共查询到20条相似文献,搜索用时 15 毫秒
1.
M Tepel H Wischniowski W Zidek 《Biochemical and biophysical research communications》1991,177(3):991-997
Using fura-2 cytosolic free calcium concentrations were measured in intact washed platelets from 9 spontaneously hypertensive rats (SHR) and from 9 age-matched normotensive Wistar-Kyoto rats (WKY). In resting platelets cytosolic free calcium concentration was significantly higher in SHR than in WKY (171.8 +/- 64.4 nM vs 93.1 +/- 59.0 nM, p less than 0.05). After preincubation with erythropoietin cytosolic free calcium concentration was significantly higher in SHR than in WKY (197.5 +/- 83.2 vs 93.0 +/- 60.1, p less than 0.01). Using platelets from SHR erythropoietin increased mean resting cytosolic free calcium concentration by 14.9% (p less than 0.05) and mean thrombin induced changes of cytosolic free calcium by 58.3% (p less than 0.01). In contrast, erythropoietin caused no significant increase in the resting calcium concentration or in thrombin induced changes of cytosolic free calcium in platelets from WKY. It is concluded that erythropoietin is involved in the pathogenesis of hypertension by elevating cytosolic free calcium concentration. 相似文献
2.
Regulation of cytosolic free calcium concentration by intrasynaptic mitochondria. 总被引:1,自引:0,他引:1
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By the use of digitonin permeabilized presynaptic nerve terminals (synaptosomes), we have found that intrasynaptic mitochondria, when studied "in situ," i.e., surrounded by their cytosolic environment, are able to buffer calcium in a range of calcium concentrations close to those usually present in the cytosol of resting synaptosomes. Adenine nucleotides and polyamines, which are usually lost during isolation of mitochondria, greatly improve the calcium-sequestering activity of mitochondria in permeabilized synaptosomes. The hypothesis that the mitochondria contributes to calcium homeostasis at low resting cytosolic free calcium concentration ([Ca2+]i) in synaptosomes has been tested; it has been found that in fact this is the case. Intrasynaptic mitochondria actively accumulates calcium at [Ca2+]i around 10(-7) M, and this activity is necessary for the regulation of [Ca2+]i. When compared with other membrane-limited calcium pools, it was found that depending on external concentration the calcium pool mobilized from mitochondria is similar or even greater than the IP3- or caffeine-sensitive calcium pools. In summary, the results presented argue in favor of a more prominent role of mitochondria in regulating [Ca2+]i in presynaptic nerve terminals, a role that should be reconsidered for other cellular types in light of the present evidence. 相似文献
3.
Transmembrane signaling by the B subunit of cholera toxin: increased cytoplasmic free calcium in rat lymphocytes 总被引:6,自引:2,他引:6
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It has previously been shown that the B subunit of cholera toxin, which binds solely to the plasma membrane ganglioside GM1, stimulates the proliferation of rat thymic lymphocytes (Spiegel, S., P. H. Fishman, and R. J. Weber, 1985, Science [Wash. DC], 230:1285-1287). The purpose of this study was to identify which transmembrane signaling system(s) are activated by the B subunit of cholera toxin. We compared the effects of B subunit and concanavalin A (Con A), a potent mitogenic lectin, on a number of second messenger systems that are putative mediators of T cell activation. Changes in the fluorescence of quin2-loaded cells revealed that mitogenic doses of either B subunit or Con A induced rapid and sustained increases in cytoplasmic free Ca2+ ([Ca2+]i). Within 5 min, [Ca2+]i increased from a basal level of 69 +/- 4 to 136 +/- 17 and 185 +/- 24 nM, respectively. The effects of B subunit and Con A were additive and largely dependent on the presence of extracellular Ca2+, though release of Ca2+ from intracellular stores could be detected for Con A, but not B subunit, using indo-1. The B subunit had no effect on either inositol phosphate levels or on the distribution of protein kinase C, indicating that, unlike Con A, the B subunit does not activate phosphoinositide hydrolysis. Fluorimetric measurements on cells loaded with bis(carboxyethyl)-5,6-carboxyfluorescein revealed that Con A induced a rapid cytoplasmic alkalinization via activation of Na+/H+ exchange, whereas B subunit had no effect on intracellular pH. Finally, by monitoring bis-oxonol fluorescence, we found that Con A induced a small hyperpolarization of the membrane potential, whereas B subunit had no acute effect. These data suggest that the biological effects of B subunit are mediated by an increase in [Ca2+]i resulting from a net influx of extracellular Ca2+. 相似文献
4.
K Tasaka K Kasahara N Masumoto J Mizuki H Kurachi A Miyake O Tanizawa 《Biochemical and biophysical research communications》1992,185(3):974-980
The effect of activin A on the cytosolic free calcium concentration ([Ca2+]i) in normal rat pituitary cells was examined using a calcium sensitive fluorescent dye, indo 1 AM, and a digital imaging fluorescent microscope system. The cells showing an increase in [Ca2+]i in response to activin A were then characterized by comparison with cells responding to growth hormone releasing hormone (GRH), thyrotropin releasing hormone (TRH), corticotropin releasing hormone (CRH), and gonadotropin releasing hormone (GnRH) in monolayer cultures of normal rat pituitary cells. Activin A increased [Ca2+]i in some cells in a mixed population of normal rat pituitary cells. The cells that responded to activin A also responded to GRH. Most of these cells were not affected by other tropic hormones (CRH, TRH, and GnRH), but a few cells responded to both GRH and TRH. None of the activin A-responding cells responded to CRH or GnRH, and none of the CRH- or GnRH-responding cells responded to activin A. In a preparation of somatotropes purified 80-90% by fluorescence-activated cell sorting, activin A increased [Ca2+]i in 30% of the cells that shows a [Ca2+]i-response to GRH. These findings suggest direct involvement of somatotropes in activin A-induced biological events in the rat pituitary gland. 相似文献
5.
Gangliosides, sialic acid-containing glycosphingolipids, are ubiquitously expressed in all eukaryotic cells and are primarily localized in the plasma membrane. Cholera toxin B subunit (Ctxb), a component of a heat-labile enterotoxin produced by Vibrio cholerae, has been frequently used as a probe to detect GM1 ganglioside because of its high affinity for this glycolipid. In this study, we evaluated the reactivity of Ctxb and the expression of GM1 in mouse embryonic neuroepithelial cells (NECs). Analysis of Ctxb reactivity of NECs based on flow cytometry revealed that about 80% of the cells are Ctxb positive. A detailed biochemical analysis, however, indicated that GM1 was expressed in NECs in barely detectable quantities. Thus, it was thought that reactivity of Ctxb in the NECs could arise from high-affinity interaction with GM1. Because Ctxb is commonly used as a reagent for flow cytometry and GM1 cell staining, we recommend that using this reagent alone would be inconclusive and that biochemical analysis of GM1 should also be performed to avoid overestimation of GM1 expression and/or mischaracterization of the ganglioside species. 相似文献
6.
Cholera toxin promotes B cell isotype differentiation 总被引:11,自引:0,他引:11
Cholera toxin (CT) is a powerful oral immunogen and adjuvant that elicits strong IgG and IgA antibody responses. In our study we investigated whether this property of CT was associated with an effect on B cell isotype differentiation. Initially, we determined the effect of CT on normal LPS-induced Peyer's patch B cells and found that whereas CT is strongly inhibitory of IgM production, it increases by approximately three-fold the number and frequency of IgG- and IgA-producing cells. Subsequently, using cell sorting technology, we demonstrated that CT acts on membrane (m)IgM+, mIgG/mIgA- B cells rather than mIgG/mIgA+ B cells. In addition, we showed that CT does not cause selective inhibition of mIgM, or enhancement of mIgG/mIgA B cell proliferation. In parallel studies we determined the effect of CT on the differentiation of a clonal B cell population, CH12.LX cells, i.e., a population comprised mainly of mIgM+ cells (98%) admixed with a small subpopulation of mIgA+ cells (2%). Here we found that CT (in the absence of LPS) causes a rapid decrease (24 h) in the intensity of mIgM expression as well as a marked increase in the size of the subpopulation expressing mIgA. In addition, we found that CT (in the presence of LPS), inhibits CH12.LX IgM production while increasing the absolute number and frequency of IgA-producing cells. In contrast, CT inhibits IgA production by CH12.LX.A2 cells, a subclone of CH12.LX cells that bears only IgA. Finally, we demonstrated that CT is equally inhibitory of the proliferation of CH12.LX cells and CH12.LX.A2 cells. Taken together, these effects of CT on normal B cells and a clonal B cell line indicate that CT induces substantial numbers of mIgM+ cells to undergo isotype differentiation into mIgG+ or mIgA+ B cells. In a final series of studies we showed that the effect of CT on isotype differentiation was mimicked by the B subunit of CT, i.e., the subunit that does not activate intracellular adenylate cyclase; thus the induction of isotype differentiation by CT is not mediated by a perturbation in cAMP level. 相似文献
7.
T R Baeker E R Simons T L Rothstein 《Journal of immunology (Baltimore, Md. : 1950)》1987,138(8):2691-2697
Cytochalasin promotes the progression of anti-immunoglobulin-treated B lymphocytes to S phase. However, the intracellular events induced by cytochalasin which may mediate signaling for progression have not been elucidated. In this study, the effect of cytochalasin on the level of intracellular free calcium in murine splenic B lymphocytes was assessed by using the fluorescent calcium indicator Indo-1. Cytochalasins A, B, D, and E induced a rapid and sustained elevation of intracellular free calcium. The calcium response to cytochalasin derived largely from the influx of extracellular calcium, although a small, transient elevation in intracellular calcium persisted when the suspension medium was made calcium-free with EGTA, implicating an intracellular source for a portion of the calcium response. Single cell fluorescence studies revealed that cytochalasin elicited a calcium response in most splenic B cells in suspension, indicating that this phenomenon is not restricted to a subpopulation of responding B cells. Phorbol esters inhibited the B cell calcium response to cytochalasin, and an established response to cytochalasin was rapidly and completely reversed by subsequently administered phorbol ester. T cells that lack the cytochalasin pathway showed a markedly diminished calcium response that was only apparent at higher cytochalasin concentration. However, B cells from xid-defective [CBA/N X DBA/2]F1 males, which fail to respond to anti-immunoglobulin plus cytochalasin, showed a calcium response to cytochalasin similar to that of phenotypically normal F1 females. These data, along with the finding that the rise in intracellular calcium occurred in naive B cells as well as B cells previously treated with anti-immunoglobulin, suggest that there is no clear association between the calcium response induced by cytochalasin and the ability of cytochalasin to stimulate progression to S phase. However, this effect of cytochalasin may suggest a connection between actin filaments and calcium influx in B cells. 相似文献
8.
Prostaglandins (PGs) are autocrine or paracrine hormones that may interact with circulating hormones such as parathyroid hormone (PTH) in bone. We examined the interaction of the PGs, PGF2 alpha, PGE2, and 6-keto-PGF1 alpha with PTH to enhance the rapid, initial transient rise in free cytosolic calcium ([Ca2+]i) and cAMP levels stimulated by PTH. Pretreatment of UMR-106, MC3T3-E1, and neonatal rat calvarial osteoblast-like cells by PGs resulted in an enhancement of the early transient rise in [Ca2+]i stimulated by PTH. PGF2 alpha was approximately 100 times more potent than PGE2. PGE2 itself was more potent than 6-keto-PGF1 alpha in enhancing PTH-stimulated rise in [Ca2+]i. Near-maximal augmentation was achieved at PGF2 alpha doses of 10 nM and PGE2 of 1 microM. The degree of augmentation in [Ca2+]i by PGF2 alpha was independent of preincubation time. PGF2 alpha pretreatment did not alter the EC50 for the PTH-induced [Ca2+]i increase but only the extent of rise in [Ca2+]i at each dose of PTH. The augmented increase in [Ca2+]i was mostly due to enhanced PTH-mediated release of Ca2+ from intracellular stores. PGF2 alpha did not stimulate an increase in PTH receptor number as assessed by [125I]-PTH-related peptide binding. PG pretreatment partially reversed PTH inhibition of cell proliferation, suggesting that an increase in [Ca2+]i may play a role in tempering the anti-proliferative effect of PTH mediated by cAMP. These studies suggest a new mode by which PGs can affect cellular activity. 相似文献
9.
We studied the effects of arachidonic acid and its metabolites on intracellular free calcium concentrations ([Ca2+]i) in highly purified bovine luteal cell preparations. Corpora lutea were collected from Holstein heifers between days 10 and 12 of the estrous cycle. The cells were dispersed and small and large cells were separated by unit gravity sedimentation and flow cytometry. The [Ca2+]i was determined by spectrofluorometry in luteal cells loaded with the fluorescent Ca2+ probe. Fura-2. Arachidonic acid elicited a dose-dependent increase in [Ca2+]i in both small and large luteal cells, having an effect at concentrations as low as 5μM; and was maximally effective at 50μM. Several other fatty acids failed to exert a similar response. Addition of nordihydroguaiaretic acid (NDGA) or indomethacin failed to suppress the effects of arachidonic acid. In fact, the presence of both inhibitors resulted in increases of [Ca2+]i, with NDGA exerting a greater stimulation of [Ca2+i than indomethacin. Prostaglandin F2α (PGF2α) as well as prostaglandin E2 (PGE2) increased [Ca2+ in the small luteal cells. These results support the idea that arachidonic acid exerts a direct action in mobilizing [Ca2+]i, in the luteal cells. Furthermore, they demonstrate that the cyclooxygenase (PGF2α and PGE2) and lipoxygenase products of arachidonic acid metabolism also play a role in increasing [Ca2+]i in bovine luteal cells. Since the bovine corpus luteum contains large quantities of arachidonic acid, these findings suggest that this compound may regulate calcium-dependent functions of the corpus luteum, including steroid and peptide hormone production and secretion. 相似文献
10.
Cholera toxin is synthesized in precursor form on free polysomes in Vibrio cholerae 569B. 总被引:2,自引:1,他引:2
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Membrane-bound and free polysomes have been isolated from Vibrio cholerae 569B. Nacent polypeptide chains were completed in a cell-free translation mixture containing Escherichia coli S-300 extracts and [3H]leucine or [35S]methionine. Cholera toxin-related polypeptides synthesized in vitro were immunologically detected after treatment with either anti-subunit A or anti-subunit B serum. Immunoreactive translation products were removed from reaction mixtures with formalinized Cowan's strain of Staphylococcus aureus, electrophoresed on sodium dodecyl sulfate-polyacrylamide gels, and visualized by fluorography. Anti-subunit A serum precipitated two major polypeptide species (molecular weights 52,000 and 45,000) from translation mixtures programed with free polysomes, whereas anti-subunit B serum precipitated only the 45,000-molecular-weight polypeptide. No cholera toxin-related polypeptides were detectable in translation mixtures programed with membrane-bound polysomes. Purified subunit A and cholera toxin competed for anti-subunit A binding sites and blocked the immunoprecipitation of the 35S-labeled 52,000- and 45,000-dalton polypeptides from in vitro translation mixtures. The data presented suggest that cholera toxin is synthesized in the cytoplasm in a precursor form on free polysomes and is secreted post-translationally. 相似文献
11.
Cholera toxin is very well characterized in terms of the activation of adenylate cyclase. In some systems, however, this cyclase activation does not seem to account for all of the physiological responses to the toxin. On the premise that cholera toxin may also exert effects through other second messenger compounds we have studied the effect of cholera toxin on the rate of Ca2+ movement across the membrane of intestinal brush border vesicles. Increasing concentrations of cholera toxin progressively accelerated the passive uptake of Ca2+ into, and the efflux of Ca2+ from, an osmotically active space in brush border membrane vesicles. This effect of cholera toxin was saturable by excess Ca2+ and was relatively specific, as the toxin did not affect vesicle permeability to an uncharged polar solute. The toxin had two high affinity Ca2+ binding sites on the A subunit as measured by equilibrium dialysis. Ca2+ transport facilitated by cholera toxin was temperature dependent, required the holotoxin, and could be inhibited by preincubation of the toxin with excess free ganglioside GM1. This increased rate of Ca2+ influx caused by the in vitro addition of cholera toxin to brush border membrane vesicles may have physiological significance as it was comparable to rates observed with the Ca ionophore A23187. Similar effects occurring in vivo could permit cholera toxin to increase cytoplasmic Ca2+ concentrations and to produce accompanying second messenger effects. 相似文献
12.
Shibata K Yoshino H Mizuno N Shinohara H Morita K Kitayama S Kurihara H Dohi T 《Prostaglandins & other lipid mediators》2000,62(4):385-394
12(R)-hydroxyeicosatetraenoic acid (HETE) shows biphasic increase in cytosolic free calcium concentration ([Ca2+]i) in rabbit and human neutrophils; the initial transient phase and the continuous falling phase. 12(S)-HETE was less potent in both species. BN50739, a platelet-activating factor (PAF) receptor antagonist, inhibited both phases of 12(R)-HETE-induced [Ca2+]i rise but did not affect leukotriene B4 (LTB4)-induced [Ca2+]i rise. N-tosyl-L-phenylalanine chloromethyl ketone (TPCK), a PAF synthesis inhibitor, and manoalide, a phospholipase A2 inhibitor, reduced 12(R)-HETE-induced [Ca2+]i rise. These blockers inhibited the continuous phase of [Ca2+]i rise induced by N-formyl-methionyl-leucyl-phenylalanine (FMLP) with little effect on the initial phase. It had no significant effect on LTB4-induced [Ca2+]i rise. SC-41930, a LTB4-receptor antagonist, did not block 12-HETE-induced [Ca2+]i rise. In 12(R)-HETE-, FMLP- and LTB4-stimulated cells, accumulations of cell-associated PAF and released PAF were detected but not in unstimulated cells. BN50739 did not affect the accumulation of cell-associated PAF and release of PAF in 12(R)-HETE-stimulated cells. These results suggest that 12(R)-HETE-induced and partially, FMLP-induced, but not LTB4-induced [Ca2+]i rise are mediated by PAF, which is produced and released by stimulation of the cells by 12(R)-HETE and FMLP, respectively. 相似文献
13.
Multiwavelength method for measuring concentration of free cytosolic calcium using the fluorescent probe indo-1 总被引:2,自引:0,他引:2
E G Popov Y Iu Gavrilov Z A Pozin EYaGabbasov 《Archives of biochemistry and biophysics》1988,261(1):91-96
It is assumed that the spectra of fluorescent probes indo-1 and fura-2 in the cytoplasm are linear combinations of the spectra of calcium-bound and free probes with weight factors proportional to the concentrations of these forms. When the concentration of calcium is measured by the dual-wavelength method, the above assumption is employed without testing. A multiwavelength method for measuring free cytosolic calcium concentration is described in the present study. The method is based on the registration of the fluorescence spectra of the probe with an optical multichannel analyzer and deconvolution of the spectra into components, corresponding to free and bound forms of the probe. A mismatch is also calculated to allow estimation of deconvolution accuracy. It was found that the spectra, recorded in aqueous calibration solution with varying calcium concentrations, can be deconvoluted into components, obtained both in the absence of calcium and at its saturating concentration. When the spectrum of the probe in the cytoplasm is deconvoluted into the same components the mismatch is higher. When aqueous calibration is used, the cytosolic calcium concentration determined by the dual-wavelength method is dependent considerably on the selected wavelengths. Our data indicate that this phenomenon may be associated with the lower polarity of cytoplasm compared to the aqueous calibration solution. Addition of either ethanol or glycerol into the calibration medium results in a considerable decrease in the mismatch. The optimal concentration of ethanol is 22-32%, and depends on the type and condition of cells tested. It is shown that the use of calibration spectra obtained in aqueous solutions leads to considerable overestimation of cytosolic calcium concentration. 相似文献
14.
Cyclic AMP inhibits platelet activation independently of its effect on cytosolic free calcium 总被引:4,自引:0,他引:4
Stimulation of platelets with the ionophore A23187, thrombin, ADP or PAF-acether resulted in a rapid increase of cytosolic free Ca2+, as measured with Quin-2, and in aggregation, 5HT secretion and - in the case of the first two agonists - thromboxane generation. PGI2 and dibutyryl cyclic AMP inhibited all these responses, except in the case of A23187, in response to which the increase in Ca2+ was unaffected, although the other responses were inhibited. The inhibition of aggregation and secretion in response to the combination of thrombin and A23187 was indistinguishable from that in response to thrombin alone. It thus appears that cAMP inhibits these responses independently of its effect in lowering cytosolic free Ca2+. 相似文献
15.
Effect of superoxide and lipid peroxide on cytosolic free calcium concentration in cultured pig aortic endothelial cells 总被引:1,自引:0,他引:1
J Hirosumi Y Ouchi M Watanabe J Kusunoki T Nakamura H Orimo 《Biochemical and biophysical research communications》1988,152(1):301-307
The effect of reactive oxygen on cytosolic free calcium concentration [( Ca++]i) in pig aortic endothelial cells (ECs) was studied. Linoleate hydroperoxide (LHO) and superoxide radicals generated from xanthine with xanthine oxidase (X-XO) were used as sources of reactive oxygen. [Ca++]i in ECs was measured with quin 2 and the value for quiescent ECs was 112 +/- 11 nM. Both LHO and X-XO increased [Ca++]i in a dose-dependent manner without accompanying the significant cellular damage. Nifedipine suppressed the increase in [Ca++]i provoked by LHO and X-XO. Thus, the biological effects of reactive oxygen might be mediated, at least in part, by the activation of voltage-dependent calcium channels in ECs. 相似文献
16.
Activation by bacterial lipopolysaccharide causes changes in the cytosolic free calcium concentration in single peritoneal macrophages 总被引:8,自引:0,他引:8
O Letari S Nicosia C Chiavaroli P Vacher W Schlegel 《Journal of immunology (Baltimore, Md. : 1950)》1991,147(3):980-983
Variations in the cytosolic free Ca2+ concentration [( Ca2+]i) upon LPS exposure were studied in single rat peritoneal macrophages loaded with fura-2 under carefully controlled conditions. Of a total of 60 cells examined, 47% responded to LPS (1 microgram/ml) with an increase in [Ca2+]i. Macrophages were heterogeneous with regard to the LPS response, with individual cells exhibiting single rapid and transient increases in [Ca2+]i, multiple transients, or slower and more sustained variations. In 62% of the responding cells, a second exposure to LPS elicited a [Ca2+]i rise, although usually to a slightly lower peak value. Thus, rapid desensitization to LPS does not occur in the majority of these macrophages. EGTA did not abolish the response of those cells that exhibited a single rapid transient in [Ca2+]i, indicating that the source of the initial [Ca2+]i rise was the intracellular stores. There was no obvious correlation between the type of response to LPS and the initial morphologic features (rounded vs polarized) of the cells. Our present work shows unequivocally that LPS induces increases in macrophage [Ca2+]i and, thereby, lends substantial support to the hypothesis that [Ca2+]i is a second messenger in LPS-mediated activation of the macrophage. 相似文献
17.
We investigated the action of cholera toxin on the intracellular ionized calcium [Ca2+]i increase induced by anti-CD2 and anti-CD3 monoclonal antibodies in the leukemic human T-cell line Jurkat. Cholera toxin inhibits in a dose-dependent manner these two pathways of human T-lymphocyte activation but with different half maximal inhibition doses (75 ng/ml for CD3, 30 ng/ml for CD2). This effect cannot be accounted for only by the increase in cAMP induced by cholera toxin because forskolin, which raises cellular cyclic adenosine monophosphate (cAMP) to the same levels, induced only a small inhibition of the [Ca2+]i increase in similar conditions. Cholera toxin induced a decrease in the surface expression of the CD3 molecule, suggesting a down-regulation of the CD3 molecules. On the other hand, the expression of CD2 remained unchanged. Cell surface disappearance of the CD3 molecule cannot account for all the inhibitory effects of cholera toxin because CD2 molecule expression was not affected (no modifications in the half maximal binding of anti-CD2 monoclonal antibodies). All together, these results suggest that cholera toxin acts on substrates, possibly G proteins, that could regulate the [Ca2+]i increase induced by anti-CD2 and anti-CD3 mAbs in Jurkat cells. In addition, the present study demonstrated that the rise in cellular cAMP partially inhibits the [Ca2+]i increase induced by anti-CD2 and anti-CD3 mAbs. 相似文献
18.
霍乱毒素B亚单位(CTB)在大肠杆菌表达体系中不能实现良好的分泌性表达。本文拟利用ctxb的自身启动子来实现CTB的高效分泌性表达。PCR方法扩增ctxb的调控序列和结构基因,克隆至pGEM-T载体,并在其下游链上肠杆菌核糖体基因的转录终止信号rmBT1T2,构建的表达质粒pGEM-T48和霍乱弧菌IEM101都实现了CTB的分泌性表达。但在pGEM-T48*TEM101)中CTB的分泌性表达量明显高于pGEM-T48(JM109)中的量,两者比较为50:1。因此,pGEM-T48(IEM101)表达体系较为理想的CTB分泌性表达体系。 相似文献
19.
Attachment to fibronectin or vitronectin makes human neutrophil migration sensitive to alterations in cytosolic free calcium concentration 总被引:2,自引:0,他引:2
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Transient increases in cytosolic free calcium concentration, [Ca2+]i, appear to be required for the migration of human neutrophils on poly-D-lysine-coated glass in the presence of dilute serum (Marks, P. W., and F. R. Maxfield. 1990. J. Cell Biol. 110:43-52). In contrast, no requirement for [Ca2+]i transients exists when neutrophils migrate on albumin-coated glass in the absence of serum. To determine the mechanism that necessitates [Ca2+]i transients on poly-D-lysine in the presence of serum, migration was examined on substrates consisting of purified adhesive glycoproteins. In the absence of external Ca2+, a treatment which causes the cessation of [Ca2+]i transients, migration on fibronectin (fn) and vitronectin (vn) was significantly inhibited. Migration was also inhibited in Ca2(+)-buffered cells on these substrates, indicating that this effect was the result of an alteration of [Ca2+]i. In the absence of external Ca2+, the inhibition of migration on fn or vn was more pronounced when soluble fn or vn was added to cells migrating on these substrates. This effect of soluble adhesive glycoprotein was specific: in the absence of external Ca2+, soluble fn did not affect the migration of cells on vn, and soluble vn did not affect the migration on fn. No additional inhibition of migration was observed in Ca2(+)-buffered cells with the addition of soluble adhesive glycoprotein. These data indicate that [Ca2+]i transients are involved in continued migration of human neutrophils on fn or vn, proteins which are part of the extracellular matrix that neutrophils encounter in vivo. 相似文献
20.
Melatonin potentiates NE-induced vasoconstriction without augmenting cytosolic calcium concentration
Vandeputte C Giummelly P Atkinson J Delagrange P Scalbert E Capdeville-Atkinson C 《American journal of physiology. Heart and circulatory physiology》2001,280(1):H420-H425
Because little is known of the intracellular mechanisms involved in the vasoconstrictor effect of melatonin (Mel), we examined the in vitro effects of Mel by using perfused cylindrical segments of the rat tail artery loaded with the intracellular Ca(2+) concentration ([Ca(2+)](i))-sensitive fluorescent dye, fura 2. Mel (10(-14) to 10(-4) M) had no effect on baseline perfusion pressure or [Ca(2+)](i) but increased, at submicromolar concentrations, the vasoconstrictor effect of norepinephrine (NE) (P = 0.0029). Mel did not modify NE-induced [Ca(2+)](i) mobilization, and thus the [Ca(2+)](i) sensitivity of NE-induced contraction increased in the presence of Mel. Mel consistently increased KCl-induced vasoconstriction and [Ca(2+)](i) sensitivity of contraction, but differences were not statistically significant. In conclusion, Mel increases the [Ca(2+)](i) sensitivity of vasoconstriction evoked by NE suggesting that Mel may amplify endogenous vasoconstrictor responses to sympathetic outflow. 相似文献