CAJ | 학술논문

目的以Ⅱ型胶原水凝胶为支架、兔骨髓基质干细胞(BMSCs)为种子细胞构建可注射型组织工程软骨,探讨其在膝关节腔内成软骨的情况. 方法以兔BMSCs为种子细胞,4℃下按105/mL的浓度接种到Ⅱ型胶原水凝胶,倒置相差显微镜下观察接种的细胞形态,并计算细胞存活率.选取2个月龄新西兰大白兔12只,随机分为2组(n=6):A组在兔左膝关节腔内植入装载Ⅱ型胶原水凝胶-BMSCs复合物的扩散盒,B组在兔左膝关节腔内植入装载单纯Ⅱ型胶原水凝胶的扩散盒.术后4周行左膝关节MRI检查;术后8周处死动物,通过组织学观察扩散盒中复合物在膝关节腔内的转归情况.结果 Ⅱ型胶原水凝胶-兔BMSCs复合物体外培养1h后细胞呈规则圆形,分布均匀,胞浆染成棕黑色;细胞存活率在95％以上.术后4周A组与B组MRI检查均可见游离的扩散盒影像位于外侧间室后方,形状完整无破损.术后8周A组HE染色见细胞数量较多,分布均匀,大部分呈圆形或椭圆形,胞核深染,基质呈均一的粉红色;蕃红O染色和甲苯胺蓝染色可见大量圆形、椭圆形细胞分布,周围基质呈中到强阳性.术后8周B组HE染色呈弱阳性粉红色云雾状,其中无清晰可辨的细胞结构;蕃红O染色及甲苯胺蓝染色均呈阴性. 结论 Ⅱ型胶原水凝胶-BMSCs复合物能够在体生成透明软骨样组织,为其下一步在体修复大动物关节软骨缺损奠定了实验基础.
목적 이Ⅱ형효원수응효위지가、토골수기질간세포(BMSCs)위충자세포구건가주사형조직공정연골,탐토기재슬관절강내성연골적정황. 방법 이토BMSCs위충자세포,4℃하안105/mL적농도접충도Ⅱ형효원수응효,도치상차현미경하관찰접충적세포형태,병계산세포존활솔.선취2개월령신서란대백토12지,수궤분위2조(n=6):A조재토좌슬관절강내식입장재Ⅱ형효원수응효-BMSCs복합물적확산합,B조재토좌슬관절강내식입장재단순Ⅱ형효원수응효적확산합.술후4주행좌슬관절MRI검사;술후8주처사동물,통과조직학관찰확산합중복합물재슬관절강내적전귀정황.결과 Ⅱ형효원수응효-토BMSCs복합물체외배양1h후세포정규칙원형,분포균균,포장염성종흑색;세포존활솔재95％이상.술후4주A조여B조MRI검사균가견유리적확산합영상위우외측간실후방,형상완정무파손.술후8주A조HE염색견세포수량교다,분포균균,대부분정원형혹타원형,포핵심염,기질정균일적분홍색;번홍O염색화갑분알람염색가견대량원형、타원형세포분포,주위기질정중도강양성.술후8주B조HE염색정약양성분홍색운무상,기중무청석가변적세포결구;번홍O염색급갑분알람염색균정음성. 결론 Ⅱ형효원수응효-BMSCs복합물능구재체생성투명연골양조직,위기하일보재체수복대동물관절연골결손전정료실험기출.
Objective To study the chondrification of the injectable tissue-engineered cartilage,which was composed of collagen type Ⅱ hydrogel as the scaffold and bone marrow stromal cells (BMSCs) as seed cells,in the knee joint cavity in rabbits.Methods The rabbit BMSCs were seeded into collagen type Ⅱ hydrogel at 4 ℃ at a concentration of 105/mL.Inverted phase contrast microscopy was used to observe the morphology and survival of the seeded cells.Twelve two-month-old New Zealand rabbits (12 left knees) were randomly divided into 2 groups (n =6).In group A,diffusion chambers containing the hydrogel-MSCs composite were placed into the left knee joint cavity while diffusion chambers containing only the hydrogel were placed into the left knee joint cavity in group B.Four weeks after surgery,the animals were subjected to MRI examinations of the left knee; 8 weeks after surgery,they were sacrificed for observation of the chondrification of the injectable engineered cartilage in vivo.Results After in vitro culture for one hour,the cells in the hydrogel-MSCs composite appeared regularly round and evenly distributed,with dark brown cytoplasm and survival of above 95％.At 4 weeks after surgery,MRI revealed images of the free intact diffusion chambers at the posterior part of the lateral compartment in both groups.At 8 weeks in group A,HE staining showed numerous evenly distributed cells most of which were round or oval,with deep stained nuclei and uniform pink matrix; safranin O and toluidine blue staining showed distribution of numerous round and oval cells and moderate to strong positivity of surrounding matrix.At 8 weeks in group B,HE staining showed weak positive pink clouds without distinct cellular structure; safranin O and toluidine blue staining was negative.Conclusion As the composite of collagen type Ⅱ hydrogel/BMSCs can grow into hyaline chondroid tissue in vivo,it can be used in animal experiments on repairing large articular cartilage defects.