首页 | 本学科首页   官方微博 | 高级检索  
相似文献
 共查询到20条相似文献,搜索用时 62 毫秒
1.
Plasma phospholipases A2 (PLA2) hydrolyze phospholipids of circulating lipoproteins or deposited in arteries producing bioactive lipids believed to contribute to the atherosclerotic inflammatory response. PLA2(s) are elevated in obesity and type 2 diabetes (T2D) but it is not clear which of these conditions is the cause since they frequently coexist. This study attempts to evaluate if high plasma PLA2(s) activities and markers of their effects in lipoproteins are associated with obesity or T2D diabetes, or with both. Total PLA2 and Ca2+‐dependent and ‐independent activities, lipids, lipoproteins, apoAI, and apoB apolipoproteins and affinity of apoB‐lipoproteins for arterial proteoglycans were measured, as well as Inflammation markers. These parameters were evaluated in plasma samples of four groups: (i) apparently healthy controls with normal BMI (nBMI), (ii) obese subjects with no T2D, (iii) patients with T2D but with nBMI, and (iv) obese patients with T2D. PLA2 activities were measured in the presence and absence of Ca2+ and in the presence of specific inhibitors. Obese subjects, with or without T2D, had high activities of total PLA2 and of Ca2+‐dependent and Ca2+‐independent enzymes. The activities were correlated with inflammation markers in obese subjects with and without diabetes and with alterations of low‐density lipoproteins (LDLs) that increased their affinity for arterial proteoglycans. Ca2+‐dependent secretory (sPLA2) enzymes were the main responsible of the obesity‐associated high activity. We speculate that augmented PLA2(s) activity that increases affinity of circulating LDL for arterial intima proteoglycans could be another atherogenic component of obesity.  相似文献   

2.
O. Pantoja  C. M. Willmer 《Planta》1988,174(1):44-50
Redox systems have been reported in the plasma membrane of numerous cell types and in cells from various species of higher plant. A search for a redox system in the plasma membrane of guard cells was therefore made in efforts to explain how blue light stimulates stomatal opening, a process which is coupled to guard cell H+ efflux and K+ uptake. The rates of O2 uptake by intact guard-cell protoplasts (GCP) of Commelina communis L., in the dark, were monitored in the presence of NAD(P)H since the stimulation of O2 consumption by reduced pyridine nucleotides is used as an indicator of the presence of a redox system in the plasma membrane. Oxygen consumption by intact GCP increased two- to threefold in the presence of NAD(P)H. The NAD(P)H-stimulation of O2 uptake was dependent on Mn2+ and was stimulated 10- to 15-fold by salicylhydroxamic acid (SHAM). Catalase, cyanide and ascorbate, a superoxide scavenger, all individually inhibited the SHAM-stimulated O2 uptake. These are all characteristics of peroxidase activity although some of these features have been used to imply the presence of a redox system located in the plasma membrane. High levels of peroxidase activity (using guaiacol as a substrate) were also detected in the GCP and in the supernatant. The activity in the supernatant increased with time indicating that peroxidase was being excreted by the protoplasts. The properties of O2 uptake by the incubation medium after separation from the protoplasts were similar to those of the protoplast suspension. It is concluded that our observations can be more readily explained by peroxidase activity associated with the plasma membrane and secreted by the GCP than by the presence of a redox system in the plasma membrane of the protoplasts.Abbreviations EDTA ethylenediaminetetraacetic acid - GCP guard cell protoplast - Mes 2-(N-morpholino)ethanesulphonic acid - SHAM salicylhydroxamic acid  相似文献   

3.
We here demonstrate the presence of a plasma membrane-associated phospholipase A2 (EC 3.1.1.4; PLA2) activity in spinach (Spinacia oleracea) leaves. The pH profile of the spinach plasma membrane PLA2 activity revealed two peaks, one at pH 4.4 and one at pH 5.5. The activity at pH 5.5 had an absolute requirement of Ca2+, with full enzyme activity at 10 μmol/L Ca2+. The Ca2+-dependent PLA2 activity was both heat sensitive and stimulated by diacylglycerol, whereas ATP completely inhibited the activity. Thus, the spinach plasma membrane contains a Ca2+-dependent PLA2 activity, which has not previously been characterised in plants. Cold acclimation of spinach resulted in a 2.2-fold higher plasma membrane PLA2 activity whereas the plasma membrane phospholipase D activity remained unaffected. Taken together, our data suggest a role of PLA2 in cold acclimation in plants.  相似文献   

4.
The epithelial Ca2+ channel TRPV5 constitutes the apical entry gate for Ca2+ transport in renal epithelial cells. Ablation of the trpv5 gene in mice leads to a reduced Ca2+ reabsorption. TRPV5 is tightly regulated by various calciotropic hormones, associated proteins, and other factors, which mainly affect channel activity via the C terminus. To further identify the role of the C terminus in TRPV5 regulation, we expressed channels harboring C-terminal deletions and studied channel activity by measuring intracellular Ca2+ concentration ([Ca2+]i) using fura-2 analysis. Removal of amino acid His712 elevated the [Ca2+]i, indicating enlarged TRPV5 activity. In addition, substitution of the positively charged His712 for a negative (H712D) or neutral (H712N) amino acid also stimulated TRPV5 activity. This critical role of His712 was confirmed by patch clamp analysis, which demonstrates increased Na+ and Ca2+ currents for TRPV5-H712D. Cell surface biotinylation studies revealed enhanced plasma membrane expression of TRPV5-H712D as compared with wild-type (WT) TRPV5. This elevated plasma membrane presence also was observed with the Ca2+-impermeable TRPV5-H712D and TRPV5-WT pore mutants, demonstrating that the elevation is not due to the increased [Ca2+]i. Finally, using an internalization assay, we demonstrated a delayed cell surface retrieval for TRPV5-H712D, likely causing the increase in plasma membrane expression. Together, these results demonstrate that His712 plays an essential role in plasma membrane regulation of TRPV5 via a constitutive endocytotic mechanism.  相似文献   

5.
H+-ATPase activity in plasma membranes isolated from Avena sativa root cells is inhibited by N-ethylmaleimide, a covalent modifier of protein sulfhydryl groups. The rate of inhibition is reduced by ADP, MgADP, and MgATP, but even at 40 millimolar ADP the enzyme is only partially protected against inactivation. When plasma membranes are treated wth N-[2-3H]ethylmaleimide and analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis, prominent radioactive bands appear at Mr=100,000 and several other positions. However, only radioactivity in the Mr=100,000 protein is reduced by the presence of MgADP. These results provide independent evidence that the Mr=100,000 polypeptide which is observed in purified preparations of the enzyme is the catalytic subunit of the H+-ATPase. When tryptic peptides are produced from N-[2-3H]ethylmaleimide labeled Mr=100,000 protein and separated by reverse phase high performance liquid chromatography, two radioactive peaks are observed for which N-[2-3H]ethylmaleimide incorporation is reduced in the presence of MgADP.  相似文献   

6.
Subcellular localizations of CoA-independent transacylase and phospholipase D enzymes have been investigated in human neutrophils performing a two-step gradient system to separate plasma membranes from internal membranes and from the bulk of granules. The internal membranes were constituted by endoplasmic reticulum and by a subpopulation of specific and tertiary granules. The enzymes activities were assayed in vitro on gradient fractions using exogenous substrates. Following cell prelabelling with [3H]alkyllyso-GPC, we also analyzed the in situ localization of labelled products involving the action of both enzymes. The CoA-independent transacylase activity, together with the CoA-dependent transacylase and acyltransferase activities were only located in the internal membranes. Following 15 min cell labelling, part of the [3H]alkylacyl-GPC was recovered in plasma membranes indicating a rapid redistribution of the acylated compound. Very high contents in arachidonate containing [3H]alkylacyl-GPC were recovered both in plasma membranes and internal membranes. Phospholipase D activity being assayed in the presence of cytosol, GTPγS and gradient fractions, only the plasma membrane fractions from resting or stimulated cells allowed the enzyme to be active. The [3H]alkylacyl-GP and [3H]alkylacyl-GPethanol, phospholipase D breakdown products from [3H]alkylacyl-GPC, obtained after neutrophil prelabelling and activation by phorbol myristate acetate, were exclusively present in the plasma membranes. In contrast, the secondary generated [3H]alkylacylglycerols were equally distributed between plasma and internal membranes. No labelled product was recovered on azurophil granules. These data demonstrate that internal membranes are the site of action of the CoA-independent transacylase and plasma membranes are the site of action of the phospholipase D. This topographical separation between CoA-independent transacylase which generated substrate and phospholipase D which degraded it, suggested that subcellular localisation and traffic of substrates within the cell can be important to regulate the enzymes. © 1996 Wiley-Liss, Inc.  相似文献   

7.
The alteration of the plasma membrane (Ca2+-Mg2+)-ATPase activity in the liver of rats administered orally carbon tetrachloride (CCl4) solution was investigated. Rats received a single oral administration of CCl4 (10, 25 and 50%, 1.0 ml/100 g body weight), and 3 or 24 h later they were sacrificed. CCl4 administration caused a remarkable elevation of liver calcium content and a corresponding increase in liver plasma membrane (Ca2+-Mg2+)-ATPase activity, indicating that the increased Ca2+ pump activity is partly involved in calcium accumulation in liver cells. Moreover, the participation in regucalcin, which is an intracellular activating factor on the enzyme, was examined by using anti-regucalcin IgG. The plasma membrane (Ca2+-Mg2+)-ATPase activity increased by CCl4 administration was not entirely inhibited by the presence of anti-regucalcin IgG (1.0 and 2.5 ug/ml) in the enzyme reaction mixture. However, the effect of regucalcin (0.25–1.0 uM) to activate (Ca2+-Mg2+)-ATPase in the liver plasma membranes of normal rats was not revealed in the liver plasma membranes obtained from CCl4-administered rats. Also, the effect of regucalcin was not seen when the plasma membranes were washed with 1.0 mM EGTA, indicating that the disappearance of regucalcin effect is not dependent on calcium binding to the plasma membranes due to liver calcium accumulation. Now, the presence of dithiothreitol (5 mM) or heparin (20 ug/ml) caused a remarkable elevation of the plasma membrane (Ca2+-Mg2+)-ATPase activity in the liver obtained from CCl4-administered rats. Thus, the regucalcin effect differed from that of dithiothreitol or heparin. The present study suggests that the impairment of regucalcin effect on Ca2+ pump activity in liver plasma membranes is partly contribute to hepatic calcium accumulation induced by liver injury with CCl4 administration.  相似文献   

8.
The inhibition of activated bovine Factors VII and X by antithrombin III has been studied by kinetic methods. The reaction between Factor Xa and antithrombin III is characterized by second-order kinetics, with a rate constant of 3.9 × 103m?1s?1 at pH 7.5 at 37 °C. Inhibition in the presence of excess antithrombin III does not proceed to completion: The decay of Factor Xa deviates from pseudo-first-order kinetics and a final equilibrium is reached, suggesting reversibility. The apparent association constant, at pH 7.5, 37 °C, is 2.3 × 109m?1. The interaction of three forms of bovine Factor VII with antithrombin III has been studied by the same methods. Factor VII and the two-chain activated form, α-Factor VIIa, and the tissue factor-Factor VIIa complex are not significantly inhibited by plasma levels of antithrombin III, in the either the presence or absence of heparin.  相似文献   

9.
Diphenylmethoxyacetic acid (DPMA) is a major metabolite of diphenhydramine in monkeys, dogs, and humans. The metabolic fate of diphenhydramine (DPHM) in sheep is not yet well understood; however, preliminary studies have demonstrated the presence of DPMA in the plasma and urine of sheep following an intravenous bolus of DPHM. Our current studies employ the simultaneous intravenous co-administration of DPHM and the stable isotope analog of DPHM to investigate the pharmacokinetics of DPHM in sheep. In these studies, in order to investigate the pharmacokinetics of the DPMA metabolite, measurement of both unlabeled and stable-isotope labeled DPMA is required. Thus, a stable isotope analog of DPMA ([2H10]DPMA) was synthesized, characterized, and purified for use as an analytical standard. The quantitative method for the gas chromatography—electron-impact mass spectrometry (GC—EI-MS) analysis of DPMA and [2H10]DPMA used a single step liquid-liquid extraction procedure using toluene for sample cleanup. The samples were derivatized with N-methyl-N-(tert.-butyldimethylsilyl) trifluoroacetamide. A 1.0-μl aliquot of the prepared sample was injected into the GC-MS system and quantitated using selected-ion monitoring (SIM). One ion was monitored for each compound, namely, m/z 165 for the internal standard diphenylacetic acid, m/z 183 for DPMA, and m/z 177 for [2H10]DPMA. The ion chromatograms were free from chromatographic peaks co-eluting with the compound of interest. The calibration curve was linear from 2.5 ng/ml (limit of quantitation) to 250.0 ng/ml in both urine and plasma. The intra-day and inter-day variabilities of this assay method were within acceptable limits (below 20% at the limit of quantitation and below 10% at all other concentrations). This method was used to measure the concentration of DPMA and [2H10]DPMA in plasma and urine samples from a ewe in which equimolar amounts of DPHM and [2H10]DPHM were administered by an intravenous bolus dose via the femoral vein. DPMA appeared to persist longer in the plasma and the urine as compared to DPHM. This method is robust and reliable for the quantitation of DPMA and [2H10]DPMA in biological samples obtained from sheep (e.g. plasma and urine).  相似文献   

10.
Using the novel radioligand, [3H]-9′-nor-fusicoccin-8′-alcohol, high affinity binding sites for fusicoccin were characterized in preparations from leaves of Arabidopsis thaliana (L.) Heynh. The binding site copartitioned with the plasmalemma marker, vanadate-sensitive K+, Mg2+-ATPase, when microsomal fractions were further purified by aqueous two-phase partitioning in polyethylene glycol-dextran phase systems and sedimented at an equilibrium density of 1.17 grams per cubic centimeter in continuous sucrose density gradients, as did the ATPase marker. The binding of [3H]-9′-nor-fusicoccin-8′-alcohol was saturable and Scatchard analysis revealed a biphasic plot with two apparent dissociation constants (KD), KD1 = 1.5 nanomolar and KD2 = 42 nanomolar, for the radioligand. Binding was optimal at pH 6, thermolabile, and was reduced by 70% when the membrane vesicles were pretreated with trypsin. The data are consistent with the presence of one or several binding proteins for fusicoccin at the plasma membrane of A. thaliana. Binding of the radioligand was unaffected by pretreatment of the sites with various alkylating and reducing agents, but was reduced by 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide, diethylpyrocarbonate, chloramine T, and periodate. A number of detergents were tested to find optimum conditions for solubilization. Nonanoyl-N-methylglucamide (50 millimolar) solubilized 70% of the radioligand-binding protein complex in undissociated form. Photoaffinity labeling of membrane preparations with a tritiated azido analog of fusicoccin resulted in the labeling of a 34 ± 1 kilodalton polypeptide. Labeling of this polypeptide, presumably the fusicoccin-binding protein, was severely reduced in the presence of unlabeled fusicoccin.  相似文献   

11.
Steroid hormones, beside their classical genomic mechanism of action, exert rapid, non genomic effects in different cell types. These effects are mediated by still poorly characterized plasma membrane receptors that appear to be distinct from the classic intracellular receptors. In the present study we evaluated the non genomic effects of estradiol (17βE2) in human sperm and its effects on sperm stimulation by extracellular ATP, a potent activator of sperm acrosome reaction. In human sperm 17βE2 induced a rapid increase of intracellular calcium (Ca2+) concentrations dependent on an influx of Ca2+ from the extracellular medium. The monitoring of the plasma membrane potential variations induced by 17βE2 showed that this steroid induces a rapid plasma membrane hyperpolarization that was dependent on the presence of Ca2+ in the extracellular medium since it was absent in Ca2+ free-medium. When sperm were pre-incubated in the presence of the K+ channel inhibitor tetra-ethylammonium, the 17βE2 induced plasma membrane hyperpolarization was blunted suggesting the involvement of K+ channels in the hyperpolarizing effects of 17βE2. Extracellular ATP induced a rapid plasma membrane depolarization followed by acrosome reaction. Sperm pre-incubation with 17βE2 inhibited the effects of extracellular ATP on sperm plasma membrane potential variations and acrosome reaction. The effects of 17βE2 were specific since its inactive steroisomer 17αE2 was inactive. Furthermore the effects of 17βE2 were not inhibited by tamoxifen, an antagonist of the classic 17βE2 intracellular receptor.  相似文献   

12.
Rat liver cells isolated by the collagenase-hyaluronidase perfusion method were treated with membrane-impermeable protein reagents (7-diazonium, 1–3-naphthalene disulfonate, diazotized sulfanilic acid, 8-anilino-naphthalene disulfonate), trypsin, phospholipase A, phospholipase C, and phospholipase D. The treated cells were incubated with [1-14C]palmitate and the 14CO2 produced was taken as a measure of fatty acid uptake by the cells. 14CO2 production by the cells was not inhibited after treatments with the membrane-impermeable protein reagents or phospholipase D. Treatments with small amounts of trypsin or phospholipases A or C caused inhibition of CO2 production from tracer amounts of palmitate. The inhibition by trypsin was partially, and that by phospholipase A was fully, reversed by increasing the amount of palmitic acid in the incubation medium. The oxidation of shorter-chain fatty acids such as octanoic acid was not decreased but increased after treating the cells with trypsin or phospholipase A. The membrane-impermeable reagents inhibited the oxidation of palmitate to CO2 by liver cells isolated by mechanical dispersion. These reagents also inhibited the long-chain acyl CoA ligase activity of liver microsomes. From these results it is suggested that the inhibition of CO2 production by intact liver cells from palmitate after enzyme treatments, is due to partial removal or modification of a normal transport component for long-chain fatty acids on the plasma membrane. The possibility of proteins (or lipoproteins) buried below the surface layer of plasma membrane in fatty acid uptake by liver cells is indicated.  相似文献   

13.
Katz A  Pick U  Avron M 《Plant physiology》1992,100(3):1224-1229
The effect of different growth conditions on the activity of the Na+/H+ antiporter in Dunaliella salina has been investigated. Adaptation of D. salina cells to ammonia at alkaline pH or to high NaCl concentrations is associated with a pronounced increase in the plasma membrane Na+/H+ exchange activity. The enhanced activity is manifested both in vivo, by stimulation of Na+ influx into intact cells in response to internal acidification, and in vitro, by a larger 22Na accumulation in plasma membrane vesicles in response to an induced pH gradient. Kinetic analysis shows that the stimulation does not result from a change of the Km for Na+ but from an increase in the Vmax. In contrast, adaptation of cells to a high LiCl concentration (0.8 m) depresses the activity of the Na+/H+ antiporter. Adaptation to ammonia is also associated with a large increase of three polypeptide bands in purified plasma membrane preparations, indicating that they may compose the antiporter polypeptides. These results suggest that adaptation to ammonia or to high salinity induces overproduction of the plasma membrane Na+/H+ antiporter in Dunaliella.  相似文献   

14.
Fractionation of preparations of rat-liver membranes on linear sucrose gradients revealed different profiles for the binding of α1-, α2- and β-adrenergic radioligands. The peaks of binding activities of [3H]prazosin and [3H]epinephrine were clearly separated from those of [3H]yohimbine and [125I]iodocyanopindolol which appeared at lower sucrose densities. Enzyme marker activities in the sucrose subfractions indicated the presence of plasma membranes in all of the subfractions. Furthermore, the binding peaks of the various adrenergic radioligands cannot be correlated with the presence of membranes derived from microsomes, lysosomes or Golgi apparatus. Pretreatment of rat livers with concanavalin A, in order to prevent the fragmentation of the plasma membranes during isolation, resulted in the shift of the binding of [3H]yohimbine and [125I]iodocyanopindolol to sucrose-gradient subfractions of higher densities, clearly separate from fractions containing microsomes and Golgi apparatus. There was no distinct separation of the binding peaks of prazosin, yohimbine, and cyanopindolol in sucrose-gradient subfractions from concanavalin A-pretreated livers. These results are consistent with the hypothesis that α1-, α2-, and β-adrenergic binding sites are associated with plasma membranes, and are heterogeneously distributed on the rat-liver cell surface.  相似文献   

15.
After capacitation of guinea pig spermatozoa in vitro, the plasma membrane was mechanically separated from the spermatozoa in the presence or absence of HgCl2 and subsequently isolated by density gradient centrifugation. Examination of the spermatozoa by electron microscopy after homogenization in the presence of HgCl2 revealed that plasma membrane was removed only from the acrosomal region and remained predominately intact posterior to the equatorial segment of the sperm head, as well as the midpiece and tail. In comparison, spermatozoa homogenized under similar buffer conditions but in the absence of HgCl2 lose the large apical segment of the acrosome and the plasma membrane is removed essentially from the entire cell. If spermatozoa were homogenized in the absence of Hg2+, analysis of plasma membrane phospholipid composition revealed a complete loss of lysophosphatidylcholine (LPC) from the plasma membrane after incubation of spermatozoa in minimal capacitating medium (MCM-PL) for 2 hours. Under these culture conditions the addition of Ca2+ (5 mM) to the capacitated spermatozoa induced approximately 78 ± 5% (n = 3) of the motile spermatozoa to undergo acrosome reactions while still maintaining sperm motility (80 ± 5%) (n = 3). If the spermatozoa were homogenized in the presence of Hg2+, a time course study revealed that plasma membrane LPC loss occurred between 60 and 90 minutes of incubation. This complete loss of LPC was evident when approximately half of the capacitated spermatozoa had undergone acrosome reactions. Incubation of the spermatozoa with the metabolic and acrosome reaction inhibitor, 2-deoxyglucose (10 mM) for 2 hours, maintained the plasma membrane phospholipid composition similar to that in the noncapacitated state. These data provide evidence that changes in the plasma membrane phospholipid composition may be associated with guinea pig sperm capacitation.  相似文献   

16.
The effect of GTP on the hydrolysis of [3H]phosphatidyinositol (PI), [3H]phosphatidylinositol-4-phosphate (PIP) and [3H]phosphatidylinositol-4,5-bisphosphate (PIP2) by phospholipase C of rat brain plasma membrane, microsomes and cytosol was determined. Moreover the regulation of PI and PIP phosphorylation by GTP in brain plasma membrane was investigated.In the presence of EGTA PIP2 was actively degradted, opposite to PI and PIP which require Ca2+ for their hydrolysis. Addition of calcium ions in each case caused stimulation of inositide phosphodiesterase(s). GTP independently of calcium ions activates by about 3 times phospholipase C acting on PIP and PIP2 exclusively in the plasma membrane. PI degradation was unaffected by GTP. In the presence of Ca2+ guanine nucleotides have synergistic stimulatory effect on plasma membrane bound phospholipase C acting on PIP2. PIP kinase of brain plasma membrane was stimulated by GTP by about 20–100% in the presence of exogenous and endogenous substrate respectively. PI kinase was negligible activated by about 20% exclusively in the presence of endogenous substrate. These results indicated that guanine nucleotide modulates the level of second messengers as diacylglycerol and IP3 through the activation of phospholipase C acting on PIP2 exclusively in brain plasma membrane. The stimulation of phospholipase C by GTP may occur directly or through the enhancement of substrate level PIP2 due to stimulation of PIP kinase.  相似文献   

17.
ATP-dependent Ca2+ uptake distinct from that of the mitochondria is found in both plasma membrane and microsomal membranes of rat kidney. Activity attributed to these fractions is enhanced by ammonium oxalate and is apparently insensitive to NaN3. In contrast, rat kidney mitochondrial Ca2+ uptake is blocked by NaN3. The pH of optimal activity is significantly higher for the mitochondrial fraction. Microsomal membrane Ca2+ uptake differs from that of the plasma membrane. Microsomal membranes are four times as active as the plasma membrane at high (5 mM) ATP levels. Apparent Km values for Mg2+-ATP differ in the two preparations with a higher affinity for Mg2+-ATP found in the plasma membrane Ca2+ uptake activity of the plasma membrane preparation is readily inhibited by Na+. Sucrose gradient density fractionation indicates that the observed microsomal membrane Ca2+ pump activity is associated with membrane vesicles derived from the endoplasmic reticulum. Ca2+ pump activity of both plasma membrane and microsomal fraction is depressed din the adrenalectomized rat. This activity is not restored by a single natriuretic dose of aldosterone.  相似文献   

18.
Human plasma fibronectin contains two latent aspartic proteinases, FN-gelatinase and FN-lamininase. Both enzymes can be generated and activated in the presence of Ca2+ from the purified cathepsin D-produced 190-kDa fibronectin fragment. We investigated the proteolytic activity and cleavage specificity of both enzymes in a range of pH from 3.5 to 9.0 using the B chain of oxidized bovine insulin and chromogenic peptides as substrates. The inhibition of the enzymes by several natural inhibitors from human plasma was also tested. The specificities of FN-gelatinase and FN-lamininase are similar to other major acidic proteinases, including pepsin, renin, cathepsin D, and HIV-proteinases. Both enzymes mainly hydrolyze three peptide bonds in the oxidized insulin B chain, namely Glu–Ala (residues 13–14), Tyr–Leu (residues 16–17), and Phe–Phe (residues 24–25). For the peptide substrates H-Pro-Thr-Glu-Phe-p-nitro-Phe-Arg-Leu-OH and H-Phe-Gly-His-p-nitro-Phe-Phe-Val-Leu-OMe that were cleaved the respective values of k cat/K M were 105.1 and 11.8 mM–1 sec–1 for cleavage by FN-gelatinase, and 123.2 and 15.5 mM–1 sec–1 for cleavage by FN-lamininase. The maximal activities of both enzymes were observed in a range between pH 5.6 and 6.3 and they became inactivated at a pH value above 8.4. Both FN-gelatinase and FN-lamininase were efficiently inhibited by 2-macroglobulin.  相似文献   

19.
Abstract

The aim of the study was to assess plasma and scales levels of interleukin (IL) 18 collected from psoriatic patients with different disease activity. IL-18 concentrations were measured using an enzyme immunoassay in the plasma and scales of 34 patients with chronic plaque type psoriasis. IL-18 levels were analysed with respect to plasma-transforming growth factor β1 (TGF-β1), the disease duration and the duration of the present relapse, and psoriasis area and severity index (PASI). Plasma IL-18 concentration varied from 90 to 1300 pg ml?1 and means (368.2±42.4 pg ml?1) were significantly elevated in comparison with healthy controls (205.9±31.8 pg ml?1). The presence of IL-18 was also demonstrated in scales from skin lesions. Treatment caused a significant decrease of plasma IL-18 concentration to 250.2±13.8 pg ml?1. There was a significant correlation between plasma IL-18 levels and PASI values (r=0.554). There was no correlation between IL-18 concentration in scales and PASI, between IL-18 concentrations in plasma and scales, and between plasma IL-18 and the disease duration or duration of present relapse. Plasma TGF-β1 concentration demonstrated a significant correlation with PASI (r=0.353), but not with IL-18 levels in plasma (r=0.063) and scales (0.141). The sum of plasma levels of IL-18 and TGF-β1 divided by the optimal coefficient demonstrated a statistically significant correlation with the highest r-value. The findings confirm an association between plasma IL-18 concentration and psoriasis severity. Moreover, it was shown that combined measurement of IL-18 and TGF-β1 in plasma can be considered as a possible biomarker of psoriasis activity.  相似文献   

20.
The mesocarp tissue of zucchini (Cucurbita pepo L. cv. Black Beauty, zucchini) fruit exhibits ATP-dependent H+-pumping activities associated with tonoplast (nitrate-sensitive) and plasma membrane (vanadate-sensitive) vesicles. The two activities are easily separated on step gradients with isopycnic densities lower than usually reported (< 20% (w/w) sucrose for tonoplast; 25–35% (w/w) sucrose for plasma membrane). The tonoplast is relatively impermeable to H+ (the half-time for equilibration of a pH gradient is 23–36 min) compared to plasma membrane (half-time of 4–6 min). Anion permeability was measured by adding ATP in the absence of an accompanying K+ salt, then measuring the increase in the pH gradient caused by the addition of a K+ salt. The increase in the pH gradient is presumably due to alleviation of the Δψ component (positive inside) and consequent increase in the Δ pH component (acid inside) of the electrochemical gradient by movement of the anion into the vesicle interior. Cl and NO3 are permeable, SO42− is not. The anion permeabilities of the tonoplast and plasma membrane were similar. This is inconsistent with the marked difference in the H+ permeabilities, but might be explained by the presence of anion channel(s) associated with tonoplast-derived vesicles.  相似文献   

设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号