Serum-free, chemically defined medium with TGF-beta(3) enhances functional properties of nucleus pulposus cell-laden carboxymethylcellulose hydrogel constructs.
Degeneration of the nucleus pulposus (NP) has been implicated as a major cause of low back pain. Tissue engineering strategies may provide a viable NP replacement therapy; however, culture conditions must be optimized to promote functional tissue development. In this study, a standard serum-containing medium formulation was compared to a chemically defined, serum-free medium to determine the effect on matrix elaboration and functional properties of NP cell-laden carboxymethylcellulose (CMC) hydrogels. Additionally, both media were further supplemented with transforming growth factor-beta 3 (TGF-beta(3)). Glycosaminoglycan (GAG) content increased in both TGF-beta(3)-treated groups and was highest for treated, serum-free constructs (9.46 +/- 1.51 microg GAG/mg wet weight), while there were no quantifiable GAGs in untreated serum-containing samples. Histology revealed uniform, interterritorial staining for chondroitin sulfate proteoglycan throughout the treated, serum-free constructs. Type II collagen content was greater in both serum-free groups and highest in treated, serum-free constructs. The equilibrium Young's modulus was highest in serum-free samples supplemented with TGF-beta(3) (18.54 +/- 1.92 kPa), and the equilibrium weight swelling ratio of these constructs approached that of the native NP tissue (22.19 +/- 0.46 vs. 19.94 +/- 3.09, respectively). Taken together, these results demonstrate enhanced functional matrix development by NP cells when cultured in CMC hydrogels maintained in serum-free, TGF-beta(3) supplemented medium, indicating the importance of medium formulation in NP construct development.
Department of Bioengineering, University of Pennsylvania, Philadelphia, 19104, USA.
Anna T
AT
Reza
Steven B
SB
Nicoll
D000818:Animals; D002266:Carboxymethylcellulose Sodium / Q000737:chemistry*; D002417:Cattle; D002478:Cells, Cultured; D016895:Culture Media, Serum-Free / Q000378:metabolism; D055119:Elastic Modulus; D006025:Glycosaminoglycans / Q000378:metabolism; D020136:Hydrogel, Polyethylene Glycol Dimethacrylate / Q000737:chemistry*; D007403:Intervertebral Disc / Q000166:cytology*; D023822:Tissue Engineering / Q000379:methods*; D053782:Transforming Growth Factor beta3 / Q000378:metabolism*
D016895:Culture Media, Serum-Free; D006025:Glycosaminoglycans; D053782:Transforming Growth Factor beta3; D020136:Hydrogel, Polyethylene Glycol Dimethacrylate; D002266:Carboxymethylcellulose Sodium