CD98 plays an important role in coordinating intestinal epithelial events, such as adhesion/polarity, amino acid transport, and direct binding o cell surface molecules. The unique molecular orientation of CD98 with a PDZ-binding domain in the extracellular C-terminal tail suggests that extracellular signaling plays a role in its multipl functions. For example, ecto-phosphorylation of CD98 enhances heterotypic cell-cell interactions, and the extracellular domain possessing serine phosphorylation sites is crucial for this effect. CD98 expression is increased in inflammatory inflammation conditions, such as inflammatory bowel disease (IBD). Earlier research supports the ability of intestinal epithelial cell-specific CD98 to induce barrier dysfunction and stimulate the production of pro-inflammatory mediators. Studies by our group have collectively shown that CD98 expression plays a role in the pathogenesis of intestinal bowel disease (IBD) in humans. Our overall hypothesis is that colonic CD98 plays a critical role in initiating and perpetuating intestinal coltis. The initial aim of this proposal is to investigate the role of CD98/bacteria interactions in the los of intestinal barrier function. We will additionally focus on establishing the role of CD98 in spontaneous colitis. The efficacy of knockdown of CD98 expression in inflamed colon will subsequently be assessed using a targeted nanotechnology approach. The project will involve a variety of biochemical, chemical, nanotechnological, molecular, in vitro, and in vivo approaches. It is envisaged that the proposed experiments will facilitate identification of the molecular mechanisms underlying the functional role of CD98 in intestinal inflammation and allow the development of therapeutic strategies targeting intestinal inflammatory conditions, including IBD.
My first aim will focus on investigating if intestinal epithelial CD98 and CD98/bacteria interactions participate in loss of intestinal barrier function. Preliminary data obtained by our group showed that overexpression of colonic CD98 increases intestinal permeability, bacterial translocation and expression of pro-inflammatory cytokines, such as IL-1? and TNF?. Our initial aim is to investigate whether bacteria initiate intestinal barrier function in mice that specificaly overexpress colonic CD98. Specifically, Villin-CD98+/+ transgenic mice will be treated with antibiotic, and the intestinal barrier function and cytokine expression in these animals assessed in relation to untreated Villin-CD98+/+ transgenic mice. Secondly, direct interactions between CD98 and bacteria and their signaling processes will be investigated via ex and in vivo approaches. These experiments should clarify the sequential role of CD98 expression and CD98/bacteria interactions in loss of intestinal barrier function. Thus, my second aim will study the colonic CD98 expression effect on the modulation of colitis. We have demonstrated that DSS-induced colitis is aggravated in villin-CD98 transgenic mice (in which CD98 is expressed in colonocytes) than wild-type animals. In contrast, mice with low colonic CD98 expression (CD98fl/+Villin-Cre) are more resistant to DSS, compared to those with higher colonic CD98 expression (CD98fl/+). We propose to establish the role of CD98 expression in chronic colitis by assessing spontaneous inflammation in IL-10-/-/CD98fl/+Villin-Cre, IL-10-/-/Villin-CD98+/+ and IL-10-/-/CD98fl/+ animals. Finally, my third aim will investigate the site-directed delivery of ant-CD98 siRNA using non-viral vectors encapsulated in nanoparticles (NPs) as carriers effectively inhibits induced colitis. Recently, we described an original technique targeting the colon with anti-inflammatory molecules loaded in NPs and encapsulated in an alginate-chitosan hydrogel. Using this technology, we obtained preliminary data showing that TNF? siRNA-loaded NPs suppress intestinal inflammation. We further propose to specifically target and deliver CD98 siRNA to inflamed colonic (epithelial and immune) cells that overexpress CD98 in mice with induced colitis.
This is a Mentored Research Scientist Development Award application for Dr. Hamed Laroui, a biochemist, nanotechnologist, and physical chemist by training with specialization in Digestive Diseases. He has initiated an independent study from his mentor on the physiologic and biologic role of CD98, a glycoprotein that is a heterodimer in intestinal epithelium. He will be using this award opportunity to differentiate himself from his mentor by shifting his research interest from basic biology to potential therapeutic treatment based on CD98. His main hypothesis is that temporary downregulation of CD98 in the intestinal epithelium under pathological conditions modulates inflammation. As a new Assistant Professor at Georgia State University, Dr. Laroui submits this five-year career development plan under the mentorship of Dr. Merlin. The award will be used to i) mechanistically develop his initial observations showing that downregulation of CD98 in the intestinal epithelium under pathological conditions modulates inflammation; ii) receive hands-on training in molecular biology, microbiology, and immunology; iii) develop the therapeutic strategy based on CD98 targeting by nanotechnology; iv) attend and present at academic seminars and conferences (e.g. DDW and FASEB), and receive guidance from a selected Career Development Research Advisory Committee.
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