overall reaction rate was independent of the initial concentration of GSH, and activities of glutathione peroxidase and reductase. Some GSH was released from mitochondria during incubation, but the rate of the decomposition could be simply expressed as kappa [GSH]/2, where kappa is the first-order rate constant of the peroxidase and [GSH] is the intramitochondrial level of GSH in the steady state. The rate of the reaction in the steady state was also dependent on the NADPH level, its reciprocal being linearly correlated with [NADPH]-1. The rate of decomposition of hydroperoxide was influenced by the respiratory state. During state 3 respiration, the rate was greatly depressed, but was still considered to exceed by far the rate of physiological generation of hydroperoxide. 相似文献
77.
Trigonelline, i.e., N-methylnicotinate, which has a zwitterionic structure similar to a substrate D-amino acid, is a useful active site probe for D-amino acid oxidase (DAO). The affinity of trigonelline for DAO in the deprotonated state at the enzyme bound FAD 3-imino group is higher than in the neutral state, contrary to in the case of benzoate, which is a competitive inhibitor and is in a monoanionic form. The time course of the absorbance change was monitored for the binding of DAO with trigonelline by means of a stopped-flow technique. The reaction, on monitoring at 507 nm, was found to be biphasic at pH 8.3, with fast and slow phases. The dissociation of the 3-imino proton of the enzyme bound FAD was observed in the same time course as the slow phase. These results suggest that the positive charge of trigonelline exists near the 3-imino group of the enzyme bound FAD and interacts repulsively with the proton of the 3-imino group. The absorption spectra of the DAO-trigonelline complex at various pHs also support this hypothesis. In the catalysis of DAO, a similar mechanism may be involved, that is, the positive charge of a D-amino acid may interact repulsively with the 3-imino proton of the enzyme bound FAD, and this interaction may be important for the catalysis. 相似文献
78.
Induction and stimulation of 92-kDa gelatinase/type IV collagenase production in osteosarcoma and fibrosarcoma cell lines by tumor necrosis factor alpha 总被引:10,自引:0,他引:10
Y Okada H Tsuchiya H Shimizu K Tomita I Nakanishi H Sato M Seiki K Yamashita T Hayakawa 《Biochemical and biophysical research communications》1990,171(2):610-617
Production of a 92-kDa gelatinase/type IV collagenase and tissue inhibitor of metalloproteinases (TIMP) was investigated with human sarcoma cell lines. Among the cytokines and growth factors examined, only human recombinant tumor necrosis factor alpha (TNF alpha) induced and stimulated the proteinase with concomitant increase in TIMP expression, but matrix metalloproteinase 2 (72-kDa gelatinase/type IV collagenase) expression was unchanged. These data suggest that gene expression of the two metalloproteinases is regulated in a different fashion and TNF alpha may be important to allow cancer cells to be more invasive and metastatic. 相似文献
79.
80.
The procedure of analyzing hormone and growth factor requirements for the growth of MPC-11 cells and of developing a serum-free medium for this cell line has been described. In this medium, MPC-11 cells grow as fast as in serum-supplemented medium, up to 50 generation. MPC-11 cells grown in serum-supplemented medium secrete IgG2b and K light chain into the medium as they do in serum-containing medium. 相似文献