Xiong Liuliu, Cui Meng, Zhou Ziye, Wu Minchen, Wang Quanming, Song Haiyan, Ding Lei
Department of Basic Medical Sciences, Jiangnan University Wuxi College of Medicine, Jiangsu 214122, China.
Department of Orthopaedic Surgery, Jiangnan University Affiliated Hospital, Jiangsu 214062, China.
Asian Biomed (Res Rev News). 2021 Apr 30;15(2):91-99. doi: 10.2478/abm-2021-0011. eCollection 2021 Apr.
Joint replacement surgery provides articular cartilage samples for chondrocyte isolation. To our knowledge, the effect of the collagenase type on releasing of chondrocytes from the extracellular matrix of cartilage is not reported.
To determine whether cartilage digested with collagenase IA yielded more chondrocytes than that digested with collagenase II and determine whether chondrocytes isolated with collagenase IA could be cultured in vitro.
Cartilage slices collected from 18 elderly patients who received joint replacement surgery (16 hips, 2 knees) were digested sequentially with 0.4% pronase E and 0.02% collagenase IA, or with 0.15% collagenase II alone, or sequentially with 0.4% pronase E and 0.02% collagenase II. We compared cell yield from each method. Cell viability by the most effective method was calculated and plotted. The morphology of cultured monolayer chondrocytes was recorded with a light microscope.
Sequential digestion with pronase E and collagenase IA yielded 2566 ± 873 chondrocytes per mg wet cartilage, which was more effective than the other isolation methods ( = 0.018). The average chondrocyte viability could reach 84% ± 8% (n = 11). Light microscopic images showed typical chondrocyte morphology in monolayer cultures.
Sequential digestion of human articular cartilage with pronase E and collagenase IA was more effective than collagenase II alone or collagenase II combined with pronase E for releasing chondrocytes from extracellular matrix of cartilage. Chondrocytes isolated with this method could be maintained in monolayer cultures for at least 2 passages with unaltered morphology.
关节置换手术可提供用于分离软骨细胞的关节软骨样本。据我们所知，目前尚无关于不同类型胶原酶从软骨细胞外基质中释放软骨细胞效果的报道。
确定用Ⅰ型胶原酶消化软骨所获得的软骨细胞是否比用Ⅱ型胶原酶消化得到的更多，并确定用Ⅰ型胶原酶分离得到的软骨细胞能否在体外培养。
收集18例接受关节置换手术患者（16例髋关节，2例膝关节）的软骨切片，分别依次用0.4%链霉蛋白酶E和0.02%Ⅰ型胶原酶消化，或仅用0.15%Ⅱ型胶原酶消化，或依次用0.4%链霉蛋白酶E和0.02%Ⅱ型胶原酶消化。比较每种方法的细胞产量。计算并绘制最有效方法的细胞活力。用光学显微镜记录培养的单层软骨细胞的形态。
依次用链霉蛋白酶E和Ⅰ型胶原酶消化，每毫克湿软骨可获得2566±873个软骨细胞，这比其他分离方法更有效（P=0.018）。软骨细胞平均活力可达84%±8%（n=11）。光学显微镜图像显示单层培养中有典型的软骨细胞形态。
对于从软骨细胞外基质中释放软骨细胞，依次用链霉蛋白酶E和Ⅰ型胶原酶消化人关节软骨比单独使用Ⅱ型胶原酶或Ⅱ型胶原酶与链霉蛋白酶E联合使用更有效。用这种方法分离得到的软骨细胞可在单层培养中维持至少2代，形态不变。
