表面修饰化明胶海绵支架与颌下腺种子细胞复合培养的研究术

目的应用表面修饰技术处理的明胶海绵与颌下腺种子细胞复合培养，研究表面修饰技术对颌下腺种子细胞在支架材料上附着、生长和分泌功能的影响。方法选用明胶海绵36块，随机分A、B、C3组，每组12块。分别用单纯明胶海绵、层粘连蛋白表面修饰的明胶海绵材料、转化生长因子B3表面修饰的明胶海绵材料与鼠颌下腺细胞在体外复合培养。培养后第3、7、15天行组织学观察、扫描电镜观察，第15天行免疫组织化学鉴定细胞来源，测定上清淀粉酶含量。结果组织学和扫描电镜观察可见培养后第3天各组细胞分散于材料表面，无细胞突起形成；第7天B组细胞数量明显多于A组和c组，细胞突起形成并锚定于胶原海绵表面；第15天B组细胞数量仍多于A组和C组，并可见细胞之间有触突联系和腺管样结构形成。免疫组织化学染色观察复合培养的颌下腺细胞BrdU呈强阳性。随着接种后培养时问的延长，各组颌下腺细胞分泌的淀粉酶含量均有不同程度的增加。第15天C组淀粉酶含量明显高于A组和B组（P〈0．05）。结论应用层粘连蛋白和转化生长因子β3对明胶海绵进行表面修饰，可促进颌下腺种子细胞在材料上的附着、增殖和分泌功能。

Experiment Research of Tissue Engineering Submandibular Gland Cells Growing on Collagen Sponge Scaffold with Surface Modification Technique

Objective To reconstitute tissue engineering submandibular gland by implant(rat submandibular gland cells,RSGCs) combined with collagen sponge scaffold used surface modification technique.Methods Thirty-six collagen sponges were divided into three groups according surface modify material(group A: blank control;group B: laminin surface modified collagen sponges;group C: TGF-β3 surface modified collagen sponges).RSGCs labeled by BrdU were seeded on collagen sponges(1 cm×1 cm×1 cm).At 3,7 and 15 days post-culture,HE and BrdU immunohistochemical examined the cultured cells.Scanning electron microscope was used to observe the growth behavior of RSGCs on collagen sponge scaffolds.Supernatant amylase was examined by conventional clinical chemistry analyses.Results Observation under histology and scanning electron microscope showed that RSGCs start to adhere on collagen fiber in group B at 7th day.At the 15th day,RSGCs can keep proliferating and differentiating on collagen fiber and form functional lumen unit in group B.In group C,the number of RSGCs was similar to group A,but less than group B.The amount of amylase secreted increased at a different degree with an extension of the culturing time in all the groups.At the 15th day after culture,the amount of amylase in group C was higher than the other two groups.Conclusion Collagen sponge scaffold with laminin and TGF-β3 surface modified can stimulate RSGCs adhere,proliferate and differentiate on collagen fiber in vivo.