Shaoyang Shenggu Fang inhibits oxidative stress and delays cartilage aging in rats with knee osteoarthritis

Abstract

BACKGROUND: Preliminary studies have demonstrated that Shaoyang Shenggu Fang can alleviate joint cartilage degeneration and promote cartilage repair, but its specific mechanism for alleviating knee osteoarthritis symptoms remains unclear. The Wnt/β-catenin pathway and oxidative stress play crucial roles in maintaining articular cartilage homeostasis. OBJECTIVE: To investigate the molecular mechanisms by which Shaoyang Shenggu Fang regulates the Wnt/β-catenin pathway to inhibit oxidative stress in cartilage and thereby delay cartilage aging in a rat model of knee osteoarthritis. METHODS: Thirty-two Sprague-Dawley rats were randomly divided into four groups: a blank control group, a model group, a Western medicine group, and a Chinese medicine group. Animal models of knee osteoarthritis were established in all groups except for the blank control group by transecting the anterior cruciate ligament and resecting the anterior horn of the medial meniscus. After 28 days of modeling, the Chinese medicine group was administered concentrated Shaoyang Shenggu Fang at a dose of 16 g/(kg·d) by gavage, the Western medicine group received glucosamine hydrochloride solution at 4 mL/d, and the blank and model groups received the same volume of normal saline. After 4 weeks, hematoxylin-eosin staining and Safranin O-fast green staining were used to observe the degree of cartilage damage and degeneration. ELISA was used to detect serum levels of inflammatory factors and oxidative stress indicators. Western blot was used to detect the expression of p21Cip1, p16INK4a, and Wnt signaling pathway-related proteins in knee cartilage. RESULTS AND CONCLUSION: Compared with the model group, the Western medicine and Chinese medicine groups showed significant improvement in cartilage defects, thinning of the cartilage layer, and decreased density, with significantly lower Mankin scores (P < 0.05). Compared with the model group, serum levels of interleukin-1β, tumor necrosis factor-α, and interleukin-6 were significantly decreased in the Western medicine and Chinese medicine groups (P < 0.05), while superoxide dismutase and glutathione peroxidase levels were increased and malondialdehyde concentration was decreased (all P < 0.05). In the Chinese medicine group, the expression levels of p21Cip1, p16INK4a, and Wnt5a proteins were significantly decreased (P < 0.05 and P < 0.01), β-catenin and C-Myc protein expression levels were decreased (P < 0.05), and glycogen synthase kinase-3β protein expression was significantly increased (P < 0.05). These results suggest that Shaoyang Shenggu Fang can significantly reduce inflammation and alleviate cartilage aging in rats with knee osteoarthritis, and the potential mechanism may be through regulation of the Wnt/β-catenin pathway to inhibit cartilage oxidative stress.

1. Introduction

Knee osteoarthritis (KOA) is a common age-related degenerative disease characterized by joint inflammation and chondrocyte senescence, with the main pathological hallmark being articular cartilage degeneration [1-2]. Its complex pathological process often originates from biochemical factors and cellular physiological changes in the synovial tissue, leading to histological and cytological alterations—structural defects in hyaline articular cartilage, loss of subchondral bone integrity, tissue hypertrophy, increased bone mass, and structural changes in the joint, ultimately resulting in whole-tissue dysfunction [3]. Knee joint tissues age over time, and the number of senescent chondrocytes is closely related to age; moreover, due to the important role of bone-cartilage coupling, the senescence tendency of bone cells also increases during aging [4].

The Wnt/β-catenin pathway plays a crucial role in maintaining articular cartilage homeostasis. When the joint is injured, the quiescent Wnt/β-catenin pathway rapidly responds and activates, with Wnt being highly expressed in osteoarthritis and promoting abnormal accumulation of β-catenin [5]. Chondrocyte senescence is a key mechanism in the pathogenesis of osteoarthritis. Studies have shown that the Wnt/β-catenin pathway can promote the senescent phenotype of chondrocytes by downregulating silent information regulator 1 and p53 acetylation [6]. Additionally, the progression of osteoarthritis is closely related to reactive oxygen species (ROS) and oxidative stress. ROS are produced by NADPH oxidases and mitochondria in chondrocytes, affecting gene expression, apoptosis, extracellular matrix synthesis, and inflammatory cytokine production. Overexpression of NADPH oxidase exacerbates oxidative stress and accelerates osteoarthritis progression [7]. The Wnt/β-catenin signaling pathway and oxidative stress interact, making the regulation of the Wnt/β-catenin signaling pathway a potential therapeutic target for KOA.

According to the classic text Lingshu·Genjie, “Shaoyang is the pivot; if the pivot is broken, the bones will be weak and unstable, so for those with flaccidity, treat Shaoyang.” The Shaoyang qi flows along the meridians to nourish the bones, promoting bone growth and joint strength, which is the concept of “Shaoyang governing bone.” While “kidney governs bone” emphasizes the “body” of bone, “Shaoyang governs bone” emphasizes its “function” [8]. The core of the “Shaoyang Sheng Gu” theory is that the Shaoyang meridian plays a dominant role in bone formation and metabolic balance. Clinically, regulating Shaoyang has been used to treat bone diseases, and the Shaoyang Shenggu Fang, derived from the classic formula Xiaochaihu Tang and the theory of “Shaoyang governing bone,” combined with the clinical experience of national renowned TCM expert Sun Tongjiao, has shown efficacy in treating KOA.