Intestinal paracellular transport or paracellular permeation in-between intestinal epithelial cells is regulated by tight junctions. Patients with Crohn's disease (CD) have a defective intestinal tight junction (TJ) barrier, manifested by an increase in paracellular permeability. The defective intestinal TJ barrier is an important pathogenic factor of CD that allows increased paracellular permeation of toxic luminal antigens, leading to intestinal inflammation. Tumor necrosis factor-? (TNF-?), a pro-inflammatory cytokine, has been shown to play a central role in the intestinal inflammation process of CD. An important pro-inflammatory action of TNF-? is to cause a functional opening of the intestinal TJ barrier, leading to an increase in paracellular permeation of noxious luminal antigens. The TNF-? induced increase in intestinal paracellular permeability has been postulated to be an important mechanism contributing to the intestinal TJ barrier defect in CD and other inflammatory conditions of the gut. The broad objectives of this grant proposal are to elucidate the cellular and molecular mechanisms that mediate the TNF-? induced increase in intestinal TJ permeability and to determine the potential therapeutic targets to prevent the TNF-? induced defect in TJ barrier and subsequent development of intestinal inflammation. To achieve these objectives, we intend to use both in-vitro (consisting of filter-grown Caco-2 intestinal epithelial cells) and in-vivo (mouse intestinal perfusion system) model systems. Our preliminary studies suggested that the TNF-? induced increase in intestinal epithelial TJ permeability was regulated in part by ERK 1/2 and p38 signaling cascade induced increase in myosin light chain kinase (MLCK) gene activity and microRNA induced down-regulation of occludin gene expression. Based on our preliminary studies, we advance a novel hypothesis that the TNF-? induced increase in intestinal TJ permeability is mediated in part by ERK1/2 and p38 signaling cascade induced activation of MLCK gene and modulation of microRNA expression. The proposed specific aims are to: 1) elucidate the intracellular and the molecular processes that mediate the TNF-? induced increase in MLCK gene activity and intestinal TJ permeability;2) delineate the cellular and molecular mechanisms that mediate the TNF-? induced down- regulation of occludin gene and protein expression;and 3) delineate the mechanisms involved in TNF-? induced increase in intestinal permeability in-vivo and determine the therapeutic implications of targeted preservation of TJ barrier function in TNF-? induced intestinal inflammation.

Public Health Relevance

Patients with Crohn's disease have a leaky gut, characterized by an increase in intestinal permeability to harmful antigens in the intestinal lumen. The studies proposed in this grant application seek to provide novel insight into the cellular processes that lead to the leaky gut of Crohn's disease. These studies also seek to identify potential therapeutic targets and strategies to induce therapeutic re-tightening or normalization of leaky gut in Crohn's disease.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK064165-09
Application #
8322029
Study Section
Special Emphasis Panel (ZRG1-DKUS-B (02))
Program Officer
Hamilton, Frank A
Project Start
2003-06-01
Project End
2014-08-31
Budget Start
2012-09-01
Budget End
2013-08-31
Support Year
9
Fiscal Year
2012
Total Cost
$310,727
Indirect Cost
$84,799
Name
University of New Mexico
Department
Internal Medicine/Medicine
Type
Schools of Medicine
DUNS #
868853094
City
Albuquerque
State
NM
Country
United States
Zip Code
87131
Al-Sadi, Rana; Guo, Shuhong; Ye, Dongmei et al. (2016) TNF-α Modulation of Intestinal Tight Junction Permeability Is Mediated by NIK/IKK-α Axis Activation of the Canonical NF-κB Pathway. Am J Pathol 186:1151-65
Nighot, Prashant; Al-Sadi, Rana; Rawat, Manmeet et al. (2015) Matrix metalloproteinase 9-induced increase in intestinal epithelial tight junction permeability contributes to the severity of experimental DSS colitis. Am J Physiol Gastrointest Liver Physiol 309:G988-97
Nighot, Prashant K; Hu, Chien-An Andy; Ma, Thomas Y (2015) Autophagy enhances intestinal epithelial tight junction barrier function by targeting claudin-2 protein degradation. J Biol Chem 290:7234-46
Guo, Shuhong; Nighot, Meghali; Al-Sadi, Rana et al. (2015) Lipopolysaccharide Regulation of Intestinal Tight Junction Permeability Is Mediated by TLR4 Signal Transduction Pathway Activation of FAK and MyD88. J Immunol 195:4999-5010
Guo, Shuhong; Al-Sadi, Rana; Said, Hamid M et al. (2013) Lipopolysaccharide causes an increase in intestinal tight junction permeability in vitro and in vivo by inducing enterocyte membrane expression and localization of TLR-4 and CD14. Am J Pathol 182:375-87
Al-Sadi, Rana; Guo, Shuhong; Ye, Dongmei et al. (2013) Mechanism of IL-1β modulation of intestinal epithelial barrier involves p38 kinase and activating transcription factor-2 activation. J Immunol 190:6596-606
Al-Sadi, Rana; Guo, Shuhong; Ye, Dongmei et al. (2013) TNF-α modulation of intestinal epithelial tight junction barrier is regulated by ERK1/2 activation of Elk-1. Am J Pathol 183:1871-84
Al-Sadi, Rana; Ye, Dongmei; Said, Hamid M et al. (2011) Cellular and molecular mechanism of interleukin-1β modulation of Caco-2 intestinal epithelial tight junction barrier. J Cell Mol Med 15:970-82
Ye, Dongmei; Guo, Shuhong; Al-Sadi, Rana et al. (2011) MicroRNA regulation of intestinal epithelial tight junction permeability. Gastroenterology 141:1323-33
Al-Sadi, Rana; Khatib, Khaldun; Guo, Shuhong et al. (2011) Occludin regulates macromolecule flux across the intestinal epithelial tight junction barrier. Am J Physiol Gastrointest Liver Physiol 300:G1054-64

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