Ethanol exerts dual effects on calcium homeostasis in CCK-8-stimulated mouse pancreatic acinar cells

Fernández-Sánchez, Marcela; Castillo-Vaquero, Angel del; Salido, Ginés M.; González, Antonio

doi:10.1186/1471-2121-10-77

Cited by 15 publications

(8 citation statements)

References 39 publications

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“…By itself, a concentration of 850 mM was required to evoke a small Ca 2ϩ transient (6). There is also evidence that ethanol either directly or through generation of reactive oxygen species causes Ca 2ϩ influx (35,36). However, several reports implicate a role for ethanol metabolites as the actual mediators of aberrant Ca 2ϩ signaling in acinar cells (34,37).…”

Section: Discussionmentioning

confidence: 99%

Ethanol Enhances Carbachol-induced Protease Activation and Accelerates Ca2+ Waves in Isolated Rat Pancreatic Acini

Orabi¹,

Shah²,

Muili

et al. 2011

Journal of Biological Chemistry

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Alcohol abuse is a leading cause of pancreatitis, accounting for 30% of acute cases and 70 -90% of chronic cases, yet the mechanisms leading to alcohol-associated pancreatic injury are unclear. An early and critical feature of pancreatitis is the aberrant signaling of Ca 2؉ within the pancreatic acinar cell. An important conductor of this Ca 2؉ is the basolaterally localized, intracellular Ca 2؉ channel ryanodine receptor (RYR). In this study, we examined the effect of ethanol on mediating both pathologic intra-acinar protease activation, a precursor to pancreatitis, as well as RYR Ca 2؉ signals. We hypothesized that ethanol sensitizes the acinar cell to protease activation by modulating RYR Ca 2؉ . Acinar cells were freshly isolated from rat, pretreated with ethanol, and stimulated with the muscarinic agonist carbachol (1 M). Ethanol caused a doubling in the carbachol-induced activation of the proteases trypsin and chymotrypsin (p < 0.02). The RYR inhibitor dantrolene abrogated the enhancement of trypsin and chymotrypsin activity by ethanol (p < 0.005 for both proteases). Further, ethanol accelerated the speed of the apical to basolateral Ca 2؉ wave from 9 to 18 m/s (p < 0.0005; n ‫؍‬ 18 -22 cells/group); an increase in Ca 2؉ wave speed was also observed with a change from physiologic concentrations of carbachol (1 M) to a supraphysiologic concentration (1 mM) that leads to protease activation. Dantrolene abrogated the ethanol-induced acceleration of wave speed (p < 0.05; n ‫؍‬ 10 -16 cells/group). Our results suggest that the enhancement of pathologic protease activation by ethanol is dependent on the RYR and that a novel mechanism for this enhancement may involve RYR-mediated acceleration of Ca 2؉ waves.Pancreatitis is a life-threatening inflammatory disorder of the pancreas that leads to more than 30,000 deaths per year (1). Alcohol-associated pancreatitis accounts for 30% of acute cases and 70 -90% of chronic cases. Further, alcoholic pancreatitis carries the highest mortality rate among all etiologies (2). However, the mechanisms by which ethanol mediates pathology are largely unknown.Alcohol can exert diverse effects on the pancreas. Ethanol exposure has been linked to abnormal blood flow, leading to ischemic changes, and increased sphincter of Oddi dysfunction, resulting in pancreatic duct hypertension (2).In addition, ethanol appears to directly predispose the acinar cell to pathological changes including oxidant stress (3), membrane fragility (4), mitochondrial uncoupling (5, 6), and basolateral exocytosis (7). Several lines of evidence also link ethanol to aberrant Ca 2ϩ 3 signaling (6). High amplitude, aberrant, intracellular Ca 2ϩ waves that propagate from the apical to basolateral region of the acinar cell predispose to early features of pancreatitis, particularly intra-acinar protease activation (8, 9). We know that intracellular Ca 2ϩ release is responsible for the onset of this aberrant Ca 2ϩ signal (10). In addition, we have previously shown that the ryanodine receptor (RYR), 4 a major intracel...

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Section: Discussionmentioning

confidence: 99%

Ethanol Enhances Carbachol-induced Protease Activation and Accelerates Ca2+ Waves in Isolated Rat Pancreatic Acini

Orabi¹,

Shah²,

Muili

et al. 2011

Journal of Biological Chemistry

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“…These patterns of changes in [Ca 2? ] c have been previously observed in both tumor and non-tumor pancreatic cells (Del Castillo-Vaquero et al 2010;Fernandez-Sanchez et al 2009;Gonzalez et al 1997).…”

Section: Analysis Of Intracellular Free-camentioning

confidence: 70%

Interferences of resveratrol with fura-2-derived fluorescence in intracellular free-Ca2+ concentration determinations

2015

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Resveratrol (3,4 0 ,5-trihydroxy-trans-stilbene) is an antioxidant widely employed in cell physiology studies. It has been reported that it interferes with fura-2-derived fluorescence, making the employment of this dye nonviable. In this work, the interference of resveratrol with fura-2 determinations of intracellular free-Ca 2? concentration ([Ca 2? ] c ) was examined. Solutions containing different concentrations of resveratrol, with or without fura-2, in the presence or in the absence of Ca 2? , were analyzed by spectrofluorimetry. AR42J tumor cells were employed to study the influence of resveratrol on fura-2 fluorescence in living cells, by single cell fluorimetry. Resveratrol impaired the detection of fura-2-fluorescence emission (510 nm) at the 340, 360 and 380 nm excitation wavelengths. Resveratrol emitted fluorescence at 510 nm when lighted at all three excitation wavelengths. In addition, resveratrol emitted fluorescence at 380 nm when excited at 340 nm. Our observations suggest that the employment of the ratiometric properties of fura-2 to follow changes in [Ca 2? ] c in the presence of resveratrol is not viable. However, we think that the 380 nm excitation light could be employed. Results could be expressed as F 0 /F 380 , where F 0 is the resting fluorescence and F 380 is the value of fluoresce at a certain time point. We could follow changes in [Ca 2? ] c evoked by CCK-8, and we also detected Ca 2? mobilization by 100 lM resveratrol in AR42J cells. This investigation presents evidence demonstrating that resveratrol interferes with fura-2 fluorescence spectra. Nevertheless, a chance still exists if the 380 nm excitation wavelength is employed in the middle or low micromolar concentrations of resveratrol.

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“…Specifically, it has been shown that after treatment of isolated pancreatic acinar cells either with EtOH or its non-oxidative metabolites, physiological concentrations of secretagogue induces sustained increases in [Ca 2ϩ ] cyto that are no longer limited to the apical pole but spread throughout the acinar cell (28,29). Associated with the prolonged aberrant Ca 2ϩ signal is an inhibition of apical zymogen secretion (30), impaired mitochondrial function (i.e.…”

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confidence: 99%

Raf-1 Kinase Inhibitory Protein (RKIP) Mediates Ethanol-induced Sensitization of Secretagogue Signaling in Pancreatic Acinar Cells

Kim

Ives

Wang

et al. 2012

Journal of Biological Chemistry

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Background:The molecular mechanism by which ethanol (EtOH) sensitizes pancreatic acinar cells to secretagogue stimulation is largely undefined. Results: Ethanol stimulates PKC-dependent RKIP phosphorylation, and RKIP ablation prevents EtOH-induced sensitization of secretagogue Ca 2ϩ signaling and aberrant chymotrypsin activation. Conclusion: RKIP mediates the cytotoxic effects of EtOH on pancreatic acinar cells. Significance: Modulation of RKIP expression may have therapeutic utility in the prevention or treatment of alcohol-associated pancreatitis.

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Ethanol exerts dual effects on calcium homeostasis in CCK-8-stimulated mouse pancreatic acinar cells

Cited by 15 publications

References 39 publications

Ethanol Enhances Carbachol-induced Protease Activation and Accelerates Ca2+ Waves in Isolated Rat Pancreatic Acini

Ethanol Enhances Carbachol-induced Protease Activation and Accelerates Ca2+ Waves in Isolated Rat Pancreatic Acini

Interferences of resveratrol with fura-2-derived fluorescence in intracellular free-Ca2+ concentration determinations

Raf-1 Kinase Inhibitory Protein (RKIP) Mediates Ethanol-induced Sensitization of Secretagogue Signaling in Pancreatic Acinar Cells

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