An accumulation of DNA damage over the entire treatment period amidst slow DNA repair and cell turnover is therefore a probable explanation for increasing levels of DNA damage inAtm/mice, taking into account the relatively long lifespan of lymphocytes

An accumulation of DNA damage over the entire treatment period amidst slow DNA repair and cell turnover is therefore a probable explanation for increasing levels of DNA damage inAtm/mice, taking into account the relatively long lifespan of lymphocytes. stability and homeostasis of the gut epithelial barrier. Keywords:Atm, dextran sulfate sodium, ulcerative colitis, DNA damage == Introduction == Long standing inflammation contributes to the development of over 20% of all human cancers, owing to increased cellular proliferation in environments favoring DNA damage and tumorigenesis (2). Ulcerative colitis (UC) is one such chronic inflammatory condition affecting millions of people worldwide, leading to increased risk of colorectal cancer. Duration and severity of inflammation correlate to cumulative probability of cancer development, ranging from 2% at 10 years of colitis to 18% after 30 years (3). A dysregulated immune response to commensal flora caused by transient breaks in the mucosal barrier is thought to be involved in pathogenesis of UC (4), though exact mechanisms remain to be elucidated. TheAtmgene codes for ataxia telangiectasia mutated (ATM), a pleiotropic kinase involved in DNA double strand break recognition, activation of DNA repair proteins, and signaling in cell cycle checkpoint control (5,6). Its deficiency leads to ataxia telangiectasia (A-T), a rare human disease involving defects in T-cell maturation, cerebellar degeneration, radiosensitivity, and increased susceptibility to lymphoma among other cancers (7).Atm/mice foster similar defects as A-T patients (810), and have allowed for studying A-T, mechanisms of DNA damage responses, and carcinogenesis. Notably,Atm/mice do not spontaneously develop colitis BIBS39 or colorectal cancer, however demonstrate elevated levels of oxidative stress compared to wildtype mice. BIBS39 Though inflammation-derived oxidative and nitrative stress leading to DNA damage have been recently implicated in colitis-associated cancers (11,12), the role of ATM in chemically induced intestinal inflammation has not been previously studied. Cyclic administration of dextran sulfate sodium (DSS), a non-genotoxic sulfated polysaccharide, in the drinking water clinically and morphologically resembles ulcerative colitis and its progression to cancer, thus allowing the exclusive study of chronic inflammation in carcinogenesis without the use of tumor promoting carcinogens (13). Direct breaks in the epithelial barrier and the innate immune response are proposed in the pathogenesis of DSS-induced colitis (13,14). Investigators have observed oxidized DNA bases in rat and mice colonic mucosa after acute DSS treatment (15,16), microsatellite instability in colon tissue ofMsh2/mice with chronic DSS treatment (17), and a protective role of alkyladenine DNA glycosylase in colon tumorigenesis (11); suggesting the importance of repairing oxidative DNA damage at the site of inflammation. We have recently found that intestinal inflammation causes systemic genotoxicity in the form of DNA single- and double- stranded breaks and oxidized bases in the peripheral blood of DSS-treated wildtype mice, as Rabbit Polyclonal to RNF149 well as in genetic models of mucosal inflammation (1). In order to explore the systemic role of ATM in pathogenesis of intestinal inflammation, we characterized the sensitivity ofAtm/mice to DSS-induced acute and chronic inflammation in terms of systemic genotoxicity and the consequentially mounted immune response. == Materials BIBS39 and Methods == == Animals == AdultAtm/mice crossed into the parental C57BL/6Jpun/punbackground as previously described (18), heterozygous (Atm+/pun/pun), and wildtype control mice (Atm+/+pun/pun), 12 to 16 weeks old, were housed in a specific pathogen free facility fed a standard rodent chow diet, provided acidified drinking water, and 12:12 light:dark cycle. Food, bedding, and water were autoclaved. All experimental procedures were in accordance with the UCLA Animal Research Committee guidelines. == Induction of experimental colitis == Acute and chronic experimental colitis was induced by administering 3% (w/v) DSS (MP Biomedicals, MW 40,000) dissolved in sterile acidified drinking waterad libitumfor 3 cycles. One cycle of treatment consisted of 7 days of treated water followed by 14 days of normal drinking water. Water was changed daily and symptoms including weight loss, stool consistency, and gross bleeding were also recorded for calculation of the disease activity index (DAI), as described further BIBS39 elsewhere (19). Briefly, a score ranging from 04 was assigned for each measure (weight loss (015% loss), stool consistency (normal to diarrhea), and blood in stool (no blood to gross bleeding)), and the average of these scores was recorded as the DAI. Mice were monitored for 31 days after the end of treatment. == Blood Collection == Peripheral blood was collected via.