ABSTRACT

Degenerative joint disease is a condition that affects joints and is commonly referred to as osteoarthritis (OA). This form of arthritis is most prevalent among women and tends to become more frequent as people age. The pathogenesis of OA involves an imbalance of cytokines in favor of pro-inflammatory cytokines. However, the steroid hormone dehydroepiandrosterone (DHEA) exerts chondroprotective effects and regulates the balance of catabolic factors such as thrombospondin motif disintegrin and metalloproteinase (ADAMTS), thereby playing a role against OA. Pro-inflammatory cytokines induce aggrecanases, such as ADAMTS5, which degrade the extracellular matrix and contribute to OA. The molecule nerve growth factor (NGF), associated with pain in OA, is important for cartilage homeostasis, and DHEA can modulate pain by interfering with NGF receptors. This review covers the roles of DHEA, ADAMTS5, and NGF in the pathogenesis of OA, their relationship with pain pathways, and their use in current treatments. We also anticipate that these pathways will be crucial in developing new strategies to prevent and treat OA, and understanding their interactions may make it possible to enhanced the quality of life of patients with OA.

Keywords:

ADAMTS5, DHEA, NGF, osteoarthritis

Introduction

Worldwide, more than 250 million people are affected by osteoarthritis (OA), which is known as the most prevalent form of arthritis and characterized as a degenerative joint disease (1-3). Factors such as continued population growth and aging are the main causes of the increasing in the prevalence of OA which makes it considered as on of the leading causes of disability among elderly (4,5). OA has a higher incidence in women and is a significant socio-economic burden in many countries globally (6). The disease is progressive and debilitating, causing pain and resulting in a loss of function, which can be so severe that it disrupts the patient’s ability to get restful sleep. At the societal level, the annual cost of OA is estimated to exceed $303 billion due to medical costs and lost earnings (7). The economic impact of OA is expected to double by 2030 on a global scale (8), emphasizing the urgency for extensive research to comprehensively understand the contributing factors in the onset and advancement of the disease. Various factors contribute to women’s susceptibility to OA, including thinner cartilage, joint instability, misalignment, and unequal mechanical loading (9). Recent studies have demonstrated that a sharp decrease in sex hormone levels during menopause can trigger OA development (10,11). Other risk factors for OA include trauma, genetics, high basal metabolic index, and structural abnormalities in the joint (12). Genetic factors have been found to be effective in primary generalized OA with Heberden’s nodule, Bouchard’s nodule, hip involvement, and knee involvement. This is particularly evident in Heberden’s nodules and is carried by an autosomal gene that is dominant in females and recessive in males. Matched twin and family-risk studies have shown that the genetic contribution to OA may be around 50-65% (13). For instance, occupational activity in professional athletes such as football players can cause the development of OA (12). OA can affect both small joints, such as those in the hand, and larger joints such as the knee and hip (14). Although pain is the primary symptom, other symptoms may accompany OA, including joint swelling, locking, cramping, decreased range of motion, and morning stiffness that resolves within thirty minutes (15). Radiographic evaluation of the joint is the gold standard in diagnosis (16). Treatment options for OA vary depending on the severity of the disease and the individual’s specific needs. Some of the available treatments include joint replacement surgery, autologous mesenchymal stem cell transplantation, and non-steroidal anti-inflammatory drugs (NSAIDs) to reduce pain (17,18). In those whose pain is not adequately controlled, first-line evidence-based analgesia, NSAIDs and acetaminophen (paracetamol) are used. Oral NSAIDs and the use of cyclooxygenase-II (COX-II) inhibitors, opiates, or intra-articular steroids are considered when first-line agents fail (13,19). Additionally, early interventions such as chondroitin sulfate and glucosamine may be recommended (17). Physical therapies can also be used to treat early-stage OA (17). Finally, a Mediterranean diet may decrease the prevalence of OA and improve patients’ quality of life (20).

In the pathogenesis of OA, an important factor is the disruption of the balance between anti-inflammatory and pro-inflammatory cytokines, with pro-inflammatory cytokines often becoming more dominant. The induction of interleukin (IL)-1β results in aggrecanases such as a Disintegrin and Metalloproteinase with ThromboSpondin motifs (ADAMTS)-4 and ADAMTS5 leading to extracellular matrix (ECM) degradation. Moreover, matrix metalloproteinase (MMP) induction can cause hypertrophy, differentiation, and apoptosis in chondrocytes due to these events (21). Among the 19 members of ADAMTS enzymes with various functions, ADAMTS-5 is more notable in arthritis (22). Dehydroepiandrosterone (DHEA) is a steroid hormone produced by the adrenal gland cortex, which regulates the balance between catabolic factors such as ADAMTS in cartilage (23). DHEA shows chondroprotective effects and reduces oxidative stress, protecting against OA (24). Although the role of DHEA in modulating OA-related pain is not confirmed, studies suggest that DHEA can interfere with nerve growth factor (NGF) receptors (25). NGF is primarily expressed in synovial fibroblasts and is associated with pain. Patients with knee OA and hip OA have higher NGF expression, as studies have demonstrated (26). There is evidence from mouse models that NGF and other neurotrophins are overexpressed in symptomatic diseases and are themselves synthesized by joint connective tissue (19). In 2019, a study emphasized the role of NGF signaling in the calcification of human joint chondrocytes and the importance of NGF signaling in articular cartilage homeostasis (27). Although conventional OA treatments alleviate pain, they cannot reverse cartilage damage (28). Therefore, it is crucial to further explore new molecular targets. This article will explain how ADAMTS5, DHEA, and NGF contribute to OA disease and the mechanisms they mediate by summarizing articles published in the last 5 years.

In humans, DHEA is synthesized in the central nervous system (CNS), gonads, and adrenal cortex, and it has been found to have anti-inflammatory effects on various tissues, including the prevention of leukocyte recruitment. DHEA has been shown to interact with various nuclear receptors, such as estrogen receptors, as well as G protein-coupled receptors found in endothelial and neuronal cells (29). In a study conducted by Lazaridis et al. (30), specific antibodies were employed to target tropomyosin-related kinase A (TrkA) and p75 in coimmunoprecipitation assays and Western blot analyses of precipitates. The study showed that DHEA can directly suppress the NGF receptors p75 and TrkA, and can eliminate both receptors from PC12 cells (30). Given this discovery, it is feasible that DHEA could potentially compete with NGF for its intended receptors, resulting in the inhibition of peripheral pain production. This is because NGF binding to these receptors located in peripheral nociceptors is responsible for initiating the downstream cascade of pain signals (25,31).

In addition, research has demonstrated that both DHEA and NGF can effectively prevent apoptosis in neuronal cells. Their antiapoptotic effects begin at the plasma membrane and involve the activation of similar prosurvival kinase cascades. They also regulate the transcription of the antiapoptotic protein B-cell lymphoma 2 (Bcl-2) through the activation of transcription factors nuclear factor kappa-B (NF-B) and cyclic AMP-responsive element binding protein. Researchers have hypothesized that NGF receptors may play a role in the antiapoptotic effects of DHEA, given the similarities observed in the signal transduction pathways triggered by both molecules (32).

DHEA Pathways in OA

DHEA is a hormone that is synthesized in the zona reticularis of the adrenal gland and serves as the precursor to all sex steroid hormones. Its levels are more closely linked to age than gender, with a marked decline in the elderly that is strongly linked to the development of age-related conditions (33,34). Numerous studies have emphasized the potential therapeutic benefits of DHEA in treating chronic degenerative joint diseases, including OA. Earlier studies have suggested that administering DHEA has a positive effect on cartilage preservation in animal models of OA, particularly in the early and mid-stages of the disease (35-37). In 2015, a study was conducted to investigate the modifying effect of DHEA in various stages of experimentally induced OA disease. The results showed that DHEA treatment significantly reduced cartilage lesions and delayed cartilage degeneration in four regions of the knee (38). These findings suggest that DHEA may play a role in the pathophysiology of OA. DHEA is produced by the zona reticularis part of adrenal gland (39). The inactive sulfate ester of DHEA is converted to sulfate DHEAS in the adrenal glands and liver (40). They are bound to albumin and together form the most abundant steroid hormones in the human circulation (41). Due to the aging-related decrease of DHEAS and DHEA levels, it has been suggested that the formation of aging-related diseases may be linked to a relative deficiency of these hormones (42,43). Therefore, it is important to consider the role of DHEA in the context of OA and explore its potential as a treatment option. Moreover, DHEA has been shown to have protective effects against various aging-related diseases such as dementia (44,45), osteoporosis (46), and atherosclerosis (47).

DHEA’s ability to positively influence chondrocyte/articular cartilage metabolism has been demonstrated in numerous animal- and cell-based studies, which supports its chondroprotective effects (36,37,48). Disruption to this role of DHEA could be considered as a predominant cause of OA related pathogenesis. Chondrocytes, a special cell type of the skeletal system, play a significant role in skeletal maturation and fracture healing, and endochondral ossification, a process that replaces the developing cartilage nidus by bone, may assist both fracture healing and skeletal maturation (49). Although there are few studies about the relationship between DHEA and chondrocytes, the literature suggests that DHEA can affect the overall performance of chondrocytes (49). By preventing the expression of matrix metalloproteinases enzymes known to catalyze cartilage degradation, DHEA has a chondroprotective impact on mature cartilage (36,37,48). Therefore, DHEA may be helpful in degenerative chronic conditions like OA. Moreover, there is evidence to suggest that Wnt/β-catenin pathway can be modulated by DHEA, which is critical for the maintenance of myogenic and cartilage homeostasis, with β-catenin levels playing a crucial role (50). A 2013 study examined the protective mechanism of DHEA in experimental models of OA and suggested that the chondroprotective effect of DHEA on the animals cartilage and chondrocytes may be due in part to its aromatase-mediated conversion to estrogens. This process may occur via the blocking of aromatase with letrozole (36). Therefore, it is understood that DHEA levels play a vital role in the mechanism of OA. In healthy cartilage, a robust collagen scaffold and high aggrecan content are necessary for its weight-bearing properties. However, massive loss of aggrecans, caused by aggrecanases, is the hallmark of most arthropathies, including OA (51). Aggrecanase-mediated degradation of aggrecan is an important event in the early stages of OA. While the mechanisms of action of DHEA in OA are not fully understood, data from in vitro studies and animal models suggest that its protective efficacy on osteoarthritic cartilage may be due to its role in the inhibition of pro-inflammatory pathways, leading to down-regulation of MMP enzymes that play a critical role in aggrecan loss in OA (37,52). Additionally, DHEA has been shown to have an anti-catabolic effect by suppressing MMPs and inducing tissue inhibitor of metalloproteinases (TIMPs), which suggests that modulating the balance between MMPs and TIMPs is another protective mechanism of DHEA on OA (53).

Researchers conducted a study in 2003 to explore the impact of DHEA on the expression of catabolic enzymes in chondrocytes. The results demonstrated that DHEA treatment suppressed the expression and protein synthesis of MMP-1, a metalloproteinase associated with cartilage degradation, while increasing the expression and synthesis of TIMP-1, an inhibitor of MMP-1. Additionally, DHEA treatment decreased the expression of type I collagen, which is a marker of ECM senescence, and increased the expression of type II collagen, a marker for chondrogenesis (37). These findings suggest that DHEA may have chondroprotective effects in osteoarthritic chondrocytes.

In 2006, a follow-up study explored the influence of DHEA on chondrocytes obtained from neonatal rats that were exposed to catabolic stimulators, such as lipopolysaccharide (LPS) and S-nitroso-N-acetyl-l-penicillamine (SNAP). The study revealed that DHEA treatment did not impact the viability of healthy chondrocytes or interfere with their glycosaminoglycan (GAG) production capabilities (49). Furthermore, the administration of DHEA was shown to suppress the expression of prostaglandin E2 caused by LPS, MMP-13, MMP-1, and MMP-3, and also hinder the NO synthesis and GAG degradation induced by SNAP in chondrocytes (49). These results suggest that DHEA possesses anti-catabolic characteristics that address inflammation and degeneration, two crucial biological processes involved in the progression and development of OA (49). Taken together, these investigations indicate that DHEA can protect cartilage by impeding catabolic enzymes, stimulating the production of extracellular matrix, and halting inflammation and degeneration in chondrocytes. These results underscore the potential therapeutic advantages of DHEA in treating OA (53).

The pathophysiology of OA involves a complex interplay of various factors, including ECM-degrading enzymes. The MMP family has been extensively studied as an important catabolic factor in the development of OA. However, other enzymes, such as the urokinase plasminogen activator (uPA), the cysteine protease family, and the disintegrin and metalloproteinase with thrombospondin motifs family, have also been implicated in the pathogenesis of OA (54). uPA has been shown to regulate the extent of ECM degradation and is believed to play a role in the development of arthritis (54). Aggrecanases, particularly ADAMTS4 and ADAMTS5 from the ADAMTS family, are the most powerful enzymes that degrade aggrecan, a crucial constituent of the cartilage ECM (55-57). Meanwhile, cathepsins K, B, L, and S from the cysteine protease family are deemed the most significant enzymes in the progression of OA (58,59).

In healthy cartilage, it is necessary to achieve a balance between anabolic and catabolic processes to preserve homeostasis (51). However, if catabolic processes outweigh the chondrocytes’ potential for regeneration, articular cartilage will degenerate. Therefore, targeting these catabolic enzymes, such as MMPs, ADAMTS, and cysteine proteases, could be a potential therapeutic approach for OA.

Several studies have investigated the association between ECM-degrading catabolic enzymes and OA. In 2014, a study investigated the impact of TIMP1, a glycoprotein, on MMPs and revealed that TIMP1 has the capacity to inhibit all MMPs by establishing complexes with high-affinity at one to one ratio (60). This discovery could potentially pave the way for novel OA treatment approaches.

Natural inhibitors of cysteine proteinase and uPA, such as cystatin C and plasminogen activator inhibitor-1 (PAI-1), respectively, have been identified (61,62). To better understand the chondroprotective function of DHEA, researchers conducted a study to investigate how DHEA affected these linked enzyme systems in the development of OA. According to in vivo conducted study, by regulating the metabolic balance of certain enzymes such as ADAMTS/TIMP-3 (63), uPA/PAI-1 (35), MMP-3/TIMP-1 (53), and cysteine proteinases/cystatin C, DHEA protects the cartilage from damage and degradation.

ADAMTS5 Interactions in OA

ADAMTS5 is a group of ADAMTS enzymes containing thrombospondin motifs and metalloproteinases. The ADAMTS family has 19 members in humans (64,65), with members 4 and 5 in the aggrecanases subgroup, which are responsible for tissue morphogenesis and pathophysiological remodeling (66). In arthritis, proteoglycan degradation is facilitated by ADAMTS4 and 5, leading to cartilage aggrecan degradation. Knee injuries increase the risk of post-traumatic OA, and while the relationship of subchondral bone and bone marrow lesions with OA is known, the response of the synovium to joint damage is not fully understood. A study conducted to determine changes in the synovium within the first 14 days of knee injury found that transcripts encoding ADAMTS4 increased in the synovium (67). In addition, the expression of aquaporin 1 (AQP 1), a channel, increased in OA chondrocytes, and the downregulation of AQP 1 was found to decrease the expression of ADAMTS4, which suppresses IL-1β-induced ADAMTS4 (68). Therefore, the reduction of AQP 1 expression in OA can suppress ADAMTS4. OA is characterized by degenerative loss of articular cartilage in synovial joints, with changes in bone and synovium. The main culprit in aggrecan cleavage, an ECM component, is ADAMTS5 (69). In rats with medial meniscal tear, the OA group had a significant increase in ADAMTS5 at 4, 6, and 8 weeks compared to healthy controls and the placebo group, leading Elsadek et al. (70) to conclude that ADAMTS5 could serve as a serological marker in OA.

ADAMTS5, also known as aggrecanase 2, is a risk factor for degenerative disorders due to its overexpression, which is the main cause of joint destruction and matrix loss in OA (71). IL-1β induces ADAMTS5 expression in human chondrocytes, while WW domain-containing protein 2 (Wwp2) overexpression down-regulates it. Thus, Wwp2 is thought to regulate ADAMTS5 expression in articular cartilage (72). A recent study conducted in 2021 showed that ADAMTS5 is highly regulated in cartilage with OA and that miR-9-3p overexpression suppresses ADAMTS5 expression, leading to inhibition of IL-1β-induced apoptosis and ECM destruction. The study also found that MIR22HG inhibition reduced ECM degradation through the miR-9-3p/ADAMTS axis (73). A research study on the genetic variation of ADAMTS5 revealed that the polymorphism of rs2830585 contributes to the risk of OA in the knee. The study found that individuals with the TT genotype had a 1.95-fold higher risk of developing OA compared to those with the CC genotype, and the presence of the rs28305885T allele increased the risk of OA by 39% compared to individuals with the C allele (74). However, it was observed that the ADAMTS5 rs226794 gene polymorphism was not associated with knee OA, and the G allele was not confirmed to be a risk factor for OA (75). In a study by Canbek et al. (64), the relationship between OA and ADAMTS5 gene polymorphisms in Turkey was examined. As a result of the study, no significant difference was found in allele frequencies between the groups for the ADAMTS5 rs226794 and rs2830585 genotypes (64).

Age-related DNA damage is a known factor in the development of OA, as it can lead to cellular aging in joint tissue (76). To investigate the role of interferon genes (STING) in the pathogenesis of OA, a study was conducted in human and mouse cartilage. The study found that STING expression increased in OA patients and overexpression of STING led to increased expression of ADAMTS5. However, the use of lenti-sh-STING to destroy STING reversed the IL-1β-stimulated expression of ADAMTS5. Therefore, the study concluded that STING induces ECM degradation, contributing to the progression of OA (77).

Betulinic acid was found to reduce OA-like changes in a collagenase-injected mouse model by inhibiting the production of pro-inflammatory cytokines and the production of ADAMTS4 and 5 (78).

The TIMP family consists of four members, TIMP1-4, which are proteins with protease inhibitory effects. While TIMP3 is bound to the ECM, the others exist in a soluble form in the ECM. Selective inhibition of members of the ADAMTS family is achieved by TIMPs (79). A study carried out in 2016 aimed to identify the primary endogenous inhibitor of ADAMTS5 and ADAMTS4. The results revealed that TIMP-3 is the most significant inhibitor of ADAMTS4 and ADAMTS5 (80).

In a study investigating the effects of DHEA on the expression of aggrecanases and endogenous inhibitors of aggrecanases in a rabbit model of OA, cartilage treated with DHEA was found to have higher expression of TIMP-3 and lower expression of ADAMTS4 and ADAMTS5 compared to the control group, as determined by real-time quantitative reverse transcription-polymerase chain reaction (RT-PCR) analysis. The results suggest that intra-articular administration of DHEA can decrease the gene expression of aggrecanases and increase the expression of the endogenous inhibitor TIMP-3, leading to a reduction in aggrecanase activity. This suggests that DHEA may be a beneficial treatment for OA by affecting the balance between aggrecanases and TIMP-3 (63).

NGF Pathways in OA

Musculoskeletal conditions such as OA and back pain remain prevalent and cause substantial distress and societal costs, despite considerable therapeutic progress in recent decades (81). To meet the pressing medical need for effective pain relief in these conditions, there is a demand for novel approaches and targeted therapies (81). NGF inhibitors offer a promising alternative to traditional drugs, which often pose risks of adverse effects on organs such as the gastrointestinal tract, heart, or kidneys (81). Although OA can manifest in various functional impairments, pain remains the principal symptom, and thus, pain management represents a key aspect of clinical care for OA patients (82).

NGF, the first growth factor to be identified, was discovered by Rita Levi-Montalcini in 1952. Subsequent studies in the 1950s with Stanley Cohen showed that this factor regulated the growth and development of the nervous system (83). NGF belongs to a family of neurotrophic factors that includes brain-derived neurotrophic factor, neurotrophin-3, and neurotrophin-4 (84). Neurotrophins signal through tyrosine receptor kinases (Trk), also known as tropomyosin receptor kinases, including TrkA, TrkB, and TrkC. Among the neurotrophins, TrkA shows significant NGF specificity and binds to NT-3 as well (85).

Initially, NGF was identified as a soluble signaling protein produced by tumor tissue that promoted the survival and proliferation of sensory neurons (84). However, in the 1990s, it was discovered that NGF also contributes to tissue damage, discomfort, and pain in adults (83). NGF is released by immune cells involved in the inflammatory response to peripheral damage. Besides immune cells, non-immune cells such as endothelial cells, pericytes, chondrocytes, and synoviocytes may also produce NGF (83,86,87). Previous research has suggested that NGF does not immediately cause pain but rather contributes to pain by inducing peripheral and potentially central nerve sprouting (88). In an original study published by Testa et al. (89) in 2019, researchers investigated the relationship between NGF and pain by creating transgenic mice with the human 661C>T NGF gene mutation. The study found that the nociception of heterozygous NGFR100W/wt mice was impaired (89). Recent studies have demonstrated that targeted inhibition of NGF is highly effective in animal models of many acute and chronic pain conditions while being notably free of side effects (90). Interestingly, research suggests that inflammation-induced NGF expression is seen in OA (84).

The Role of DHEA, NGF, and ADAMTS5 in the Treatment of OA and Current Treatments

Currently, there is no cure for OA. However, there are several forms of treatment available that can be grouped into the categories of reducing modifiable risk factors, intra-articular therapy, physical modalities, alternative therapies, and surgical treatments. In the early stages of OA, the primary goal of treatment is to alleviate stiffness and pain. As the disease progresses, the focus of treatment shifts towards maintaining physical function and preventing further damage (15).

OA is typically managed with first-line treatments such as NSAIDs and acetaminophen. However, if these options fail or are not appropriate, stronger medications like weak opioids and narcotic analgesics may be considered. Steroid injections into the affected joint can also be used for the management of inflammatory flares, although their effectiveness is limited and temporary. Intra-articular administration of hyaluronic acid and other viscosupplements can provide longer-lasting treatment, but their short-term efficacy is questionable and results are inconsistent (91-94). Conservative treatment approaches for OA primarily aim to alleviate symptoms, while arthroplasty is usually recommended for individuals with moderate to severe OA. For young patients seeking to preserve their knee function, osteotomy around the knee may be considered as an alternative surgical procedure, as it focuses on “knee preservation” (28). The effectiveness of these interventions may vary based on the severity of the condition (95). The effectiveness of OA treatment can be limited due to various factors, such as an individual’s response to medications like NSAIDs and acetaminophen. Although these drugs can offer pain relief, their prolonged use can lead to serious negative effects and must be used with caution under medical supervision (96). While corticosteroid therapy can significantly improve outcomes in the short term, its regular use may promote cartilage deterioration, joint damage, or tissue atrophy (96). Unlike traditional treatments, in a study conducted in Turkey, autologous conditioned serum (ACS) treatment was tried. For this purpose, they performed bilateral knee injections. The results showed that the use of ACS resulted in significant improvement in pain severity and knee community scoring (97).

Additionally, lifestyle changes like weight loss and exercise can slow disease progression and reduce symptoms, but may not fully alleviate advanced stage symptoms (98). Surgeries, such as joint replacement, can provide significant function improvement and relief of pain, but they are invasive and carry risks like aseptic loosening, stiffness, prosthesis failure, instability, infection, and misalignment (99). OA treatment often involves a gradual process of multiple approaches, and it may not completely eliminate symptoms or restore function.

NGF

Studies using various OA models have demonstrated that anti-NGF drugs can effectively reduce pain-related behaviors, however, preclinical testing of NGF antibodies and TrkA inhibitors in OA models have lagged behind clinical trials (100). Utilizing preclinical models is important in understanding the mechanisms behind the analgesic relief provided by inhibiting the NGF/TrkA pathway in OA, as well as identifying the reasons and risk factors for rapid progression of the disease, which is often observed in clinical trials (100). Despite the lack of human use approval, anti-NGF monoclonal antibodies (mAbs) are being developed as potential therapies for pain management in various conditions (101). Tanezumab (Pfizer in collaboration with Eli Lilly), fulranumab (Amgen), and fasinumab (Regeneron Pharmaceuticals in partnership with Sanofi) are currently humanized mAbs that have been developed to target free NGF as treatments (101). Clinical trials using NGF inhibition have been conducted in patients with hip and knee OA, which have shown that anti-NGF antibodies significantly reduce pain and improve function (102). A recent study by Ohashi et al. (26) 2021 aimed to investigate the relationship between pain, central sensitization, and synovial NGF expression in patients with hip OA who had undergone total hip replacements. The study found that in hip OA patients, synovial NGF expression is linked to both pain intensity and central sensitization, which supports the association between NGF molecule and pain (26). These findings demonstrate the potential for NGF inhibition as a promising therapeutic approach for OA pain management. In a 2019 study by Dakin et al. (103), 342 patients were given fasinumab, an anti-NGF monoclonal antibody drug, over a 36-week period to assess its efficacy and safety in OA. Results showed that fasinumab produced statistically significant and clinically meaningful pain reductions in comparison to placebo at all four dosages from baseline to week 16 (103).

The phase 3 study by Berenbaum et al. (104) in 2020 demonstrated significant pain relief, physical function improvement, and positive physician global assessment scores in patients with moderate-to-severe OA who had not responded to or could not tolerate standard-of-care analgesics after receiving subcutaneous tanezumab at a dose of 5 mg every 8 weeks. It is noteworthy that tanezumab is the most extensively researched anti-NGF drug and the most advanced agent, making it the likeliest candidate for regulatory approval among the anti-NGF drugs studied in OA (105). In 2019, Krupka et al. (106) published an original article evaluating the effectiveness and safety of GZ389988A, TrkA inhibitor, in participants with painful knee OA. Several pre-clinical OA models have demonstrated reduced pain behavior with TrkA inhibition, as TrkA is one of the receptors for NGF. This approach focuses on antagonizing both TrkA and p75NTR, the two NGF receptors, to decrease NGF-induced pain (105).

In a study by Krupka et al. (106) in 2019, 104 participants with moderate to severe knee OA pain were administered a single intra-articular injection of either GZ389988A, a TrkA inhibitor, or a placebo. The authors found that the injection of the TrkA inhibitor resulted in a sustained reduction in pain and a quantifiable functional improvement compared to the placebo. Additionally, the TrkA inhibitor demonstrated an acceptable safety profile (106). In 2020, Ishiguro et al. (107) conducted a study to evaluate the efficacy, safety, and tolerability of ONO-4474, a Pan-Tropomyosin Receptor Kinase inhibitor, in Japanese patients with knee OA. The study demonstrated that the drug effectively reduced pain in individuals with knee OA, supporting the association between blocking Trk and pain reduction in patients with OA, and further supporting the relationship between NGF molecule and pain (107). In contrast, a 2019 study by another research team investigated the efficacy of ASP7962, another TrkA inhibitor, in treating pain in knee OA. Despite a suitable study design, the oral small-molecule TrkA inhibitor did not improve pain in individuals with knee OA, and the authors confirmed the reliability of their results because of the significant improvement in pain observed between naproxen and placebo in the same study (108). According to the authors of the study, the discrepancy in results between their findings and other studies showing pain reduction after TrkA inhibitor administration may be due to the possibility that a higher dose of ASP7962 could have a stronger pharmacological effect, but this would need to be balanced against an increased risk of toxicity (108). Furthermore, the authors suggest that the slightly higher baseline pain levels in the ASP7962 group compared to the placebo group may have hindered the drug’s ability to achieve a statistically significant effect (108). While the effectiveness of DHEA in reducing OA-related discomfort has not yet been confirmed, recent research suggests a potential relationship between DHEA’s pharmacological actions and pain generation in OA (Figure 1). Given the lack of effective pharmaceutical options for treating OA, understanding the molecular pathways underlying DHEA’s pain-relieving effects may pave the way for the development of novel anti-OA medications (25).

DHEA

In 2004, a study was conducted on a rabbit model to investigate the effects of DHEA treatment. Based on gross morphological examination and histological analysis, the results indicated that the femoral condyles that received DHEA treatment exhibited lower levels of cartilage damage compared to the untreated condyles (52). Additional evaluation to the overall structure of the cartilage, Safranin O staining, and thickness showed reduced damage in the DHEA group in comparison to the placebo group. The findings were further corroborated by the RT-PCR based analysis of gene expression. These analysis has demonstrated a reduction in MMP-1/3 mRNA and IL-1 expression. Furthermore, TIMP-1 mRNA levels increase has also been detected in a treated knee joint cartilage with DHEA. These results indicate that DHEA may impede the catabolic degradation of MMPs in the OA process in vivo (109).

In 2015, a study using a rabbit model of OA examined the modifying effects of DHEA on the structure. The results demonstrated that DHEA treatment was capable of halting the advancement of pre-existing cartilage degeneration in various areas of the knee joint at different stages of OA, with certain variations possibly linked to the severity of the disease (38). In particular, in moderate OA, DHEA administration was found to prevent deterioration in both compartments of the knee. However, in advanced OA, DHEA was found to be effective only in inhibiting the deterioration of the lateral knee compartment. This site-specific and time dependent efficacy suggests that DHEA’s structure-modifying effects against OA may vary depending on the location and stage of the disease. The multitargeted protective features of DHEA are illustrated in Figure 2. These findings suggest that understanding the molecular pathways of DHEA’s protective effects on OA may lead to the development of new anti-OA medications, as there are currently no effective pharmaceuticals for treating OA.

ADAMTS5

Current treatments for OA are mainly aimed at relieving symptoms, such as the use of intra-articular drugs like NSAIDs and hyaluronic acid. Surgical options are considered for patients with end-stage OA. Disease-modifying osteoarthritis drugs (DMOADs) have the potential to alter the course of the disease by preventing structural changes in the joint and improving symptoms. One promising DMOAD is the anti-ADAMTS5 Nanobody M6495, which belongs to the proteinase inhibitor family (110). In an ex vivo cartilage model, M6495 demonstrated high affinity for the ADAMTS5 target and did not bind to ADAMTS4. In addition, it was observed that M6495 was able to completely inhibit the enzymatic activity of its target in a concentration-dependent manner (111).

A study conducted on rats with OA, bone marrow mesenchymal stem cell-derived exosomes were found to significantly reduce the upregulation of the IL-1β derived ADAMTS5 proteolytic enzyme, which researchers hypothesized was due to exosomes protecting chondrocytes from IL-1β induced damage. The protective effect was also observed to be dose-dependent (112). Another potential therapeutic agent is hyperoside, a bioactive flavonoid with anti-inflammatory properties that has been shown to reduce ADAMTS5 expression and have anti-arthritic effects (113).

Aptamers are single-stranded DNA or RNAs with 3-dimensional structures that enable them to selectively bind to specific molecular targets (114). Recently, two new DNA aptamer inhibitors, apt 21 and apt 25, were developed, both of which demonstrated high binding affinity and specificity towards ADAMTS5. These aptamers exclusively inhibit ADAMTS5 activity and do not bind to ADAMTS4, making them potential candidates for the treatment of OA (115). A study conducted on rats with knee OA caused by monosodium iodoacetate (MIA) investigated the therapeutic potential of fibroblast growth factor-2 (FGF-2). The researchers overexpressed FGF-2 via rAAV-mediated gene transfer and found that inhibiting toll-like receptor 4 (TLR4) signaling and activating TIMP-1 downregulated ADAMTS5 mRNA and MMP13, markers of knee joint degradation. These results suggest that FGF-2 may have therapeutic benefits for MIA-induced knee OA (116).

In another study, Jia et al. (117) isolated and used murine primary chondrocytes to investigate the effect of cell-free fat extract (CEFFE) on ADAMTS5 expression. The researchers found that when primary chondrocytes were co-cultured with inflammation factors, ADAMTS5 expression increased, while treatment with CEFFE led to a reduction in ADAMTS5 expression. These findings suggest that CEFFE may be a promising therapeutic strategy for the treatment of OA (117).

In a study conducted in 2022, miR-17 was found to be expressed highly in the middle and superficial regions of articular cartilage and was observed to protect against the destruction of cartilage caused by destabilization of the medial meniscus by targeting pathological catabolic factors, including ADAMTS5. In the context of OA, miR-17 downregulation has been observed, which results in an increase in catabolic factors like ADAMTS5. The targeting of these genes by miR-17 suppresses the function of these catabolic factors and helps in maintaining cartilage homeostasis. Therefore, miR-17 could be a potential therapeutic target for the treatment of OA (118).

In a separate study conducted in 2022, researchers utilized IL-1β-treated chondrocytes as a cellular OA in vitro model to assess the inhibitory impact of microRNA-613 (miR-613) on ADAMTS5. The researchers discovered the strong inhibitory effect of miR-613 on ADAMTS5 in this model, and the expression of ADAMTS5 was inversely correlated with the expression of miR-613. These results suggest that miR-613 may be a potential therapeutic target for OA by inhibiting ADAMTS5 expression in chondrocytes (119).

Conclusion

OA is a common condition that affects the joints and can cause pain and disability. Several molecules have been identified as being involved in the pathogenesis of OA, including:

• Pro-inflammatory cytokines such as TNF-α IL-1, which can promote the breakdown of cartilage and stimulate the formation of osteophytes (bone spurs).

• MMPs, enzymes that break down the ECM of cartilage.

• Reactive oxygen species, which can cause damage to cells and contribute to the development of OA.

• Aggrecanases, enzymes that break down the proteoglycan aggrecan, a major component of cartilage.

• Proteolytic enzymes such as ADAMTS4 and ADAMTS5 that can break down the ECM of cartilage.

• Cytokines (IL-1, IL-6, TNF-α) that are involved in promoting inflammation and pain in OA.

In treating OA, various methods such as therapy, medication, and in some cases, joint replacement surgery may be recommended to alleviate pain, improve joint function, and slow down the progression of the disease. Our review has predominantly focused on recent studies that aim to understand the role of DHEA, NGF, and ADAMTS5 in the pathophysiology of OA. These molecules play a significant role in distinct aspects of OA, such as cartilage breakdown, inflammation, and bone remodeling.

Currently, anti-NGF monoclonal antibodies are not a substitute for commonly used medications like NSAIDs. While ongoing research is exploring the effectiveness of targeting NGF as a treatment strategy for OA, there are no approved treatments that specifically target this molecule. On the other hand, targeting ADAMTS5 has shown promise as a treatment strategy for OA. Several ADAMTS5 inhibitors are currently under development, which specifically target the enzyme and inhibit its activity. As a result, these inhibitors slow down the destruction of cartilage. Additionally, it’s worth noting that ADAMTS5 inhibitors are still in pre-clinical and clinical trial phases and not yet available on the market. Meanwhile, DHEA supplements have been suggested as a possible treatment for OA, but their effectiveness remains uncertain due to limited evidence, requiring more research to determine their efficacy in treating the condition. DHEA therapy is not commonly utilized for OA treatment as it lacks FDA approval for that indication, and further studies are necessary to evaluate its safety and effectiveness. The investigation of the molecular mechanisms underlying OA is still ongoing, and new molecules are continuously being discovered.

Ethics

Peer-review: Internally and externally peer-reviewed.

Authorship Contributions

Drafting Manuscript: D.S.A., N.H., O.A., E.B.Y., M.M.A-J., A.K., Critical Revision of Manuscript: D.S.A., N.H., O.A., E.B.Y., M.M.A-J., A.K., Final Approval and Accountability: D.S.A., N.H., O.A., E.B.Y., M.M.A-J., A.K., Writing: D.S.A., N.H., O.A., E.B.Y., M.M.A-J., A.K.

Conflict of Interest: No conflict of interest was declared by the authors.

Financial Disclosure: The authors declared that this study received no financial support.

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