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[This corrects the article DOI: 10.3389/fimmu.2024.1353695.].
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Objectives: This study aimed to analyze active compounds and signaling pathways of CH applying network pharmacology methods, and to additionally verify the molecular mechanism of CH in treating AP. Materials and methods: Network pharmacology and molecular docking were firstly used to identify the active components of CH and its potential targets in the treatment of AP. The pancreaticobiliary duct was retrogradely injected with sodium taurocholate (3.5%) to create an acute pancreatitis (AP) model in rats. Histological examination, enzyme-linked immunosorbent assay, Western blot and TUNEL staining were used to determine the pathway and mechanism of action of CH in AP. Results: Network pharmacological analysis identified 168 active compounds and 276 target proteins. In addition, there were 2060 targets associated with AP, and CH had 177 targets in common with AP. These shared targets, including STAT3, IL6, MYC, CDKN1A, AKT1, MAPK1, MAPK3, MAPK14, HSP90AA1, HIF1A, ESR1, TP53, FOS, and RELA, were recognized as core targets. Furthermore, we filtered out 5252 entries from the Gene Ontology(GO) and 186 signaling pathways from the Kyoto Encyclopedia of Genes and Genomes(KEGG). Enrichment and network analyses of protein-protein interactions predicted that CH significantly affected the PI3K/AKT signaling pathway, which played a critical role in programmed cell death. The core components and key targets showed strong binding activity based on molecular docking results. Subsequently, experimental validation demonstrated that CH inhibited the phosphorylation of PI3K and AKT in pancreatic tissues, promoted the apoptosis of pancreatic acinar cells, and further alleviated inflammation and histopathological damage to the pancreas in AP rats. Conclusion: Apoptosis of pancreatic acinar cells can be enhanced and the inflammatory response can be reduced through the modulation of the PI3K/AKT signaling pathway, resulting in the amelioration of pancreatic disease.
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Medicamentos Herbarios Chinos , Simulación del Acoplamiento Molecular , Farmacología en Red , Pancreatitis , Transducción de Señal , Animales , Pancreatitis/tratamiento farmacológico , Pancreatitis/metabolismo , Pancreatitis/patología , Medicamentos Herbarios Chinos/farmacología , Medicamentos Herbarios Chinos/uso terapéutico , Medicamentos Herbarios Chinos/química , Ratas , Transducción de Señal/efectos de los fármacos , Masculino , Modelos Animales de Enfermedad , Apoptosis/efectos de los fármacos , Ratas Sprague-Dawley , Mapas de Interacción de ProteínasRESUMEN
Acute pancreatitis, marked by sudden inflammation of the pancreas, presents a complex spectrum of causative factors including gallstone obstruction, alcohol abuse, and viral infections. Recent studies have illuminated the emergence of vaccine-induced acute pancreatitis, notably associated with COVID-19 vaccinations, presenting diverse mechanisms ranging from direct viral-mediated injury to autoimmune reactions. Understanding this link is pivotal for public health, yet challenges persist in identifying and managing cases post-vaccination. Comprehensive literature reviews employing the PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) statement outline the potential pathways and mechanisms leading to vaccine-induced pancreatitis, emphasizing the need for deeper investigations into underlying health conditions and modifications to vaccine components. Notably, the rare occurrences of vaccine-induced pancreatitis extend beyond COVID-19 vaccines, with reports also documenting associations with measles, mumps, and rubella (MMR), human papillomavirus (HPV), and other viral vaccinations. Mechanistically, hypotheses such as molecular mimicry and immunologic injury have been proposed, necessitating ongoing vigilance and exploration. Regulatory agencies play a crucial role in monitoring and communicating vaccine safety concerns, emphasizing transparency to address potential risks and maintain public trust. Understanding and communicating these rare adverse events with transparency remain integral for informed vaccination policies and to allay concerns surrounding vaccine safety.
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Acute pancreatitis (AP) can lead to death; however, there is no specific treatment for AP. Screening of drugs for AP treatment is rarely performed. Compounds were screened in a primary pancreatic acinar cell and peritoneal macrophage coculture system. Compounds were used in vitro and in vivo. Compound targets were predicted and validated. Among the 18 nitrogen-containing heterocycles, Z10 was shown to decrease the cerulein plus lipopolysaccharide (CL)-induced secretion of both acinar digestive enzymes and macrophage cytokines. Z10 was also shown to ameliorate CL-induced or sodium taurocholate-induced AP in mice. Proteomics analysis and enzyme linked immunosorbent assay (ELISA) revealed that Z10 decreased the levels of D-dopachrome tautomerase (Ddt) within macrophages and those in the extracellular milieu under CL treatment. Z10 also decreased Ddt expression in AP mice. Moreover, exogenous Ddt induced cytokine and digestive enzyme secretion, which could be inhibited by Z10. Ddt knockdown inhibited CL-induced cytokine secretion. Medium from CL-treated macrophages induced the release of amylase by acinar cells, and Ddt knockdown medium decreased amylase secretion. The target of Z10 was predicted to be ERK2. Z10 increased the thermostability of ERK1/2 but not ERK1 K72A/ERK2 K52A. The docking poses of ERK1 and ERK2 with Z10 were similar. Z10 inhibited ERK1/2 phosphorylation, and Ddt levels and cytokines were regulated by ERK1/2 during AP. Additionally, Z10 could not further inhibit cytokines under ERK1/2 knockdown with CL. Thus, this study revealed that Z10-mediated ERK1/2 inhibition decreased Ddt expression and secretion by macrophages. Ddt inhibition decreased cytokine release and digestive enzyme secretion.
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Pancreatitis , Ratones , Animales , Pancreatitis/inducido químicamente , Pancreatitis/tratamiento farmacológico , Pancreatitis/metabolismo , Enfermedad Aguda , Citocinas , Amilasas/efectos adversos , PirazolesRESUMEN
During organ formation, progenitor cells need to acquire different cell identities and organize themselves into distinct structural units. How these processes are coordinated and how tissue architecture(s) is preserved despite the dramatic cell rearrangements occurring in developing organs remain unclear. Here, we identified cellular rearrangements between acinar and ductal progenitors as a mechanism to drive branching morphogenesis in the pancreas while preserving the integrity of the acinar-ductal functional unit. Using ex vivo and in vivo mouse models, we found that pancreatic ductal cells form clefts by protruding and pulling on the acinar basement membrane, which leads to acini splitting. Newly formed acini remain connected to the bifurcated branches generated by ductal cell rearrangement. Insulin growth factor (IGF)/phosphatidylinositol 3-kinase (PI3K) pathway finely regulates this process by controlling pancreatic ductal tissue fluidity, with a simultaneous impact on branching and cell fate acquisition. Together, our results explain how acinar structure multiplication and branch bifurcation are synchronized during pancreas organogenesis.
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Fosfatidilinositol 3-Quinasa , Fosfatidilinositol 3-Quinasas , Ratones , Animales , Fosfatidilinositol 3-Quinasas/metabolismo , Fosfatidilinositol 3-Quinasa/metabolismo , Páncreas , Células Acinares/metabolismo , Morfogénesis/fisiología , Péptidos y Proteínas de Señalización Intercelular/metabolismoRESUMEN
Objective: With the number of patients with acute pancreatitis (AP) increasing year by year, it is pressing to explore new key genes and markers for the treatment of AP. miR-455-3p/solute carrier family 2 member 1 (Slc2a1) obtained through bioinformatics analysis may participate in the progression of AP. Materials and Methods: The C57BL/6 mouse model of AP was constructed for subsequent studies. Through bioinformatics analysis, the differentially expressed genes related to AP were screened and hub genes were identified. A caerulein-induced AP animal model was constructed to detect the pathological changes of mouse pancreas by HE staining. The concentrations of amylase and lipase were measured. Primary mouse pancreatic acinar cells were isolated and subjected to microscopy to observe their morphology. The enzymatic activities of trypsin and amylase were detected. The secretion of inflammatory cytokines in mouse were measured with the ELISA kits of TNF-α, IL-6 and IL-1ß to determine pancreatic acinar cell damage. A binding site between the Slc2a1 3' UTR region and the miR-455-3p sequence was verified by dual-luciferase reporter assay. The expression of miR-455-3p was quantified by qRT-PCR, and Slc2a1 were detected by western blot. Results: A total of five (Fyn, Gadd45a, Sdc1, Slc2a1, and Src) were identified by bioinformatics analysis, and miR-455-3p/Slc2a1 were further studied. HE staining results showed that the AP models were successfully established by caerulein induction. In mice with AP, the expression of miR-455-3p was reduced, while that of Slc2a1 was increased. In the caerulein-induced cell model, the expression of Slc2a1 was significantly reduced after intervention of miR-455-3p mimics, whereas increased after miR-455-3p inhibitor treatment. miR-455-3p decreased the secretion of inflammatory cytokines in the cell supernatant, reduced the activity of trypsin and amylase, and alleviated the cell damage induced by caerulein. In addition, Slc2a1 3'UTR region was bound by miR-455-3p, and its protein expression was also regulated. Conclusion: miR-455-3p alleviated caerulein-induced mouse pancreatic acinar cell damage by regulating the expression of Slc2a1.
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MicroARNs , Pancreatitis , Animales , Ratones , Células Acinares , Enfermedad Aguda , Amilasas/efectos adversos , Ceruletida/efectos adversos , Citocinas/efectos adversos , Ratones Endogámicos C57BL , MicroARNs/genética , Pancreatitis/inducido químicamente , Tripsina/efectos adversosRESUMEN
The exocrine pancreas secretes fluid and digestive enzymes in response to parasympathetic release of acetylcholine (ACh) via the vagus nerve and the gut hormone cholecystokinin (CCK). Both secretion of fluid and exocytosis of secretory granules containing enzymes and zymogens are dependent on an increase in the cytosolic [Ca2+ ] in acinar cells. It is thought that the specific spatiotemporal characteristics of the Ca2+ signals are fundamental for appropriate secretion and that these properties are disrupted in disease states in the pancreas. While extensive research has been performed to characterize Ca2+ signalling in acinar cells, this has exclusively been achieved in ex vivo preparations of exocrine cells, where it is difficult to mimic physiological conditions. Here we have developed a method to optically observe pancreatic acinar Ca2+ signals in vivo using a genetically expressed Ca2+ indicator and imaged with multi-photon microscopy in live animals. In vivo, acinar cells exhibited baseline activity in fasted animals, which was dependent on CCK1 receptors (CCK1Rs). Both stimulation of intrinsic nervous input and administration of systemic CCK induced oscillatory activity in a proportion of the cells, but the maximum frequencies were vastly different. Upon feeding, oscillatory activity was also observed, which was dependent on CCK1Rs. No evidence of a vago-vagal reflex mediating the effects of CCK was observed. Our in vivo method revealed the spatial and temporal profile of physiologically evoked Ca2+ signals, which will provide new insights into future studies of the mechanisms underlying exocrine physiology and that are disrupted in pathological conditions. KEY POINTS: In the exocrine pancreas, the spatiotemporal properties of Ca2+ signals are fundamentally important for the appropriate stimulation of secretion by the neurotransmitter acetylcholine and gut hormone cholecystokinin. These characteristics were previously defined in ex vivo studies. Here we report the spatiotemporal characteristics of Ca2+ signals in vivo in response to physiological stimulation in a mouse engineered to express a Ca2+ indicator in acinar cells. Specific Ca2+ 'signatures' probably important for stimulating secretion are evoked in vivo in fasted animals, by feeding, neural stimulation and cholecystokinin administration. The Ca2+ signals are probably the result of the direct action of ACh and CCK on acinar cells and not indirectly through a vago-vagal reflex.
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Células Acinares , Páncreas Exocrino , Ratones , Animales , Acetilcolina/farmacología , Páncreas , Colecistoquinina/farmacología , Calcio/farmacologíaRESUMEN
The type 2 Ca2+-dependent activator protein for secretion (CAPS2/CADPS2) regulates dense-core vesicle trafficking and exocytosis and is involved in the regulated release of catecholamines, peptidergic hormones, and neuromodulators. CAPS2 is expressed in the pancreatic exocrine acinar cells that produce and secrete digestive enzymes. However, the functional role of CAPS2 in vesicular trafficking and/or exocytosis of non-regulatory proteins in the exocrine pancreas remains to be determined. Here, we analyzed the morpho-pathological indicators of the pancreatic exocrine pathway in Cadps2-deficient mouse models using histochemistry, biochemistry, and electron microscopy. We used whole exosome sequencing to identify CADPS2 variants in patients with chronic pancreatitis (CP). Caps2/Cadps2-knockout (KO) mice exhibited morphophysiological abnormalities in the exocrine pancreas, including excessive accumulation of secretory granules (zymogen granules) and their amylase content in the cytoplasm, deterioration of the fine intracellular membrane structures (disorganized rough endoplasmic reticulum, dilated Golgi cisternae, and the appearance of empty vesicles and autophagic-like vacuoles), as well as exocrine pancreatic cell injury, including acinar cell atrophy, increased fibrosis, and inflammatory cell infiltration. Pancreas-specific Cadps2 conditional KO mice exhibited pathological abnormalities in the exocrine pancreas similar to the global Cadps2 KO mice, indicating that these phenotypes were caused either directly or indirectly by CAPS2 deficiency in the pancreas. Furthermore, we identified a rare variant in the exon3 coding region of CADPS2 in a non-alcoholic patient with CP and showed that Cadps2-dex3 mice lacking CAPS2 exon3 exhibited symptoms similar to those exhibited by the Cadps2 KO and cKO mice. These results suggest that CAPS2 is critical for the proper functioning of the pancreatic exocrine pathway, and its deficiency is associated with a risk of pancreatic acinar cell pathology.
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BACKGROUND: The microarray data analysis predicted that Rbpjl is poorly expressed in acute pancreatitis (AP). Activated IL-6/STAT3 signaling is further known to contribute to the progression of AP through immune regulation, and both IL-6 and STAT3 were bioinformatically predicted to interact with Arid5a. Accordingly, we aimed to investigate the potential involvement of the Arid5a/IL-6/STAT3 axis in the regulatory role of Rbpjl in the inflammation of AP. METHODS: Pancreatic acinar cells were exposed to lipopolysaccharide (LPS) to induce the pancreatic cell damage, and mice were subjected to supramaximal cerulein stimulation to induce AP. Expression patterns of Rbpjl and the Arid5a/IL-6/STAT3 axis were measured in mouse and cell models. Their expression was further manipulated to explore their effects on pancreatic cell injury and inflammation, as reflected by cell viability and apoptosis as well as reactive oxygen species (ROS) accumulation and proinflammatory cytokine secretion. Moreover, ChIP, EMSA, and dual-luciferase reporter assays were carried out to identify the interactions between Rbpjl and Arid5a. RESULTS: Rbpjl was found to be down-regulated in pancreatic tissues of AP mice and LPS-induced pancreatic acinar cells, while re-expression of Rbpjl led to enhanced cell viability, suppressed LPS-induced inflammation and ROS accumulation, and alleviation of AP-induced damage. Mechanistically, Rbpjl could bind to the promoter region of Arid5a and down-regulated its expression, thus repressing the activation of the IL-6/STAT3 signal axis. Furthermore, Rbpjl impaired Arid5a-dependent IL-6/STAT3 activation, hence alleviating pancreatic acinar cell inflammation. Furthermore, these effects were validated with in vivo experiments. CONCLUSION: Collectively, our findings highlight that Rbpjl attenuates AP by down-regulating Arid5a and inactivating the IL-6/STAT3 pathway.
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BACKGROUND & AIMS: Mitochondrial dysfunction disrupts the synthesis and secretion of digestive enzymes in pancreatic acinar cells and plays a primary role in the etiology of exocrine pancreas disorders. However, the transcriptional mechanisms that regulate mitochondrial function to support acinar cell physiology are poorly understood. Here, we aim to elucidate the function of estrogen-related receptor γ (ERRγ) in pancreatic acinar cell mitochondrial homeostasis and energy production. METHODS: Two models of ERRγ inhibition, GSK5182-treated wild-type mice and ERRγ conditional knock-out (cKO) mice, were established to investigate ERRγ function in the exocrine pancreas. To identify the functional role of ERRγ in pancreatic acinar cells, we performed histologic and transcriptome analysis with the pancreas isolated from ERRγ cKO mice. To determine the relevance of these findings for human disease, we analyzed transcriptome data from multiple independent human cohorts and conducted genetic association studies for ESRRG variants in 2 distinct human pancreatitis cohorts. RESULTS: Blocking ERRγ function in mice by genetic deletion or inverse agonist treatment results in striking pancreatitis-like phenotypes accompanied by inflammation, fibrosis, and cell death. Mechanistically, loss of ERRγ in primary acini abrogates messenger RNA expression and protein levels of mitochondrial oxidative phosphorylation complex genes, resulting in defective acinar cell energetics. Mitochondrial dysfunction due to ERRγ deletion further triggers autophagy dysfunction, endoplasmic reticulum stress, and production of reactive oxygen species, ultimately leading to cell death. Interestingly, ERRγ-deficient acinar cells that escape cell death acquire ductal cell characteristics, indicating a role for ERRγ in acinar-to-ductal metaplasia. Consistent with our findings in ERRγ cKO mice, ERRγ expression was significantly reduced in patients with chronic pancreatitis compared with normal subjects. Furthermore, candidate locus region genetic association studies revealed multiple single nucleotide variants for ERRγ that are associated with chronic pancreatitis. CONCLUSIONS: Collectively, our findings highlight an essential role for ERRγ in maintaining the transcriptional program that supports acinar cell mitochondrial function and organellar homeostasis and provide a novel molecular link between ERRγ and exocrine pancreas disorders.
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Páncreas Exocrino , Pancreatitis Crónica , Células Acinares/patología , Animales , Estrógenos/metabolismo , Humanos , Ratones , Ratones Noqueados , Páncreas/patología , Páncreas Exocrino/metabolismo , Pancreatitis Crónica/patologíaRESUMEN
In alcoholic pancreatitis, alcohol increases gut permeability, which increases the penetration of endotoxins, such as lipopolysaccharides (LPS). LPS act as clinically significant triggers to increase pancreatic damage in alcoholic pancreatitis. Ethanol or LPS treatment increases reactive oxygen species (ROS) production in pancreatic acinar cells. ROS induce inflammatory cytokine production in pancreatic acinar cells, leading to pancreatic inflammation. The nuclear erythroid-2-related factor 2 (Nrf2) pathway is activated as a cytoprotective response to oxidative stress, and induces the expression of NAD(P)H quinone oxidoreductase 1 (NQO1) and heme oxygenase-1 (HO-1). Lycopene exerts anti-inflammatory and antioxidant effects in various cells. We previously showed that lycopene inhibits NADPH oxidase to reduce ROS and IL-6 levels, and zymogene activation in ethanol or palmitoleic acid-treated pancreatic acinar cells. In this study, we examined whether lycopene inhibits IL-6 expression by activating the Nrf2/NQO1-HO-1 pathway, and reducing intracellular and mitochondrial ROS levels, in ethanol and LPS-treated pancreatic AR42J cells. Lycopene increased the phosphorylated and nuclear-translocated Nrf2 levels by decreasing the amount of Nrf2 sequestered in the cytoplasm via a complex formation with Kelch-like ECH1-associated protein 1 (Keap1). Using exogenous inhibitors targeting Nrf2 and HO-1, we showed that the upregulation of activated Nrf2 and HO-1 results in lycopene-induced suppression of IL-6 expression and ROS production. The consumption of lycopene-rich foods may prevent the development of ethanol and LPS-associated pancreatic inflammation by activating Nrf2-mediated expression of NQO1 and HO-1, thereby decreasing ROS-mediated IL-6 expression in pancreatic acinar cells.
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Acute pancreatitis (AP) is a common gastrointestinal disease that affects 1 million individuals worldwide. Inflammation and apoptosis are considered to be important pathogenic mechanisms of AP, and high mobility group box 1 (HMGB1) has been shown to play a particularly important role in the etiology of this disease. MicroRNAs (miRs) are emerging as critical regulators of gene expression and, as such, they represent a promising area of therapeutic target identification and development for a variety of diseases, including AP. Using the online database query (microRNA.org), the current study identified a site in the 3' untranslated region of HMGB1 mRNA that was a viable target for miR-340-5p. The present study aimed to investigate the association between miR-340-5p and HMGB1 expression in pancreatic acinar cells following lipopolysaccharide (LPS) treatment by performing luciferase, western blotting and reverse transcription-quantitative PCR assays. The results suggest that miR-340-5p attenuates the induction of HMGB1 by LPS, thereby inhibiting inflammation and apoptosis via blunted activation of Toll-like receptor 4 and enhanced AKT signaling. Thus, the therapeutic application of miR-340-5p may be a useful strategy in AP via upregulation of HMGB1 and subsequent promotion of inflammation and apoptosis.
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The inositol 1,4,5-triphosphate receptor-associated 2 (IRAG2) is also known as Jaw1 or lymphoid-restricted membrane protein (LRMP) and shares homology with the inositol 1,4,5-triphosphate receptor-associated cGMP kinase substrate 1 (IRAG1). IRAG1 interacts with inositol trisphosphate receptors (IP3 receptors /IP3R) via its coiled-coil domain and modulates Ca2+ release from intracellular stores. Due to the homology of IRAG1 and IRAG2, especially in its coiled-coil domain, it is possible that IRAG2 has similar interaction partners like IRAG1 and that IRAG2 also modulates intracellular Ca2+ signaling. In our study, we localized IRAG2 in pancreatic acinar cells of the exocrine pancreas, and we investigated the interaction of IRAG2 with IP3 receptors and its impact on intracellular Ca2+ signaling and exocrine pancreatic function, like amylase secretion. We detected the interaction of IRAG2 with different subtypes of IP3R and altered Ca2+ release in pancreatic acinar cells from mice lacking IRAG2. IRAG2 deficiency decreased basal levels of intracellular Ca2+, suggesting that IRAG2 leads to activation of IP3R under unstimulated basal conditions. Moreover, we observed that loss of IRAG2 impacts the secretion of amylase. Our data, therefore, suggest that IRAG2 modulates intracellular Ca2+ signaling, which regulates exocrine pancreatic function.
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Células Acinares/metabolismo , Calcio/metabolismo , Receptores de Inositol 1,4,5-Trifosfato/metabolismo , Inositol 1,4,5-Trifosfato/metabolismo , Proteínas de la Membrana/metabolismo , Páncreas Exocrino/metabolismo , Amilasas/metabolismo , Animales , Señalización del Calcio/fisiología , Femenino , Masculino , Ratones , Ratones NoqueadosRESUMEN
Acute pancreatitis (AP) is one of the most frequent gastrointestinal diseases and has no specific treatment. It has been shown that dysfunction of pancreatic acinar cells can lead to AP progression. Emodin is a natural product, which can alleviate the symptoms of AP. However, the mechanism by which emodin regulates the function of pancreatic acinar cells remains unclear. Thus, the present study aimed to investigate the mechanism by which emodin modulates the function of pancreatic acinar cells. To mimic AP in vitro, pancreatic acinar cells were cotreated with caerulein and lipopolysaccharide (LPS). Exosomes were isolated using the ExoQuick precipitation kit. Western blot analysis, Nanosight Tracking analysis and transmission electron microscopy were performed to detect the efficiency of exosome separation. Gene expression was detected by reverse transcriptionquantitative PCR. The levels of IL1ß and TNFα were detected by ELISA. The data indicated that emodin significantly decreased the levels of IL1ß and TNFα in the supernatant samples derived from AR42J cells cotreated with caerulein and LPS. In addition, emodin significantly promoted the proliferation of AR42J cells cotreated with caerulein and LPS, and inhibited apoptosis, while the effect of emodin was reversed by long noncoding (lnc)RNA taurine upregulated 1 (TUG1) overexpression. The expression level of TUG1 in AR42J cells or exosomes derived from AR42J cells was significantly increased following treatment of the cells with LPS and caerulein, while this effect was notably reversed by emodin treatment. In addition, exosomes derived from caerulein and LPS cotreated AR42J cells inhibited the differentiation and antiinflammatory function of regulatory T cells, while treatment of the cells with emodin significantly decreased this effect. In conclusion, the data indicated that emodin inhibited the induction of inflammation in AR42J cells by regulating the expression of cellular and exosomal lncRNA. Therefore, emodin may be used as a potential agent for the treatment of AP.
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Emodina/farmacología , Pancreatitis/tratamiento farmacológico , ARN Largo no Codificante/metabolismo , Células Acinares/efectos de los fármacos , Células Acinares/metabolismo , Células Acinares/patología , Animales , Apoptosis/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Exosomas/genética , Humanos , Masculino , Ratones , Pancreatitis/genética , Pancreatitis/metabolismo , Pancreatitis/patología , Inhibidores de Proteínas Quinasas/farmacología , ARN Largo no Codificante/genética , Ratas WistarRESUMEN
The pancreatic ß cells can synthesize dopamine by taking L-dihydroxyphenylalanine, but whether pancreatic acinar cells synthesize dopamine has not been confirmed. By means of immunofluorescence, the tyrosine hydroxylase -immunoreactivity and aromatic amino acid decarboxylase (AADC)- immunoreactivity were respectively observed in pancreatic acinar cells and islet ß cells. Treatment with L-dihydroxyphenylalanine, not tyrosine, caused the production of dopamine in the incubation of INS-1 cells (rat islet ß cell line) and primary isolated islets, which was blocked by AADC inhibitor NSD-1015. However, only L-dihydroxyphenylalanine, but not dopamine, was detected when AR42J cells (rat pancreatic acinar cell line) were treated with tyrosine, which was blocked by tyrosine hydroxylase inhibitor AMPT. Dopamine was detected in the coculture of INS-1 cells with AR42J cells after treatment with tyrosine. In an in vivo study, pancreatic juice contained high levels of L-dihydroxyphenylalanine and dopamine. Both L-dihydroxyphenylalanine and dopamine accompanied with pancreatic enzymes and insulin in the pancreatic juice were all significantly increased after intraperitoneal injection of bethanechol chloride and their increases were all blocked by atropine. Inhibiting TH with AMPT blocked bethanechol chloride-induced increases in L-dihydroxyphenylalanine and dopamine, while inhibiting AADC with NSD-1015 only blocked the dopamine increase. Bilateral subdiaphragmatic vagotomy of rats leads to significant decreases of L-dihydroxyphenylalanine and dopamine in pancreatic juice. These results suggested that pancreatic acinar cells could utilize tyrosine to synthesize L-dihydroxyphenylalanine, not dopamine. Islet ß cells only used L-dihydroxyphenylalanine, not tyrosine, to synthesize dopamine. Both L-dihydroxyphenylalanine and dopamine were respectively released into the pancreatic duct, which was regulated by the vagal cholinergic pathway. The present study provides important evidences for the source of L-dihydroxyphenylalanine and dopamine in the pancreas.
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Células Acinares/metabolismo , Dihidroxifenilalanina/biosíntesis , Dopamina/biosíntesis , Islotes Pancreáticos/metabolismo , Tirosina/metabolismo , Animales , Inhibidores de Descarboxilasas de Aminoácidos Aromáticos/farmacología , Descarboxilasas de Aminoácido-L-Aromático/inmunología , Descarboxilasas de Aminoácido-L-Aromático/metabolismo , Atropina/farmacología , Betanecol/farmacología , Línea Celular , Dihidroxifenilalanina/análisis , Dopamina/análisis , Hidrazinas/farmacología , Islotes Pancreáticos/citología , Ratas , Ratas Sprague-Dawley , Tirosina 3-Monooxigenasa/antagonistas & inhibidores , Tirosina 3-Monooxigenasa/inmunología , Tirosina 3-Monooxigenasa/metabolismoRESUMEN
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.
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Adenosina Trifosfato/fisiología , Calcio/fisiología , Páncreas Exocrino/fisiología , Enfermedades Pancreáticas/fisiopatología , Animales , Señalización del Calcio , Humanos , Páncreas Exocrino/fisiopatologíaRESUMEN
BACKGROUND: Alcoholic chronic pancreatitis (ACP) is a serious inflammatory disorder of the exocrine pancreatic gland. A previous study from this laboratory showed that ethanol (EtOH) causes cytotoxicity, dysregulates AMPKα and ER/oxidative stress signaling, and induces inflammatory responses in primary human pancreatic acinar cells (hPACs). Here we examined the differential cytotoxicity of EtOH and its oxidative (acetaldehyde) and nonoxidative (fatty acid ethyl esters; FAEEs) metabolites in hPACs was examined to understand the metabolic basis and mechanism of ACP. METHODS: We evaluated concentration-dependent cytotoxicity, AMPKα inactivation, ER/oxidative stress, and inflammatory responses in hPACs by incubating them for 6 h with EtOH, acetaldehyde, or FAEEs at clinically relevant concentrations reported in alcoholic subjects using conventional methods. Cellular bioenergetics (mitochondrial stress and a real-time ATP production rate) were determined using Seahorse XFp Extracellular Flux Analyzer in AR42J cells treated with acetaldehyde or FAEEs. RESULTS: We observed concentration-dependent increases in LDH release, inactivation of AMPKα along with upregulation of ACC1 and FAS (key lipogenic proteins), downregulation of p-LKB1 (an oxidative stress-sensitive upstream kinase regulating AMPKα) and CPT1A (involved in ß-oxidation of fatty acids) in hPACs treated with EtOH, acetaldehyde, or FAEEs. Concentration-dependent increases in oxidative stress and ER stress as measured by GRP78, unspliced XBP1, p-eIF2α, and CHOP along with activation of p-JNK1/2, p-ERK1/2, and p-P38MAPK were present in cells treated with EtOH, acetaldehyde, or FAEEs, respectively. Furthermore, a significant decrease was observed in the total ATP production rate with subsequent mitochondrial stress in AR42J cells treated with acetaldehyde and FAEEs. CONCLUSIONS: EtOH and its metabolites, acetaldehyde and FAEEs, caused cytotoxicity, ER/oxidative and mitochondrial stress, and dysregulated AMPKα signaling, suggesting a key role of EtOH metabolism in the etiopathogenesis of ACP. Because oxidative EtOH metabolism is negligible in the exocrine pancreas, the pathogenesis of ACP could be attributable to the formation of FAEEs and related pancreatic acinar cell injury.
Asunto(s)
Células Acinares/efectos de los fármacos , Depresores del Sistema Nervioso Central/farmacología , Estrés del Retículo Endoplásmico/efectos de los fármacos , Etanol/farmacología , Mitocondrias/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Páncreas/citología , Quinasas de la Proteína-Quinasa Activada por el AMP/efectos de los fármacos , Quinasas de la Proteína-Quinasa Activada por el AMP/metabolismo , Proteínas Quinasas Activadas por AMP/efectos de los fármacos , Proteínas Quinasas Activadas por AMP/metabolismo , Acetaldehído/farmacología , Acetil-CoA Carboxilasa/efectos de los fármacos , Acetil-CoA Carboxilasa/metabolismo , Células Acinares/metabolismo , Carnitina O-Palmitoiltransferasa/efectos de los fármacos , Carnitina O-Palmitoiltransferasa/metabolismo , Supervivencia Celular/efectos de los fármacos , Ésteres/farmacología , Humanos , Mitocondrias/metabolismo , Proteína Quinasa 1 Activada por Mitógenos/efectos de los fármacos , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos/efectos de los fármacos , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Proteína Quinasa 8 Activada por Mitógenos/efectos de los fármacos , Proteína Quinasa 8 Activada por Mitógenos/metabolismo , Proteína Quinasa 9 Activada por Mitógenos/efectos de los fármacos , Proteína Quinasa 9 Activada por Mitógenos/metabolismoRESUMEN
Multiorgan failure may not be completely resolved among people living with HIV despite HAART use. Although the chances of organ dysfunction may be relatively low, alcohol may potentiate HIV-induced toxic effects in the organs of alcohol-abusing, HIV-infected individuals. The pancreas is one of the most implicated organs, which is manifested as diabetes mellitus or pancreatic cancer. Both alcohol and HIV may trigger pancreatitis, but the combined effects have not been explored. The aim of this review is to explore the literature for understanding the mechanisms of HIV and alcohol-induced pancreatotoxicity. We found that while premature alcohol-inducing zymogen activation is a known trigger of alcoholic pancreatitis, HIV entry through C-C chemokine receptor type 5(CCR5)into pancreatic acinar cells may also contribute to pancreatitis in people living with HIV (PLWH). HIV proteins induce oxidative and ER stresses, causing necrosis. Furthermore, infiltrative immune cells induce necrosis on HIV-containing acinar cells. When necrotic products interact with pancreatic stellate cells, they become activated, leading to the release of both inflammatory and profibrotic cytokines and resulting in pancreatitis. Effective therapeutic strategies should block CCR5 and ameliorate alcohol's effects on acinar cells.
RESUMEN
Immune cells were identified in intact live mouse pancreatic lobules and their Ca2+ signals, evoked by various agents, characterized and compared with the simultaneously recorded Ca2+ signals in neighboring acinar and stellate cells. Immunochemistry in the live lobules indicated that the pancreatic immune cells most likely are macrophages. In the normal pancreas the density of these cells is very low, but induction of acute pancreatitis (AP), by a combination of ethanol and fatty acids, markedly increased the number of the immune cells. The principal agent eliciting Ca2+ signals in the pancreatic immune cells was ATP, but these cells also frequently produced Ca2+ signals in response to acetylcholine and to high concentrations of bradykinin. Pharmacological studies, using specific purinergic agonists and antagonists, indicated that the ATP-elicited Ca2+ signals were mediated by both P2Y1 and P2Y13 receptors. The pancreatic immune cells were not electrically excitable and the Ca2+ signals generated by ATP were primarily due to release of Ca2+ from internal stores followed by store-operated Ca2+ entry through Ca2+ release-activated Ca2+ channels. The ATP-induced intracellular Ca2+ liberation was dependent on both IP3 generation and IP3 receptors. We propose that the ATP-elicited Ca2+ signal generation in the pancreatic immune cells is likely to play an important role in the severe inflammatory response to the primary injury of the acinar cells that occurs in AP.