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THE ROLE OF FIBROBLAST GROWTH FACTOR 9 (FGF9) IN ESOPHAGITIS
DJ Mulder, N Mak, RJ MacLeod, I Pacheco, CJ Justinich
GI Diseases Research Unit, Departments of Pediatrics, Physiology and Medicine, Queen’s University, Kingston, Ontario
BACKGROUND: Fibroblast growth factor 9 (FGF9) is expressed in a variety of tissues and is implicated in epithelial proliferation in response to tissue injury. Esophagitis is a common disorder that may result from acid reflux, allergy or other injury. Esophagitis is characterized by basal hyperplasia and inflammatory cells, including eosinophils, which release mediators including major basic protein(MBP). MBP and its biological mimetic, polyarginine, may activate the calcium sensing receptor (CaSR) on esophageal epithelium. The aim of this study is to characterize FGF9 in esophageal epithelium and explore its role in esophagitis.
METHODS: RT-PCR was used to detect mRNA for FGF9, FGF9 receptors FGFR2 and 3, and genes regulating proliferation, bone morphogenetic protein (BMP)-2, BMP-4 and noggin. ELISA was used to detect FGF9 production from the esophageal epithelial cell line, HET-1A, and from esophageal biopsies. HET-1A cells were transfected with short interfering RNA (siRNA) directed against the CaSR. The proliferative effect of FGF9 on HET-1A cells was studied using BrdU and MTT assays.
RESULTS: FGF9 is constitutively expressed by HET-1A cells and by human esophageal mucosa. FGF9 mRNA in HET-1A increased as a result of treatment with polyarginine or MBP, and secretion of FGF9 protein increased from 141(±33) ng/mL to 348(±70) ng/mL with polyarginine. This is dependent on the CaSR; in cells transfected with siRNA directed against the CaSR, FGF9 is not increased by incubation with MBP. In the HET-1A cells, mRNA is constitutively produced for the major FGF9 receptors, FGFR2 and 3. Treatment with rhFGF9 increased epithelial proliferation 2.1(±0.1) times by MTT and 1.9(±0.1) times by BrdU assay. FGF9 from tissue homogenates (n=5 for each) from esophagitis patients was increased: reflux esophagitis (630±238 ng/mL/mg protein), eosinophilic esophagitis (916±176 ng/mL/mg protein) versus normal patients (382±108 ng/mL/mg protein). In addition, FGF9 treatment increased transcription of proliferative genes (noggin and BMP4) and decreased transcription of the apoptotic gene (BMP2).
CONCLUSION: FGF9 is expressed in esophageal epithelium and may play a role in esophagitis. FGF9 is produced in response to eosinophil products and FGF9 leads to proliferation of HET-1A cells. FGF9 is increased in patients with esophagitis.