Caerulein-induced Pancreatitis Research Articles

Pancreas-directed AAV8-hSPINK1 gene therapy safely and effectively protects against pancreatitis in mice

ObjectiveCurrently, there is no cure for chronic pancreatitis (CP). Germline loss-of-function variants in SPINK1 (encoding trypsin inhibitor) are common in patients with CP and are associated with acute attacks and...

Abstract 4267: Inflammation mediated expression of integrin αvβ3 potentiates KRAS driven PDAC oncogenesis

Abstract Understanding how pancreatic epithelial cells respond to inflammation and why some become tumorigenic and aggressive could identify new methods of early detection and novel therapeutic targets to control disease progression. Here, we leveraged an in vivo model in which mice with pancreas-specific expression of mutant Kras progress from pre-malignant intraepithelial neoplasias (PanINs) to pancreatic ductal adenocarcinoma (PDAC) following caerulein-induced chronic pancreatitis. During the transition from PanINs to PDAC, we observe two classes of neoplastic lesions: classically-described PanIN tubular structures that are highly organized and previously uncharacterized atypical highly disorganized lesions that are marked by robust integrin αvβ3 expression, observations which are consistent with human pancreatitis and PDAC. In vitro, we report that immune-, stroma-, and tumor cell-derived inflammatory cytokines induce β3 expression in a STAT3-dependent manner leading to increased integrin αvβ3. Considering that we previously established αvβ3 as a driver of cancer cell stress tolerance, drug resistance, and a stem-like phenotype, the acquisition of αvβ3 during chronic inflammation may promote tumorigenesis. Indeed, using the spontaneous tumor mouse model and xenograft models of human PDAC, we find that cells with an inability to upregulate β3 during tumor initiation fail to form tumors. These findings implicate integrin αvβ3 as a critical driver of inflammation-mediated pancreatic tumorigenesis. Citation Format: Ryan M. Shepard, Alejandro D. Campos, Zachary M. Ortega, Hiromi I. Wettersten, Tami Von Schalscha, Sara M. Weis, David A. Cheresh. Inflammation mediated expression of integrin αvβ3 potentiates KRAS driven PDAC oncogenesis [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 4267.

Interleukin-22 Alleviates Caerulein-Induced Acute Pancreatitis by Activating AKT/mTOR Pathway.

Acute pancreatitis (AP) is one of the most common acute abdominal disorders; due to the lack of specific treatment, the treatment of acute pancreatitis, especially serious acute pancreatitis (SAP), is difficult and challenging. We will observe the changes of Interleukin -22 levels in acute pancreatitis animal models, and explore the mechanism of Interleukin -22 in acute pancreatitis. This study aims to assess the potential protective effect of Interleukin -22 on caerulein-induced acute pancreatitis and to explore its mechanism. Blood levels of amylase and lipase and Interleukin -22 were assessed in mice with acute pancreatitis. In animal model and cell model of caerulein-induced acute pancreatitis, the mRNA levels of P62 and Beclin-1 were determined using PCR, and the protein expression of P62, LC3-II, mTOR, AKT, p-mTOR, and p-AKT were evaluated through Western blot analysis. Interleukin -22 administration reduced blood amylase and lipase levels and mitigated tissue damage in acute pancreatitis mice model. Interleukin -22 inhibited the relative mRNA levels of P62 and Beclin-1, and the Interleukin -22 group showed a decreased protein expression of LC3-II and P62 and the phosphorylation of the AKT/mTOR pathway. Furthermore, we obtained similar results in the cell model of acute pancreatitis. This study suggests that Interleukin -22 administration could alleviate pancreatic damage in caerulein-induced acute pancreatitis. This effect may result from the activation of the AKT/mTOR pathway, leading to the inhibition of autophagy. Consequently, Interleukin -22 shows potential as a treatment.

Abstract C094: Tff2 defines pancreatic TA progenitors that are protective against Kras-driven carcinogenesis

Abstract Background: Progenitor or stem cells play a pivotal role in maintaining tissue homeostasis and are often the source of tumor progression. While genuine stem cells are absent in the pancreatic exocrine organ, a variety of progenitor cells are present that contribute to normal homeostasis and regeneration process post-injury. In a previous study, we identified a pancreatic facultative progenitor (FP) that was marked by the gene doublecortin kinase like-1 (Dclk1). These Dclk1+ FP are rare, long-lived, quiescent cells that are critical for regeneration following injury. In contrast, cells marked by Trefoil factor 2 (Tff2) are active progenitors within the gastrointestinal tract, playing a crucial role in homeostasis and tumor progression. However, the role of Tff2+ cells within the pancreas remains relatively underexplored. Method & Results: To explore the function of Tff2 cells, we developed a Tff2 BAC transgenic mouse line (Tff2-DTR-CreERT2), which was then crossed with report mice. Employing lineage tracing and single cell RNA sequencing, we uncovered a new type of progenitor, distinct from the FP - Tff2+ transit amplifying progenitors (TAP). Tff2+ acinar cells showed much greater proliferation (>100-fold) than all other acinar cells. Single cell RNA-seq of the Tff2-expressing lineage showed that one cluster (cluster 0) identified acinar cells with a low differentiated, progenitor-like state. These cells were found to contribute to tissue homeostasis, exhibiting expansion in the early stages, giving rise to more differentiated acinar cells, peaking around six months, and then gradually diminishing until they disappeared. To understand the role of Tff2 in PanIN lesions, we developed a compound mouse model: Tff2-DTR-CreERT; Mist1-CreERT; KrasG12D(TMK). When Tff2+ cells were then ablated using Diphtheria Toxin in the setting of caerulein-induced pancreatitis, the TMK mice developed significantly more PanIN lesions. Tff2-CreERT; KrasG12D mice were generally more resistant to developing advanced PanINs, but in the setting of caerulein pancreatitis, the Tff2; KrasG12D model developed more pancreatic ductal adenocarcinoma (PDAC). To explore the role of the TFF2 peptide in tumor progression, we used an adenovirus expressing TFF2 (Ad-TFF2), and introduced it through tail vein injections. We found that Ad-Tff2 helped prevent the development of advanced PanIN lesions. Conclusion: In summary, our research indicated that Tff2 cells play a significant role in maintaining pancreatic homeostasis and act in paracrine fashion to suppress the development of pancreatic adenocarcinoma. Citation Format: Feijing Wu, Zhengyu Jiang, Xiaofei Zhi, Yosuke Ochiai, Ruth A. White, Ermanno Malagola, Jin Qian, Biyun Zheng, Ruhong Tu, Hiroki Kobayashi, Quin Waterbury, Leah B. Zamechek, Timothy C. Wang. Tff2 defines pancreatic TA progenitors that are protective against Kras-driven carcinogenesis [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Pancreatic Cancer; 2023 Sep 27-30; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(2 Suppl):Abstract nr C094.

Abstract C080: Decoding the MYC-PDAC nexus: Unveiling the impact of PP2A-C subunit methylation on tumor onset and advancement

Abstract Introduction: Persistent disruptions in MYC regulation and the inactivation of the serine/threonine protein phosphatase 2A (PP2A) are prevalent among individuals diagnosed with pancreatic ductal adenocarcinoma (PDAC). While it is known that PP2A influences MYC activity by facilitating its degradation, the precise mechanism by which this occurs remains unclear. Small molecule activators PP2A (SMAPs) regulate the holoenzyme assembly consisting of a scaffolding-A subunit and a catalytic-C subunit, and over 40 distinct substrate directing regulatory-B subunits. PP2A heterotrimerization is regulated by post-translational modifications of the catalytic C subunit, notably methylation of L309, which impacts B subunit binding affinity to the AC dimer and thus the substrate selectivity. In this study, we aim to investigate whether the deregulation of MYC, attributed to the loss of PP2A-C subunit methylation, contributes to the initiation and progression of PDAC. Methods: We investigated the influence of SMAPs on MYC’s stability using clinical specimens, cellular, and mouse models. Clinical specimens consisted of tissue microarray (TMA) from patients with pancreatitis and PDAC. Cellular models involved ex-vivo acinar cell assays and proximity ligation assays. Mouse models included caerulein-induced pancreatitis, genetic mouse models like Ptf1a-Cre; LSL-KRASG12D (KC) and LSL-Kras G12D;P53 R172H/+;Pdx1-Cre (KPC), along with pharmacological inhibition studies with SMAPs. Results and discussion: Our TMA data revealed a consistent pattern of Methyl-PP2A-C (Methyl-C) loss and an increase in the stable and active phosphoform of MYC (p62MYC) throughout the sequential stages of PDAC progression, starting from normal tissue and advancing to acinar-to-ductal metaplasia, PanINs, and finally PDAC. In ex-vivo ADM assays, treatment with small molecule activators of PP2A (SMAPs), specifically DT061, significantly reduced the transformation of acinar cells to duct-like cells compared to untreated cells from both wild-type (WT) and KC mice. This suggests that PP2A plays a crucial role in the early events that drive the formation of pancreatic lesions. Moreover, SMAPs demonstrated the ability to alleviate caerulein-induced inflammation in normal mice, as well as suppress the development of pancreatic PanIN lesions and collagen deposition in caerulein-treated KC mice. These effects were associated with decreased levels of p62MYC and increased Methyl-C staining in both WT and KC mice treated with caerulein. Additionally, in KPC mice, treatment with DT061 resulted in reduced PDAC tumor growth, accompanied by decreased levels of p62MYC. Conclusion: Our results provide compelling evidence that small molecule activators of PP2A (SMAPs), through their ability to downregulate MYC, hold significant promise as a therapeutic strategy for individuals at high risk of developing PDAC, particularly those with chronic pancreatitis. This approach has the potential to effectively prevent PDAC development and mitigate its onset in this high-risk population. Citation Format: Vidhi Shah, Alexander Smith, Gabriel Cohn, Motoyuki Tsuda, Goutham Narla, Jonathan Brody, Brett Sheppard, Rosalie Sears. Decoding the MYC-PDAC nexus: Unveiling the impact of PP2A-C subunit methylation on tumor onset and advancement [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Pancreatic Cancer; 2023 Sep 27-30; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84(2 Suppl):Abstract nr C080.

Tectoridin alleviates caerulein-induced severe acute pancreatitis by targeting ERK2 to promote macrophage M2 polarization

Severe acute pancreatitis (SAP) is an inflammatory disease of the pancreas with a high mortality rate. Macrophages play a crucial role in the pathogenesis of pancreatitis. Tectoridin (Tec) is a highly active isoflavone with anti-inflammatory pharmacological activity. However, the role of Tec in the SAP process is not known. The purpose of this study was to investigate the therapeutic effect and potential mechanism of Tec on SAP. To establish SAP mice by intraperitoneal injection of caerulein and Lipopolysaccharide (LPS), the role of Tec in the course of SAP was investigated based on histopathology, biochemical indicators of amylase and lipase and inflammatory factors. The relationship between Tec and macrophage polarization was verified by immunofluorescence, real-time quantitative PCR and Western blot analysis. We then further predicted the possible targets and signal pathways of action of Tec by network pharmacology and molecular docking, and validated them by in vivo and in vitro. In this study, we demonstrated that Tec significantly reduced pancreatic injury in SAP mice, and decreased serum levels of amylase and lipase. The immunofluorescence and Western blot analysis showed that Tec promoted macrophage M2 polarization. Network pharmacology and molecular docking predicted that Tec may target ERK2 for the treatment of SAP, and in vivo and in vitro experiments proved that Tec inhibited the ERK MAPK signal pathway. In summary, Tec can target ERK2, promote macrophage M2 polarization and attenuate pancreatic injury, Tec may be a potential drug for the treatment of SAP.

Decreased neutrophil counts prolong inflammation in acute pancreatitis and cause inflammation spillover to distant organs

Background/objectiveAcute pancreatitis is an aseptic inflammation caused by pathologically activated pancreatic enzymes and inflammatory mediators produced secondarily by neutrophils and other inflammatory cells and is one of the most difficult diseases to treat. This study aimed to investigate the role of neutrophils in pancreatitis by examining tissue dynamics. MethodsWe created a model of caerulein-induced pancreatitis in 12-week-old male granulocyte colony-stimulating factor knockout mice (G–CSF–KO) and wild-type littermate control mice (six intraperitoneal injections of caerulein [80 μg/kg body weight] at hourly intervals for 2 days). Mice were sacrificed 0, 3, 6, 12, 24, 36, 48, 72, and 168 h after caerulein administration and examined histologically. ResultsThe survival rate after one week of caerulein administration was 100 % in the control mice, whereas it was significantly lower (10 %) in the G–CSF–KO mice. Histological examination revealed significant hemorrhage and inflammatory cell migration in the G–CSF–KO mice, indicating prolonged inflammation. ConclusionProlonged inflammation was observed in the G–CSF–KO mice. Tissue cleanup by neutrophils during the acute phase of inflammation may influence healing through the chronic phase.

Galantamine ameliorates experimental pancreatitis

BackgroundAcute pancreatitis is a common and serious inflammatory condition currently lacking disease modifying therapy. The cholinergic anti-inflammatory pathway (CAP) is a potent protective anti-inflammatory response activated by vagus nerve-dependent α7 nicotinic acetylcholine receptor (α7nAChR) signaling using splenic CD4+ T cells as an intermediate. Activating the CAP ameliorates experimental acute pancreatitis. Galantamine is an acetylcholinesterase inhibitor (AChEI) which amplifies the CAP via modulation of central muscarinic ACh receptors (mAChRs). However, as mAChRs also activate pancreatitis, it is currently unknown whether galantamine would be beneficial in acute pancreatitis.MethodsThe effect of galantamine (1–6 mg/kg-body weight) on caerulein-induced acute pancreatitis was evaluated in mice. Two hours following 6 hourly doses of caerulein (50 µg/kg-body weight), organ and serum analyses were performed with accompanying pancreatic histology. Experiments utilizing vagotomy, gene knock out (KO) technology and the use of nAChR antagonists were also performed.ResultsGalantamine attenuated pancreatic histologic injury which was mirrored by a reduction in serum amylase and pancreatic inflammatory cytokines and an increase the anti-inflammatory cytokine IL-10 in the serum. These beneficial effects were not altered by bilateral subdiaphragmatic vagotomy, KO of either choline acetyltransferase+ T cells or α7nAChR, or administration of the nAChR ganglionic blocker mecamylamine or the more selective α7nAChR antagonist methyllycaconitine.ConclusionGalantamine improves acute pancreatitis via a mechanism which does not involve previously established physiological and molecular components of the CAP. As galantamine is an approved drug in widespread clinical use with an excellent safety record, our findings are of interest for further evaluating the potential benefits of this drug in patients with acute pancreatitis.

Tenascin C deficiency impairs epithelial regeneration after caerulein-induced acute pancreatitis

Tenascin C deficiency impairs epithelial regeneration after caerulein-induced acute pancreatitis

Evaluation of Serum miR-216a, miR-216b, miR-217, miR-92b, miR-375 and miR-148a as Potential Biomarkers for Acute Pancreatitis and the Role of miR-92b in Attenuating Caerulein-induced Injury and Inflammatory Responses in AR42J Cells.

Acute pancreatitis can eventually lead to morbidity and mortality. The present study aimed to identify the differentially expressed microRNAs (miRNAs) that are related to acute pancreatitis and explore the in vitro functional role of miR-92b in acute pancreatitis. Bioinformatics analysis was used to identify differentially expressed miRNAs in caerulein- induced acute pancreatitis samples when compared to normal controls. The role of miR-92b in acute pancreatitis was examined by in vitro functional assays. MiRNA-network analysis revealed 12 miRNAs that function as "core regulatory miRNAs". Further validation studies revealed that six miRNAs (miR-216a, miR-216b, miR-217, miR- 92b, miR-375 and miR-148a) were differentially expressed in the serum samples from patients with acute pancreatitis. These six miRNAs have fair diagnostic potential for severe acute pancreatitis. Caerulein induced cell injury and inflammatory response and repressed miR-92b expression in AR42J cells. MiR-92b overexpression attenuated caerulein-induced cell injury and inflammatory responses in AR42J cells. Luciferase reporter assay showed that mitogen-activated protein kinase 4 (MAP2K4) was a direct target of miR-92b. MiR-92b overexpression repressed MAP2K4 expression, while caerulein up-regulated MAP2K4 expression in AR42J cells. The rescue experiments showed that enforced expression of MAP2K4 partially reversed the miR-92b-mediated protective effects on caerulein-induced AR42J cell injury. In conclusion, we identified miR-216a, miR-216b, miR217, miR-92b, miR-375 and miR-148a as new candidate biomarkers for acute pancreatitis. Further in vitro functional studies revealed that miR-92b attenuated caerulein-induced cell injury and inflammatory responses in AJ42R cells partially via targeting MAP2K4.

Anti-Necroptotic Effects of Itaconate and its Derivatives.

Itaconate is an unsaturated dicarboxylic acid that is derived from the decarboxylation of the Krebs cycle intermediate cis-aconitate and has been shown to exhibit anti-inflammatory and anti-bacterial/viral properties. But the mechanisms underlying itaconate's anti-inflammatory activities are not fully understood. Necroptosis, a lytic form of regulated cell death (RCD), is mediated by receptor-interacting protein kinase 1 (RIPK1), RIPK3, and mixed lineage kinase domain-like protein (MLKL) signaling. It has been involved in the pathogenesis of organ injury in many inflammatory diseases. In this study, we aimed to explore whether itaconate and its derivatives can inhibit necroptosis in murine macrophages, a mouse MPC-5 cell line and a human HT-29 cell line in response to different necroptotic activators. Our results showed that itaconate and its derivatives dose-dependently inhibited necroptosis, among which dimethyl itaconate (DMI) was the most effective one. Mechanistically, itaconate and its derivatives inhibited necroptosis by suppressing the RIPK1/RIPK3/MLKL signaling and the oligomerization of MLKL. Furthermore, DMI promoted the nuclear translocation of Nrf2 that is a critical regulator of intracellular redox homeostasis, and reduced the levels of intracellular reactive oxygen species (ROS) and mitochondrial superoxide (mtROS) that were induced by necroptotic activators. Consistently, DMI prevented the loss of mitochondrial membrane potential induced by the necroptotic activators. In addition, DMI mitigated caerulein-induced acute pancreatitis in mice accompanied by reduced activation of the necroptotic signaling in vivo. Collectively, our study demonstrates that itaconate and its derivatives can inhibit necroptosis by suppressing the RIPK1/RIPK3/MLKL signaling, highlighting their potential applications for treating necroptosis-associated diseases.

Pancreatic acinar cell fate relies on system xC- to prevent ferroptosis during stress

Acinar cell dedifferentiation is one of the most notable features of acute and chronic pancreatitis. It can also be the initial step that facilitates pancreatic cancer development. In the present study, we further decipher the precise mechanisms and regulation using primary human cells and murine experimental models. Our RNAseq analysis indicates that, in both species, early acinar cell dedifferentiation is accompanied by multiple pathways related to cell survival that are highly enriched, and where SLC7A11 (xCT) is transiently upregulated. xCT is the specific subunit of the cystine/glutamate antiporter system xC-. To decipher its role, gene silencing, pharmacological inhibition and a knock-out mouse model were used. Acinar cells with depleted or reduced xCT function show an increase in ferroptosis relating to lipid peroxidation. Lower glutathione levels and more lipid ROS accumulation could be rescued by the antioxidant N-acetylcysteine or the ferroptosis inhibitor ferrostatin-1. In caerulein-induced acute pancreatitis in mice, xCT also prevents lipid peroxidation in acinar cells. In conclusion, during stress, acinar cell fate seems to be poised for avoiding several forms of cell death. xCT specifically prevents acinar cell ferroptosis by fueling the glutathione pool and maintaining ROS balance. The data suggest that xCT offers a druggable tipping point to steer the acinar cell fate in stress conditions.

Sex-dependent modulation of caerulein-induced acute pancreatitis in C57BL/6J mice: histopathological and immunohistochemical characterization

Sex-dependent modulation of caerulein-induced acute pancreatitis in C57BL/6J mice: histopathological and immunohistochemical characterization

Optogenetic stimulation of the brainstem dorsal motor nucleus ameliorates acute pancreatitis.

Inflammation is an inherently self-amplifying process, resulting in progressive tissue damage when unresolved. A brake on this positive feedback system is provided by the nervous system which has evolved to detect inflammatory signals and respond by activating anti-inflammatory processes, including the cholinergic anti-inflammatory pathway mediated by the vagus nerve. Acute pancreatitis, a common and serious condition without effective therapy, develops when acinar cell injury activates intrapancreatic inflammation. Prior study has shown that electrical stimulation of the carotid sheath, which contains the vagus nerve, boosts the endogenous anti-inflammatory response and ameliorates acute pancreatitis, but it remains unknown whether these anti-inflammatory signals originate in the brain. Here, we used optogenetics to selectively activate efferent vagus nerve fibers originating in the brainstem dorsal motor nucleus of the vagus (DMN) and evaluated the effects on caerulein-induced pancreatitis. Stimulation of the cholinergic neurons in the DMN significantly attenuates the severity of pancreatitis as indicated by reduced serum amylase, pancreatic cytokines, tissue damage, and edema. Either vagotomy or silencing cholinergic nicotinic receptor signaling by pre-administration of the antagonist mecamylamine abolishes the beneficial effects. These results provide the first evidence that efferent vagus cholinergic neurons residing in the brainstem DMN can inhibit pancreatic inflammation and implicate the cholinergic anti-inflammatory pathway as a potential therapeutic target for acute pancreatitis.

Celastrol inhibits necroptosis by attenuating the RIPK1/RIPK3/MLKL pathway and confers protection against acute pancreatitis in mice.

Necroptosis is a necrotic form of regulated cell death, which is primarily mediated by the receptor-interacting protein kinase 1 (RIPK1), RIPK3, and mixed lineage kinase domain-like (MLKL) pathway in a caspase-independent manner. Necroptosis has been found to occur in virtually all tissues and diseases evaluated, including pancreatitis. Celastrol, a pentacyclic triterpene extracted from the roots of Tripterygium wilfordii (thunder god vine), possesses potent anti-inflammatory and anti-oxidative activities. Yet, it is unclear whether celastrol has any effects on necroptosis and necroptotic-related diseases. Here we showed that celastrol significantly suppressed necroptosis induced by lipopolysaccharide (LPS) plus pan-caspase inhibitor (IDN-6556) or by tumor-necrosis factor-α in combination with LCL-161 (Smac mimetic) and IDN-6556 (TSI). In these in vitro cellular models, celastrol inhibited the phosphorylation of RIPK1, RIPK3, and MLKL and the formation of necrosome during necroptotic induction, suggesting its possible action on upstream signaling of the necroptotic pathway. Consistent with the known role of mitochondrial dysfunction in necroptosis, we found that celastrol significantly rescued TSI-induced loss of mitochondrial membrane potential. TSI-induced intracellular and mitochondrial reactive oxygen species (mtROS), which are involved in the autophosphorylation of RIPK1 and recruitment of RIPK3, were significantly attenuated by celastrol. Moreover, in a mouse model of acute pancreatitis that is associated with necroptosis, celastrol administration significantly reduced the severity of caerulein-induced acute pancreatitis accompanied by decreased phosphorylation of MLKL in pancreatic tissues. Collectively, celastrol can attenuate the activation of RIPK1/RIPK3/MLKL signaling likely by attenuating mtROS production, thereby inhibiting necroptosis and conferring protection against caerulein-induced pancreatitis in mice.

Use of a dual genetic system to decipher exocrine cell fate conversions in the adult pancreas

Unraveling cell fate plasticity during tissue homeostasis and repair can reveal actionable insights for stem cell biology and regenerative medicine. In the pancreas, it remains controversial whether lineage transdifferentiation among the exocrine cells occur under pathophysiological conditions. Here, to address this question, we used a dual recombinase-mediated genetic system that enables simultaneous tracing of pancreatic acinar and ductal cells using two distinct genetic reporters, avoiding the “ectopic” labeling by Cre-loxP recombination system. We found that acinar-to-ductal transdifferentiation occurs after pancreatic duct ligation or during caerulein-induced pancreatitis, but not during homeostasis or after partial pancreatectomy. On the other hand, pancreatic ductal cells contribute to new acinar cells after significant acinar cell loss. By genetic tracing of cell proliferation, we also quantify the cell proliferation dynamics and deduce the turnover rate of pancreatic exocrine lineages during homeostasis. Together, these results suggest that the lineage transdifferentiation happens between acinar cells and ductal cells in the pancreatic exocrine glands under specific conditions.

The Kruppel-like factor 4-signal transducer and activator of transcription 5A axis promotes pancreatic fibrosis in mice with caerulein-induced chronic pancreatitis.

Pancreatic fibrosis (PF) is a hallmark of chronic pancreatitis (CP), but its molecular mechanism remains unclear. This study was conducted to explore the role of Kruppel-like factor 4 (KLF4) in PF in CP mice. The CP mouse model was established using caerulein. After KLF4 interference, pathological changes in pancreatic tissues and fibrosis degree were observed by hematoxylin-eosin staining and Masson staining, and levels of Collagen I, Collagen III, and alpha-smooth muscle actin, inflammatory cytokines, KLF4, signal transducer and activator of transcription 5A (STAT5) in pancreatic tissues were measured by enzyme-linked immunosorbent assay, quantitative real-time polymerase chain reaction, Western blot assay, and immunofluorescence. The enrichment of KLF4 on the STAT5 promoter and the binding of KLF4 to the STAT5 promoter were analyzed. The rescue experiments were performed by co-injection of sh-STAT5 and sh-KLF4 to confirm the regulatory mechanism of KLF4. KLF4 was upregulated in CP mice. Inhibition of KLF4 effectively attenuated pancreatic inflammation and PF in mice. KLF4 was enriched on the STAT5 promoter and enhanced the transcriptional and protein levels of STAT5. Overexpression of STAT5 reversed the inhibitory role of silencing KLF4 in PF. In summary, KLF4 promoted the transcription and expression of STAT5, which further facilitated PF in CP mice.

Effect of lncRNA H19 targeting miR-30a-5p on cell apoptosis, secretion of inflammatory factors, and autophagy in caerulein-induced acute pancreatitis

Effect of lncRNA H19 targeting miR-30a-5p on cell apoptosis, secretion of inflammatory factors, and autophagy in caerulein-induced acute pancreatitis

Biochanin A ameliorates caerulein-induced acute pancreatitis and associated intestinal injury in mice by inhibiting TLR4 signaling

Acute pancreatitis (AP) is an acute inflammatory abdominal disease frequently associated with intestinal barrier dysfunction. Biochanin A (BCA), a dietary isoflavone, has gained increasing interest with its pronounced biological activities. However, its potential beneficial effects on AP have not been demonstrated. Herein, we explored the protective effect of BCA on caerulein-induced AP in BALB/c mice and underlying mechanisms. BCA alleviated AP as evidenced by reduced serum amylase and lipase levels, pancreatic edema, pancreatic myeloperoxidase activity, and improved pancreatic morphology. Amelioration of pancreatic damage by BCA was associated with reduced levels of tumor necrosis factor-α, interleukin (IL)-1β, IL-6, and monocyte chemotactic protein-1 in both pancreas and colon. Moreover, BCA attenuated AP-associated barrier damage by upregulating the expression of tight junction proteins zonulin occluding (ZO)-1, ZO-2, occludin, and claudin-1. Concomitantly, the translocation of pathogenic bacteria Escherichia coli (E. coli) to pancreas was reduced by BCA. More importantly, reduction of E. coli dissemination by BCA inhibited the TLR4-MAPK/NF-κB signaling and NLRP3 inflammasome activation, thereby protecting against AP and related intestinal injury. Consistently, TLR4 inhibition by TAK-242 pre-treatment counteracted the anti-inflammatory effects of BCA in acinar cells. Taken together, our study extends beneficial effects of BCA to AP prevention, and dietary BCA supplement may be a potential strategy to safeguard AP.

Abstract A053: Inhibition of the MEK5-ERK5 signaling pathway blocks pancreatic tumorigenesis

Abstract Mutant KRAS represents one of the most common driving events in pancreatic ductal adenocarcinomas (PDAC), yet therapies targeting this pathway have proven largely ineffective. We identified ERK5 activation as critical component of KRAS-driven pancreatic tumorigenesis. ERK5 belongs to the MAP kinase family and is phosphorylated and activated by the MAPK kinase MEK5. Therefore, the aim of this study is to elucidate the function of the MEK5-ERK5 signaling axis in PDAC pathogenesis. Several novel genetically engineered mouse models with pancreas-specific deletion or activation of the MEK5-ERK5 signaling were generated to study pancreatic cancer initiation and progression. Caerulein-induced pancreatitis was performed in p48Cre/+ KrasG12D/+ (Kras) and p48Cre/+ KrasG12D/+ Erk5fl/fl (Kras;Erk5) mice to study early steps in pancreatic tumorigenesis. To investigate if ERK5 ablation impacts tumor progression a two-stage tumor model utilizing dual recombinase (Cre/Flp) mouse model was utilized. Next, the impact of ERK5 depletion or pharmacological inhibition on survival of pancreatic cancer mice was analyzed. Furthermore, human patient-derived PDAC xenograft models in immunodeficient mice were utilized to evaluate efficacy of a small molecule ERK5 inhibitor which does not exhibit off-target activity on bromodomains. Deletion of ERK5 or inhibition of its kinase activity resulted in blocked tumorigenesis in Kras mutant mice using caerulein-induced pancreatitis. Upon pancreatitis induction pancreas in Kras;Erk5 mice exhibit normal tissue architecture with significantly fewer proliferating cells compared to control mice. Intriguingly, pancreas-specific expression of constitutively active MEK5 (MEK5DD) even without oncogenic Kras mutations was sufficient in promoting development of precancerous lesions after pancreatitis induction. Next, we observed that genetic depletion or pharmacological inhibition of ERK5 significantly prolonged survival of the KPC PDAC mouse model, whereas expression of MEK5DD in KPC mice resulted in significantly accelerated tumorigenesis and reduced survival compared to control mice. Furthermore, depletion of ERK5 in already established PDAC resulted in significantly prolonged survival and reduced tumor burden indicating an important role of ERK5 in tumor maintenance. Finally, ERK5 inhibitor treatment of patient-derived PDAC xenografts in immunocompromised mice resulted in tumor growth arrest. Our studies demonstrate that the MEK5-ERK5 signaling pathway plays an important role in pancreatic cancer initiation and progression. Deletion of ERK5 or pharmacological inhibition of ERK5 kinase activity results in significantly reduced tumor burden and prolonged survival in autochthonous PDAC models and patient-derived xenografts. Conversely, activation of MEK5-ERK5 signaling is sufficient to drive pancreatic tumorigenesis indicating an essential role of the ERK5 pathway as downstream effector of oncogenic KRAS. Further studies will be performed to identify the mechanisms of ERK5 action including downstream kinase targets. Citation Format: Simone Hausmann, Asli Ece Kasapoglu, Ana Benitez Morales, Shane Lofgren, Pawel K. Mazur. Inhibition of the MEK5-ERK5 signaling pathway blocks pancreatic tumorigenesis [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer; 2022 Sep 13-16; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2022;82(22 Suppl):Abstract nr A053.