Regulation of the regeneration of intestinal stem cells after irradiation
Liping Liang, Lijun Shen, Guoxiang Fu, Ye Yao, Guichao Li, Yun Deng, Hui Zhang, Menglong Zhou, Yang Wang, Guoqiang Hua, Zhen Zhang
Abstract
BACKGROUND: Radiation enteritis is common in cancer patients with abdominal and pelvic malignant tumors that have received radiotherapy. Regeneration of intestinal stem cells is a critical process for intestine self-repairing post-irradiation. In this study, we attempted to find out the molecules that promote the regeneration of intestinal stem cells to repair the irradiation damage. METHODS: cultured organoids were given 6 Gy X-rays to construct the regeneration of intestinal stem cells. Hematoxylin and eosin (H&E) staining was performed for morphological observation. In situ hybridization was used to detect the expression of Lgr5, and immunofluorescence staining was adopted to detect the expression of CD44. FACS was used to sort CD44 positive cells of crypts. RNA was then extracted, and RNA-Seq was performed. The Wnt11 over-expression cell line was constructed to collect the Wnt11 conditioned medium (CM). RESULTS: The results showed both Lgr5 and CD44 located at the bottom of normal crypts. The expression of Lgr5 was lower at day 3.5, 5, but recovered at day 10 post-irradiation compared with the control. However, the expression of CD44 was higher at day 3.5, 5, but recovered at day 10 post-irradiation compared with the control group. The quantitative real-time polymerase chain reaction (qRT-PCR) assay showed consistent results. RNA-Seq results showed that Wnt11 was over-expressed in the irradiation group. After irradiation adding Wnt11 condition medium to culture, the intestinal organoids resulted in a bigger size and more buddings of the newborn organoids compared with the control group. CONCLUSIONS: The expression of CD44 increases during the radiation-induced regeneration of intestinal stem cells while Lgr5 decreases, adding Wnt11 CM can facilitate the proliferation of the newborn organoids after irradiation. Wnt11 is a potential target to promote the regeneration of intestinal stem cells to repair the radiation injury.