## 1 Introduction  
Osteoarthritis (OA) is the most prevalent articular disease among the elderly [1]. It involves alterations in joint structure and function, primarily resulting from a degenerative inflammatory process occurring in the articular cartilage (AC) [2, 3]. OA affects nearly 70% of individuals at some point in their lives, leading to significant economic and social consequences for both patients and healthcare systems [4, 5]. Moreover, OA is characterized by the degradation of AC [6] and is the most prevalent clinical syndrome associated with joint pain, frequently causing various degrees of functional disability and a diminished quality of life [7]. The extracellular matrix (ECM) of AC is primarily composed of collagen and proteoglycans (PGs), including aggrecan and glycosaminoglycans (GAGs) [8; 9; 10; 11]. As an effect of OA, AC components, particularly PGs, vanish, resulting in tissue destruction and hypocellularity and ultimately loss of joint function [12]. Notably, proinflammatory cytokines promote the breakdown of cartilage and hinder the production of new matrix components in the articular tissue [13]. Cytokines are also known to trigger an increase in prostaglandin E2 (PGE2) and the release of nitric oxide from chondrocytes [14; 15]. Both PGE2 and nitric oxide are reported to play roles in the process of cartilage matrix turnover [16; 17]. Hence, developing disease-modifying OA drugs is necessary [18]. Presently, the main clinical strategy for managing OA is based on the use of nonsteroidal anti-inflammatory drugs (NSAIDs), analgesics, and hyaluron. Although these treatments provide symptom relief, they do not induce a significant disease-modifying effect [19]. In certain cases, NSAIDs can potentially have harmful effects as they impede the synthesis of PG, a substance that plays a crucial role in preserving cartilage function [19]. Therefore, we should explore the development of alternative agents aimed at preventing cartilage degradation or promoting its effective repair. Thus, numerous efforts have been undertaken to investigate potent cartilage-preserving agents derived from natural sources and combinations with medicinal foods, with the goal of achieving more powerful anti-arthritic treatments.  
Natural eggshell membrane (NEM) is a naturally occurring byproduct of egg processing, often overlooked and discarded as industrial waste, thereby adding to the environmental burden [20]. However, NEM contains an abundance of fibrous proteins, collagen, hyaluronic acid, chondro sulfate, and glucosamine [21]; thus, it holds the promise of being a functional bioactive compound in the field of health science. In rodent studies, oral supplementation of eggshell membrane (ESM) is generally considered safe, as it has demonstrated both anti-inflammatory and anti-obesity effects [22; 23]. Moreover, NEM contains bioactive glycosaminoglycans, including dermatan sulfate, chondro sulfate, and hyaluronic acid [24], making it similar to the components found in human joints, particularly in the synovial membrane. Due to its composition, NEM has the potential to enhance joint health, alleviate pain, and reduce the stiffness often associated with OA. It achieves this by supplementing crucial anabolic factors that are often lacking in the typical Western diet and by mitigating inflammation through immunomodulatory effects [25].  
Thus, the purpose of this study was to explore the anti-OA effect of NEM and its underlying mechanisms in primary chondrocytes and a knee OA rodent surgery model.