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1.
We have examined the effects of retinoids on growth of cultured human skin fibroblasts from four individuals. Retinoic acid and retinol both produce a dose-dependent inhibition of growth in the four strains examined; retinoic acid was more potent than retinol in this respect. The growth inhibitory effect of retinoic acid is characterized by a decrease in the exponential growth rate, which is reversible upon removal of retinoic acid from the growth medium; the final saturation density, however, is not modified by retinoic acid treatment. No alterations of cell morphology, viability, or adhesiveness to substratum are induced by the retinoid concentrations utilized. The inhibitory effect of 10−6 M retinoic acid on cell growth is not affected by the concentration of fetal calf serum (FCS) in the medium. In all four human fibroblast strains examined, specific binding of [3H]retinoic acid to cytosol is present as determined by sucrose-density gradient centrifugation. Despite the effects of retinol on fibroblast growth, no cytoplasmic binding of [3H]retinol could be demonstrated in these cells. 相似文献
2.
We have recently demonstrated that gonadotropin releasing hormone (GnRH) acts directly on ovarian granulosa cells to inhibit the follicle stimulating hormone (FSH)-induced increase in granulosa cell steroidogenesis . A GnRH antagonist, [D-pGlu1, D-Phe2, D-Trp3,6] GnRH (A), which is known to antagonize GnRH-stimulated gonadotropin release by cultured pituitary cells, was tested in the granulosa cell system. GnRH (10?8M) inhibited estrogen and progesterone production by FSH-treated granulosa cells , whereas the antagonist A (10?6M) did not affect FSH stimulation of steroidogenesis. Antagonist A, when added together with GnRH and FSH, blocked the GnRH inhibition of FSH-induced steroidogenesis. Estrogen and progesterone production by granulosa cells was increased by 50% at a molar ratio (IDR50) of ([antagonist]/[GnRH]), respectively. At 10?6M, antagonist A completely prevented the GnRH (10?8M) inhibition. A similar effect of antagonist A was seen in FSH-induced increase of luteinizing hormone (LH) receptor content. FSH treatment for 2 days induced an 8-fold increase in LH receptor content in cultured granulosa cells; concomitant treatment with 10?8M GnRH completely inhibited the FSH effect. Antagonist A (10?6M), by itself, had no effect on the FSH action. However, when added together with FSH and GnRH, antagonist A completely abolished the inhibitory effect of GnRH. These results demonstrate that the direct inhibitory effect of GnRH on granulosa cell function can be prevented by a GnRH antagonist and that the GnRH action at the ovarian level may require stringent stereospecific interactions of these peptides with putative GnRH recognition sites. 相似文献
3.
Hiromichi Hemmi Theodore R. Breitman 《Biochemical and biophysical research communications》1982,109(3):669-674
HL-60, a human promyelocytic leukemia cell line, is induced to differentiate by retinoic acid to mature granulocytes. We have now found that after the addition of 1 μM retinoic acid to HL-60 cultures an increase in NAD+-glycohydrolase (NADase) activity is detected by 6 hr and after a 33-fold increase in activity reaches a plateau by 24 hr. Cycloheximide inhibits completely the retinoic acid-induced increase in NADase activity indicating that enzyme induction requires protein synthesis . An increase of NADase activity was found not only in HL-60 cells but also in two human monoblast cell lines (U-937 and THP-1) and fresh cells in primary culture from two patients with acute promyelocytic leukemia. An increase in synthesis of NADase does not appear to be obligatory for differentiation of HL-60 because there was no increase of NADase activity in HL-60 cells induced to differentiate with either dimethylsulfoxide, hypoxanthine, butyrate, or 1, 25-dihydroxycholecalciferol and there were marked increases in NADase activity at concentrations of retinoic acid having little or no effect on differentiation. 相似文献
4.
Paul Russell Barbara Wiggert Gerald J. Chader 《Biochimica et Biophysica Acta (BBA)/General Subjects》1978,543(4):586-589
Receptor proteins for [3H] retinol and [3H] retinoic acid in cultured human retinoblastoma cells have been separated rapidly and reproducibly by two different methods. By isoelectric focusing, the isoelectric point of the retinol receptor is at pH 4.0; the retinoic acid receptor has a higher isoelectric point of 4.3. Polyacrylamide slab gel electrophoresis revealed a slower migration rate for the [3H] retinoic acid receptor compared to the [3H] retinol receptor. The separate nature of the two proteins has thus been established in this unique human cell line. 相似文献
5.
Corneas were mounted in flux chambers and endothelial bicarbonate fluxes were determined following sensitization of endothelial cells with 5 · 10?6 M rose bengal and exposure to light. Corneas exposed to light demonstrated an increased passive bicarbonate flux compared to corneas not photosensitized. Active bicarbonate flux was reduced after 5 min of light exposure, but not after 1 min of light exposure. The increase in passive bicarbonate flux was prevented by the addition of 200 μg/ml catalase to the bathing solution; however, catalase had no effect on the photodynamic alteration of active flux. Neither 10 mM ascorbic acid nor 1.012 g/l glutathione prevented the photodynamically induced increase in passive flux. Perfusion of corneas with 5 · 10?6 M rose bengal dissolved in a sucrose-substituted Krebs-Ringer bicarbonate solution with a of and exposed to light swelled at rates more rapid than corneas treated in a similar fashion but perfused with a solution with a of . This study demonstrates that photodynamically induced corneal endothelial cell alteration results in increased passive bicarbonate flux, a time-dependent decrease in active bicarbonate flux, is oxygen dependent, and is at least in part secondary to H2O2 produced by the dismutation reaction of the superoxide free radical. 相似文献
6.
Regulation of histamine-mediated prostacyclin synthesis in cultured human vascular endothelial cells
Histamine stimulates the production of prostacyclin (PGI2) in cultured human endothelial cells. We have examined the cell specificity of histamine-mediated PGI2 synthesis in primary and subcultured human cells. Venous and arterial smooth muscle cells and skin fibroblasts synthesized PGI2 from exogenous arachidonic acid, but they did not synthesize a significant amount of PGI2 when treated with histamine. Endothelial cells, however, produced similar amounts of PGI2 in response to histamine and arachidonic acid. Thrombin also stimulates PGI2 production in endothelial cells. Histamine and thrombin yielded an additive production of PGI2 when added simultaneously to endothelial cells. When histamine and thrombin were added sequentially, the amount of PGI2 produced was not additive but equaled the amount characteristic of the first agonist alone. Following an initial treatment with histamine, endothelial cells were unable to respond to histamine for 3 hr, after which the PGI2 biosynthetic response rapidly returned to normal by hr. When the initial histamine treatment was carried out under mildly alkaline conditions, the complete return of activity was delayed to 8 hr after treatment. The synthesis of PGI2 from exogenous arachidonic acid was unaffected by prior treatment with histamine. Recovery of histamine-mediated PGI2 production was not dependent on protein synthesis but required a component of fetal calf serum that is nondialyzable and moderately heat stable. Thus endothelial cell PGI2 synthesis in response to a physiologic agonist is subject to several levels of regulation, reflecting not only intracellular events but also the extracellular environment. 相似文献
7.
We studied the effect of vitamin A and its analogues (retinoids) on the clonal growth in vitro of normal human myeloid progenitor cells. Normal human bone marrow cells were cultured in soft gel in the presence of a source of colony-stimulating factor (CSF), and various retinoids, and the number of granulocyte-macrophage colonies (CFU-GM) were scored. The addition of 3 × 10?8 to 3 × 10?6 M retinoic acid to culture plates containing CSF markedly increased the number of myeloid colonies as compared with culture plates containing CSF alone. Maximal stimulation occurred at a concentration of 3 × 10?7 M retinoic acid which increased the mean number of colonies by 213 ± 8 % (±S.E.) over plates containing CSF alone. Retinal or retinyl acetate was less potent than retinoic acid, and retinol (vitamin A) had no effect on colony growth. Retinoic acid had no direct CSF activity nor did it stimulate CSF production by the cultured bone marrow cells. Our studies show for the first time that retinoids can stimulate granulopoiesis in vitro and we suggest that this stimulation may be mediated by increased responsiveness of the granulocyte-macrophage progenitors to the action of CSF. 相似文献
8.
A component which can bind retinol and fatty acids was detected in the rat's intestinal cell cytosol following intestinal perfusion with 3H-all-trans retinol. Following Sephadex G-100 filtration of the cytosol, the void volume concentrate was treated with 2-mercaptoethanol and SDS. Sephadex G-100 filtration of the concentrate disclosed the presence of a cytosol binder of an approximate molecular weight of 12,000–17,000. The binder contained most of the 3H-retinol eluted off the column. incubation experiments disclosed that 3H-retinol could be displaced from its bindinf cytosol fraction by the addition of nonradioactive retinol, retinyl acetate, and the fatty acids octanoic, linoleic, and linolenic. Butyric acid addition did not displace 3H-retinol from its binding fraction. The intestinal cytosol binding fraction may be involved in the trans-cytosol transport of lipid compounds from the lipid cell membrane to the intracellular organelles. 相似文献
9.
T Haneji M Maekawa Y Nishimune 《Biochemical and biophysical research communications》1982,108(3):1320-1324
Artificially-induced cryptorchid testes of adult mice were cultured in order to study the effects of retinoic acid, a compound considered to be a biochemically inactive in mammalian reporductive function, on testicular germ cell differentiation, especially that of type A spermatogonia. Retinoic acid activated cell division in type A spermatogonia. and stimulate them to differentiate. It is suggested that retinoic acid might play an essential role in the control of early stages in spermatogenesis. 相似文献
10.
In previous reports from various laboratories, the levels of the microsomal cytochromes b5 and P-450 in hepatocytes in primary culture have been found to be very low and difficult to measure. The studies reported in this paper demonstrate that cytochromes b5 and P-450 in hepatocytes cultured on floating collagen membranes for periods of at least 10 days are maintained at levels readily measured by conventional techniques and comparable to those of liver . Addition of high levels of hydrocortisone (10?4M) to the culture medium for periods up to 10 days resulted in further increases in the levels of these cytochromes. Cells cultured in the presence of hydrocortisone exhibited the appearance of cytochrome P-448, in contrast to the cells cultured in the absence of hydrocortisone, where cytochrome P-450 was maintained. 相似文献
11.
Cavazzini D Catizone A Galdieri M Ottonello S 《Molecular and cellular biochemistry》2003,252(1-2):165-171
Sertoli and peritubular myoid cells, the somatic cells of the seminiferous tubule, support growth and differentiation of developing germ cells. This action strictly depends on the availability of in situ synthesized retinoic acid and we have previously documented the ability of Sertoli, but not peritubular cell extracts, to support the oxidation of retinol to retinoic acid. Using primary cultures of somatic cells treated with a physiological concentration of free retinol, we show here that the same is essentially true also for whole cultured cells. Sertoli cells are capable of producing not only retinoic acid, but are also the major site of retinyl ester (mainly, retinyl palmitate) formation. Compared with retinyl palmitate accumulation, retinoic acid synthesis was both faster and positively influenced by prior exposure to retinol. This increase in retinoic acid synthesis was further augmented by treatment with the retinoic acid catabolic inhibitor liarozole, thus indicating that enhanced synthesis, rather than reduced catabolism, is responsible for such an effect. Myoid cells had a higher capacity to incorporate exogenously supplied retinol, yet retinoic acid synthesis, and even more so retinyl palmitate formation, were considerably lower than in Sertoli cells. Retinoic acid synthesis in myoid cells was not only depressed, but also very little influenced by prior retinol exposure and totally insensitive to liarozole. These data further support the view that myoid cells are involved in retinol uptake from the blood and its transfer to other cells, rather than in metabolic interconversion or long-term storage of vitamin A, two processes that mainly take place in Sertoli cells. 相似文献
12.
Batch and continuous production of high fructose syrup from Jerusalem artichoke tubers has been studied using yeast cells immobilized in open pore gelatin matrix. In a batch reactor, the hydrolysis was 93% () and 42 mg d-fructose per ml was produced from the artichoke tuber extract by immobilized cells in 3 h. The same immobilized cells were recycled and used repeatedly for 10 batch cycles starting with fresh juice at the beginning of each cycle. It was found that immobilized cells were extremely stable and the percent hydrolysis was almost constant for all 10 batch cycles. In a continuous reactor using an immobilized cell concentration of 65.7 g (dry wt) l?1 of total working bioreactor volume, the percent hydrolysis was found to remain constant at ~100% at dilution rates <1.26 h?1, but beyond that it decreased. Volumetric productivity attained its maximum value at D = 2.08 h?1 and was found to be 100 g l?1 h?1. This was achieved at a feed sugar conversion of 80%. At 90% conversion and D = 1.66 h?1, the productivity was found to be 90 g l?1 h?1. Continuous operation of the immobilized cell bioreactor at a constant dilution rate of 1.65 h?1 for 240 h resulted in only 2% loss of original activity. 相似文献
13.
John C. Saari Sidney Futterman 《Biochimica et Biophysica Acta (BBA)/General Subjects》1976,444(3):789-793
Binding proteins for retinoic acid and retinol were separated from a supernatant prepared from bovine retina. Fraction IV from DEAE-cellulose chromatography bound exogenous [3H] retinoic acid which could not be effectively displayed by retinol, retinal, retinyl acetate or palmitate, but which was readily displaced with excess retinoic acid. [3H] Retinol was bound by fraction V from DEAE-cellulose chromatography and was not displaced by retinal, retinoic acid, retinyl acetate or retinyl palmitate, but was readily displaced by excess retinol. Unlike bovine serum retinol-binding protein, neither intracellular binding protein formed a complex with purified human serum prealbumin. The supernatant from bovine retinas was estimated to contain five times more retinoic acid binding than retinol binder. 相似文献
14.
Prostaglandin D2 was found to be a potent inhibitor of B-16 melanoma cell replication . The inhibition was dose-dependent between 3×10?9M and 3×10?6M (IC50~ 0.3 μM after 6 days). On a molar basis, PGD2 was a better inhibitor than PGA2 or 16,16-dimethyl-PGE2-methyl ester (di-M-PGE2) and in higher concentrations (10?6?10?7M), comparable to retinoic acid. In higher concentrations, PGD2 inhibited DNA, RNA and protein synthesis. The B-16 melanoma cell line which we used synthesized arachidonic acid metabolites which comigrated with PGA2, PGD2, PGE2 and PGF2α on a thin layer chromatography system. 相似文献
15.
The influence of retinol on the growth of retinal cells cannot be studied as such since when used alone it is toxic for the cells. To overcome this difficulty we obtained a serum enriched in vitamin A by feeding roosters with retinol acetate. Such a serum, in which retinol is linked to its biological carrier: the retinol binding protein, induces a maximal three folds increase in the rate of proliferation of the retinal cells versus controls. 相似文献
16.
Tetsuhiro Sakai Toshitaka Ikehara Hiroshi Miyamoto Kenichi Kaniike 《Life sciences》1981,29(23):2429-2436
Growth of HeLa cells cultured with a chemically defined medium was slightly stimulated in the presence of 5% dialyzed calf serum. The growth-promoting action of serum was more conspicuous when cell growth was suppressed in the same medium, in which K+ was replaced by Rb+ to various ratios. The growth-promoting factors(s) of serum was heat-labile. Upon addition of dialyzed serum, passive K+ or Rb+ influx was increased, whereas the active cation uptake was unaffected and cell K+ was rather decreased. The serum did not alter uptake of [3H] amino acids. Also, protein synthesis inhibited in the Rb+-substituted medium was not stimulated significantly, except that observed only when the external K+/Rb+ ratio was . From the distinct effects of serum on cell growth and protein synthesis, we conclude that (i) the serum-induced stimulation of cell growth, which is suppressed in the Rb+-substituted medium, is not a result of the direct effect of serum on synthesis of bulk protein, but a reflection of the effect on another mechanism(s) required for cell growth; and that (ii) this action is basically identical with the growth-promoting action on cells cultured in the normal medium. 相似文献
17.
Sera from hypertransfused polycythemic rabbits were found to significantly inhibit 59Fe incorporation into heme in erythroid cells in normal rabbit bone marrow cultures when compared with that of normal serum controls suggesting a higher concentration of this inhibitor in polycythemic serum. This serum inhibitor delayed the time of peak cumulative heme synthesis and the delay in peak cumulative heme synthesis was increase with increasing concentrations of polycythemic serum. It is suggested from these studies that this serum inhibitor may be involved in a negative feedback system in the control of erythropoiesis and may act specifically on differentiated nucleated erythroid cells to delay their entry into the cell cycle, consequently inhibiting heme synthesis. 相似文献
18.
This study elucidates the biochemical response of rabbit corneal keratocytes (fibroblasts) to retinol and retinoic acid in their production of collagen, fibronectin, sulfated glycosaminoglycans, collagenase, and [3H]thymidine incorporation. The morphologic appearance of cultured keratocytes was not altered by retinoid treatment. Collagen production and [3H]thymidine incorporation demonstrated a parallel decline in response to retinoids. Collagen type was unaffected as was collagenase activity. Retinoids had minimal effect on cell layer-associated 35S-labeled glycosaminoglycans, however medium-soluble 35S-glycosaminoglycans were increased. The most dramatic effect was in fibronectin synthesis which was increased 2-3-fold. These data demonstrate that rabbit keratocytes alter their biosynthesis of extracellular matrices in response to retinoids. This may be significant in corneal pathology, since the delicate balance of these extracellular macromolecules is responsible for corneal integrity and stability. 相似文献
19.
Tetradecanoyl phorbol acetate stimulates latent collagenase production by cultured human endothelial cells 总被引:15,自引:0,他引:15
Angiogenesis is associated with the fragmentation of blood vessel basement membranes. Since collagen is a major constituent of basement membranes, cultured human endothelial cells derived from umbilical cord veins were assayed for their ability to produce collagenase. Unstimulated cultured human endothelial cells did not secrete detectable levels of active collagenase into the culture medium. However, if the post-culture medium was treated with trypsin or plasmin, low levels of collagenolytic activity were detected, indicating that endothelial cells secrete small amounts of latent collagenase. Addition of the tumor promoter 12-O-tetradecanoyl phorbol-13-acetate (TPA) to the culture medium stimulated the secretion of collagenase by endothelial cells 5–30 fold. More than 90% of the collagenase was secreted in the latent form. Stimulation of collagenase production was detected at 10?9 M TPA and was maximal at 10?8 M TPA. An increase in the rate of collagenase production could be detected within 3 hr after the addition of TPA, and full induction occurred by 12 hr. Cycloheximide (3 μg/ml) or actinomycin D (0.1 μg/ml) inhibited both basal levels of collagenase production and the stimulation of collagenase production by TPA. Phorbol-12,13-didecanoate (PDD), a tumor-promoting analog of TPA, also stimulated collagenase production when administered at the same concentrations that were effective for TPA. However, 4-O-methyl TPA and 4-αPDD, two analogs of TPA which are not tumor promoters, did not stimulate collagenase production at concentrations up to 10?7 M. The collagenase produced by endothelial cells was a typical vertebrate collagenase as judged by the following criteria: it cleaved collagen into only two fragments which were three quarters and one quarter of the length of the intact molecule; it was inhibited by EDTA and human serum; it was not inhibited by inhibitors of serine, thiol or aspartate proteases. Thus TPA causes an increase in the production of latent collagenase by cultured human endothelial cells. 相似文献
20.
Ryegrass, harvested at the pre-ear emergence stage of growth, was ensiled in laboratory silos, either fresh (175 g dry matter kg?1) or wilted to five DM levels ranging from 216–432 g DM kg?1, with and without additive treatment. The additives used were “Sylade” containing sulphuric acid (15%) and formaldehyde (23%) applied at 4.6 l t?1 and an “ADD-F” mixture (7:3 by volume) applied at a similar rate (4.8 l t?1). An additional treatment included application of the mixture at a constant rate related to the DM content of the ensiled crop (25 l t?1 DM).In the untreated silages, the water-soluble carbohydrates (WSC) varied, respectively (over the DM range 175–432), from 0–32 g kg?1 DM compared with 197-6 g kg?1 DM for the “Sylade” treated silages and 256-50 g kg?1 DM for the formic acid/formalin silages treated at an additive rate of 4.8 l t?1. Corresponding ranges of protein N for the control and treatments (expressed as g kg?1 total N) were 302–447, 624-502 and 620-505, respectively. When the formic acid/formalin additive was applied at a constant level related to the DM content of the crop, although the WSC content decreased with increasing DM (247-158 g kg?1 DM), the protein N content (612 g kg?1 total N) remained constant.Grass from the same field was ensiled fresh, treated with “ADD-F” at the rate of 3.4 l t?1 fresh grass, at the rate of 4.8 l t?1 fresh grass and “Sylade” at the rate of 4.6 l t?1 fresh grass. The silages were given to Suffolk-cross wether lambs in digestibility and intake trials. Digestibility coefficients of DM and energy of the silage treated with “Sylade” were significantly lower (P < 0.05) than those of the other three silages. The DM intakes of all the silages were high, ranging from 27.7 g kg?1 live weight for the “Sylade” silage to 30.7 g kg?1 live weight for the silage treated with . Live weight gains ranged from 200 g/day for the control silage to 267 g/day for the silage. 相似文献