RESUMO
Pancreatic stellate cells (PSC) are one source of cancer-associated fibroblasts (CAF) and play, therefore, an essential role in pancreatic ductal adenocarcinoma (PDA). Paracrine signalling between PDA cells and CAF has been widely studied, yet external influences on paracrine crosstalk are poorly understood. This study aimed to gain a deeper insight into the communication of PSC and cancer cells under different co-culture conditions via analysis of PSC gene expression profiles. Two contactless co-culture models with tumor cells from the p48-Cre; lox-stop-lox-KrasG12D/+; lox-stop-lox-Trp53R172H/+ mouse model (KPC) and murine PSC separated through a microporous membrane and grown in different compartments (standard co-culture) or on different sides of the same membrane (inverse co-culture), were established. RNA-Sequencing analysis of PSC mRNA was performed 24 h and 72 h after co-culture with KPC cells. For selected genes, results were confirmed by quantitative RT-PCR and immunocytochemistry. Standard co-culture displayed 19 differentially expressed genes (DEG) at 24 h and 52 DEG at 72 h. In inverse co-culture, 800 DEG at 24 h and 2213 DEG at 72 h were enriched. PSC showed great heterogeneity in their gene expression profiles; however, mutually regulated genes of both co-cultures, such as VCAN and CHST11, could be identified. VCAN-protein-protein interaction-network analysis revealed several shared genes between co-culture models, such as SDC4 and FN1. In conclusion, PSC show a varying susceptibility to cancer cell signals depending on the co-culture method, with intensified transcriptome changes with closer proximity.
Assuntos
Carcinoma Ductal Pancreático , Técnicas de Cocultura , Neoplasias Pancreáticas , Células Estreladas do Pâncreas , Comunicação Parácrina , Transcriptoma , Células Estreladas do Pâncreas/metabolismo , Células Estreladas do Pâncreas/patologia , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/patologia , Carcinoma Ductal Pancreático/metabolismo , Animais , Camundongos , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Neoplasias Pancreáticas/metabolismo , Regulação Neoplásica da Expressão Gênica , Linhagem Celular Tumoral , Humanos , Fibroblastos Associados a Câncer/metabolismo , Fibroblastos Associados a Câncer/patologia , Perfilação da Expressão GênicaRESUMO
BACKGROUND: Pancreatic fibrosis is an early diagnostic feature of the common inherited disorder cystic fibrosis (CF). Many people with CF (pwCF) are pancreatic insufficient from birth and the replacement of acinar tissue with cystic lesions and fibrosis is a progressive phenotype that may later lead to diabetes. Little is known about the initiating events in the fibrotic process though it may be a sequela of inflammation in the pancreatic ducts resulting from loss of CFTR impairing normal fluid secretion. Here we use a sheep model of CF (CFTR-/-) to examine the evolution of pancreatic disease through gestation. METHODS: Fetal pancreas was collected at six time points from 50-days of gestation through to term, which is equivalent to ~ 13 weeks to term in human. RNA was extracted from tissue for bulk RNA-seq and single cells were prepared from 80-day, 120-day and term samples for scRNA-seq. Data were validated by immunochemistry. RESULTS: Transcriptomic evidence from bulk RNA-seq showed alterations in the CFTR-/- pancreas by 65-days of gestation, which are accompanied by marked pathological changes by 80-days of gestation. These include a fibrotic response, confirmed by immunostaining for COL1A1, αSMA and SPARC, together with acinar loss. Moreover, using scRNA-seq we identify a unique cell population that is significantly overrepresented in the CFTR-/- animals at 80- and 120-days gestation, as are stellate cells at term. CONCLUSION: The transcriptomic changes and cellular imbalance that we observe likely have pivotal roles in the evolution of CF pancreatic disease and may provide therapeutic opportunities to delay or prevent pancreatic destruction in CF.
Assuntos
Biomarcadores , Regulador de Condutância Transmembrana em Fibrose Cística , Fibrose Cística , Modelos Animais de Doenças , Células Estreladas do Pâncreas , Fibrose Cística/genética , Fibrose Cística/metabolismo , Fibrose Cística/patologia , Animais , Células Estreladas do Pâncreas/metabolismo , Células Estreladas do Pâncreas/patologia , Regulador de Condutância Transmembrana em Fibrose Cística/genética , Regulador de Condutância Transmembrana em Fibrose Cística/metabolismo , Feminino , Ovinos , Pâncreas/metabolismo , Pâncreas/patologia , Gravidez , Pancreatopatias/genética , Pancreatopatias/metabolismo , Pancreatopatias/patologia , Transcriptoma , Humanos , Perfilação da Expressão GênicaRESUMO
Chronic pancreatitis (CP) is marked by progressive fibrosis and the activation of pancreatic stellate cells (PSCs), accompanied by the destruction of pancreatic parenchyma, leading to the loss of acinar cells (ACs). Few research studies have explored the mechanism by which damaged ACs (DACs) contribute to PSCs activation and pancreatic fibrosis. Currently, there are no effective drugs for curing CP or limiting the progression of pancreatic fibrosis. In this research, co-culture with intact acinar cells (IACs) suppressed PSC activation, while co-culture with DACs did the opposite. Krüppel-like factor 4 (KLF4) was significantly upregulated in DACs and was established as the key molecule that switches ACs from PSCs-suppressor to PSCs-activator. We revealed the exosomes of IACs contributed to the anti-activated function of IACs-CS on PSCs. MiRNome profiling showed that let-7 family is significantly enriched in IAC-derived exosomes (>30% miRNome), which partially mediates IACs' suppressive impacts on PSCs. Furthermore, it has been observed that the enrichment of let-7 in exosomes was influenced by the expression level of KLF4. Mechanistic studies demonstrated that KLF4 in ACs upregulated Lin28A, thereby decreasing let-7 levels in AC-derived exosomes, and thus promoting PSCs activation. We utilized an adeno-associated virus specifically targeting KLF4 in ACs (shKLF4-pAAV) to suppress PSCs activation in CP, resulting in reduced pancreatic fibrosis. IAC-derived exosomes hold potential as potent weapons against PSCs activation via let-7s, while activated KLF4/Lin28A signaling in DACs diminished such functions. ShKLF4-pAAV holds promise as a novel therapeutic approach for CP.
Assuntos
Células Acinares , Exossomos , Fibrose , Fator 4 Semelhante a Kruppel , MicroRNAs , Células Estreladas do Pâncreas , Pancreatite Crônica , Fator 4 Semelhante a Kruppel/metabolismo , Animais , Células Estreladas do Pâncreas/metabolismo , Células Estreladas do Pâncreas/patologia , Exossomos/metabolismo , Pancreatite Crônica/metabolismo , Pancreatite Crônica/genética , Pancreatite Crônica/patologia , MicroRNAs/genética , Células Acinares/metabolismo , Células Acinares/patologia , Dependovirus/genética , Camundongos , Humanos , Fatores de Transcrição Kruppel-Like/metabolismo , Fatores de Transcrição Kruppel-Like/genética , Modelos Animais de Doenças , Vetores Genéticos/genética , Vetores Genéticos/administração & dosagem , Masculino , Técnicas de Cocultura , Pâncreas/metabolismo , Pâncreas/patologia , Terapia Genética/métodosRESUMO
Pancreatic ductal adenocarcinoma (PDAC) is an extremely aggressive form of cancer with a low survival rate. A successful treatment strategy should not be limited to targeting cancer cells alone, but should adopt a more comprehensive approach, taking into account other influential factors. These include the extracellular matrix (ECM) and immune microenvironment, both of which are integral components of the tumor microenvironment. The present review describes the roles of pancreatic stellate cells, differentiated cancerassociated fibroblasts and the interleukin family, either independently or in combination, in the progression of precursor lesions in pancreatic intraepithelial neoplasia and PDAC. These elements contribute to ECM deposition and immunosuppression in PDAC. Therapeutic strategies that integrate interleukin and/or stromal blockade for PDAC immunomodulation and fibrogenesis have yielded inconsistent results. A deeper comprehension of the intricate interplay between fibrosis, and immune responses could pave the way for more effective treatment targets, by elucidating the mechanisms and causes of ECM fibrosis during PDAC progression.
Assuntos
Carcinoma Ductal Pancreático , Fibrose , Interleucinas , Neoplasias Pancreáticas , Células Estreladas do Pâncreas , Microambiente Tumoral , Humanos , Carcinoma Ductal Pancreático/imunologia , Carcinoma Ductal Pancreático/patologia , Carcinoma Ductal Pancreático/metabolismo , Células Estreladas do Pâncreas/metabolismo , Células Estreladas do Pâncreas/patologia , Microambiente Tumoral/imunologia , Neoplasias Pancreáticas/imunologia , Neoplasias Pancreáticas/patologia , Neoplasias Pancreáticas/metabolismo , Interleucinas/metabolismo , Interleucinas/imunologia , Animais , Matriz Extracelular/metabolismo , Fibroblastos Associados a Câncer/metabolismo , Fibroblastos Associados a Câncer/imunologia , Fibroblastos Associados a Câncer/patologiaRESUMO
INTRODUCTION: Chronic pancreatitis (CP) is a persistent, recurrent, and progressive disorder that is characterized by chronic inflammation and irreversible fibrosis of the pancreas. It is associated with severe morbidity, resulting in intense abdominal pain, diabetes, exocrine and endocrine dysfunction, and an increased risk of pancreatic cancer. The etiological factors are diverse and the major risk factors include smoking, chronic alcoholism, as well as other environmental and genetic factors. The treatment and management of CP is challenging, and no definitive curative therapy is currently available. AREAS COVERED: This review paper aims to provide an overview of the different cell types in the pancreas that is known to mediate disease progression and outline potential novel therapeutic approaches and drug targets that may be effective in treating and managing CP. The information presented in this review was obtained by conducting a NCBI PubMed database search, using relevant keywords. EXPERT OPINION: In recent years, there has been an increased interest in the development of novel therapeutics for CP. A collaborative multi-disciplinary approach coupled with a consistent funding for research can expedite progress of translating the findings from bench to bedside.
Assuntos
Macrófagos , Células Estreladas do Pâncreas , Pancreatite Crônica , Pancreatite Crônica/terapia , Humanos , Células Estreladas do Pâncreas/metabolismo , Células Estreladas do Pâncreas/efeitos dos fármacos , Células Estreladas do Pâncreas/patologia , Animais , Macrófagos/metabolismo , Terapia de Alvo MolecularRESUMO
As a pluripotent cell, activated pancreatic stellate cells (PSCs) can differentiate into various pancreatic parenchymal cells and participate in the secretion of extracellular matrix and the repair of pancreatic damage. Additionally, PSCs characteristics allow them to contribute to pancreatic inflammation and carcinogenesis. Moreover, a detailed study of the pathogenesis of activated PSCs in pancreatic disease can offer promise for the development of innovative therapeutic strategies and improved patient prognoses. Therefore, the present study review aimed to examine the involvement of activated PSCs in pancreatic diseases and elucidate the underlying mechanisms to provide a viable therapeutic strategy for the management of pancreasrelated diseases.
Assuntos
Pâncreas , Pancreatopatias , Células Estreladas do Pâncreas , Humanos , Células Estreladas do Pâncreas/metabolismo , Células Estreladas do Pâncreas/patologia , Pâncreas/metabolismo , Pâncreas/patologia , Pâncreas/citologia , Pancreatopatias/patologia , Pancreatopatias/metabolismo , Animais , Matriz Extracelular/metabolismo , Diferenciação Celular , Neoplasias Pancreáticas/patologia , Neoplasias Pancreáticas/metabolismoRESUMO
Small extracellular vesicles (sEVs) are cell-derived vesicles evolving as important elements involved in all stages of cancers. sEVs bear unique protein signatures that may serve as biomarkers. Pancreatic cancer (PC) records a very poor survival rate owing to its late diagnosis and several cancer cell-derived proteins have been reported as candidate biomarkers. However, given the pivotal role played by stellate cells (PSCs, which produce the collagenous stroma in PC), it is essential to also assess PSC-sEV cargo in biomarker discovery. Thus, this study aimed to isolate and characterise sEVs from mouse PC cells and PSCs cultured alone or as co-cultures and performed proteomic profiling and pathway analysis. Proteomics confirmed the enrichment of specific markers in the sEVs compared to their cells of origin as well as the proteins that are known to express in each of the culture types. Most importantly, for the first time it was revealed that PSC-sEVs are enriched in proteins (including G6PI, PGAM1, ENO1, ENO3, and LDHA) that mediate pathways related to development of diabetes, such as glucose metabolism and gluconeogenesis revealing a potential role of PSCs in pancreatic cancer-related diabetes (PCRD). PCRD is now considered a harbinger of PC and further research will enable to identify the role of these components in PCRD and may develop as novel candidate biomarkers of PC.
Assuntos
Vesículas Extracelulares , Neoplasias Pancreáticas , Células Estreladas do Pâncreas , Proteômica , Animais , Neoplasias Pancreáticas/metabolismo , Neoplasias Pancreáticas/patologia , Células Estreladas do Pâncreas/metabolismo , Células Estreladas do Pâncreas/patologia , Camundongos , Vesículas Extracelulares/metabolismo , Proteômica/métodos , Biomarcadores Tumorais/metabolismo , Linhagem Celular Tumoral , Proteoma/análise , Proteoma/metabolismoRESUMO
PURPOSE: To evaluate the effect and mechanism of 1,25(OH)2D3 on pancreatic stellate cells (PSCs) in type 2 diabetes mellitus (T2DM). METHODS: A mouse model of T2DM was successfully established by high-fat diet (HFD) /streptozotocin (STZ) and administered 1,25(OH)2D3 for 3 weeks. Fasting blood glucose (FBG), glycated hemoglobin A1c (GHbA1c), insulin (INS) and glucose tolerance were measured. Histopathology changes and fibrosis of pancreas were examined by hematoxylin and eosin staining and Masson staining. Mouse PSCs were extracted, co-cultured with mouse insulinoma ß cells (MIN6 cells) and treated with 1,25(OH)2D3. ELISA detection of inflammatory factor expression. Tissue reactive oxygen species (ROS) levels were also measured. Immunofluorescence or Western blotting were used to measure fibrosis and inflammation-related protein expression. RESULTS: PSCs activation and islets fibrosis in T2DM mice. Elevated blood glucose was accompanied by significant increases in serum inflammatory cytokines and tissue ROS levels. 1,25(OH)2D3 attenuated islet fibrosis by reducing hyperglycemia, ROS levels, and inflammatory factors expression. Additionally, the co-culture system confirmed that 1,25(OH)2D3 inhibited PSCs activation, reduced the secretion of pro-inflammatory cytokines, down-regulated the expression of fibrosis and inflammation-related proteins, and promoted insulin secretion. CONCLUSION: Our findings identify that PSCs activation contributes to islet fibrosis and ß-cell dysfunction. 1,25(OH)2D3 exerts beneficial effects on T2DM potentially by inhibiting PSCs activation and inflammatory response, highlighting promising control strategies of T2DM by vitamin D.
Assuntos
Calcitriol , Diabetes Mellitus Experimental , Diabetes Mellitus Tipo 2 , Secreção de Insulina , Insulina , Células Estreladas do Pâncreas , Animais , Células Estreladas do Pâncreas/efeitos dos fármacos , Células Estreladas do Pâncreas/metabolismo , Células Estreladas do Pâncreas/patologia , Camundongos , Diabetes Mellitus Tipo 2/metabolismo , Calcitriol/farmacologia , Masculino , Secreção de Insulina/efeitos dos fármacos , Insulina/metabolismo , Insulina/sangue , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patologia , Espécies Reativas de Oxigênio/metabolismo , Glicemia/metabolismo , Glicemia/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Fibrose , Técnicas de CoculturaRESUMO
BACKGROUND: Pancreatic fibrosis is one of the most important pathological features of chronic pancreatitis (CP) and pancreatic stellate cells (PSCs) are the key cells of fibrosis. As an extracellular matrix (ECM) glycoprotein, cartilage oligomeric matrix protein (COMP) is critical for collagen assembly and ECM stability and recent studies showed that COMP exert promoting fibrosis effect in the skin, lungs and liver. However, the role of COMP in activation of PSCs and pancreatic fibrosis remain unclear. We aimed to investigate the role and specific mechanisms of COMP in regulating the profibrotic phenotype of PSCs and pancreatic fibrosis. METHODS: ELISA method was used to determine serum COMP in patients with CP. Mice model of CP was established by repeated intraperitoneal injection of cerulein and pancreatic fibrosis was evaluated by Hematoxylin-Eosin staining (H&E) and Sirius red staining. Immunohistochemical staining was used to detect the expression changes of COMP and fibrosis marker such as α-SMA and Fibronectin in pancreatic tissue of mice. Cell Counting Kit-8, Wound Healing and Transwell assessed the proliferation and migration of human pancreatic stellate cells (HPSCs). Western blotting, qRT-PCR and immunofluorescence staining were performed to detect the expression of fibrosis marker, AKT and MAPK family proteins in HPSCs. RNA-seq omics analysis as well as small interfering RNA of COMP, recombinant human COMP (rCOMP), MEK inhibitors and PI3K inhibitors were used to study the effect and mechanism of COMP on activation of HPSCs. RESULTS: ELISA showed that the expression of COMP significantly increased in the serum of CP patients. H&E and Sirius red staining analysis showed that there was a large amount of collagen deposition in the mice in the CP model group and high expression of COMP, α-SMA, Fibronectin and Vimentin were observed in fibrotic tissues. TGF-ß1 stimulates the activation of HPSCs and increases the expression of COMP. Knockdown of COMP inhibited proliferation and migration of HPSCs. Further, RNA-seq omics analysis and validation experiments in vitro showed that rCOMP could significantly promote the proliferation and activation of HPSCs, which may be due to promoting the phosphorylation of ERK and AKT through membrane protein receptor CD36. rCOMP simultaneously increased the expression of α-SMA, Fibronectin and Collagen I in HPSCs. CONCLUSION: In conclusion, this study showed that COMP was up-regulated in CP fibrotic tissues and COMP induced the activation, proliferation and migration of PSCs through the CD36-ERK/AKT signaling pathway. COMP may be a potential therapeutic candidate for the treatment of CP. Interfering with the expression of COMP or the communication between COMP and CD36 on PSCs may be the next direction for therapeutic research.
Assuntos
Pancreatopatias , Pancreatite Crônica , Animais , Humanos , Camundongos , Proteína de Matriz Oligomérica de Cartilagem/metabolismo , Proteína de Matriz Oligomérica de Cartilagem/farmacologia , Proteína de Matriz Oligomérica de Cartilagem/uso terapêutico , Células Cultivadas , Colágeno Tipo I/metabolismo , Fibronectinas/metabolismo , Fibrose , Pancreatopatias/metabolismo , Células Estreladas do Pâncreas/metabolismo , Células Estreladas do Pâncreas/patologia , Pancreatite Crônica/tratamento farmacológico , Pancreatite Crônica/metabolismo , Pancreatite Crônica/patologia , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de SinaisRESUMO
Pancreatic cancer is characterized by a densely fibrotic stroma. The fibrotic stroma hinders the intratumoral penetration of nanomedicine and diminishes therapeutic efficacy. Fibrosis is characterized by an abnormal organization of extracellular matrix (ECM) components, namely the abnormal deposition and/or orientation of collagen and fibronectin. Abnormal ECM organization is chiefly driven by pathological signaling in pancreatic stellate cells (PSCs), the main cell type involved in fibrogenesis. However, whether targeting signaling pathways involved in abnormal ECM organization improves the intratumoral penetration of nanomedicines is unknown. Here, we show that targeting transforming growth factor-ß (TGFß)/Rho-associated kinase (ROCK) 1/2 signaling in PSCs normalizes ECM organization and concomitantly improves macromolecular permeability of the fibrotic stroma. Using a 3-dimensional cell culture model of the fibrotic pancreatic cancer microenvironment, we found that pharmacological inhibition of TGFß or ROCK1/2 improves the permeation of various macromolecules. By using an isoform-specific pharmacological inhibitor and siRNAs, we show that targeting ROCK2, but not ROCK1, alone is sufficient to normalize ECM organization and improve macromolecular permeability. Moreover, we found that ROCK2 inhibition/knockdown attenuates Yes-associated protein (YAP) nuclear localization in fibroblasts co-cultured with pancreatic cancer cells in 3D. Finally, pharmacological inhibition or siRNA-mediated knockdown of YAP normalized ECM organization and improved macromolecular permeability. Our results together suggest that the TGFß/ROCK2/YAP signaling axis may be therapeutically targeted to normalize ECM organization and improve macromolecular permeability to augment therapeutic efficacy of nanomedicines in pancreatic cancer.
Assuntos
Neoplasias Pancreáticas , Células Estreladas do Pâncreas , Permeabilidade , Fator de Crescimento Transformador beta , Microambiente Tumoral , Quinases Associadas a rho , Quinases Associadas a rho/metabolismo , Quinases Associadas a rho/antagonistas & inibidores , Neoplasias Pancreáticas/patologia , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/metabolismo , Humanos , Células Estreladas do Pâncreas/metabolismo , Células Estreladas do Pâncreas/efeitos dos fármacos , Células Estreladas do Pâncreas/patologia , Fator de Crescimento Transformador beta/metabolismo , Fibrose , Matriz Extracelular/metabolismo , Proteínas de Sinalização YAP/metabolismo , Linhagem Celular Tumoral , Fatores de Transcrição/metabolismo , Transdução de Sinais , Proteínas Adaptadoras de Transdução de Sinal/metabolismoRESUMO
In the realm of fibroinflammatory conditions, chronic pancreatitis (CP) stands out as a particularly challenging ailment, lacking a dedicated, approved treatment. The potential of Pirfenidone (PFD), a drug originally used for treating idiopathic pulmonary fibrosis (IPF), in addressing CP's fibrotic aspects has sparked new interest. This investigation focused on the role of PFD in diminishing fibrosis and immune response in CP, using a mouse model induced by caerulein. The research extended to in vitro studies examining the influence of PFD on pancreatic stellate cells' (PSCs) behavior and the polarization of macrophages into M1 and M2 types. Advanced techniques like RNA sequencing and comprehensive data analyses were employed to decode the molecular interactions of PFD with PSCs. Supplementary experiments using techniques such as quantitative real-time PCR, western blotting, and immunofluorescence were also implemented. Results showed a notable reduction in pancreatic damage in PFD-treated mice, manifested through decreased acinar cell atrophy, lower collagen deposition, and a reduction in macrophage presence. Further investigation revealed PFD's capacity to hinder PSCs' migration, growth, and activation, alongside a reduction in the production and secretion of extracellular matrix proteins. This effect is primarily achieved by interfering with signaling pathways such as TGF-ß/Smad, Wnt/ß-catenin, and JAK/STAT. Additionally, PFD selectively hampers M1 macrophage polarization through the STAT3 pathway, without impacting M2 polarization. These outcomes highlight PFD's dual mechanism in moderating PSC activity and M1 macrophage polarization, positioning it as a promising candidate for CP therapy.
Assuntos
Células Estreladas do Pâncreas , Pancreatite Crônica , Piridonas , Humanos , Células Estreladas do Pâncreas/metabolismo , Células Estreladas do Pâncreas/patologia , Pancreatite Crônica/tratamento farmacológico , Pancreatite Crônica/induzido quimicamente , Pâncreas/patologia , Macrófagos/metabolismo , FibroseRESUMO
The effect of 2,2',4,4'-tetrabromodiphenyl ether (BDE-47), a persistent environmental pollutant commonly used as a flame retardant in various consumer products, on pancreatitis has not been clearly elucidated, although it has been reported to be toxic to the liver, nervous system, and reproductive system. Acute pancreatitis (AP) and chronic pancreatitis (CP) models were induced in this study by intraperitoneal injection of caerulein. The aim was to investigate the impact of BDE-47 on pancreatitis by exposing the animals to acute (1 week) or chronic (8 weeks) doses of BDE-47 (30 mg/kg in the low-concentration group and 100 mg/kg in the high-concentration group). Additionally, BDE-47 was utilized to stimulate mouse bone marrow-derived macrophages, pancreatic primary stellate cells, and acinar cells in order to investigate the impact of BDE-47 on pancreatitis. In vivo experiments conducted on mice revealed that chronic exposure to BDE-47, rather than acute exposure, exacerbated the histopathological damage of AP and CP, leading to elevated fibrosis in pancreatic tissue and increased infiltration of inflammatory cells in the pancreas. In vitro experiments showed that BDE-47 can promote the expression of the inflammatory cytokines Tnf-α and Il-6 in M1 macrophages, as well as promote acinar cell apoptosis through the activation of the PERK and JNK pathways via endoplasmic reticulum stress. The findings of this study imply chronic exposure to BDE-47 may exacerbate the progression of both AP and CP by inducing acinar cell apoptosis and dysregulating inflammatory responses.
Assuntos
Células Acinares , Apoptose , Éteres Difenil Halogenados , Pancreatite Crônica , Pancreatite , Animais , Éteres Difenil Halogenados/toxicidade , Apoptose/efeitos dos fármacos , Pancreatite Crônica/induzido quimicamente , Pancreatite Crônica/patologia , Células Acinares/efeitos dos fármacos , Células Acinares/patologia , Células Acinares/metabolismo , Masculino , Pancreatite/induzido quimicamente , Pancreatite/patologia , Macrófagos/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Camundongos , Ceruletídeo/toxicidade , Pâncreas/efeitos dos fármacos , Pâncreas/patologia , Inflamação/induzido quimicamente , Inflamação/patologia , Células Estreladas do Pâncreas/efeitos dos fármacos , Células Estreladas do Pâncreas/patologia , Células Estreladas do Pâncreas/metabolismo , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Retardadores de Chama/toxicidade , Células CultivadasRESUMO
Pancreatic stellate cells (PSCs) are activated following loss of cytoplasmic vitamin A (retinol)-containing lipid droplets, which is a key event in the process of fibrogenesis of chronic pancreatitis (CP) and pancreatic ductal adenocarcinoma (PDCA). PSCs are the major source of cancer-associated fibroblasts (CAFs) that produce stroma to induce PDAC cancer cell growth, invasion, and metastasis. As an active metabolite of retinol, retinoic acid (RA) can regulate target gene expression in PSCs through its nuclear receptor complex (RAR/RXR or RXR/RXR) or transcriptional intermediary factor. Additionally, RA also has extranuclear and non-transcriptional effects. In vitro studies have shown that RA induces PSC deactivation which reduces extracellular matrix production through multiple modes of action, such as inhibiting TßRâ ¡, PDGFRß, ß-catenin and Wnt production, downregulating ERK1/2 and JNK phosphorylation and suppressing active TGF-ß1 release. RA alone or in combination with other reagents have been demonstrated to have an effective anti-fibrotic effect on cerulein-induced mouse CP models in vivo studies. Clinical trial data have shown that repurposing all-trans retinoic acid (ATRA) as a stromal-targeting agent for human pancreatic cancer is safe and tolerable, suggesting the possibility of using RA for the treatment of CP and PDCA in humans. This review focuses on RA signaling pathways in PSCs and the effects and mechanisms of RA in PSC-mediated fibrogenesis as well as the anti-fibrotic and anti-tumor effects of RA targeting PSCs or CAFs in vitro and in vivo, highlighting the potential therapies of RA against CP and PDAC.
Assuntos
Carcinoma Ductal Pancreático , Neoplasias Pancreáticas , Pancreatite Crônica , Camundongos , Humanos , Animais , Tretinoína/uso terapêutico , Células Estreladas do Pâncreas/metabolismo , Células Estreladas do Pâncreas/patologia , Vitamina A/metabolismo , Transdução de Sinais , Neoplasias Pancreáticas/patologia , Pancreatite Crônica/tratamento farmacológico , Pancreatite Crônica/metabolismo , Pancreatite Crônica/patologia , Carcinoma Ductal Pancreático/tratamento farmacológicoRESUMO
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 , RegeneraçãoRESUMO
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/patologiaRESUMO
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 PancreáticasRESUMO
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 , FibroseRESUMO
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 PancreáticasRESUMO
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/patologiaRESUMO
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.