Mechanisms Driving Fasting-Induced Protection from Genotoxic Injury in the Small Intestine

Genotoxic agents like doxorubicin (DXR) can cause damage to the intestines that can be ameliorated by fasting. How fasting is protective and the optimal timing of fasting and refeeding remain unclear. Here, our analysis of fasting/refeeding-induced global intestinal transcriptional changes revealed metabolic shifts and implicated the cellular energetic hub mTORC1 in protecting from DXR-induced DNA damage. Our analysis of specific transcripts and proteins in intestinal tissue and tissue extracts showed that fasting followed by refeeding at the time of DXR administration reduced damage and caused a spike in mTORC1 activity. However, continued fasting after DXR prevented the mTORC1 spike and damage reduction. Surprisingly, the mTORC1 inhibitor, rapamycin, did not block fasting/refeeding-induced reduction in DNA damage, suggesting that increased mTORC1 is dispensable for protection against the initial DNA damage response. In Ddit4 -/- mice (DDIT4 functions to regulate mTORC1 activity), fasting reduced DNA damage and increased intestinal crypt viability versus ad libitum-fed Ddit4 -/- mice. Fasted/refed Ddit4 -/- mice maintained body weight, with increased crypt proliferation by 5 days post-DXR, while ad libitum-fed Ddit4 -/- mice continued to lose weight and displayed limited crypt proliferation. Genes encoding epithelial stem cell and DNA repair proteins were elevated in DXR-injured fasted vs ad libitum Ddit4 -/- intestines. Thus, fasting strongly reduced intestinal damage when normal dynamic regulation of mTORC1 was lost. Overall, the results confirm that fasting protects the intestines against DXR and suggests that fasting works by pleiotropic - including both mTORC1-dependent and independent - mechanisms across the temporally dynamic injury response.