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1.
J Pathol ; 262(1): 61-75, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-37796386

RESUMO

Pancreatic stellate cells (PSCs) are stromal cells in the pancreas that play an important role in pancreatic pathology. In chronic pancreatitis (CP) and pancreatic ductal adenocarcinoma (PDAC), PSCs are known to get activated to form myofibroblasts or cancer-associated fibroblasts (CAFs) that promote stromal fibroinflammatory reactions. However, previous studies on PSCs were mainly based on the findings obtained using ex vivo expanded PSCs, with few studies that addressed the significance of in situ tissue-resident PSCs using animal models. Their contributions to fibrotic reactions in CP and PDAC are also lesser-known. These limitations in our understanding of PSC biology have been attributed to the lack of specific molecular markers of PSCs. Herein, we established Meflin (Islr), a glycosylphosphatidylinositol-anchored membrane protein, as a PSC-specific marker in both mouse and human by using human pancreatic tissue samples and Meflin reporter mice. Meflin-positive (Meflin+ ) cells contain lipid droplets and express the conventional PSC marker Desmin in normal mouse pancreas, with some cells also positive for Gli1, the marker of pancreatic tissue-resident fibroblasts. Three-dimensional analysis of the cleared pancreas of Meflin reporter mice showed that Meflin+ PSCs have long and thin cytoplasmic protrusions, and are localised on the abluminal side of vessels in the normal pancreas. Lineage tracing experiments revealed that Meflin+ PSCs constitute one of the origins of fibroblasts and CAFs in CP and PDAC, respectively. In these diseases, Meflin+ PSC-derived fibroblasts showed a distinctive morphology and distribution from Meflin+ PSCs in the normal pancreas. Furthermore, we showed that the genetic depletion of Meflin+ PSCs accelerated fibrosis and attenuated epithelial regeneration and stromal R-spondin 3 expression, thereby implying that Meflin+ PSCs and their lineage cells may support tissue recovery and Wnt/R-spondin signalling after pancreatic injury and PDAC development. Together, these data indicate that Meflin may be a marker specific to tissue-resident PSCs and useful for studying their biology in both health and disease. © 2023 The Pathological Society of Great Britain and Ireland.


Assuntos
Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Pancreatite Crônica , Animais , Humanos , Camundongos , Carcinoma Ductal Pancreático/patologia , Fibrose , Pâncreas/patologia , Neoplasias Pancreáticas/patologia , Células Estreladas do Pâncreas/patologia , Pancreatite Crônica/genética , Pancreatite Crônica/metabolismo , Pancreatite Crônica/patologia , Regeneração
2.
Int J Mol Sci ; 24(21)2023 Nov 02.
Artigo em Inglês | MEDLINE | ID: mdl-37958889

RESUMO

Pancreatic ductal adenocarcinoma (PDAC), a highly malignant neoplasm, is classified as one of the most severe and devastating types of cancer. PDAC is a notable malignancy that exhibits a discouraging prognosis and a rising occurrence. The interplay between diabetes and pancreatic cancer exhibits a reciprocal causation. The identified metabolic disorder has been observed to possess noteworthy consequences on health outcomes, resulting in elevated rates of morbidity. The principal mechanisms involve the suppression of the immune system, the activation of pancreatic stellate cells (PSCs), and the onset of systemic metabolic disease caused by dysfunction of the islets. From this point forward, it is important to recognize that pancreatic-cancer-related diabetes (PCRD) has the ability to increase the likelihood of developing pancreatic cancer. This highlights the complex relationship that exists between these two physiological states. Therefore, we investigated into the complex domain of PSCs, elucidating their intricate signaling pathways and the profound influence of chemokines on their behavior and final outcome. In order to surmount the obstacle of drug resistance and eliminate PDAC, researchers have undertaken extensive efforts to explore and cultivate novel natural compounds of the next generation. Additional investigation is necessary in order to comprehensively comprehend the effect of PCRD-mediated apoptosis on the progression and onset of PDAC through the utilization of natural compounds. This study aims to examine the potential anticancer properties of natural compounds in individuals with diabetes who are undergoing chemotherapy, targeted therapy, or immunotherapy. It is anticipated that these compounds will exhibit increased potency and possess enhanced pharmacological benefits. According to our research findings, it is indicated that naturally derived chemical compounds hold potential in the development of PDAC therapies that are both safe and efficacious.


Assuntos
Produtos Biológicos , Carcinoma Ductal Pancreático , Diabetes Mellitus Tipo 2 , Neoplasias Pancreáticas , Humanos , Diabetes Mellitus Tipo 2/metabolismo , Produtos Biológicos/farmacologia , Produtos Biológicos/uso terapêutico , Produtos Biológicos/metabolismo , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/etiologia , Neoplasias Pancreáticas/metabolismo , Carcinoma Ductal Pancreático/tratamento farmacológico , Carcinoma Ductal Pancreático/etiologia , Carcinoma Ductal Pancreático/metabolismo , Células Estreladas do Pâncreas/metabolismo , Microambiente Tumoral , Neoplasias Pancreáticas
3.
Front Immunol ; 14: 1248547, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-38035115

RESUMO

Activation of pancreatic stellate cells (PSCs) to cancer-associated fibroblasts (CAFs) is responsible for the extensive desmoplastic reaction observed in PDAC stroma: a key driver of pancreatic ductal adenocarcinoma (PDAC) chemoresistance leading to poor prognosis. Specialized pro-resolving mediators (SPMs) are prime modulators of inflammation and its resolution, traditionally thought to be produced by immune cells. Using liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based lipid mediator profiling PSCs as well as primary human CAFs express enzymes and receptors to produce and respond to SPMs. Human PSC/CAF SPM secretion profile can be modulated by rendering these cells activated [transforming growth factor beta (TGF-ß)] or quiescent [all-trans retinoic acid (ATRA)]. ATRA-induced nuclear translocation of arachidonate-15-lipoxygenase (ALOX15) was linked to increased production of n-3 docosapentaenoic acid-derived Resolvin D5 (RvD5n-3 DPA), among other SPMs. Inhibition of RvD5n-3 DPA formation increases cancer cell invasion, whereas addback of this molecule reduced activated PSC-mediated cancer cell invasion. We also observed that circulating concentrations of RvD5n-3 DPA levels were decreased in peripheral blood of metastatic PDAC patients when compared with those measured in plasma of non-metastatic PDAC patients. Together, these findings indicate that RvD5n-3 DPA may regulate cancer-stroma cross-talk and invasion.


Assuntos
Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Humanos , Araquidonato 15-Lipoxigenase/metabolismo , Células Estreladas do Pâncreas/metabolismo , Células Estreladas do Pâncreas/patologia , Cromatografia Líquida , Espectrometria de Massas em Tandem , Neoplasias Pancreáticas/patologia , Carcinoma Ductal Pancreático/patologia , Tretinoína/metabolismo , Invasividade Neoplásica/patologia , Neoplasias Pancreáticas
4.
Int J Mol Sci ; 24(19)2023 Sep 27.
Artigo em Inglês | MEDLINE | ID: mdl-37834100

RESUMO

Pancreatic ductal adenocarcinoma (PDAC) cells display extensive crosstalk with their surrounding environment to regulate tumor growth, immune evasion, and metastasis. Recent advances have attributed many of these interactions to intercellular communication mediated by small extracellular vesicles (sEVs), involving cancer-associated fibroblasts (CAF). To explore the impact of sEVs on monocyte lineage transition as well as the expression of checkpoint receptors and activation markers, peripheral blood monocytes from healthy subjects were exposed to PDAC-derived sEVs. Additionally, to analyze the role of sEV-associated HA in immune regulation and tissue-resident fibroblasts, monocytes and pancreatic stellate cells were cultured in the presence of PDAC sEVs with or depleted of HA. Exposure of monocytes to sEVs resulted in unique phenotypic changes in HLA-DR, PD-L1, CD86 and CD64 expression, and cytokine secretion that was HA-independent except for IL-1ß and MIP1ß. In contrast, monocyte suppression of autologous T cell proliferation was reduced following exposure to HA-low sEVs. In addition, exposure of stellate cells to sEVs upregulated the secretion of various cytokines, including MMP-9, while removal of HA from PDAC-derived sEVs attenuated the secretion of MMP-9, demonstrating the role of sEV-associated HA in regulating expression of this pro-tumorigenic cytokine from stellate cells. This observation lends credence to the findings from the TCGA database that PDAC patients with high levels of enzymes in the HA synthesis pathway had worse survival rates compared with patients having low expression of these enzymes. PDAC-derived sEVs have an immune modulatory role affecting the activation state of monocyte subtypes. However, sEV-associated HA does not affect monocyte phenotype but alters cytokine secretion and suppression of autologous T cell proliferation and induces secretion of pro-tumorigenic factors by pancreatic stellate cells (PSC), as has been seen following the conversion of PSCs to cancer-associated fibroblasts (CAFs). Interruption of the hexosamine biosynthetic pathway, activated in PDAC producing the key substrate (UDP-GlcNAc) for HA synthesis, thus, represents a potential clinical interception strategy for PDAC patients. Findings warrant further investigations of underlying mechanisms involving larger sample cohorts.


Assuntos
Carcinoma Ductal Pancreático , Vesículas Extracelulares , Neoplasias Pancreáticas , Humanos , Metaloproteinase 9 da Matriz/metabolismo , Neoplasias Pancreáticas/patologia , Carcinoma Ductal Pancreático/patologia , Citocinas/metabolismo , Vesículas Extracelulares/metabolismo , Microambiente Tumoral , Células Estreladas do Pâncreas , Neoplasias Pancreáticas
5.
Acta Biomater ; 169: 451-463, 2023 10 01.
Artigo em Inglês | MEDLINE | ID: mdl-37572982

RESUMO

Pancreatic ductal adenocarcinoma (PDAC) is characterized by a dense extracellular matrix flooded with immune suppressive cells, resulting in extremely poor clinical response to immunotherapy. It has been revealed that the activation of pancreatic stellate cells (PSCs) makes considerable contributions to the immunological "cold" tumor microenvironment (TME). Herein, we developed a polyamino acid-based nanodrug incorporating the PSC activation inhibitor calcipotriol and anti-CXCL12 siRNA. The nanodrug was easily prepared with a small particle size and is capable of penetrating pancreatic tumors to inactivate PSCs and downregulate CXCL12. The in vivo results of orthotopic pancreatic tumor treatment demonstrated that codelivery of calcipotriol and anti-CXCL12 siRNA remodeled the PDAC TME with reduced extracellular matrix and decreased immunosuppressive T cells. Eventually, the infiltration of cytotoxic T cells was increased, thereby acting with immune checkpoint blockade (ICB) therapy for immunologically "cold" pancreatic tumors. In the present study, we propose a promising paradigm to improve the immunotherapy outcome of PDAC using nanodrugs that synchronously inhibit PSC activation and regulatory T-cell infiltration. STATEMENT OF SIGNIFICANCE: Pancreatic ductal adenocarcinoma (PDAC) is characterized by a dense extracellular matrix (ECM) that impedes the tumor infiltration of therapeutic agents and cytotoxic T lymphocytes, resulting in a poor clinical response to immunotherapy. In the present study, we proposed a promising approach for enhanced immunotherapy of pancreatic cancer. Specifically, a nanodrug incorporating calcipotriol and anti-CXCL12 siRNA was synthesized to synchronously inactivate matrix-producing pancreatic stellate cells and suppress the infiltration of regulatory T cells. The reduced ECM removed the pathological barrier, preventing nanodrug penetration and effector T-cell infiltration, leading to a conversion of the immunosuppressive "cold" microenvironment to a "hot" microenvironment, which eventually boosted the immunotherapy of anti-PD-1 antibodies in pancreatic cancer.


Assuntos
Carcinoma Ductal Pancreático , Nanopartículas , Neoplasias Pancreáticas , Humanos , Linfócitos T Reguladores/patologia , Células Estreladas do Pâncreas/patologia , Neoplasias Pancreáticas/tratamento farmacológico , Carcinoma Ductal Pancreático/tratamento farmacológico , Imunoterapia , Nanopartículas/uso terapêutico , Microambiente Tumoral , Neoplasias Pancreáticas
6.
Pflugers Arch ; 475(10): 1225-1240, 2023 10.
Artigo em Inglês | MEDLINE | ID: mdl-37566113

RESUMO

Pancreatic stellate cells (PSCs) that can co-metastasize with cancer cells shape the tumor microenvironment (TME) in pancreatic ductal adenocarcinoma (PDAC) by producing an excessive amount of extracellular matrix. This leads to a TME characterized by increased tissue pressure, hypoxia, and acidity. Moreover, cells within the tumor secrete growth factors. The stimuli of the TME trigger Ca2+ signaling and cellular Na+ loading. The Na+/Ca2+ exchanger (NCX) connects the cellular Ca2+ and Na+ homeostasis. The NCX is an electrogenic transporter, which shuffles 1 Ca2+ against 3 Na+ ions over the plasma membrane in a forward or reverse mode. Here, we studied how the impact of NCX activity on PSC migration is modulated by cues from the TME. NCX expression was revealed with qPCR and Western blot. [Ca2+]i, [Na+]i, and the cell membrane potential were determined with the fluorescent indicators Fura-2, Asante NaTRIUM Green-2, and DiBAC4(3), respectively. PSC migration was quantified with live-cell imaging. To mimic the TME, PSCs were exposed to hypoxia, pressure, acidic pH (pH 6.6), and PDGF. NCX-dependent signaling was determined with Western blot analyses. PSCs express NCX1.3 and NCX1.9. [Ca2+]i, [Na+]i, and the cell membrane potential are 94.4 nmol/l, 7.4 mmol/l, and - 39.8 mV, respectively. Thus, NCX1 usually operates in the forward (Ca2+ export) mode. NCX1 plays a differential role in translating cues from the TME into an altered migratory behavior. When NCX1 is operating in the forward mode, its inhibition accelerates PSC migration. Thus, NCX1-mediated extrusion of Ca2+ contributes to a slow mode of migration of PSCs.


Assuntos
Células Estreladas do Pâncreas , Trocador de Sódio e Cálcio , Humanos , Trocador de Sódio e Cálcio/metabolismo , Células Estreladas do Pâncreas/metabolismo , Proteínas de Membrana Transportadoras/metabolismo , Transdução de Sinais , Hipóxia , Cálcio/metabolismo
7.
Dig Dis Sci ; 68(9): 3644-3659, 2023 09.
Artigo em Inglês | MEDLINE | ID: mdl-37526905

RESUMO

BACKGROUND: Nintedanib (Ninte) has been approved for the treatment of pulmonary fibrosis, and whether it can ameliorate chronic pancreatitis (CP) is unknown. AIMS: This study was conducted to investigate the effect and molecular mechanism of Ninte on pancreatic fibrosis and inflammation in vivo and in vitro. METHODS: The caerulein-induced CP model of murine was applied, and Ninte was orally administered. Pathological changes in pancreas were evaluated using hematoxylin & eosin, Sirius Red, Masson's trichrome, and anti-Ki-67 staining. For in vitro studies, the effects of Ninte on cell viability, apoptosis, and migration of pancreatic stellate cells (PSCs) were determined by CCK-8, flow cytometry, and wound healing assays, respectively. The potential molecular mechanisms of the effects of Ninte on PSCs were analyzed by RNA-Seq and verified at the gene expression and protein activity levels by qRT-PCR and Western Blot. RESULTS: Ninte significantly alleviated the weight loss in mice with caerulein-induced CP and simultaneously attenuated the pancreatic damage, as evidenced by reduced acinar atrophy, collagen deposition, infiltration of inflammatory cells, and inhibited cell proliferation/regeneration. Besides, Ninte markedly suppressed the transcription of fibrogenic and proinflammatory genes in pancreatic tissues. Further in vitro studies showed that Ninte significantly inhibited the transcription and protein expression of genes corresponding to fibrogenesis and proliferation in PSCs. The results of RNA-Seq analysis and subsequent verification assays indicated that Ninte inhibited the activation and proliferation of PSCs via the JAK/STAT3 and ERK1/2 pathways. CONCLUSIONS: These findings indicate that Ninte may be a potential anti-inflammatory and anti-fibrotic therapeutic agent for CP.


Assuntos
Sistema de Sinalização das MAP Quinases , Pancreatite Crônica , Camundongos , Animais , Células Estreladas do Pâncreas/patologia , Ceruletídeo/toxicidade , Pancreatite Crônica/induzido quimicamente , Pancreatite Crônica/tratamento farmacológico , Pancreatite Crônica/metabolismo , Pâncreas/patologia , Fibrose
8.
Gastroenterology ; 165(6): 1488-1504.e20, 2023 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-37634735

RESUMO

BACKGROUND & AIMS: Studies have demonstrated that activated pancreatic stellate cells (PSCs) play a crucial role in pancreatic fibrogenesis in chronic pancreatitis (CP); however, the precise mechanism for PSCs activation has not been fully elucidated. We analyzed the role of injured pancreatic acinar cells (iPACs) in the activation of PSCs of CP. METHODS: Sphingosine kinase 1 (SPHK1)/sphingosine-1-phosphate (S1P) signaling was evaluated in experimental CP induced by cerulein injection or pancreatic duct ligation, as well as in PACs injured by cholecystokinin. The activation of PSCs and pancreatic fibrosis in CP samples was evaluated by immunohistochemical and immunofluorescence analyses. In vitro coculture assay of iPACs and PSCs was created to evaluate the effect of the SPHK1/S1P pathway and S1P receptor 2 (SIPR2) on autophagy and activation of PSCs. The pathogenesis of CP was assessed in SPHK1-/- mice or PACs-specific SPHK1-knockdown mice with recombinant adeno-associated virus serotypes 9-SPHK1-knockdown, as well as in mice treated with inhibitor of SPHK1 and S1P receptor 2 (S1PR2). RESULTS: SPHK1/S1P was remarkably increased in iPACs and acinar cells in pancreatic tissues of CP mice. Meanwhile, the pathogenesis, fibrosis, and PSCs activation of CP was significantly prevented in SPHK1-/- mice and recombinant adeno-associated virus serotypes 9-SPHK1-knockdown mice. Meanwhile, iPACs obviously activated PSCs, which was prevented by SPHK1 knockdown in iPACs. Moreover, iPACs-derived S1P specifically combined to S1PR2 of PSCs, by which modulated 5' adenosine monophosphate-activated protein kinase/mechanistic target of rapamycin pathway and consequently induced autophagy and activation of PSCs. Furthermore, hypoxia-inducible factor 1-α and -2α promoted SPHK1 transcription of PACs under hypoxia conditions, which is a distinct characteristic of the CP microenvironment. Coincidently, inhibition of SPHK1 and S1PR2 activity with inhibitor PF-543 and JTE-013 obviously impeded pancreatic fibrogenesis of CP mice. CONCLUSIONS: The activated SPHK1/S1P pathway in iPACs induces autophagy and activation of PSCs by regulating the S1PR2/5' adenosine monophosphate-activated protein kinase/mammalian target of rapamycin pathway, which promotes fibrogenesis of CP. The hypoxia microenvironment might contribute to the cross talk between PACs and PSCs in pathogenesis of CP.


Assuntos
Células Acinares , Pancreatite Crônica , Animais , Camundongos , Receptores de Esfingosina-1-Fosfato , Células Estreladas do Pâncreas , Pancreatite Crônica/induzido quimicamente , Autofagia , Proteínas Quinases Ativadas por AMP , Fibrose , Monofosfato de Adenosina , Hipóxia , Mamíferos
9.
JCI Insight ; 8(19)2023 Oct 09.
Artigo em Inglês | MEDLINE | ID: mdl-37643024

RESUMO

Pancreatic ductal adenocarcinoma (PDAC) progresses in an organ with a unique pH landscape, where the stroma acidifies after each meal. We hypothesized that disrupting this pH landscape during PDAC progression triggers pancreatic stellate cells (PSCs) and cancer-associated fibroblasts (CAFs) to induce PDAC fibrosis. We revealed that alkaline environmental pH was sufficient to induce PSC differentiation to a myofibroblastic phenotype. We then mechanistically dissected this finding, focusing on the involvement of the Na+/H+ exchanger NHE1. Perturbing cellular pH homeostasis by inhibiting NHE1 with cariporide partially altered the myofibroblastic PSC phenotype. To show the relevance of this finding in vivo, we targeted NHE1 in murine PDAC (KPfC). Indeed, tumor fibrosis decreased when mice received the NHE1-inhibitor cariporide in addition to gemcitabine treatment. Moreover, the tumor immune infiltrate shifted from granulocyte rich to more lymphocytic. Taken together, our study provides mechanistic evidence on how the pancreatic pH landscape shapes pancreatic cancer through tuning PSC differentiation.


Assuntos
Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Camundongos , Animais , Células Estreladas do Pâncreas/patologia , Linhagem Celular Tumoral , Neoplasias Pancreáticas/patologia , Carcinoma Ductal Pancreático/patologia , Fenótipo , Homeostase , Fibrose , Neoplasias Pancreáticas
10.
Eur J Pharmacol ; 957: 176000, 2023 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-37604222

RESUMO

OBJECTIVE: To evaluate the inhibitory effect of vitamin D3 analogue calcipotriol (Cal) on the fibrosis of pancreatic stellate cells (PSCs) induced by TGF-ß1 and the rationality of Cal use in alcoholic chronic pancreatitis (ACP). MATERIAL AND METHODS: Double-labeling immunofluorescence was used for the identification of VDR+PSCs in the pancreas of healthy controls (HC) and ACP patients. Van Gieson staining for examination of collagen fibers. RT-qPCR and Western Blot for determining the mRNAs and proteins of VDR, TGF-ß1 and COL1A1 in the pancreas of ACP or in vitro PSCs. ELISA or LC-MS/MS for detection of serum TGF-ß1 and COL1A1 or 25(OH)D3. The PSC line (RP-2 cell) was used for the determination of proteomic alterations in Cal plus TGF-ß1 versus TGF-ß1 and to examine the effect of VDR gene knockdown. RESULTS: Enhanced expression of VDR was detected in RP-2 cells stimulated with alcohol (ALC) plus Cal versus Cal alone and in PSCs in the pancreas of ACP versus HC. The increased VDR+PSCs were positively correlated with the levels of COL1A1 mRNAs or areas of collagen deposition in the pancreas of ACP. TGF-ß1 was overexpressed in the pancreas of ACP and ALC-treated RP-2 cells while 25(OH)D3 level in serum was significantly decreased in ACP versus HC. Through a VDR-dependent mechanism, Cal antagonized 16 profibrotic proteins in TGF-ß1-induced RP-2 cells that included 7 extracellular matrix components, 2 cytoskeletal proteins, 2 fibrosis-associated factors (RUNX1 and TRAF2), TIMP-1, CCN1, integrin α11, an adhesion scaffold protein (TGFB1i1) and an enzyme mediating TGF-ß1-induced fibrogenesis (ENPP1). CONCLUSION: This study suggests that Cal administration may be a potential antifibrotic strategy via inhibiting TGF-ß1-mediated PSC action during the development of ACP.


Assuntos
Colecalciferol , Fator de Crescimento Transformador beta1 , Humanos , Colecalciferol/farmacologia , Cromatografia Líquida , Células Estreladas do Pâncreas , Proteômica , Espectrometria de Massas em Tandem , Fatores de Crescimento Transformadores
11.
BMJ Open Gastroenterol ; 10(1)2023 07.
Artigo em Inglês | MEDLINE | ID: mdl-37433685

RESUMO

OBJECTIVE: Stellate cells are responsible for liver and pancreas fibrosis and strictly correlate with tumourigenesis. Although their activation is reversible, an exacerbated signalling triggers chronic fibrosis. Toll-like receptors (TLRs) modulate stellate cells transition. TLR5 transduces the signal deriving by the binding to bacterial flagellin from invading mobile bacteria. DESIGN: Human hepatic and pancreatic stellate cells were activated by the administration of transforming growth factor-beta (TGF-ß). TLR5 was transiently knocked down by short-interference RNA transfection. Reverse Transcription-quantitativePCR and western blot were performed to analyse the transcript and protein level of TLR5 and the transition players. Fluorescence microscopy was performed to identify these targets in spheroids and in the sections of murine fibrotic liver. RESULTS: TGF-ß-activated human hepatic and pancreatic stellate cells showed an increase of TLR5 expression. TLR5 knockdown blocked the activation of those stellate cells. Furthermore, TLR5 busted during murine liver fibrosis and co-localised with the inducible Collagen I. Flagellin suppressed TLR5, COL1A1 and ACTA2 expression after the administration of TGF-ß. Instead, the antagonist of TLR5 did not block the effect of TGF-ß. Wortmannin, a specific AKT inhibitor, induced TLR5 but not COL1A1 and ACTA2 transcript and protein level. CONCLUSION: TGF-ß-mediated activation of hepatic and pancreatic stellate cells requires the over-expression of TLR5. Instead, its autonomous signalling inhibits the activation of the stellate cells, thus prompting a signalling through different regulatory pathways.


Assuntos
Flagelina , Células Estreladas do Pâncreas , Receptor 5 Toll-Like , Animais , Humanos , Camundongos , Flagelina/farmacologia , Cirrose Hepática , Receptor 5 Toll-Like/metabolismo
12.
J Cell Mol Med ; 27(17): 2533-2546, 2023 09.
Artigo em Inglês | MEDLINE | ID: mdl-37488774

RESUMO

The pancreatic stellate cells (PSCs) play an important role in the development of pancreatic cancer (PC) through mechanisms that remain unclear. Exosomes secreted from PSCs act as mediators for communication in PC. This study aimed to explore the role of PSC-derived exosomal small RNAs derived from tRNAs (tDRs) in PC cells. Exosomes from PSCs were extracted and used to detect their effects on PC cell proliferation, migration and invasion. Exosomal tDRs profiling was performed to identify PSC-derived exosomal tDRs. ISH and qRT-PCR were used to examine the tRF-19-PNR8YPJZ levels and clinical value in clinical samples. The biological function of exosomal tRF-19-PNR8YPJZ was determined using the CCK-8, clone formation, wound healing and transwell assays, subcutaneous tumour formation and lung metastatic models. The relationship between the selected exosomal tRF-19-PNR8YPJZ and AXIN2 was determined by RNA sequencing, luciferase reporter assay. PSC-derived exosomes promoted the proliferation, migration, and invasion of PC cells. Novel and abundant tDRs are found to be differentially expressed in PANC-1 cells after treatment with PSC-derived exosomes, such as tRF-19-PNR8YPJZ. PC tissue samples showed markedly higher levels of tRF-19-PNR8YPJZ than normal controls. Patients with PC exhibiting high tRF-19-PNR8YPJZ expression had a highly lymph node invasion, metastasis, perineural invasion, advanced clinical stage and poor overall survival. Exosomal tRF-19-PNR8YPJZ from PSCs targeted AXIN2 in PC cells and decreased its expression, thus activating the Wnt pathway and promoting proliferation and metastasis. Exosomal tRF-19-PNR8YPJZ from PSCs promoted proliferation and metastasis in PC cells via AXIN2.


Assuntos
Exossomos , MicroRNAs , Neoplasias Pancreáticas , Humanos , Células Estreladas do Pâncreas/metabolismo , Linhagem Celular Tumoral , Movimento Celular/genética , Neoplasias Pancreáticas/patologia , Exossomos/metabolismo , Proliferação de Células/genética , MicroRNAs/genética , MicroRNAs/metabolismo , Proteína Axina/genética , Proteína Axina/metabolismo , Neoplasias Pancreáticas
13.
Molecules ; 28(14)2023 Jul 22.
Artigo em Inglês | MEDLINE | ID: mdl-37513458

RESUMO

Chronic pancreatitis (CP) is a disease characterized by inflammatory recurrence that accompanies the development of pancreatic fibrosis. As the mystery of CP pathogenesis is gradually revealed, accumulating evidence suggests that the activation of pancreatic stellate cells (PSCs) and the appearance of a myofibroblast-like phenotype are the key gatekeepers in the development of CP. Targeting PSCs to prevent their activation and conversion to a myofibroblast-like phenotype, as well as increasing antioxidant capacity to counteract ongoing oxidative stress, are effective strategies for preventing or treating CP. Therefore, we reviewed the crosstalk between CP and pancreatic fibrosis, summarized the activation mechanisms of PSCs, and investigated potential CP therapeutic strategies targeting PSCs, including, but not limited to, anti-fibrosis therapy, antioxidant therapy, and gene therapy. Meanwhile, the above therapeutic strategies are selected in order to update the available phytopharmaceuticals as novel complementary or alternative approaches for the prevention and treatment of CP to clarify their potential mechanisms of action and their relevant molecular targets, aiming to provide the most comprehensive therapeutic treatment direction for CP and to bring new hope to CP patients.


Assuntos
Pâncreas , Pancreatite Crônica , Humanos , Pâncreas/patologia , Células Estreladas do Pâncreas/patologia , Antioxidantes/uso terapêutico , Pancreatite Crônica/tratamento farmacológico , Pancreatite Crônica/patologia
14.
Sci Rep ; 13(1): 12201, 2023 07 27.
Artigo em Inglês | MEDLINE | ID: mdl-37500741

RESUMO

Chronic pancreatitis (CP) is a disease characterized by the inflammation and destruction of pancreatic tissue, leading to the replacement of functional tissue with fibrotic tissue. The regenerating gene (Reg) family proteins have recently been implicated in the repair and regeneration of inflamed pancreatic tissue, though the exact mechanisms of their involvement in the pathogenesis of CP are not yet fully understood. To investigate the role of Reg family proteins in CP, we generated global knockout mice (Reg-/-) for Reg1-3 (Reg1,2,3a,3b,3d,3g) genes using the CRISPR/Cas9 system. We then investigated the effect of Reg family protein deficiency in a genetic model of CP (X-SPINK1) mice by knocking out Reg1-3 genes. We examined pancreatic morphology, inflammatory cytokines expression, and activation of pancreatic stellate cells (PSCs) at different ages. Reg-/- mice showed no abnormalities in general growth and pancreas development. Deficiency of Reg1-3 in CP mice led to a reduction in pancreatic parenchymal loss, decreased deposition of collagen, and reduced expression of proinflammatory cytokines. Additionally, Reg proteins were found to stimulate PSCs activation. Overall, our study suggests that Reg1-3 deficiency can lead to the remission of CP and Reg family proteins could be a potential therapeutic target for the treatment of CP.


Assuntos
Células Estreladas do Pâncreas , Pancreatite Crônica , Camundongos , Animais , Células Estreladas do Pâncreas/metabolismo , Pancreatite Crônica/metabolismo , Pâncreas/metabolismo , Inflamação/patologia , Camundongos Knockout , Colágeno/metabolismo , Citocinas/metabolismo , Regeneração , Fibrose , Litostatina/genética , Litostatina/metabolismo
15.
Indian J Gastroenterol ; 42(4): 558-561, 2023 08.
Artigo em Inglês | MEDLINE | ID: mdl-37418051

RESUMO

Pancreatic fibrosis is characterized by the activation of pancreatic stellate cells leading to the expression of smooth muscle actin (α-SMA). Normal pancreatic tissue has predominantly quiescent stellate cells in periductal and perivascular locations, which do not express α-SMA. We aimed at studying the immunohistochemistry (IHC) expression pattern of α-SMA, platelet-derived growth factor (PDGF-BB) and transforming growth factor (TGF-ß) in the resected specimen of chronic pancreatitis. Twenty biopsies from resected specimens of patients with chronic pancreatitis were included. The expression was measured in comparison to positive control biopsies (breast carcinoma for PDGF-BB and TGF-ß and appendicular tissue for α-SMA) and scored based on a semi-quantitative system based on staining intensity. The percentage of positive cells was used for objective scoring, which ranged from 0 to 15. The scoring was done separately for acini, ducts, stroma and islet cell. All patients had undergone surgery for refractory pain and the median duration of symptoms was 48 months. On IHC, α-SMA was not expressed in the acini, ducts or islets, but had high expression in the stromal regions (vs. acini, ducts and islet, p < 0.05), TGF-ß1 was also expressed maximally in islet cells; however, the distribution among all locations was statistically similar. α-SMA expression in the pancreatic stroma is an indicator of the concentration of activated stellate cells in the stroma, a site for genesis of fibrosis under the influence of growth factors in the local milieu.


Assuntos
Células Estreladas do Pâncreas , Pancreatite Crônica , Humanos , Becaplermina/metabolismo , Células Estreladas do Pâncreas/metabolismo , Células Estreladas do Pâncreas/patologia , Pancreatite Crônica/cirurgia , Pancreatite Crônica/patologia , Fator de Crescimento Transformador beta1/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Fibrose
16.
Acta Biochim Biophys Sin (Shanghai) ; 55(9): 1393-1403, 2023 Jun 19.
Artigo em Inglês | MEDLINE | ID: mdl-37337632

RESUMO

Since the prognosis of patients with pancreatic cancer is very poor and there is a lack of treatment methods, this study is performed to investigate the function of PITX2 in pancreatic stellate cells (PSCs) in the progression of pancreatic cancer. Scientific hypotheses are proposed according to bioinformatics analysis and tissue microarray analysis. Stable knockdown of PITX2 in PSCs is achieved through lentiviral infection. The relative expressions of PITX2, α-SMA, vimentin, CTNNB1, AXIN1 and LEF1 are measured in wild-type PSCs and PITX2-knockdown PSCs. Proliferative capacity is measured by EdU assay. After coculture with PSCs, the proliferation, invasion and migration capacity of pancreatic cancer cells are tested. EMT and Wnt/ß-catenin downstream genes of pancreatic cancer cells are investigated to reveal the potential mechanism. Bioinformatics analysis reveals that the PITX2 gene is highly expressed in stromal cells in pancreatic cancer and is correlated with squamous-type PDAC. Analysis of PDAC tissue microarray further demonstrates that high PITX2 level in stromal cells is correlated with poor prognosis in PDAC. After stable knockdown of PITX2 in PSCs, the relative protein levels of α-SMA, vimentin, CTNNB1, AXIN1 and LEF1 are decreased, and the proliferative capacity of PSCs is also decreased. After coculture with PSCs, in which PITX2 expression is downregulated, the proliferation, invasion and migration capacities of pancreatic cancer cells are inhibited. Thus, our results show that PITX2-silenced PSCs inhibit the growth, migration and invasion of pancreatic cancer cells via reduced EMT and Wnt/ß-catenin signaling.


Assuntos
Neoplasias Pancreáticas , beta Catenina , Humanos , beta Catenina/genética , beta Catenina/metabolismo , Vimentina/genética , Vimentina/metabolismo , Células Estreladas do Pâncreas/metabolismo , Movimento Celular/genética , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Via de Sinalização Wnt/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Neoplasias Pancreáticas
17.
Chem Biol Interact ; 380: 110541, 2023 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-37169277

RESUMO

Sphingosine-1-phosphate (S1P) is a bioactive lipid molecule that governs various functions by embedding its receptor, S1PR, in different cells. Chronic pancreatitis (CP) is characterized by pancreatic fibrosis via activation of pancreatic stellate cells (PSCs). However, the effect of S1P on CP and PSC activation is still unknown. Here, we conducted a series of experiments to explore the effect of S1P on a CP rat model and primary cultured PSCs. In vivo, CP was induced by intravenous injection of dibutyltin dichloride. S1P was administered at a dosage of 200 µg/kg body weight per day by intraperitoneal injection. After 4 weeks, serum, plasma and pancreas samples were collected for molecular analysis and histological detection. In vitro, PSCs were isolated and cultured for treatment with different doses of S1P. 3MA and MCC950 were used to determine the effect of S1P on PSC activation by regulating autophagy and the NLRP3 inflammasome. JTE013 and Si-S1PR2 were applied to verify that the functions of S1P were realized by combining with S1PR2. Cells were collected for RT‒PCR, western blotting and immunofluorescence. The results showed that S1P was increased in the plasma and pancreatic tissue of CP rats. When S1P was administered to CP rats, the function and histomorphology of the pancreas were severely impaired. In addition, S1P promoted PSC activation, heightened autophagy and enhanced the NLRP3 inflammasome in vivo and in vitro. Moreover, S1PR2 mediated the effect of S1P on PSC activation by regulating autophagy and the NLRP3 inflammasome sequentially. In conclusion, S1P binding to S1PR2 promoted PSC activation and pancreatic fibrosis in CP by regulating autophagy and the NLRP3 inflammasome. These findings provide a theoretical basis for targeting S1P/S1PR2 to treat pancreatic fibrosis and further suggest that considering the role of autophagy and the NLRP3 inflammasome may help with the treatment pancreatic fibrosis.


Assuntos
Inflamassomos , Pancreatite Crônica , Ratos , Animais , Inflamassomos/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Células Estreladas do Pâncreas , Fibrose , Pancreatite Crônica/induzido quimicamente , Autofagia
18.
Mol Cell Endocrinol ; 572: 111947, 2023 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-37150285

RESUMO

Hypoxia in pancreatic islets (islet hypoxia) can occur in type 2 diabetes mellitus. Previously, our in vitro experiments demonstrated that pancreatic stellate cells (PSCs) within the islet are activated in hypoxia, promoting pancreatic ß-cell death. Here, we aimed to demonstrate the in vivo activation of intra-islet PSCs and investigate the mechanism of PSC-induced ß-cell death in hypoxia. A novel in vivo model of islet hypoxia was established by injecting fluorescent microspheres into a carotid artery of Balb/c mice (Microsphere mice). The intraperitoneal glucose tolerance (IPGTT) was performed, and pancreatic tissues were stained for insulin expression after tissue clearing. Pimonidazole staining was also performed in the pancreas to detect the presence of hypoxia in islets. Next, primary PSCs were isolated and cultured from Balb/c mice. Exosomes were isolated from culture media from PSCs cultured in hypoxia (1% oxygen). MicroRNAs (miRNAs) were prepared from exosomes from PSCs, and miRNA expression profiles were analyzed by miRNA sequencing. Several miRNAs were overexpressed in islets using miRNA mimics. Two weeks after injection of microspheres, the Microsphere mice showed worsening of glucose tolerance in IPGTT. Later, cataracts were developed in the eyes of the mice. The pancreas showed that the areas, perimeters, and diameters of insulin-positive cells decreased in Microsphere mice. Pimonidazole adducts were detected in the islets of these mice, indicating the presence of islet hypoxia. In addition, α-smooth muscle actin-positive cell numbers per islet were higher in Microsphere mice, confirming the in vivo activation of intra-islet PSCs in hypoxia. Mouse islets incubated with exosomes isolated from PSCs cultured in hypoxia showed a decrease in cell viability. The exosomes contained a variety of miRNAs, of which miR-23a-3p was found to notably increase ß-cell death through apoptosis. Together, our in vivo and in vitro data provide evidence to support that PSCs within the islets are activated in hypoxia and promote ß-cell death through exosomal miRNA transfer, which may contribute to the progression of type 2 diabetes mellitus.


Assuntos
Diabetes Mellitus Tipo 2 , Ilhotas Pancreáticas , MicroRNAs , Camundongos , Animais , MicroRNAs/genética , MicroRNAs/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Células Estreladas do Pâncreas/metabolismo , Ilhotas Pancreáticas/metabolismo , Insulina/metabolismo , Glucose/metabolismo , Hipóxia/metabolismo , Morte Celular
19.
Front Immunol ; 14: 1155085, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37205118

RESUMO

The dense tumor stroma of pancreatic ductal adenocarcinoma (PDAC) and its secreted immune active molecules provide a barrier for chemotherapy treatment as well as for immune cell infiltration to the tumor core, providing a challenge for immunotherapeutic strategies. Consequently, the investigation of processes underlying the interaction between the tumor stroma, particularly activated pancreatic stellate cells (PSCs), and immune cells may offer new therapeutic approaches for PDAC treatment. In this study, we established a 3D PDAC model cultured under flow, consisting of an endothelial tube, PSCs and PDAC organoids. This was applied to study the role of the tumor microenvironment (TME) on immune cell recruitment and its effect on partly preventing their interaction with pancreatic cancer cells. We observed that stromal cells form a physical barrier, partly shielding the cancer cells from migrating immune cells, as well as a biochemical microenvironment, that seems to attract and influence immune cell distribution. In addition, stromal targeting by Halofuginone led to an increase in immune cell infiltration. We propose that the here developed model setups will support the understanding of the cellular interplay influencing the recruitment and distribution of immune cells, and contribute to the identification of key players in the PDAC immunosuppressive TME as well as support the discovery of new strategies to treat this immune unresponsive tumor.


Assuntos
Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Humanos , Neoplasias Pancreáticas/patologia , Carcinoma Ductal Pancreático/terapia , Células Estreladas do Pâncreas/patologia , Dispositivos Lab-On-A-Chip , Microambiente Tumoral , Neoplasias Pancreáticas
20.
Semin Cancer Biol ; 93: 114-122, 2023 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-37225047

RESUMO

Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive cancer that is characterised by a prominent collagenous stromal reaction/desmoplasia surrounding tumour cells. Pancreatic stellate cells (PSCs) are responsible for the production of this stroma and have been shown to facilitate PDAC progression. Recently, extracellular vesicles (EVs), in particular, small extracellular vesicles (exosomes) have been a topic of interest in the field of cancer research for their emerging roles in cancer progression and diagnosis. EVs act as a form of intercellular communication by carrying their molecular cargo from one cell to another, regulating functions of the recipient cells. Although the knowledge of the bi-directional interactions between the PSCs and cancer cells that promote disease progression has advanced significantly over the past decade, studies on PSC-derived EVs in PDAC are currently rather limited. This review provides an overview of PDAC, pancreatic stellate cells and their interactions with cancer cells, as well as the currently known role of extracellular vesicles derived from PSCs in PDAC progression.


Assuntos
Carcinoma Ductal Pancreático , Vesículas Extracelulares , Neoplasias Pancreáticas , Humanos , Células Estreladas do Pâncreas/patologia , Neoplasias Pancreáticas/patologia , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/patologia , Vesículas Extracelulares/patologia , Neoplasias Pancreáticas
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