Monosodium urate crystals induce oxidative stress in human synoviocytes
The current study revealed that MSU crystals are able to decrease cell viability through apoptosis induction in FLS. Although the definitive mechanism for MSU-induced apoptosis has not been established, it has been demonstrated that MSU crystals exert different apoptotic effects depending on the cell type interacting with the crystals. While some studies have reported that MSU crystals inhibit neutrophil apoptosis [20], others have shown that they do not induce any change in the percentage of apoptosis for osteoblast-like cells [21]. Recently, MSU crystals have been shown to promote renal cell apoptosis through a mechanism involving ROS generation [22]. However, no data were available on their influence on FLS. According to one report, apoptosis is induced in chondrocytes isolated from patients with RA following stimulation with MSU crystals [23]. The association of this apoptotic state with the loss of cartilage repair and regeneration capacity could highlight a link between FLS apoptosis and the tissue damage observed in gouty patients. Moreover, the relationship between the increment of ROS and NO and the loss of FLS viability caused by MSU crystals is consistent with published findings [24–26].
In addition, we established that crystal-exposed FLS produced H2O2, O2– and, to a lesser extent, NO, promoting a state of cellular oxidation. One mechanism involved in ROS production is the NADPH oxidase system in THP-1 cells stimulated with MSU crystals [27]. This mechanism of ROS generation has also been shown in FLS from patients with OA and RA that were exposed to TNF-? and IL-1?, exhibiting a heightened state of cellular oxidation [28]. Our experiments proved, via an increase in ROS/RNS, that MSU crystals activated an oxidative state in FLS. The increase in H2O2 observed in FLS exposed to MSU crystals for 24 h is similar to that reported for FLS stimulated with advanced oxidation proteins products; a threefold to eightfold increase in H2O2 was observed compared to unstimulated control cells [29]. This suggests that MSU crystal-mediated ROS overproduction in FLS is involved in the disturbance of homeostasis within the joint microenvironment, which can damage all cellular components, including DNA, lipids and proteins [30]. However, proteins are possibly the most immediate vehicle for inflicting oxidative damage on cells because they are often catalysts. Therefore, we assessed the influence of ROS in oxidized protein content of FLS affected by MSU crystals.
The impact of ROS on the proteins of FLS with MSU crystals was clearly seen on images because there were more spots and with higher intensities than in control cells, indicating increased carbonyl content. While there are no reports that can be directly compared to our data, accumulation of protein carbonyls [31] has been observed in some rheumatic diseases (including RA and psoriasis), but it is known that exposure of proteins to ROS leads to denaturalization, loss of function, crosslinking, aggregation, and fragmentation. Under these conditions, it is suggested that accumulation of some compounds in the joint, like glycosaminoglycans and hyaluronic acid, cause damage by reducing joint viscosity [32]. However, there are no studies of the underlying mechanism. We suspected that the increase in OS might be contributing to synovial cell damage altering the functional and structural integrity. Therefore, we visualized OS-induced ultrastructural changes triggered by MSU-crystals in gout. In our model, we observed an increase in rough ER and in the presence of MP aggregates due to cellular stress in the FLS. These findings are similar to those described for synoviocytes exposed to an adjuvant used for treating arthritis (i.e., a reduction of the Golgi apparatus, mitochondria and ER [33]), and to the ones describing the appearance of vacuoles in FLS cytoplasm due to the internalization of particles. In addition, intracellular lysosomes and other cytoplasmatic formations were found [34], and these morphological changes suggest the induction of autophagy in the cells [35].
An important unanswered question is the mechanism responsible for activating OS in response to MSU crystals in FLS. We can speculate that this effect might be related to the mechanism involved in the overproduction of ROS and the decrease of anti-oxidative enzymes caused by lead-induced OS [36]. However, the molecular pathways involved in MSU-induced OS in FLS are not yet completely understood. Inhibition studies of these pathways may be helpful to understand the signaling network behind MSU crystals.