摘要
Objective：Although chondroprotective activities have been documented for polysaccharides,the potential target of different polysaccharide may differ.The study was aimed to explore the effect of glucan HBP-A in chondrocyte monolayer culture and chondrocytes-alginate hydrogel constructs in vivo,especially on the expression of type Ⅱ collagen.Methods：Chondrocytes isolated from rabbit articular cartilage were cultured and verified by immunocytochemical staining of type Ⅱ collagen.Chondrocyte viability was assessed after being treated with HBP-A in different concentrations.Morphological status of chondrocytes-alginate hydrogel constructs in vitro was observed by scanning electron microscope（SEM）.The constructs were treated with HBP-A and then injected to nude mice subcutaneously.Six weeks after transplantation,the specimens were observed through transmission electron microscopy（TEM）.The mRNA expressions of disintegrin and metalloproteinase with thrombospondin motifs 5（ADAMTs-5）,aggrecan and type Ⅱ collagen in both monolayer culture and constructs were determined by real time polymerase chain reaction（PCR）.The expression of typeⅡ collagen and matrix metalloproteinases-3（MMP-3） in chondrocyte monolayer culture was also tested through Western blot and enzyme linked immunosorbent assay（ELISA）,respectively.Results：MMP-3 secretion and ADAMTs-5 mRNA expression in vitro were inhibited by HBP-A at 0.3 mg/mL concentration.In morphological study,there were significant appearance of collagen in those constructs treated by HBP-A.Accordingly,in both chondrocyte monolayer culture and chondrocytes-alginate hydrogel constructs,the expression of type Ⅱcollagen was increased significantly in HBP-A group when compared with control group（P〈0.001）.Conclusions：The study documented that the potential pharmacological target of glucan HBP-A in chondrocytes monolayer culture and tissue engineered cartilage in vivo may be concerned with the inhibition of catabolic enzymes MMP-3,ADAMTs-5,and increasing of type Ⅱ collagen expression.
Objective：Although chondroprotective activities have been documented for polysaccharides,the potential target of different polysaccharide may differ.The study was aimed to explore the effect of glucan HBP-A in chondrocyte monolayer culture and chondrocytes-alginate hydrogel constructs in vivo,especially on the expression of type Ⅱ collagen.Methods：Chondrocytes isolated from rabbit articular cartilage were cultured and verified by immunocytochemical staining of type Ⅱ collagen.Chondrocyte viability was assessed after being treated with HBP-A in different concentrations.Morphological status of chondrocytes-alginate hydrogel constructs in vitro was observed by scanning electron microscope（SEM）.The constructs were treated with HBP-A and then injected to nude mice subcutaneously.Six weeks after transplantation,the specimens were observed through transmission electron microscopy（TEM）.The mRNA expressions of disintegrin and metalloproteinase with thrombospondin motifs 5（ADAMTs-5）,aggrecan and type Ⅱ collagen in both monolayer culture and constructs were determined by real time polymerase chain reaction（PCR）.The expression of typeⅡ collagen and matrix metalloproteinases-3（MMP-3） in chondrocyte monolayer culture was also tested through Western blot and enzyme linked immunosorbent assay（ELISA）,respectively.Results：MMP-3 secretion and ADAMTs-5 mRNA expression in vitro were inhibited by HBP-A at 0.3 mg/mL concentration.In morphological study,there were significant appearance of collagen in those constructs treated by HBP-A.Accordingly,in both chondrocyte monolayer culture and chondrocytes-alginate hydrogel constructs,the expression of type Ⅱcollagen was increased significantly in HBP-A group when compared with control group（P〈0.001）.Conclusions：The study documented that the potential pharmacological target of glucan HBP-A in chondrocytes monolayer culture and tissue engineered cartilage in vivo may be concerned with the inhibition of catabolic enzymes MMP-3,ADAMTs-5,and increasing of type Ⅱ collagen expression.
基金
Supported by the National Natural Science Foundation of China(No.30300459,81072830)
Shanghai Leading Academic Discipline Project(No.T0303)
the Shanghai Youth Phospherus Project(No.08QA14063)
