Dietary Transforming Growth Factor-Beta 2 (TGF-β2) Supplementation Reduces Methotrexate-Induced Intestinal Mucosal Injury in a Rat

Background/Aims: Dietary supplementation with transforming growth factor-beta (TGF-beta) has been proven to minimize intestinal damage and facilitate regeneration after mucosal injury. In the present study, we evaluated the effects of oral TGF-beta 2 supplementation on intestinal structural changes, enterocyte proliferation and apoptosis following methotrexate (MTX)-induced intestinal damage in a rat and in a cell culture model. Methods: Caco-2 cells were treated with MTX and were incubated with increasing concentrations of TGF-beta 2. Cell apoptosis was assessed using FACS analysis by annexin staining and cell viability was monitored using Trypan Blue assay. Male rats were divided into four experimental groups: Control rats, CONTR-TGF-beta rats were treated with diet enriched with TGF-beta 2, MTX rats were treated with a single dose of methotrexate, and MTX-TGF-beta rats were treated with diet enriched with TGF-b beta 2. Intestinal mucosal damage, mucosal structural changes, enterocyte proliferation and enterocyte apoptosis were determined at sacrifice. Real Time PCR and Western blot were used to determine bax and bcl-2 mRNA, p-ERK, beta-catenin, IL-1B and bax protein expression. Results: Treatment of MTX-pretreated Caco-2 cells with TGF-B2 resulted in increased cell viability and decreased cell apoptosis. Treatment of MTX-rats with TGF-beta 2 resulted in a significant increase in bowel and mucosal weight, DNA and protein content, villus-height (ileum), crypt-depth (jejunum), decreased intestinal-injury score, decreased level of apoptosis and increased cell proliferation in jejunum and ileum compared to the untreated MTX group. MTX-TGF-beta 2 rats demonstrated a lower bax mRNA and protein levels as well as increased bcl-2 mRNA levels in jejunum and ileum compared to MTX group. Treatment with TGF-beta 2 also led to increased pERK, IL-1B and beta-catenin protein levels in intestinal mucosa. Conclusions: Treatment with TGF-beta 2 prevents mucosal-injury, enhances p-ERK and beta-catenin induced enterocyte proliferation, inhibits enterocyte apoptosis and improves intestinal recovery following MTX-induced intestinal-mucositis in rats.