Principios en el aislamiento y cultivo de condrocitos de
acuerdo a buenas prácticas de laboratorio
José Manuel Muñoz Olmedo¹, Denisa Harvanová¹, Tímea Špaková¹, Ján Rosocha¹, Marek Lacko ² , Darina Bacenková¹ and Judita Amrichová¹
(1) Associated Tissue Bank of Faculty of Medicine of
P. J. Šafárik University and L. Pasteur University Hospital, 041 66 Košice, Slovakia.
(2) Department of Orthopaedics and Traumatology of Locomotory Apparatus, Faculty of Medicine of
P. J. Šafárik University and L. Pasteur University Hospital, 041 66 Košice, Slovakia.
OBJEC TIVES : The objective of the present study was to determine if chondrocytes isolated from human cartilage of five elderly patients (mean age 63) with osteoarthritis (stage 3) maintain their proliferation and chondrogenic potential. Isolation and cultivation of chondrocytes was performed according to good laboratory practice (GLP ) standards. Methods: Chondrocytes were isolated from cartilage biopsy by enzymatic digestion. Cultivation of cells was performed in a controlled environment (cleanroom). Phenotype characterization of chondrocytes was achieved by flow cytometry analysis. Results: Three weeks after cultivation, polygonal
structures typical of chondrocytes were observed, but spindle/fibroblast like morphology was also detected in cultured cells. Flow cytometric
analysis showed that chondrocytes were positive for CD 44 (98.35% ± 0.50), CD 90 (97.15% ± 0.13) after first passage (P1) and the cells were negative
for hematopoietic marker CD 45 (0.21% ± 0.11). ConclusionS: Human articular chondrocytes obtained from five elderly patients with
osteoarthritis maintained a chondrocyte phenotype and could be potentially used for autologous implantation. We have standardized the conditions for cultivation according to GLP standards to minimize the risk of in vitro cell contamination.
Keywords: autologous implantation, cartilage, cleanroom, chondrocytes.
Palabras clave: IMPLANTACIÓN AUTÓLOGA , CARTÍLAGO , sala blanca, condrocitos.