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. 1999 Apr;44(4):534–541. doi: 10.1136/gut.44.4.534

Pancreatic stellate cells are activated by proinflammatory cytokines: implications for pancreatic fibrogenesis

M Apte 1, P Haber 1, S Darby 1, S Rodgers 1, G McCaughan 1, M Korsten 1, R Pirola 1, J Wilson 1
PMCID: PMC1727467  PMID: 10075961

Abstract

BACKGROUND—The pathogenesis of pancreatic fibrosis is unknown. In the liver, stellate cells play a major role in fibrogenesis by synthesising increased amounts of collagen and other extracellular matrix (ECM) proteins when activated by profibrogenic mediators such as cytokines and oxidant stress. 
AIMS—To determine whether cultured rat pancreatic stellate cells produce collagen and other ECM proteins, and exhibit signs of activation when exposed to the cytokines platelet derived growth factor (PDGF) or transforming growth factor β (TGF-β). 
METHODS—Cultured pancreatic stellate cells were immunostained for the ECM proteins procollagen III, collagen I, laminin, and fibronectin using specific polyclonal antibodies. For cytokine studies, triplicate wells of cells were incubated with increasing concentrations of PDGF or TGF-β. 
RESULTS—Cultured pancreatic stellate cells stained strongly positive for all ECM proteins tested. Incubation of cells with 1, 5, and 10 ng/ml PDGF led to a significant dose related increase in cell counts as well as in the incorporation of 3H-thymidine into DNA. Stellate cells exposed to 0.25, 0.5, and 1 ng/ml TGF-β showed a dose dependent increase in α smooth muscle actin expression and increased collagen synthesis. In addition, TGF-β increased the expression of PDGF receptors on stellate cells. 
CONCLUSIONS—Pancreatic stellate cells produce collagen and other extracellular matrix proteins, and respond to the cytokines PDGF and TGF-β by increased proliferation and increased collagen synthesis. These results suggest an important role for stellate cells in pancreatic fibrogenesis. 



Keywords: pancreatic fibrosis; stellate cell activation; cytokines

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Figure 1 .

Figure 1

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Pancreatic stellate cells in culture stained strongly positive for procollagen III, collagen I, laminin, and fibronectin (panels B, D, F, and H respectively) compared with control cells incubated with the appropriate non-immune sera (panels A, C, E, and G respectively). Staining for the collagens and for laminin was perinuclear in distribution; that for fibronectin was diffuse. 


Figure 2 .

Figure 2

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Effect of PDGF on cell counts (five separate cell preparations). Results are expressed as a percentage of control values (cells not incubated with PDGF). Increasing concentrations of PDGF resulted in a dose related increase in cell numbers (*p<0.025, †p<0.001). 


Figure 3 .

Figure 3

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Effect of PDGF on DNA synthesis (five separate cell preparations). DNA synthesis was estimated by measuring the incorporation of 3H-thymidine into TCA precipitable material. Results are expressed as a percentage of control values observed in cells not incubated with PDGF. **p<0.01, ***p<0.001. 


Figure 4 .

Figure 4

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Western blot analysis of cell lysates for αSMA expression. The figure shows a representative immunoblot for expression in cells incubated with and without TGF-β1. A single band was detected in each lane corresponding to the known molecular weight (42 kDa) of αSMA. 


Figure 5 .

Figure 5

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Effect of TGF-β1 on collagen synthesis (five separate cell preparations). *p<0.005. 


Figure 6 .

Figure 6

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Effect of TGF-β1 on expression of the β subunit of the PDGF receptor (PDGF-Rβ). Panels A and B show stellate cells (not exposed to TGF-β1) incubated with non-immune serum (negative control) and an antibody to PDGF-Rβ respectively. Panels C and D show TGF-β1 treated stellate cells incubated with non-immune serum (negative control) and an antibody to PDGF-Rβ respectively. 


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