共查询到20条相似文献,搜索用时 15 毫秒
1.
The regulation of intracellular pH (pHi) in a renal epithelial cell line, LLC-PK1/Cl4, during re-acidification from an alkaline load was studied by 31P-NMR. Intracellular alkalinization was induced by 10 mM ammonium glucuronate or by preloading with and subsequent removal of 20% CO2; the rate of re-acidification was found to be 0.047 pH units/min and 0.053 pH units/min, respectively. This rate of re-acidification was inhibited by 83% if Cl- was removed from the extracellular medium. A similar inhibition was found in the presence of 1 mM 4-acetamido-4'-isothiocyanostilbene-2,2'-disulphonic acid (SITS) (76% inhibition) and 1 mM bumetanide (81% inhibition). No change in recovery was found after removing sodium from the extracellular medium, indicating that LLC-PK1/Cl4 cells recover from an intracellular alkaline load by a Cl-/HCO3- exchanger, which is SITS- and bumetanide-sensitive and has no requirement for sodium. In addition, the steady-state pHi in Cl4 cells was monitored by 31P-NMR. Removal of Cl- from the extracellular medium introduced an increase in pHi by 0.33 pH units, whereas 1 mM SITS and 1 mM bumetanide caused an increase in pHi by 0.14 or 0.13 pH units. In the presence of 1 mM amiloride, an inhibitor of the Na+/H+ exchanger, the steady-state pHi did not change significantly. These results indicate that at pHo 7.4 the steady-state intracellular pH of LLC-PK1/Cl4 cells strongly depends on the activity of the Cl-/HCO3- exchanger. Under the same conditions the activity of the Na+/H+ exchanger seems to be negligible. 相似文献
2.
Characterization of the renal epithelial cell line, PKE 5, using 31P- and 13C-NMR spectroscopy 总被引:1,自引:0,他引:1
In order to characterize intracellular pH regulation and cellular metabolism in PKE 5 cells, a mutant of the renal epithelial cell line LLC-PK1 supposed to lack Na+-H+ exchanger activity, 31P and 13C-NMR studies were conducted. The 31P studies on intact cell suspensions revealed that these cells have an ATP content and an ATP/ADP ratio similar to the parent cell line. Their intracellular pH, in the presence of 5 mM HCO3-, was 7.17 +/- 0.04 (n = 5) - identical to that of LLC-PK1 cells. After acid loading the cells with 15% CO2, the initial rate of realkalinization was 0.027 pH units/min (n = 6), 50% lower than in the parent cells. The recovery rate was not affected by the removal of extracellular sodium or by the addition of 1 mM amiloride. These results indicate that PKE 5 cells are devoid of Na+-H+ exchange activity, but are able to regulate their intracellular pH by amiloride-insensitive, sodium-independent mechanisms. Extracts prepared from PKE 5 cells incubated with [13C]lactate showed 13C spectra identical to those of the parent cell line. In particular, no synthesis of 13C-labeled D-glucose was observed. 相似文献
3.
R. J. T. Corbett A. R. Laptook R. L. Nunnally A. Hassan J. Jackson 《Journal of neurochemistry》1988,51(5):1501-1509
Sequential 31P and 1H nuclear magnetic resonance spectra were measured for neonatal piglets (n = 7) to determine the relationship between brain intracellular pH (pHi), lactate, and phosphorylated energy metabolites during partial ischemia. Simultaneous determinations of arterial and cerebral venous blood gases, pH, O2 content, and plasma concentrations of glucose and lactate were also made. Ischemia, induced by bilateral carotid artery ligation plus hemorrhagic hypotension for 35 min, resulted in variable reductions in ATP, phosphocreatine, and increases in Pi, H+, and lactate relative to control levels. In four piglets, whose arterial blood glucose rose above control, brain lactate exceeded 20 mumol g-1 with corresponding decreases in pHi of greater than 0.7 units compared to control levels. The extents of brain acidosis and lactosis showed a strong linear correlation with each other (r = 0.94). Maximal changes in brain lactate, pHi, and ATP at the end of ischemia showed significant positive linear correlations with the control levels of arterial blood glucose, but did not correlate with arterial glucose or arterial cerebral-venous glucose difference values during ischemia. The relationship between pHi and buffer base deficit was comparable to results reported for adult animals up to 20 mumol ml-1. However, in contrast to models proposed for adult brain, the continued linear relationship between pH and higher buffer base levels is most consistent with a theoretical model that assumes the presence of weak acid buffers with pKa values from 6.7 to 5.2. 相似文献
4.
Bugianesi R Serafini M Simone F Wu D Meydani S Ferro-Luzzi A Azzini E Maiani G 《Analytical biochemistry》2000,284(2):296-300
There is mounting evidence emphasizing the importance of intracellular antioxidant levels for maintenance of the immune function. The flavonoid quercetin, a natural antioxidant, has been shown to modulate enzymes involved in the regulation of the inflammatory response. However, up to now, there have been no studies describing quercetin levels in cells of the immune system. A gradient reversed-phase HPLC technique to identify and quantify intracellular levels of quercetin and its application in mice splenocytes are described. Mobile phases were a 0.01 M sodium phosphate monobasic solution adjusted to pH 2.8 with 85% orthophosphoric acid (buffer, Solvent A) and methanol (Solvent B) with a flow rate of 1 ml/min. An eight-channel coulometric electrode array detector was used. In vitro supplementation with increasing concentration of quercetin (25, 50, and 100 microM) raises intracellular quercetin levels in a dose-dependent manner. The method has the required features of specificity and sensitivity for monitoring quercetin uptake in cells of the immune system. 相似文献
5.
Transient increase in Ca2+ influx in Saccharomyces cerevisiae in response to glucose: effects of intracellular acidification and cAMP levels 总被引:4,自引:0,他引:4
Influx of 45Ca2+ into Saccharomyces cerevisiae was measured under experimental conditions which enabled measurements of initial rate of transport across the plasma membrane, without interference by the vacuolar Ca2+ transport system. Addition of glucose or glycerol to the cells, after pre-incubation in glucose-free medium for 5 min, caused a rapid, transient increase in 45Ca2+ influx, reaching a peak at 3-5 min after addition of substrate. Ethanol, or glycerol added with antimycin A, had no effect on 45Ca2+ influx. We have shown previously that this increase is not mediated by an effect of the substrates on intracellular ATP levels. Changes in membrane potential accounted for only a part of the glucose-stimulated 45Ca2+ influx. The roles of intracellular acidification and changes in cellular cAMP in mediating the effects of glucose on 45Ca2+ influx were examined. After a short preincubation in glucose-free medium addition of glucose caused a decrease in the intracellular pH, [pH]i, which reached a minimum value after 3 min. A transient increase in the cellular cAMP level was also observed. Addition of glycerol also caused intracellular acidification, but ethanol or glycerol added with antimycin A had no effect on [pH]i. Artificial intracellular acidification induced by exposure to isobutyric acid or to CCCP caused a transient rise in Ca2+ influx but the extent of the increase was smaller than that caused by glucose, and the time-course was different. We conclude that intracellular acidification may be responsible for part of the glucose stimulation of Ca2+ influx.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
6.
Coakley RJ Taggart C Canny G Greally P O'Neill SJ McElvaney NG 《American journal of physiology. Lung cellular and molecular physiology》2000,279(1):L66-L74
Cystic fibrosis (CF) is a condition characterized by neutrophil-mediated lung damage and bacterial colonization. The physiological basis for reported functional alterations in CF neutrophils, including increased release of neutrophil elastase, myeloperoxidase, and oxidants, is unknown. These processes are, however, regulated by intracellular pH (pH(i)). We demonstrate here that pH(i) regulation is altered in neutrophils from CF patients. Although resting pH(i) is similar, pH(i) after acid loading and activation (N-formyl-methionyl-leucyl-phenylalanine and phorbol 12-myristate 13-acetate) is more acidic in CF cells than in normal cells. Furthermore, patients with non-CF-related bronchiectasis handle acid loading and activation in a fashion similar to subjects with normal neutrophils, suggesting that chronic pulmonary inflammation alone does not explain the difference in pH(i). This is further supported by data showing that normal neutrophils exposed to the CF pulmonary milieu respond by increasing pH(i) as opposed to decreasing pH(i) as seen in activated CF neutrophils. These pH(i) differences in activated or acid-loaded CF neutrophils are abrogated by ZnCl(2) but not by amiloride and bafilomycin A(1), suggesting that passive proton conductance is abnormal in CF. In addition, DIDS, which inhibits HCO(3)(-)/Cl(-) exchange, causes alkalinization of control but not of CF neutrophils, suggesting that anion transport is also abnormal in CF neutrophils. In summary, we have shown that pH(i) regulation in CF neutrophils is intrinsically abnormal, potentially contributing to the pulmonary manifestations of the condition. 相似文献
7.
Intracellular pH (pHi) was measured in the insulin-secreting HIT-T15 cell line using the pH-sensitive fluorescent dye, 2',7'-bis(carboxyethyl)-5'(6')-carboxyfluorescein (BCECF). It was observed that the addition of a weak acid (e.g., acetate or propionate) caused a rapid decrease in pHi, followed by a slower recovery to the resting pH value. Conversely the addition of N4Cl caused an increase in pHi followed by recovery. The addition of amiloride caused a fall in pHi; however, in this case no recovery to basal pH levels was observed. Subsequent addition of a weak acid caused a further fall in pHi with no recovery. The addition of glucose caused a transient acidification followed by alkalinization. When glucose was added to cells which had been pretreated with amiloride, the initial acidification was not followed by recovery or alkalinization. Addition of glyceraldehyde, alpha-ketoisocaproate, lactate or pyruvate to HIT cells also resulted in intracellular acidification followed by recovery. Similarly, depolarisation of HIT cells by treatment with high K+ or with Ba2+ was associated with a pronounced fall in pHi, followed by a gradual recovery. Insulin secretion from HIT cells was stimulated by glucose, glyceraldehyde, alpha-ketoisocaproate, lactate, pyruvate and KCl, whilst amiloride and weak acids exerted only modest effects in the absence of glucose, but amiloride in particular markedly potentiated glucose-induced insulin release. Thus, HIT cells appear to have an amiloride-sensitive mechanism for the extrusion of protons, probably Na+-H+ exchange. Whilst intracellular acidification appears to potentiate secretory responses to nutrient stimuli, it seems unlikely that the activation of HIT cells by these nutrients occurs as a result of intracellular acidification. The mechanisms by which various nutrient and non-nutrient stimuli might exert distinct effects on pHi are discussed. 相似文献
8.
Evidence for a novel growth factor in Xenopus oocytes 总被引:1,自引:0,他引:1
C Théry P Jullien D A Lawrence 《Biochemical and biophysical research communications》1989,160(2):615-622
Xenopus oocytes contain a novel growth factor, as determined by its effect on the anchorage-dependent and -independent growth of various rat cells and on a Xenopus cell line; it is destroyed by trypsin, acidification (pH 2.0), heating (100 degrees C, 3 min), 8 M urea but not by dithiothreitol. Gel filtration of this activity in nondissociating conditions suggests the existence of aggregates and the presence of a very high (approximately 2000 Kd) and a much lower (approximately 30 Kd) form. 相似文献
9.
The equilibrium distribution of 5,5-dimethyloxazolidine-2,4-dione (DMO) between intra- and extracellular volume was used to estimate the intracellular pH in Yoshida rat ascites hepatoma AH-130 cells under different growth conditions (log, midlog and stationary). The cells were suspended in a Krebs-Ringer 25 mM phosphate buffer and the effects of variation of external pH, of glucose and amiloride addition on intracellular pH were measured. Proliferating cells had higher intracellular pH than stationary phase cells and this difference was inhibited by amiloride. On addition of glucose the fall in external pH was similar in all conditions and corresponded to lactate production. However, the intracellular pH decreased only in proliferating cells. Stationary phase cells showed an amiloride-sensitive cytoplasmic alkalinization with glucose. Glucose addition also caused prompt recovery to a normal polysomal pattern in these cells that might suggest increased efficiency of the initiation step of protein synthesis under these conditions. The data thus suggest that the increased intracellular pH of proliferating and of glucose-treated stationary phase cells is linked to the rate of protein synthesis and is mediated by the amiloride-sensitive Na+/H+ exchange system. This could lead to increased intracellular Na+ concentration under these conditions and to initiation of growth. 相似文献
10.
右旋糖酐蔗糖酶是一种以蔗糖为唯一底物,将蔗糖分子中D-葡萄糖基催化转移到受体分子上的葡萄糖基转移酶。利用右旋糖酐蔗糖酶的转糖基作用,以蔗糖为葡萄糖糖基供体,槲皮素为糖基受体,对槲皮素糖苷的酶法合成进行了探索。通过对该酶催化反应体系、催化反应条件及产物分析的研究,结果表明:在25℃下,右旋糖酐蔗糖酶能够在30%DMSO-70%乙酸-乙酸钙(0.02 mol/L,pH值5.4)的反应体系中催化合成一种槲皮素葡萄糖苷,在这个反应体系下,以10%的蔗糖作为糖基供体,槲皮素为糖基受体,右旋糖酐蔗糖酶活力为40 U/mL,转速为150 r/min,槲皮素糖苷的转化率最高,可达39.5%。通过质谱分析确定是一种槲皮素单糖苷,分子量为464。该研究结果为黄酮类物质的糖基化修饰奠定了基础。 相似文献
11.
J M Thevelein M Beullens F Honshoven G Hoebeeck K Detremerie B Griewel J A den Hollander A W Jans 《Journal of general microbiology》1987,133(8):2197-2205
Addition of glucose to derepressed cells of the yeast Saccharomyces cerevisiae is known to cause a rapid, transient increase in the cAMP level, which lasts for 1-2 min and induces a cAMP-dependent protein phosphorylation cascade. The glucose-induced cAMP signal cannot be explained solely on the basis of an increased ATP level. Transient membrane depolarization and transient intracellular acidification have been suggested as possible triggers for the cAMP peak. Addition of glucose to cells in which the plasma membrane had been depolarized still produced the increase in the cAMP level excluding membrane depolarization as the possible trigger. Using in vivo 31P NMR-spectroscopy we followed phosphate metabolism and the time course of the drop in the intracellular pH after addition of glucose with a time resolution of 15 s. Under aerobic conditions the initial pH and ATP level were high. On addition of glucose, they both showed a rapid, transient drop, which lasted for about 30 s. Under anaerobic conditions, the initial pH and ATP level were low and on addition of glucose they both increased relatively slowly compared to aerobic conditions. Several conditions were found in which the pH drop which occurs under aerobic conditions could be blocked completely without effect on the cAMP signal or without completely preventing it: addition of NH4Cl together with glucose at high extracellular pH and addition of a low concentration of glucose before a high concentration. Also, when glucose was added twice to the same cells no consistent relationship was observed between the pH drop and the cAMP peak. These results appear to exclude transient intracellular acidification as the trigger for the cAMP signal. Hence, we conclude that the effect of glucose cannot be explained on the basis of effects known to be caused by the membrane depolarizing compounds which cause increases in the cAMP level. A new, more specific kind of interaction appears to be involved. 相似文献
12.
Joseph D Tirmizi O Zhang XL Crandall ED Lubman RL 《American journal of physiology. Lung cellular and molecular physiology》2002,282(4):L675-L683
We investigated acid-base permeability properties of electrically resistive monolayers of alveolar epithelial cells (AEC) grown in primary culture. AEC monolayers were grown on tissue culture-treated polycarbonate filters. Filters were mounted in a partitioned cuvette containing two fluid compartments (apical and basolateral) separated by the adherent monolayer, cells were loaded with the pH-sensitive dye 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein, and intracellular pH was determined. Monolayers in HCO-free Na(+) buffer (140 mM Na(+), 6 mM HEPES, pH 7.4) maintained a transepithelial pH gradient between the two fluid compartments over 30 min. Replacement of apical fluid by acidic (6.4) or basic (8.0) buffer resulted in minimal changes in intracellular pH. Replacement of basolateral fluid by acidic or basic buffer resulted in transmembrane proton fluxes and intracellular acidification or alkalinization. Intracellular alkalinization was blocked > or =80% by 100 microM dimethylamiloride, an inhibitor of Na(+)/H(+) exchange, whereas acidification was not affected by a series of acid/base transport inhibitors. Additional experiments in which AEC monolayers were grown in the presence of acidic (6.4) or basic (8.0) medium revealed differential effects on bioelectric properties depending on whether extracellular pH was altered in apical or basolateral fluid compartments bathing the cells. Acid exposure reduced (and base exposure increased) short-circuit current from the basolateral side; apical exposure did not affect short-circuit current in either case. We conclude that AEC monolayers are relatively impermeable to transepithelial acid/base fluxes, primarily because of impermeability of intercellular junctions and of the apical, rather than basolateral, cell membrane. The principal basolateral acid exit pathway observed under these experimental conditions is Na(+)/H(+) exchange, whereas proton uptake into cells occurs across the basolateral cell membrane by a different, undetermined mechanism. These results are consistent with the ability of the alveolar epithelium to maintain an apical-to-basolateral (air space-to-blood) pH gradient in situ. 相似文献
13.
Persistent infection of a rat kidney cell line with Rauscher murine leukemia virus 总被引:107,自引:0,他引:107
Duc-Nguyen, Huu (National Cancer Institute, Bethesda, Md.), Edith N. Rosenblum, and Robert F. Zeigel. Persistent infection of a rat kidney cell line with Rauscher murine leukemia virus. J. Bacteriol. 92:1133-1140. 1966.-The propagation of a murine leukemia virus (Rauscher) in a kidney cell line, derived from a rat with lymphoid leukemia, was studied. A complement-fixing (CF) antigen reacting with Rauscher immune sera was detected at various passage levels, which correlated with the visualization by use of electron microscopy of viral buds and viral particles in different stages of maturation in all passages. Five-month-old monolayers continued to shed virus and to yield high CF antigen titers. The cell-free supernatant fluid from cultures of the 14th passage was shown to be infectious for a normal rat kidney cell line, as evidenced by the appearance of the CF antigen in this line. Interferon production was not demonstrated in infected cultures. The overall data indicated that rat kidney cells could be used to propagate Rauscher virus in a carrier state. 相似文献
14.
Murphy R Cherny VV Morgan D DeCoursey TE 《American journal of physiology. Lung cellular and molecular physiology》2005,288(2):L398-L408
Voltage-gated proton channels are expressed highly in rat alveolar epithelial cells. Here we investigated whether these channels contribute to pH regulation. The intracellular pH (pH(i)) was monitored using BCECF in cultured alveolar epithelial cell monolayers and found to be 7.13 in nominally HCO(3)(-)-free solutions [at external pH (pH(o)) 7.4]. Cells were acid-loaded by the NH(4)(+) prepulse technique, and the recovery was observed. Under conditions designed to eliminate the contribution of other transporters that alter pH, addition of 10 microM ZnCl(2), a proton channel inhibitor, slowed recovery about twofold. In addition, the pH(i) minimum was lower, and the time to nadir was increased. Slowing of recovery by ZnCl(2) was observed at pH(o) 7.4 and pH(o) 8.0 and in normal and high-K(+) Ringer solutions. The observed rate of Zn(2+)-sensitive pH(i) recovery required activation of a small fraction of the available proton conductance. We conclude that proton channels contribute to pH(i) recovery after an acid load in rat alveolar epithelial cells. Addition of ZnCl(2) had no effect on pH(i) in unchallenged cells, consistent with the expectation that proton channels are not open in resting cells. After inhibition of all known pH regulators, slow pH(i) recovery persisted, suggesting the existence of a yet-undefined acid extrusion mechanism in these cells. 相似文献
15.
The relationship between intracellular and extracellular pH was investigated in a murine tumor and normal tissue, prior to and following glucose injection. Isotransplants of the murine tumor FSa-II in the dorsum of the hind foot and leg muscle, were investigated in nonanesthetized mice. Extracellular pH was measured with a glass microelectrode, with a tip diameter of approximately 80 microns. Intracellular pH was evaluated by 31P-NMR spectroscopy, using a wide-bore NT-150 spectrometer operating at 60.75 MHz. Five grams per kilogram intraperitoneal glucose led to small changes in extracellular pH of muscle (-0.13 unit) measured with a microelectrode, and no change in intracellular pH measured by NMR. In contrast, tumor intracellular pH decreased by 0.34 units and tumor extracellular pH by 1.13 units. The differential effect of glucose on tumor vs normal tissue, and pronounced pH gradient which develops in tumor cells should markedly affect the intracellular:extracellular distribution of drugs which are weak acids or bases. 相似文献
16.
The effect of lactic acid on intracellular pH and adenosine output from superfused rat soleus muscle fibres 总被引:1,自引:0,他引:1
We investigated the effects of graded doses of lactic acid on the intracellular pH and adenosine output from superfused bundles of about 15 skeletal muscle fibres. Intracellular pH was determined using the fluorescent intracellular dye, 2',7'-bis-(2-carboxyethyl)-5-(and,6-) carboxyfluorescein (BCECF), and adenosine efflux was measured by HPLC. Intracellular pH was 7.07 +/- 0.05 under control conditions, which was around 0.35 units lower than extracellular pH, and adenosine output was 63 +/- 10 pmol/min/g. Lactic acid produced dose-dependent decreases in intracellular pH and dose-dependent increases in adenosine output: 10 mM lactic acid decreased intracellular pH to 6.57 +/- 0.04 and increased adenosine output to 159 +/- 34 pmol/min/g. The adenosine output and the intracellular pH were well correlated (r2 = 0.988; P < 0.01). 相似文献
17.
A R Neves A Ramos M C Nunes M Kleerebezem J Hugenholtz W M de Vos J Almeida H Santos 《Biotechnology and bioengineering》1999,64(2):200-212
The metabolism of glucose by nongrowing cells of L. lactis strain MG5267 was studied under controlled conditions of pH, temperature, and gas atmosphere (anaerobic and aerobic) using a circulating system coupled to nuclear magnetic resonance (NMR) detection that allowed a noninvasive determination of intracellular pools of intermediate metabolites by 13C-NMR with a time resolution of 30 seconds. In addition, intracellular parameters, such as pH, NTP levels, and concentration of inorganic phosphate in the cytoplasm, could be monitored on-line by 31P-NMR with a time resolution of approx. 3 min. The time course for the concentrations of intracellular fructose 1,6-bisphosphate (FBP), 3-phosphoglycerate (3-PGA), and phosphoenolpyruvate (PEP), together with kinetic measurements of substrate consumption and endproducts formation, were used as a basis for the construction of a mechanistic model for glycolysis. In vivo measurements were complemented with determinations of phosphorylated metabolites in perchloric acid extracts. A top-down model was developed by simplifying the metabolism to the resolution allowed by the experimental data collected by in vivo NMR (grouped in seven metabolic steps). This simplified mechanistic model was adjusted to the metabolite concentrations determined by in vivo NMR. The results obtained led to the rationalization of the dynamics of glucose metabolism as being driven largely by ATP surplus. This excess causes accumulation of FBP due to NAD+ limitation, whose regeneration is dependent on downstream pyruvate reduction. The model was capable of predicting qualitative shifts in the metabolism of glucose when changing from anaerobic to aerobic conditions. 相似文献
18.
James P. Garnett Michael A. Gray Robert Tarran Malcolm Brodlie Christopher Ward Emma H. Baker Deborah L. Baines 《PloS one》2013,8(10)
People with cystic fibrosis (CF) who develop related diabetes (CFRD) have accelerated pulmonary decline, increased infection with antibiotic-resistant Pseudomonas aeruginosa and increased pulmonary exacerbations. We have previously shown that glucose concentrations are elevated in airway surface liquid (ASL) of people with CF, particularly in those with CFRD. We therefore explored the hypotheses that glucose homeostasis is altered in CF airway epithelia and that elevation of glucose flux into ASL drives increased bacterial growth, with an effect over and above other cystic fibrosis transmembrane conductance regulator (CFTR)-related ASL abnormalities. The aim of this study was to compare the mechanisms governing airway glucose homeostasis in CF and non-CF primary human bronchial epithelial (HBE) monolayers, under normal conditions and in the presence of Ps. aeruginosa filtrate. HBE-bacterial co-cultures were performed in the presence of 5 mM or 15 mM basolateral glucose to investigate how changes in blood glucose, such as those seen in CFRD, affects luminal Ps. aeruginosa growth. Calu-3 cell monolayers were used to evaluate the potential importance of glucose on Ps. aeruginosa growth, in comparison to other hallmarks of the CF ASL, namely mucus hyperviscosity and impaired CFTR-dependent fluid secretions. We show that elevation of basolateral glucose promotes the apical growth of Ps. aeruginosa on CF airway epithelial monolayers more than non-CF monolayers. Ps. aeruginosa secretions elicited more glucose flux across CF airway epithelial monolayers compared to non-CF monolayers which we propose increases glucose availability in ASL for bacterial growth. In addition, elevating basolateral glucose increased Ps. aeruginosa growth over and above any CFTR-dependent effects and the presence or absence of mucus in Calu-3 airway epithelia-bacteria co-cultures. Together these studies highlight the importance of glucose as an additional factor in promoting Ps. aeruginosa growth and respiratory infection in CF disease. 相似文献
19.
The use of functional fluorescent dyes has allowed us to monitor intracellular pH in individually identified cells in renal epithelia. Using video microscopy we simultaneously measured the change in intracellular pH in several contiguous cells in response to various maneuvers. The video equipment included a silicon intensified target camera, a VHS videocassette recorder, a high resolution monochrome monitor, a video photometric analyzer and a 2-channel chart recorder. This equipment had a spatial resolution of 1 micron by light microscopy and a response time of less than 200 ms; it allowed us to perform double fluorescent labeling and obtain reliable measurements of intracellular pH, independent of gain, regardless of the location of the image on the screen. Using this video system we have shown that there is substantial heterogeneity in activity of H+/HCO3- transport pathways among adjacent cells in a monolayer of cells cultured from the rat renal inner medullary collecting duct. In isolated perfused rabbit renal cortical collecting ducts, video microscopy allowed us to show that there are two different types of intercalated cells: one that exhibits apical Cl-/HCO3- exchange and one that does not. Both show alkaline intracellular pH with respect to non-acid-base transporting epithelia. Video microscopy has several advantages over conventional microspectrophotometry. It provides rapid data acquisition along with increased sensitivity and the capacity for some subcellular analyses. One is able to analyze several individually identified cells during an experimental maneuver. The present video system was assembled for less than $15,000 and permits a more complete analysis of an epithelium than either single-cell photometry or spectrophotometric analysis of thousands of cells in suspension or monolayers. 相似文献
20.
Quercetin interaction with the chloroplast ATPase complex 总被引:1,自引:0,他引:1
1. Quercetin, a flavonoid which acts as an energy transfer inhibitor in photophosphorylation is shown to inhibit the P-ATP exchange activity of membrane-bound CF1 and the ATPase activity of isolated CF1. Quercetin, affects also the proton uptake in chloroplasts in a manner similar to that of dicyclohexylcarbodiimide. 2. The light-dependent proton uptake in EDTA-treated chloroplasts is stimulated by quercetin. In untreated chloroplasts quercetin has a dual effect: it enhances at pH above 7.5 while at lower pH values it decreases the extent of H+ uptake. Similar effects were obtained with dicyclohexylcarbodiimide. 3. Like quercetin, dicyclohexylcarbodiimide was also found to inhibit the ATPase activity of isolated CF1. 4. Quercetin inhibits uncoupled electron transport induced by either EDTA-treatment of chloroplasts or by addition of uncouplers. Quercetin restores H+ uptake in both types of uncoupled chloroplasts. 5. The mode of action of quercetin and dicyclohexylcarbodiimide in photophosphorylation is discussed, and interaction with both CF1 and F0 is suggested. 相似文献