Establishment and Evaluation of a Moderate-to-Severe Knee Osteoarthritis Model in Rats Induced by Surgery

doi:10.12300/j.issn.1674-5817.2024.066

Abstract

Abstract:

Objective To establish a rat model of moderate-to-severe knee osteoarthritis, laying the foundation for studying the pathogenesis of moderate-to-severe knee osteoarthritis and its prevention and treatment methods. Methods Thirty male SD rats were randomly divided into three groups: a sham surgery group, an 8-week model group, and a 20-week model group, with 10 rats in each group. Rats in the 8-week and 20-week model groups underwent surgery to cut the anterior and posterior cruciate ligaments and medial collateral ligament of the right knee joint, and remove the medial and lateral menisci. After surgery, the rats were allowed to move freely. The rats in the sham surgery group had only skin incisions to expose the joint without any surgical treatment. At 8 and 20 weeks post-surgery, Micro-CT scans were performed to analyze the femoral osteoporosis in the rats. After euthanizing the rats, gross observations of the knee joints were made, and the cartilage of the joint surface was scored using the Pelletier scoring system. The knee joints were collected for hematoxylin and eosin (HE) staining and safranin O-fast green staining to observe changes in cartilage morphology. The modified Mankin's scoring system was used to assess the tissue pathology of the joint surface. Immunohistochemical staining was used to detect the expression of type II collagen and matrix metalloproteinase 13 (MMP13), reflecting the anabolic and catabolic metabolism of the knee joint cartilage. Results The knee joint cartilage in the 8-week and 20-week model groups was severely damaged, with Pelletier and modified Mankin's scores significantly higher than those in the sham surgery group (both P<0.01). The Pelletier and modified Mankin's scores in the 20-week model group were significantly higher than those in the 8-week model group (P<0.01). Micro-CT observations revealed irregular joint surfaces, osteophyte formation, and signs of osteoporosis in both the 8-week and 20-week model groups, with the 20-week model group showing more loose bodies around the knee joints. Immunohistochemical staining showed increased expression of MMP13 and decreased expression of type II collagen in the knee joint tissues of the model groups, indicating that the balance of anabolic and catabolic metabolism in the joint cartilage was disrupted. MMP13 increased while type II collagen decreased. Conclusion The surgical method of cutting the anterior and posterior cruciate ligaments and medial collateral ligament and removing the medial and lateral menisci successfully creates a moderate-to-severe knee osteoarthritis model in rats. Imaging examinations reveal osteophytes, osteoporosis, and loose bodies in the knee joints, while pathological observations show a reduction or even disappearance of joint cartilage, with a disruption in the balance of cartilage anabolic and catabolic metabolism.

Key words: Animal model, Knee osteoarthritis, Micro-CT, Histopathology, Rat

CLC Number:

R684.3

SUN Xiaorong,SU Dan,GUI Wenjuan,et al. Establishment and Evaluation of a Moderate-to-Severe Knee Osteoarthritis Model in Rats Induced by Surgery[J]. Laboratory Animal and Comparative Medicine, 2024, 44(6): 597-604. DOI: 10.12300/j.issn.1674-5817.2024.066.

Figures/Tables 5

Figure 1 Micro-CT images of knee joint in different groups of ratsNote： A， Micro-CT of knee joint in sham surgery group； B， Micro-CT of knee joint in 8-week model group； C， Micro-CT of knee joint in 20-week model group. The sequence of images in each group， from left to right， is as follows： CT scan on sagittal plane， CT scan on coronal plane， X-ray transmission image， and 3D reconstruction image. Red arrows show the osteophytes； White arrows show the joint loose bodies.

Figure 2 Micro-CT femur morphological analysis in different groups of ratsNote： BMD， Bone mineral density； BV/TV， Bone volume fraction； Tb.Th， Trabecular thickness； Tb.Sp， Trabecular separation； Tb.N， Trabecular number. Compared with sham surgery group， *P<0.05； n=10.

Figure 3 Pelletier score of the knee joint in different groups of ratsNote：Compared with sham surgery group， **P<0.01； Compared with 8-week model group， ##P<0.01； n=10.

Figure 4 Histopathological changes of the knee joint in different groups of ratsNote： A， HE staining （×100）； B， Safranin O-Fast Green staining （×100）； C-D， Immunohistochemical staining （×100）. MMP13， matrix metalloproteinase 13； Collagen II， type II collagen.

Figure 5 Mankin’s score of the knee joint in different groups of ratsNote：Compared with sham surgery group， **P<0.01； Compared with 8-week model group， ##P<0.01； n=10.

References 24

1	BRUYÈRE O, HONVO G, VERONESE N, et al. An updated algorithm recommendation for the management of knee osteoarthritis from the European Society for Clinical and Economic Aspects of Osteoporosis, Osteoarthritis and Musculoskeletal Diseases (ESCEO)[J]. Semin Arthritis Rheum, 2019, 49(3):337-350. DOI: 10.1016/j.semarthrit.2019.04.008 .
2	HUNTER D J, BIERMA-ZEINSTRA S. Osteoarthritis[J]. Lancet, 2019, 393(10182): 1745-1759. DOI: 10.1016/S0140-6736(19)30417-9 .
3	DAINESE P, WYNGAERT K V, DE MITS S, et al. Association between knee inflammation and knee pain in patients with knee osteoarthritis: a systematic review[J]. Osteoarthritis Cartilage, 2022, 30(4):516-534. DOI: 10.1016/j.joca.2021.12.003 .
4	WOOLF A D, PFLEGER B. Burden of major musculoskeletal conditions[J]. Bull World Health Organ, 2003, 81(9):646-656.DOI: 10.1590/S0042-96862003000900007 .
5	SHARMA L. Osteoarthritis of the knee[J]. N Engl J Med, 2021, 384(1): 51-59. DOI: 10.1056/NEJMcp1903768 .
6	NÜESCH E, DIEPPE P, REICHENBACH S, et al. All cause and disease specific mortality in patients with knee or hip osteoarthritis: population based cohort study[J]. BMJ, 2011, 342: d1165. DOI: 10.1136/bmj.d1165 .
7	MINTARJO J A, POERWANTO E, TEDYANTO E H. Current non-surgical management of knee osteoarthritis[J]. Cureus, 2023, 15(6): e40966. DOI: 10.7759/cureus.40966 .
8	TSOKANOS A, LIVIERATOU E, BILLIS E, et al. The efficacy of manual therapy in patients with knee osteoarthritis: a systematic review[J]. Medicina (Kaunas), 2021, 57(7):696. DOI: 10.3390/medicina57070696 .
9	KATZ J N, ARANT K R, LOESER R F. Diagnosis and treatment of hip and knee osteoarthritis: a review[J]. JAMA, 2021, 325(6):568-578. DOI: 10.1001/jama.2020.22171 .
10	SELLAM J, COURTIES A, EYMARD F, et al. Recommendations of the French Society of Rheumatology on pharmacological treatment of knee osteoarthritis[J]. Joint Bone Spine, 2020, 87(6):548-555. DOI: 10.1016/j.jbspin.2020.09.004 .
11	ATIK O Ş, HANGODY L R, TURAN S. Total versus unicompartmental knee arthroplasty[J]. Jt Dis Relat Surg, 2023, 34(2):235-236. DOI: 10.52312/jdrs.2023.57913 .
12	DOSSETT H G, ARTHUR J R, MAKOVICKA J L, et al. A randomized controlled trial of kinematically and mechanically aligned total knee arthroplasties: long-term follow-up[J]. J Arthroplasty, 2023, 38(6S): S209-S214. DOI: 10.1016/j.arth.2023.03.065 .
13	DREVET S, FAVIER B, BRUN E, et al. Mouse models of osteoarthritis: a summary of models and outcomes assessment[J]. Comp Med, 2022, 72(1):3-13. DOI: 10.30802/AALAS-CM-21-000043 .
14	ALVES-SIMÕES M. Rodent models of knee osteoarthritis for pain research[J]. Osteoarthritis Cartilage, 2022, 30(6):802-814. DOI: 10.1016/j.joca.2022.01.010 .
15	HULTH A. Does osteoarthrosis depend on growth of the mineralized layer of cartilage?[J]. Clin Orthop Relat Res, 1993(287):19-24. DOI: 10.1097/00003086-199302000-00004 .
16	ZEIGHAMI A, DUMAS R, AISSAOUI R. Knee loading in OA subjects is correlated to flexion and adduction moments and to contact point locations[J]. Sci Rep, 2021, 11(1):8594. DOI: 10.1038/s41598-021-87978-2 .
17	VAN TUNEN J A, DELL'ISOLA A, JUHL C, et al. Biomechanical factors associated with the development of tibiofemoral knee osteoarthritis: protocol for a systematic review and meta-analysis[J]. BMJ Open, 2016, 6(6): e011066. DOI: 10.1136/bmjopen-2016-011066 .
18	ZHANG K J, LI L, YANG L F, et al. Effect of degenerative and radial tears of the Meniscus and resultant meniscectomy on the knee joint: a finite element analysis[J]. J Orthop Translat, 2019, 18:20-31. DOI: 10.1016/j.jot.2018.12.004 .
19	LEEMAN M F, CURRAN S, MURRAY G I. The structure, regulation, and function of human matrix metalloproteinase-13[J]. Crit Rev Biochem Mol Biol, 2002, 37(3):149-166. DOI: 10.1080/10409230290771483 .
20	FAN A Y, WU G B, WANG J F, et al. Inhibition of fibroblast activation protein ameliorates cartilage matrix degradation and osteoarthritis progression[J]. Bone Res, 2023, 11(1):3. DOI: 10.1038/s41413-022-00243-8 .
21	CHELLINI F, TANI A, PARIGI M, et al. HIF-1α/MMP-9 axis is required in the early phases of skeletal myoblast differentiation under normoxia condition in vitro [J]. Cells, 2023, 12(24):2851. DOI: 10.3390/cells12242851 .
22	GROEN S S, SINKEVICIUTE D, BAY-JENSEN A C, et al. A serological type II collagen neoepitope biomarker reflects cartilage breakdown in patients with osteoarthritis[J]. Osteoarthr Cartil Open, 2021, 3(4):100207. DOI: 10.1016/j.ocarto.2021.100207 .
23	HU Q C, ECKER M. Overview of MMP-13 as a promising target for the treatment of osteoarthritis[J]. Int J Mol Sci, 2021, 22(4):1742. DOI: 10.3390/ijms22041742 .
24	YAO X D, SUN K, YU S N, et al. Chondrocyte ferroptosis contribute to the progression of osteoarthritis[J]. J Orthop Translat, 2021, 27:33-43. DOI: 10.1016/j.jot.2020.09.006 .

[1]	LIU Lida, CHEN Bing, XIE Na, LIU Li, ZHUANG Siqi, ZOU Yixing. Survey Report Analysis on Parasitic and Microbial Quality of Laboratory Animals in Sichuan Province, 2017-2023 [J]. Laboratory Animal and Comparative Medicine, 2024, 44(6): 654-660.
[2]	LIU Yishu, CAI Liping. Advances and Challenges of Using Experimental Pigs in Da Vinci Surgical Robot Training [J]. Laboratory Animal and Comparative Medicine, 2024, 44(6): 667-674.
[3]	ZHAO He, ZHANG Fan, XIAO Yuzhou, AN Xuefang, ZHANG Tao, LI Li. Preliminary Diagnosis and Characterization of a Spontaneous Immature Testicular Teratoma in an Interferon Receptor-Deficient Mouse Model [J]. Laboratory Animal and Comparative Medicine, 2024, 44(6): 691-694.
[4]	YANG Jiahao, DING Chunlei, QIAN Fenghua, SUN Qi, JIANG Xusheng, CHEN Wen, SHEN Mengwen. Research Progress on Animal Models of Sepsis-Related Organ Injury [J]. Laboratory Animal and Comparative Medicine, 2024, 44(6): 636-644.
[5]	ZHAO Lijuan, XIAO Chunlan, SHENG Yajie, LU Xi, ZHOU Zhengyu. Challenges and Development in Suzhou Laboratory Animal Industry Over the Past Five Decades [J]. Laboratory Animal and Comparative Medicine, 2024, 44(6): 645-653.
[6]	TIAN Fang, PAN Bin, SHI Jiayi, XU Yanyi, LI Weihua. Advances in Development of PM_2.5-Exposed Animal Models and Their Application in Reproductive Toxicity Research [J]. Laboratory Animal and Comparative Medicine, 2024, 44(6): 626-635.
[7]	ZHAO Xiaona, WANG Peng, YE Maoqing, QU Xinkai. Establishment of a New Hyperglycemic Obesity Cardiac Dysfunction Mouse Model with Triacsin C [J]. Laboratory Animal and Comparative Medicine, 2024, 44(6): 605-612.
[8]	YIN Yulian, MA Lina, TU Siyuan, CHEN Ling, YE Meina, CHEN Hongfeng. Establishment and Evaluation of a Rat Model of Non-Puerperal Mastitis [J]. Laboratory Animal and Comparative Medicine, 2024, 44(6): 587-596.
[9]	DU Xiaoyan, LIU Yunbo. Analysis of the Progress in Identification and Evaluation of Laboratory Animal Resources in China [J]. Laboratory Animal and Comparative Medicine, 2024, 44(5): 469-474.
[10]	TU Yingxin, JI Yilan, WANG Fei, YANG Dongming, WANG Dongdong, SUN Zhixin, DAI Yuexin, WANG Yanji, Guanghan KAN, WU Bin, ZHAO Deming, YANG Lifeng. Evaluation of Simulated Weightlessness Model of Hindlimb Unloading Miniature Pigs and Their Tissue Damage [J]. Laboratory Animal and Comparative Medicine, 2024, 44(5): 475-486.
[11]	ZHU Chan, ZHANG Dongliang, ZHAO Deli, SHI Xueqin, QIAN Lei, ZHANG Xuan, JIN Yan, DUAN Wei, QI Ruocheng, LIU Chaohua, YANG Xuekang, HAN Juntao, PAN Dengke. Perioperative Animal Care for Xenotransplantation from Genetically Edited Pigs to Monkeys [J]. Laboratory Animal and Comparative Medicine, 2024, 44(5): 495-501.
[12]	YANG Jin, YU Shiya, LIN Nan, FANG Yongchao, ZHAO Hu, QIU Jinwei, LIN Hongming, CHEN Huiyan, WANG Yu, WU Weihang. Effect of Modified Duodenal Exclusion Surgery on Glucose Metabolism in Rats with Type 2 Diabetes Mellitus [J]. Laboratory Animal and Comparative Medicine, 2024, 44(5): 523-530.
[13]	QI Longju, CHEN Shiyuan, LIAO Zehua, SHI Yuanhu, SUN Yuyu, WANG Qinghua. Transcriptomic Analysis of Menstrual Blood-Derived Stem Cells Transplantation Combined with Exercise Training in Promoting Spinal Cord Injury Recovery in Rats [J]. Laboratory Animal and Comparative Medicine, 2024, 44(5): 531-542.
[14]	ZHANG Naiqun, YUAN Piaopiao, CAO Linrong, YING Na, YANG Taotao. Application of PNR Detection in the Diagnosis and Drug-efficacy Evaluation of Diabetic Kidney Disease in Rats [J]. Laboratory Animal and Comparative Medicine, 2024, 44(5): 543-549.
[15]	HUANG Dongyan, WU Jianhui. Establishment Methods and Application Evaluation of Animal Models in Reproductive Toxicology Research [J]. Laboratory Animal and Comparative Medicine, 2024, 44(5): 550-559.