There is significant literature that IL1RN polymorphisms affect clinical outcomes in a variety of diseases, including osteoarthritis, diabetes, juvenile arthritis and obesity. The precise mechanisms that impart IL1RN's effects are unknown. We have shown that the IL1RN TTG haplotype (rs419598, rs315952 and rs9005) predicts both age-related radiographic severity [Kellgren-Lawrence grade (KL), medial joint-space width (JSW)] and is associated with a 3-fold increased risk for incident knee OA. Moreover, this pro-inflammatory genotype is not limited to OA but can be demonstrated in patients with rheumatoid arthritis (RA) and in obese patients who carry the IL1RN TTG risk haplotype. There are a number of functional elements in the IL1RN gene, and it is unclear which of these confers increased inflammatory function by modulating IL1RN expression and influencing other genes in the haplotype block. The central objective of this grant, therefore, will be to define the effects of the IL1RN risk haplotypes in vitro and in vivo using transgenic technologies.
In Aim 1 : Use synthetic human haplotypes to determine which functional elements are critical for the inflammatory phenotype conferred by the IL1RN risk haplotype, in vitro. We will construct risk and protective haplotypes in the context of the flanking human genes using ?Assemblon? technology, which allows the construction and precision delivery of mammalian gene loci of up to 200 kb in length. Using this technology, we will select informative mouse embryonic stem cell (mESC) lines and will be differentiated in vitro to monocytes and macrophages which will be assayed for expression of IL1RN and the other genes in the haplotype block, cytokine production, surface marker expressions.
Aim 2 : Generate a humanized mouse expressing the IL1RN TTG-2 risk or TTG-0 non-risk haplotypes for study in the surgically induced destabilization of the medial meniscus (DMM) model of OA. Since recent studies also indicate an association of IL1RN genetic polymorphism in obesity and insulin resistance, we will study high-fat diet-induced obesity models in transgenic humanized mice. These studies will elucidate novel mechanisms by which the IL1RN risk haplotype influences cellular responses and inflammatory gene regulation, and will reveal previously unrecognized various functional linked genes, which are crucial for the inflammatory function of the risk haplotype.
Genetic differences in interleukin genes between populations of patients with knee osteoarthritis (OA) and other disease affect disease risk and severity. The grant will focus on the elucidation of specific gene regulatory networks differentially modulated by each risk gene, enabling a deeper mechanistic understanding of disease pathogenesis. By elucidating the molecular events and understanding the role of genes that regulate inflammation, the studies can help identify targets for disease prevention or treatment.