Mucosal inflammation results in altered contractility, calcium mobilization, and CaMKII signaling in colon smooth muscles.
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Ulcerative colitis is an inflammatory bowel disease that causes malnourishment, weight loss, and bloody diarrhea in patients. Inflammation of the gut is associated with colonic dysmotility that changes mucosal functions and contributes to increased urgency and frequency of defecation in ulcerative colitis patients. Since the etiology of IBD is unclear and with incidence on the rise, much research in IBD has focused on understanding the underlying pathophysiology to uncover novel, specific therapeutic targets to accommodate the large population affected.Altered colonic smooth muscle calcium mobilization has been implicated in the development of colonic dysmotility in inflammatory bowel disease. Alterations in key Ca<super>2+</super>-transporter proteins, including the sarcoplasmic reticulum Ca<super>2+</super>-ATPase (SERCA2), have been reported for animal models of colitis. Changes in fura-2 Ca<super>2+</super> signals that indicate reduced intracellular Ca<super>2+</super> stores have been observed in patients with ulcerative colitis and in animal models of the disease. Therefore, the overall goal of this dissertation was to investigate the mechanisms by which disrupted intracellular Ca<super>2+</super> signals contributes to the impaired contractility of colonic circular smooth muscles. Acute colitis was induced in male C57Bl/6 mice with dextran sulfate sodium (DSS) in the drinking water. Proximal and distal colon smooth muscles from non-treated controls and the first five days after DSS treatment were compared. Mechanical and electrical activities of colonic smooth muscles were studied in the presence of agonists and other drugs. Intracellular calcium waves and activity of the Ca<super>2+</super>/calmodulin-dependent protein kinase II (CaMKII) activity were explored. Protein expression of SERCA2, phospholamban (PLB), and γCaMKII was also examined in the DSS murine model. Spontaneous and agonist-evoked contractions of circular smooth muscle strips from dextran sulfate sodium-treated mice were reduced compared to controls. Control levels of contractile activity were not restored with tetrodotoxin. SERCA2 and PLB expression was reduced in colonic smooth muscles from DSS-colitis mice. Intracellular Ca<super>2+</super> waves were altered in DSS-colitis smooth muscle cells in the presence of caffeine. Expression of γCaMKII was increased in distal colon smooth muscle cells, but remained unchanged in the proximal colon from DSS-colitis mice. In addition, total and autonomous CaMKII activity was increased in colonic smooth muscles in DSS-colitis. In distal colon smooth muscles, cytosolic levels of HDAC4 were increased and IκBα levels were decreased in DSS-colitis. The results in this dissertation suggest that disruptions in intracellular Ca<super>2+</super> mobilization due to down-regulation of SERCA2 and PLB expression lead to increased CaMKII activity and cytosolic HDAC4 that may contribute to the dysmotility of colonic smooth muscles in colitis by enhancing NF-κB activity. In addition, CaMKII can signal to other downstream targets that may interfere with smooth muscle cell function via gene transcription or ion channel activation/deactivation, thus contributing to dysmotility. The findings in this dissertation demonstrate a link between smooth muscle Ca<super>2+</super> physiology and inflammatory cytokine signaling and provide novel information to the underlying pathogenesis of colonic dysmotility in ulcerative colitis.