Hypoxia-Inducible Factor 1–Dependent Induction of Intestinal Trefoil Factor Protects Barrier Function during Hypoxia

Abstract

Mucosal organs such as the intestine are supported by a rich and complex underlying vasculature. For this reason, the intestine, and particularly barrier-protective epithelial cells, are susceptible to damage related to diminished blood flow and concomitant tissue hypoxia. We sought to identify compensatory mechanisms that protect epithelial barrier during episodes of intestinal hypoxia. Initial studies examining T84 colonic epithelial cells revealed that barrier function is uniquely resistant to changes elicited by hypoxia. A search for intestinal-specific, barrier-protective factors revealed that the human intestinal trefoil factor (ITF) gene promoter bears a previously unappreciated binding site for hypoxia-inducible factor (HIF)-1. Hypoxia resulted in parallel induction of ITF mRNA and protein. Electrophoretic mobility shift assay analysis using ITF-specific, HIF-1 consensus motifs resulted in a hypoxia-inducible DNA binding activity, and loading cells with antisense oligonucleotides directed against the α chain of HIF-1 resulted in a loss of ITF hypoxia inducibility. Moreover, addition of anti-ITF antibody resulted in a loss of barrier function in epithelial cells exposed to hypoxia, and the addition of recombinant human ITF to vascular endothelial cells partially protected endothelial cells from hypoxia-elicited barrier disruption. Extensions of these studies in vivo revealed prominent hypoxia-elicited increases in intestinal permeability in ITF null mice. HIF-1–dependent induction of ITF may provide an adaptive link for maintenance of barrier function during hypoxia.

Neutrophil priming by hypoxic preconditioning protects against epithelial barrier damage and enteric bacterial translocation in intestinal ischemia/reperfusion

“…57 Previous studies showed that adaptive hypoxia protects the intestinal epithelial barrier integrity via transcriptional regulation of hypoxiainducible factor 1. [22][23][24] Further investigation of the molecular Figure 9 Depletion of blood neutrophils blocked the hypoxic preconditioning (HPC) protection against ischemia/reperfusion (I/R) injury. Rats exposed to HPC were administered anti-PMN or isotype antibodies, and then subjected to mesenteric I/R challenge.…”

“…[18][19][20][21] Other reports have indicated that adaptive hypoxia protects the intestinal epithelial barrier integrity via transcriptional regulation of hypoxia-inducible factor 1 in experimental colitis models. [22][23][24] It remains unclear whether HPC attenuates mesenteric I/R-induced intestinal mucosal injury and BT.…”

Hypoxia and gastrointestinal disease

“…A number of studies have revealed that HIF elicits a barrier protective program in the intestine [17][18][19][20]. While originally guided by microarray analysis of differentially expressed messenger ribonucleic acid (mRNA) in cultured epithelial cells subjected to hypoxia, these studies have proven robust in a number of animal models of inflammation.…”

“…Third, HIF-dependent epithelial barrier-protective pathways driven by hypoxia tend to be more "nonclassical" regulators of barrier function. Rather than classic junctional proteins such as occludin or claudin(s), hypoxia-induced enhancement of barrier function occurs through diverse pathways, ranging from increased mucin production [21] and molecules that modify mucins (e.g., intestinal trefoil factor) [17], to xenobiotic clearance (P-glycoprotein) [18] to nucleotide metabolism (ecto-5′-nucleotidase, CD73) [19][20] and nucleotide signaling (adenosine A2B receptor) [20] (Fig. 2).…”

AKAP13, a RhoA GTPase-specific Guanine Exchange Factor, Is a Novel Regulator of TLR2 Signaling

“…After 60 h, cells were stimulated with 1 g/ml of palmitoyl-3-cysteine-serine-lysine 4 (pam3CSK4) (Invivogen, San Diego, CA) for 15, 30, or 60 min or with tumor necrosis factor-␣ for 30 min (R & D Systems, Inc., Minneapolis, MN) as a positive control. Nuclear extracts were prepared according to a protocol previously reported (14). Briefly, cells were washed in 1 ml of ice-cold phosphate-buffered saline, collected by centrifugation (3000 rpm for 5 min at 4°C), and incubated in buffer A (10 mM HEPES, 1.5 mM MgCl 2 , 10 mM KCl, 0.1 mM EDTA, 0.1 mM EGTA, 1 mM dithiothreitol, 0.3 mM Na 3 VO 4 , protease inhibitor (complete Mini; Roche Applied Science)) for 10 min.…”