Open AccessChondrocytes isolation from hyaline cartilage by continuous monitoring method
Author(s) -
Vitalie Cobzac,
AUTHOR_ID,
Liliana Vereştiuc,
Mariana Jian,
Viorel Nacu,
AUTHOR_ID,
AUTHOR_ID,
AUTHOR_ID
Publication year - 2021
Publication title -
the moldovan medical journal
Language(s) - English
Resource type - Journals
eISSN - 2537-6381
pISSN - 2537-6373
DOI - 10.52418/moldovan-med-j.64-6.21.03
Subject(s) - cartilage , collagenase , chondrocyte , andrology , hyaline cartilage , anatomy , staining , laboratory flask , viability assay , extracellular matrix , chemistry , matrix (chemical analysis) , decellularization , tissue engineering , pathology , cell , medicine , osteoarthritis , biomedical engineering , articular cartilage , biochemistry , chromatography , alternative medicine , enzyme
Background: Articular cartilage has poor regenerative capacities. Numerous cartilage repair techniques are known, including implantation of autologous chondrocytes. Material and methods: From 18 rabbits pieces of cartilage were harvested from femoral condyle. Minced cartilage was treated with 0.25% trypsin-EDTA. In the 1st group (n=9) the cartilage was digested with 0.6% collagenase in 15 ml tubes by shaking in incubator at 37°C, 5%CO2 . In the 2nd group (n=9) digestion was performed in 25cm2 cell culture flasks placed on the lateral side, monitoring the process under a microscope after 120 minutes. The isolated cells were cultured to a 80-90% confluence. The chondrocytes were identified using histochemical staining after culturing for 16 days in overconfluence. Results: Chondrocytes isolation in the 1st group lasted a fixed 360 minutes, in the 2nd group – 140±10 minutes. In the 1stgroup were isolated 9.2x104 ±3.1x104 chondrocytes with a viability of 85.36±16.41%, but in the 2nd group – 1.6x105 ±3.4x104 chondrocytes with a viability of 98.09±3.85%. The mean period of cell culture in the 1st group was 15±2 days, in the 2nd group – 11±3 days. In first passage of the 1st group were obtained – 1.2x106 ±4.3x105 chondrocytes and in the 2nd group – 2.92x106 ±3.6x105 chondrocytes. The secreted extracellular matrix by chondrocytes was stained specifically for cartilaginous tissue. Conclusions: The method used for chondrocytes isolation has a direct impact on the number of isolated cells, their viability, but also upon the culture period and the number of cells obtained during the first passage.