Molecular Comparison of Macrophage Foreign Body Response

描述(由申请人提供)： 目前使用生物材料炎症和异物检测所需的人或动物来源的吞噬细胞来源是不方便的、昂贵的，而且可能没有必要。次级巨噬细胞系细胞系在这些体外试验中的等价性在很大程度上是未经证实的，尽管存在许多关于它们在生物材料毒性或炎症试验中的效用的未发表的轶事。这项建议的总体目标是，利用现代分子和细胞生物学定量方法，确定某些巨噬细胞来源的细胞系培养在准确复制人和动物来源的吞噬细胞表型行为方面的有效性，这些细胞通常用于模拟生物材料植入产生的异物反应的各个方面。 我们的工作假设是，在旨在监测生物材料上细胞介导性炎症反应的培养中，使用选定的次级巨噬细胞衍生细胞系在几种模型生物材料上的表型反应和异物巨细胞形成将与使用宿主来源的初级巨噬细胞观察到的表型反应和异物巨细胞形成显著相似。研究寻求围绕巨噬细胞培养系统的适当使用和限制的重要、长期存在的问题的答案，以分析所有植入的生物材料普遍存在的异物反应。新的数据将与成熟的原代单核/巨噬细胞培养方法进行比较，这些方法目前普遍用于异物巨细胞生物材料的炎症评估。 提出了以下具体目标：(1)在几种受控表面化学的模型生物材料上，确定代表培养的次级巨噬细胞衍生细胞系Th1和Th2表型反应的选定细胞产生的细胞因子；(2)通过巨噬细胞衍生细胞培养中细胞因子产生的变化和作为表面化学函数的反应，确定培养中的短期和长期表型稳定性；(3)量化异物巨细胞形成的几个不同的已知表型特征，作为巨噬细胞衍生细胞系和原代巨噬细胞随着培养时间的表面化学变化的函数；(4)建立可靠、定量的基于微阵列的基因分析，以加速对巨噬细胞活化和炎症行为的特定已知标记物的分析，特别是细胞因子的表达和凋亡标记物；以及(5)利用体外培养的巨噬细胞和成纤维细胞的共培养系统，研究巨噬细胞衍生的细胞因子产生对成纤维细胞产生胶原和纤维前体的影响，这些胶原和纤维前体与体内常见的异物包裹反应有关。

DESCRIPTION (provided by applicant): Current use of human- or animal-derived phagocytic cell sources required for biomaterials inflammation and foreign body assay is inconvenient, costly, and potentially unnecessary. Equivalence of secondary macrophage-lineage cell lines in these in vitro assays is largely unproven, although many unpublished anecdotes exist regarding their utility in biomaterials toxicity or inflammatory assays. This proposal's overall objective is to establish, using quantitative modem molecular and cell biological methods, the validity of certain macrophage-derived cell line cultures in accurately duplicating phenotypic behavior of human- and animal-derived phagocytes commonly used to model aspects of the biomaterial implant-generated foreign body response. Our working hypothesis is that phenotypic response and foreign body giant cell formation on several model biomaterials using select secondary macrophage-derived cell lines in cultures designed to monitor cell-mediated inflammatory responses on biomaterials will be measurably similar to that observed using host-derived primary macrophages. Studies seek answers to important, long-standing questions surrounding the appropriate use and limitations of macrophage culture systems to analyze the foreign-body response ubiquitous for all implanted biomaterials. New data will be compared against well-developed primary monocyte/macrophage culture assays now common for biomaterials inflammatory assessments with foreign body giant cells. The following Specific Aims are proposed: (1) Determine selected cell-produced cytokines representing the Th1 and Th2 phenotypic responses in cultured secondary macrophage-derived cell lines on several model biomaterials of controlled surface chemistry; (2) Identify short and long term phenotypic stability in culture through changes in cytokine production and responses in macrophage derived cell cultures as a function of surface chemistry; (3) Quantify several distinct known phenotypic features of foreign body giant cell formation as a function of surface chemistry for macrophage-derived cell lines and primary macrophages over time in culture; (4) Establish reliable, quantitative microarray-based gene analysis to accelerate profiling of specific known markers of macrophage activation and inflammatory behavior, specifically for cytokine expression and apoptosis markers; and (5) Exploit an in vitro co culture system containing cultured macrophages and fibroblasts to study affects of macrophage-derived cytokine production on fibroblast production of collagen and fibrous precursors relevant to encapsulation reactions common to foreign bodies in vivo.