The production of yellow pond turtle plastron refers to a type of polypeptide with the anti-inflammation effects, antioxidant activity, and immune system enhancing effect and may help strengthen the bone [1–3]. Many peptides also exhibit important functions in response to diseases through the anti-inflammatory pathway[6–8]. However, there has been no report on the cartilage protective effect of YPTP in KOA and the related mechanism. KOA caused by chronic synovial inflammation is a common chronic disease characterized by joint destruction, which is an irreversible pathological change. Currently, there is no curative therapeutic strategy for KOA, and anti-inflammatory agents and physical therapy are the most common treatments. When KOA develops to a certain stage, it may be inevitable to conduct an arthroplasty or osteotomy[26, 27]. Therefore, it is of great significance to dissect the mechanism for the positive effect of nutraceutical and medicinal food on KOA.
Network pharmacology analysis results implied that YPTP could alleviate KOA through various pathways, such as pathways in cancer, TNF signaling pathway, and PI3K-AKT signaling pathway[28]. The results indicated that the anti-inflammatory targets of YPTP were distributed in multiple signaling pathways, among which COX-2 and iNOS are the most important targets. IL-1β and TNF-α could activate each other, resulting in the activation of the NF-κB pathway. Meanwhile, the biosynthesis of COX-2 and iNOS was activated by IL-1β, IL-6, and TNF-α, which will then promote the expression of PGE2 and NO[29].
To confirm the effect of YPTP on collagen degradation and synthesis, CTX-II and COMP were measured in the serum samples from rats. With the degradation of cartilage, CTX-II and COMP fragments would be increased in the serum and synovial fluid of patients with rheumatoid arthritis. The levels of CTX-II and COMP have been proposed as effective biomarkers of joint injury progression, and their levels in serum can be used to evaluate the severity and progression of KOA[30]. In this study, papain could induce KOA in rats, and the KOA group showed the highest severity of KOA, followed by the LYPTP group and then HYPTP group. In this regard, YPTP might have certain suppressive effect on joint injury progression of KOA.
TNF-α, IL-1β, IL-6, NO, and PGE2 are inflammatory factors closely related to KOA, which play vital roles in the NF-κB/iNOS-COX-2 signaling pathway. TNF-α directly promotes the degradation of cartilage matrix and the destruction of chondrocytes, and induces the production of COX-2, iNOS, IL-6 and IL-1β, aggravating the inflammatory reaction of joint synovium. The production of PGE2 and NO is mainly regulated by inducible COX-2 and iNOS, respectively[31]. NF-κB is a group of protein dimers involved in immune and inflammatory responses[32]. In this study, YPTP showed dose-dependent effects on serum TNF-α, IL-1β, IL-6, NO, and PGE2 levels, which was consistent with the results of knee joint swelling and Mankin scoring. These results revealed that YPTP might regulate IL-1β, IL-6, TNF-α, PGE2, NO, COX-2, and iNOS in the NF-κB/iNOS-COX-2 signaling pathway, and the inhibition of these inflammatory factors could stimulate the repair of cartilage tissue in osteoarthritis patients.
MMP-3 can activate other MMPs to inhibit the degradation of COLLII. Therefore, MMP-3 is a major enzyme for cartilage degradation[33]. COLLII can help maintain the normal structure of cartilage, relieve osteoarthritis, and inhibit the degeneration of bone and joint[34]. These facts are consistent with our research results that YPTP can not only inhibit the degradation of extracellular matrix and reduce the structural damage of articular cartilage by suppressing the expression of MMP-3, but also increase the expression of COLLII to promote cartilage self-repair and control the degeneration of bone.
The results of network pharmacology and in vivo pharmacological evaluation suggested that inhibition of COX-2 or iNOS expression might regulate the normal expression of MMP-3 and COLLII. In addition, our findings revealed the inhibition efficacy of YPTP on cartilage damage and bone resorption. Therefore, COX-2, iNOS, MMPs, and COLLII might be potential targets for the prevention of KOA. Further in vitro experiments are needed to verify these targets and how COX-2 and iNOS mediate the activation of MMPs. The reults of this study provide a good foundation for understanding the protective effect of nutraceutical and medicinal food on KOA.