Objective To explore the methods of repairing cartilagedefects and to introduce the clinical experience with the autologous osteochondral transplantation. Methods Twenty-five patients with chondral and osteochondral defects of the weight-bearing surfaces were treated by the autologous osteochondral transplantation for the repair of the chondral and osteochondral defects of the unweightbearing surfaces under arthroscope. According to the shape of the defects, the different dimensions of the osteochondral autograft were selected. All the patients began the training of the continuous passive motion after operation. Six weeks after operation, the patients began to walk in the weightbearing habitus. However, in the control group, another 25 patients were retrospectively analyzed, who had chondral and osteochondral defects of the weight-bearing surfaces but were treated only by the cleaning and drilling procedures. The scores evaluated bythe Brittberg-Peterson scoring scale of the 2 group were 98.65±9.87 and 96.98±8.94 respectively. Results The follow-upfor 3-24 months after operation revealed that the treated knee joint had a goodmotion extent. The pain was obviously alleviated. Based on the longitudinal study with the three-dimensional spoiled magnetic resonance imaging (MRI), the signal intensity of the repaired tissues approached to the normal condition. The scores evaluated by the Brittberg-Peterson scoring scale were almost zero 3 monthsafter operation in the experimental group, and the scores were 58.48±6.98 inthe control group. There were significant differences between the experimental group and the control group(P＜0.01). Conclusion Autologous osteochondral transplanation under arthroscope is a good curative method for the cartilage defects, with advantages of minimal invasiveness and avoidanceofrejections resulting from allografts. However, its long-term effect needs to befurther studied. The conventional therapies including cleaning and drilling are useful in alleviating the symptoms.
Objective To explore the relationship of the limited resource of the autologous bone marrow mesenchymal stem cells (MSCs) in articularcavity to the treatment results of full-thickness articular cartilage defect, and to investigate whether the extrogenous sodium hyaluronate(SH) promotes the migration of MSCs cultured in vitro tothe articular defect in vivo. Methods Sixty-six Japan rabbits were made the model of the full-thickness articular cartilage defect (5 mm width and 4 mm depth).The autologous MSCs were extracted from the rabbit femur, cultured in vitro, labeledby Brdu, and injected into the injured articular cavity with or without SH. Theexperiment was divided into 4 groups; group A (MSCs and SH, n=15); group B (MSCs, n=15); group C (SH, n=18); and group D (non-treatment, n=18). The morphologic observation was made by HE staining, Mallory staining and immunohistochemical staining after 5 weeks, 8 weeks and 12 weeks of operation. Results There were significant differences in the thickness of repairing tissue between group A and group B(Plt;0.01); but there were no significant differences between group A and group C, and between group B and group D(P＞0.05). Thehistological observation showed that the main repairing tissue was fibrocartilage in group A and fiber tissue in group B. Conclusion MSCs cultured in vitro and injected into the articular cavity can not improve the treatment results of the articular cartilage defect. Extrogenous SH has effect on repairing cartilage defect. The extrogenous SH has no effect on the chemotaxis of the MSCs, and on the collection of MSCs into the joint defect.
Objective To study the biological characteristic and potential of chondrocytes grafting cultured on fascia in repairing large defect of articular cartilage in rabbits. Methods Chondrocytes of young rabbits were isolated and subcultured on fascia. The large defect of articular cartilage was repaired by grafts of freeze-preserved and fresh chondrocytes cultured on fascia, and free chondrocytes respectively; the biological characteristic and metabolism were evaluated bymacroscopic, histological and immunohistochemical observations, autoradiography method and the measurement of nitric oxide content 6, 12, 24 weeks after grafting. Results The chondrocytes cultured on fascia maintained normal growth feature and metabolism, and there was no damage to chondrocytes after cryopreservation; the repaired cartilage was similar to the normal cartilage in cellular morphology and biological characteristics. Conclusion Chondrocytes could be cultured normally on fascia, which could be used as an ideal carrier of chondrocytes.
Objective To investigate the clinical application of periosteal autograft in repair of cartilage defect caused by osteoarthritis of knee. Methods From 1996 to 1999, 36 knees of cartilage defect of knee joint in 28 cases were treated. In the operation, the cracked degenerative cartilage was removed before free periosteum from tibia was transplanted to repair the defect, and the meniscuses in 8 knees of the 36 knees were reconstructed. After operation, early continuous passive movement was adopted for 4 weeks, and 8 knees with reconstruction ofthe meniscus were immobilized by plaster splint for 7 days after operation and before passive movement. All of the cases were followed up for 1 to 4 years before clinical evaluation in symptoms, signs and radiological findings. Results The general satisfactory rate was 86.1%, in which the function was excellent in 22 knees and good in 9 knees. Conclusion The periosteal autograft is a good choice for repairing cartilage defect due to osteoarthritis, with a satisfactory outcomein the short term.
Objective To investigate the effect of homograft of marrow mesenchymal stem cells (MSCs) seeded onto poly-L-lactic acid (PLLA)/gelatin on repair of articular cartilage defects. Methods The MSCs derived from36 Qingzilan rabbits, aging 4 to 6 months and weighed 2.5-3.5 kg were cultured in vitroand seeded onto PLLA/gelatin. The MSCs/ PLLA/gelatin composite was cultured and transplanted into full thickness defects on intercondylar fossa. Thirty-six healthy Qingzilan rabbits were made models of cartilage defects in the intercondylar fossa. These rabbits were divided into 3 groups according to the repair materials with 12 in each group: group A, MSCs and PLLA/gelatin complex(MSCs/ PLLA/gelatin); group B, only PLLA/gelatin; and group C, nothing. At 4,8 and 12 weeks after operation, the gross, histological and immunohistochemical observations were made, and grading scales were evaluated. Results At 12 weeks after transplantation, defect was repaired and the structures of the cartilage surface and normal cartilage was in integrity. The defects in group A were repaired by the hylinelike tissue and defects in groups B and C were repaired by the fibrous tissues. Immunohistochemical staining showed that cells in the zones of repaired tissues were larger in size, arranged columnedly, riched in collagen Ⅱ matrix and integrated satisfactorily with native adjacent cartilages and subchondral bones in group A at 12 weeks postoperatively. In gross score, group A(2.75±0.89) was significantly better than group B (4.88±1.25) and group C (7.38±1.18) 12 weeks afteroperation, showing significant differences (P＜0.05); in histological score, group A (3.88±1.36) was better than group B (8.38±1.06) and group C (13.13±1.96), and group B was better than group C, showing significant differences (P＜0.05). Conclusion Transplantation of mesenchymal stem cells seeded onto PLLA/gelatin is a promising way for the treatment of cartilage defects.
ObjectiveTo study the effect and feasibility of poly-lactide-co-glycolide (PLGA) loaded with recombinant human bone morphogenetic protein 2 (rhBMP-2) on repairing articular cartilage defect in rabbits. Methods PLGA was made into cylinders which were 4 mm in diameter and 3 mm in thickness. rhBMP-2 was fully homogenated before used. PLGA combined with 0.5 mg rhBMP-2 under the condition of vacuum(700 mmHg),and then lyophilized, packed ,sterilized with ethylene oxide and reserved. Defects of 4 mm in diameter and reaching medullary cavity were made in femoral condyles of 72 two-month-old New Zealand white rabbits. The 36 right defects were repaired with PLGA-rhBMP-2 composites as the experimental group, the 36 left defects with PLGA only as PLGA group, the other 36 left defects were left untreated as control group, and the other 36 right defects with PLGA-MSCs composites as cell group. At 4, 8, 12, 24, 36 and 48 weeks after operation, macroscopical and microscopical observations were made, and the histological grade wasdone.Results After 4 weeks of operation: In the experimental group and cell group, defects were filled with white translucent tissue which appeared smooth and soft; the matrix around chondrocytes was weakly metachromatic, the newly formed cartilage tissue was thicker than normal cartilage tissue; there was no formed tissue in the PLGA group and the blank control group. After 8 weeks of operation: In the experimental group and cell group, the new tissue was white, translucent, tenacious and smooth. The boundary with normal cartilage became vague. New cartilage cells distributed evenly. The cells of the surface layerparalleled, but the deeper layer lost directivity. The matrix dyed weakly. The new cartilage gradually became thinner, but it still thicker than the normal cartilage ones. The PLGA degraded besides some drops.In the blank control group and PLGA group, a little white membrane formed at the bottom of the defect. After 1224 weeks of operation: In the experimental group and cell group, defects were filled with new tissues which were white, translucent, tenacious and smooth. The boundary disappeared.The thickness of the new cartilage was similar to that of the normal ones. The cells of the surface layer paralleled to each other,but the cells of the deeper layer tended to arrange vertically. The matrix around chondrocytes was metachromatic,but the color was lighter than that of the normal cartilage. Bone under the cartilage and the tide mark recovered. The new cartilage linked with nomal cartilage finely.In the blank control group and PLGA group, there was a little fibrous tissue at the bottom of the defect withe obvious boundary. After 36 weeks and 48 weeks of operation：in the experimental group and the cell group, the new cartilage was slightly white，continuous and less smooth. The boundary disappeared. There was no proliferated synovial membrane.The thickenss of the new cartilage was thinner than that of the normal ones. The matrix around chondrocytes was weakly metachromatic. In the blank control group and PLGA group, the defect still existed, but became smaller.At the bottom of the defect, fibrous tissues formed. Some cartilage denudated and became less smooth.Some bone under cartilage exposed,and the synovial membrane became thick. The histologic grade of the repair tissue at 12 weeks and 24 weeks of operation in experimental group and cell group was significantly different from that at 4, 8 and 48 weeks of operation(Plt;0.01). There was also significant difference in the experimental group and cell group compared with the blank control group and PLGA group at each time after operation(Plt;0.01). But there was no significant difference between the experimental group and the cell group. Conclusion In the course of degradation。。。。。。.
ObjectiveTo investigate the effect of phosphorylatable short peptide (pSP) conjugated chitosan (CS) (pSP-CS) mediated insul in-l ike growth factor 1 (IGF-1) gene and human interleukin 1 receptor antagonist (IL-1Ra) gene local transfection on the repair of articular cartilage defect. MethodsCo-expression plasmid pBudCE4.1-IL-1Ra+IGF-1, single gene expression plasmid pBudCE4.1-IL-1Ra and pBudCE4.1-IGF-1 were constructed and combined with pSP-CS to form pSP-CS/ pDNA complexes. Thirty 3-month-old healthy male New Zealand white rabbits, weighing 2.0-2.5 kg, double legs were randomly divided into 5 groups (n=12). Lateral femoral condyle articular surface was only exposed in sham-operated group (group A); full-thickness cartilage defects were created in the articular surface of the lateral femoral condyle of the knee in 4 intervention groups: pSP-CS/pBudCE4.1 (group B), pSP-CS/pBudCE4.1-IL-1Ra (group C), pSP-CS/pBudCE4.1-IGF-1(group D), and pSPCS/ pBudCE4.1-IL-1Ra+IGF-1 (group E). At 1 week after operation, intra-articular injection of pSP-CS/pDNA complexes was administrated 2 times a week for 7 weeks in each intervention group, the same volume normal sal ine in group A. The general condition of animal was observed after operation, and rabbits were sacrificed at 8 weeks. Knee joint synovial fluid was collected to measure the concentrations of the IL-1Ra and IGF-1 by ELISA; mRNA expressions of Aggrecan, matrix metalloproteinase 3 (MMP-3), and MMP inhibitor 1 (TIMP-1) were detected by real-time fluorescent quantitative PCR; the chondrogenic phenotype of nascent cells in the damage zone was identified by alcian blue-periodic acid/schiff (AB-PAS) histochemistry and Aggrecan immunohistochemistry staining. ResultsThirty experimental rabbits all survived to the end of experiment, without infection and death. Large amounts of exogenous proteins of IGF-1 and IL-1Ra were detected in the synovial fluid of 4 intervention groups. There were significant differences between groups D, E and group A in IGF-1 protein expression, and between goups C, E and group A in IL-1Ra protein expression (P < 0.05). Aggrecan and TIMP-1 mRNA expressions were significantly up-regulated in group E, simultaneously MMP-3 mRNA expression was significantly down-regulated when compared with groups C and D (P < 0.05). Varying degrees of cartilage repair appeared in groups C, D, and E, showing positive staining of AB-PAS and Aggrecan, and group E had better results than groups C and D (P < 0.05); inflammatory cell infiltration and fibrous tissue prol iferation were seen in the defect region of group B, without significant cartilage repairing. ConclusionpSP-CS is an ideal gene del ivery system for cartilage defect gene therapy; IL-1Ra and IGF-1 double gene transfection has better biologic effect on cartilage defect repair.
Objective To investigate the effect of allogeneic chondrocytes-calcium alginate gel composite under the intervention of low intensive pulsed ultrasound (LIPUS) for repairing rabbit articular cartilage defects. Methods Bilateral knee articular cartilage were harvested from 8 2-week-old New Zealand white rabbits to separate the chondrocytes by mechanical-collagen type II enzyme digestion. The 3rd passage chondrocytes were diluted by 1.2% sodium alginate to 5 × 106 cells/mL, then mixed with CaCl2 solution to prepare chondrocytes-calcium alginate gel composite, which was treated with LIPUS for 3 days (F0: 1 MHz; PRF: 1 kHz; Amp: 60 mW/cm2; Cycle: 50; Time: 20 minutes). An articular cartilage defect of 3 mm in diameter and 3 mm in thickness was established in both knees of 18 New Zealand white rabbits (aged 28-35 weeks; weighing, 2.1-2.8 kg), and divided into 3 groups randomly, 6 rabbits in each group: LIPUS group, common group, and model group. Defect was repaired with LIPUS-intervention gel composite, non LIPUS-intervention gel composite in LIPUS group and common group, respectively; defect was not treated in the model group. The general condition of rabbits was observed after operation. The repair effect was evaluated by gross and histological observations, immunohistochemical staining, and Wakitani score at 8 and 12 weeks after operation. Results Defect was filled with hyaline chondroid tissue and white chondroid tissue in LIPUS and common groups, respectively. LIPUS group was better than common group in the surface smooth degree and the degree of integration with surrounding tissue. Defect was repaired slowly, and the new tissue had poor elasticity in model group. Histological observation and Wakitani score showed that LIPUS group had better repair than common group at 8 and 12 weeks after operation; the repair effect of the 2 groups was significantly better than that of model group (P lt; 0.05); and significant differences in repair effect were found between at 8 and 12 weeks in LIPUS and common groups (P lt; 0.05). The collagen type II positive expression area and absorbance (A) value of LIPUS and common groups were significantly higher than those of model group (P lt; 0.05) at 8 and 12 weeks after operation, and the expression of LIPUS group was superior to that of common group at 12 weeks (P lt; 0.05); and significant differences were found between at 8 and 12 weeks in LIPUS group (P lt; 0.05), but no significant difference between 2 time points in common and model groups (P gt; 0.05). Conclusion Allogeneic chondrocytes-calcium alginate gel composite can effectively repair articular cartilage defect. The effect of LIPUS optimized allogeneic chondrocytes-calcium alginate gel composite is better.
【Abstract】 Objective To compare the effect of PLGA and collagen sponge combined with rhBMP-2 on repairing ofarticular cartilage defect in rabbits respectively. Methods PLGA and collagen sponge were made into cyl inders which were 4 mm in diameter and 3 mm in thickness, and compounded with rhBMP-2 (0.5 mg). Defect 4 mm in diameter were made in both of femoral condyles of 24 two-month-old New Zealand white rabbits. The defects in right 18 knees were treated with PLGA/rhBMP-2 composites (experimental group 1), and the left 18 knees were treated with collagen sponge/rhBMP-2 composites (experimental group 2), the other 12 knees were left untreated as control group. At 4, 12 and 24 weeks after operation, the animals were sacrificed and the newly formed tissues were observed macroscopically and microscopically, graded histologically and analyzed statistically. Results From the results of macroscopical and microscopical observation, in the experimental group 1, the defects were filled with smooth and translucent cartilage; while in the experimental group 2, the white translucent tissues did notfill the defects completely; and in the two experimental groups, the new cartilage tissues demarcated from the surrounding cartilage,chondrocytes distributed uniformly but without direction; a l ittle fibrous tissue formed in the control group 4 weeks postoperatively. In the experimental group 1, the defects were filled completely with white, smooth and translucent cartilage tissue without clear l imit with normal cartilage; while in the experimental group 2, white translucent tissues formed, the boundary still could be recognized; in the two experimental groups, the thickness was similar to that of the normal cartilage; the cells paralleled to articular surface in the surface layer, but in the deep layer, the cells distributed confusedly, the staining of matrix was positive but a l ittle weak; subchondral bone and tide mark recovered and the new tissue finely incorporated with normal cartilage;however, in the control group, there was a l ittle of discontinuous fibrous tissue, chondrocytes maldistributed in the border andthe bottom of the defects 12 weeks postoperatively. In the experimental group 1, white translucent cartilage tissues formed, the boundary disappeared; in the experimental group 2, the color and the qual ity of new cartilage were similar to those of 12 weeks; in the two experimental groups, the thickness of the new cartilage, which appeared smooth, was similar to that of the normal cartilage, the chondrocytes arranged uniformly but confusedly; the staining of matrix was positive and subchondral bone and tide mark recovered, the new tissue finely incorporated with normal cartilage; in the control group, a layer of discontinuous fibrous tissue formed in the bottom of the defects 24 weeks postoperatively. Results of histological grade showed that there were significantdifference between experimental group (1 and 2) and control group at any time point (P lt; 0.01); the scores of 12 weeks and 24 weeks in experimental group 1 and 2 had a significant difference compared with that of 4 weeks (P lt; 0.01), there was no significant difference between 12 weeks and 24 weeks (P gt; 0.05), and there were no significant difference between the two experimental groups at the same time point (P gt; 0.05). Conclusion Both PLGA and collagen sponge as a carrier compounded with rhBMP-2 can repair articular cartilage defects.
Objective To evaluate the immunological reaction and the outcome of allogeneic chondrocyte transplantation in repairing articular cartilage defects in porcins. Methods Full articular cartilage from the knee of two Shanghai white porcins about one-month-old was removed and cut mechanically, digested by 0.25% trypsin and 0.2% type Ⅱ collagenase and cultured in 10％ DMEM medium. Defects of 0.5 cm×0.5 cm involving the subchodral bone were created in both the left and right femur condyloid in 8 two-month-old Yunnai bama porcins. Allogeneic chondrocyte transplantation were implanted in defects at a density of (1.0-2.0)×106,0.2 ml. The lymphocytes from the receivers’ blood were collected before transplantation and after 3, 5, 7 and 12 weeks of transplantation, then mixed with allogeneic chondrocytes to determin the lymphocyte stimulation index(SI) in vitro. The histological observation in vivo was made after 5, 7 and 24 weeks of transplantation. Results Lymphocyte SI at 3, 5, 7 and 12 weeks(1.457±0.062,1.739±0.142,1.548±0.047，1.216±0.028) after transplantation was higher than that before transplantation(1.102±0.034，Plt;0.05). SI began to increase in the 3rd week and reached the peak value in the 5th week, then gradually declined at the 7th and 12th weeks, showing significant differences when compared with in the 5th week (Plt;0.05). Inflammation and lymphocytes infiltration could be seen in subchondral bone and the intergration area between repair tissue and normal cartilage in the 5th week, and then decreased and limited in subchondral bone in the 7th week. Defects were filled with cartilage tissue, which had good intergration with subchondral bone at 24 weeks after transplantation. Conclusion Immunological reactions can be found at early stage of allogeneic chondrocyte transplantation and then decreased with the time, the fullthickness articular cartilage defects could be repaired mainlywith hyaline cartilage by the allogeneic chondrocyte transplantation. This may provide a new method to repair articular cartilage defects clinically.