Closing the gap: sex-related differences in osteoarthritis and the ongoing need for translational studies

Clinical insights into sex-related differences in osteoarthritis

Initially, it was believed that molecular and cellular mechanism found in a species were ubiquitous amongst its members, regardless of age, sex or other inter-individual factors. Over the last few decades, a paradigm shift has occurred with researchers and clinicians backing away from generalisation and moving towards a personalized approach. Ongoing research efforts have focused on identifying differences in the pathogenesis, prevalence, incidence, and severity of a given disease based on inter-individual factors. A sexual dimorphism has been established across a wide variety of processes (including genetic, molecular, cellular, clinical and psychological) during both health and disease. Unfortunately, in the field of degenerative joint disease, and particularly osteoarthritis (OA), the understanding of the influence of sex on disease is still limited (1). This is partially due to a historical perception that OA was simply caused by “wear and tear” with the majority of research focusing on biomechanical causes of disease. More recently, several studies have explored the intricate biological processes that contribute towards the pathogenesis of OA; findings support that it is a far more complex pathology than initially believed (2).

For almost four decades, researchers have reported sex-related differences in the clinical presentation and prevalence of OA (3). Clinical evidence has emerged that women over the age of 55 years have a higher prevalence of knee OA than men (4). In addition, women suffer from more debilitating pain (4), rapidly progressing annual articular cartilage loss (4 times the rate in comparison to men) (5) and a more severe radiographic OA phenotype than men (4). Sex hormones, such as estrogen, have been shown to mitigate pain and exert protective roles on articular cartilage biochemistry (5). Therefore, researchers postulate estrogen loss in postmenopausal women may explain the exacerbated progression of OA (5).

To date, no prophylactic or disease modifying drug is available for clinical use; the only established long-term treatment remains joint replacement using arthroplasty. Consequently, the Osteoarthritis Research Society International (OARSI) still considers OA to be an incurable disease (6). However, recent studies have shown that men achieve better and faster functional recovery, while women have a favourable prognosis with respect to implant survival (5). Findings from a meta-analysis conducted in 2005 (4) and a systematic review conducted in 2021 (5) investigating the role of sex differences in OA are surprisingly similar, indicating a lack of novel insights in the last decade. Published reports have heterogenous patient populations and outcome measures. Further, there is a persistent male dominated sex bias for enrolment in clinical trials (5). In fact, only 32–50% of publications concerning knee OA evaluated sex-specific outcomes (6).

Sex differences may be attributed to multiple factors such as biomechanical properties, gene expression, behavior, and sex hormones levels. An ongoing lack of studies investigating potential mechanisms of action, genetic determinants for sexual dimorphism in OA as well as molecular pathways underlying the observed clinical differences exists. Thus, studies on a preclinical level are needed to improve the understanding of sex differences in OA which is a crucial step in developing new sex-based approaches ranging from disease modifying drugs to diagnostics, treatment, and physical rehabilitation programs.

Finding evidence-based explanations for gender differences in the aetiology of OA

Researchers use pre-clinical models of OA to understand both the aetiology of OA and investigate novel treatments for OA. These models include in vitro models of cell culture coupled with in vivo animal models utilizing a variety of species from rodents (mouse, rat, rabbit) to large animal (dogs, sheep, goats, horses) models of disease. Interestingly, animal models of disease also display sex differences. However, these sex differences may be contrary to those documented in humans and, therefore, can further complicate OA research. For example, it is well established that male mice experience more significant OA than female mice (7). Further, male mice seem to experience greater OA associated pain (7). Several publications have implicated sex as a determinant of both innate and adaptive immune responses (8). This coupled with recent advances indicating the immune system’s role in OA (2), and significant immune variations between sexes provide an exciting avenue for explaining the sex differences in OA.

Pathomechanisms associated with the development of OA cause an imbalance between anabolic and catabolic factors affecting chondrocytes and the integrity of the extracellular matrix (9). Both inflammatory and mechanical drivers of OA have been identified. On a cellular level, a pro-inflammatory T-helper cell polarization has been associated with OA development (2). While on a molecular level, an increased release of pro-inflammatory (such as IL1β, IL6, IL8, and TNF-α) as well as catabolic cytokines (MMPs) has been implicated.

Sex hormones appear to be crucial in understanding the sex differences associated with OA development. Although sex hormones, themselves, have been extensively studied the individual pathways of these interactions in context with OA are still being investigated. Further, therapeutic interventions are not simple; long-term use of estrogen not only fails in reversing OA progression but also increases the risk of breast and endometrial cancer (10). Thus, alternative targets are needed for novel treatment modalities.

Sex differences are evident both in vitro and in vivo. Female chondroblasts and chondrocytes had higher mRNA levels of pro-inflammatory cytokines (IL1A, IL6, and IL8) and lower expression of signalling molecules (WNT5A and DKK2) compared to male chondroblasts, and female osteoblasts exhibited higher levels of estrogen receptors (ER) compared to their male counterpart during culturing (9). ER signalling in the cartilage has been established as a connection between sexual dimorphism and OA development. Specifically, studies suggest there is crosstalk between ER and the extracellular matrix receptor integrin a1β1: epidermal growth factor receptor (EGFR) axis (11). Upregulation of EGFR activity in articular cartilage results in production of reactive oxygen species and catabolic enzymes promoting cartilage destruction. Further, EGFR antagonism by erlotinib is protective against the development of post-traumatic OA in female mice exclusively (11).

Sex-related differences are even evident in the cellular and molecular composition of the synovial fluid. Studies showed female patients had higher levels of monocytes/macrophages as well as a significantly higher ratio of CD4⁺/CD8⁺ T-cells and consequently higher levels of inflammatory cytokines (IL2a, IL3, IL12p40, IL16, IL18, and TNF-α) as well as pro-inflammatory mediators (MCP-3) (12). In contrast, synovial fluid from male patients had higher levels of sGAGs, TGF-β1 and TGF-β2 (9). Furthermore, differing effects of the biological milieu on disease progression have been observed. Increased IL-17A and IL-23 levels, both of which are related to Th17 cells, double the risk for OA associated bone marrow lesions in females but not males (11).

Despite the important findings made on a preclinical level it is imperative to assess the limitations associated with in-vitro experiments and in-vivo animal studies. There is a general lack of consistency amongst individual studies investigating sex-differences in OA development (9). While in-vitro studies have partly investigated molecular mechanisms, in-vivo studies have relied largely on histopathological differences with minimal analysis of molecular mechanisms. Studies are needed that reduce experimental variability by boosting sample size. Studies should investigate molecular mechanisms in the onset and development of OA including multiomic analysis (9) and attempt to translate this information to cellular and ultimately clinical processes. Bioinformatics approaches can be used to screen for sex-related differences in OA development and resulting differentially expressed genes (DEGs). DEGs can be used as a starting point to decipher biological pathways and mechanisms contributing which can be used to close the gap in understanding gender differences in OA.

A 2020 study was one of the first to investigate sex differences in OA using bioinformatics. The study identified specific hub genes in either female or male patients and the authors concluded that it is likely that sex differences exist in the pathomechanisms of OA (13). Likewise, a similar study published in 2019 identified pathways mediating sex-related OA differences between males and post-menopausal females (10). The authors identified hub genes associated with PI3K-Akt signalling, focal adhesion and osteoclast differentiation all in agreement with previous data acquired in-vitro. EGF and EGFR were identified as two hub genes corresponding to sex-differences in osteoarthritis (11). Building on the evidence for sex specific pathomechanisms of OA, a consecutive study investigated potential cartilage targets. (6) Females exhibited a higher elevation of COL1A1, COL1A2, and CD90 (an inflammatory regulator) in response to OA indicating a more robust inflammatory and cartilage response to OA induction (6). These results provide a partial explanation for the more severe OA symptoms found in females. In addition, the importance of the PI3K-Akt signalling pathway, which also regulates subchondral bone dysfunction and synovial inflammation, was further corroborated (6). Although these studies further support the fundamental role of sex-related differences in the pathogenesis of OA, the datasets are derived from patients with end stage OA. While insights gained are invaluable for understanding sex differences, samples from earlier stages of disease will be important for investigating pathomechanisms with the intent of prevention or treatment of disease. In addition, genomic level differences do not necessarily translate into available biological targets, thus further studies are necessary to identify tangible therapeutical targets.

A recent study by Feng et al. screened some of the same datasets as previous studies for hub genes (14). The study confirmed previously identified genes including EGF, EGFR, CDC42, AQP4, and STAT 1 in addition to NTRK3 and CaMK4; neither NTRK3 nor CaMK4 have been previously linked to OA pathogenesis. Polymerase chain reaction (PCR) analysis confirmed a significantly higher expression of CaMK4 in osteoarthritic females compared to males and healthy females, establishing CaMK4 as a key sex-related gene in OA. To further investigate the role of CaMK4, OA was induced in mice by medial meniscal destabilization and the efficacy of early KN-93, a potent CaMK4 inhibitor, was evaluated. KN-93 administration was chondroprotective, further supporting the role of CaMK4 in OA (14). CaMK4 is an important member of the calmodulin-dependent kinase family known to be mainly distributed in the brain, thymus and bone marrow and not yet linked to musculoskeletal diseases. CaMK4 plays an intricate role in orchestrating the immune response, is a known inhibitor of IL-2 (a cytokine essential to the differentiation of regulatory T-helper cells), and promotes inflammatory cytokine and chemokine expression (14). Interestingly, IL-2 acts via downstream signalling pathways including the PI3K-Akt pathway thereby providing a potential mechanism of OA pathogenesis (15).

While CaMK4 is a promising potential target for a sex-specific disease modifying drug, this is only the first step in bringing a disease modifying therapeutic to market (16). The extensive resources (financial and temporal) needed to advance a drug from bench to bedside thwart many promising therapeutics. Approximately 2 billion USD and 20 years are needed for an end-to-end translational process and even then, the success rate is under 1% (17). Scientific discoveries are only one side of the coin in closing the gap and providing gender-specific interventions for the prevention or reversal of OA. In the future it will be necessary to actually advance and translate promising findings derived from robust data. This will require an interdisciplinary team working across medical fields and breaching the wall between academia and industry.

Acknowledgments

Funding: None.

Footnote

Provenance and Peer Review: This article was commissioned by the editorial office, Annals of Translational Medicine. The article did not undergo external peer review.

Conflicts of Interest: Both authors have completed the ICMJE uniform disclosure form (available at https://atm.amegroups.com/article/view/10.21037/atm-23-1546/coif). The authors have no conflicts of interest to declare.

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