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Introduction Acute pancreatitis (AP) is an inflammatory disease that originates in the exocrine pancreas, where inactive pancreatic proenzymes become prematurely activated inside the pancreatic acinar cells (PACs), digesting the pancreas [...], Acute pancreatitis (AP), a human disease in which the pancreas digests itself, has substantial mortality with no specific therapy. The major causes of AP are alcohol abuse and gallstone complications, but it also occurs as an important side effect of the standard asparaginase-based therapy for childhood acute lymphoblastic leukemia. Previous investigations into the mechanisms underlying pancreatic acinar cell death induced by alcohol metabolites, bile acids, or asparaginase indicated that loss of intracellular ATP generation is an important factor. We now report that, in isolated mouse pancreatic acinar cells or cell clusters, removal of extracellular glucose had little effect on this ATP loss, suggesting that glucose metabolism was severely inhibited under these conditions. Surprisingly, we show that replacing glucose with galactose prevented or markedly reduced the loss of ATP and any subsequent necrosis. Addition of pyruvate had a similar protective effect. We also studied the effect of galactose in vivo in mouse models of AP induced either by a combination of fatty acids and ethanol or asparaginase. In both cases, galactose markedly reduced acinar necrosis and inflammation. Based on these data, we suggest that galactose feeding may be used to protect against AP.

Here we describe a technique that allows us to visualize in real time the formation and dynamics (fusion, changes of shape, and translocation) of vacuoles in living cells. The technique involves infusion of a dextran-bound fluorescent probe into the cytosol of the cell via a patch pipette, using the whole-cell patch-clamp configuration. Experiments were conducted on pancreatic acinar ceils stimulated with supramaximal concentrations of cholecystokinin (CCK). The vacuoles, forming in the cytoplasm of the cell, were revealed as dark imprints on a bright fluorescence background, produced by the probe and visualized by confocal microscopy. A combination of two dextran-bound probes, one infused into the cytosol and the second added to the extracellular solution, was used to identify endocytic and nonendocytic vacuoles. The cytosolic dextran-bound probe was also used together with a Golgi indicator to illustrate the possibility of combining the probes and identifying the localization of vacuoles with respect to other cellular organelles in pancreatic acinar cells. Combinations of cytosollc dextran-bound probes with endoplasmic reticulum (ER) or mitochondrial probes were also used to simultaneously visualize vacuoles and corresponding organelles. We expect that the new technique will also be applicable and useful for studies of vacuole dynamics in other cell types. pancreatic acinar cells; confocal microscopy; patch clamp; cholecystokinin

Cell-death programs executed in the pancreas under pathological conditions remain largely undetermined, although the severity of experimental pancreatitis has been found to depend on the ratio of apoptosis to necrosis. We have defined mechanisms by which apoptosis is induced in pancreatic acinar cells by the oxidant stressor menadione. Real-time monitoring of initiator caspase activity showed that caspase-9 (66% of cells) and caspase-8 (15% of cells) were activated within 30 min of menadione administration, but no activation of caspase-2, -10, or -12 was detected. Interestingly, when caspase-9 activation was inhibited, activation of caspase-8 was increased. Half-maximum activation ([t.sub.0.5]) of caspase-9 occurred within ~2 min and was identified at or in close proximity to mitochondria, whereas [t.sub.0.5] for caspase-8 occurred within ~26 min of menadione application and was distributed homogeneously throughout cells. Caspase-9 but not caspase-8 activation was blocked completely by the calcium chelator BAPTA or bongkrekic acid, an inhibitor of the mitochondrial permeability transition pore. In contrast, caspase-8 but not caspase-9 activation was blocked by the destruction of lysosomes (preincubation with Gly-Phe [beta]-naphthylamide, a cathepsin C substrate), loss of lysosomal acidity (bafilomycin A1), or inhibition of cathepsin L or D. Using pepstatin A-BODIPY FL conjugate, we confirmed translocation of cathepsin D out of lysosomes in response to menadione. We conclude that the oxidative stressor menadione induces two independent apoptotic pathways within pancreatic acinar cells: the classical mitochondrial calcium-dependent pathway that is initiated rapidly in the majority of cells, and a slower, caspase-8-mediated pathway that depends on the lysosomal activities of cathepsins and is used when the caspase-9 pathway is disabled. pancreas; acinar cells; menadione doi:10.1152/ajpgi.00103.2007

The S protein subunit 1 (S1) of SARS-CoV-2 is known to be responsible for the binding of the virus to host cell receptors, but the initial intracellular signalling steps following receptor activation of cells in the exocrine pancreas are unknown. Using an intact live mouse pancreatic lobule preparation, we observed that S1 elicited Ca2+ signals in stellate cells and macrophages, but not in the dominant acinar cells. The Ca2+ signals occurred mostly in the form of repetitive Ca2+ spikes. The probability of observing Ca2+ signals depended on the S1 concentration. The threshold was close to 70 nM, whereas at 600 nM, all cells responded. The SARS-Cov-2 nucleocapsid protein did not elicit any Ca2+ signals in any of the three cell types tested. The S1-induced Ca2+ signals in stellate cells started much faster (122 ± 37s) than those in macrophages (468 ± 68s). Furthermore, the interleukin-18 binding protein (IL-18BP) abolished the responses in macrophages without affecting the Ca2+ signals in stellate cells. The S1-elicited Ca2+ signals were completely dependent on the presence of external Ca2+ and were abolished by a selective inhibitor (CM4620) of Orai1 Ca2+ Release Activated Ca2+ channels. SARS-CoV-2 may contribute to acute pancreatitis, an often fatal inflammatory human disease. The S1-elicited Ca2+ signals we have observed in the pancreatic stellate cells and endogenous macrophages may play an important part in the development of the inflammatory process. [ABSTRACT FROM AUTHOR]

The article explores the intricate relationship between diabetes mellitus (DM) and pancreatic function, focusing on the impact of type 1 diabetes (T1DM) on pancreatic ductal function. It highlights how T1DM affects both the endocrine and exocrine functions of the pancreas, leading to conditions like exocrine pancreatic insufficiency and acute pancreatitis. The study by Ébert et al. challenges conventional views by showing that ductal secretion is affected at the initial stage of DM, emphasizing increased ductal secretion mainly due to increased CFTR activity. The authors suggest that ductal HCO3− secretion could serve as a potential therapeutic target for the prevention or treatment of DM, calling for further research in this area. [Extracted from the article]

This review deals with the roles of calcium ions and ATP in the control of the normal functions of the different cell types in the exocrine pancreas as well as the roles of these molecules in the pathophysiology of acute pancreatitis. Repetitive rises in the local cytosolic calcium ion concentration in the apical part of the acinar cells not only activate exocytosis but also, via an increase in the intramitochondrial calcium ion concentration, stimulate the ATP formation that is needed to fuel the energy-requiring secretion process. However, intracellular calcium overload, resulting in a global sustained elevation of the cytosolic calcium ion concentration, has the opposite effect of decreasing mitochondrial ATP production, and this initiates processes that lead to necrosis. In the last few years it has become possible to image calcium signaling events simultaneously in acinar, stellate, and immune cells in intact lobules of the exocrine pancreas. This has disclosed processes by which these cells interact with each other, particularly in relation to the initiation and development of acute pancreatitis. By unraveling the molecular mechanisms underlying this disease, several promising therapeutic intervention sites have been identified. This provides hope that we may soon be able to effectively treat this often fatal disease. [ABSTRACT FROM AUTHOR]

This article discusses the role of calcium signaling and ATP production in pancreatic cancer. Pancreatic cancer is a significant health burden and has a low survival rate. Specific gene mutations, as well as risk factors such as aging, obesity, and inflammatory diseases, contribute to the development of pancreatic cancer. The study mentioned in the article investigates calcium signaling and ATP levels in pancreatic acinar cells from a genetic model of pancreatic cancer. The results suggest that aberrant calcium signaling and metabolic changes play a crucial role in pancreatic cancer progression. Further research in these areas could lead to the identification of new therapeutic targets for this deadly disease. [Extracted from the article]

Background and Purpose: Many cancer cells depend on anti-apoptotic B-cell lymphoma 2 (Bcl-2) proteins for their survival. Bcl-2 antagonism through Bcl-2 homology 3 (BH3) mimetics has emerged as a novel anti-cancer therapy. ABT-199 (Venetoclax), a recently developed BH3 mimetic that selectively inhibits Bcl-2, was introduced into the clinic for treatment of relapsed chronic lymphocytic leukaemia. Early generations of Bcl-2 inhibitors evoked sustained Ca2+ responses in pancreatic acinar cells (PACs) inducing cell death. Therefore, BH3 mimetics could potentially be toxic for the pancreas when used to treat cancer. Although ABT-199 was shown to kill Bcl-2-dependent cancer cells without affecting intracellular Ca2+ signalling, its effects on PACs have not yet been determined. Hence, it is essential and timely to assess whether this recently approved anti-leukaemic drug might potentially have pancreatotoxic effects.Experimental Approach: Single-cell Ca2+ measurements and cell death analysis were performed on isolated mouse PACs.Key Results: Inhibition of Bcl-2 via ABT-199 did not elicit intracellular Ca2+ signalling on its own or potentiate Ca2+ signalling induced by physiological/pathophysiological stimuli in PACs. Although ABT-199 did not affect cell death in PACs, under conditions that killed ABT-199-sensitive cancer cells, cytosolic Ca2+ extrusion was slightly enhanced in the presence of ABT-199. In contrast, inhibition of Bcl-xL potentiated pathophysiological Ca2+ responses in PACs, without exacerbating cell death.Conclusion and Implications: Our results demonstrate that apart from having a modest effect on cytosolic Ca2+ extrusion, ABT-199 does not substantially alter intracellular Ca2+ homeostasis in normal PACs and should be safe for the pancreas during cancer treatment.Linked Articles: This article is part of a themed section on Mitochondrial Pharmacology: Featured Mechanisms and Approaches for Therapy Translation. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v176.22/issuetoc. [ABSTRACT FROM AUTHOR]

Key points: Ca2+ signalling in different cell types in exocrine pancreatic lobules was monitored simultaneously and signalling responses to various stimuli were directly compared. Ca2+ signals evoked by K+‐induced depolarization were recorded from pancreatic nerve cells. Nerve cell stimulation evoked Ca2+ signals in acinar but not in stellate cells. Stellate cells are not electrically excitable as they, like acinar cells, did not generate Ca2+ signals in response to membrane depolarization. The responsiveness of the stellate cells to bradykinin was markedly reduced in experimental alcohol‐related acute pancreatitis, but they became sensitive to stimulation with trypsin. Our results provide fresh evidence for an important role of stellate cells in acute pancreatitis. They seem to be a critical element in a vicious circle promoting necrotic acinar cell death. Initial trypsin release from a few dying acinar cells generates Ca2+ signals in the stellate cells, which then in turn damage more acinar cells causing further trypsin liberation. Abstract: Physiological Ca2+ signals in pancreatic acinar cells control fluid and enzyme secretion, whereas excessive Ca2+ signals induced by pathological agents induce destructive processes leading to acute pancreatitis. Ca2+ signals in the peri‐acinar stellate cells may also play a role in the development of acute pancreatitis. In this study, we explored Ca2+ signalling in the different cell types in the acinar environment of the pancreatic tissue. We have, for the first time, recorded depolarization‐evoked Ca2+ signals in pancreatic nerves and shown that whereas acinar cells receive a functional cholinergic innervation, there is no evidence for functional innervation of the stellate cells. The stellate, like the acinar, cells are not electrically excitable as they do not generate Ca2+ signals in response to membrane depolarization. The principal agent evoking Ca2+ signals in the stellate cells is bradykinin, but in experimental alcohol‐related acute pancreatitis, these cells become much less responsive to bradykinin and then acquire sensitivity to trypsin. Our new findings have implications for our understanding of the development of acute pancreatitis and we propose a scheme in which Ca2+ signals in stellate cells provide an amplification loop promoting acinar cell death. Initial release of the proteases kallikrein and trypsin from dying acinar cells can, via bradykinin generation and protease‐activated receptors, induce Ca2+ signals in stellate cells which can then, possibly via nitric oxide generation, damage more acinar cells and thereby cause additional release of proteases, generating a vicious circle. [ABSTRACT FROM AUTHOR]