Abstract
Introduction. Tissue engineering is a new methodology for addressing meniscal injury or loss. Synovium may be an ideal source of cells for in vitro meniscal fibrocartilage formation, however, favorable in vitro culture conditions for synovium must be established in order to achieve this goal. The objective of this study was to determine cellularity, cell distribution, and extracellular matrix (ECM) formation of equine fibroblast-like synoviocytes (FLS) cultured on synthetic scaffolds, for potential application in synovium-based meniscal tissue engineering. Scaffolds included open-cell poly-L-lactic acid (OPLA) sponges and polyglycolic acid (PGA) scaffolds cultured in static and dynamic culture conditions, and PGA scaffolds coated in poly-L-lactic (PLLA) in dynamic culture conditions.Materials and Methods. Equine FLS were seeded on OPLA and PGA scaffolds, and cultured in a static environment or in a rotating bioreactor for 12 days. Equine FLS were also seeded on PGA scaffolds coated in 2% or 4% PLLA and cultured in a rotating bioreactor for 14 and 21 days. Three scaffolds from each group were fixed, sectioned and stained with Masson’s Trichrome, Safranin-O, and Hematoxylin and Eosin, and cell numbers and distribution were analyzed using computer image analysis. Three PGA and OPLA scaffolds from each culture condition were also analyzed for extracellular matrix (ECM) production via dimethylmethylene blue (sulfated glycosaminoglycan) assay and hydroxyproline (collagen) assay. PLLA coated PGA scaffolds were analyzed using double stranded DNA quantification as areflection of cellularity and confocal laser microscopy in a fluorescent cell viability assay.Results. The highest cellularity occurred in PGA constructs cultured in a rotating bioreactor, which also had a mean sulfated glycosaminoglycan content of 22.3 µg per scaffold. PGA constructs cultured in static conditions had the lowest cellularity. Cells had difficulty adhering to OPLA and the PLLA coating of PGA scaffolds; cellularity was inversely proportional to the concentration of PLLA used. PLLA coating did not prevent dissolution of the PGA scaffolds. All cell scaffold types and culture conditions produced non-uniform cellular distribution.Discussion/Conclusion. FLS-seeding of PGA scaffolds cultured in a rotating bioreactor resulted in the most optimal cell and matrix characteristics seen in this study. Cells grew only in the pores of the OPLA sponge, and could not adhere to the PLLA coating of PGA scaffold, due to the hydrophobic property of PLA. While PGA culture in a bioreactor produced measureable GAG, no culture technique produced visible collagen. For this reason, and due to the dissolution of PGA scaffolds, the culture conditions and scaffolds described here are not recommended for inducing fibrochondrogenesis in equine FLS for meniscal tissue engineering.
Highlights
The knee menisci are semilunar-shaped fibrocartilages with extracellular matrix (ECM)composed primarily of types I and II collagen, glycosaminoglycans (GAGs), and water (Fithian et al 1990)
The second objective was to compare cell viability, distribution, and ECM formation of fibroblast-like synoviocytes (FLS) cultured on 2% vs 4% Poly-L –Lactic acid (PLLA) coated PGA scaffolds, cultured for 14 or 21 days
The current study analyzed the effect of scaffold type, biomechanical stimuli, and culture duration on FLS seeding and production of specific meniscal ECM constituents
Summary
The knee menisci are semilunar-shaped fibrocartilages with extracellular matrix (ECM)composed primarily of types I and II collagen, glycosaminoglycans (GAGs), and water (Fithian et al 1990). Avascular portion of the meniscus has a limited ability to heal spontaneously, (Arnoczky & Warren 1983; Kobayashi et al 2004), the majority of meniscal injuries are treated with partial menisectomy. This results in eventual articular cartilage damage of the tibia and femoral condyles, and progression of debilitating osteoarthritis (Arnoczky & Warren 1983; Cox et al 1975). Tissue engineering new meniscal fibrocartilage is being investigated as a treatment for avascular meniscal injuries
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
