A porous PCL scaffold promotes the human chondrocytes redifferentiation and hyaline-specific extracellular matrix protein synthesis.
The redifferentiation, proliferation, and hyaline-specific extracellular matrix (ECM) protein synthesis of chondrocytes cultured in a polycaprolactone (PCL) scaffold were analyzed. Gene expression of the type II collagen and aggrecan was assessed by real-time PCR in cells from PCL scaffolds, monolayer, and pellet cultures. The proliferative activity was assessed using Ki-67 immunodetection, and the chondrocytic differentiation was evaluated using S-100 immunodetection. The synthesis and deposition into scaffold pores of type II collagen and glycosaminoglycan were analyzed by immunohistochemistry and Alcian blue staining, respectively. All parameters were assessed throughout 28 days of cultures maintained in either fetal bovine serum-containing medium (FCM) or Insulin-Transferrin-Selenium-containing medium (ICM). Expression of the type II collagen gene was lower in FCM cultures than in ICM cultures for all culture systems (p < 0.05). Moreover, PCL scaffolds cultured in ICM were able to induce collagen gene expression more efficiently than pellet and monolayer cultures. Aggrecan gene expression did not vary significantly between mediums and three-dimensional system cultures, but in ICM cultures, the monolayer cultures had significantly higher levels of aggrecan gene expression than did either the PCL or pellet cultures. Chondrocytes cultured in PCL scaffolds or pellets with FCM did not proliferate to a great extent but did maintain their differentiated phenotype for 28 days. Levels of cartilage ECM protein synthesis and deposition into the scaffold pores were similar among PCL and pellet cultures grown in FCM and in ICM. In conclusion, chondrocytes seeded into PCL scaffolds, cultured in ICM, efficiently maintained their differentiated phenotype and were able to synthesize cartilage-specific ECM proteins.
Garcia-Giralt
N
N
URFOA-IMIM, Hospital del Mar, Universitat Autònoma de Barcelona, C/Doctor Aiguader 88, E-08003 Barcelona, Spain. ngarcia@imim.es
Izquierdo
R
R
Nogués
X
X
Perez-Olmedilla
M
M
Benito
P
P
Gómez-Ribelles
J L
JL
Checa
M A
MA
Suay
J
J
Caceres
E
E
Monllau
J C
JC
D053674:Aggrecans / Q000235:genetics / Q000378:metabolism; D000423:Alcian Blue; D002454:Cell Differentiation / Q000187:drug effects*; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D019902:Chondrocytes / Q000166:cytology* / Q000187:drug effects; D024043:Collagen Type II / Q000235:genetics / Q000378:metabolism; D016326:Extracellular Matrix Proteins / Q000096:biosynthesis* / Q000235:genetics; D005786:Gene Expression Regulation / Q000187:drug effects; D006416:Hematoxylin; D006801:Humans; D006818:Hyalin / Q000378:metabolism*; D007150:Immunohistochemistry; D011091:Polyesters / Q000494:pharmacology*; D016062:Porosity / Q000187:drug effects; D014176:Protein Biosynthesis / Q000187:drug effects*; D054457:Tissue Scaffolds
D053674:Aggrecans; D024043:Collagen Type II; D016326:Extracellular Matrix Proteins; D011091:Polyesters; C016240:polycaprolactone; D000423:Alcian Blue; D006416:Hematoxylin