ABSTRACT
Background and study aims The prognosis for pancreatic cancer remains poor. Molecular diagnostics and customized therapies are becoming increasingly important in clinical routine. Patient-derived, predictive model systems such as organoids have the potential to substantially increase the depth of information from biopsy material by functional and molecular characterization. We compared the extent to which the use of fine-needle aspiration needles (FNA, 22G) or fine-needle biopsy needles (FNB, 22G) influences the generation of pancreatic cancer patient-derived organoids (PDOs) to establish endoscopic standards of organoid technology. Patients and methods Endoscopic ultrasound (EUS)-guided punctures by EUS-FNA and EUS-FNB of pancreatic masses highly suspicious for adenocarcinoma (detected by computed tomography and/or magnetic resonance imaging) were prospectively evaluated. Consecutive patients received EUS-FNA and EUS-FNB in a randomized order without the need to exchange the needle shaft (only the inner needle type (FNA/-B) was exchanged) between the passes. With each needle type, the specimens for histological analysis and for PDOs were obtained separately. Results Fifty patients were enrolled in the study. Histology revealed malignancy in 42 of 50 cases (84%). In total PDOs were generated from 17 patients (34%). Of these, nine were established by FNB only, two by FNA only, and six by both FNA and FNB. Histology revealed malignancy in 13 of 17 PDO cases (76%). In two histologically false-negative cases, PDOs could be established. Conclusions EUS-FNB was superior to EUS-FNA in terms of successful generation of PDOs, although it failed to show statistical significance.
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INTRODUCTION: Hepatic fibrosis is a progressive pathological process involving the exhaustion of hepatocellular regenerative capacity and ultimately leading to the development of cirrhosis and even hepatocellular carcinoma. Brg1, the core subunit of the SWI/SNF chromatin-remodeling complex, was recently identified as important for liver regeneration. This study investigates the role of Brg1 in hepatic fibrosis development. METHODS: Hepatocyte-specific Brg1 knockout mice were generated and injected with carbon tetrachloride (CCl4) for 4, 6, 8, and 12 weeks to induce liver fibrosis. Afterwards, liver fibrosis and liver damage were assessed. RESULTS: Brg1 expression was significantly increased in the fibrotic liver tissue of wild-type mice, as compared to that of untreated wild-type mice. The livers of the Brg1 knockout animals showed reduced liver inflammation, extracellular matrix accumulation, and liver fibrosis. TNF-α and NF-κB-mediated inflammatory response was reduced in Brg1 knockout animals. CONCLUSION: Brg1 promotes the progression of liver fibrosis in mice and may therefore be used as a potential therapeutic target for treating patients with liver fibrosis due to chronic injury.
Subject(s)
Carcinoma, Hepatocellular , Hepatitis , Liver Neoplasms , Animals , Mice , Carbon Tetrachloride/toxicity , Carcinoma, Hepatocellular/pathology , Extracellular Matrix/metabolism , Fibrosis , Hepatitis/pathology , Inflammation/pathology , Liver/metabolism , Liver Cirrhosis/chemically induced , Liver Cirrhosis/genetics , Liver Cirrhosis/metabolism , Liver Neoplasms/pathology , Mice, KnockoutABSTRACT
Stringent regulation of the inflammatory response is crucial for normal tissue regeneration. Here, we analyzed the role of Toll-like receptor 3 (TLR3) in pancreatic regeneration after acute pancreatitis (AP). AP was induced by caerulein treatment in mice with global TLR3 deficiency (TLR3OFF ) or in mice re-expressing TLR3 exclusively in the myeloid cell lineage (TLR3Mye ). Compared to WT mice, TLR3OFF mice had a markedly increased formation of acinar-to-ductal metaplasia (ADM) that persisted until day 7 after initiation of AP. Pancreatic tissue of WT mice was completely regenerated after 5 days with no detectable ADM structures. The enhancing effect of TLR3-deficiency on ADM formation was closely linked with an increased and prolonged accumulation of macrophages in pancreata of TLR3OFF mice. Importantly, the phenotype of TLR3OFF mice was rescued in TLR3Mye mice, demonstrating the causative role of myeloid cell selective TLR3 signaling. Moreover, in vitro stimulation of macrophages through TLR3 initiated cell death by a caspase-8-associated mechanism. Therefore, these findings provide evidence that TLR3 signaling in myeloid cells is sufficient to limit inflammation and ADM formation and to promote regeneration after AP. Notably, resolution of inflammation after AP was associated with macrophage sensitivity to TLR3-mediated cell death.
Subject(s)
Gene Expression , Myeloid Cells/metabolism , Pancreatitis/genetics , Pancreatitis/metabolism , Toll-Like Receptor 3/genetics , Acute Disease , Animals , Biomarkers , Cell Proliferation , Cytokines/metabolism , Disease Models, Animal , Immunohistochemistry , Macrophages/immunology , Macrophages/metabolism , Mice , Mice, Knockout , Myeloid Cells/immunology , Pancreatitis/immunology , Pancreatitis/pathology , Regeneration/genetics , Signal Transduction , Toll-Like Receptor 3/metabolismABSTRACT
Ductal pancreatic carcinoma is expected to become one of the most common malignant diseases worldwide in the coming decades. However, the prognosis of the disease remains very poor and has improved only slightly over the last decade. The 5-year survival rate of all patients with ductal adenocarcinoma has been increased to approximately 10 percent. The reasons for the very poor prognosis are the advanced stage of the disease at diagnosis with metastases already present in many cases, the anatomical location of the pancreas and the tumor biology. Therapeutically, chemotherapy remains the basis of systemic therapy. Intensive combinations with FOLFIRINOX (oxaliplatin, irinotecan, leucovorin/5-FU) and nanoparticel albumin bound (nab)paclitaxel/gemcitabine lead to an improvement in overall survival in the palliative situation; used preoperatively, they can increase the rate of secondary resections. Targeted therapies and immune checkpoint inhibitors could not be established. In patients with a proven germline mutation in the BRCA gene, a therapy with the PARP inhibitor olaparib is in the approval process.This article provides an overview of differential diagnoses, meaningful diagnostics, therapeutic concepts to improve surgical treatment, possibilities of palliative chemotherapy and targeted therapy in the presence of a BRCA mutation.
Subject(s)
Carcinoma, Pancreatic Ductal , Pancreatic Neoplasms , Antineoplastic Agents/administration & dosage , Antineoplastic Agents/therapeutic use , Antineoplastic Combined Chemotherapy Protocols/administration & dosage , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Carcinoma, Pancreatic Ductal/diagnosis , Carcinoma, Pancreatic Ductal/therapy , Fluorouracil/administration & dosage , Fluorouracil/therapeutic use , Humans , Irinotecan/administration & dosage , Irinotecan/therapeutic use , Leucovorin/administration & dosage , Leucovorin/therapeutic use , Oxaliplatin/administration & dosage , Oxaliplatin/therapeutic use , Pancreatic Neoplasms/diagnosis , Pancreatic Neoplasms/therapyABSTRACT
BACKGROUND & AIMS: Angiocrine signaling by liver sinusoidal endothelial cells (LSECs) regulates hepatic functions such as growth, metabolic maturation, and regeneration. Recently, we identified GATA4 as the master regulator of LSEC specification during development. Herein, we studied the role of endothelial GATA4 in the adult liver and in hepatic pathogenesis. METHODS: We generated adult Clec4g-icretg/0xGata4fl/fl (Gata4LSEC-KO) mice with LSEC-specific depletion of Gata4. Livers were analyzed by histology, electron microscopy, immunohistochemistry/immunofluorescence, in situ hybridization, and LSECs were isolated for gene expression profiling, ChIP- and ATAC-sequencing. Partial hepatectomy was performed to assess regeneration. We used choline-deficient, l-amino acid-defined (CDAA) diet and chronic carbon tetrachloride exposure to model liver fibrosis. Human single cell RNA-seq data sets were analyzed for endothelial alterations in healthy and cirrhotic livers. RESULTS: Genetic Gata4 deficiency in LSECs of adult mice caused perisinusoidal liver fibrosis, hepatopathy and impaired liver regeneration. Sinusoidal capillarization and LSEC-to-continuous endothelial transdifferentiation were accompanied by a profibrotic angiocrine switch involving de novo endothelial expression of hepatic stellate cell-activating cytokine PDGFB. Increased chromatin accessibility and amplification by activated MYC mediated angiocrine Pdgfb expression. As observed in Gata4LSEC-KO livers, CDAA diet-induced perisinusoidal liver fibrosis was associated with GATA4 repression, MYC activation and a profibrotic angiocrine switch in LSECs. Comparison of CDAA-fed Gata4LSEC-KO and control mice demonstrated that endothelial GATA4 indeed protects against dietary-induced perisinusoidal liver fibrosis. In human cirrhotic livers, GATA4-positive LSECs and endothelial GATA4 target genes were reduced, while non-LSEC endothelial cells and MYC target genes including PDGFB were enriched. CONCLUSIONS: Endothelial GATA4 protects against perisinusoidal liver fibrosis by repressing MYC activation and profibrotic angiocrine signaling at the chromatin level. Therapies targeting the GATA4/MYC/PDGFB/PDGFRß axis offer a promising strategy for prevention and treatment of liver fibrosis. LAY SUMMARY: The liver vasculature is supposed to play a major role in the development of liver fibrosis and cirrhosis, which can lead to liver failure and liver cancer. Herein, we discovered that structural and transcriptional changes induced by genetic deletion of the transcription factor GATA4 in the hepatic endothelium were sufficient to cause liver fibrosis. Activation of the transcription factor MYC and de novo expression of the "angiocrine" growth factor PDGFB were identified as downstream drivers of fibrosis and as potential therapeutic targets for this potentially fatal disease.
Subject(s)
Endothelial Cells/metabolism , GATA4 Transcription Factor/metabolism , Liver Cirrhosis , Liver , Lymphokines , Platelet-Derived Growth Factor , Animals , Chromatin/metabolism , Drug Discovery , Gene Expression Profiling , Hepatic Stellate Cells/metabolism , Humans , Liver/blood supply , Liver/metabolism , Liver/pathology , Liver Cirrhosis/metabolism , Liver Cirrhosis/prevention & control , Liver Regeneration/physiology , Lymphokines/genetics , Lymphokines/metabolism , Mice , Platelet-Derived Growth Factor/genetics , Platelet-Derived Growth Factor/metabolism , Signal Transduction/drug effects , Zinc FingersABSTRACT
One of the major challenges in using pancreatic cancer patient-derived organoids (PDOs) in precision oncology is the time from biopsy to functional characterization. This is particularly true for endoscopic ultrasound-guided fine-needle aspiration biopsies, typically resulting in specimens with limited tumor cell yield. Here, we tested conditioned media of individual PDOs for cell-free DNA to detect driver mutations already early on during the expansion process to accelerate the genetic characterization of PDOs as well as subsequent functional testing. Importantly, genetic alterations detected in the PDO supernatant, collected as early as 72 hours after biopsy, recapitulate the mutational profile of the primary tumor, indicating suitability of this approach to subject PDOs to drug testing in a reduced time frame. In addition, we demonstrated that this workflow was practicable, even in patients for whom the amount of tumor material was not sufficient for molecular characterization by established means. Together, our findings demonstrate that generating PDOs from very limited biopsy material permits molecular profiling and drug testing. With our approach, this can be achieved in a rapid and feasible fashion with broad implications in clinical practice.
Subject(s)
Biomarkers, Tumor/genetics , Cell-Free Nucleic Acids/analysis , Cell-Free Nucleic Acids/genetics , Organoids/pathology , Pancreatic Neoplasms/pathology , Precision Medicine , Animals , Apoptosis , Biomarkers, Tumor/analysis , Cell Proliferation , Female , Humans , Mice , Mice, Nude , Organoids/metabolism , Pancreatic Neoplasms/genetics , Tumor Cells, Cultured , Xenograft Model Antitumor AssaysABSTRACT
Between 2007 and 2016, 140 consecutive patients who underwent resection of colorectal cancer with simultaneous liver metastases at a single university hospital were retrospectively analysed. In order to gather information regarding potential survival differences for nâ¯=â¯68 simultaneous versus nâ¯=â¯72 staged resections of the colorectal primary tumor and the liver metastases, Clinical, histopathological, serological, and survival data were compared for those two patient groups. The rate of simultaneous tumor resections increased from approximately 25% in 2007 to >75% in 2016. There was no difference in tumor specific survival for patients with simultaneous vs. staged resection (pâ¯=â¯0.631). This effect continued after excluding patients with extrahepatic metastases (pâ¯=â¯0.440). Further, neoadjuvant treatment did not lead to differences in the tumor-specific survival (pâ¯=â¯0.123). Factors associated with an increased tumor-specific survival were low ASA score (p < 0.001), low number of tumor-affected lymph nodes (p < 0.001), histological grading G1/2 (pâ¯=â¯0.001), and a low number of liver metastases (pâ¯=â¯0.044). There was no significant survival difference for the primary tumor stage (pT), the Clavien-Dindo complication rate, the resection status (R0), and minor versus major hepatectomies.
ABSTRACT
BACKGROUND: For patients with colorectal cancer and synchronous liver metastasis, either a simultaneous, or a two-staged resection of the primary tumor and the liver metastases is possible. There are currently no guidelines preferring one approach to the other. MATERIAL AND METHODS: Consecutive patients who underwent hepatic resection at our university hospital from 2007-2016 were included. Clinical, histopathologic, serologic, and survival data were analyzed. The primary end point was tumor-specific survival for patients with simultaneous versus staged resections. RESULTS: Of all 140 patients, 68 underwent simultaneous resection and 72 underwent staged resection. The characteristics of both groups were comparable. Patients with simultaneous resections had a shorter duration of cumulative operation time (299 versus 460 min; P = 0.003) and a shorter cumulative length of hospital stay (23 versus 43 d; P = 0.002). Perioperative mortality (P = 0.257) did not differ significantly; however, patients with simultaneous resections had higher rates of grade 2 complications according to Clavien-Dindo (P < 0.001). Tumor-specific 1-y survival was 85 ± 5% for simultaneous and 83 ± 5% for staged resection (P = 0.631). On multivariable analysis, pT4 (P = 0.038), pN3 (P = 0.003), and G3/4 (P = 0.041) of the primary tumor and postoperative complications (Clavien-Dindo 3/4/5, P = 0.003) were poor prognostic factors regarding tumor-specific survival. CONCLUSIONS: This is one of the largest and most thoroughly documented retrospective single-center studies of consecutive patients with synchronous hepatic metastases. Simultaneous resection of colorectal cancer together with hepatic metastases is a safe procedure in selected patients and does not have a significant influence on long-term survival.
Subject(s)
Carcinoma/surgery , Colorectal Neoplasms/surgery , Digestive System Surgical Procedures/statistics & numerical data , Liver Neoplasms/surgery , Adult , Aged , Aged, 80 and over , Carcinoma/mortality , Carcinoma/secondary , Colorectal Neoplasms/mortality , Colorectal Neoplasms/pathology , Female , Germany/epidemiology , Humans , Liver/pathology , Liver/surgery , Liver Neoplasms/mortality , Liver Neoplasms/secondary , Male , Middle Aged , Postoperative Complications/epidemiology , Retrospective Studies , Young AdultABSTRACT
Several studies have shown that over 70 different microRNAs are aberrantly expressed in pancreatic ductal adenocarcinoma (PDAC), affecting proliferation, apoptosis, metabolism, EMT and metastasis. The most important genetic alterations driving PDAC are a constitutive active mutation of the oncogene Kras and loss of function of the tumour suppressor Tp53 gene. Since the MicroRNA 34a (Mir34a) is a direct target of Tp53 it may critically contribute to the suppression of PDAC. Mir34a is epigenetically silenced in numerous cancers, including PDAC, where Mir34a down-regulation has been associated with poor patient prognosis. To determine whether Mir34a represents a suppressor of PDAC formation we generated an in vivo PDAC-mouse model harbouring pancreas-specific loss of Mir34a (KrasG12D; Mir34aΔ/Δ). Histological analysis of KrasG12D; Mir34aΔ/Δ mice revealed an accelerated formation of pre-neoplastic lesions and a faster PDAC development, compared to KrasG12D controls. Here we show that the accelerated phenotype is driven by an early up-regulation of the pro-inflammatory cytokines TNFA and IL6 in normal acinar cells and accompanied by the recruitment of immune cells. Our results imply that Mir34a restrains PDAC development by modulating the immune microenvironment of PDAC, thus defining Mir34a restauration as a potential therapeutic strategy for inhibition of PDAC development.
Subject(s)
Carcinoma, Pancreatic Ductal/pathology , Down-Regulation , MicroRNAs/genetics , Pancreatic Neoplasms/pathology , Animals , Carcinoma, Pancreatic Ductal/genetics , Gene Expression Regulation, Neoplastic , High-Throughput Nucleotide Sequencing , Interleukin-6/metabolism , Mice , Pancreatic Neoplasms/genetics , Phenotype , Proto-Oncogene Proteins p21(ras)/genetics , Sequence Analysis, RNA , Tumor Necrosis Factor-alpha/metabolismABSTRACT
Several studies have suggested an oncogenic role for the neural Wiskott-Aldrich syndrome protein (N-WASP, encoded by the Wasl gene), but thus far, little is known about its function in pancreatic ductal adenocarcinoma (PDAC). In this study, we performed in silico analysis of WASL expression in PDAC patients and found a correlation between low WASL expression and prolonged survival. To clarify the role of Wasl in pancreatic carcinogenesis, we used 2 oncogenic Kras-based PDAC mouse models with pancreas-specific Wasl deletion. In line with human data, both mouse models had an increased survival benefit due to either impaired tumor development in the presence of the tumor suppressor Trp53 or the delayed tumor progression and senescent phenotype upon genetic ablation of Trp53. Mechanistically, loss of Wasl resulted in cell-autonomous senescence through displacement of the N-WASP binding partners WASP-interacting protein (WIP) and p120ctn; vesicular accumulation of GSK3ß, as well as YAP1 and phosphorylated ß-catenin, which are components of the destruction complex; and upregulation of Cdkn1a(p21), a master regulator of senescence. Our findings, thus, indicate that Wasl functions in an oncogenic manner in PDAC by promoting the deregulation of the p120-catenin/ß-catenin/p21 pathway. Therefore, strategies to reduce N-WASP activity might improve the survival outcomes of PDAC patients.
Subject(s)
Neoplasms, Experimental/metabolism , Pancreatic Neoplasms/metabolism , Wiskott-Aldrich Syndrome Protein, Neuronal/deficiency , Animals , Humans , Mice , Mice, Transgenic , Neoplasms, Experimental/genetics , Neoplasms, Experimental/pathology , Pancreatic Neoplasms/genetics , Pancreatic Neoplasms/pathology , Tumor Suppressor Protein p53/genetics , Tumor Suppressor Protein p53/metabolism , Wiskott-Aldrich Syndrome Protein, Neuronal/metabolismABSTRACT
Liver sinusoidal endothelial cells (LSECs) control organ functions, metabolism, and development through the secretion of angiokines. LSECs express hepatocyte growth factor (Hgf), which is involved in prenatal development, metabolic homeostasis, and liver regeneration. This study aimed to elucidate the precise contribution of LSEC-derived Hgf in physiological homeostasis and liver regeneration. Stab2-iCretg/wt;Hgffl/fl (HgfΔLSEC) mice were generated to abrogate Hgf expression selectively in LSECs from early fetal development onwards, to study global development, metabolic and endothelial zonation, and organ functions as well as liver regeneration in response to 70% partial hepatectomy (PH). Although zonation and liver/body weight ratios were not altered, total body weight and total liver weight were reduced in HgfΔLSEC. Necrotic organ damage was more marked in HgfΔLSEC mice, and regeneration was delayed 72 hours after PH. This was associated with decreased hepatocyte proliferation at 48 hours after PH. Molecularly, HgfΔLSEC mice showed down-regulation of Hgf/c-Met signaling and decreased expression of Deptor in hepatocytes. In vitro knockdown of Deptor was associated with decreased proliferation. Therefore, angiocrine Hgf controls hepatocyte proliferation and susceptibility to necrosis after partial hepatectomy via the Hgf/c-Met axis involving Deptor to prevent excessive organ damage.
Subject(s)
Body Size , Cell Proliferation , Hepatocyte Growth Factor/physiology , Hepatocytes/cytology , Liver Diseases/prevention & control , Liver Regeneration , Organogenesis/physiology , Animals , Cell Adhesion Molecules, Neuronal/physiology , Endothelium/cytology , Endothelium/metabolism , Female , Hepatectomy , Hepatocytes/physiology , Homeostasis , Liver Diseases/metabolism , Liver Diseases/pathology , Male , Mice , Mice, Knockout , Paracrine Communication , Signal TransductionABSTRACT
BACKGROUND: Cap-assisted colonoscopy is frequently used to facilitate adenoma detection during endoscopy. However, data on how cap assistance influences polyp resection are scarce. We aimed to evaluate the impact of cap assistance with the Endocuff vision device (EVD) on the resection time for colorectal polyps in patients undergoing colonoscopy. METHODS : A randomized, prospective study was performed in a university hospital in Germany. A total of 250 patients were randomly assigned 1:1 to undergo either colonoscopy with the EVD (EVD arm) or standard colonoscopy without the use of a cap (standard arm). The primary outcome was the average duration of polypectomy. Secondary outcomes included adenoma detection rate, cecal and ileal intubation times, and propofol dosage. RESULTS: The use of EVD led to a significant reduction in the median polypectomy time in the EVD vs. standard arm (54 vs. 80 seconds, respectively; Pâ=â0.02). This effect was strongest for polyps ≥â6âmm. Compared with the standard group, Endocuff assistance also resulted in a shorter cecal intubation time (6 vs. 8 minutes; Pâ=â0.03) and overall colonoscopy time (23 vs. 27 minutes; Pâ=â0.02). In contrast, no difference in withdrawal time was observed. The polyp and adenoma detection rates did not differ significantly between the two groups. CONCLUSION: Endocuff-assisted colonoscopy reduces the duration of polypectomy, which may be due to a more stable scope position during resection. Further studies are needed to investigate whether comparable effects will be seen for other interventions, such as clipping or biopsy sampling.
Subject(s)
Colonic Polyps , Colonoscopy , Cecum , Colonic Polyps/surgery , Germany , Humans , Ileum , Prospective Studies , Reference StandardsABSTRACT
OBJECTIVES: The importance of the Calcitonin-gene-related-peptide-pathway (CGRP) as neuronal modulator of innate immune responses in mice has been previously demonstrated. The CGRP-receptor is composed of two subunits: the receptor-activity-modifying-protein-1 (RAMP1) and the calcitonin-receptor-like-receptor (CLR). CGRP can influence immune cells and their capacity of producing inflammatory cytokines. Using a RAMP1 knockout-mouse (RAMP1-/-) we examined the role of the CGRP-receptor in the acute-phase of cerulein-induced pancreatitis. METHODS: Hourly cerulein-injections for a period of 8â¯h in RAMP1-/- and wild-type mice were performed. To compare severity and extent of inflammation in RAMP1-/- and wild-type mice, histological analyses were done and cytokine levels were assessed using qRT-PCR 8â¯h, 24â¯h, 2 days, and 7 days post-cerulein-treatment. Furthermore, serum activities of LDH and lipase were determined. RESULTS: After 8â¯h RAMP1-/- mice showed a higher pancreas-to-body-weight-ratio, increased tissue edema and immune cell infiltration with higher amount of F4/80-positive cells as compared to wild-type mice. Overall infiltration of immune cells at 24â¯h was increased in RAMP1-/- mice and composed predominantly of MPO-positive neutrophils. In addition, after 24â¯h RAMP1-/- mice presented a higher pancreas-to-body-weight-ratio, higher expression of Ccl3, Il6, and Il1b and increased number of cleaved caspase 3 positive cells. Serum lipase correlated with the extent of tissue damage in RAMP1-/- compared to wild-type mice 24â¯h post-cerulein treatment. CONCLUSION: Mice lacking RAMP1 showed increased inflammation, tissue edema, and pancreas injury particularly in the early phase of acute pancreatitis. This study highlights the essential role of CGRP for dampening the innate immune response in acute pancreatitis.
Subject(s)
Immunity, Innate/genetics , Pancreatitis/genetics , Pancreatitis/immunology , Receptor Activity-Modifying Protein 1/genetics , Acute Disease , Animals , Ceruletide , Cytokines/blood , Female , Inflammation/chemically induced , Inflammation/pathology , L-Lactate Dehydrogenase/metabolism , Lipase/analysis , Lipase/metabolism , Mice , Mice, Inbred C57BL , Mice, Knockout , Neutrophil Infiltration , Organ Size , Pancreatitis/chemically induced , Receptor Activity-Modifying Protein 1/immunologyABSTRACT
Brahma-related gene 1 (Brg1), a catalytic subunit of the SWItch/Sucrose Non-Fermentable (SWI/SNF) complex, is known to be involved in proliferative cell processes. Liver regeneration is initiated spontaneously after injury and leads to a strong proliferative response. In this study, a hepatocyte-specific Brg1 gene knockout mouse model was used to analyse the role of Brg1 in liver regeneration by performing a 70% partial hepatectomy (PH). After PH, Brg1 was significantly upregulated in wildtype mice. Mice with hepatocyte-specific Brg1 gene knockout showed a significantly lower liver to body weight ratio 48 h post-PH concomitant with a lower hepatocellular proliferation rate compared to wildtype mice. RNA sequencing demonstrated that Brg1 controlled hepatocyte proliferation through the regulation of the p53 pathway and several cell cycle genes. The data of this study reveal a crucial role of Brg1 for liver regeneration by promoting hepatocellular proliferation through modulation of cell cycle genes and, thus, identify Brg1 as potential target for therapeutic approaches.
Subject(s)
Cell Cycle , DNA Helicases/metabolism , Hepatectomy , Liver Regeneration , Nuclear Proteins/metabolism , Transcription Factors/metabolism , Animals , Cell Proliferation , Liver/growth & development , Male , Mice, Knockout , Signal Transduction , Tumor Suppressor Protein p53/metabolismABSTRACT
Peroxisome Proliferator-Activated Receptor gamma (PPARγ) is a nuclear receptor demonstrated to play an important role in various biological processes. The aim of this study was to determine the effect of PPARγ on liver regeneration upon partial hepatectomy (PH) in mice. Mice were subjected to two-thirds PH. Before surgery, mice were either treated with the PPARγ agonist rosiglitazone, the PPARγ antagonist GW9662 alone, or with the c-met inhibitor SGX523. Liver-to-body-weight ratio, lab values, and proliferation markers were assessed. Components of the PPARγ-specific signaling pathway were identified by western blot and qRT-PCR. Our results show that liver regeneration is being inhibited by rosiglitazone and accelerated by GW9662. Inhibition of c-Met by SGX523 treatment abrogates GW9662-induced liver regeneration and hepatocyte proliferation. Hepatocyte growth factor (HGF) protein levels were significantly downregulated after rosiglitazone treatment. Activation of HGF/c-Met pathways by phosphorylation of c-Met and ERK1/2 were inhibited in rosiglitazone-treated mice. In turn, blocking phosphorylation of c-Met significantly abrogated the augmented effect of GW9662 on liver regeneration. Our data support the concept that PPARγ abrogates liver growth and hepatocellular proliferation by inhibition of the HGF/c-Met/ERK1/2 pathways. These pathways may represent potential targets in response to liver disease and could impact on the development of molecular therapies.
Subject(s)
Hepatocyte Growth Factor/metabolism , Liver Regeneration/physiology , MAP Kinase Signaling System/physiology , PPAR gamma/metabolism , Proto-Oncogene Proteins c-met/metabolism , Signal Transduction/physiology , Anilides/pharmacology , Animals , Cell Proliferation/drug effects , Cell Proliferation/physiology , Down-Regulation/drug effects , Female , Hepatectomy/methods , Hepatocytes/drug effects , Hepatocytes/metabolism , Hepatocytes/physiology , Liver/drug effects , Liver/metabolism , Liver/physiology , Liver Regeneration/drug effects , MAP Kinase Signaling System/drug effects , Mice , Mice, Inbred C57BL , Pyridazines/pharmacology , Rosiglitazone/pharmacology , Signal Transduction/drug effects , Triazoles/pharmacologyABSTRACT
The chromatin remodeler complex SWI/SNF plays an important role in physiological and pathological processes. Brahma related gene 1(BRG1), a catalytic subunit of the SWI/SNF complex, is known to be mutated in hepatocellular carcinoma (HCC). However, its role in HCC remains unclear. Here, we investigate the role of BRG1 on cell growth and invasiveness as well as its effect on the expression of putative target genes. Expression of BRG1 was examined in human liver tissue samples and in HCC cell lines. In addition, BRG1 was silenced in human HCC cell lines to analyse cell growth and invasiveness by growth curves, colony formation assay, invasion assay and the expression of putative target genes. BRG1 was found to be significantly increased in HCC samples compared to non-HCC samples. In addition, a declined proliferation rate of BRG1-silenced human HCC cell lines was associated with a decrease of expression of cyclin family members. In line with a decreased invasiveness of BRG1-siRNA-treated human HCC cell lines, down-regulation of MMP7 was detected. These results support the hypothesis that overexpression of BRG1 increases cell growth and invasiveness in HCC. Furthermore, the data highlight cyclin B, E and MMP7 to be associated with BRG1 during hepatocarcinogenesis.
Subject(s)
Carcinoma, Hepatocellular/metabolism , Cell Proliferation , DNA Helicases/genetics , Liver Neoplasms/metabolism , Nuclear Proteins/genetics , Transcription Factors/genetics , Carcinogenesis , Carcinoma, Hepatocellular/genetics , Carcinoma, Hepatocellular/pathology , Cyclins/genetics , Cyclins/metabolism , DNA Helicases/metabolism , Gene Expression Regulation, Neoplastic , Hep G2 Cells , Humans , Liver/embryology , Liver Neoplasms/genetics , Liver Neoplasms/pathology , Matrix Metalloproteinase 7/genetics , Matrix Metalloproteinase 7/metabolism , Neoplasm Invasiveness , Nuclear Proteins/metabolism , Transcription Factors/metabolism , Up-RegulationABSTRACT
OBJECTIVES: Pancreatic ductal adenocarcinoma (PDAC) is thought to derive from different precursor lesions including the recently identified atypical flat lesions (AFL). While all precursor lesions and PDAC share ductal characteristics, there is an ongoing debate about the cellular origin of the different PDAC precursor lesions. In particular, pancreatic acinar cells have previously been shown to display a remarkable plasticity being able to undergo ductal dedifferentiation in the context of oncogenic stimuli. METHODS: Histological analyses were performed in a murine PDAC model that specifically expresses oncogenic Kras in adult pancreatic acinar cells. Occurrence, characterization, and lineage tracing of AFLs were investigated. RESULTS: Upon expression of oncogenic Kras in adult pancreatic acinar cells, AFLs with typical morphology and expression profile arise. Lineage tracing confirmed that the AFLs were of acinar origin. CONCLUSIONS: Using a murine PDAC model, this study identifies pancreatic acinar cells as a cellular source for AFLs.
Subject(s)
Acinar Cells/pathology , Carcinoma, Pancreatic Ductal/pathology , Pancreatic Neoplasms/pathology , Animals , Carcinoma, Pancreatic Ductal/chemically induced , Carcinoma, Pancreatic Ductal/metabolism , Cell Differentiation , Cell Transformation, Neoplastic/pathology , Estrogen Antagonists , Immunohistochemistry , Mice , Pancreatic Neoplasms/chemically induced , Pancreatic Neoplasms/metabolism , Proto-Oncogene Proteins p21(ras)/biosynthesis , Proto-Oncogene Proteins p21(ras)/genetics , TamoxifenABSTRACT
Perturbation of pancreatic acinar cell state can lead to acinar-to-ductal metaplasia (ADM), a precursor lesion to the development of pancreatic ductal adenocarcinoma (PDAC). In the pancreas, Notch signaling is active both during development and in adult cellular differentiation processes. Hes1, a key downstream target of the Notch signaling pathway, is expressed in the centroacinar compartment of the adult pancreas as well as in both preneoplastic and malignant lesions. In this study, we used a murine genetic in vivo approach to ablate Hes1 in pancreatic progenitor cells (Ptf1a+/Cre; Hes1fl/fl). Using this model, we studied the role of Hes1 in both acinar cell plasticity and pancreatic regeneration after caerulein-induced pancreatitis and in KrasG12D-driven PDAC development. We show that, although pancreatic development is not perturbed on the deletion of Hes1, terminal acinar differentiation in the adult pancreas is compromised. Moreover, the loss of Hes1 leads to the impaired regeneration of the exocrine compartment, accelerated fatty metaplasia, and persistent ADM after acute caerulein-induced pancreatitis. In KrasG12D-driven carcinogenesis, Hes1 ablation resulted in increased ADM, decreased formation of high-grade pancreatic intraepithelial neoplasias, and accelerated development of PDAC with shortened survival time. In conclusion, Hes1 plays a key role in acinar cell integrity and plasticity on cellular insults. Furthermore, Hes1 is an essential component of the pancreatic intraepithelial neoplasias-to-PDAC route in KrasG12D-driven mouse pancreatic carcinogenesis.