Effects and Mechanisms of Sinew Stagnation and Bone Dislocation Manipulation on Cartilage Degeneration in Rabbit Knee Osteoarthritis via the Extracellular Signal-Regulated Kinase Pathway

ObjectiveTo investigate the effects of the Sinew Stagnation and Bone Dislocation Manipulation on cartilage degeneration in rabbit with knee osteoarthritis (KOA) via the Extracellular Signal-Regulated Kinase (ERK) pathway and to elucidate its underlying mechanisms.MethodsA total of 30 healthy male New Zealand White rabbits were randomly divided into blank group (n=10), and modeling group (n=20). The blank group received no treatment, while the modeling group followed the Videman modeling method to establish an early KOA model. Due to modeling failure and death during the intervention period, 8 and 16 cases were finally included in the blank group and the modeling group, respectively. The modeling group was further divided into a model group and a manipulation group, with eight cases in each group. The blank group and the model group were allowed free activity and diet without any intervention; the manipulation group received sinew stagnation and bone dislocation manipulation technique after successful modeling, including manipulation of regulating tendon stagnation (manipulation along the three Yang meridians of foot and three Yin meridians of foot; pressing acupoints such as Futu (ST32), Liangqiu (ST34), medial and lateral Xiyan (EX-LE4, EX-LE5), Xuehai (SP10), Yanglingquan (G34), Ashi points, 60~80 times per minute, two minutes per acupoint, 12 minutes per session, once a day, for a total of 21 days and bone dislocation correcting technique once a week, three times a course, with a total intervention of 21 days. The Lequesne Algofunctional Index (Lequesne MG score) was used to assess rabbit behavior and gait; Hematoxylin-Eosin (HE) staining was used to observe pathological changes in rabbit knee joints; Safranin O-Fast Green staining was used to evaluate structural alterations in articular cartilage and subchondral bone; Western blot analysis was conducted to determine the protein expression levels of ERK1/2, phosphorylated ERK1/2 (pERK1/2), matrix metalloproteinase-13 (MMP-13), and tumor necrosis factor-alpha (TNF-α); quantitative reverse transcription polymerase chain reaction (qRT-PCR) was employed to measure the mRNA transcription levels of ERK1/2, MMP13, interleukin-1 beta (IL-1β), and TNF-α; Micro-CT was utilized to assess trabecular connection density (Conn.D) and trabecular number (Tb.N) in rabbit knee joints.Results(1) Lequesne MG score: compared with the blank group, the Lequesne MG scores of the model group and the manipulation group increased significantly after intervention (P<0.05). Compared with the model group, the Lequesne MG score in the manipulation group decreased significantly after intervention (P<0.05). (2) Cartilage pathological changes: the cartilage surface of the medial plateau was uneven in the model group, with local loss of cartilage, decreased superficial chondrocytes showing hypertrophic differentiation, and deep chondrocytes clustered into groups; obvious cartilage fissures or defects were observed, and cartilage matrix staining was uneven. The superficial cartilage in the manipulation group was more regular than that of the model group, chondrocytes arrangement and distribution were more uniform, and cartilage matrix staining was more even. (3) TNF-α, MMP-13, ERK1/2 protein expression levels and pERK1/2 proportion: compared with the blank group, TNF-α, ERK1/2 protein expression levels and pERK1/2 proportion in the model group significantly increased after intervention (P<0.05), and MMP-13 protein expression level significantly increased (P<0.05). Compared with the model group, TNF-α, MMP13, ERK1/2 protein expression levels and pERK1/2 proportion in the manipulation group significantly decreased after intervention (P<0.05). (4) MMP-13, IL-1β, TNF-α, and ERK1/2 mRNA transcription levels: compared with the blank group, TNF-α, MMP13, and IL-1β mRNA transcription levels in the model group significantly increased after intervention (P<0.05). Compared with the model group, mRNA transcription levels of MMP-13, IL-1β, TNF-α, and ERK1/2 in the manipulation group significantly decreased after intervention (P<0.05). (5) Bone mineral density and trabecular number: compared with the blank group, Conn.D and Tb.N in the model group significantly decreased after intervention (P<0.05); trabecular structure was irregular, and osteophyte formation significantly increased. Compared with the model group, Conn.D and Tb.N in the manipulation group significantly increased after intervention (P<0.05), trabecular structure was more regular, and osteophyte formation significantly decreased.ConclusionThe sinew stagnation and bone dislocation manipulation technique can improve knee joint structure and knee joint function of KOA model rabbits, and its mechanism may be related to regulating the ERK pathway to inhibit inflammatory responses and extracellular matrix degradation, and alleviating endoplasmic reticulum stress damage.