Chronic hyperglycemia induces trans-differentiation of human pancreatic stellate cells and enhances the malignant molecular communication with human pancreatic cancer cells

BACKGROUND: Diabetes mellitus is linked to pancreatic cancer. We hypothesized a role for pancreatic stellate cells (PSC) in the hyperglycemia induced deterioration of pancreatic cancer and therefore studied two human cell lines (RLT-PSC, T3M4) in hyperglycemic environment. METHODOLOGY/PRINCIPAL FINDINGS: The effect of chronic hyperglycemia (CHG) on PSCs was studied using mRNA expression array with real-time PCR validation and bioinformatic pathway analysis, and confirmatory protein studies. The stress fiber formation (IC: αSMA) indicated that PSCs tend to transdifferentiate to a myofibroblast-like state after exposure to CHG. The phosphorylation of p38 and ERK1/2 was increased with a consecutive upregulation of CDC25, SP1, cFOS and p21, and with downregulation of PPARγ after PSCs were exposed to chronic hyperglycemia. CXCL12 levels increased significantly in PSC supernatant after CHG exposure independently from TGF-β1 treatment (3.09-fold with a 2.73-fold without TGF-β1, p<0.05). The upregualtion of the SP1 transcription factor in PSCs after CHG exposure may be implicated in the increased CXCL12 and IGFBP2 production. In cancer cells, hyperglycemia induced an increased expression of CXCR4, a CXCL12 receptor that was also induced by PSC's conditioned medium. The receptor-ligand interaction increased the phosphorylation of ERK1/2 and p38 resulting in activation of MAP kinase pathway, one of the most powerful stimuli for cell proliferation. Certainly, conditioned medium of PSC increased pancreatic cancer cell proliferation and this effect could be partially inhibited by a CXCR4 inhibitor. As the PSC conditioned medium (normal glucose concentration) increased the ERK1/2 and p38 phosphorylation, we concluded that PSCs produce other factor(s) that influence(s) pancreatic cancer behaviour. CONCLUSIONS: Hyperglycemia induces increased CXCL12 production by the PSCs, and its receptor, CXCR4 on cancer cells. The ligand-receptor interaction activates MAP kinase signaling that causes increased cancer cell proliferation and migration

Recruitment and Activation of Pancreatic Stellate Cells from the Bone Marrow in Pancreatic Cancer: A Model of Tumor-Host Interaction

BACKGROUND AND AIMS: Chronic pancreatitis and pancreatic cancer are characterised by extensive stellate cell mediated fibrosis, and current therapeutic development includes targeting pancreatic cancer stroma and tumor-host interactions. Recent evidence has suggested that circulating bone marrow derived stem cells (BMDC) contribute to solid organs. We aimed to define the role of circulating haematopoietic cells in the normal and diseased pancreas. METHODS: Whole bone marrow was harvested from male β-actin-EGFP donor mice and transplanted into irradiated female recipient C57/BL6 mice. Chronic pancreatitis was induced with repeat injections of caerulein, while carcinogenesis was induced with an intrapancreatic injection of dimethylbenzanthracene (DMBA). Phenotype of engrafted donor-derived cells within the pancreas was assessed by immunohistochemistry, immunofluorescence and in situ hybridisation. RESULTS: GFP positive cells were visible in the exocrine pancreatic epithelia from 3 months post transplantation. These exhibited acinar morphology and were positive for amylase and peanut agglutinin. Mice administered caerulein developed chronic pancreatitis while DMBA mice exhibited precursor lesions and pancreatic cancer. No acinar cells were identified to be donor-derived upon cessation of cerulein treatment, however rare occurrences of bone marrow-derived acinar cells were observed during pancreatic regeneration. Increased recruitment of BMDC was observed within the desmoplastic stroma, contributing to the activated pancreatic stellate cell (PaSC) population in both diseases. Expression of stellate cell markers CELSR3, PBX1 and GFAP was observed in BMD cancer-associated PaSCs, however cancer-associated, but not pancreatitis-associated BMD PaSCs, expressed the cancer PaSC specific marker CELSR3. CONCLUSIONS: This study demonstrates that BMDC can incorporate into the pancreas and adopt the differentiated state of the exocrine compartment. BMDC that contribute to the activated PaSC population in chronic pancreatitis and pancreatic cancer have different phenotypes, and may play important roles in these diseases. Further, bone marrow transplantation may provide a useful model for the study of tumor-host interactions in cancer and pancreatitis

Tenascin-C Enhances Pancreatic Cancer Cell Growth and Motility and Affects Cell Adhesion through Activation of the Integrin Pathway

Background: Pancreatic cancer (PDAC) is characterized by an abundant fibrous tissue rich in Tenascin-C (TNC), a large ECM glycoprotein mainly synthesized by pancreatic stellate cells (PSCs). In human pancreatic tissues, TNC expression increases in the progression from low-grade precursor lesions to invasive cancer. Aim of this study was the functional characterization of the effects of TNC on biologic relevant properties of pancreatic cancer cells. Methods: Proliferation, migration and adhesion assays were performed on pancreatic cancer cell lines treated with TNC or grown on a TNC-rich matrix. Stable transfectants expressing the large TNC splice variant were generated to test the effects of endogenous TNC. TNC-dependent integrin signaling was investigated by immunoblotting, immunofluorescence and pharmacological inhibition. Results: Endogenous TNC promoted pancreatic cancer cell growth and migration. A TNC-rich matrix also enhanced migration as well as the adhesion to the uncoated growth surface of poorly differentiated cell lines. In contrast, adhesion to fibronectin was significantly decreased in the presence of TNC. The effects of TNC on cell adhesion were paralleled by changes in the activation state of paxillin and Akt. Conclusion: TNC affects proliferation, migration and adhesion of poorly differentiated pancreatic cancer cell lines and migh

Mononuclear cells modulate the activity of pancreatic stellate cells which in turn promote fibrosis and inflammation in chronic pancreatitis

Background: Interactions between mononuclear cells and activated pancreatic myofibroblasts (pancreatic stellate cells; PSC) may contribute to inflammation and fibrosis in chronic pancreatitis (CP). Methods: Markers of fibrosis and inflammation were concomitantly analysed by immunohistochemistry in chronic pancreatitis tissues. In vitro, PSC were stimulated with TNFalpha and LPS. Primary human blood mononuclear cells (PBMC) and PSC were cocultured, followed by analysis of cytokines and extracellular matrix (ECM) proteins. PBMC were derived from healthy donors and CP and septic shock patients. Results: In areas of mononuclear cell infiltration in chronic pancreatitis tissues, there was decreased immunoreactivity for collagen1 and fibronectin, in contrast to areas with sparse mononuclear cells, although PSC were detectable in both areas. LPS and TNFalpha induced collagen1 and fibronectin levels as well as the matrix degradation enzyme MMP-1. Coculture experiments with PSC and PBMC revealed increased fibronectin secretion induced by PBMC. In addition, donor and CP PBMC significantly induced an increase in IL-6, MCP-1 and TGFbeta levels under coculture conditions. Determination of the source of cytokines and ECM proteins by mRNA expression analysis confirmed PSC as major contributors of ECM production. The increase in cytokine expression was PBMC- and also PSC-derived. Conclusion: Mononuclear cells modulate the activity of pancreatic stellate cells, which may in turn promote fibrosis and inflammation

The α2β1 integrin mediates the malignant phenotype on type I collagen in pancreatic cancer cell lines

Pancreatic cancer is characterised by a hallmark desmoplastic response that includes upregulated expression of the extracellular matrix, and type I collagen in particular. Recent studies indicate that pancreatic cancer cells stimulate type I collagen synthesis in adjacent stellate cells, and that this upregulated type I collagen expression promotes the malignant phenotype in tumour cells as defined by increased proliferation, resistance to chemically induced apoptosis, and increased tumorigenesis. The integrin specificity of this interaction between type I collagen and tumour cells was not identified, however. In the present study, we examined eight pancreatic cancer cell lines for adhesion, proliferation, and migration, on types I and IV collagen, fibronectin, laminin, and vitronectin, as well as integrin expression. Our results indicate, for the overwhelming majority of cell lines, that type I collagen promotes the strongest adhesion, proliferation, and migration relative to the other substrates tested. Utilising function-blocking monoclonal antibodies directed against particular integrin subunits in cell adhesion and migration inhibition assays, we demonstrate further that the malignant phenotype on type I collagen is mediated specifically by the α2β1 integrin. These results identify α2β1 integrin-mediated adhesion to type I collagen as a potential therapeutic target in the treatment of pancreatic cancer

Predicting In Vivo Anti-Hepatofibrotic Drug Efficacy Based on In Vitro High-Content Analysis

Background/Aims Many anti-fibrotic drugs with high in vitro efficacies fail to produce significant effects in vivo. The aim of this work is to use a statistical approach to design a numerical predictor that correlates better with in vivo outcomes. Methods High-content analysis (HCA) was performed with 49 drugs on hepatic stellate cells (HSCs) LX-2 stained with 10 fibrotic markers. ~0.3 billion feature values from all cells in >150,000 images were quantified to reflect the drug effects. A systematic literature search on the in vivo effects of all 49 drugs on hepatofibrotic rats yields 28 papers with histological scores. The in vivo and in vitro datasets were used to compute a single efficacy predictor (Epredict). Results We used in vivo data from one context (CCl4 rats with drug treatments) to optimize the computation of Epredict. This optimized relationship was independently validated using in vivo data from two different contexts (treatment of DMN rats and prevention of CCl4 induction). A linear in vitro-in vivo correlation was consistently observed in all the three contexts. We used Epredict values to cluster drugs according to efficacy; and found that high-efficacy drugs tended to target proliferation, apoptosis and contractility of HSCs. Conclusions The Epredict statistic, based on a prioritized combination of in vitro features, provides a better correlation between in vitro and in vivo drug response than any of the traditional in vitro markers considered.Institute of Bioengineering and Nanotechnology (Singapore)Singapore. Biomedical Research CouncilSingapore. Agency for Science, Technology and ResearchSingapore-MIT Alliance for Research and Technology Center (C-185-000-033-531)Janssen Cilag (R-185-000-182-592)Singapore-MIT Alliance Computational and Systems Biology Flagship Project (C-382-641-001-091)Mechanobiology Institute, Singapore (R-714-001-003-271

Temperature stress differentially modulates transcription in meiotic anthers of heat-tolerant and heat-sensitive tomato plants

Contains fulltext : 92405.pdf (publisher's version ) (Open Access

Feedback within the Inter-Cellular Communication and Tumorigenesis in Carcinomas

The classical somatic mutation theory (SMT) of carcinogenesis and metastasis postulates that malignant transformation occurs in cells that accumulate a sufficient amount of mutations in the appropriate oncogenes and/or tumor suppressor genes. These mutations result in cell-autonomous activation of the mutated cell and a growth advantage relative to neighboring cells. However, the SMT cannot completely explain many characteristics of carcinomas. Contrary to the cell-centered view of the SMT with respect to carcinogenesis, recent research has revealed evidence that the tumor microenvironment plays a role in carcinogenesis as well. In this review, we present a new model that accommodates the role of the tumor microenvironment in carcinogenesis and complements the classical SMT. Our “feedback” model emphasizes the role of an altered spatiotemporal communication between epithelial and stromal cells during carcinogenesis: a dysfunctional intracellular signaling in tumorigenic epithelial cells leads to inappropriate cellular responses to stimuli from associated stromal or inflammatory cells. Thus, a positive feedback loop of the information flow between parenchymal and stromal cells results. This constant communication between the stromal cells and the tumor cells causes a perpetually activated state of tumor cells analogous to resonance disaster

Pathways to Injury in Chronic Pancreatitis: Decoding the Role of the High-Risk SPINK1 N34S Haplotype Using Meta-Analysis

Background: The complex interactions between recurrent trypsin-mediated pancreatic injury, alcohol-associated pancreatic injury and SPINK1 polymorphisms in chronic pancreatitis (CP) are undefined. We hypothesize that CP occurs as a result of multiple pathological mechanisms (pathways) that are initiated by different metabolic or environmental factors (etiologies) and may be influenced differentially by downstream genetic risk factors. We tested this hypothesis by evaluating the differences in effect size of the high risk SPINK1 N34S haplotype on CP from multiple etiologies after combining clinical reports of SPINK1 N34S frequency using meta-analysis. Methods and Findings: The Pubmed and the Embase databases were reviewed. We studied 24 reports of SPINK1 N34S in CP (2,421 cases, 4,857 controls) using reported etiological factors as surrogates for pathways and multiple meta-analyses to determine the differential effects of SPINK1 N34S between alcoholic and non-alcoholic etiologies. Using estimates of between-study heterogeneity, we sub-classified our 24 studies into four specific clusters. We found that SPINK1 N34S is strongly associated with CP overall (OR 11.00; 95% CI: 7.59-15.93), but the effect of SPINK1 N34S in alcoholic CP (OR 4.98, 95% CI: 3.16-7.85) was significantly smaller than in idiopathic CP (OR 14.97, 95% C.I. = 9.09-24.67) or tropical CP (OR 19.15, 95% C.I. = 8.83-41.56). Studies analyzing familial CP showed very high heterogeneity suggestive of a complex etiology with an I2 = 80.95%. Conclusion: The small effect of SPINK1 N34S in alcoholic subjects suggests that CP is driven through a different pathway that is largely trypsin-independent. The results also suggest that large effect sizes of SPINK1 N34S in small candidate gene studies in CP may be related to a mixture of multiple etiologic pathways leading to the same clinical endpoint. © 2008 Aoun MD et al