Soluble galectin-3 is a strong, colonic epithelial-cell-derived, lamina propria fibroblast-stimulating factor
- E Lippert1,
- W Falk1,
- F Bataille2,
- T Kaehne3,
- M Naumann3,
- M Goeke4,
- H Herfarth1,
- J Schoelmerich1,
- G Rogler1
- 1Department of Internal Medicine I, University of Regensburg, Regensburg, Germany
- 2Insitute of Pathology, University of Regensburg, Regensburg, Germany
- 3Institute of Experimental Internal Medicine, Otto-von-Guericke University, Magdeburg, Germany
- 4Department of Gastroenterology, Hepatology and Endocrinology, Medizinische Hochschule, Hannover, Germany
- Correspondence to:
Dr G Rogler
Department of Internal Medicine I, University of Regensburg, Regensburg 93042, Germany; gerhard.rogler{at}klinik.uni-regensburg.de
- Received 31 March 2006
- Accepted 10 May 2006
- Published Online First 18 May 2006
Abstract
Background: Colonic lamina propria fibroblasts (CLPFs) play an important role in the pathogenesis of fibrosis and strictures in Crohn’s disease.
Aim: To identify colonic epithelial cell (CEC)-derived factors that activate CLPFs.
Methods: Primary human CECs and CLPFs were isolated from control mucosa and interleukin 8 (IL8) of CLPF cultures was quantified by ELISA. Activation of nuclear factor κB (NF-κB) was shown, and translocation of NF-κB was inhibited by a dominant-negative IκB-expressing adenovirus. The major CLPF-activating and IL8 inducing protein was purified using fast-performance liquid chromatography (HiPrep 16/60 Sephacryl S-200 High Resolution Column) and sodium dodecyl sulphate gel electrophoresis.
Results: A considerable increase in IL8 secretion by CLPFs cultured in CEC-conditioned media compared with that in unconditioned media (155.00 (10.00) pg/µg v 1.434 (0.695) pg/µg) was found. The effect of CEC-conditioned media on CLPF IL8 secretion was NF-κB dependent. A protein or DNA array confirmed the involvement of NF-κB and activator protein-1. Purification of a candidate band isolated with the use of sodium dodecyl sulphate-polyacrylamide gel electrophoresis and subsequent sequencing showed soluble galectin-3 to be a strong CLPF-activating factor. Depletion of galectin-3 from conditioned media by immunoprecipitation abolished the CLPF stimulatory effect.
Conclusions: Using a classical biochemical approach, soluble galectin-3 was identified as a strong activator of CLPFs produced by CEC. Galectin-3 induced NF-κB activation and IL8 secretion in these cells and may be a target for future therapeutic approaches to reduce or avoid stricture formation.
- CEC, colonic epithelial cell
- CLPF, colonic lamina propria fibroblast
- CRD, carbohydrate recognition domain
- DMEM, Dulbecco’s modified Eagle’s medium
- FCS, fetal calf serum
- FPLC, fast-performance liquid chromatography
- IEC, intestinal epithelial cell
- IκB, inhibitor protein κB
- MALDI-TOF, matrix-assisted laser desorption/ionization time-of-flight
- MOI, multiplicity of infection
- NF-κB, nuclear factor κB
- PBS, phosphate buffered saline
- SDS-PAGE, sodium dodecyl sulphate-polyacrylamide gel electrophoresis
- TGF, transforming growth factor
Footnotes
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Published Online First 18 May 2006
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Funding: This work was supported by the Bundesministerium für Bildung und Forschung, BMBF (Kompetenznetz CED) and the Deutsche Forschungsgemeinschaft (SFB 585).
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Competing interests: None declared.
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The isolation of fibroblasts from biopsies and surgical specimens was approved by the University of Regensburg ethics committee.
