Review: Collagen markers in early arthritic diseases

Abstract

In arthritic diseases e.g. osteoarthritis (OA) and rheumatoid arthritis (RA), the stability of the collagen type II (CII) fibers, a major component of articular cartilage, is compromised with extensive proteolytic breakdown leading to cartilage erosion and joint deterioration. A clinical need for molecular markers that give instantaneous measure of rate of joint deterioration has developed, as other measurements e.g. arthroscopy, and joint space narrowing are insensitive to small changes in disease status over short periods of time. Owing to its exclusive presence in cartilaginous tissues, markers of CII synthesis and degradation have been extensively studied. Assays that measure these markers in biological fluids e.g. synovial fluid (SF), serum, and urine have been developed and applied to detect early disease onset, monitor disease progression, and response to anti-arthritic drugs. CII synthesis markers include the procollagen type II C-propeptide (PIICP) and the procollagen type IIA N-propeptide (PIIANP). CII degradation markers include CII C-telopeptide (CII-X), CII neoepitope (TIINE), helix II, C2C, CNBr 9.7, Coll 2-1, and Coll 2-1 NO₂. Most of these markers differentiate between early stages of OA, RA and reference controls. The best correlations with structural changes occur when measurements are made in SF while serum measurement frequently did not correlate with structural changes. Although the selection of an optimal marker or a set of markers is still problematic, few markers are of considerable utility in early detection and monitoring of arthritic diseases. The current challenge is to improve the discriminatory power of these markers so they can be used to guide therapeutic decisions.

Introduction

The two main components of articular cartilage are fibril-forming collagen type II (CII) and the cartilage specific, large proteoglycan, aggrecan [1]. CII is found exclusively in cartilagenous tissues where it constitutes over 60% of cartilage's dry weight [2]. Most of the physiological properties of cartilage are dependent on an intact CII network. These characteristics suggest that monitoring CII metabolism might provide a method for assessing the health of articular cartilage.

CII is a triple helical protein composed of three identical alpha chains. Following their synthesis, procollagen alpha chains are modified intracellularly via hydroxylation of proline and lysine residues by prolyl and lysyl hydroxylases, O-glycosylation of certain hydroxylysine residues, chain association and inter-chain disulfide bond formation to form the triple helical structure. Following secretion, the procollagen triple helix is further modified by removal of the N- and C-terminal propeptides by specific proteases, ordered arrangement of the mature fibrils and cross-linking. The helices are intra- and inter-molecularly cross-linked predominantly by hydroxylysyl pyridinoline cross-links [3], [4], a process mediated by the activity of lysyl oxidase enzyme. Two minor collagens, type IX (CIX) and type XI (CXI), join with CII in forming a heterofibril. CXI is composed of three different alpha chains and is believed to regulate fibril size. CIX [5], [6] is a FACIT collagen (fibril associated collagen with interrupted triple helices) with covalently linked proteoglycans. It constitutes around 1% of the collagen fibril and decorates its surface [7], [8].

In arthritic diseases, the stability of collagen fibrils is compromised with extensive proteolytic breakdown of CII fibrils leading to cartilage erosion and joint deterioration. Initial cleavage of CII is attributed to the collagenase sub-family of matrix metalloproteinases (MMPs) e.g. collagenase 1, 2, 3, and MT1-MMP also known as MMP-1, 8, 13, and 14, respectively [9], [10]. These collagenases preferentially cleave CII between Gly⁷⁹⁴ and Leu⁷⁹⁵ generating two fragments that are 3/4 and 1/4 the size of the collagen precursor [11]. Following initial cleavage, the triple helix of CII fragments unwind, providing a denatured substrate susceptible to further degradation by a variety of proteolytic enzymes.

Arthritic diseases are characterized by compromise of articular cartilage leading to loss of its physical properties accompanied by joint pain and loss of mobility for the patient. On gross examination, the smooth glistening articular cartilage surface becomes fibrillated progressing on to fissuring, ulceration and irreversible loss of full-thickness articular cartilage. Osteoarthritis (OA) has multiple etiologies including joint trauma [12] and genetic mutations in fibrillar collagen genes [13]. Rheumatoid arthritis (RA) is an autoimmune disorder characterized by persistent joint inflammation, polymorphonuclear cell infiltration, and synovial hyperplasia resulting in cartilage loss and joint deformity [14], [15].

The severity and progression of OA have been evaluated using magnetic resonance imaging (MRI), arthroscopy and radiographic measurement of joint space narrowing (JSN). All three techniques require a baseline measurement and a subsequent measurement after a 6 month to two year period in order to determine the rate of cartilage loss. Thus, there is a great clinical need for molecular markers which can give an instantaneous measure of the rate of joint deterioration.

Arrays of molecular markers, that reflect metabolism of cartilage and synovial tissues in OA and RA, have been investigated and validated [16], [17], [18], [19], [20], [21]. Among the most promising are markers of CII metabolism in OA and RA. Owing to its extensive and exclusive presence in cartilage, markers of CII metabolism in synovial fluid (SF), serum, and urine can reflect joint status marking the onset, and progression of OA and RA. Since alteration in CII metabolism occurs prior to detectable radiographic changes, it is a specific and early marker of arthritic joint diseases.

Cartilage components such as the telopeptides and breakdown fragments of CII diffuse into the SF. Once in the SF, they can undergo further proteolytic processing and can be cleared by the lymphatics. Some processing, particularly of carbohydrates, can occur in the lymph nodes, before entering the blood. There, further processing may occur in the liver and kidney before elimination in the urine.

The SF measurements of CII markers more directly reflect CII turnover in cartilage than either serum or urine measurements. However, SF aspiration from arthritic joints is a non-routine invasive process so routine measurements have been focused on serum and urine. To measure CII metabolism markers in arthritic diseases, immunoassays based on monoclonal or polyclonal antibodies against specific CII sequences (Fig. 1) were used to quantitate CII fragments to infer the degree of CII synthesis and degradation compared to a control population. In this review, we will focus on CII assays and their application in early detection of OA and RA.