Matrix molecule influence on chondrocyte phenotype and proteoglycan 4 expression by alginate‐embedded zonal chondrocytes and mesenchymal stem cells

Abstract

Articular cartilage resists load and provides frictionless movement at joint surfaces. The tissue is organized into the superficial, middle, deep, and calcified zones throughout its depth, each which serve distinct functions. Proteoglycan 4 (PRG4), found in the superficial zone, is a critical component of the joint's lubricating mechanisms. Maintenance of both the chondrocyte and zonal chondrocyte phenotype remain challenges for in vitro culture and tissue engineering. Here we investigate the expression of PRG4 mRNA and protein by primary bovine superficial zone chondrocytes, middle/deep zone chondrocytes, and mesenchymal stem cells encapsulated in alginate hydrogels with hyaluronic acid (HA) and chondroitin sulfate (CS) additives. Chondrogenic phenotype and differentiation markers are evaluated by mRNA expression, histochemical, and immunohistochemical staining. Results show middle/ deep cells express no measurable PRG4 mRNA by day 7. In contrast, superficial zone cells express elevated PRG4 mRNA throughout culture time. This expression can be significantly enhanced up to 15-fold by addition of both HA and CS to scaffolds. Conversely, PRG4 mRNA expression is downregulated (up to 5-fold) by CS and HA in differentiating MSCs, possibly due to build up of entrapped protein.HA and CS demonstrate favorable effects on chondrogenesis by upregulating transcription factor Sox9 mRNA (up to 4.6-fold) and downregulating type I collagen mRNA (up to 18-fold). Results highlight the important relationship between matrix components and expression of critical lubricating proteins in an engineered cartilage scaffold.

Extracellular-Matrix-Based and Arg-Gly-Asp–Modified Photopolymerizing Hydrogels for Cartilage Tissue Engineering

“…37 Additionally recent study indicated that ECM, such as CS and HA, added to alginate hydrogel induced lubricin expression. 38 Our findings confirmed that incorporation of these ECMs along with cell-adhesions moieties enhanced lubricin secretion of encapsulated chondrocytes. ECM molecule included hydrogel with cell-adhesive characteristic enhanced lubricin secretion.…”

Engineering Superficial Zone Chondrocytes from Mesenchymal Stem Cells

“…Reports from the literature support that mesechymal progenitor cells express PRG4 during TGF-b-induced chondrogensis, however, the mechanisms and pathways involved in this expression are not clear. 12,26,30 As evidence now supports a population of progenitor cells within the superficial zone, stem cell differentiation may be a normal biological process within this zone, and thus signals derived from it provide favorable signals for chondrogenesis. What is not clear from our data is the source, or identity, of the factors that induce chondrogenesis in the superficial zone coculture groups.…”

“…25,26 Briefly, cartilage plugs (4 mm diameter, 3-6 mm height) were harvested from the femoral condyles of 20-week-old calves using a Sklar Tru-Punch disposable biopsy punch (Sklar Instruments). The top 10% (0.3-0.6 mm) was taken as the superficial zone, and remaining tissue up was defined as the middle and deep tissue zones.…”

Pronounced Biomaterial Dependency in Cartilage Regeneration Using Nonexpanded Compared with Expanded Chondrocytes

“…HyStem consists of HA, a naturally occurring polysaccharide in cartilage with many prochondrogenic functions [1], which makes it a promising biomaterial for cartilage engineering. Indeed, addition of HA to other biomaterials such as alginate, chitosan and fibrin has been shown to enhance extracellular matrix formation [32]. Depending on the degree of polymerization, HA has been shown to have either inflammatory or anti-inflammatory effects [33].…”