Role of CD98 Glycoprotein in Intestinal Inflammation

CD98 糖蛋白在肠道炎症中的作用

• 批准号: 8424676
• 负责人: Hamed Laroui
• 金额: $ 10.01万
• 依托单位: GEORGIA STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-09-24 至 2017-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8424676
• 关键词: Adhesions; Advisory Committees; Alginates; Amino Acids; Animals; Anti-Inflammatory Agents; Anti-inflammatory; Antibiotics; Ants; Award; Bacteria; Bacterial Translocation; Binding; Biochemical; Biology; C-terminal; Cell Communication; Cell surface; Cells; Chemicals; Chitosan; Chronic; Colitis; Colon; Data; Development Plans; Digestive System Disorders; Disease; Down-Regulation; Encapsulated; Epithelial; Epithelial Cells; Event; Extracellular Domain; Functional disorder; Glycoproteins; Human; Hydrogels; Immune; Immunology; In Vitro; Inflammation; Inflammation Mediators; Inflammatory; Inflammatory Bowel Diseases; Inflammatory disease of the intestine; Interleukin-1; Interleukin-10; Intestines; Mentored Research Scientist Development Award; Mentors; Mentorship; Microbiology; Molecular; Molecular Biology; Mus; Nanotechnology; Neurofibromin 2; Non-Viral Vector; Pathogenesis; Permeability; Phosphorylation; Physiological; Play; Production; Research; Research Support; Resistance; Role; Serine Phosphorylation Site; Signal Transduction; Site; Small Interfering RNA; TNF gene; Tail; Techniques; Technology; Therapeutic; Training; Transgenic Mice; Universities; base; career development; computerized data processing; cytokine; extracellular; in vivo; interest; intestinal epithelium; nanoparticle; overexpression; professor; research and development; research study; symposium; therapeutic development; villin

DESCRIPTION (provided by applicant): 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. PUBLIC HEALTH RELEVANCE: 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.

描述（由申请人提供）：CD98在协调肠上皮事件中起重要作用，如粘附/极性，氨基酸运输和与细胞表面分子的直接结合。CD98独特的分子取向在细胞外c端尾部有一个pdz结合域，这表明细胞外信号在其多种功能中起作用。例如，CD98的外磷酸化增强了异型细胞-细胞相互作用，而拥有丝氨酸磷酸化位点的细胞外结构域对这种作用至关重要。CD98在炎症性炎症条件下表达增加，如炎症性肠病（IBD）。早期的研究支持肠上皮细胞特异性CD98诱导屏障功能障碍和刺激促炎介质产生的能力。本课题组的研究共同表明，CD98表达在人类肠道疾病（IBD）的发病机制中起作用。我们的总体假设是结肠CD98在引发和延续肠结肠炎中起关键作用。本提案的最初目的是研究CD98/细菌相互作用在肠屏障功能丧失中的作用。我们将进一步关注CD98在自发性结肠炎中的作用。随后将使用靶向纳米技术方法评估降低炎症结肠中CD98表达的效果。该项目将涉及各种生物化学、化学、纳米技术、分子、体外和体内方法。设想所提出的实验将有助于确定CD98在肠道炎症中的功能作用的分子机制，并允许开发针对肠道炎症疾病（包括IBD）的治疗策略。我的第一个目标是研究肠上皮CD98和CD98/细菌的相互作用是否参与肠屏障功能的丧失。我们小组获得的初步数据显示，结肠CD98的过表达增加了肠道通透性、细菌易位和促炎细胞因子如IL-1¿和TNF¿的表达。我们最初的目的是研究细菌是否在特异性过表达结肠CD98的小鼠中启动肠道屏障功能。具体来说，我们将用抗生素治疗Villin-CD98+/+转基因小鼠，并评估这些动物的肠道屏障功能和细胞因子表达与未治疗的Villin-CD98+/+转基因小鼠的关系。其次，CD98与细菌及其信号传导过程之间的直接相互作用将通过体外和体内方法进行研究。这些实验将阐明CD98表达和CD98/细菌相互作用在肠屏障功能丧失中的顺序作用。因此，我的第二个目标是研究结肠CD98表达对结肠炎的调节作用。我们已经证明，与野生型动物相比，dss诱导的结肠炎在绒毛蛋白CD98转基因小鼠（CD98在结肠细胞中表达）中加重。相比之下，与结肠CD98表达（CD98fl/+）较高的小鼠相比，结肠CD98表达（CD98fl/+）较低的小鼠对DSS的抗性更强。我们建议通过评估IL-10-/-/CD98fl/+Villin-Cre， IL-10-/-/Villin-CD98+/+和IL-10-/-/CD98fl/+动物的自发炎症来确定CD98表达在慢性结肠炎中的作用。最后，我的第三个目标是研究使用包裹在纳米颗粒（NPs）中的非病毒载体作为载体有效抑制诱导结肠炎的靶向递送抗cd98 siRNA。最近，我们描述了一种针对结肠的原始技术，该技术将抗炎分子装载在NPs中并包裹在海藻酸-壳聚糖水凝胶中。利用这项技术，我们获得了初步数据，显示TNF - sirna负载的NPs抑制肠道炎症。我们进一步提出在诱导结肠炎小鼠中特异性靶向并将CD98 siRNA递送到过度表达CD98的炎症结肠（上皮和免疫）细胞。