Evaluation of rabbit auricular chondrocyte isolation and growth parameters in cell culture.
Auricular cartilage is an attractive potential source of cells for many tissue engineering applications. However, there are several requirements that have to be fulfilled in order to develop a suitable tissue engineered implant. Animal experiments serve as important tools for validating novel concepts of cartilage regeneration; therefore rabbit auricular chondrocytes were studied. Various parameters including isolation procedures, passage number, rate of proliferation and gene expression profile for major extracellular matrix components were evaluated in order to assess the potential use of elastic chondrocytes for tissue engineering. Chondrocytes were isolated from rabbit ear cartilage and grown in monolayer cultures over four passages. Yields of harvested cells and proliferation were analysed from the digestion step to the fourth passage, and changes in phenotype were monitored. The proliferation capacity of cell cultures decreased during cultivation and was accompanied by enlargement of cells, this phenomenon being especially evident in the third and fourth passages. The expression of cartilage specific genes for collagen type II, aggrecan and cartilage non-specific collagen type I was determined. The mRNA levels for all three genes were obviously lower in the primo culture than immediately after isolation. During subsequent cultivation the expression of collagen type II decreased further, while there were only slight changes in expression of aggrecan and collagen type I. This study provides a valuable basis for testing of different tissue engineering applications in rabbit model, where auricular chondrocytes are considered as cell source.
Fröhlich
M
M
Educell d.o.o., Letaliska cesta 33, SI-1000 Ljubljana, Slovenia. mirjam.froehlich@educell.si
Malicev
E
E
Gorensek
M
M
Knezević
M
M
Kregar Velikonja
N
N
D053674:Aggrecans / Q000235:genetics / Q000378:metabolism; D000818:Animals; D002452:Cell Count; D018929:Cell Culture Techniques; D049109:Cell Proliferation; D002469:Cell Separation; D002470:Cell Survival; D019902:Chondrocytes / Q000166:cytology*; D024042:Collagen Type I / Q000235:genetics / Q000378:metabolism; D024043:Collagen Type II / Q000235:genetics / Q000378:metabolism; D004425:Ear Cartilage / Q000166:cytology*; D005434:Flow Cytometry; D005786:Gene Expression Regulation; D011817:Rabbits; D013997:Time Factors
D053674:Aggrecans; D024042:Collagen Type I; D024043:Collagen Type II