The G1 domain of cartilage proteoglycan (PG) aggrecan is responsible for the binding to hyaluronan (HA), and this macromolecular interaction is stabilized with cartilage link protein (LP). This interaction (in normal cartilage) forms multimillion-Dalton-size aggregates entrapped within the type II collagen meshwork, and the glycosaminoglycan (GAG)-substituted regions of the aggrecan core protein retain water responsible for the compression-resisting resilience of cartilage. Therefore, our working hypothesis is that the in vivo constitutive (over)expression of recombinant HA-binding molecules (such as cartilage LP and G1 domain of aggrecan) should disrupt the natural in vivo homeostasis of cartilage, as the recombinant molecules compete with the endogenous (wild-type) molecules for HA-binding sites. We have generated transgenic mice overexpressing either LP or G1 domain (hG1) of human aggrecan in cartilage. We selected high and low transgene expression lines of each, and backcrossed them into BALB/c background. Here, we propose to test our hypothesis that joint/cartilage abnormalities might be due to the consequence of the slow, but continuous accumulation of the G1 domain and LP in aging mice, and these HA-binding structures may competitively inhibit desired repair processes in cartilage by occupying the binding sites on the HA backbone from newly synthesized aggrecan and LP. A similar process may occur in humans, which is slow in normal aging tissue, but it is """"""""accelerated"""""""" in osteoarthritic cartilage. The two transgenic models (hG1-Tg and LP-Tg), although generated for different purposes, may be useful models to test this hypothesis. We will simultaneously perform morphological, biochemical (transcriptional and translational) and biomechanical studies of knee and hip joints of these transgenic animals. We will identify the consequence of the constitutive (over)expression of individual (hG1 or LP) transgene, and the combination (co-expression) of both HA-binding proteins during the aging processes of transgenic animals. The ultimate goal of this proposal is to gather sufficient information on the in vivo role of HA-binding proteins and create a solid basis for subsequent transgenic and knockout approaches for studying osteoarthritic events in cartilage. Cartilage proteoglycan (PG) aggrecan binds to hyaluronan (HA), and this aggrecan-HA interaction is stabilized by a third component called """"""""link protein"""""""" (LP). The major physiological function of aggrecan is to immobilize water, via its glycosaminoglycan (GAG) side chains, providing resilience for the weight-bearing articular cartilage. Although the differences between normal aging and osteoarthritic tissues are significant, there are also similarities in (patho)physiological mechanisms. For example, degradation products are either lost (GAG-binding domains) or accumulate (HA-binding domain) in both aging and osteoarthritic cartilages, which results in cartilage degradation and loss of function. Although resident chondrocytes attempt to repair cartilage damage, the newly synthesized aggrecan is also lost as virtually no binding site is available on the G1/LP-saturated HA backbone. To date, no systemic study has been performed for measuring simultaneously the expression and synthesis of HA, aggrecan and LP. We propose to investigate the in vivo consequence of manipulating aggrecan, LP and HA-binding interactions through transgenic approaches. We have generated two different transgenic colonies constitutively (over)expressing either the G1 domain of human aggrecan, and LP. We propose to study """"""""accelerated aging"""""""" in cartilage, which may lead to early OA-like abnormalities. We propose to achieve this goal by in vivo overexpression of the aggrecan G1 domain (the HA-binding region) and LP in transgenic mice. These transgene products should compete for the binding sites on HA with naturally synthesized aggrecan and LP. Transgenic animals overexpressing G1 domain and LP will be intercrossed to create double transgenic mice, possibly simulating various degrees of aging and/or osteoarthritic processes. ? ? ?
|Ghassemi-Nejad, S; Kobezda, T; Rauch, T A et al. (2011) Osteoarthritis-like damage of cartilage in the temporomandibular joints in mice with autoimmune inflammatory arthritis. Osteoarthritis Cartilage 19:458-65|