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1.
A chicken pectoralis muscle membrane fraction enriched in a Mg2+- or Ca2+-activated (‘basic’) ATPase was obtained by sucrose gradient centrifugation. Enzymatic properties of the ‘basic’ ATPase were determined and used to localize its enzymatic activity in situ by ultrastructural cytochemistry. The enzyme was activated by Mg2+ or Ca2+ but not by Sr2+, Ba2+, Co2+, Ni2+ or Pb2+. It was present in a membranous fraction with a buoyant density of 1.10-1.12 (24–27.5% (w/w) sucrose). ‘Basic’ ATPase activity had a sedimentation pattern similar to the putative plasma membrane enzymes, 5′-nucleotidase and leucyl β-naphthylamidase, but different from that of sarcoplasmic reticulum Ca2+ ATPase. Also unlike sarcoplasmic reticulum Ca2+ ATPase, ‘basic’ ATPase was resistant to N-ethylmaleimide and aldehyde fixatives, was active in a medium containing a high Ca2+ concentration (3 mM), and was lost when exposed to Triton X-100 or deoxycholate. In cytochemical studies, a low Pb2+ concentration was used to capture the enzymatically released phosphate ions. Under conditions which eliminated interfering (Na+ + K+) ATPase and sarcoplasmic reticulum Ca2+ ATPase activities, electron-dense lead precipitates were present at the plasmalemma and T-system membranes. These studies suggest that ‘basic’ ATPase activity is associated with plasmalemma and T-system membranes of skeletal muscle. 相似文献
2.
[Na] in the subsarcolemmal ‘fuzzy’ space and modulation of [Ca]i and contraction in cardiac myocytes
The strength of the heart beat depends on the amplitude and time course of the transient increase in [Ca2+] in the myocytes with each cycle. [Na+]i modulates cardiac contraction through its effect on the Ca2+ flux through the Na/Ca exchanger. Cardiac excitation–contraction coupling has been postulated to occur in a microdomain or ‘fuzzy’ space at the junction of the T-tubules and the sarcoplasmic reticulum. This ‘fuzzy’ space is well described for the Ca2+ fluxes and the interaction between the L-type Ca2+ channel, the Ca2+ release channel of the sarcoplasmic reticulum and the Na/Ca exchanger. Co-localization of the Na+ transporters, in particular the Na/K pump and the Na+ channel, within this ‘fuzzy’ space is not as well established. The functional and morphological characteristics of the ‘fuzzy’ space for Na+ and its interaction with the Ca2+ handling suggest that this space is not strictly co-inciding with the Ca2+ microdomain. In this space [Na+] can be several-fold higher or lower than [Na+] in the bulk cytosol. This has implications for modulation of [Ca2+]i during a single beat as well as during alterations in Na+ fluxes seen in pathological conditions. 相似文献
3.
Mounting evidence shows that neuronal death is an important and essential component of brain tissue homeostasis, with two major forms of cell death occurring: necrosis and apoptosis. No general consensus exists as to whether these two forms of neuronal death represent separate cellular processes or just two different forms of a common ‘death pathway’. One difference between them is the role played by intracellular Ca2+: central and obligatory, in necrosis and possible, but not always necessary in triggering apoptosis. Furthermore, the same assessment of the involvement of Ca2+ signalling could also distinguish between two poss ible apoptotic states in the nervous system: one, the ‘developmental apoptosis’, involving immature and developing neurons, in which Ca2+ plays mainly an apoprotector role, and another one, associated mainly with pathological instances and involving fully matured and established neurons, in which Ca2+ plays an apo-inducing role. 相似文献
4.
Cardiac cellular calcium (Ca2+) handling is the well-investigated mediator of excitation–contraction coupling, the process that translates cardiac electrical activation into mechanical events. The reverse—effects of mechanical stimulation on cardiomyocyte Ca2+ handling—are much less well understood, in particular during the inter-beat period, called ‘diastole’. We have investigated the effects of diastolic length changes, applied axially using a pair of carbon fibres attached to opposite ends of Guinea pig isolated ventricular myocytes, on the availability of Ca2+ in the main cellular stores (the sarcoplasmic reticulum; SR), by studying the rest-decay of SR Ca2+ content [Ca2+]SR, and the reloading of the SR after prior depletion of Ca2+ from the cell.Cells were loaded with Fura-2 AM (an indicator of the cytosolic ‘free’ Ca2+ concentration, [Ca2+]i), and pre-conditioned by field-stimulation (2 Hz) at 37 °C, while [Ca2+]i transients and sarcomere length (SL) were recorded simultaneously. After reaching a steady state in the behaviour of observed parameters, stimulation was interrupted for between 5 and 60 s, while cells were either held at resting length, or stretched (controlled to cause a 10% increase in SL, to aid inter-individual comparison). Thereafter, each cell was returned to its original resting length, followed by swift administration of 10 mM of caffeine (in Na+/Ca2+-free solution), which causes the release of Ca2+ from the SR (caffeine), but largely prevents extrusion of Ca2+ from the cytosol to the cell exterior (Na+/Ca2+-free solution). By comparing the [Ca2+]i in cells exposed/not exposed to diastolic stretch of different duration, we assessed the rest-decay dynamics of [Ca2+]SR. To assess SR reloading after initial Ca2+ depletion, the same stretch protocol was implemented after prior emptying of the cell by application of 10 mM of caffeine in normal Tyrode solution (which causes Ca2+ to be released from the SR and extruded from the cell via the Na+/Ca2+ exchanger; NCX).Axial stretch enhanced the rate of both rest-decay and reloading of [Ca2+]SR. Application of 40 μM streptomycin, a blocker of stretch-activated ion channels, did not affect the stretch-induced increase in SR reloading. This behaviour was reproduced in a computer simulation study, using a modified version of the 2006 Iribe–Kohl–Noble model of single cardiac myocyte Ca2+ handling, suggesting that stretch increases both Ca2+ leak from the SR and Ca2+ influx via the sarcolemma. This may have important implications for the mobilisation of Ca2+ in stretched cells, and could contribute to the regional ‘matching’ of individual cardiomyocyte contractility to dynamic, and regionally varying, changes in mechanical loads, such as diastolic pre-load, of cardiac tissue. 相似文献
5.
We have examined intracellular calcium buffer capacity of cytoplasm from the giant axon of the marine invertebrate Myxicola infundibulum by photolytically releasing calcium from ‘caged’ compounds, while monitoring free calcium, [Ca2+], with Ca-sensing electrodes. In cytoplasm containing intact organelles, two features of the [Ca2+] response were seen upon light exposure: an initial spike from basal [Ca2+], followed by a slower phase recovery. Both the amplitude of the spike in [Ca2+] and the recovery were reduced by removal of MgATP. If organelles were removed from the cytoplasm, light exposure caused only a step-like change in [Ca2+] with no recovery.Apparent buffer capacities (Δ bound Ca/Δ free Ca) were unaffected by changing pH from 7.0 to 7.5; however, raising basal free calcium above 3 μM significantly reduced this parameter. The buffer capacity measured after the initial spike varied by as much as an order of magnitude from one giant axon to another but averaged −50 in the absence and 100 in the presence of 1 mM MgATP for [Ca2+] below 3 μM. 相似文献
6.
J. Nagel-Volkmann C. Plieth D. Becker H. Lüthen K. Drffling 《Journal of plant physiology》2009,166(17):1955-1960
Relatively little is known about changes in the cytosolic free calcium ion concentration ([Ca2+]c) in monocotyledonous plants. Therefore, we produced transgenic winter wheat lines stably expressing the calcium-sensitive photoprotein aequorin constitutively in the cytosol. [Ca2+]c was detected in vivo by luminometry, and [Ca2+]c elevations were imaged at video rate. Experiments with the transgenic seedlings focused on potential changes in [Ca2+]c during cold exposure. Temperature-induced changes in [Ca2+]c were found to be more dependent on the change in temperature (dT dt−1) than on the absolute value of temperature. [Ca2+]c increased only at cooling rates higher than 8°C min−1, indicating that an overall cellular [Ca2+]c increase is of minor relevance as a signal for cold acclimation in wheat under ecological conditions. The results are discussed with regard to the so-called ‘calcium signature hypothesis’. 相似文献
7.
The existence of two types of binding sites for ouabain in human erythrocyte membranes is described. Receptor sites designated as ‘type I’, which may be identical to the K+-insensitive sites of intact cells, were detected at concentrations of ouabain as low as 10−7 M. The ‘type II’ receptor sites require the inclusion of Mg2+ + Pi to form complexes with ouabain; they may be identical to the K+-sensitive sites of intact cells. These sites were saturated at approx. 5 · 10−7 M ouabain but could not be detected at higher concentrations. The range of ouabain concentrations at which ‘type I’ receptors start to predominate (i.e. 5 · 10−8–5 · 10−7 M) was termed ‘critical digitalis concentrations’. The process of binding reached equilibrium within 1 and 4 h for ‘type I’ and ‘type II’ sites, respectively. The dissociation constant for ‘type II’ receptor-ouabain complexes was 7.6 · 10−9 M.Under similar experimental conditions, rat erythrocyte membranes exhibited only non-saturable sites.Alterations in the proportions of the two types of receptors were demonstrated by preincubation of the membranes, in the presence or absence of Mg2+ + Pi, prior to the addition of ouabain. In the first case, ‘type II receptor-ouabain’ complexes were stabilized at about 50% of the untreated membranes and ‘type I-ouabain’ complexes slowly approached equilibrium over a period of 24 h. In the latter instance, ‘type I’ receptors were not detected, and only ‘type II-ouabain’ complexes prevailed. 相似文献
8.
The particulate fraction from osmotically shocked synaptosomes (‘synaptosomal membranes’) sequesters Ca when incubated with ATP-containing solutions. This net accumulation of Ca can reduce the free [Ca2+] of the bathing medium to sub-micromolar levels (measured with arsenazo III). Two distinct types of Ca sequestration site are responsible for the Ca2+ buffering. One site, presumed to be smooth endoplasmic reticulum, operates at low [Ca2+] (less than 1 μM), and has a relatively small capacity. Ca sequestration at this site is prevented by the Ca2+ ionophore, A-23187, but not by mitochondrial poisons. The second (mitochondrial) site, in contrast, is blocked by the mitochondrial uncoupler, carbonyl cyanide p-trifluoromethoxyphenylhydrazone, and oligomycin. Since the intraterminal organelles can buffer [Ca2+] to about 0.3–0.5 μM, this may be an upper limit to the normal resting level of [Ca2+]i in nerve terminals. In the steady state, total cell Ca and [Ca2+]i will be governed principally by Ca transport mechanisms in the plasmalemma; the intracellular organelle transport systems then operate in equilibrium with this [Ca2+]. During activity, however, Ca rapidly enters the terminals and [Ca2+]i rises. The intracellular buffering mechanisms then come into play and help to return [Ca2+]i toward the resting level; the non-mitochondrial Ca sequestration mechanism probably plays the major role in this Ca buffering. 相似文献
9.
Consideration of the principal current models for agonist-induced activation of Ca2+ entry in electrically non-excitable cells suggests that it may be possible to distinguish between them on the basis of predicted differences in the temporal relationship(s) between intracellular Ca2+ release and the activation of Ca2+ entry. Measurements of changes in [Ca2+]i and Mn2+ quench in individual exocrine cells from the avian nasal gland indicate that, whereas Ins(1,4,5)P3-induced release of intracellular Ca2+ occurs within 3–5 s, the increase in Mn2+ quench is delayed by some 20–30 s. Mn2+ quench rate is similarly increased by thapsigargin, and is blocked by SK&F 96365, indicating that the increased Mn2+ quench observed genuinely reflects agonist-enhanced activity of the divalent cation entry pathway normally traversed by Ca2+. Additional experiments indicate that the observed delay is not due to inhibition of this pathway by elevated [Ca2+]i. Furthermore, the delay cannot be explained by the time required for Ins(1,3,4,5)P4 generation, which is essentially maximal within 10 s of agonist addition. It is concluded that the observed delay in the activation of the Ca2+ entry pathway is best explained by ‘capacitative’ models where increased entry requires the generation, and transmission to the plasma membrane, of an unknown messenger as a direct result of the depletion of intracellular Ca2+ stores. 相似文献
10.
We studied the effect of extracellular Ca2+ concentration ([Ca2+]e) on adipocyte differentiation. Preadipocytes exposed to continuous [Ca2+]e higher than 2.5 mmol/l accumulated little or no cytoplasmic lipid compared to controls in 1.8 mmol/l [Ca2+]e. Differentiation was monitored by Oil Red O staining of cytoplasmic lipid and triglyceride assay of accumulated lipid, by RT-PCR analysis of adipogenic markers, and by the activity of glycerol-3-phosphate dehydrogenase (GPDH). Elevated [Ca2+]e inhibited expression of peroxisome proliferator-activated receptor γ, CCAAT/enhancer binding protein α, and steroid regulatory binding element protein. High [Ca2+]e significantly inhibited differentiation marker expression including adipocyte fatty acid binding protein, and GPDH. The decrease in Pref-1 expression that accompanied differentiation also was prevented by high [Ca2+]e. Treatment of 3T3-L1 cells with high [Ca2+]e did not significantly affect cell number or viability and did not trigger apoptosis. Levels of intracellular Ca+2 remained unchanged in various [Ca2+]e. Treatment of 3T3-L1 with pertussis toxin (PTX) partially restored lipid accumulation and increased differentiation markers in cells treated with 5 mmol/l [Ca2+]e. ‘Classical’ parathyroid cell Ca2+ sensing receptors (CaSR) were not detected either by RT-PCR or by Western blotting. These results suggest that continuos exposure to high [Ca2+]e inhibits preadipocyte differentiation and that this may involve a G-protein-coupled mechanism mediated by a novel Ca2+ sensor or receptor. 相似文献
11.
Agonist-induced oscillations in the concentration of intracellular free calcium ([Ca2+]1) display a wide variety of temporal and spatial patterns. In non-excitable cells, typical oscillatory patterns are somewhat cell-type specific and range from frequency-encoded, repetitive Ca2+ spikes to oscillations that are more sinusoidal in shape. Although the response of a cell population, even to the same stimulus, is often extremely heterogeneous, the response of the same cell to successive exposures can be remarkably similar. We propose that such ‘Ca 2+ fingerprints’ can be a consequence of cell-specific morphological properties. The hypothesis is tested by means of a stochastic computer simulation of a two-dimensional model for oscillatory Ca 2+ waves which encompasses the basic elements of the two-pool oscillator introduced by Goldbeter et al. (Goldbeter A., Dupont G., Berridge M.J. Minimal model for signal-induced Ca2+-oscillations and for their frequency encoding through protein phosphorylation. Proc Natl Acad Sci USA 1990; 87: 1461–1465). In the framework of our extended spatiotemporal model, single cells can display various oscillation patterns which depend on the agonist dose, Ca2+ diffusibility, and several morphological parameters. These are, for example, size and shape of the cell and the cell nucleus, the amount and distribution of Ca2+ stores, and the subcellular location of the inositol(1,4,5)-trisphosphate-generating apparatus. 相似文献
12.
Transient receptor potential melastatin 2 (TRPM2) is a thermosensitive Ca2+-permeable cation channel expressed by pancreatic β cells where channel function is constantly affected by body temperature. We focused on the physiological functions of redox signal-mediated TRPM2 activity at body temperature. H2O2, an important molecule in redox signaling, reduced the temperature threshold for TRPM2 activation in pancreatic β cells of WT mice but not in TRPM2KO cells. TRPM2-mediated [Ca2+]i increases were likely caused by Ca2+ influx through the plasma membrane because the responses were abolished in the absence of extracellular Ca2+. In addition, TRPM2 activation downstream from the redox signal plus glucose stimulation enhanced glucose-induced insulin secretion. H2O2 application at 37 °C induced [Ca2+]i increases not only in WT but also in TRPM2KO β cells. This was likely due to the effect of H2O2 on KATP channel activity. However, the N-acetylcysteine-sensitive fraction of insulin secretion by WT islets was increased by temperature elevation, and this temperature-dependent enhancement was diminished significantly in TRPM2KO islets. These data suggest that endogenous redox signals in pancreatic β cells elevate insulin secretion via TRPM2 sensitization and activity at body temperature. The results in this study could provide new therapeutic approaches for the regulation of diabetic conditions by focusing on the physiological function of TRPM2 and redox signals. 相似文献
13.
Nutrient-induced increases in intracellular free Ca2+ concentrations are the key trigger for insulin release from pancreatic islet β-cells. These Ca2+ changes are tightly regulated temporally, occurring as Ca2+ influx-dependent baseline oscillations. We explore here the concept that locally high [Ca2+] concentrations (i.e. Ca2+ microdomains) may control exocytosis via the recruitment of key effector proteins to sites of exocytosis. Importantly, recent advances in the development of organelle- and membrane-targeted green fluorescent protein (GFP-) or aequorin-based Ca2+ indicators, as well as in rapid imaging techniques, are providing new insights into the potential role of these Ca2+ microdomains in β-cells. We summarise here some of the evidence indicating that Ca2+ microdomains beneath the plasma membrane and at the surface of large dense core vesicles may be important in the normal regulation of insulin secretion, and may conceivably contribute to “ATP-sensitive K+-channel independent” effects of glucose. We also discuss evidence that, in contrast to certain non-excitable cells, direct transfer of Ca2+ from the ER to mitochondria via localised physical contacts between these organelles is relatively less important for efficient mitochondrial Ca2+ uptake in β-cells. Finally, we discuss evidence from single cell imaging that increases in cytosolic Ca2+ are not required for the upstroke of oscillations in mitochondrial redox state, but may underlie the reoxidation process. 相似文献
14.
A. Gamberucci R. Fulceri P. Tarroni R. Giunti P. Marcolongo V. Sorrentino A. Benedetti 《Cell calcium》1995,17(6)
To investigate the presence and the size of different non-mitochondria) Ca2+ pools of Ehrlich ascites tumor cells (EATCs) , digitonin-permeabilized cells were allowed to accumulate Ca2+ in the presence of mitochondrial inhibitors and treated with the reticular Ca2+-ATPase inhibitor thapsigargin, IP3 and the Ca2+ ionophore A23187. Emptying of thapsigargin-sensitive Ca2+ stores prevented any Ca2+ release by IP3, and, after IP3 addition, little or no Ca2+ was released by thapsigargin. In both instances, a further Ca2+ release was accomplished by A23187. The IP3-thapsigargin-sensitive pool and the residual A23187-sensitive one corresponded to approximately 60 and 37% of non-mitochondria) stored Ca2+, respectively. In intact EATCs, IP3-dependent agonists and thapsigargin discharged Ca 2+ pools almost completely overlapping, and A32187 released a minor residual Ca2+ pool. The IP3-insensitive pool appeared to have a relatively low affinity for Ca2+ (below 600 nM). The high affinity, IP3-sensitive Ca2+ pool was discharged in a ‘quantal’ manner following step additions of sub maximal [IP3], and the IP3-induced fractional Ca2+ release was more marked at higher concentrations of stored (luminal) Ca2+, The IP3-sensitive Ca2+ pool appeared to be devoid of the Ca2+-activated Ca2+ release channel since caffeine did not released any Ca2+ in intact and permeabilized EATCs, and Western blot analyses of EATC microsomal membranes failed to detect any known ryanodine receptor isoform. 相似文献
15.
Calcium-activated potassium channels in liver cells 总被引:1,自引:0,他引:1
Activation of certain membrane receptors increases the concentration of Ca2+ in the cytosol of hepatocytes. Since in most species these cells possess a PK(Ca) mechanism, the outcome is a rise in PK. This can be blocked by quinine, apamin and certain neuromuscular blocking agents. The binding of labelled apamin to hepatocytes has been studied under physiological conditions, and the relationship between the binding sites and K+ channels is discussed. The physiological role of the PK(Ca) mechanism in hepatocytes is unclear, though it is largely responsible for ‘adrenaline hyperkalaemia’. 相似文献
16.
NADPH is a key reductant carrier that maintains internal redox and antioxidant status, and that links biosynthetic, catabolic and signalling pathways. Plants have a mitochondrial external NADPH oxidation pathway, which depends on Ca2+ and pH in vitro, but concentrations of Ca2+ needed are not known. We have determined the K0.5(Ca2+) of the external NADPH dehydrogenase from Solanum tuberosum mitochondria and membranes of E. coli expressing Arabidopsis thaliana NDB1 over the physiological pH range using O2 and decylubiquinone as electron acceptors. The K0.5(Ca2+) of NADPH oxidation was generally higher than for NADH oxidation, and unlike the latter, it depended on pH. At pH 7.5, K0.5(Ca2+) for NADPH oxidation was high (≈100 μM), yet 20-fold lower K0.5(Ca2+) values were determined at pH 6.8. Lower K0.5(Ca2+) values were observed with decylubiquinone than with O2 as terminal electron acceptor. NADPH oxidation responded to changes in Ca2+ concentrations more rapidly than NADH oxidation did. Thus, cytosolic acidification is an important activator of external NADPH oxidation, by decreasing the Ca2+-requirements for NDB1. The results are discussed in relation to the present knowledge on how whole cell NADPH redox homeostasis is affected in plants modified for the NDB1 gene. 相似文献
17.
T. J. Shuttleworth 《Cell calcium》1995,17(6)
It has been suggested that the release of Ca2+ from intracellular stores by inositol 1,4,5-trisphosphate (InsP3) is modulated by the luminal Ca2+ content of the stores and that such an effect could underlie the apparent ‘quantal’ nature of InsP3-induced release. Although initial studies failed to find evidence in support of such a modulation, several subsequent reports have indicated luminal Ca2+ effects that become apparent only after a greater than 70–75% depletion of Ca2+ stores. In these studies, Ca2+ release was expressed as a percentage of an A23187-releasable pool which comprised both InsP3-sensi-tive and InsP3-insensitive components. In model calculations we have found that the presence of even a minor InsP3-insensitive component in the total Ca2+ pool significantly distorts interpretation of the data. We show that the published results can be accurately duplicated without any requirement for a shift in the true InsP3 sensitivity of Ca2+ release if either: (a) the InsP3-insensitive component does not remain a constant proportion of the total pool during depletion (i.e. depletion disproportionally affects the InsP3-sensitive component); or (b) during generation of InsP3-response curves, additional Cal 2+ is released from the InsP3-insensitive component as the InsP3-sensitive component is progressively emptied. Examination indicates that either, or both, of these conditions apply in the published reports and we conclude that the demonstrated effects of luminal Ca2+ may be artifacts. 相似文献
18.
Can elevated CO(2) improve salt tolerance in olive trees? 总被引:2,自引:0,他引:2
We compared growth, leaf gas exchange characteristics, water relations, chlorophyll fluorescence, and Na+ and Cl− concentration of two cultivars (‘Koroneiki’ and ‘Picual’) of olive (Olea europaea L.) trees in response to high salinity (NaCl 100 mM) and elevated CO2 (eCO2) concentration (700 μL L−1). The cultivar ‘Koroneiki’ is considered to be more salt sensitive than the relatively salt-tolerant ‘Picual’. After 3 months of treatment, the 9-month-old cuttings of ‘Koroneiki’ had significantly greater shoot growth, and net CO2 assimilation (ACO2) at eCO2 than at ambient CO2, but this difference disappeared under salt stress. Growth and ACO2 of ‘Picual’ did not respond to eCO2 regardless of salinity treatment. Stomatal conductance (gs) and leaf transpiration were decreased at eCO2 such that leaf water use efficiency (WUE) increased in both cultivars regardless of saline treatment. Salt stress increased leaf Na+ and Cl− concentration, reduced growth and leaf osmotic potential, but increased leaf turgor compared with non-salinized control plants of both cultivars. Salinity decreased ACO2, gs, and WUE, but internal CO2 concentrations in the mesophyll were not affected. eCO2 increased the sensitivity of PSII and chlorophyll concentration to salinity. eCO2 did not affect leaf or root Na+ or Cl− concentrations in salt-tolerant ‘Picual’, but eCO2 decreased leaf and root Na+ concentration and root Cl− concentration in the more salt-sensitive ‘Koroneiki’. Na+ and Cl− accumulation was associated with the lower water use in ‘Koroneiki’ but not in ‘Picual’. Although eCO2 increased WUE in salinized leaves and decreased salt ion uptake in the relatively salt-tolerant ‘Koroneiki’, growth of these young olive trees was not affected by eCO2. 相似文献
19.
Donald A. McCarthy Ryan R. Clark Toni R. Bartling Mohamed Trebak J. Andres Melendez 《The Journal of biological chemistry》2013,288(45):32149-32159
Senescent cells accumulate in aged tissue and are causally linked to age-associated tissue degeneration. These non-dividing, metabolically active cells are highly secretory and alter tissue homeostasis, creating an environment conducive to metastatic disease progression. IL-1α is a key senescence-associated (SA) proinflammatory cytokine that acts as a critical upstream regulator of the SA secretory phenotype (SASP). We established that SA shifts in steady-state H2O2 and intracellular Ca2+ levels caused an increase in IL-1α expression and processing. The increase in intracellular Ca2+ promoted calpain activation and increased the proteolytic cleavage of IL-1α. Antioxidants and low oxygen tension prevented SA IL-1α expression and restricted expression of SASP components IL-6 and IL-8. Ca2+ chelation or calpain inhibition prevented SA processing of IL-1α and its ability to induce downstream cytokine expression. Conditioned medium from senescent cells treated with antioxidants or Ca2+ chelators or cultured in low oxygen markedly reduced the invasive capacity of proximal metastatic cancer cells. In this paracrine fashion, senescent cells promoted invasion by inducing an epithelial-mesenchymal transition, actin reorganization, and cellular polarization of neighboring cancer cells. Collectively, these findings demonstrate how SA alterations in the redox state and Ca2+ homeostasis modulate the inflammatory phenotype through the regulation of the SASP initiator IL-1α, creating a microenvironment permissive to tumor invasion. 相似文献
20.
β-cells in the pancreatic islet respond to elevated plasma glucose by secreting insulin to maintain glucose homeostasis. In addition to glucose stimulation, insulin secretion is modulated by numerous G-protein coupled receptors (GPCRs). The GPCR ligands Kisspeptin-10 (KP) and glucagon-like peptide-1 (GLP-1) potentiate insulin secretion through Gq and Gs-coupled receptors, respectively. Despite many studies, the signaling mechanisms by which KP and GLP-1 potentiate insulin release are not thoroughly understood. We investigated the downstream signaling pathways of these ligands and their affects on cellular redox potential, intracellular calcium activity ([Ca2+]i), and insulin secretion from β-cells within intact murine islets. In contrast to previous studies performed on single β-cells, neither KP nor GLP-1 affect [Ca2+]i upon stimulation with glucose. KP significantly increases the cellular redox potential, while no effect is observed with GLP-1, suggesting that KP and GLP-1 potentiate insulin secretion through different mechanisms. Co-treatment with KP and the Gβγ-subunit inhibitor gallein inhibits insulin secretion similar to that observed with gallein alone, while co-treatment with gallein and GLP-1 does not differ from GLP-1 alone. In contrast, co-treatment with the Gβγ activator mSIRK and either KP or GLP-1 stimulates insulin release similar to mSIRK alone. Neither gallein nor mSIRK alter [Ca2+]i activity in the presence of KP or GLP-1. These data suggest that KP likely alters insulin secretion through a Gβγ-dependent process that stimulates glucose metabolism without altering Ca2+ activity, while GLP-1 does so, at least partly, through a Gα-dependent pathway that is independent of both metabolism and Ca2+. 相似文献