Montembault A, Tahiri K, Korwin-Zmijowska C, Chevalier X, Corvol M-T, Domard A
CNRS UMR 5627, 69622 Villeurbanne, France.
Biochimie. 2006 May;88(5):551-64. doi: 10.1016/j.biochi.2006.03.002. Epub 2006 Mar 31.
The cartilage tissue has a limited self-regenerative capacity. Tissue-engineering represents a promising trend for cartilage repair. The present study was aimed to develop a biomaterial formulation by combining fragments of chitosan hydrogel with isolated rabbit or human chondrocytes. We first reported the properties of the constructs elaborated with rabbit chondrocytes and pure chitosan physical hydrogels with defined molecular weight, acetylation degree and polymer concentration. Morphological data showed that chondrocytes were not penetrating the hydrogels but tightly bound to the surface of the fragments and spontaneously formed aggregates of combined cell/chitosan. A significant amount of neo-formed cartilage-like extracellular matrix (ECM) was first accumulated in-between cells and hydrogel fragments and furthermore was widely distributed within the neo-construct. The optimal biological response was obtained with hydrogel fragments concentrated at 1.5% (w/w) of polymer made from a chitosan with a degree of acetylation between 30 and 40%. Such hydrogels were then mixed with human chondrocytes. The phenotype of the cells was analyzed by using chondrocytic (mRNA expression of mature type II collagen and aggrecan as well as secretion of proteoglycans of high molecular weight) and non chondrocytic (mRNA expression of immature type II collagen and type I collagen) molecular markers. As compared with human chondrocytes cultured without chitosan hydrogel which rapidly dedifferentiated in primary culture, cells mixed with chitosan rapidly loose the expression of type I and immature type II collagen while they expressed mature type II collagen and aggrecan. In these conditions, chondrocytes maintained their phenotype for as long as 45 days, thus forming cartilage-like nodules. Taken together, these data suggest that a chitosan hydrogel does not work as a scaffold, but could be considered as a decoy of cartilage ECM components, thus favoring the binding of chondrocytes to chitosan. Such a biological response could be described by the concept of reverse encapsulation.
软骨组织的自我再生能力有限。组织工程是软骨修复的一个有前景的发展趋势。本研究旨在通过将壳聚糖水凝胶片段与分离的兔或人软骨细胞相结合来开发一种生物材料配方。我们首次报道了用兔软骨细胞和具有确定分子量、乙酰化程度和聚合物浓度的纯壳聚糖物理水凝胶制备的构建体的特性。形态学数据表明，软骨细胞没有穿透水凝胶，而是紧密结合在片段表面，并自发形成细胞/壳聚糖组合聚集体。大量新形成的软骨样细胞外基质（ECM）首先在细胞和水凝胶片段之间积累，并且进一步广泛分布在新构建体中。当由乙酰化程度在30%至40%之间的壳聚糖制成的聚合物以1.5%（w/w）的浓度浓缩水凝胶片段时，可获得最佳生物学反应。然后将这种水凝胶与人类软骨细胞混合。通过软骨细胞（成熟II型胶原蛋白和聚集蛋白聚糖的mRNA表达以及高分子量蛋白聚糖的分泌）和非软骨细胞（未成熟II型胶原蛋白和I型胶原蛋白的mRNA表达）分子标记物来分析细胞的表型。与在原代培养中迅速去分化的未使用壳聚糖水凝胶培养的人类软骨细胞相比，与壳聚糖混合的细胞迅速失去I型和未成熟II型胶原蛋白的表达，同时表达成熟II型胶原蛋白和聚集蛋白聚糖。在这些条件下，软骨细胞保持其表型长达45天，从而形成软骨样结节。综上所述，这些数据表明壳聚糖水凝胶不作为支架起作用，但可被视为软骨ECM成分的诱饵，从而有利于软骨细胞与壳聚糖的结合。这种生物学反应可以用反向包封的概念来描述。
