The Role of Wnt/β-Catenin Signaling in Enterocyte Turnover during Methotrexate-Induced Intestinal Mucositis in a Rat

Background/Aims: Intestinal mucositis is a common side-effect in patients who receive aggressive chemotherapy. The Wnt signaling pathway is critical for establishing and maintaining the proliferative compartment of the intestine. In the present study, we tested whether Wnt/beta-catenin signaling is involved in methotrexate (MTX)-induced intestinal damage in a rat model. Methods: Non-pretreated and pretreated with MTX Caco-2 cells were evaluated for cell proliferation and apoptosis using FACS analysis. Adult rats were divided into three experimental groups: Control rats; MTX-2 animals were treated with a single dose of MTX given IP and were sacrificed on day 2, and MTX-4 rats were treated with MTX similar to group B and were sacrificed on day 4. Intestinal mucosal damage, mucosal structural changes, enterocyte proliferation, and enterocyte apoptosis were measured at sacrifice. Real Time PCR and Western blot was used to determine the level of Wnt/beta-catenin related genes and protein expression. Results: In the vitro experiment, treatment with MTX resulted in marked decrease in early cell proliferation rates following by a 17-fold increase in late cell proliferation rates compared to early proliferation. Treatment with MTX resulted in a significant increase in early and late apoptosis compared to Caco-2 untreated cells. In the vivo experiment, MTX-2 and MTX-4 rats demonstrated intestinal mucosal hypoplasia. MTX-2 rats demonstrated a significant decrease in FRZ-2, Wnt 3A Wnt 5A, beta-catenin, c-myc mRNA expression and a significant decrease in b-catenin and Akt protein levels compared to control animals. Four days following MTX administration, rats demonstrated a trend toward a restoration of Wnt/beta-catenin signaling especially in ileum. Conclusions: Wnt/beta-catenin signaling is involved in enterocyte turnover during MTX-induced intestinal mucositis in a rat.