title: Radiation persistently promoted oxidative stress, activated mTOR via PI3K/Akt, and downregulated autophagy pathway in mouse intestine.
abstract: While acute effects of toxic radiation doses on intestine are well established, we are yet to acquire a complete spectrum of sub-lethal radiation-induced chronic intestinal perturbations at the molecular level. We investigated persistent effects of a radiation dose (2 Gy) commonly used as a daily fraction in radiotherapy on oxidants and anti-oxidants, and autophagy pathways, which are interlinked processes affecting intestinal homeostasis. Six to eight weeks old C57BL/6J mice (n=10) were exposed to 2 Gy γ-ray. Mice were euthanized two or twelve months after radiation, intestine surgically removed, and flushed using sterile PBS. Parts of the intestine from jejunal-ilial region were fixed, frozen, or used for intestinal epithelial cell (IEC) isolation. While oxidant levels and mitochondrial status were assessed in isolated IEC, autophagy and oxidative stress related signaling pathways were probed in frozen and fixed samples using PCR-based expression arrays and immunoprobing. Radiation exposure caused significant alterations in the expression level of 26 autophagy and 17 oxidative stress related genes. Immunoblot results showed decreased Beclin1 and LC3-II and increased p62, PI3K/Akt, and mTOR. Flow cytometry data showed increased oxidant production and compromised mitochondrial integrity in irradiated samples. Immunoprobing of intestinal sections showed increased 8-oxo-dG and nuclear PCNA, and decreased autophagosome marker LC3-II in IEC after irradiation. We show that sub-lethal radiation could persistently downregulate anti-oxidants and autophagy signaling, and upregulate oxidant production and proliferative signaling. Radiation-induced promotion of oxidative stress and downregulation of autophagy could work in tandem to alter intestinal functions and have implications for post-radiation chronic gastrointestinal diseases.
authors.affiliation: Department of Biochemistry and Molecular & Cellular Biology, Georgetown University, Washington, DC 20057, USA; Lombardi Comprehensive Cancer Center, Georgetown University, Washington, DC 20057, USA. Electronic address: kd257@georgetown.edu.
authors.forename: Kamal
authors.initials: K
authors.lastname: Datta
authors.affiliation: Department of Biochemistry and Molecular & Cellular Biology, Georgetown University, Washington, DC 20057, USA; Lombardi Comprehensive Cancer Center, Georgetown University, Washington, DC 20057, USA.
authors.forename: Shubhankar
authors.initials: S
authors.lastname: Suman
authors.affiliation: Department of Biochemistry and Molecular & Cellular Biology, Georgetown University, Washington, DC 20057, USA; Lombardi Comprehensive Cancer Center, Georgetown University, Washington, DC 20057, USA; Center of Excellence In Genomic Medicine Research (CEGMR), King Abdulaziz University, Jeddah, Saudi Arabia.
authors.forename: Albert J
authors.initials: AJ
authors.lastname: Fornace
mesh_terms: D000818:Animals; D001343:Autophagy / Q000528:radiation effects; D015536:Down-Regulation / Q000528:radiation effects; D005260:Female; D007413:Intestinal Mucosa / Q000378:metabolism*; D007422:Intestines / Q000528:radiation effects*; D051379:Mice; D008810:Mice, Inbred C57BL; D008928:Mitochondria / Q000378:metabolism / Q000528:radiation effects; D018384:Oxidative Stress / Q000502:physiology / Q000528:radiation effects*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism*; D011833:Radiation Injuries, Experimental / Q000378:metabolism; D015398:Signal Transduction; D058570:TOR Serine-Threonine Kinases / Q000378:metabolism*
chemical_list: D019869:Phosphatidylinositol 3-Kinases; D051057:Proto-Oncogene Proteins c-akt; D058570:TOR Serine-Threonine Kinases; C546843:mTOR protein, mouse
keywords: Autophagy; Intestine; Mitochondria; Oxidative stress; Radiation