Introduction

Osteoarthritis in humans is most commonly a multifactorial degenerative joint disease. It is incurable, costly and responds poorly to treatment. The disease process of OA is characterized by the progressive erosion of articular cartilage, leading to joint space narrowing, subchondral sclerosis, subchondral cysts, synovial inflammation, and marginal osteophyte formation [1]. A number of drugs such as nonsteroidal anti-inflammatory drugs (NSAIDs), cyclooxygenase 2 inhibitors, and steroids for the prevention and treatment of OA act by inhibiting joints degradation. However, the effectiveness of OA prevention in clinical practice is limited. Therefore, there is a great interest in finding a novel potential biotherapeutic approaches for the treatment of OA.

Lubricin, a 227.5-kDa mucinous glycoprotein which was originally identified as a lubricating glycoprotein present in synovial fluid [2], is recognized to have an important role in preventing cartilage wear and synovial cell adhesion and proliferation now [3]. Lubricin had previously been shown to be a proteoglycan specifically synthesized and expressed by articular chondrocytes of the superficial zone [4], which was known to interact with to the articular surface and function as a boundary lubricant in articular joints and reduces the coefficient of friction of the articualr cartilage surface [5]. It is possible that normal accumulation of lubricin is impaired in OA, leading to a net loss of protein. The content of this protein produced by the superficial layer of cartilage is damaged with aging and OA, and the lubricin gene is differently expressed in the synovium of RA and OA implying a possible role in the pathogenesis of these disease [6]. The role of lubricin such as lubrication and cytoprotection have been scientific investigated. Lubricin plays an important role in articualr joint physiology, and the loss of accumulation of lubricin may have a role in the pathology of OA. Therefore, a recent study by Flannery et al. [7] demonstrated the recombinant lubricin played significant disease-modifying and chondroprotective effects during the progression of animal OA model, which suggested the potential use of recombinant lubricin molecules in novel biotherapeutic approaches for the treatment of OA and associated cartilage abnormalities. This review focuses on the lubricin, emphasizing its role in joint disease and its currently in use, and potential future strategies.

Lubricin

Lubricin was originally identified as a lubricating glycoprotein present in synovial fluid [2], which was encoded by the Prg4 gene [8] and it is recognized to have a major protective role in preventing cartilage wear and synovial cell adhesion and proliferation recently [3]. Lubricin is also known as proteoglycan 4 (PRG4) or superficial zone protein (SZP), which was originally isolated and purified from the culture media of explants derived from the superficial zone of bovine articular cartilage [9]. In subsequent studies, lubricin and a related homologue, megakaryocyte-stimulating factor precursor protein (MSF), first detected in the urine of patients with acute thrombocytopenia undergoing bone marrow transplantation, was cloned and its primary amino acid sequence determined [10]. Besides, lubricin is also identified as an analogue with hemangiopoietin (HAPO) [11]. This protein has a molecular mass of approximately 227.5-kDa, which contains multiple protein domains that likely contribute to its diverse biologic properties. In addition, lubricin is well known to be 50% w/w O-glycosylated with β(1-3)-Gal-GalNAc and nonuniformly capped with NeuAc. The former posttranslational modification is essential to the protein’s lubricating ability [5], hydration force [12], conferring steric [13] and repulsion. The lubricin’s 1,404 amino acid sequence was analyzed and has revealed to be related to vitronectin in both the N-terminus and the C-terminus [14]. Both lubricin and vitronectin contain somatomedin B (SMB) and hemopexin-like (PEX) domains, which have approximately 60% sequence similarity in these regions. SAM and PEX domains have been suggested to regulate the complement and coagulation systems, mediate extracellular matrix attachment, and promote cell attachment and proliferation [15–17]. Purified hemopexin interacts with hyaluronan (HA) [18], suggesting that the hemopexin-like domain could also mediate lubricin binding to HA present at or near the articular surface [19]. In addition, lubricin attachment to the articular surface may be maintained by disulfide bond formation, via the unmatched cysteine residue near the C-terminus [20].

Lubricin distribution

Lubricin was first isolated based on its role as a lubricant in SF [21, 22], and this protein had previously been shown to be a proteoglycan specifically synthesized and expressed by articular chondrocytes of the superficial zone [23]. Flannery et al. [10] and Schumacher et al. [24] both found that some synovial lining cells such as synoviocytes and cells bordering the joint cavity synthesize lubricin. Another study by Schumacher et al. [25] showed lubricin was visualized in sections of bovine calf meniscus by immunohistochemistry. In their study, lubricin was detected in two regions including surfaces of meniscus and within and near cells along the radial fibers and circumferential fibers. Rees et al. [26] demonstrated that lubricin was present predominantly at the surface of fibrocartilaginous regions of tendon using immunohistochemical analyses, with the intensity of immunoreactivity in this region increasing with age. The lubricin was also present with infrapatellar fat pad (IFP) of the knee and was synthesized and secreted by IFP stromal cells, while IFP produced lubricin into synovial fluid and therefore contributed to maintain healthy joint function and homeostasis [27]. Moreover, a recent study by Sun et al. [28] demonstrated that lubricin was present in many joint tissues and for the first time they concluded there were lubricin present in the anterior cruciate ligament (ACL) and knee lateral collateral ligament (LCL). Meanwhile, they found the distribution of lubricin varied by different tissue type, and this variations in splicing and physical distribution suggested the variants of lubricin may play different roles in different locations. The distribution of lubricin in joint cavity is shown in Table 1.

Table 1 Distribution of lubricin in joint cavity
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Lubricin function and regulation

The discrete functional domains in lubricin suggest multiple biological functions, such as matrix binding, lubrication, cytoprotection and growth promotion. Based on biochemical composition and localization at the surface layer of articular cartilage, lubricin is known to function as a boundary lubricant in articular joints and reduces the coefficient of friction of the articualr cartilage surface [29–31]. Furthermore, mutations of PRG4 gene which located in chromosome 1q25 can cause CACP in humans [32], which is an autosomal recessive disease characterized by synovial hyperplasia with nonflammatory early-onset joint failure [33]. Expression of the mouse lubricin gene increased with progression of ectopic ossification [8]. Mice lacking the PRG4 gene showed abnormal protein deposits on the cartilage surface, disappearance of the underlying superficial zone, synovial hyperplasia, and precocious failure of joint function [3]. In addition, as the mice aged, intimal cells in the synovium surrounding the joint space became hyperplastic and further contributed to joint failure, while purified or recombinant lubricin could inhibit the growth of these synoviocytes in vitro [3]. Furthermore, as a splicing variant of lubricin, HAPO, was reported to be a novel growth factor acting on the primitive cells of both hematopoietic and endothelial cell lineage [11].

The synthesis and secretion of lubricin were determined by a complex combination of chemical and mechanical stimuli. Schmidt et al. [34] found that inclusion of fetal bovine serum and ascorbate in culture media up-regulated lubricin secretion levels. Later in their recent study, they demonstrated lubricin expression by chondrocytes near the articular surface was markedly stimulated by TGF-β1, decreased by IL-1α, but not affected by IGF-1 [35], which findings were consistent with previous studies on the regulation of lubricin by growth factors except IGF-1, who has been shown to increase the lubricin expression [36, 37]. Moreover, Niikura et al. [38] demonstrated that the regulation of lubricin accumulation by TGF-β/BMP superfamily members was regulated differently in articular chondrocytes and synoviocytes. TGF-β was a critical regulator of lubricin in both superficial zone articular chondrocytes and synoviocytes, while synoviocytes were more sensitive to BMP family members than were superficial zone articular chondrocytes. TGF-β and BMP were also found have an additive effect on the accumulation of lubricin expression [38]. In addition, various other cytokines and growth factors such as Oncostatin M (OSM), FGF-2 and PDGF were also proven to be the potent stimulators of lubricin synthesis while proinflammatory cytokines such as IL-1β and TNF-α were shown to suppress lubricin levels in monolayer chondrocytes cultures [36, 39]. Moreover, lubricin has been shown to be proteolytically susceptible to the effects of cysteine proteases, such as cathepsin B, and serine proteases, such as neutrophil elastase [40]. Mechanical stimulation can also affect the biosynthesis and secretion of lubricin. During embryonic development of the mouse elbow joint, the mRNA expression of lubricin begins at the onset of joint cavitation, suggesting that lubricin expression might be induced by the intiation of relative motion between the articular surfaces [3]. The certain mechanical stimuli including load-induced flow by interstitial fluid [41], cyclical tension [42], articular surface motion [43, 44] and chondrocyte subpopulations surface motion [45, 46] can up-regulate chondrocyte lubricin mRNA expression by chondrocyte-seeded cartilaginous constructs. Nugent et al. [47, 48] found that both static and dynamic shear stimulation increased cartilage secretion of lubricin, and the dynamic shear stimulation increased lubricin’s secretion to 3–4 times that of unloaded controls and statically compressed samples [48]. The number of cells expressing lubricin was higher in dynamically sheared cartilage and lubricin expressed at the depths of 200–400 μm from the articualr surface where were lubricin-negative in normal cartilage [48]. Furthermore, they suggested there would be a possible feedback mechanism for mechanical regulation of the production of lubricin molecule in vivo, that the tissue strains imparted to cartilage and depleted boundary lubricant molecules at the surface of articular simultaneously signal the nearby chondrocytes to secrete more lubricin [48]. Another study by Das et al. [49] demonstrated that stretch led to an up-regulation of lubricin in gene expression in primary chondrocytes. The regulation of lubricin is shown in Table 2.

Table 2 Regulation of lubricin
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Role of lubricin in articular cartilage

Lubricin is secreted predominatly by superficial zone chondrocytes, while cells from the middle and deep zone synthesize very low amounts of this protein. The biosynthesis of lubricin by articular chondrocytes has therefore been used to demarcate the superficial zone of cartilage [4]. As a surface-active mucin glycoprotein that physiosorbs to surfaces, lubricin thus providing joint lubrication, antiadhesive properties, chondroprotective and prevention of wear of articular cartilage [3]. Lubricin has been proposed to be a key factor for joint lubrication, a in vitro study had found that treatment with lubricin reduced the adhesive strength by more than 10-fold, and lubricin at a concentration necessary for boundary lubrication markedly reduced cartilage-cartilage integration [50]. Recent work revealed a correlation between lubricin expression level at the articular surface and short-term friction coefficient at the cartilage-glass interface [31]. Moreover, DuRaine et al. [51] demonstrated that the localization of lubricin in different zone would affect the friction coefficient of articular cartilage. During normal joint articulation, expression of lubricin has an important role for both preventing cell attachment to the articular surface as well as maintaining lubrication properties at the cartilage-synovial fluid interface [10]. Loss of lubricin influences the functional properties of synovial joints and could have a role in the pathogenesis of cartilage degeneration [52]. Jay and coworkers have demonstrated that lubricin provides essential chondroprotective properties to articular cartilage, and can interact with HA in synovial fluid to enhance its chondroprotective properties via the dissipation of shear-induced energy [40, 53]. Nevertheless, in addition to its function as a boundary lubricant, other studies found that lubricin appeared to have a detrimental effect in damaged joints by coating the surfaces of damaged cartilage and inhibiting integrative repair of articular cartilage lacerations [54].

Role of lubricin in joint disease

Justen et al. [6] have demonstrated that PRG4 gene expressed differently in the synovium of rheumatoid arthritis and osteoarthritis, which implied a possible role in the pathogenesis of these disease. SF lubricin concentrations were significantly reduced at an early stage following anterior cruciate ligament (ACL) injury when compared with those in the contralateral joint [40], and within 12 months, the lubricin concentration in the injured knee approached that in the contralateral knee, which did not change with time [40]. Therefore the decrease in SF lubricin concentrations following ACL injury may place the joint at an increased risk of wear-induced damage as a consequence of lack of boundary lubrication, potentially leading to secondary OA. In a rabbit knee injury model, Elsaid and coworkers found that the articular cartilage degradation were associate with the loss of boundary-lubricating ability of SF, while the lubricin concentrations in SF were markedly decreased after injury [55]. They also found that the association existed in patients with acute knee injuries or progressive chronic inflammatory arthritis. Another study by Jay et al. [56] demonstrated that SF lacking lubricin failed to reduce friction in the boundary mode, and joints of lubricin-null mice showed early and higher friction than joints from their wild-type counterparts, while friction was coupled with wear at the cartilage surface in vivo. They suggested that acquired lubricin degradation occurring in inflammatory joint disease predisposed the cartilage to damage [56]. Additionally, decreasing lubricin synthesis by synovial fibroblasts and superficial zone chondrocytes were found in a rat antigen-induced arthritis model [57], while the authors suggested restoring chondroprotection and preventing potential wear-induced cartilage degradation may require lubricin supplementation in synovial fluid. Lubricin also expressed in the cartilaginous deposits and osteoarthritic cartilage in patients with advanced OA [58]. All these results indicated that lubricin was involved in the joint disease and may play a beneficial role against the degradation of articular cartilage.

Potential biotherapeutic approaches for osteoarthritis

The current pharmaceutical treatments for OA include NSAIDs, cyclooxygenase 2 inhibitors, and HA-based injectables, which are promoted for symptomatic relief during the process of OA. These drugs have focused mainly on the broadspectrum targeting of matrix metalloproteinase activities, however, such inhibitors have been proven ineffective or due to several side effects such as soft tissue fibroplasia, that restricted the use of these drugs in clinical [59].

Based on the function of lubricin in the joint cavity (i.e., articular cartilage) and joint disease, lubricin would be as a potential therapeutic agent in the joint disease such as OA. The synthesis and localization of lubricin have been found down-regulated in rat, sheep, and guinea pig [52, 60] models of OA, which further suggesting that augmentation of lubricin levels in OA joints could be a potential therapeutic for OA. Additionally, the stimulation of lubricin expression in chondrocytes near the articular surface may be useful for creating tissue engineered cartilage from isolated subpopulation with a surface that is bioactive and functional in lubrication [61]. In fact, recent study by Flannery and colleagues demonstrated that intraarticular treatment with recombinant lubricin could prevent the degeneration of cartilage in a rat OA model [7]. In their study, a novel recombinant lubricin construct named LUB:1 was expressed in Chinese hamster ovary (CHO) cells and isolated at high purity, and LUB:1 represented a modified version of human lubricin and exhibited ~70% amino acid sequence identity to rat lubricin. Also, this novel recombinant protein contained important domains as lubricin, and in ex vivo functional assays, LUB:1 were found efficiently binded and localized to articular cartilage surfaces. Moreover, LUB:1 was found significantly improved cartilage boundary lubrication in a custom friction testing apparatus, and LUB:1 was also shown to inhibit the binding/adhesion of synovial sarcoma cells, which was similar to the full-length lubricin [7]. After intraarticular treatment with this recombinant in the rat OA model either 3 times per week or once per week over the subsequent 4 weeks, the tibial cartilage degeneration scores, total joint scores, and cartilage lesion and degeneration width measurements were significantly lower than that PBS-treated animals, suggesting that lubricin had significant disease-modifying and chondroprotective effects during the progression of OA.

It’s worth noting that another high molecular weight polysaccharide, hyaluronic acid (HA), which has been studied for the treatment of OA over the past 2 decades, was stated played an important role during the OA treatment [62]. As an important component of synovial fluid and extracellular matrix of articular cartilage, it acts as a fluid shock absorber and it helps to maintain the structural and functional characteristics of the cartilage matrix. It also inhibits the formation and release of prostaglandins, induces proteoglycan aggregation and synthesis, and modulates the inflammatory response [63, 64]. According to Balazs and coworkers [65, 66], the injection of HA into osteoarthritic joints could restore the viscoelasticity of the synovial fluid, augment the flow of joint fluid, normalize endogenous hyaluronate synthesis, inhibit hyaluronate degradation, reduce joint pain, and improve joint function. Moreover, viscosupplementation with intra-articular HA was approved by the Food and Drug Administration (FDA) in 1997. As a novel molecular, there has been increasing interest of potential biotherapeutic approaches for the treatment of OA, and the beneficial synergy between lubricin and HA with respect to both boundary lubrication and mechanical attributes of SF has been highlighted. Schmidt et al. [67] found that lubricin and HA in combination produced a greater boundary-lubricating effect than either constituent alone. Another study by Jay et al. [53] demonstrated that lubricin can interact with HA in synovial fluid to enhance its chondroprotective properties via the dissipation of shear-induced energy.

Conclusions

Many researchers have demonstrated the role of lubricin in the joint cavity and diseases. Several studies suggested that lubricin has a major protective role in preventing cartilage wear and synovial cell adhesion and proliferation during the process of OA. Moreover, several recent studies found that treatment with recombinant lubricin could protect articular cartilage and prevent the process of OA in animal model. This suggests that lubricin is a novel potential biotherapeutic approaches to the treatment of OA. However, some concerns about technical problems for local delivery of lubricin in the joint cavity, as well as its potential side effect still remain. Further in vivo studies will increase our understanding of the ture significance of lubricin and provide the foundations for the development of effective therapy for virous joint disease, especially the OA.