Impaired Integrity of DNA After Recovery From Inflammation Causes Persistent Dysfunction of Colonic Smooth Muscle

BACKGROUND & AIMS: Patients with inflammatory bowel disease who are in remission and those who developed irritable bowel syndrome after enteric infection continue to have symptoms of diarrhea or constipation in the absence of overt inflammation, indicating motility dysfunction. We investigated whether oxidative stress during inflammation impairs integrity of the promoter of Cacna1c, which encodes the pore-forming alpha(1C) subunit of Ca(v)1.2b calcium channels. METHODS: We used long-extension polymerase chain reaction to evaluate DNA integrity in tissues from distal colons of rats; trinitrobenzene sulfonic acid was used to induce inflammation. RESULTS: The H2O2 increased in the muscularis externa 1-7 days after inflammation was induced with trinitrobenzene sulfonic acid. The oxidative stress significantly impaired DNA integrity in 2 specific segments of the Cacna1c promoter: -506 to -260 and -2193 to -1542. The impairment peaked at day 3 and recovered partially by day 7 after induction of inflammation; expression of the products of Cacna1c followed a similar time course. Oxidative stress suppressed the expression of nuclear factor-erythroid-2-related factor 2 (Nrf2), an important regulator of anti-oxidant proteins. Intraperitoneal administration of sulforaphane significantly reversed the suppression of Nrf2, oxidative damage in the promoter of Cacna1c, and suppression of Cacna1c on day 7 of inflammation. The inflammation subsided completely by 56 days after inflammation was induced; however, impairment of DNA integrity, expression of Nrf2 and Cacna1c, and smooth muscle reactivity to acetylcholine remained suppressed at this time point. CONCLUSIONS: Oxidative stress during inflammation impairs the integrity of the promoter of Cacna1c; impairment persists partially after inflammation has subsided. Reduced transcription of Cacna1c contributes to smooth muscle dysfunction in the absence of inflammation.