Solute Carrier Transporters in Synovial Membrane and Hoffa’s Pad of Patients with Rheumatoid Arthritis

Rheumatoid arthritis (RA) is a complex autoimmune disease that causes destruction of the small joints of the hands and feet. The pathogenesis of the disease is due to the effects of the immune system on the cells that make up joint tissues such as synovial membrane (SM), cartilage tissue, and periarticular fatty tissue (Finckh et al. 2022). Previous studies have shown that the inflammatory process takes place within all tissues that make up the joint, and they can be a source of cytokines and chemokines that cause inflammation and destruction of joint structures (Jang et al. 2022). It is also important that the drugs used in the treatment of RA reach all the tissues of the joint, thus inhibiting the development of inflammation. The transport of drugs, as well as many endogenous compounds involved in the development of inflammation in RA patients, is mediated by membrane transporters. Membrane transporters are proteins that mediate the transfer of substrates across biological membranes (Giacomini et al. 2010). The presence of membrane transporters has been found in several tissues such as liver, intestine, kidney, brain, testis, and placenta (Hediger et al. 2013; Schlessinger et al. 2013). Previous studies have shown that the expression of these transporters varies in different tissues (Hediger et al. 2013; Schlessinger et al. 2013). These studies indicate that membrane transporters exhibit tissue specificity. These transporters play a role in the delivery and exchange of various substrates and metabolites, and thus regulate cellular homeostasis. Many transporters also facilitate the transport and elimination of endogenous and xenobiotic compounds (Fotiadis et al. 2013).

The solute carrier (SLC) gene superfamily currently consists of 458 genes grouped based on the primary amino acid sequence that encodes the corresponding proteins (Hagenbuch and Stieger 2013). Some studies have demonstrated the key functions of these transporters in the transport of drugs, xenobiotics, nutrients, ions, and many metabolites (Koepsell and Endou 2004; Hagenbuch and Stieger 2013). It is recognized that the SLC family of transporters plays a key role in the process of maintaining homeostasis under physiological conditions, as well as influencing the development of some diseases and also the effectiveness of their therapy (Bröer 2008; Sanna et al. 2009; Franke et al. 2010; Jostins et al. 2012; Köttgen et al. 2013; Mueckler and Thorens 2013). The presence of SLC transporters has been confirmed in various tissues, but to date there are only few studies examining the expression of SLC transporters in joint tissues. To our knowledge, to date, there are no studies investigating the expression of SLC transporters in periarticular fatty tissue, which plays an important role in the development of inflammation in RA patients. This study aimed to evaluate the expression of individual SLC family transporters in the SMs and periarticular fatty tissue (Hoffa’s pad) in patients with RA.

This study aimed to examine the expression of SLC family transporters in joint tissues: SMs and periarticular fatty tissue (Hoffa’s pad). To date, there are only few studies that have confirmed the constitutive expression of some SLC family transporters in SMs; however, according to our knowledge, there are no studies examining the expression of SLC transporters in periarticular fatty tissue, which plays an important role in the inflammatory response in RA patients. SLC transporters have been shown to play an important role in a number of physiological processes, but their expression has also been investigated to influence the concentration of various metabolites and drugs in cells and their involvement in the pathogenesis of various diseases, including RA and OA (Beckmann et al. 2015; Seki et al. 2017; Sohn et al. 2021). Figure 8 shows the molecules transported by this family of transporters. Previous studies indicated that the expression of some transporters may be increased in RA patients as compared with OA patients, suggesting the involvement of these transporters in transport of inflammatory mediators in RA (Xu et al. 2020).

We demonstrated the expression of 17 membrane transporters of the SLC family in the SM and periarticular fatty tissue (Hoff’s pad) of RA and OA patients. We showed differences in the expression of these transporters between RA and OA patients, as well as between the SM and periarticular fatty tissue. Moreover, the expression of some SLC transporters correlated significantly with disease activity parameters. These findings suggest that the inflammation present in RA patients may affect the expression of SLC transporters, which likely influences disease activity and progression. Differences in the expression of SLC transporters between SM and periarticular adipose tissue were also demonstrated. This confirms the results of previous studies showing that the expression of SLC transporters varies in different tissues and is tissue-specific (Hediger et al. 2013; Schlessinger et al. 2013).

RA is an autoimmune chronic inflammatory disease leading to the destruction of joints and the development of a number of extra-articular complications such as sicca syndrome and amyloidosis. The ongoing inflammatory process primarily involves the small joints of the hands and feet leading to their destruction. The inflammatory process has been shown to affect all joint structures, including the periarticular adipose tissue, which can be a source of many mediators that enhance and sustain the development of the inflammatory process and the destruction of joint structures (Kuca-Warnawin et al. 2011; Komatsu and Takayanagi 2022). However, it seems that the key site of ongoing inflammation is the SM. The presence of protein transporters in these joint structures is very important due to the transport of many mediators involved in the development of inflammation, the transport of various metabolites as well as drugs used in the treatment of RA. It is very important that the drugs used reach all tissues of the joint involved in inflammation. Previous studies have confirmed the presence of SLC family transporters in many tissues such as the liver, kidney, intestine, and brain tissue (Koepsell and Endou 2004). Knowing the localization of these transporters in synovial and periarticular adipose tissue can be important in selecting therapies for RA patients, as well as in understanding the processes that occur within these joint structures and lead to inflammation and joint destruction.

The important role of SLC transporters in the pathogenesis of RA, as well as the efficacy of RA therapy, seems to be indirectly confirmed by studies of the polymorphism of these genes in RA patients (Tokuhiro et al. 2003; Martínez et al. 2006; Lima et al. 2014; Eektimmerman et al. 2018; Pawlik et al. 2019; Wang et al. 2020). The results of our previous study suggest that the SLC22A5 polymorphism may be associated with the development of extra-articular manifestations in RA (Pawlik et al. 2019). Lima et al. (2014) have shown that SLC19A1, SLC46A1, and SLCO1B1 genotypes were associated with increased risk of MTX-related overall toxicity, whereas SLC19A1 and SLCO1B1 genotypes and SLC19A1 haplotypes were associated with increased risk of MTX-related gastrointestinal toxicity. Eektimmerman et al. (2018) have found the significant association between Methotrexate (MTX) toxicity and rs624249, and rs1060896. However, this association has not been confirmed in other study groups, in which no association was observed between methotrexate efficacy and SLC04A1, SLC22A2, or SLC28A2 variants (Eektimmerman et al. 2018). Wang et al. (2020) examined the association between the SLCO1A2 gene (G550A, G553A, A775C, and G862A) polymorphisms and MTX-related toxicity in RA patients. The A775C and G862A polymorphisms were not detected in RA patients, but the 550AA genotype was associated with a high risk of MTX adverse effects (Wang et al. 2020). In our study, we demonstrated the expression of 17 membrane transporters from the SLC family in the SMs and Hoffa’s pad of RA and OA patients. In addition, we showed the different expressions of some transporters in the SM and Hoffa’s pad of RA patients compared to OA. It has been indicated that the SLC transporters play an important role in regulating cellular metabolism, especially in the SM, affecting the development of inflammation in RA patients (Torres et al. 2022a, b). Differences in epigenetic factors between RA and OA patients have been shown to affect the expression of SLC transporters in the synovial fibroblasts (Torres et al. 2022a, b). This affects changes in cellular metabolism, which may also affect the immune response and development of inflammation in RA.

In our study, the presence of SLC transporters in RA and OA patients was detected at the mRNA level, as well as confirmed at the protein level by IHC. We have detected statistically significant differences in the expression of some transporters between RA and OA patients. The observed differences may be primarily due to the different pathogenesis of the two diseases or the influence of different epigenetic factors. Previous studies have also shown that the expression of SLC transporters can be modulated by a number of mediators and metabolites (Bonaventura et al. 2016; Bustamante et al. 2018; Amrhein et al. 2020; Gallagher et al. 2020). Regardless of the reason, however, the differences we observed in the expression of SLC transporters between patients with RA and OA suggest the involvement of these transporters in their pathogenesis, in metabolic processes, transport of pro-inflammatory mediators, or transport of drugs.

We also observed correlations between certain transporters and CRP, WBC, and DAS28 values, suggesting that the inflammation present in RA patients may both increase and decrease the expression of some SLC transporters. RA is a complex autoimmune disease that results from interactions between immune cells such as T and B lymphocytes and macrophages with cells in joint tissues (synovial cells, chondrocytes, or periarticular fatty tissue). This leads to the enhanced synthesis of a number of pro-inflammatory mediators, such as cytokines and chemokines, which increase inflammation within the joint tissues (Song et al. 2020). SLC transporters may be involved in the transport of these mediators, thus influencing the development of inflammation (Diao et al. 2010; Foster et al. 2013; Liu et al. 2015; Song et al. 2020; Mirdamadi et al. 2021). Pro-inflammatory mediators can affect the expression of some membrane transporters. This regulatory effect may be related to the influence of various inflammatory mediators, such as pro-inflammatory cytokines and chemokines (Bonaventura et al. 2016; Bustamante et al. 2018; Amrhein et al. 2020; Gallagher et al. 2020). Pro-inflammatory cytokines such as interleukin (IL)-1β, IL-6, tumor necrosis factor (TNF)-α, and interferon (IFN)-γ affect the expression of membrane transporters (Bonaventura et al. 2016; Bustamante et al. 2018; Amrhein et al. 2020; Gallagher et al. 2020). On the other hand, it has been shown that upregulated SLC7A5 activates mTOR-P70S6K signaling and enhances MMP3 and MMP13 expression in fibroblast-like synoviocytes from RA patients (Xu et al. 2020).

In our study, we demonstrated the expression of SLC transporters in the SMs and periarticular fatty tissue of patients with RA and OA. Our results confirmed the presence of SLC transporters not only in SMs, but also in periarticular fatty tissue, which plays an important role in the development of the inflammatory state in RA. We showed differences in the expression of these transporters between RA and OA patients. Regardless of the reasons for these differences, this suggests the possible involvement of certain transporters in the pathogenesis of these diseases, whether by affecting the transport of metabolites, pro-inflammatory mediators, or drugs. We also showed an association between the expression of some transporters and the parameters of clinical activity in RA. This may indicate the involvement of these transporters in the course of the inflammatory process and the influence of pro-inflammatory mediators on the expression of some transporters. However, conclusive confirmation of the involvement of SLC transporters in processes leading to the development of RA requires studies in cellular or animal models to assess signaling pathways and metabolic processes in which SLC transporters may be involved.

A limitation of our study is the lack of determination of their expression in healthy subjects and the lack of explanation of the role of SLC transporters in RA pathogenesis. SLC transporters are involved in numerous processes that may underlie the development of RA and affect the efficacy of treatment. However, explaining their involvement in RA pathogenesis requires numerous studies, especially in cellular models.

We hope that the confirmation of the expression of a number of SLC transporters in the SM of joints and periarticular adipose tissue will contribute to further studies aimed at explaining the role of particular SLC transporters in the pathogenesis of RA.

In conclusion, our results show the expression of 17 membrane transporters from the SLC family in the SMs and the periarticular fatty tissue (Hoffa’s pad) of RA and OA patients. To our knowledge, no studies have been conducted to date on the presence of expression of these transporters in periarticular fatty tissue. The confirmed expression of these transporters in the SMs as well as Hoffa’s pad of patients with RA and OA, and the differences in their expression between these groups, suggest the involvement of SLC transporters in both the maintenance of homeostasis under physiological conditions in the tissues of the joints, as well as in the inflammatory process in RA. It is likely that these transporters may also play a role in the efficacy of RA treatment. We hope that our results will prompt further studies aimed at understanding the role of SLC transporters in joint tissues under physiological conditions as well as in the pathogenesis of RA.