Tendinopathy is a common musculoskeletal disorder characterized by pain, inflammation, and degenerative changes in tendons. Globally, tendinopathy is becoming more common, and this can lead to long-term or permanent functional limitations in people of all ages, including athletes and non-athletes [7]. Furthermore, the incidence and prevalence of tendinopathy varies widely between different parts of the body, as well as based on factors including age, sex, the kind of physical activity, occupational setting and specific disease condition. Tendinopathies are commonly observed in the lower and the upper extremities of the body. The incidence of lower limb tendinopathy is reported to be 10.52 per 1,000 person-years [8, 9]. In the upper extremity, tendinopathies in the supraspinatus within the shoulder and the elbow are the most common. Collagen fiber disarray, increased microvasculature and sensory nerve innervation, dysregulated extracellular matrix homeostasis, elevated immune cell presence and inflammatory mediators, and enhanced cellular apoptosis are important histological and molecular characteristics of tendinopathy [1]. Tendinopathy is also a multifaceted and intricate tendon pathology. There are many factors that can induce this kind of disorder. The hypothesized pathological and physiological mechanisms of tendinopathy include the mechanical hypothesis, inflammation hypothesis, apoptosis hypothesis, vascular or neural origin hypothesis, as well as the continuum model hypothesis [1].
TCM compounds have multi-target and multi-pathway biological effects. As a traditional classic prescription, THSWD is currently widespread in the fields of internal medicine, surgery, gynecology, and orthopedics [10, 11]. It is widely used in clinical practice for cardiovascular diseases [12], osteoporosis and bone fracture[13–16] and exhibits excellent therapeutic effects. In animal models, THSWD alleviates ischemic strokes and myocardial fibrosis [17–19]. In long-term clinical practice, THSWD is a representative prescription of TCM that promotes blood circulation and eliminates stasis.
This study investigated the underlying mechanisms of THSWD in the treatment of tendinopathy using an integrative approach combining bioinformatics, network pharmacology, and molecular docking. Based on bioinformatics and network pharmacology, we screened the key core targets and effective ingredients of THSWD for treating tendinopathy, and conducted PPI, GO and KEGG enrichment analysis to clarify the possible mechanisms and pathways involved. Meanwhile, the binding activity between target proteins and compounds was evaluated by molecular docking analysis, scientific evidences for the clinical application of THSWD in treating tendinopathy were found.
The study identified 45 active components in the THSWD and 216 potential targets for treating tendinopathy. The key targets PTGS2, IL6, and AKT1 may serve as potential biomarkers and therapeutic targets for tendinopathy. PTGS2, also known as COX-2, is an enzyme protein that plays a crucial role in the synthesis of PGH2 and is involved in inflammation reactions and other physiological processes [20]. Researches have shown that ruptured Achilles tendons highly express NF-κB target genes, PTGS2 and IL-8, exhibiting vascular dilation [21]. This suggests a close association between ruptured tendons, inflammation reactions, and vascular dilation. In cases of tendon rupture, inflammatory responses are activated along with the NF-κB signaling pathway, leading to increased expression of various inflammation-related genes. At the same time, IL-8 is a pro-inflammatory cytokine that promotes recruitment and activation of inflammatory cells, further intensifying the inflammatory response [22, 23]. The cytokine IL-6 plays a pivotal role in immune responses and acute phase reactions, being secreted by various cell types including monocytes/macrophages, T cells, and endothelial cells [24]. Furthermore, IL-6 significantly contributes to the growth and development of tendons while facilitating their healing process post-injury[25]. Andersen, et al (2011) confirmed that mechanically impaired tendons in IL-6 knockout transgenic mice exhibited lower mechanical properties and tissue characteristics compared to untreated control mice [26]. This may be related to the activation initiation of the STAT-3 signaling pathway induced by IL-6 since the STAT-3 signaling pathway is closely associated with cell proliferation and survival [27]. The AKT1 protein, belonging to the AKT kinase family, exerts a crucial influence on intracellular signal transduction and predominantly governs processes such as cell proliferation, differentiation, and survival [28].
The results of molecular docking suggest that luteolin and quercetagetin, which are present in various vegetables and traditional Chinese herbs, may serve as potential key active compounds for the treatment of tendinopathy. The two compounds belong to the flavonoid class and exhibit a wide range of biochemical and pharmacological activities, encompassing antioxidant, antibacterial, anti-inflammatory, antiallergic, and vasodilatory properties [29]. De Bioquímica et al. (2010) verified that flavonoids can inhibit the activation of MAPK and AKT signaling pathways induced by lipoteichoic acid (LTA) stimulation in H9c2 cells [29]. Previous research has also shown that luteolin and quercetagetin can suppress phosphorylation of ERK 1/2, p38, and JNK in human gingival fibroblasts induced by LTA stimulation [30]. Kang et al.(2014) confirmed that quercetagetin may be an effective inhibitor of STAT1 signaling, potentially serving as a new molecular target for anti-inflammatory therapy and having therapeutic applications as an immunomodulator in inflammatory diseases such as atopic dermatitis (AD) [31]. Recent studies have also found that luteolin can protect skeletal muscles from inflammation-induced damage and downregulate the expression of proteins associated with muscle breakdown metabolism [32]. Liu et al. (2024) demonstrated that luteolin not only enhances the structural integrity of cartilage and promotes tissue repair in mice with arthritis but also suppresses the expression of COX-2 and matrix metalloproteinase 1 (Mmp1), while simultaneously upregulating collagen protein expression [33]. Furthermore, luteolin not only enhances muscle strength in fatigued subjects and improves skeletal muscle contraction during ischemia-reperfusion [34], but also inhibits VEGFA and affects microvascular network formation during mouse angiogenesis [35].
The KEGG enrichment analysis revealed that the key pathways implicated in the treatment of tendinopathy with THSWD encompassed the IL-17 signaling pathway, TNF signaling pathway, and AGE-RAGE signaling pathway in diabetic complications. In the IL-17 signaling pathway, IL-17 binds to its receptor and activates extracellular signal-regulated kinases (ERK), c-jun N-terminal kinases (JNK), and p38/MAPK pathways, resulting in the upregulation of IL-6, IL-1, and NF-kB [36]. The IL-17 signaling pathway plays a crucial role in host defense against extracellular bacterial and fungal infections as well as the pathogenesis of various autoimmune diseases [37]. TNFα is a cytokine that exerts direct tumoricidal effects with minimal cytotoxicity towards normal cells [38]. It actively participates in systemic inflammatory responses and represents one of the key cytokines involved in acute phase reactions, predominantly produced by activated macrophages [39]. The AGE-RAGE signaling pathway refers to the binding of protein glycation products (AGEs) caused by high blood sugar with their receptor (RAGE), which then triggers a series of reactions in signal transduction pathways [40]. The signaling pathway plays an important role in the occurrence and development of diabetic complications [40]. AGEs are compounds formed by the interaction between sugars and biomacromolecules such as proteins or nucleic acids, which increase abnormally under high blood sugar conditions. These compounds can activate various signaling pathways, such as NF-κB, MAPK, and PI3K-AKT-mTOR, through binding with RAGE, leading to adverse effects such as inflammation response, oxidative stress, endothelial dysfunction, and cell apoptosis [41]. In patients with diabetes mellitus, activation of the AGE-RAGE signaling pathway accelerates the occurrence and development of diabetes-related complications such as neuropathy, renal damage,and cardiovascular diseases[40]. The activation of the AGE-RAGE signaling pathway is implicated in neuronal injury, inflammatory response, and oxidative stress in diabetic neuropathy. Moreover, it contributes to structural and functional impairments such as glomerulosclerosis and interstitial fibrosis in diabetic nephropathy, while also inducing endothelial dysfunction and vascular smooth muscle cell proliferation with detrimental effects on diabetic cardiovascular diseases.
In summary, the study utilized bioinformatics, network pharmacology, and molecular docking analysis to explore the potential targets and mechanisms of action of THSWD in treating tendinopathy. The study identified luteolin and quercetagetin as potential core active compounds in THSWD for treating tendon diseases. The potential key targets were found to be PTGS2, IL6, AKT1, and the key signaling pathways involved IL-17 signaling pathway, TNF signaling pathway, and AGE-RAGE signaling pathway in diabetic complications. THSWD exerts its therapeutic effects on complex molecular mechanisms of tendinopathy through a multi-component, multi-targeted approach involving multiple pathways. It is an important medication for clinical treatment of tendon diseases and further experimental and clinical research should focus on its key targets and signaling pathways.