**4. Discussion**

The serum concentration of sRAGE seems to be modulated by a complex group of factors such as genetic factors, internal and environmental stimuli [27–33]. It has also been suggested that serum levels of sRAGE are influenced by gender, age, ethnicity, the imbalance between antioxidants and prooxidants and inflammatory processes [27]. There are studies which attribute to sRAGE the role of a potential biomarker of oxidative stress [34]. Some reports have suggested that the overexpression of sRAGE reflects the excessive inflammatory response involved in the progression of endothelial lesions and coagulopathy associated with severe infection. Low levels of sRAGE were associated with increased levels of IL-6, VCAM-1 and PAI-1 as well as with thrombocytopenia [35]. At the same time, in patients with type 1 diabetes, sRAGE overexpression in response to increased levels of AGEs was interpreted as a modality of protection against cell damage. Under these conditions, sRAGE acts as a negative feedback mediator for eliminating AGEs. The overexpression of sRAGE can be considered a weapon against cell damage and a mechanism to regulate the receptor synthesis by modulating the synthesis of enzymes that produce a proteolytic cleavage [36].

In our study, we have found that patients with palmoplantar warts had lower serum levels of sRAGE compared to controls. sRAGE downregulation may be a factor involved in HPV pathogenesis; it can be speculated that sRAGE acts as a negative regulator of warts occurrence and could represent an early mediator involved in the onset and development of warts. The decrease in sRAGE levels in patients with palmoplantar warts could be explained by different mechanisms. HPV induces increased proliferation of keratinocytes resulting in a higher rate of glucose metabolism in the infected cells, which stimulates the synthesis of AGEs. Thus, AGEs accumulate in extracellular spaces and interact with sRAGE [37]. Another possible explanation is the disruption of the AGEs-sRAGE axis that might induce a low synthesis of soluble receptors [18,36,38]. sRAGE is cleaved on cell surface through the action of matrix metalloproteinases. The activity of these enzymes is modulated by oxidized lipoproteins [32]. It has also been reported that advanced glycosylation of high density lipoproteins leads to endogenous sRAGE sequestration [32,33].

The relationship between RAGE expression in the skin and the level of its ligands remains unclear. In human skin, sRAGE was positively correlated with the expression of genes encoding for ligands of RAGE such as tumor necrosis factor (TNF) alpha, IL-1 alpha, S100B, proapoptotic factors (Fas, Bax), epidermal differentiation markers (involucrin), and proliferating cell nuclear antigen [17]. Another factor that could influence sRAGE activity is the presence of a group of cell surface receptors, AGE-R1, AGE-R2, and AGE-R3, which seem to modulate the endocytosis and degradation of AGEs, thus counteracting the effects of RAGE. AGE-R1 has been shown to reduce oxidative stress induced by AGEs through the inhibition of RAGE signaling pathway [37].

Another point analyzed in our study was the investigation of potential mechanisms by which sRAGE is involved in the pathogenesis of warts. To demonstrate this hypothesis, we have performed a complete, simultaneous and comparable analysis of the axis sRAGE – markers of oxidative stress – markers of inflammation in patients with warts and in a control group. First, we have investigated oxidative stress markers (TOS, TAS, OSI) and confirmed the presence of an imbalance between oxidant load and antioxidant defense in patients with warts. Previous studies have suggested that the balance between oxidants and antioxidants plays an important role in the spontaneous regression of HPV infection, and the antioxidant system prevents the effects of oxidative stress and mediates the immune response [39,40]. A recent study has shown that oxidative stress plays an important role in recalcitrant warts [41]. Excessive amounts of oxidants lead to destructive effects, materialized in the structural and functional alteration of lipids, proteins and nucleotides [42]. In this case, the antioxidant systems may become deficient, favoring the perpetuation of oxidative stress. We consider that sRAGE can participate in the restoration of the oxidant/antioxidant balance in patients with warts. This hypothesis is supported by the strong negative correlation between sRAGE and TOS, respectively OSI, and the positive association between sRAGE and TAS. Based on these results, we assign to sRAGE the role of a potential biomarker of oxidative stress in patients with warts. Therefore, the modulation of sRAGE level in HPV patients might influence the progression of the disease.

In our study, the evaluation of a panel of markers of inflammation did not reveal an inflammatory systemic process in patients with warts. The sRAGE level was not correlated with the levels of the markers of inflammation (IL-6, fibrinogen and ESR) excepting hs-CRP. However, we do not exclude the presence of a proinflammatory environment in infected tissues. The association between sRAGE and hs-CRP has also been proven in several pathological conditions [43]. CRP is synthesized by hepatocytes in response to TNF alpha, IL-1, and IL-6 [44]. The AGEs-RAGE interaction increases the expression of these cytokines; sRAGE and RAGE compete for the same ligands. As a result, low sRAGE levels increase the AGEs-mRAGE interaction, which leads to increased cytokine production. It is known that sRAGE modulates the synthesis of hs-CRP in patients with acute coronary syndrome [45].

We have identified that none of the examined parameters was influenced by the extension of warts and disease duration. Our results are in concordance with the study by Sasmaz et al., which has also revealed that markers of oxidative stress (catalase, glucose-6-phosphate dehydrogenase, superoxide dismutase, and malondialdehyde) did not correlate with the duration and the number of the lesions [46]. We consider that the level of sRAGE cannot be used as a biomarker for the severity of warts. The molecular mechanisms by which sRAGE could be involved in the etiopathogenesis of warts are complex and could include the interference between oxidative stress and inflammation.

In our study, we have shown changes of serum levels of sRAGE in patients with palmoplantar warts compared to the control group. Given that warts are produced by HPV we have suggested a possible role of sRAGE in the pathogenesis of HPV infection. Further studies investigating the presence of the virus, its type, and its viral load in the examined patients are needed, in order to establish the exact role of sRAGE in HPV infection. Our findings open new perspectives and pave the way for the investigation of sRAGE in HPV infection.

Currently, there is no data available in the literature on the implication of sRAGE in the deep mechanisms that mediate the appearance and evolution of warts. These findings could help broaden the therapeutic options for HPV lesions. Some studies have shown that sRAGE could be an effective therapeutic target and might be used as a biological agen<sup>t</sup> [21]. It has been suggested that increased concentrations of sRAGE may contribute to the inhibition of the inflammatory signaling pathways [47].
