Characterization of primary chondrocytes harvested from hips with femoroacetabular impingement.

OBJECTIVE: Acetabular chondral lesions are common in patients with femoroacetabular impingement (FAI) syndrome. The aim of this study was (1) to evaluate the proliferation potential of primary human chondrocytes (hC) derived from both acetabular and femoral site and (2) to validate cellular differentiation during three-dimensional (3D) cultivation as a prerequisite for autologous matrix-assisted cartilage regeneration of the hip joint. METHODS: hC were isolated from cartilage samples obtained from N = 6 patients during offset reconstruction. Proteoglycan content was assessed by Safranin-O staining. Proliferation and cell viability were quantified by microscopic cell counting and Trypan Blue exclusion. Messenger ribonucleic acid (mRNA) expression levels of collagen type 1 and 2, aggrecan (ACAN), and interleukin-1ß (IL-1ß) genes were assessed upon monolayer cultivation, after 48 h/4-10°C - transport simulation and after 14 days of 3D hydrogel cultivation. RESULTS: Primary hC from acetabular and femoral damaged sites were viable. No significant intergroup differences were observed concerning cell viability (>95%) after monolayer cultivation and transport simulation. Harvest yields from acetabular and femoral cartilage samples were comparable to that known from knee joints (mean ± standard deviation (SD), 13.4 × 10(6) ± 5 × 10(6) cells per culture vs 20 × 10(6) cells). Redifferentiation was induced during 3D hydrogel cultivation as observed by increased levels of collagen II (1000-fold) and ACAN (10-fold) gene vs monolayer cultivation (P < 0.001). CONCLUSION: hC derived from damaged acetabular and femoral site are qualified for autologous matrix-assisted cartilage transplantation paving the way for cell-based cartilage regeneration in FAI patients.

Genomic chondrocyte culture profiling by array-CGH, interphase-FISH and RT-PCR.

OBJECTIVE: In vitro expansion is an important step to acquire sufficient cells in human tissue engineering technologies. The high number of chondrocytes needed for human articular cartilage implants requires in vitro expansion of the primary cells, bearing a theoretical risk of in vitro induced changes in the genomes. To gain more insights into this situation, model cultures were prepared and analyzed. DESIGN: 25 chondrocyte cell DNA samples from nine donors were analyzed by array comparative genomic hybridization (aCGH) on whole genome level and 28 chondrocyte cell samples from 16 individuals were analyzed by fluorescence in situ hybridization (FISH) on single cell level. The expanded cells were further characterized upon the chondrocytic mRNA phenotype by reverse-transciptase polymerase chain reaction (RT-PCR). RESULTS: The molecular karyotyping results revealed autosomal stability, but all male samples analyzed by aCGH displayed a variable loss of the Y-chromosome. These data were confirmed by FISH-experiments and suggest an age dependant effect toward the loss of the Y-chromosome in cultured chondrocytes. RT-PCR data for the mRNAs from collagen types I, II, and aggrecan and the pro-inflammatory cytokine interleukin-1ß (IL-1ß) did not reveal any correlation of transcriptional activity in cultures with Y-chromosome losses, nor were there statistically significant differences between cells from female and male donors. CONCLUSIONS: While cells of male origin may suffer from an age-related loss of the Y-chromosome, there was no indication of a functional impairment. The data suggest some caution toward applying proliferative steps when considering chondrocytes from elderly male patients for tissue engineering approaches.