Intra-Articular Atelocollagen Injection for the Treatment of Articular Cartilage Defects in Rabbit Model

Suh Dong Sam,Yoo Ji Chul,Woo Sang Hun,Kwak Andrew S. 한국조직공학과 재생의학회 2021 조직공학과 재생의학 Vol.18 No.4

Background: Atelocollagen is widely recognized as a biomaterial for regenerative medicine because of its good compatibility and low antigenicity. Injury of the outermost layer of articular cartilage, known as the lamina splendens, can lead to osteoarthritis (OA) and eventually full-thickness cartilage loss. The intra-articular injection of atelocollagen has been designed to restore the cartilage layer and cartilage defects in OA joints. In this study, we investigated the efficacy of atelocollagen as a cartilage supplement for joint defects. Methods: In this study, we evaluated the therapeutic effects of atelocollagen in animals with cartilage defects. Femoral groove defects were artificially created in 12 male New Zealand white rabbits, which were treated with intra-articular injection of either atelocollagen (experimental) or normal saline (control). The results were observed 3, 6, 9, and 12 weeks following macroscopic and histological evaluations. Results: At 3 weeks, cartilage tissue was restored in the experimental group, whereas the control group did not show signs of restoration. At 12 weeks, defects in both groups were filled with regenerated tissue, but the experimental group displayed a morphologically better appearance. Histologically, the regenerated tissue in the experimental group showed statistically significant improvement compared to the control group, with a structure similar to that of normal articular cartilage. Conclusion: The results showed that the intra-articular injection of atelocollagen enhanced cartilage regeneration following rabbit patellar groove defects. Therefore, intra-articular injection of atelocollagen can be used as an effective supplement for joint defects. Background: Atelocollagen is widely recognized as a biomaterial for regenerative medicine because of its good compatibility and low antigenicity. Injury of the outermost layer of articular cartilage, known as the lamina splendens, can lead to osteoarthritis (OA) and eventually full-thickness cartilage loss. The intra-articular injection of atelocollagen has been designed to restore the cartilage layer and cartilage defects in OA joints. In this study, we investigated the efficacy of atelocollagen as a cartilage supplement for joint defects. Methods: In this study, we evaluated the therapeutic effects of atelocollagen in animals with cartilage defects. Femoral groove defects were artificially created in 12 male New Zealand white rabbits, which were treated with intra-articular injection of either atelocollagen (experimental) or normal saline (control). The results were observed 3, 6, 9, and 12 weeks following macroscopic and histological evaluations. Background: Atelocollagen is widely recognized as a biomaterial for regenerative medicine because of its good compatibility and low antigenicity. Injury of the outermost layer of articular cartilage, known as the lamina splendens, can lead to osteoarthritis (OA) and eventually full-thickness cartilage loss. The intra-articular injection of atelocollagen has been designed to restore the cartilage layer and cartilage defects in OA joints. In this study, we investigated the efficacy of atelocollagen as a cartilage supplement for joint defects. Methods: In this study, we evaluated the therapeutic effects of atelocollagen in animals with cartilage defects. Femoral groove defects were artificially created in 12 male New Zealand white rabbits, which were treated with intra-articular injection of either atelocollagen (experimental) or normal saline (control). The results were observed 3, 6, 9, and 12 weeks following macroscopic and histological evaluations. Results: At 3 weeks, cartilage tissue was restored in the experimental group, whereas the control group did not show signs of restoration. At 12 weeks, defects in both groups were filled with regenerated tissue, but the experimental group displayed a morphologically better appearance. Histologically, the regenerated tissue in the experimental group showed statistically significant improvement compared to the control group, with a structure similar to that of normal articular cartilage. Conclusion: The results showed that the intra-articular injection of atelocollagen enhanced cartilage regeneration following rabbit patellar groove defects. Therefore, intra-articular injection of atelocollagen can be used as an effective supplement for joint defects. Results: At 3 weeks, cartilage tissue was restored in the experimental group, whereas the control group did not show signs of restoration. At 12 weeks, defects in both groups were filled with regenerated tissue, but the experimental group displayed a morphologically better appearance. Histologically, the regenerated tissue in the experimental group showed statistically significant improvement compared to the control group, with a structure similar to that of normal articular cartilage. Conclusion: The results showed that the intra-articular injection of atelocollagen enhanced cartilage regeneration following rabbit patellar groove defects. Therefore, intra-articular injection of atelocollagen can be used as an effective supplement for joint defects.

인체 퇴행성 슬관절염의 관절연골의 Nitrotyrosine, Fas발현과 세포자 멸사

전대철,김상우,서승석 인제대학교 백병원 2002 仁濟醫學 Vol.23 No.1

■ Purpose The purpose of this experiment is to examine whether articular cartilage degeneration in human osteoarthritis(OA) are related with the occurrence of apoptosis, the expression of nitotyrosine and Fas. ■ Materials and Methods Articular cartilage was obtained from OA patients undergoing total knee replacement arthroplasty and from normal subjects who were operated the above knee amputation. According to histologic degeneration, articular cartilage was graded to grade 1, grade 2, grade 3 and grade 4. Apoptotic cells were identified by TUNEL method and electron microscopy. Articular cartilage sections were analyzed by immunohistochemistry for the presence of nitrotyrosine and Fas expression. ■ Results The number of apoptotic cells was significantly in OA articular cartilage compared with normal articular cartilage(p<0.05). The number of apoptotic cells was increased with tissue degeneration. On electron microscopy, an apoptotic cell showed the typical chromatin condensation in the OA articular cartilage. The incidence of Fas-expressing cells was significantly higher in the OA articular cartilage(p<0.05). Nitrotyrosine immunoreactivity was prominent in the degenerative articular cartilage(p<0.05). Fas and nitrotyrosine expression was increased with tissue degeneration. The increase in a number of apoptotic cells was correlated with tissue degeneration. ■ Conclusions Apoptosis was the one of the causes in the tissue degeneration of the human OA articular cartilage. The development of apoptosis in the articular cartilage was related with Fas and nitrotyrosine expression.

가토 슬관절연골의 대기노출 시간에 따른 변성 및 재생

김익동,김풍택,박병철,최영욱,이상복 慶北大學校 醫科大學 1990 慶北醫大誌 Vol.31 No.2

Orthopedic procedures expose articular cartilage to operating room air for varying periods of time as long as two hours. Articular cartilage is dense white, glossy and smooth, and is composed of chondrocytes, collagen, and a hyaline matrix made up of proteoglycan molecules. Articular cartilage is a specialized tissue containing 75 to 85% water. Most of the water is trapped in a gel of matrix collagen and proteoglycan and is freely exchangeable with the synovial fluid and blood. Only about 6% of cartilage water is tightly bound. Theoretically, since cartilage gel is hyperhydrated, the water content is important in maintaining cartilage resiliency and lubrication of the articular surface. If a pathologic state is associated with reduction on water content, these properties may be altered, leading to impaired joint function. We reproduced the O.R. situation by exposing one knee of 24 mature New-Zealand white rabbits to ambient air (20-22℃) for 30 minutes to 2 hours, the opposite knee serving as a control. Following exposure, the joints were closed and the animal resumed cage activity for 1 day prior to sacrifice, at that time cartilage was removed from both exposed and control joints and prepared for gross, light microscopy and electron microscopy. We observed cartilages which were sacrificed at 1 week after operation, 2 weeks after and 4 weeks after for varying periods of exposing time in same method. The results were as follow; 1. A distinct color change was seen in the exposed cartilage in groups which were sacrificed postoperative at 1 day ; However minimal change was detected in each group sacrificed at 1 week & 2 weeks later, and no change was seen in each group sacrificed postoperative at 4 weeks. 2. Exposure to room air for 30 minutes produced chondrocyte necrosis in the superficial zone of rabbit articular cartilage. Exposure for 1 hour or longer produced chondrocyte necrosis up to radial zone. 3. Chondrocyte necrosis was not seen in each group for varying periods of air exposure microscopically, which sacrificed postoperative at 1 week. 4. Proteoglycan content was minimal at 1 week postoperatively in each group ; However proteoglycan content increased progressively with time. 5. Change in cellularity were consistent with cartilage damage. Seemingly, exposing articular cartilage to air for 2 hours caused reversible cartilage damage.

가토의 관절 연골 결손 부위에서 미세 골절과 골형성 단백-2가 연골의 치유에 미치는 영향

이승준,김학준,양희석,김병수,박정호 한국조직공학과 재생의학회 2009 조직공학과 재생의학 Vol.6 No.13

Articular cartilage has a limited capacity for repair. The lack of repair in partial cartilage injury may result from the avascular nature of cartilage. Several methods have been developed to accelerate cartilage healing. Microfracture is an operative method that can facilitate access to stem cells in bone marrow and stimulate cartilage regeneration. However, microfracture has been shown to result in regeneration with fibrocartilage. The bone morphogenetic protein-2 has been shown to induce the expression of cartilage and bone marker. These findings prompted us to investigate rhBMP-2’s ability to modulate the repair of full-thickness defect of articular cartilage. The full thickness articular cartilage defects of 6x3mm size were created in the trochlear groove of forty-eight rabbits. Twelve defects were left empty, and other thirty-six defects were treated with microfracture. Among those thirty-six defects, twelve defects were filled with fibrin glue, and twelve defects with fibrin glue and rhBMP-2. The animals were killed at 8 weeks after surgery. The repair tissue was examined histologically and evaluated with real time PCR to analysis collagen type. After microfracture at the cartilage defect, treatment with fibrin glue and rhBMP-2 accelerated cartilage healing and the formation of new subchondral bone, and showed more hyaline-like cartilage components than other groups on the histological findings. Real-time PCR showed higher amounts of collage type 1 and type 2 in the group filled with fibrin glue and rhBMP-2. Articular cartilage has a limited capacity for repair. The lack of repair in partial cartilage injury may result from the avascular nature of cartilage. Several methods have been developed to accelerate cartilage healing. Microfracture is an operative method that can facilitate access to stem cells in bone marrow and stimulate cartilage regeneration. However, microfracture has been shown to result in regeneration with fibrocartilage. The bone morphogenetic protein-2 has been shown to induce the expression of cartilage and bone marker. These findings prompted us to investigate rhBMP-2’s ability to modulate the repair of full-thickness defect of articular cartilage. The full thickness articular cartilage defects of 6x3mm size were created in the trochlear groove of forty-eight rabbits. Twelve defects were left empty, and other thirty-six defects were treated with microfracture. Among those thirty-six defects, twelve defects were filled with fibrin glue, and twelve defects with fibrin glue and rhBMP-2. The animals were killed at 8 weeks after surgery. The repair tissue was examined histologically and evaluated with real time PCR to analysis collagen type. After microfracture at the cartilage defect, treatment with fibrin glue and rhBMP-2 accelerated cartilage healing and the formation of new subchondral bone, and showed more hyaline-like cartilage components than other groups on the histological findings. Real-time PCR showed higher amounts of collage type 1 and type 2 in the group filled with fibrin glue and rhBMP-2.

주사전자현미경으로 관찰한 관절연골의 초미세구조

권건영,강창수,이영국 啓明大學校 醫科大學 1992 계명의대학술지 Vol.11 No.3

The authors observed morphological changes with light microscopy and scanning electron microscopy of 8 patients with osteoarthritis caused by avascular necrosis whose articular cartilage of femoral head was classified by the degree of injury. In light microscopy, the articular cartilage of stage Ⅱ patient appeared homogenous in structure, composed of a dense ground substance and chondrocytes. But in the other atient of stage Ⅱ could be visualized partial degeneration change and loss of articular surface and chondrocytes. In the case of stage Ⅲ, the chondrocyte cluster could be seen just underneath the surface which enabled us to observe and irregular articular surface. The number of chondrocytes were also increased on the superficial and middle portions. In the case of stage Ⅳ, sever degenerative change of articular cartilage and subchondral bone were observed. In the mild osteoarthritic articular cartilage, there were observed marked ultrastructural differences in the articular surface, the subsurface fibrillar framework of the matrix and chondrocytes. It was estimated that the organization of collagen fibrils in articular cartilage shows a three-dimensional network with a special system in each zone. In the severe osteoarthritic cartilage, it is observed degenerated chondroid materials, destroyed and irregular oriented collagen fibers and amorphous conglomeration. In the mild osteoarthritis caused by avascular necrosis of femoral head is initially developed from minor disruption of ultrastructural collagen fibrils due to an excessive tensile stressed than eventually progressed to osteoarthritis. The result of this study was acquired from a small number of specimens but such a study may provides very helpful information in the analysis of other diseases related to articular cartilage if similar continued research is carried out in the future as there are few reports as yet published on the morphological structure of articular cartilage.

The Effect of Hyaluronic Acid on Full Thickness Defect of Articular Cartilage in Rabbit Knee

Kim, Yong Beum,Woo, Young Kyun CATHOLIC MEDICAL CENTER 1996 Bulletin of the Clinical Research Institute Vol.24 No.2

The hyaluronic acid, a high molecular weight glycoprotein ubiquitous in connective tissues, has protective functions as it plays role in the aggregation for proteoglycan, keeps synovial fluid viscoelasticity for lubrication, absorbs stress on cartilage, and protects articular cartilage surface layer. Recently scarless healing effect of hyaluronic acid on tissue has been also noted. The purpose of this study is to observe the hyaluronic acid effect on full thickness defected articular cartilage of rabbit kness. All experimental animals were subjected to operation to create full thickness defect at their distal femoral condyles. The experimental 48 animals were divided into 4 groups containing 12 animals in each. In group Ⅰ, 5mm diameter full thickness defect was created at their distal femoral condyle articular surface and left empty. In group Ⅱ, the same defect was created and grafted with autogenous perichondrium from the right costal cartilage. In group Ⅲ, the knee was injected with hyaluronic acid every week after creating same defect until the time of their sacrifice. In group Ⅳ, the same defect was created and grafted with perichondrium from the autogenous costal cartilage and hyaluronic acid was injected every week until the time of their sacrifice. Gross, histologic, histochemical, transmission electron microscopic and scanning electron microscopic observations were done for 12 weeks. The results were as follows; 1. Grossly at the 12th week, in group Ⅰ, the repaired tissue was rough and its margin was irregular but distinguishable from the adjacent normal cartilage. In group Ⅱ, the grafted tissue was well taken with even surface but margin was distinguishable from the adjacent normal cartilage. In group Ⅲ, the reparative tissue surface was regular and margin was smooth. In group Ⅳ, the grafted tissue was well integrated with smooth surface and its margin was undistinguishable from neighboring tissue. 2. Histologically, in group Ⅰ, fibrinoid arcade and red blood cells appeared in the defect site at the 4th week. The spindle shaped mesenchymal cell filled the cartilagenous defect as well as subchondral defect by the 8th week. At the 12th week, cartilagenous defect was filled mostly with fibroblast like cells. In group Ⅱ, grafted perichondria were integrated to surrounding tissue at the 8th week. And the defect was filled with tissue containing many chondrocytes at the 12th week. Group Ⅲ and Ⅳ showed almost complete healing of the articular cartilage by the 12th week. 3. Histochemical staining with safranin-O showed staining of matrix at the base of chondrocyte layer in group Ⅲ. In group Ⅳ, the whole layers of chondrocyte matrices were strongly stained. 4. Transmission electron microscope examination was carried out with 12 week samples. In group Ⅰ, the surface layer was irregular with disintegration of lamina splendes, and the arrangement of superficial and tangential layers of articular surface was also irregular. The underlying tissue contained spindle shape cells that resembled fibroblast. In group Ⅱ, the lamina splendes was absent, but underlying tissue contained normal appearing chondrocyte. In group Ⅲ and Ⅳ, preserved lamina splendes and regularly arranged collagen layer containing many chondrocytes were observed. And some chondrocytes at basal layer of repaired tissue exhibited well developed Golgi apparatus. 5. Scanning electron microscope was examined with tissue from the 12th week. In group Ⅰ, the regenerated tissue had rough surface with distinguishable margin, but it seemed well integrated. In group Ⅱ, the regenerated tissue was distinguished from the surrounding cartilage by a thin gap, and it had no lamina splendens with rough surface compared to normal articular cartilage. In group Ⅲ and Ⅳ, the margin was indistinguishable from normal cartilage and the surface was amooth and regular. These results suggest that the hyaluronic acid may play an important role in regenerating cartilage in full thickness defect of the rabbit knee joint.

자가골막 이식을 이용한 관절연골의 재생에 관한 실험적 연구 : An Experimental Investigation the Rabbit

최장석,김영창,김기훈 인제대학교 1999 仁濟醫學 Vol.20 No.2

The articular cartilage can be damaged by trauma or disease. Because of the limited potential of damaged articular cartilage for either repair or regeneration, there is a need for biological resurfacing of large articualr cartilage defects in diseased or damage joints by transplantation of a tissue with significant chondrogenic potential. The purpose of this study was to investigate the use of free periosteum in articular defects and to characterize the newly formed cartilage. The periosteum was used for repair of articular cartilage defect in femoral condyle. The process of repair was evaluated grossly or histologically at from 2, 4, 6 and 8 weeks after grafted process. The results were follows : 1. At 2 weeks after operation, neochondrogenesis was rarely seen and the defects was potentially filled with some fibrous tissue. 2. At 4 weeks, the hyaline cartilage appeared focally 75% in grafted group and 25% in control group. 3. At 6 weeks, the defects were filled with more dove loped hyaline cartilage cells in all grafted group, but only 38% (6weeks), 44% (8weets) of the control group. 4. The articular cartilage defect can be repaired by autogenous periosteal chondro-genesis cartilage and the newly formed tissue 8 were originated from the cambium layer of periosteal grafts.

The Re-Expression of Collagen Type 2, Aggrecan and Sox 9 in Tissue-Engineered Human Articular Cartilage

( Munirah Sha`ban ),( Aminuddin Bin Saim ),( Samsudin Osman Cassim ),( Chua Kien Hui ),( Fuzina Nor Hussein ),( Ruszymah Bt Hj Idrus ) 한국조직공학과 재생의학회 2005 조직공학과 재생의학 Vol.2 No.4

This study was designed to verify the optimal · basic culture media that promote chondrocytes proliferation in vitro in order to facilitate adequate amount of chondrocytes for cartilage reconstructionas well as maintaining cartilage specific phenotype. Human articular chondrocytes were cultured in three types of basic culture media Ham`s F12, DMEM and the equivalent mixture of F12:DMEM. Cultured chondrocytes were trypsinized as they reached confluency. The viability and total number of cell were recorded at every passage. Large-scale culture expansion was used to reconstruct tissue-engineered cartilage. Quantitative RT-PCR analysis was used to evaluate the expression of collagen Type II, collagen Type I, aggrecan and Sox 9 gene, both in monolayer culture and in the engineered cartilage. The mixture of F12:DMEM promotes significantly greater (p<0.05) chondrocytes proliferation at every passage compared to the individual medium. Monolayer cultured chondrocytes exhibited down-regulation expression pattern of collagen Type II gene, aggrecan and Sox 9, whilst the expression of collagen Type I is up-regulated. Tissue-engineered cartilage morphologically and histologically resembled normal hyaline cartilage. Moreover, tissue-engineered cartilage re-expressed the specific chondrogenesis markers; collagen Type II, aggrecan and Sox 9. In conclusion, the mixture of F12:DMEM enhanced human articular chondrocytes proliferation thus provided adequate amount of chondrocytes for cartilage reconstruction. The new cartilage formed phenotypically resembles native cartilage. This results hold promise for the use of tissue-engineered cartilage implant for future orthopaedic reconstructive surgery.

Tissue Engineering of Articular Cartilage: From Bench to Bed-side

Rozlin Abdul Rahman,Muhammad Aa’zamuddin Ahmad Radzi,Norhamiza Mohamad Sukri,Noorhidayah Md Nazir,Munirah Sha’ban 한국조직공학과 재생의학회 2015 조직공학과 재생의학 Vol.12 No.1

Degeneration or defect of articular cartilage is a major predicament and if it is left untreated, it may leadto progressive damage and disability affecting every one disregard of their age. Although nonsurgical managementof articular cartilage injury has remained largely the same over many years, surgical treatment keeps on evolving. Restorative techniques, mainly the cell-based therapies and autologous or allograft transplants continue to expand,giving surgeons more options for biologic reconstruction of the articular surfaces. Hence the field of articular cartilagetissue engineering which seeks to repair, restore and improve injured or diseased articular cartilage functionalityhas aroused deep interest and holds great potential for improving articular cartilage therapy. However despite thisgreat evolution, therapeutic uncertainty in the restoration of damaged cartilage using tissue engineering approachesstill remains unclear for the surgeon treating patients to make evidence-based decisions. This paper will give a generalidea to different level of audiences in understanding the concept of tissue engineering from bench to bed-sideregarding recent developments in this exciting field.

Effect of Cinnamomum Cassia on Cartilage Protection in Rabbit and Human Articular Cartilage

Baek, Yong-Hyeon,Huh, Jeong-Eun,Lee, Jae-Dong,Choi, Do-Young,Park, Dong-Suk The Society of Korean Medicine 2007 대한한의학회지 Vol.28 No.4

Background & Objective: Articular cartilage is a potential target for drugs designed to inhibit the activity of matrix metalloproteinases (MMPs) to stop or slow the destruction of the proteoglycanand collagen in the cartilage extracellular matrix. The purpose of this study was to investigate the effects of Cinnamomum cassia in inhibiting the release of glycosaminoglycan (GAG), the degradation of collagen, and MMP activity in rabbit and human articular cartilage explants. Methods: The cartilage-protective effects of Cinnamomum cassia were evaluated by using glycosaminoglycan degradation assay, collagen degradation assay, colorimetric analysis of MMP activity, measurement of lactate dehydrogenase activity and histological analysis in rabbit cartilage explants culture. Results: Interleukin-1a (IL-1a) rapidly induced GAG, but collagen was much less readily released from cartilage explants. Cinnamomum cassia significantly inhibited GAG and collagen release in a concentration-dependent manner. Cinnamomum cassia dose-dependently inhibited MMP-1, MMP-3 and MMP-13 activities from IL-1a-treated cartilage explants culture when tested at concentrations ranging from 0.02 to 1 mg/ml. Conclusion : These results indicate that Cinnamomum cassia inhibits the degradation of proteoglycan and collagen through the down regulation of MMP-1, MMP-3 and MMP-13 activities of IL-1a-stimulated rabbit and human articular cartilage explants.