The basic research on functionalization of the natural polysaccharide and application to biomedical materials.

本实验研究了硫酸多糖复合物（PEC）对硬组织细胞（骨细胞和软骨细胞）的细胞特异性功能（增殖和分化）的调控作用。另一方面，在含有羧甲基化多糖（OPEC）的PEG包被的培养皿上培养的细胞增殖比在TCD上培养的细胞慢，并呈现聚集体形式。钙化的初始分化标志物--ALctic活性的表达和钙的沉积主要在细胞聚集体部分观察到。成骨细胞分化标志物（骨钙素和骨桥蛋白）mRNA的表达在OPEC上早于SPEC和TCD。软骨细胞（ATDC 5）在SPEC上生长良好，与TCD相似，呈铺展状。另一方面，ATDC 5在OPEC上的生长受到显著抑制，并表现出细胞聚集体的形式。这些聚集体的尺寸随着时间的推移而变大。这些现象与成骨细胞在这些佩奇上的培养是一致的。阴离子多糖能促进软骨细胞基质的分泌和Ⅱ型胶原mRNA的表达，提示阴离子多糖的化学结构主要控制细胞的增殖和分化;羧酸残基的存在诱导细胞分化，磺酸残基促进细胞增殖。总之，多糖型PEC是用于硬组织细胞培养的优良生物材料。

In this investigation, we have examined the control of cell specific functions (proliferation and differentiation) of the hard tissue cells (bone and chondrocyte) by polyelectrolyte complex (PEC), which are composed of polysaccharides.Osteoblasts cultured on PEC containing sulfated polysaccharide (SPEC) were proliferated similar to tissue culture dish (TCD) and exhibited spreading form. On the other hand, the proliferation of cell cultured on PEG containing carboxymethylated polysaccharide (OPEC) coated dishes was slower than those on TCD and exhibited aggregate form. Expression of ALPase activity and deposition of calcium, which were initial differentiation markers of calcification, were observed mainly at the part of cell aggregate. Further, expression of mRNA of osteoblastic differentiation markers (osteocalcin and osteopontin) were observed on OPEC earlier than on SPEC and TCD. N-cadherin, cell-cell adhesion protein, expression was strong and localized at the cell-cell interfaces in the cell aggregates.Chondrocyte (ATDC5) cultured on SPEC grew well in the similar manner to TCD with spreading form. On the other hand, the growth of ATDC5 on OPEC was remarkably suppressed and exhibited cell aggregate form. The sizes of these aggregates were made bigger with time. These phenomena are well coincident with the culture of osteoblasts on these PECs. Secretion of chondrocyte matrix and expression of mRNA of typeII collagen were promoted on the OPEC.These results suggested that the cell functions, proliferation and differentiation, are mainly controlled by the chemical structures of anionic polysaccharides ; the existence of carboxylic acid residues induces the differentiation and sulfonic acid residues promotes the proliferation of cells.In conclusion, polysaccharides-type PEC are excellent biomaterials for hard tissue cell culture.