Repair Effects of Xiaoyusan New Formula on Cartilage Injury and MMP-13 Expression in Knee Osteoarthritis Model Rabbits

doi:10.12300/j.issn.1674-5817.2022.099

Abstract

Abstract:

Objective To study the effect of Xiaoyusan new formula on the articular cartilage of knee osteoarthritis (KOA) rabbits and its mechanism. Methods A total of 42 New Zealand white rabbits aged 6 months were randomly divided into normal group, model group, ointment of Xiaoyusan group, and ointment of Xiaoyusan new formula group, with 10 rabbits in each group (the other 2 rabbits were used for model validation). Except for the normal group, the right knee joints of all rabbits in the other groups were prepared as KOA models according to the modified Hulth method. After 5 weeks of molding, the rabbits in ointment of Xiaoyusan group, ointment of Xiaoyusan New Formula group were given corresponding ointments for knee arthritis treatment, once a day, each time for 10 hours. After 2-week continuous administration and treatment, the knee joint cartilage of the four groups of rabbits was taken and the cartilage damage of each group was evaluated by Outerbridge grading method. The pathological changes of the cartilage, calcified layer and subchondral bone of the knee joint of rabbits in each group were observed by HE staining method under the light microscope, and the degree of cartilage degeneration was evaluated by Mankin's method. The expression of matrix metalloproteinase-13 (MMP-13) in the cartilage of rabbit knee joint in each group was deteced by immunohistochemistry. Results After the general observation of articular cartilage, the Outerbridge grading showed that the number of high-grade animals in ointment of Xiaoyusan group was reduced compared with the model group (P<0.05), and the number of high-grade animals in ointment of Xiaoyusan new formula group was also reduced (P<0.05) compared with ointment of Xiaoyusan group. HE staining showed that Mankin's scores of articular cartilage in the four groups ranked from high to low: model group (10.82±1.76), ointment of Xiaoyusan group (6.19±1.23), ointment of Xiaoyusan new formula group (2.64±1.18) and normal group (0.28±0.17). The difference among four groups was statistically significant (P<0.05). Immunohistochemical detection showed that the positive rates of MMP-13 expression in rabbit articular cartilage tissues in each group were (67.90±13.94)% of model group, (37.10±19.16)% of ointment of Xiaoyusan group, (13.60±3.10)% of ointment of Xiaoyusan new formula group and (3.20±2.39) % of normal group, ranking from high to low, and the difference among four groups was statistically significant (P<0.05). Conclusion Xiaoyusan new formula can repair articular cartilage degeneration in KOA rabbits and decrease the expression of MMP-13 in cartilage, which may be one of the mechanisms of the treatment.

Key words: Xiaoyusan new formula, Knee osteoarthritis, Articular cartilage, Matrix metalloproteinase-13

CLC Number:

Q95-33

Ruolin CUI, Qing WANG, Ling YANG, Wenchang FENG, Zhiwei LIU, Weiran TENG, Bitao MA, Leyang WANG, Liping QIN, Lilun JIN. Repair Effects of Xiaoyusan New Formula on Cartilage Injury and MMP-13 Expression in Knee Osteoarthritis Model Rabbits[J]. Laboratory Animal and Comparative Medicine, 2023, 43(1): 30-38.

Figures/Tables 4

Figure 1 The results of gross observation and HE staining of rabbit knee cartilage after modelingNote：A, a pair of knee joints from a rabbit in normal group; B, a pair of knee joints from a rabbit in model group (the left one is healthy side, right one is affected side); C-D, the HE staining pathological images of a pair of knee cartilage tissues from a rabbit in the model group (C is the affected side, D is the healthy side; the scale bar is 100 μm).

Table 1 Comparison of cartilage Outbridge grade and Mankin's score among different groups

分组 Group	动物数量 n	Outerbridge分级 Outerbridge grade						Mankin评分 Mankin's score $x ¯$ ±s
分组 Group	动物数量 n	0	Ⅰ	Ⅱ	Ⅲ	Ⅳ	最高级 (Ⅲ-Ⅳ)	Mankin评分 Mankin's score $x ¯$ ±s
正常组 Normal group	10	9	1	0	0	0	0	0.30±0.68
模型组 Model group	10	0	0	2	5	3	8^*	10.20±3.12^*
消瘀散组 Xiaoyusan group	10	0	3	2	5	0	5^#	6.10±2.64^*#
消瘀散新组方组 Xiaoyusan new formula group	10	0	7	2	1	0	1^#※	3.20±1.69^*#※

Table 1 Comparison of cartilage Outbridge grade and Mankin's score among different groups

分组 Group	动物数量 n	Outerbridge分级 Outerbridge grade						Mankin评分 Mankin's score $x ¯$ ±s
分组 Group	动物数量 n	0	Ⅰ	Ⅱ	Ⅲ	Ⅳ	最高级 (Ⅲ-Ⅳ)	Mankin评分 Mankin's score $x ¯$ ±s
正常组 Normal group	10	9	1	0	0	0	0	0.30±0.68
模型组 Model group	10	0	0	2	5	3	8^*	10.20±3.12^*
消瘀散组 Xiaoyusan group	10	0	3	2	5	0	5^#	6.10±2.64^*#
消瘀散新组方组 Xiaoyusan new formula group	10	0	7	2	1	0	1^#※	3.20±1.69^*#※

Figure 2 HE staining of rabbits articular cartilage among different groupsNote：A, normal group; B, the model group of knee arthritis by surgery; C, ointment of Xiaoyusan group; D, ointment of Xiaoyusan new formula group. The scale bar is 100 μm.

Figure 3 Comparison of matrix metalloproteinase-13 （MMP-13） expression of rabbits articular cartilage among different groupsNote：A, normal group; B, the model group of knee arthritis by surgery; C, ointment of Xiaoyusan group; D, ointment of Xiaoyusan new formula group. Detected by immunohistochemistry with DAB staining, the scale bar is 100 μm.

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