The effects of protein-coated surfaces on passaged porcine TMJ disc cells

Biotech & Bioengineering

2009

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The knee meniscus, a fibrocartilaginous tissue located in the knee joint, is characterized by heterogeneity in extracellular matrix and biomechanical properties. To recreate these properties using a tissue engineering approach, co-cultures of meniscus cells (MCs) and articular chondrocytes (ACs) were seeded in varying ratios (100:0, 75:25, 50:50, 25:75, and 0:100) on poly-L-lactic acid (PLLA) scaffolds and cultured in serum-free medium for 4 weeks. Histological, biochemical, and biomechanical tests were used to assess constructs at the end time point. Strong staining for collagen and glycosaminoglycan (GAG) was observed in all groups. Constructs with 100% MCs were positive for collagen I and constructs cultured with 100% ACs were positive for collagen II, while a mixture of collagen I and II was observed in other co-culture groups. Total collagen and GAG per construct increased as the percentage of ACs increased (27 +/- 8 microg, 0% AC to 45 +/- 8 microg, 100% ACs for collagen and 12 +/- 4 microg, 0% ACs to 40 +/- 5 microg, 100% ACs for GAG). Compressive modulus (instantaneous and relaxation modulus) of the constructs was significantly higher in the 100% ACs group (63 +/- 12 and 22 +/- 9 kPa, respectively) when compared to groups with higher percentage of MCs. No differences in tensile properties were noted among groups. Specific co-culture ratios were identified mimicking the GAG/DW of the inner (0:100, 25:75, and 50:50) and outer regions (100:0) of the meniscus. Overall, it was demonstrated that co-culturing MCs and ACs on PLLA scaffolds results in functional tissue engineered meniscus constructs with a spectrum of biochemical and biomechanical properties.

Section: Introductionmentioning

confidence: 97%

Effects of co‐cultures of meniscus cells and articular chondrocytes on PLLA scaffolds

Gunja

Biotech & Bioengineering

2009

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“…The expansion of these cells, however, results in phenotypic loss with rapid drop in cellular collagen II and COMP expression (Allen and Athanasiou, 2007; Darling and Athanasiou, 2005; Gunja and Athanasiou, 2007b). Researchers have employed several vehicles to recover losses in gene expression by culturing cells in three dimensional alginate gels, plating cells on two dimensional surface coatings and adding growth factors to the expansion medium (Allen et al ., 2008; Benya and Shaffer, 1982; Darling and Athanasiou, 2005; Gunja and Athanasiou, 2007b; Martin et al ., 1999). These techniques have been generally successful in reversing the effects of markers such as COMP and collagen I; however, collagen II expression reversal has been particularly challenging.…”

Section: Introductionmentioning

confidence: 99%

Additive and synergistic effects of bFGF and hypoxia on leporine meniscus cell-seeded PLLA scaffolds

Gunja

2010

J Tissue Eng Regen Med

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Injuries to avascular regions of menisci do not heal and result in significant discomfort to patients. Current treatments, such as partial meniscectomy, alleviate these symptoms in the short-term, but lead to premature osteoarthritis as a result of compromised stability and changes in knee biomechanics. Thus, tissue engineering of the meniscus may provide an alternative treatment modality to overcome this problem. In this experiment, a scaffold-based tissue engineering approach was utilized to regenerate the meniscus. Meniscus cells were cultured on poly-l-lactic acid scaffolds in normoxic (~21% oxygen) or hypoxic (~2% oxygen) conditions in the presence or absence of the growth factor, basic fibroblast growth factor (bFGF). At t = 4 wks, histological sections of constructs showed presence of collagen and glycosaminoglycan (GAG) in all groups. Immunohistochemical staining showed presence of collagen I in all groups, and collagen II in groups cultured in hypoxic conditions. bFGF in the culture medium significantly increased cell number/construct by 25%, regardless of culturing condition. For GAG/construct, synergistic increases were observed in constructs cultured in hypoxic conditions and bFGF (2-fold) when compared to constructs cultured in normoxic conditions. Compressive tests showed synergistic increases in the relaxation modulus and coefficient of viscosity, and additive increases in the instantaneous modulus for constructs cultured in hypoxic conditions and bFGF when compared to constructs cultured in normoxic conditions. Overall, these results demonstrate that bFGF and hypoxia can significantly enhance the ability of meniscus cells to produce GAGs and improve the compressive properties of tissue engineered meniscus constructs in vitro.

“…Despite numerous attempts, these cells have yet to approach the quantitative biochemical content or mechanical strength necessary to function in a tissue replacement. Additionally, these cells are difficult to obtain and very limited in quantity, which is not likely remedied through in vitro cell expansion and passaging 15, 16. Recent work with costal chondrocytes (CCs), however, suggests their potential in TMJ disc tissue engineering both in functionality and clinical translatability 17.…”

Section: Introductionmentioning

confidence: 99%

A comparison of primary and passaged chondrocytes for use in engineering the temporomandibular joint

Anderson

Archives of Oral Biology

2009

Objective-This study examines the tissue engineering potential of passaged (P3) and primary (P0) articular chondrocytes (ACs) and costal chondrocytes (CCs) from skeletally-mature goats for use in the temporomandibular joint (TMJ).Design-These four cell types were assembled into scaffoldless tissue engineered constructs and cultured for 4 wks. The constructs were then tested for cell, collagen, and glycosaminoglycan (GAG) content with biochemical assays, and collagen types I and II with enzyme-linked immunosorbent assays. Constructs were also tested under tension and compression to determine biomechanical properties.Results-Both primary and passaged CC constructs had greater GAG/wet weight than AC constructs. Primary AC constructs had significantly less total collagen and contained no collagen type I. AC P3 constructs had the largest collagen I/collagen II ratio, which was also greater in passaged CC constructs relative to primary groups. Primary AC constructs were not mechanically testable, while passaged AC and CC constructs had significantly greater tensile properties than primary CC constructs. Conclusions-PrimaryCCs are considerably better than primary ACs and have potential use in tissue engineering when larger quantities of collagen type II are desired. The poor performance of the ACs, in this study, which contradicts the results seen with previous studies using immature bovine ACs, may thus be attributed to the animals' maturity. However, CC P3 cells appear particularly wellsuited for tissue engineering fibrocartilage of the TMJ due to the high quantity of collagen and GAG, and tensile and compressive mechanical properties.