Joint degeneration: Somatic mosaic analysis in a transgenic mouse

描述(申请人提供)：骨关节炎(OA)是最常见的关节炎形式，也是老年人活动受限和身体残疾的主要原因：65岁以上的人中有80%-90%患有骨关节炎。目前治疗骨性关节炎的方法大多是姑息性的，特异性有限。事实上，最近发现的一些口服COX-2选择性抑制剂(如万络、BExtra)的严重副作用强调了开发新的抗炎疗法治疗OA的必要性。本研究的目的是描述有条件地诱导成年小鼠关节内低水平IL-1β表达后的退行性关节疾病的小鼠模型，并探索基于siRNA的抗炎治疗在治疗颞下颌关节骨关节炎中的应用。为此，我们将在我们实验室最近建立的col1-IL1β-XAT转基因小鼠模型中应用体细胞嵌合体分析，该模型利用基于Cre/loxP的分子遗传学方法在时间和空间上激活含有休眠的IL-1β转录单位的种系传递的重组底物。通过组织病理学、免疫组织化学和行为学研究，初步数据表明，经关节内注射Cre后，col1-IL1β-XAT转基因小鼠的膝关节和TMJ发生了类似骨关节炎的病理改变。为了进一步开发和鉴定这一模型，提出了以下具体目标：(1)通过体细胞镶嵌分析，从关节病理和口面部疼痛的角度表征COL1-IL1beta-XAT转基因小鼠TMJ关节炎的发展；(2)评价关节内抗mPGES-1和抗COX-2 siRNA治疗对COL1-IL1beta-XAT转基因小鼠TMJ的有效性。我们最近开发了能够减弱COX-2以及环氧合酶-前列腺素途径的其他成员的小抑制RNA(SiRNA)，包括COX-1、mPGES-1和cPGES。我们建议使用猫科动物免疫缺陷病毒平台FIV(SiRNA)在关节内表达这些siRNA结构。拟议的研究将有助于评估新的治疗靶点(即mPGES-1)，并有助于开发新的治疗工具(SiRNA)。我们的提案是针对NIDCR项目公告PA-04-139《联合变性：小鼠模型》提出的，该项目征集采用基因定义和转基因小鼠模型的研究提案，以探索OA和siRNA技术等新疗法的生物学机制。为此，我们召集了罗切斯特大学的一个多学科专家小组，他们过去曾成功合作并发表过论文，他们将为完成这一项目提供必要的专业知识。

DESCRIPTION (provided by applicant): Osteoarthritis (OA) is the most common form of arthritis and a major cause of activity limitation and physical disability in the elderly: 80%-90% of humans over the age of 65 suffer from OA. Available treatments for OA are mostly palliative with limited specificity. In fact, the recently identified serious side effects of some intra- oral COX-2 selective inhibitors (i.e. Vioxx, Bextra) underscore the need for the development of novel anti- inflammatory therapies for OA. The purpose of this research proposal is to characterize a mouse model of degenerative joint disease following the conditional induction of intra-articular, low level IL-1beta expression in the adult mouse and to explore the utility of siRNA-based anti-inflammatory therapy in the management of temporomandibular joint (TMJ) osteoarthritis. To this end, we will employ somatic mosaic analysis in the Col1-IL1beta-XAT transgenic mouse model recently developed in our laboratory, which utilizes a Cre/loxP based molecular genetic method to temporally and spatially activate a germline-transmitted recombinational substrate containing a dormant IL-1beta transcription unit. Preliminary data demonstrate the development of OA-like pathology in the knees and TMJ of Col1-IL1beta-XAT transgenic mice after intra-articular Cre administration as assessed by histopathological, immunohistochemical and behavioral studies. The following specific aims are proposed to further develop and characterize this model: (1) Characterize the development of TMJ arthritis following somatic mosaic analysis in the Col1- IL1beta-XAT transgenic mouse in terms of joint pathology and orofacial pain; (2) evaluate the effectiveness of intra-articular anti-mPGES-1 and anti-COX-2 siRNA therapy versus available pharmacologic inhibitors in the TMJ of Col1-IL1beta-XAT transgenic mice. We have recently developed small inhibitory RNA (siRNA) constructs capable of attenuating COX-2, as well as other members of the cyclooxygenase-prostaglandin pathway, including COX-1, mPGES-1 and cPGES. We propose to express these siRNA constructs intra-articulalry using a feline immuno-deficiency viral platform, FIV(siRNA). The proposed studies will contribute in the evaluation of novel therapeutic targets (i.e. mPGES- 1) as well as contribute to the development of new therapeutic tools (siRNA). Our proposal is submitted in response to the NIDCR program announcement PA-04-139 entitled "Joint degeneration: Mouse models", which solicits research proposals employing genetically defined and genetically modified mouse models to explore the biological mechanisms underlying OA and novel therapeutics such as siRNA technology. To this end, we have brought together a multidisciplinary team of experts from the University of Rochester that have successfully collaborated and published in the past and who will contribute the necessary expertise for the completion of this project.