Tissue fibrosis is one of the characteristics of chronic pancreatitis and pancreatic adenocarcinoma. Activated pancreatic stellate cells (PSC) play a central role in this process. However, analysis of the molecular mechanisms leading to PSC activation is hampered by the lack of an established human PSC line. To overcome this problem, we immortalized and characterized primary human PSC. The cells were isolated by the outgrowth method and were immortalized by transfection with SV40 large T antigen and human telomerase (hTERT). Primary human PSC served as controls. An immortalized line, RLT-PSC, was analyzed for the expression of stellate cell markers. Moreover, the effects of transforming growth factor b 1(TGFb1) or platelet-derived growth factor stimulation and of cultivation on basement membrane components or N-acetylcysteine (NAC) treatment on gene and protein expression and proliferation were analyzed. Immortal RLT-PSC cells retained the phenotype of activated PSC proven by the expression of a-smooth muscle actin (aSMA), vimentin, desmin and glial fibrillary acidic protein (GFAP). TGFb1 treatment upregulated the expression of aSMA, collagen type I (Col I), fibronectin and TGFb1. Incubation of RLT-PSC cells and primary human activated PSC on Matrigel plus NAC treatment resulted in a deactivated phenotype as evidenced by a decrease of aSMA, connective tissue growth factor and Col I expression and by a decreased proliferation of the cells. Moreover, this treatment restored the ability of the cells to store vitamin A in cytoplasmic vesicles. In conclusion, we have established an immortal pancreatic stellate cell line, without changing the characteristic phenotype. Importantly, we were able to demonstrate that besides soluble factors, the matrix surrounding PSC plays a pivotal role in the maintenance of the activation process of PSC. Cultivation of activated PSC on a reconstituted basement membrane plus treatment with NAC was able to deactivate the cells, thus pointing to the possibility of an antifibrosis therapy in chronic pancreatitis.

The prognosis of pancreatic adenocarcinoma is poor and agent ifosfamide to toxic metabolites. Administration of current treatment ineffective. A novel treatment strategy is ifosfamide to tumour-bearing mice that were recipients of described here using a mouse model system for pancreatic implanted encapsulated cells results in partial or even comcancer. Cells that have been genetically modified to plete tumour ablation. These results suggest that in situ express the cytochrome P450 2B1 enzyme are encapsuchemotherapy with genetically modified cells in an lated in cellulose sulphate and implanted into pre-estabimmunoprotected environment may prove useful for lished tumours derived from human pancreatic cells. application in man. Cytochrome P450 2B1 converts the chemotherapeutic