Antifibrogenic effects of vitamin D derivatives on mouse pancreatic stellate cells

AIM: To study the molecular effects of three different D-vitamins, vitamin D2, vitamin D3 and calcipotriol, in pancreatic stellate cells (PSCs). METHODS: Quiescent PSCs were isolated from mouse pancreas and activated in vitro by seeding on plastic surfaces. The cells were exposed to D-vitamins as primary cultures (early-activated PSCs) and upon re-culturing (fully-activated cells). Exhibition of vitamin A-containing lipid droplets was visualized by oil-red staining. Expression of α-smooth muscle actin (α-SMA), a marker of PSC activation, was monitored by immunofluorescence and immunoblot analysis. The rate of DNA synthesis was quantified by 5-bromo-2’-deoxyuridine (BrdU) incorporation assays. Real-time PCR was employed to monitor gene expression, and protein levels of interleukin-6 (IL-6) were measured by ELISA. Uptake of proline was determined using (18)F-proline. RESULTS: Sustained culture of originally quiescent PSCs induced cell proliferation, loss of lipid droplets and exhibition of stress fibers, indicating cell activation. When added to PSCs in primary culture, all three D-vitamins diminished expression of α-SMA (to 32%-39% of the level of control cells; P < 0.05) and increased the storage of lipids (scores from 1.97-2.15 on a scale from 0-3; controls: 1.49; P < 0.05). No such effects were observed when Dvitamins were added to fully-activated cells, while incorporation of BrdU remained unaffected under both experimental conditions. Treatment of re-cultured PSCs with Dvitamins was associated with lower expression of IL-6 (-42% to -49%; P < 0.05; also confirmed at the protein level) and increased expression of the vitamin D receptor gene (209%-321% vs controls; P < 0.05). There was no effect of Dvitamins on the expression of transforming growth factor-β1 and collagen type 1 (chain α1). The lowest uptake of proline, a main component of collagen, was observed in calcipotriol-treated PSCs. CONCLUSION: The three D-vitamins inhibit, with similar efficiencies, activation of PSCs in vitro, but cannot reverse the phenotype once the cells are fully activated.