Electrostatic forces and M cell uptake at mucosal surfaces

粘膜表面的静电力和 M 细胞摄取

• 批准号: 8268776
• 负责人: DAVID D LO
• 金额: $ 22.8万
• 依托单位: UNIVERSITY OF CALIFORNIA RIVERSIDE
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-02-09 至 2014-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8268776
• 关键词: Adhesions; Affect; Animal Model; Anthrax disease; Apical; Bacteria; Binding; Binding Proteins; Biological Process; Brush Border; Buffers; Cell Culture Techniques; Cell membrane; Cell model; Cell surface; Cells; Characteristics; Charge; Complement; Defect; Electrostatics; Employee Strikes; Enterocytes; Epithelial; Epithelial Cells; Epithelium; Gastric Emptying; HIV; Immune; Immunity; Immunologic Receptors; Immunologic Surveillance; In Vitro; Intestines; Ionic Strengths; Liquid substance; Lymphoid Tissue; M cell; Measures; Mediating; Microbe; Modeling; Mucous body substance; Osmolar Concentration; Pathogenesis; Peptides; Peristalsis; Property; Salmonella; Small Intestines; Stream; Surface; Suspension substance; Suspensions; System; Testing; Tissues; Villus; Virulence Factors; Virus; Water; Wit; Work; apical membrane; cellular microvillus; chemical property; density; design; in vivo; intestinal epithelium; microbial; monolayer; mouse model; mucosal uptake; mucosal vaccine; particle; pathogen; receptor; uptake; vaccine delivery; zeta potential

DESCRIPTION (provided by applicant): The mucosal epithelium is a primary barrier against invasive microbes, so understanding the physicochemical forces involved in mucosal immune surveillance may be critical to understanding pathogenesis versus immunity. Mucosal M cells overlying lymphoid tissues are central to immune surveillance, using mechanisms for active uptake of mucosal pathogens. Ironically, many invasive pathogens ranging from Salmonella to Anthrax are also adept at using M cells for entry across the epithelial barrier. How is this uptake mediated? Recently, we showed that low ionic strength in the liquid of the airway lumen significantly enhanced particle uptake by M cells. Low ionic strength should increase long-range electrostatic interactions and increase the repulsion of negatively charged particles from negatively charged epithelium; curiously, we found a paradoxically increased uptake at M cell apical membranes in vivo. This finding has given rise to our working hypothesis: in contrast to neighboring mucosal epithelial cells, the M cell is electrostatically optimized for interaction wit microparticles in suspension as an emergent property of its smooth apical membrane. The differences in surface charge establishes an electrostatic field at the boundary between enterocytes and M cells that helps draw negatively charged particles to the M cell apical membrane. We propose two aims to test this hypothesis: (1) We will use cell culture models and a modified tangential flow chamber to model the binding of particles at the apical surface of the epithelial cells with different surface charge characteristics. We will compare fluorescent microbes and PLGA microparticles produced with different surface zeta potentials to assess binding to enterocyte versus "M cell-like" smooth apical membranes. (2) Using mouse models of M cell deficiency, we will study the effect of particle surface charge on uptake in the intestine, and the effect of electrostatic forces on M cell- dependent versus -independent particle uptake to immune tissues. An understanding of the effect of these electrostatic forces in the intestine will be important in defining the mechanisms of M cell mucosal immune surveillance. In addition, these forces would be expected to be important in the pathogenesis of invasive microbes such as Salmonella. Finally, our studies will help define requirements for designing mucosal vaccine delivery systems, whether designed for M cell targeted uptake or for nonspecific adhesion and M cell-independent uptake. PUBLIC HEALTH RELEVANCE: Electrostatic forces and M cell uptake at mucosal surfaces Narrative: The mucosal epithelium is a primary barrier against invasive microbes, so understanding the physicochemical forces involved in mucosal immune surveillance may be critical to understanding pathogenesis versus immunity. Mucosal M cells overlying lymphoid tissues are central to immune surveillance, using mechanisms for active uptake of mucosal pathogens. Ironically, many invasive pathogens ranging from Salmonella to Anthrax are also adept at using M cells for entry across the epithelial barrier. Recently, we discovered evidence that charge interactions between particles such as bacteria and the intestinal epithelium may be a major factor in M cell uptake. The charge interactions between bacteria and intestinal epithelium may be modulated by the surface features of M cells, which are smoother than the neighboring intestinal epithelium. That is, the local differences between M cells and intestinal epithelium may set up charge effects that drive particles to the M cell membrane. If true, this would be a striking and surprising example of how biological processes of cellular differentiation can modify local physico-chemical properties. These effects appear to be designed for optimal interactions with particles with charge properties specific to viruses and bacteria, and would appear to complement the array of innate immune receptors for detecting pathogens. We propose to test these charge effects using well-defined cell culture models to mimic M cell charge properties, and animal models with specific M cell defects. An understanding of the effect of these forces in the intestine will be important in defining the mechanisms of M cell mucosal immune surveillance. In addition, these forces would be expected to be important in the pathogenesis of invasive microbes such as Salmonella. Finally, our studies will help define requirements for designing mucosal vaccine delivery systems, whether designed for M cell targeted uptake or for nonspecific adhesion and M cell-independent uptake.

描述（由申请人提供）：粘膜上皮是侵入性微生物的主要障碍，因此了解粘膜免疫监测涉及的物理化学力对于理解发病机理与免疫力可能至关重要。粘膜M细胞上覆盖淋巴组织的粘膜细胞是免疫监测的核心，使用了主动摄取粘膜病原体的机制。具有讽刺意味的是，许多从沙门氏菌到炭疽的侵入性病原体也擅长于使用M细胞进入上皮屏障。这种吸收如何 介导？最近，我们表明气道管腔液体中的低离子强度显着增强了M细胞的颗粒摄取。低离子强度应增加远程静电相互作用，并增加带负电荷上皮的负电荷颗粒的排斥；奇怪的是，我们发现体内M细胞顶膜的矛盾摄取增加。这一发现引起了我们的工作假设：与邻近的粘膜上皮细胞相反，M细胞是静电优化的，用于悬浮液中的微粒相互作用，作为其光滑顶端膜的新兴特性。表面电荷的差异在肠细胞和M细胞之间的边界上建立了一个静电场，有助于将带负电的颗粒吸引到M细胞顶膜上。我们提出了两个旨在检验这一假设的目的：（1）我们将使用细胞培养模型和修改的切向流动室来对具有不同表面电荷特征的上皮细胞顶部表面的颗粒结合进行建模。我们将比较具有不同表面Zeta电位的荧光微生物和PLGA微粒，以评估与肠球菌与“ M细胞样”平滑顶膜的结合。 （2）使用M细胞缺乏的小鼠模型，我们将研究颗粒表面电荷对肠道摄取的影响，以及静电力对M细胞依赖性与非依赖性颗粒对免疫组织的影响。对这些静电力在肠道中的影响的理解对于定义M细胞粘膜免疫监测的机制至关重要。此外，这些力在侵入性微生物（如沙门氏菌）的发病机理中有望很重要。最后，我们的研究将有助于定义设计粘膜疫苗输送系统的要求，无论是针对M细胞靶向摄取还是针对非特异性粘附和M细胞无关的摄取的设计。 公共卫生相关性：粘膜表面的静电力和细胞摄取叙事：粘膜上皮是针对侵入性微生物的主要障碍，因此了解粘膜免疫监测涉及的物理化学力对于理解病原体与免疫可能至关重要。粘膜M细胞上覆盖淋巴组织的粘膜细胞是免疫监测的核心，使用了主动摄取粘膜病原体的机制。具有讽刺意味的是，许多从沙门氏菌到炭疽的侵入性病原体也擅长于使用M细胞进入上皮屏障。最近，我们发现了证据表明，细菌和肠上皮等颗粒之间的电荷相互作用可能是M细胞摄取的主要因素。细菌与肠上皮之间的电荷相互作用可以由M细胞的表面特征调节，M细胞的表面特征比相邻的肠上皮更顺畅。也就是说，M细胞和肠上皮之间的局部差异可能会建立驱动颗粒到M细胞膜的电荷效应。如果是真的，这将是一个惊人而令人惊讶的例子，说明细胞分化的生物学过程如何改变局部的物理化学特性。这些效果似乎是设计用于与特定于病毒和细菌的电荷特性的颗粒的最佳相互作用，并且似乎可以补充用于检测病原体的先天免疫受体阵列。我们建议使用定义明确的细胞培养模型来测试这些电荷效应，以模仿细胞电荷的特性，以及具有特定M细胞缺损的动物模型。 对这些力在肠道中的影响的理解对于定义M细胞粘膜免疫监测的机制至关重要。此外，这些力在侵入性微生物（如沙门氏菌）的发病机理中有望很重要。最后，我们的研究将有助于定义设计粘膜疫苗输送系统的要求，无论是针对M细胞靶向摄取还是针对非特异性粘附和M细胞无关的摄取的设计。