Pancreatic Acinar Cells Research Articles

Berberine inhibits intracellular Ca2+ signals in mouse pancreatic acinar cells through M3 muscarinic receptors: Novel target, mechanism, and implication

Berberine, a natural isoquinoline alkaloid, exhibits a variety of pharmacological effects, but the pharmacological targets and mechanisms remain elusive. Here, we report a novel finding that berberine inhibits acetylcholine (ACh)-induced intracellular Ca2+ oscillations, mediated through an inhibition of the muscarinic subtype 3 (M3) receptor. Patch-clamp recordings and confocal Ca2+ imaging were applied to acute dissociated pancreatic acinar cells prepared from CD1 mice to examine the effects of berberine on ACh-induced Ca2+ oscillations. Whole-cell patch-clamp recordings showed that berberine (from 0.1 to 10 µM) reduced ACh-induced Ca2+ oscillations in a concentration-dependent manner, and this inhibition also depended on ACh concentrations. The inhibitory effect of berberine neither occurred in intracellular targets nor extracellular cholecystokinin (CCK) receptors, chloride (Cl-) channels, and store-operated Ca2+ channels. Together, the results demonstrate that berberine directly inhibits the muscarinic M3 receptors, further confirmed by evidence of the interaction between berberine and M3 receptors in pancreatic acinar cells.

Stretching the role of TMEM63a to gatekeeping Ca2+ release in pancreatic acinar cells

Stretching the role of TMEM63a to gatekeeping Ca2+ release in pancreatic acinar cells

Algal Oil Mitigates Sodium Taurocholate-Induced Pancreatitis by Alleviating Calcium Overload, Oxidative Stress, and NF-κB Activation in Pancreatic Acinar Cells.

Acute pancreatitis (AP) is characterized by elevated intracellular Ca2+ concentrations, mitochondrial dysfunction, and oxidative stress in pancreatic acinar cells. Algal oil (AO) has demonstrated antioxidant and anti-inflammatory properties. This study aims to explore the effects of algal oil on the microenvironment of AP. Rat pancreatic acinar AR42J cells were pretreated with AO containing 0, 50, 100, or 150 μM of docosahexaenoic acid (DHA) 2 h prior to AP induction using sodium taurocholate (STC). After 1 h of STC treatment, AR42J cells exhibited a significant increase in intracellular Ca2+ concentration and the production of amylase, lipase, reactive oxygen species, and pro-inflammatory mediators, including tumor necrosis factor-α and interleukin-6. These STC-induced increases were markedly reduced in cells pretreated with AO. In comparison to cells without AO, those treated with a high dose of AO before STC exposure demonstrated a significant increase in mitochondrial membrane potential and a decrease in lipid peroxidation. Furthermore, STC-activated nuclear factor kappa-B (NF-κB) was attenuated in AO-pretreated cells, as evidenced by a significant decrease in activated NF-κB. In conclusion, AO may prevent damage to pancreatic acinar cells by alleviating intracellular Ca2+ overload, mitigating mitochondrial dysfunction, reducing oxidative stress, and attenuating NF-κB-targeted inflammation.

Material basis and molecular mechanisms of Chaihuang Qingyi Huoxue Granule in the treatment of acute pancreatitis based on network pharmacology and molecular docking-based strategy.

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. 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. 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. 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.

Tumor Protein D52 Regulates Paligenosis in Exocrine Pancreatic Acinar Cells

Background: Paligenosis is a conserved cellular plasticity program in which fully differentiated cells reenter the cell cycle in order to regenerate lost tissue during injury. This process is utilized by cells that have a slow turnover rate and lack dedicated stem cells, as is the case with exocrine pancreatic acinar cells. Acinar cells, which synthesize and secrete digestive enzymes necessary for macronutrient digestion, undergo paligenosis in response to damage induced by the inflammatory disease pancreatitis. During paligenosis, acinar cells first downscale their secretory apparatus by upregulating autophagy, and then take on a mucinous-ductal-progenitor-like identity in a process called acinar to ductal metaplasia (ADM) prior to proliferation. This phenomenon can be recapitulated in rodent models in vivo utilizing repeated episodes of cerulein-induced pancreatitis or in 18-hour suspension culture during which primary isolated acinar cells dedifferentiate to a progenitor-like state. We have previously reported that markers of the endolysosomal secretory pathway in acinar cells are rapidly lost during models of experimental pancreatitis and suspension culture. Tumor Protein D52, which regulates endolysosomal traffcking and secretion in acinar cells, undergoes lysosomal degradation at the onset of experimental pancreatitis; furthermore, restoring D52 expression by adenovirus in isolated acinar cells maintains secretion during experimental pancreatitis as well as maintains acinar differentiation in suspension culture. Hypothesis: Degradation of D52, and loss of the endolysosomal secretory pathway, are pivotal events during the initial stages of paligenosis. Methods: To study the role of D52 and the regulation of paligenosis, we generated tamoxifen-inducible, acinar specific D52 knockout mice (D52KOac), and markers of paligenosis (ADM, proliferation) were assessed by immunoblot, qPCR, and immunostaining. Mice were given tamoxifen at 2-3 months of age to ensure normal pancreatic development, and tissues were harvested 21 days after tamoxifen. Results: Compared to controls, D52KOac exhibited 2.5-fold upregulation of the ductal markers Sox9 and CK19, 50% reduction of Ptf1α, and a 5-fold increase in β-catenin; these markers signify the presence of ADM and cells with progenitor-like characteristics. Cell cycle regulation and proliferation were also observed in D52KOac by a nearly 3-fold increase in cyclin D and increased Ki67+ nuclei. Conclusions: These data support that loss of D52 in pancreatic acinar cells induces spontaneous paligenosis in the absence of cellular injury and inflammation. These findings point to a previously unrecognized role for D52 in the regulation of acinar cell differentiation and plasticity. Given that paligenosis is a key pathway involved in the development of pancreatic neoplasms and adenocarcinoma, studying D52-regulated paligenosis will reveal additional insights into this critical protective mechanism. American Pancreatic Association Foundation Young Investigator Award. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Screening of Amino Acids as a Safe Energy Source for Isolated Rat Pancreatic Acini.

Amino acids play an essential role in protein synthesis, metabolism, and survival of pancreatic acini. Adequate nutritional support is important for acute pancreatitis treatment. However, high concentrations of arginine and lysine may induce acute pancreatitis. The study aimed to identify the most suitable l -amino acids as safe energy sources for pancreatic acinar cells. Pancreatic acini were isolated from male Wistar rats. Effects of amino acids (0.1-20 mM) on uncoupled respiration of isolated acini were studied with a Clark electrode. Cell death was evaluated with fluorescent microscopy and DNA gel electrophoresis. Among the tested amino acids, glutamate, glutamine, alanine, lysine, and aspartate were able to stimulate the uncoupled respiration rate of isolated pancreatic acini, whereas arginine, histidine, and asparagine were not. Lysine, arginine, and glutamine (20 mM) caused complete loss of plasma membrane integrity of acinar cells after 24 hours of incubation. Glutamine also caused early (2-4 hours) cell swelling and blebbing. Aspartate, asparagine, and glutamate only moderately decreased the number of viable cells, whereas alanine and histidine were not toxic. DNA fragmentation assay and microscopic analysis of nuclei showed no evidence of apoptosis in cells treated with amino acids. Alanine and glutamate are safe and effective energy sources for mitochondria of pancreatic acinar cells.

Impaired Long-Chain Fatty Acid Metabolism in Carnitine Palmitoyltransferase II Knockout Mouse Leads to Chronic Pancreatitis

Background: To support high rates of protein synthesis, pancreatic acinar cells must generate large quantities of ATP. Mitochondrial dysfunction and impaired energy metabolism play a significant role in the development of pancreatitis. Patients with elevated plasma triglycerides are at increased risk for development of severe acute pancreatitis. Carnitine palmitoyltransferase II (CPT2) is required for importing and thus metabolizing long-chain fatty acids (LCFAs) in mitochondria. To inhibit LCFA metabolism, we utilized the CPT2acinarKO mouse to probe the consequences of impaired LCFA import on pancreatic function. Hypothesis: Loss of CPT2 causes an accumulation of long-chain acylcarnitines and disruption of the tissue acylcarnitine/free carnitine ratio which is thought to alter cellular function. Given acinar cells’ high metabolic requirement, impaired LCFA utilization would reduce energy production, secretory function, and alter pancreatic lipid composition. Methods: We generated mice lacking CPT2 specifically in acinar cells (CPT2acinarKO) by crossing tamoxifen-inducible, Elastase-Cre mice with CPT2 floxed mice. Mice were fed a high fat (60% kcal fat), low fat (10% kcal fat), or chow (14% kcal fat) diet for 12 weeks. Analysis of fat accumulation, edema, and fibrosis was performed with H&E and picrosirius staining. Mass Spec. lipid profiling was performed in pancreas and serum. Acinar cells were isolated from wildtype, CPT2 floxed, and CPT2acinarKO mice to assess secretory function, ATP levels, and necrosis via lactate dehydrogenase (LDH) release. Results: Histologic examination revealed CPT2acinarKO mice on all three diets displayed pancreatic fat accumulation. Strikingly, CPT2acinarKO mice fed a chow or low fat diet developed chronic pancreatitis with a 25% loss in pancreatic weight, immune cell infiltration, edema, and fibrosis. As anticipated, long-chain acylcarnitines were significantly elevated in CPT2acinarKO pancreas. Assessment of secretion in 2 weeks post tamoxifen cells revealed significantly reduced maximal (10 pM) and supramaximal (1 nM) cholecystokinin (CCK) stimulated amylase release as a percent of total cellular amylase. Acini isolated at 4 weeks post tamoxifen revealed significantly increased basal secretion and reduced maximal and supramaximal CCK stimulated amylase release. When normalized to cellular DNA, secretion was similar between CPT2acinarKO and CPT2 floxed acinar cells indicating CPT2acinarKO cells had reduced amylase levels. Wildtype acinar cells treated with unsaturated and saturated free fatty acid concentrations as low as 100 μM for 3 hours caused significant necrosis. CPT2acinarKO pancreas had significantly decreased ATP levels indicative of mitochondrial dysfunction. Conclusion: Loss of CPT2 in the pancreatic acinar cells impairs LCFA import, reduces ATP levels, disrupts the secretory pathway, promoting chronic pancreatitis. In contrast to other CPT2 knockout models, pancreatitis occurs independent of high fat feeding in CPT2acinarKO mice. Further investigation is needed to identify metabolic alterations leading to acinar damage and mitochondrial dysfunction following the loss of CPT2. UW-Madison Comprehensive Diabetes Center Pilot Grant. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Inhibition of xanthine oxidase alleviated pancreatic necrosis via HIF-1α-regulated LDHA and NLRP3 signaling pathway in acute pancreatitis

Acute pancreatitis (AP) is a potentially fatal condition with no targeted treatment options. Although inhibiting xanthine oxidase (XO) in the treatment of AP has been studied in several experimental models and clinical trials, whether XO is a target of AP and what its the main mechanism of action is remains unclear. Here, we aimed to re-evaluate whether XO is a target aggravating AP other than merely generating reactive oxygen species that trigger AP. We first revealed that XO expression and enzyme activity were significantly elevated in the serum and pancreas of necrotizing AP models. We also found that allopurinol and febuxostat, as purine-like and non-purine XO inhibitors, respectively, exhibited protective effects against pancreatic acinar cell death in vitro and pancreatic damage in vivo at different doses and treatment time points. Moreover, we observed that conditional Xdh overexpression aggravated pancreatic necrosis and severity. Further mechanism analysis showed that XO inhibition restored the hypoxia-inducible factor 1-alpha (HIF-1α)-regulated lactate dehydrogenase A (LDHA) and NOD-like receptor family pyrin domain containing 3 (NLRP3) signaling pathways and reduced the enrichment of 13C6-glucose to 13C3-lactate. Lastly, we observed that clinical circulatory XO activity was significantly elevated in severe cases and correlated with C-reactive protein levels, while pancreatic XO and urate were also increased in severe AP patients. These results together indicated that proper inhibition of XO might be a promising therapeutic strategy for alleviating pancreatic necrosis and preventing progression of severe AP by downregulating HIF-1α-mediated LDHA and NLRP3 signaling pathways.

Successful curative treatment for a ruptured pancreatic acinar cell carcinoma by radical resection following modified FOLFIRINOX: a case report and literature review.

Pancreatic acinar cell carcinoma (PACC) is a rare pancreatic tumor type, and ruptured pancreatic tumors are rarer. Computed tomography (CT) in a 48-year-old man incidentally revealed a raptured pancreatic tail tumor. The patient was treated conservatively because he was asymptomatic, and his general condition was stable. After a detailed examination, the pancreatic tumor was diagnosed as raptured PACC. Considering the potential infiltration of tumor cells into the hematoma within the omental sac, our decision is to initiate chemotherapy as the primary course of action. A liquid biopsy was performed, and comprehensive genomic profiling of circulating tumor DNA showed a tumor BRCA2 mutation. Chemotherapy with modified FOLFIRINOX (mFFX) was selected as the first treatment. After seven courses of mFFX, the primary tumor diminished remarkably. At this time, the radical resection was performed via distal pancreatectomy with simultaneous resection of the gastric wall and colon, which had adhered strongly to the tumor. Histopathological examination revealed that the tumor had shrunk to less than 5% of its original size due to chemotherapy (Grade 3 of Evans Classification). Devising treatment strategies for ruptured pancreatic malignant tumors is challenging due to the worsening general condition caused by severe abdominal symptoms and intra-abdominal bleeding. In this context, this case-report documents a rare instance of raptured PACC with a tumor BRCA2 mutation that underwent radical resection following mFFX treatment.

HP-MSCs attenuate severe acute pancreatitis in mice via inhibiting NLRP3 inflammasome-mediated acinar cell pyroptosis.

Severe acute pancreatitis (SAP) is a serious gastrointestinal disease that is facilitated by pancreatic acinar cell death. The protective role of human placental mesenchymal stem cells (hP-MSCs) in SAP has been demonstrated in our previous studies. However, the underlying mechanisms of this therapy remain unclear. Herein, we investigated the regularity of acinar cell pyroptosis during SAP and investigated whether the protective effect of hP-MSCs was associated with the inhibition of acinar cell pyroptosis. A mouse model of SAP was established by the retrograde injection of sodium taurocholate (NaTC) solution in the pancreatic duct. For the hP-MSCs group, hP-MSCs were injected via the tail vein and were monitored in vivo. Transmission electron microscopy (TEM) was used to observe the pyroptosis-associated ultramorphology of acinar cells. Immunofluorescence and Western blotting were subsequently used to assess the localization and expression of pyroptosis-associated proteins in acinar cells. Systemic inflammation and local injury-associated parameters were evaluated. Acinar cell pyroptosis was observed during SAP, and the expression of pyroptosis-associated proteins initially increased, peaked at 24h, and subsequently showed a decreasing trend. hP-MSCs effectively attenuated systemic inflammation and local injury in the SAP model mice. Importantly, hP-MSCs decreased the expression of pyroptosis-associated proteins and the activity of the NOD-, LRR-, and pyrin domain-containing protein 3 (NLRP3) inflammasome in acinar cells. Our study demonstrates the regularity and important role of acinar cell pyroptosis during SAP. hP-MSCs attenuate inflammation and inhibit acinar cell pyroptosis via suppressing NLRP3 inflammasome activation, thereby exerting a protective effect against SAP.

Comparative transcriptomic analysis reveals the molecular changes of acute pancreatitis in experimental models

BACKGROUND Acute pancreatitis (AP) encompasses a spectrum of pancreatic inflammatory conditions, ranging from mild inflammation to severe pancreatic necrosis and multisystem organ failure. Given the challenges associated with obtaining human pancreatic samples, research on AP predominantly relies on animal models. In this study, we aimed to elucidate the fundamental molecular mechanisms underlying AP using various AP models. AIM To investigate the shared molecular changes underlying the development of AP across varying severity levels. METHODS AP was induced in animal models through treatment with caerulein alone or in combination with lipopolysaccharide (LPS). Additionally, using Ptf1α to drive the specific expression of the hM3 promoter in pancreatic acinar cells transgenic C57BL/6J- hM3/Ptf1α(cre) mice were administered Clozapine N-oxide to induce AP. Subsequently, we conducted RNA sequencing of pancreatic tissues and validated the expression of significantly different genes using the Gene Expression Omnibus (GEO) database. RESULTS Caerulein-induced AP showed severe inflammation and edema, which were exacerbated when combined with LPS and accompanied by partial pancreatic tissue necrosis. Compared with the control group, RNA sequencing analysis revealed 880 significantly differentially expressed genes in the caerulein model and 885 in the caerulein combined with the LPS model. Kyoto Encyclopedia of Genes and Genomes enrichment analysis and Gene Set Enrichment Analysis indicated substantial enrichment of the TLR and NOD -like receptor signaling pathway, TLR signaling pathway, and NF-κB signaling pathway, alongside elevated levels of apoptosis-related pathways, such as apoptosis, P53 pathway, and phagosome pathway. The significantly elevated genes in the TLR and NOD -like receptor signaling pathways, as well as in the apoptosis pathway, were validated through quantitative real-time PCR experiments in animal models. Validation from the GEO database revealed that only MYD88 concurred in both mouse pancreatic tissue and human AP peripheral blood, while TLR1 , TLR7 , RIPK3 , and OAS2 genes exhibited marked elevation in human AP. The genes TUBA1A and GADD45A played significant roles in apoptosis within human AP. The transgenic mouse model hM3/Ptf1α(cre) successfully validated significant differential genes in the TLR and NOD -like receptor signaling pathways as well as the apoptosis pathway, indicating that these pathways represent shared pathological processes in AP across different models. CONCLUSION The TLR and NOD receptor signaling pathways play crucial roles in the inflammatory progression of AP, notably the MYD88 gene. Apoptosis holds a central position in the necrotic processes of AP, with TUBA1A and GADD45A genes exhibiting prominence in human AP.

Post-therapy emergence of an NBN reversion mutation in a patient with pancreatic acinar cell carcinoma.

Pancreatic acinar cell carcinoma (PACC) is a rare form of pancreatic cancer that commonly harbors targetable alterations, including activating fusions in the MAPK pathway and loss-of-function (LOF) alterations in DNA damage response/homologous recombination DNA repair-related genes. Here, we describe a patient with PACC harboring both somatic biallelic LOF of NBN and an activating NTRK1 fusion. Upon disease progression following 13 months of treatment with folinic acid, fluorouracil, irinotecan, and oxaliplatin (FOLFIRINOX), genomic analysis of a metastatic liver biopsy revealed the emergence of a novel reversion mutation restoring the reading frame of NBN. To our knowledge, genomic reversion of NBN has not been previously reported as a resistance mechanism in any tumor type. The patient was treated with, but did not respond to, targeted treatment with a selective NTRK inhibitor. This case highlights the complex but highly actionable genomic landscape of PACC and underlines the value of genomic profiling of rare tumor types such as PACC.

Isorhamnetin Alleviates Mitochondrial Injury in Severe Acute Pancreatitis via Modulation of KDM5B/HtrA2 Signaling Pathway.

Severe acute pancreatitis (SAP), a widespread inflammatory condition impacting the abdomen with a high mortality rate, poses challenges due to its unclear pathogenesis and the absence of effective treatment options. Isorhamnetin (ISO), a naturally occurring flavonoid, demonstrates robust antioxidant and anti-inflammatory properties intricately linked to the modulation of mitochondrial function. However, the specific protective impact of ISO on SAP remains to be fully elucidated. In this study, we demonstrated that ISO treatment significantly alleviated pancreatic damage and reduced serum lipase and amylase levels in the mouse model of SAP induced by sodium taurocholate (STC) or L-arginine. Utilizing an in vitro SAP cell model, we found that ISO co-administration markedly prevented STC-induced pancreatic acinar cell necrosis, primarily by inhibiting mitochondrial ROS generation, preserving ATP production, maintaining mitochondrial membrane potential, and preventing the oxidative damage and release of mitochondrial DNA. Mechanistically, our investigation identified that high-temperature requirement A2 (HtrA2) may play a central regulatory role in mediating the protective effect of ISO on mitochondrial dysfunction in STC-injured acinar cells. Furthermore, through an integrated approach involving bioinformatics analysis, molecular docking analysis, and experimental validation, we uncovered that ISO may directly impede the histone demethylation activity of KDM5B, leading to the restoration of pancreatic HtrA2 expression and thereby preserving mitochondrial function in pancreatic acinar cells following STC treatment. In conclusion, this study not only sheds new light on the intricate molecular complexities associated with mitochondrial dysfunction during the progression of SAP but also underscores the promising value of ISO as a natural therapeutic option for SAP.

ALKBH5 suppresses autophagic flux via N6-methyladenosine demethylation of ZKSCAN3 mRNA in acute pancreatitis.

Increasing evidence has demonstrated that N6-methyladenosine (m6A) RNA modification plays an essential role in a wide range of pathological conditions. Impaired autophagy is a critical hallmark of acute pancreatitis (AP). To explore the role of the m6A modification of ZKSCAN3 in the regulation of autophagy in AP. The AP mouse cell model was established by cerulein-treated mouse pancreatic acinar cells (MPC-83), and the results were confirmed by the levels of amylase and inflammatory factors. Autophagy activity was evaluated by specific identification of the autophagy-related microstructure and the expression of autophagy-related genes. ZKSCAN3 and ALKBH5 were knocked down to study the function in AP. A m6A RNA binding protein immunoprecipitation assay was used to study how the m6A modification of ZKSCAN3 mRNA is regulated by ALKBH. The increased expression of amylase and inflammatory factors in the supernatant and the accumulation of autophagic vacuoles verified that the AP mouse cell model was established. The downregulation of LAMP2 and upregulation of LC3-II/I and SQSTM1 demonstrated that autophagy was impaired in AP. The expression of ZKSCAN3 was upregulated in AP. Inhibition of ZKSCAN3 increased the expression of LAMP2 and decreased the expression of the inflammatory factors, LC3-II/I and SQSTM1. Furthermore, ALKBH5 was upregulated in AP. Knockdown of ALKBH5 downregulated ZKSCAN3 expression and restored decreased autophagic flux in AP. Notably, the bioinformatic analysis revealed 23 potential m6A modification sites on ZKSCAN3 mRNA. The m6A modification of ZKSCAN3 mRNA was significantly decreased in AP. Knockdown of ALKBH5 increased the modification of ZKSCAN3 mRNA, which confirmed that ALKBH5 upregulated ZKSCAN3 expression in a m6A-dependent manner. ALKBH5 inhibits autophagic flux through m6A demethylation of ZKSCAN3 mRNA in AP, thereby aggravating the severity of the disease.

Blocking CIRP protects against acute pancreatitis by improving mitochondrial function and suppressing pyroptosis in acinar cells

Acute pancreatitis (AP) continues to pose a major challenge as targeted therapeutic interventions are absent. Mitochondrial dysfunction and inflammasome-dependent pyroptosis are involved in the pathogenic mechanisms of AP. CIRP is a stress-response protein and a damage-associated molecular pattern (DAMP) molecule. In our previous studies, we discovered that excessive CIRP can directly damage pancreatic acinar cells. Nonetheless, the precise involvement of CIRP in AP is still unexplored. The primary aim of this study was to examine the potential involvement of CIRP in the development of pyroptosis and mitochondrial dysfunction in AP. To study this, an L-arginine-induced AP mouse model was used. Our results showed that Caspase-1-mediated pyroptosis and mitochondria-derived reactive oxygen species (ROS) were crucial factors in the occurrence of tissue damage and inflammation in AP. A substantial increase in the CIRP serum levels was observed in AP mice. Blocking CIRP by either CIRP gene knockout or systemic administration of C23, a competing inhibitor of CIRP, reduced ROS accumulation and pyroptosis in AP mice. These effects were associated with attenuated pancreatic injury and inflammation. In addition, CIRP-triggered mitochondrial dysfunction, autophagy impairment, and pyroptosis in pancreatic acinar cells were prevented by TAK242, an inhibitor of CIRP receptor TLR4. In conclusion, CIRP can induce mitochondrial dysfunction and pyroptosis in pancreatic acinar cells, and blocking CIRP may be a valuable approach to treating patients with AP.

Exosomes Derived From Cerulein-Stimulated Pancreatic Acinar Cells Mediate Peritoneal Macrophage M1 Polarization and Pyroptosis via an miR-24-3p/MARCH3/NLRP3 Axis in Acute Pancreatitis.

Acute pancreatitis (AP) has a high incidence of hospitalizations, morbidity, and mortality worldwide. A growing number of studies on AP pathogenesis are based on cerulein-induced experimental model, which simulates human AP in vivo. It has been demonstrated that both pancreatic acinar cells and peritoneal macrophages are involved in pancreatic inflammation and damage. However, their connection has not been well understood. A cerulein-induced AP model was established on the pancreatic acinar cell line AR42J. Rat macrophages were isolated from the peritoneal cavity. The effects of cerulein-induced pancreatic exosomes on the peritoneal macrophage and pancreas in vivo and in vitro were examined. The underlying molecular mechanism was investigated by exploring the regulatory role of downstream molecules. We found that exosomes derived from cerulein-treated AR42J cells induced rat peritoneal macrophage M1 polarization and pyroptosis. miR-24-3p was upregulated in cerulein-stimulated exosomes, whereas the miR-24-3p inhibitor counteracted the effect of pancreatic exosomes on peritoneal macrophage M1 polarization and pyroptosis. Furthermore, miR-24-3p inhibited March3 expression, whereas MARCH3 mediated NLRP3 ubiquitination in rat peritoneal macrophages, which, in turn, contributed to the apoptosis, reactive oxygen species production, and inflammation in AR42J cells. Exosomes derived from cerulein-stimulated pancreatic acinar cells mediate peritoneal macrophage M1 polarization and pyroptosis via an miR-24-3p/MARCH3/NLRP3 axis in AP.

Effectiveness of minimally invasive surgery in the treatment of pancreatic acinar cell carcinoma

Objective: To explore the effectiveness of minimally invasive surgical treatment for pancreatic acinar cell carcinoma (PACC). Methods: Six patients with PACC diagnosed in Peking University Third Hospital from January 2010 to September 2022 were retrospectively selected. Preoperative evaluation was performed on whether the lesions were eligible for surgery, including whether radical resection of liver metastases could be performed. Laparoscopic or Da Vinci robot-assisted resection was performed on six patients, and spleen retention was determined according to the original lesions and the relationship with peripheral blood vessels and tissues, while simultaneous resection was performed on cases of peripheral organ tissue invasion. The patients' basic information, preoperative general conditions, preoperative diagnosis and tumor stage, minimally invasive surgery methods, postoperative complications, pathological results, tumor stage and follow-up data were collected and analyzed to explore the effectiveness of minimally invasive surgery. Results: Among the six patients, four were males and two were females, with the age of 25-69 years. Five patients had abdominal pain and distension before surgery, five patients had tumors located at the tail of the pancreatic body, and one patient had tumors located at the head of the pancreas. Preoperative imaging (enhanced CT and MRI) was performed to measure the tumor diameter (2.8-10.0 cm). Tumor markers were elevated in two patients before surgery, and six patients underwent surgery through laparoscopy or robotic platform. No complications such as postoperative pancreatic fistula and bleeding were clinically relevant during and after surgery. There were two cases with concurrent or heterochronous liver metastasis, two cases with lymph node metastasis and nodular metastasis, four cases with tumor invasion of surrounding organs (stomach, spleen or duodenum), and three cases with vascular cancer thrombi. The follow-up time of the six patients was 12 to 165 months, and one patient underwent three operations due to postoperative liver metastasis and residual pancreatic recurrence, and the results were satisfactory. All the six patients survived at the last follow-up. Conclusions: PACC is prone to invade the surrounding organs, and has a large tumor diameter. Radical surgery for PACC can be completed through minimally invasive surgery, and satisfactory oncology prognosis can be obtained. In addition, some PACC patients with recurrence and metastasis can still be treated by surgery.

Metaplastic regeneration in the mouse stomach requires a reactive oxygen species pathway

In pyloric metaplasia, mature gastric chief cells reprogram via an evolutionarily conserved process termed paligenosis to re-enter the cell cycle and become spasmolytic polypeptide-expressing metaplasia (SPEM) cells. Here, we use single-cell RNA sequencing (scRNA-seq) following injury to the murine stomach to analyze mechanisms governing paligenosis at high resolution. Injury causes induced reactive oxygen species (ROS) with coordinated changes in mitochondrial activity and cellular metabolism, requiring the transcriptional mitochondrial regulator Ppargc1a (Pgc1α) and ROS regulator Nf2el2 (Nrf2). Loss of the ROS and mitochondrial control in Ppargc1a-/- mice causes the death of paligenotic cells through ferroptosis. Blocking the cystine transporter SLC7A11(xCT), which is critical in lipid radical detoxification through glutathione peroxidase 4 (GPX4), also increases ferroptosis. Finally, we show that PGC1α-mediated ROS and mitochondrial changes also underlie the paligenosis of pancreatic acinar cells. Altogether, the results detail how metabolic and mitochondrial changes are necessary for injury response, regeneration, and metaplasia in the stomach.

Abstract 5325: Cytokine CCL9, new downstream target of oncogene Kras to modulate acinar-to-ductal metaplasia during pancreatic cancer initiation

Abstract Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal types of cancer with a current five-year survival rate at 12% in the US, and is dominated by oncogenic Kras mutations. Accumulating evidence showed that PDAC can be originated from acinar-to-ductal metaplasia (ADM). In this process, pancreatic acini harboring oncogenic Kras mutations are transdifferentiated to a duct-like phenotype that further progresses to become pancreatic intraepithelial neoplasia (PanIN) lesions, giving rise to PDAC. Although ADM formation is commonly observed in KrasG12D transgenic mouse models of PDAC, the exact mechanisms of how oncogenic KrasG12D modulates this process remain unclear. Here, we unveil a new downstream target of oncogenic Kras, cytokine CCL9 during ADM formation. Either exogenously treating primary pancreatic acini with recombinant CCL9 or ectopically expressing CCL9 in primary pancreatic acini resulted in ADM formation in a 3D organoid culture system. Furthermore, knockdown of CCL9 in KrasG12D-expressed acini reduced KrasG12D-induced ADM. Higher levels of reactive oxygen species (ROS) were detected in the KrasG12D-expressed pancreatic acini, and knockdown of CCL9 in these acini decreased ROS levels. Overexpression of CCL9 in primary pancreatic acini elevated cellular ROS of pancreatic acinar cells. Furthermore, depletion of ROS in 3D organoid culture of CCL9-expressed acini reduced CCL9-induced ADM formation. In transgenic mice of p48cre:KrasG12D, blockade of CCL9 through its specific neutralizing antibody attenuated ADM formation as well as PanIN structures. Altogether, our results indicated oncogenic KrasG12D initiates PDAC through a new downstream target molecule, CCL9, to enhance ROS levels of pancreatic acinar cells. Citation Format: Geou-Yarh (Stancy) Liou, Justin K. Messex, Crystal J. Byrd. Cytokine CCL9, new downstream target of oncogene Kras to modulate acinar-to-ductal metaplasia during pancreatic cancer initiation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5325.

Abstract 3378: Acinar cell susceptibility for tumor development in the pancreas: Targeting the critical regulators for therapy

Abstract Acinar to ductal metaplasia (ADM) or acinar cell dedifferentiation is one of the most notable features of chronic pancreatitis. It is also considered the initial step of pancreatic cancer development when oncogenic mutations accumulate. However, its precise mechanism and regulatory pathways remain unclear. This study profiled the transcriptome of dedifferentiated acinar cells from both humans and mice. Differential expression analysis integrated with a systematic literature search revealed several potential regulators (e.g., GDF15, GFRA1, MXRA5, CXCL17), of which the expression pattern over time was validated by qRT-PCR. We also focused on multiple conserved pathways of cell survival, among which SLC7A11 (xCT) is transiently upregulated in both species. The xCT subunit, integral to the cystine/glutamate antiporter system xc-, plays a critical role in cell survival across various cell types. To elucidate its role in acinar dedifferentiation, we employed gene silencing, pharmacological inhibition, and a knock-out mouse model. Acinar cells with diminished xCT function exhibit increased ferroptosis linked to lipid peroxidation. Antioxidants such as N-acetylcysteine and ferroptosis inhibitors like Ferrostatin-1 can mitigate lower glutathione levels and lipid ROS accumulation. Similarly, in mouse acute pancreatitis, xCT prevents lipid peroxidation. Our findings indicate that xCT fuels the glutathione pool and maintains ROS balance during ADM, thereby preventing pancreatic acinar cells from ferroptosis, a form of cell death rarely reported in relation to ADM. Next to xCT, a future and similar comprehensive study of the aforementioned genes could provide valuable insights into the dynamic changes that occur during acinar cell dedifferentiation and pave the way to the discovery of new intervention targets to suppress tumorigenesis in chronic pancreatitis. Citation Format: Zhaolong Pan, Jan-Lars Van den Bossche, Eva Rodriguez-Aznar, Pauline Janssen, Olaya Lara, Gamze Ates, Ann Massie, Diedert Luc De Paep, Isabelle Houbracken, Marco Mambretti, Ilse Rooman. Acinar cell susceptibility for tumor development in the pancreas: Targeting the critical regulators for therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3378.