Objective: To investigate the effect of autophagy expression levels of different weight-bearing states and different stages of osteoarthritis in animal models, as well as the corresponding mechanisms.
Methods: We used the male Sprague-Dawley (SD) rats (12-week-old, SPF) to establish the OA animal models by modified Hulth method, and grouped animal models according to the length of time after surgery and different weight-bearing areas. RT-qPCR was carried out for detection of autophagy-related genes such as Atg7, Atg12, P62, etc. Western blot analysis was used to detect the expression levels of corresponding autophagy-related proteins such as LC3B, P62, etc. T test was performed for statistical analysis to compare different groups, while the differences were deemed statistically significant with P < 0.05. Transmission electron microscopy was used to observe the autophagosome to demonstrate the level of autophagy expression and the status of the chondrocytes.
Results: The results of the RT-qPCR testing showed that when the weight-bearing cartilage of the 4-week group (relatively mild) was compared with that of the 10-week group (relatively severe), there were statistically significant differences in all the genes tested, in detail: Atg3 (P < 0.01), Atg7 (P < 0.01), Atg12 (P < 0.01), P62 (P < 0.0001). The expression of autophagy-related mRNA in the 4-week group is increased compared with that of the 10-week group. As for the expression of proteins, Western blotting showed that in the comparison between the 4- and the 10-week groups, statistically significant results include Atg12 (P < 0.01) in the non-weight-bearing area, with decreased autophagy in the 10-week group compared with that of the 4-week group, while expression of LC3B (P < 0.05) protein was significantly higher in the 4-week group than in the control in the non-weight-bearing area. The expression of LC3B (P < 0.0001) and P62 (P < 0.05) in the 10-week group were higher than that of the control. Transmission electron microscope showed that autophagy in the weight-bearing area is stronger than that in the non-weight-bearing area, and autophagy in the 4-week group is stronger than in the 10-week group for the weight-bearing area.
Conclusions: The expression of autophagy varies during different stages of osteoarthritis, in which the autophagy is stronger in the early stage of osteoarthritis, and gradually decreases with the progression of the disease. Autophagy in different weight-bearing areas may also be different.
Keywords: Animal models; Autophagy; Autophagy-related proteins; Chondrocytes; Osteoarthritis.
© 2022 The Authors. Orthopaedic Surgery published by Chinese Orthopaedic Association and John Wiley & Sons Australia, Ltd.