K. Thanki1, M. Nicholls1, M. Maskey1, C. Phillips1, P. Johnson1, J. R. Zatarain1, K. Modis1, S. Qiu3, I. V. Pinchuk2, M. R. Hellmich1, C. Chao1 1University Of Texas Medical Branch,Surgery,Galveston, TX, USA 2University Of Texas Medical Branch,Internal Medicine,Galveston, TX, USA 3University Of Texas Medical Branch,Pathology,Galveston, TX, USA
Introduction: Hydrogen sulfide (H2S) is an important gasotransmitter that is anti-inflammatory at physiological levels. H2S is produced, in large part, by two enzymes in the reverse transsulfuration pathway: cystathionine-γ-lyase (CSE) and cystathionine-β-synthase (CBS). In mouse models of acute colitis, the activities of CSE and CBS are upregulated. Pharmacological inhibition or gene knockout (KO) of CSE worsens inflammation and delays tissue repair. However the role of CSE in chronic inflammatory conditions, such as ulcerative colitis (UC) and UC-associated carcinogenesis (UCAC) is unknown.
Methods: C57BL/6 CSE knockout (CSE KO) mice and wild-type (WT) controls (N = 148) were given a single injection of the mutagen azoxymethane (AOM) and then randomized to receive either 1, 2 or 3 cycles of the colitis-inducer dextran sodium sulfate (DSS). Each cycle of colitis was induced by administration of 2% DSS in drinking water for a period of seven days followed by 14 days of regular drinking water. Tissues (colon, liver, and lymph nodes) were harvested at 21, 42, 63 and 80 days post AOM injection. Total tumor burden and inflammation were quantified by gross and histological examination using an established scoring system measuring inflammatory infiltrate, mucosal structure (crypt height, epithelial cell loss) and ulceration. CSE mRNA and protein levels from human normal mucosal and UC specimens were determined using qPCR and western blotting, respectively.
Results: Colonic mucosa from UC patients (n=10) exhibited a 2.9-fold decrease in CSE mRNA (qPCR, p<0.05) and protein expression when compared to normal mucosa (n=8). CSE KO mice demonstrate comparable inflammation to WT mice in response to repeated DSS treatment. KO mice demonstrate accelerated UCAC and disease progression as reflected by increased number and area of tumors (i.e., tumor burden) at 21 days (Fig 1A,B). At all other later time points, there were no significant differences in tumor burden. The AOM treatment alone did not cause carcinogenesis in either WT or CSE KO mice.
Conclusion: Our data shows that the colon mucosa from UC patients expresses significantly less CSE, an important producer of H2S. In a mouse model of UCAC, we show that absence of CSE does not protect colon mucosa from inflammation but does accelerate the time to colitis-associated tumor formation, and increases total tumor burden. Theses data suggest that CSE does not impact the inflammatory response but is important in normal mucosal restitution after injury, protecting the colon from UCAC.
