A new role for reticulon-4B/NOGO-B in the intestinal epithelial barrier function and inflammatory bowel disease

Abstract

Inflammatory bowel disease (IBD) is characterized by an impaired intestinal barrier function. We aimed to investigate the role of reticulon-4B (RTN-4B/NOGO-B), a structural protein of the endoplasmic reticulum, in intestinal barrier function and IBD. We used immunohistochemistry, confocal microscopy, real-time PCR, and Western blotting to study tissue distribution and expression levels of RTN-4B/NOGO-B in control and IBD samples from mouse and humans. We also targeted RTN-4B/NOGO-B using siRNAs in cultured human intestinal epithelial cell (IECs). Epithelial barrier permeability was assessed by transepithelial electrical resistance (TEER) measurement. RTN-4B/NOGO-B is expressed in the intestine mainly by IECs. Confocal microscopy revealed a colocalization of RTN-4B, E-cadherin, and polymerized actin fibers in tissue and cultured IECs. RTN-4B mRNA and protein expression were lower in the colon of IL-10(-/-) compared with wild-type mice. Colocalization of RTN-4B/E-cadherin/actin was reduced in the colon of IL-10(-/-) mice. Analysis of endoscopic biopsies from IBD patients showed a significant reduction of RTN-4B/NOGO-B expression in inflamed mucosa compared with control. Treatment of IECs with H2O2 reduced TEER values and triggered phosphorylation of RTN-4B in serine 107 residues as well as downregulation of RTN-4B expression. Acute RTN-4B/NOGO-B knockdown by siRNAs resulted in a decreased TEER values and reduction of E-cadherin and α-catenin expression and in the amount of F-actin-rich filaments in IECs. Epithelial RTN-4B/NOGO-B was downregulated in human and experimental IBD. RTN-4B participates in the intestinal epithelial barrier function, most likely via its involvement in E-cadherin, α-catenin expression, and actin cytoskeleton organization at sites of cell-to-cell contacts.

Inflammation and the Intestinal Barrier: Leukocyte–Epithelial Cell Interactions, Cell Junction Remodeling, and Mucosal Repair

“…Decreased reticulon 4 expression has been observed in the intestinal mucosa of people with CD and in IL10-knockout mice with spontaneous colitis. 162 Another study highlighted the role of a polarity protein Par3 in the regulation of TJ protein occludin trafficking and barrier function that was disrupted after exposure to inflammatory cytokines. 163 …”

Disruption of the epithelial barrier during intestinal inflammation: Quest for new molecules and mechanisms

“…Recent studies shed light on some stages of junctional protein exocytosis in the intestinal mucosa and suggest that trafficking defects may significantly contribute to the disruption of the gut barrier during mucosal inflammation. One study targeted a reticulon-4B/NOGO-B protein, which is essential for formation of ER tubules [117]. Downregulation of reticulon-4B in intestinal epithelial cell monolayers attenuated the establishment of the paracellular barrier and decreased expression of E-cadherin, α-catenin, and occludin.…”

“…Downregulation of reticulon-4B in intestinal epithelial cell monolayers attenuated the establishment of the paracellular barrier and decreased expression of E-cadherin, α-catenin, and occludin. Importantly, reticulon-4B expression was found to be downregulated in the intestinal mucosa of CD patients and IL-10 knockout mice that develop spontaneous colitis [117]. These results highlight a possible contribution of impaired ER structure in the disruption of epithelial junctions during intestinal inflammation.…”

Molecular Pathophysiology of Epithelial Barrier Dysfunction in Inflammatory Bowel Diseases

“…A handful of studies have investigated the role of specific proteins in the regulation of intestinal permeability from human tissue samples ( Table 3 ).Some affect the paracellular pathway by acting on the junctional proteins (e.g., Protein C pathway [ 73 ], prion protein (PrPc) [ 23 ], PECAM1) and the actin cytoskeleton (RTN-4B [ 74 ]), while others target epithelial homeostasis e.g., epithelial apoptosis (JAM-A). The studies included in this review focus on specific proteins previously known to contribute to the intestinal mucosal barrier, i.e., whole proteins have been quantified using immunohistochemistry rather than a rigorous quantitative proteomic approach that include quantification of peptide fragments.…”