Tumour necrosis factor alpha and interferon gamma induce transcriptional downregulation of aquaporin 3 RNA expression through distinct mechanisms (893.28)

Although aquaporins (AQP) are known to be involved in water movement, the role of AQP3 in the barrier dysfunction that characterizes inflammatory bowel diseases remains unknown. We hypothesized that TNFα and IFNγ were involved in the transcriptional inhibition of AQP3 RNA expression in intestinal inflammation. Methods: AQP3 expression was assessed in C57Bl/6 mice administered dextran sodium sulfate (DSS) to induce colonic inflammation. In vitro , the human adenocarcinoma cell line HT29 was treated with either TNFα or IFNγ and AQP3 pre‐mRNA and mRNA expression were assessed by real‐time RT‐PCR. Cytokine‐responsive transcriptional elements were elucidated using AQP3 promoter driven firefly luciferase expression. Results: AQP3 was decreased early in DSS colitis, with diminished basolateral membrane staining in epithelial cells lining colonic crypts. Similarly, TNFα or IFNγ‐treated HT29 cells had decreased AQP3 pre‐mRNA expression at 2 hr, followed by decreased mRNA expression at 6‐12 hr. IFNγ‐induced transcriptional inhibition of AQP3 expression was reversed using a broad‐spectrum JAK inhibitor (JAK Inhibitor I, 10 µM), but not a JAK2 specific inhibitor (JAK2 Inhibitor II, 10 µM). Truncated AQP3 promoter constructs revealed an IFNγ‐responsive transcriptional element in the 251bp region upstream of the TATA box and a transcriptional suppressor in the 251 ‐ 666bp region upstream of the TATA box. In contrast, TNFα‐induced transcriptional inhibition of AQP3 expression was not reversed by inhibitors of the PI3K, AKT, NF‐κB, ERK/MAPK and p38 MAPK pathways (LY294002, 10 µM; Triciribine, 1 µM; BAY11‐7082, 30 µM; U0126, 10 µM and SB203580, 10 µM respectively). Grant Funding Source : CCFC, CIHR & AIHS