EFFECT OF ALLOGENEIC CHONDROCYTES-CALCIUM ALGINATE GEL COMPOSITE UNDER INTERVENTION OF LOW INTENSIVE PULSED ULTRASOUND FOR REPAIRING RABBIT KNEE ARTICULAR CARTILAGE DEFECT_Chinese Journal of Reparative and Reconstructive Surgery

Authors：

GUO Yang ¹ , MA Yong ^1,2 , DONG Rui ¹ , LIU Shanglun ¹ , TU Juan ³

1Institute of Traumatology &;
Orthopedics, Nanjing University of Chinese Medicine, Nanjing Jiangsu, 210023, P.R.China;;
2Department of Traumatology &;
Orthopedics, Affiliated Hospital of Nanjing University of Chinese Medicine;;
3Institute of Acoustics, Nanjing University. Corresponding author: MA Yong, E-mail: zhongyi-my@263.net;

Corresponding author：

Keywords：

Tissue engineered cartilage; Chondrocytes; Calcium alginate gel; Low intensive pulsed ultrasound; Cartilage defect; Rabbit

DOI：

10.7507/1002-1892.20130204

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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.

Citation： GUO Yang,MA Yong,DONG Rui,LIU Shanglun,TU Juan. EFFECT OF ALLOGENEIC CHONDROCYTES-CALCIUM ALGINATE GEL COMPOSITE UNDER INTERVENTION OF LOW INTENSIVE PULSED ULTRASOUND FOR REPAIRING RABBIT KNEE ARTICULAR CARTILAGE DEFECT. Chinese Journal of Reparative and Reconstructive Surgery, 2013, 27(8): 928-934. doi: 10.7507/1002-1892.20130204 Copy

1.	Hunziker EB. Articular cartilage repair: basic science and clinical progress. A review of the current status and prospects. Osteoarthritis Cartilage, 2002, 10(6): 432-463.
2.	吴迪, 赵智君, 朱晓松, 等. 藻酸钙凝珠复合自体软骨细胞修复成年兔关节软骨缺损. 中国组织工程研究与临床康复, 2010, 14(21): 3811-3814.
3.	马勇, 陈金飞, 张允申, 等. 可注射性壳聚糖/β-甘油磷酸二钠凝胶复合同种异体软骨细胞修复兔膝关节软骨缺损及威灵仙的干预效应. 中国组织工程研究与临床康复, 2010, 14(16): 2864-2869.
4.	Naito K, Watari T, Muta T, et al. Low-intensity pulsed ultrasound (LIPUS) increases the articular cartilage type II collagen in a rat osteoarthritis model. J Orthop Res, 2010, 28(3): 361-369.
5.	Loyola Sánchez A, Ramirez Wakamatzu MA, Vazquez Zamudio J, et al. Effect of low-intensity pulsed ultrasound on regeneration of joint cartilagen patients with second and third degree osteoarthritis of the knee. Reumatol Clin, 2009, 5(4): 163-167.
6.	高锋. 低强度脉冲超声波促进人髓核细胞合成胞外基质功能性成分的体外实验研究. 镇江: 苏州大学, 2011.
7.	Wakitani S, Nawata M, Tensho K, et al. Repair of articular cartilage defect in the patello-femoral joint with autologous bone marrow mesenchymal cell transplantation: three case reports involving nine defects in five knees. J Tissue Eng Regen Med, 2007, 1(1): 74-79.
8.	陈康, 王太平. 组织工程软骨修复关节软骨损伤研究新进展. 中国矫形外科杂志, 2012, 20(8): 721-723.
9.	刘尚仑, 董睿, 马勇. 组织工程化软骨构建的研究进展. 中国组织工程研究, 2012, 16(46): 8716-8720.
10.	罗文, 范建楠, 叶川. 软骨组织工程学方法修复关节软骨缺损. 中国组织工程研究, 2012, 16(37): 7009-7014.
11.	Danisovic L, Varga I, Zamborsky R, et al. The tissue engineering of articular cartilage: cells, scaffolds and stimulating factors. Exp Biol Med, 2012, 237(1): 10-17.
12.	刘清宇, 王富友, 杨柳. 关节软骨组织工程支架的研究进展. 中国修复重建外科杂志, 2012, 26(10): 1247-1250.
13.	Wang CC, Yang KC, Lin KH, et al. Cartilage regeneration in SCID mice using a highly organized three- dimensional alginate scaffold. Biomaterials, 2012, 33(1): 120-127.
14.	Tay LX, Ahmad RE, Dashtdar H, et al. Treatment outcomes of alginate-embedded allogenic mesenchymal stem cells versus autologous chondrocytes for the repair of focal articular cartilage defects in a rabbit model. Am J Sports Med, 2012, 40(1): 83-90.
15.	Dhollander AA, Verdonk PC, Lambrecht S, et al. Midterm results of the treatment of cartilage defects in the knee using alginate beads containing human mature allogenic chondrocytes. Am J Sports Med, 2012, 40(1): 75-82.
16.	蒋恺, 崔维顶, 任科伟, 等. 低强度脉冲式超声波对大鼠软骨细胞增殖和基质合成的影响. 江苏医药, 2011, 37(12): 1391-1393.
17.	Mukai S, Ito H, Nakagawa Y, et al. Transforming growth factor-beta1 mediates the effects of low-intensity pulsed ultrasound in chondrocytes. Ultrasound Med Biol, 2005, 31(12): 1713-1721.
18.	Kobayashi Y, Sakai D, Iwashina T, et al. Low-intensity pulsed ultrasound stimulates cell proliferation, proteoglycan synthesis and expression of growth factor-related genes in human nucleus pulposus cell line. Eur Cell Mater, 2009, 17: 15-22.
19.	Suzuki A, Takayama T, Suzuki N, et al. Daily low-intensity pulsed ultrasound stimulates production of bone morphogenetic protein in ROS 17/2. 8 cells. J Oral Sci, 2009, 51(1): 29-36.
20.	Li X, Li J, Cheng K, et al. Effect of low-intensity pulsed ultrasound on MMP-13 and MAPKs signaling pathway in rabbit knee osteoarthritis. Cell Biochem Biophys, 2011, 61(2): 427-434.
21.	于海生. 低强度脉冲超声促进骨髓来源细胞增殖的研究. 重庆: 重庆医科大学, 2012.
22.	高翔, 宋锦璘, 邓锋, 等. 低强度脉冲超声波联合引导骨再生术对Beagle犬牙周骨开窗缺损修复效应的研究. 中国生物医学工程学报, 2012, 31(6): 895-901.
23.	Romano CL, Romano D, Logoluso N. Low-intensity pulsed ultrasound for the treatment of bone delayed union or nonunion: a review. Ultrasound Med Biol, 2009, 35(4): 529-536.

1. Hunziker EB. Articular cartilage repair: basic science and clinical progress. A review of the current status and prospects. Osteoarthritis Cartilage, 2002, 10(6): 432-463.
2. 吴迪, 赵智君, 朱晓松, 等. 藻酸钙凝珠复合自体软骨细胞修复成年兔关节软骨缺损. 中国组织工程研究与临床康复, 2010, 14(21): 3811-3814.
3. 马勇, 陈金飞, 张允申, 等. 可注射性壳聚糖/β-甘油磷酸二钠凝胶复合同种异体软骨细胞修复兔膝关节软骨缺损及威灵仙的干预效应. 中国组织工程研究与临床康复, 2010, 14(16): 2864-2869.
4. Naito K, Watari T, Muta T, et al. Low-intensity pulsed ultrasound (LIPUS) increases the articular cartilage type II collagen in a rat osteoarthritis model. J Orthop Res, 2010, 28(3): 361-369.
5. Loyola Sánchez A, Ramirez Wakamatzu MA, Vazquez Zamudio J, et al. Effect of low-intensity pulsed ultrasound on regeneration of joint cartilagen patients with second and third degree osteoarthritis of the knee. Reumatol Clin, 2009, 5(4): 163-167.
6. 高锋. 低强度脉冲超声波促进人髓核细胞合成胞外基质功能性成分的体外实验研究. 镇江: 苏州大学, 2011.
7. Wakitani S, Nawata M, Tensho K, et al. Repair of articular cartilage defect in the patello-femoral joint with autologous bone marrow mesenchymal cell transplantation: three case reports involving nine defects in five knees. J Tissue Eng Regen Med, 2007, 1(1): 74-79.
8. 陈康, 王太平. 组织工程软骨修复关节软骨损伤研究新进展. 中国矫形外科杂志, 2012, 20(8): 721-723.
9. 刘尚仑, 董睿, 马勇. 组织工程化软骨构建的研究进展. 中国组织工程研究, 2012, 16(46): 8716-8720.
10. 罗文, 范建楠, 叶川. 软骨组织工程学方法修复关节软骨缺损. 中国组织工程研究, 2012, 16(37): 7009-7014.
11. Danisovic L, Varga I, Zamborsky R, et al. The tissue engineering of articular cartilage: cells, scaffolds and stimulating factors. Exp Biol Med, 2012, 237(1): 10-17.
12. 刘清宇, 王富友, 杨柳. 关节软骨组织工程支架的研究进展. 中国修复重建外科杂志, 2012, 26(10): 1247-1250.
13. Wang CC, Yang KC, Lin KH, et al. Cartilage regeneration in SCID mice using a highly organized three- dimensional alginate scaffold. Biomaterials, 2012, 33(1): 120-127.
14. Tay LX, Ahmad RE, Dashtdar H, et al. Treatment outcomes of alginate-embedded allogenic mesenchymal stem cells versus autologous chondrocytes for the repair of focal articular cartilage defects in a rabbit model. Am J Sports Med, 2012, 40(1): 83-90.
15. Dhollander AA, Verdonk PC, Lambrecht S, et al. Midterm results of the treatment of cartilage defects in the knee using alginate beads containing human mature allogenic chondrocytes. Am J Sports Med, 2012, 40(1): 75-82.
16. 蒋恺, 崔维顶, 任科伟, 等. 低强度脉冲式超声波对大鼠软骨细胞增殖和基质合成的影响. 江苏医药, 2011, 37(12): 1391-1393.
17. Mukai S, Ito H, Nakagawa Y, et al. Transforming growth factor-beta1 mediates the effects of low-intensity pulsed ultrasound in chondrocytes. Ultrasound Med Biol, 2005, 31(12): 1713-1721.
18. Kobayashi Y, Sakai D, Iwashina T, et al. Low-intensity pulsed ultrasound stimulates cell proliferation, proteoglycan synthesis and expression of growth factor-related genes in human nucleus pulposus cell line. Eur Cell Mater, 2009, 17: 15-22.
19. Suzuki A, Takayama T, Suzuki N, et al. Daily low-intensity pulsed ultrasound stimulates production of bone morphogenetic protein in ROS 17/2. 8 cells. J Oral Sci, 2009, 51(1): 29-36.
20. Li X, Li J, Cheng K, et al. Effect of low-intensity pulsed ultrasound on MMP-13 and MAPKs signaling pathway in rabbit knee osteoarthritis. Cell Biochem Biophys, 2011, 61(2): 427-434.
21. 于海生. 低强度脉冲超声促进骨髓来源细胞增殖的研究. 重庆: 重庆医科大学, 2012.
22. 高翔, 宋锦璘, 邓锋, 等. 低强度脉冲超声波联合引导骨再生术对Beagle犬牙周骨开窗缺损修复效应的研究. 中国生物医学工程学报, 2012, 31(6): 895-901.
23. Romano CL, Romano D, Logoluso N. Low-intensity pulsed ultrasound for the treatment of bone delayed union or nonunion: a review. Ultrasound Med Biol, 2009, 35(4): 529-536.

Chinese Journal of Reparative and Reconstructive Surgery

EFFECT OF ALLOGENEIC CHONDROCYTES-CALCIUM ALGINATE GEL COMPOSITE UNDER INTERVENTION OF LOW INTENSIVE PULSED ULTRASOUND FOR REPAIRING RABBIT KNEE ARTICULAR CARTILAGE DEFECT

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