| Articular cartilage is hyaline cartilage tissue that contains no blood supply and nerve tissue. It is this lack of neurovascular supply that prevents damaged cartilage repairing by itself. Recent research efforts have focused on tissue engineering as a promising approach for cartilage regeneration and repair. Mesenchymal stem cell (MSC) and chondrocytes are the two main cells in cartilage tissue engineering. Currently,it has been reported that MSC resides in peripheral blood and can be isolated and culture in vitro, but little has been reported that peripheral blood derived MSC was used on repairing articular cartilage defect on the domestic and oversea. During the past decade, intraarticular injection of hyaluronate has been used in the treatment of patients with osteoarthritis in the knees.It can lubricate joint and nourish articular cartilage, but literatures with regard to whether hyaluronate sodium (HS) can promoted the repaire of acute articular cartilage defect or not are seldom reported. Our study observed the promotion on repairing articular cartilage defect in rabbits with the application of peripheral blood derived MSC and HS or only using hyaluronate sodium. Objective: Observing the effect on repairing articular cartilage defect in rabbits with the application of cell suspension mixed with peripheral blood derived MSC and HS, and the promotion for cartilage repair with the application of hyalurona- te sodium in vivo.We hope providing a new approach to heal the articular cartilage injure.Methods: Twenty-three healthy New Zealand White rabbits were selectd in all(8 rabbits was 2 months and 15 rabbits was 3 months ).Eight two-month aged rabbits selected randomly from them were used to mobilize peripheral blood by applying SCF and G-CSF. Then, MSCs were separated from their blood and cultured in vitro, including primary and passage culture. After passage cells were identified by histologically observation and flow cytometer, the P3 generation cells were collected and freezed.Fifteent three-month aged rabbits selected randomly were used to study the effect of cartilage repair.The full-thickness cartilage defects that the diameter is 4mm and the depth is 3mm were created on both medial and lateral femoral condyles in every knee joint of all rabbits. Hyaluronate sodium served as carrying agent material was mixed with MSC to make cell suspension.Ten rabbits were selected randomly from the fifteen's. MSC cell suspension was injected into their left knee joints and hyaluronate sodium into the right. Both knee joints of the other five rabbits were injected normal saline. All rabbits received intra-articular injections (0.5 ml) once a week for 3 weeks in every knee joint, started at one week after injury.12 weeks later after injection,all rabbits were killed to observe the state of repairing in the defects,the type of repair tissue and the articular immune response .Results1 The primary cultured MSC cells floated in the culture medium in the first 3 days,and little MSCs attached. Then the attached cells began to increase, and fusiform shape cells elongated gradually.About 10 days cell clones appeared, but the size was not uniformity. Subsquently, clones expanded more obviously,and cell's growth begin to enter the faster stage. About 25 days, attached cells can confluen for 80% on the bottom of culture flask. Passage cells would attach after 48 to 72 hours, and growth velocity is faster than primary cells. These cells can passage again during 16 to 20 days. After this, the purity of next two generations would rise. The achievement ratio of primary culture is 73.53% and survival rate of first generation passage cell is 72.0%.2 MSC HE staining indicated : MSCs showed long fusiform shape and some had ecphyma, minority was polygon. The cell nucleus which shows blue was similar to round and situated the center of cell body; endochylema was salmon pink. Purity quotient of P3 generation tested by flow cytometer was over 87%.3 After the full-thickness cartilage defects were created, rabbits' knee joints had a little swell and redness in the first 3 days. Then swell lessened gradully. Knee joints were free from infection. After injection, no inflammatory reaction and infection were seen.4 Twelve weeks later after injection, defcets were stuffed full by repaired tissue which was made of chondroid tissue histologically in experimental group. The surface in defects was smooth, but the boundary between repair tissue and surrounding could been seen. And the superficial layer's fibrosis was more obviously. In the HS control, defect filling was incomplete and the repair tissue showed severe surface fibrillation and the surface was lack of smooth. The repair cells were more similar to fibrocyte. In normal saline control, defect filling which was made of fibrous tissue was incomplete obviously.5 Effcet of articular cartilage repair of experimental group is superior to the HS contro(lt'=11.81, P<0.01)and the normal saline control ( t'=12.60, P<0.01 ) .But no significant differences were seen in the quantity or quality of the repair tissue between the two control groups(t'=1.95, P>0.05).Conclusion1 Intra-articular injection of cell suspension mixed with peripheral blood derived MSCs and hyaluronate sodium was able to improve the quality of the repair tissue which was chondroid tissue in the articular cartilage defects.But the superficial layer's degeneration is more obvious.2 Xenoma peripheral blood derived MSCs could escape body immune system, avoiding articular rejection;In addition, chondroid tissue was not been seen in synovial membrane within short term. Therefore, it was relatively safety.3 No significant differences were seen in the quantity or quality of the repair tissue between the HS control and the normal saline control,so hyaluronate sodium did not seem to promote improving quality of repair tissue in the treatment of acute rabbit full-thickness cartilage injuries in rabbits by intra-articular injection of hyaluronate sodium simply.4 Compared with bone mesenchymal stem cell, peripheral blood derived MSCs had the advantages of tiny trauma and acquisition conveniently. Furthermore, it could be satisfied with the demand of cartilage repair in quantity and quality. |
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