The Role of Epigenetic Factors in Psoriasis

Psoriasis is a chronic, systemic, immune-mediated disease with an incidence of approximately 2%. The pathogenesis of the disease is complex and not yet fully understood. Genetic factors play a significant role in the pathogenesis of the disease. In predisposed individuals, multiple trigger factors may contribute to disease onset and exacerbations of symptoms. Environmental factors (stress, infections, certain medications, nicotinism, alcohol, obesity) play a significant role in the pathogenesis of psoriasis. In addition, epigenetic mechanisms are considered result in modulation of individual gene expression and an increased likelihood of the disease. Studies highlight the significant role of epigenetic factors in the etiology and pathogenesis of psoriasis. Epigenetic mechanisms in psoriasis include DNA methylation, histone modifications and non-coding RNAs. Epigenetic mechanisms induce gene expression changes under the influence of chemical modifications of DNA and histones, which alter chromatin structure and activate transcription factors of selected genes, thus leading to translation of new mRNA without affecting the DNA sequence. Epigenetic factors can regulate gene expression at the transcriptional (via histone modification, DNA methylation) and posttranscriptional levels (via microRNAs and long non-coding RNAs). This study aims to present and discuss the different epigenetic mechanisms in psoriasis based on a review of the available literature.

Due to its pathomechanism, psoriasis is considered to be a systemic disease in which levels of multiple proinflammatory interleukins are elevated. Approximately 30% of psoriatic patients develop psoriatic arthritis. The disease is also associated with increased cardiovascular risk and frequent comorbidities such as obesity, metabolic syndrome, nonalcoholic fatty liver disease, diabetes, inflammatory bowel disease and depression [4,5].
The pathogenesis of the disease is complex and not yet fully understood. Activated plasmacytoid dendritic cells secrete Interferon-alfa (IFN-alpha), which activates myeloid dendritic cells and proinflammatory cytokines including IFN-gamma, TNF-a, IL-1b and IL-6. Subsequently, stimulated myeloid dendritic cells produce IL-12 and IL-23, which results in activation of Th1 and Th17 helper T-cells and synthesis of further cytokines such as TNF-a, IL-17A, IL-17F and IL-22.Novel biological drugs used on the management of this condition aim to block selectively some of these cytokines [6,7]. This is followed by activation of keratinocytes and further stimulation of cytokines, chemokines and antimicrobial peptides, resulting in increased inflammation [3,8].
Genetic factors play a significant role in the pathogenesis of the disease. Psoriasis is more common in first-and second-degree relatives, and the risk of developing the disease in monozygotic twins is 2-3 times higher compared to dizygotic twins [3,9]. Multiple gene loci responsible for psoriasis susceptibility have been described and named PSORS1-PSORS10 (psoriasis susceptibility locus 1-10). The strongest linkage to psoriasis is shown by the PSORS1 locus, located within a 220 kb segment of the major histocompatibility complex on chromosome 6p21 [9][10][11]. The major allele associated with the development of psoriasis is the HLA-Cw * 06 allele, which is associated with early onset and severe course of the disease [8,9] In predisposed individuals, multiple trigger factors may contribute to disease onset and exacerbations of symptoms. Environmental factors that play a significant role in the pathogenesis of psoriasis include stress, infections, certain medications, nicotinism, alcohol consumption, and obesity [12,13]. Given the complex etiopathogenesis of psoriasis, in which genetic and environmental factors play a role, epigenetic mechanisms are considered result in modulation of individual gene expression and an increased likelihood of the disease [14,15]. Studies highlight the significant role of epigenetic factors in the etiology and pathogenesis of psoriasis. Epigenetic mechanisms induce gene expression changes under the influence of chemical modifications of DNA and histones, which alter chromatin structure and activate transcription factors of selected genes, thus leading to translation of new mRNA without affecting the DNA sequence [14]. Epigenetic factors can regulate gene expression in several mechanisms at the transcriptional (via histone modification, DNA methylation) and posttranscriptional levels (via microRNAs-miRNAs and long non-coding RNAs-lncRNA) ( Figure 1). Recent studies highlight the role of epigenetic processes in inflammatory diseases, including psoriasis [15]. This study aims to present and discuss the different epigenetic mechanisms in psoriasis based on a review of the available literature. The list of the studies of epigenetic mechanisms in psoriasis is stored as Table 1.

Searching Strategy
Electronic literature searches were performed in the PubMed, medRxiv, Medline, Scopus, Cochrane Library, and the Cochrane Central Register of Controlled Trials databases. The articles for this review were published between September 2004 and April 2021. The following search phrases were used: "(Psoriasis) AND ((Epigenetics) OR (DNA methylation) OR (non-coding RNA) OR (long non-coding RNA) OR (microRNA) OR (histones)) as key words or MeSH terms. A number of 780 results records were found through database searches. Duplicates and non-English language articles were excluded.
Full-text articles were screened independently by two authors (K.D. and P.C.) for the following inclusion criteria: experimental studies characterizing epigenetic mechanisms, differences between subjects with psoriatic disease and a control group, studies measuring the activity or expression of a component of the epigenetic machinery in subjects with psoriatic disease.
From 435 potentially appropriate articles that underwent evaluation, 107 full-text articles were found to be suitable for analysis (hypothesis articles, publications with duplicated patients, clinical reports, non-epigenetic marks, and non-psoriatic skin diseases were excluded).

DNA Methylation
One of the crucial mechanisms promoting chromatin condensation, and thus epigenetic gene silencing, is DNA methylation [16]. The DNA methylation pattern can be duplicated during replication of the genome causing this regulation to be permanent. DNA methylation involves covalent bonding of methyl groups to cytosine within CpG islands in the 5 region of the selected gene, the product of which is 5-methylcytosine (5-mC).
Cytosine methylation is the most common post-replication epigenetic modification of DNA. This reaction is catalyzed by DNA methyltransferases (DNMT, deoxyribonucleic acid methyltransferase). Increased methylation of CpG islands, which are present in promoter regions of the gene, results in restricting access of transcription factors to DNA, and consequently reducing expression and silencing of gene function [17,18]. DNA methylation is particularly important for the regulation of developmental and tissue-specific gene expression. Studies assessing DNA methylation in psoriasis have evaluated the degree of methylation at a global level, i.e., the total 5-mC content of the genome, or for specific genes, where levels of methylated cytosines located in a specific gene or promoter were determined [18]. In several studies deregulation of DNA methylation has been observed at a global level in lesional psoriatic skin compared to healthy skin [19][20][21].
In the genome-wide study of alternated CpG methylation in psoriatic lesions, Roberson et al. analyzed the methylation status of 27,578 CpG sites in skin samples from patients with psoriasis and skin samples from healthy controls. The results showed that CpG methylation of psoriatic lesions differed from control skin at 1108 sites-twelve mapped to the epidermal differentiation complex, upstream or within genes that are highly upregulated in psoriasis. The investigators analyzed 50 of the top differentially methylated (DM) sites separated psoriatic from control skin samples with uninvolved skin exhibiting intermediate methylation. The results revealed a subset of DM CpG sites that correlated significantly with the differential expression of nearby genes including those of KYNU, OAS2, S100A12, and SERPINB3, whose strong transcriptional upregulation is important in psoriasis. Moreover, the study revealed return of methylation levels toward the non-psoriatic state after one month of anti-TNF-a therapy [19].
Zhang et al. demonstrated overall hypermethylation in both lesional psoriatic skin and PMBCs (peripheral mononuclear blood cells) compared to the control group. In addition, the overall level of 5-mC evaluated in skin lesions positively correlated with disease severity as assessed based on PASI, but no such correlation was shown for PBMC. This study also demonstrated increased expression of DNMT1 in psoriasis patients compared to the control group, while methyl-DNA binding domain genes MBD2 and MeCP2 were significantly downregulated in peripheral mononuclear blood cells in psoriasis patients compared to the control group [20]. In another study, Zhang et al. also observed differences in genome-wide DNA methylation profiling in active psoriatic lesions and skin without lesions in patients with psoriasis compared to healthy skin based on methylated DNA immunoprecipitationsequencing. The results showed that the promoter methylation levels of PDCD5 and TIMP2, both of which were related to proliferation of keratinocytes, were increased significantly and negatively correlated with mRNAs expression in psoriatic skin compared with normal controls [22]. In a study by Chandra et al., the enrichment of differentially methylated CpGs in several psoriasis susceptibility regions (PSORS) was observed with the use of genomic DNA methylation profiling in psoriasis patients (top differentially methylated genes overlapped with PSORS regions including S100A9, SELENBP1, CARD14, KAZN and PTPN22). An inverse correlation between methylation and expression of these genes has been observed. This study also demonstrated an interesting correlation between differentially methylated genes and histopathological abnormalities typical of psoriasis (Munro's microabscesses, parakeratosis, neutrophil infiltration) [23].
Another study of alternated DNA methylation in psoriasis involves SHP-1, which is a regulator in growth and proliferation processes. The study showed significant hypomethylation of promoter 2 of the SHP-1 locus in psoriatic skin samples compared to the skin from healthy controls [24]. Furthermore, the hypomethylation of the promotors of the genes involved in the regulation of the cell cycle such as p15, p21, p16 was observed in hematopoietic stem cells of psoriatic patients [25,26]. On the other hand, the inhibitor of differentiation 4 (ID4) locus was hypermethylated in skin samples from patients with psoriasis, eczema, and squamous cell carcinoma, demonstrate that hypermethylation at this locus is a general characteristic of parakeratotic skin diseases, but not specific for psoriasis [27]. In addition, it has been observed that the promotor of the other negative regulator of the pathway involved in cell proliferation and differentiationsecreted frizzled related protein 4 (SFRP4) is hypermethylated in psoriatic lesions [28].

Non-Coding RNA
Genome-wide analysis studies have identified multiple chromosome loci associated with psoriasis [22]. However, most of the signals in genome-wide association studies (GWAS) are found in non-coding regions of the human genome [18,[29][30][31]. The results of several studies show that non-coding regions of DNA play a significant role in the genetics and epigenetics of many diseases, providing a solution to the "missing heritability" problem.
Psoriasis is caused by excessive proliferation of keratinocytes and impaired interactions between keratinocytes and T lymphocytes. Numerous studies have demonstrated that patients with psoriasis have significantly higher levels of miRNA expression than healthy individuals and that these molecules may be involved in the pathogenesis of psoriasis. It was found that under disease conditions, miR-146a, miR-203, miR-21, miR-31, miR-184, miR-221 and miR-222 were upregulated, whereas miR-99a, miR-424 and miR-125b could be downregulated. Thus, miRNA may become an important target for psoriasis treatment [32,[36][37][38][39].

miRNAs in Psoriasis
4.1.1. Upregulation miRNA-146a, which is highly expressed in keratinocytes of psoriasis-affected skin, has been identified as a negative regulator of the innate immune response and inflammation. This miRNA was found to affect TNF-receptor associated factor 6 and IL-1 receptorassociated kinase I (IRAK-1) [40,41], two major mediators involved in the production of pro-inflammatory cytokines, and thus inhibit the immune response. Hence it follows that silencing miR-146a may help reduce inflammation, and miR-146a inhibitors may be effective in alleviating the symptoms of psoriasis [40,42].
The role of miR-146b in the skin is not fully understood. The results of a study by H. Hermann et al. confirmed increased expression of miR-146a and miR-146b (miR-146a/b) in psoriatic lesions. In healthy human skin, miR-146a expression was approximately twice as high as miR-146b and was upregulated by proinflammatory cytokines in keratinocytes and fibroblasts. In the skin of patients with psoriasis, target genes for miR-146a/b that regulate inflammatory responses or proliferation were altered, with FERMT1 identified as a direct target of miR-146a. Transfection of miR-146a/b inhibited keratinocyte proliferation and expression of psoriasis-related target genes, whereas inhibition of these microRNAs enhanced the above processes [40,43].
Furthermore, increased expression of miRNA-203 was observed in psoriasis-affected keratinocytes. The increase in miR-203 is consistent with the downregulation of suppressor of cytokine signaling-3 (SOCS-3) and subsequent upregulation of signal transducer and activator of transcription-3 (STAT-3) [44,45]. As a result, the interaction between cytokines and keratinocytes became more intense. The above properties suggest the possibility of using miRNA-203 inhibitors in the treatment of psoriasis [46].
Feng et al. developed a reconstituted high-density lipoprotein nanocarrier gel with antisense miR-210 and evaluated its efficacy in a mouse model of imiquimod-induced psoriasis. The results indicated that the expression of miR-210 was decreased in both skin lesions and CD4+ T lymphocytes. Additionally, it resulted in a reduction in erythema, scaling, hyperkeratosis and inflammatory cell infiltration. Levels of interleukin-17A mRNA, c-interferon, and the percentages of Th-1 and Th-17 lymphocytes were found to be decreased in skin and spleen cells [47]. MiR-210 has been shown to inhibit Th2 differentiation by suppressing the expression of STAT6 and LYN, resulting in immune imbalance during psoriasis development [48]. By inhibiting miR-210, antagomir -210 reduces inflammation and resolves the immune imbalance associated with psoriasis-like inflammation caused by imiquimod or IL-23. TGF-and IL-23 increased miR-210 expression through induction of HIF-1, which is responsible for histone H3 acetylation in the miR-210 promoter region.
In conclusion, the study by Feng et al. demonstrated that miR-210 inhibition effectively reduces psoriasis-like inflammation, suggesting that this may be a potential therapeutic target [47,48].
Yan et al. investigated the role of miR-145-5p in psoriasis skin lesions. The study revealed that miR-145-5p overexpression is required to inhibit proliferation of normal human epidermal keratinocytes and chemokines secretion. miR-145-5p regulates the function of nuclear factor B and STAT-3 by targeting a mixed-coupling kinase. The study demonstrates that a decrease in miR-145-5p level results in epidermal proliferation and psoriasis-like inflammation [49].
miR-21 is a pro-inflammatory oncogene that inhibits apoptosis. Expression of miR-21 is increased in psoriatic skin lesions due to T-cell infiltration. Additionally, inhibitors of miR-21 have effects on non-malignant T-cells, which can be used to modify activated T-cells to treat psoriasis [50]. In addition, miR-21-5p, a critical regulator of epidermal inflammation, inhibits tissue inhibitor of metalloproteinase-3 (TIMP-3). This reduction in TIMP-3 levels results in increased levels of tumor necrosis factor (TNF) converting enzyme (TACE) and increased TNF release, leading to epidermal inflammation [51].
miR-31 is a miRNA with high expression in keratinocytes. Xu et al. showed that it acts by inhibiting serine/threonine kinase 40 and protein phosphatase 6. Serine/threonine kinase 40 is involved in controlling NfkB-I signaling. Thus, miR-31 disrupts the normal Nf kB-I signaling mechanism, as well as IL-8 and IL-1β production. According to Xu et al., miR-31 inhibition can reduce inflammation [52].
It is worth noting that there was a significant correlation between miR-155 expression levels in peripheral blood mononuclear cells and PASI score at the beginning of the study, and that a difference in the expression of this miRNA was shown in samples at the beginning of treatment and after successful psoriasis therapy [56].

Down-Regulated miRNAs in Psoriasis
A study by Meng Z. showed that the expression of miR-99a and miR-424 (which regulate epidermal homeostasis through direct inhibition of mRNA targets) is decreased in psoriatic lesions [50]. Huang R.Y. found that miR-125b was one of the most downregulated miRNAs in these lesions. This provides a basis for further research on activators of the above miRNAs, which may be effective drugs for the treatment of psoriasis [44].
Xu et al. found that miR-125b is downregulated in psoriatic lesions. Overexpression of this miRNA in primary human keratinocytes inhibits proliferation and upregulates multiple markers of differentiation. Biochemical studies showed that miR-125b binds to FGFR2 [57]. It has also been shown to interact with BRD4 and inhibit Jagged-1 ligand expression. In addition, miR-125b reduces Notch signaling, inhibiting "psoriatic cell" proliferation [58].
Let-7b expression is also decreased in the skin of patients with psoriasis. This miRNA is responsible for inhibiting hyperkeratosis and reducing disease activity during treatment with imiquimod. Furthermore, inhibition of IL-6 by Let-7b promotes normal differentiation of keratinocytes [59].
In previous studies, the role of miRNAs in psoriasis has been best defined in psoriasis vulgaris. There are still very few reliable data on the importance of these molecules/structures in other types of psoriasis, such as psoriatic arthritis, inverse psoriasis, or pustular psoriasis. Although much progress has been made in identifying the miRNA responsible for psoriasis, there is still a need for research into the use of miRNAs in the diagnosis, treatment and prevention of the disease.

Long Non-Coding RNA (lncRNA)
To date, the complex mechanisms underlying psoriasis have not been fully elucidated. Long noncoding RNAs (lncRNAs) are defined as non-protein-coding RNA transcripts greater than 200 nucleotides in length [60] that may be involved in pre-transcriptional regulation such as histone modification and DNA methylation as well as transcriptional regulation regarding enhancer activities, transcriptional interference, regulatory transcription factors, variable splicing, competing endogenous RNA (ceRNA), other post-transcriptional regulation, and regulation of protein translation [61,62].
lncRNAs play a role in epigenetic silencing, splicing regulation, translation control, apoptosis, and cell cycle regulation. Additionally, the expression levels of various lncRNAs are strongly correlated with epidermal differentiation and immunoregulation. Numerous examples demonstrate that long non-coding RNAs also regulate a variety of skin pathologies, including skin cancer, wound healing, or psoriasis [32][33][34]63,64]. Tsoi et al. analyzed lncRNA expression in lesional and non-lesional psoriatic skin, identifying 4022 skin-specific lncRNAs (including 1080 previously unknown ones). Their findings indicate that a large number of lncRNAs, particularly those with differential expression, are coexpressed with genes involved in the immune response. In addition, the authors identified novel lncRNAs with different tissue-specific expression patterns, including those involved in epidermal differentiation [32].
In conclusion, the results of studies by Tang L. et al. and Tsoi l. show that numerous lncRNAs play a role in the immunopathogenesis of psoriasis [32,33].
Two important lncRNAs, ANCR and TINCR, are involved in the control of epidermal differentiation. ANCR (antidifferentiation non-coding RNA) downregulates epidermal differentiation. Lack of ANCR in progenitor cells rapidly initiates the differentiation program, thus its presence is necessary to inhibit premature differentiation of epidermal basal layer cells. In comparison, TINCR (terminal differentiation-induced non-coding RNA) is abundant in the differentiated layers of the epidermis and promotes keratinocyte differentiation [34].
PRINS (Psoriasis Associated Non-Protein Coding RNA Induced By Stress) is an lncRNA that is upregulated in both affected and non-affected psoriatic epidermis. Szegedi et al. demonstrated that PRINS silencing alters the morphology and gene expression profile of cells. PRINS regulates the anti-apoptotic gene G1P3 in keratinocytes. The expression of G1P3 is significantly increased in epidermal cells from patients with psoriasis (in both lesional and non-lesional epidermis). The results of this group of researchers showed that dysregulation of the PRINS gene can reduce the susceptibility of keratinocytes to spontaneous apoptosis through modulation of G1P3 [65].

Histones
Historically, histone proteins were thought to function solely as DNA packaging materials, with no role in gene expression. However, in the 1990s, modifications of histones were proven to play an important role in the epigenetic regulation of human cells. In eukaryotic cells, each histone octamer is wrapped around 147 DNA base pairs to form a nucleosome, which is the basic subunit of chromatin. Histone modifications have so far been studied mainly in cancer. However, there are numerous reports on their role in dermatological diseases, inter alia, in psoriasis. Psoriasis is associated with abnormal expression of histone acetyltransferases (HATs) and histone deacetylases (HDACs), which regulate the balance between histone acetylation and deacetylation [66,67].
Histone methylation is usually observed on the lysine and arginine side chains, and multiple methyl groups can be added to histones: mono-, di-, and tri-methylation are observed [68].This process can result in an active or suppressed transcriptional state, and the outcome depends on both the methylation site and the number of methyl groups added [69]. Histone methylation plays a crucial role in psoriasis in terms of regulation of cytokine production and drug response [70]. A study by Li H. et al. showed that H3K9me2 regulates IL-23 expression in keratinocytes and that keratinocyte-derived IL-23 is sufficient to induce the psoriasis phenotype in a mouse model of psoriasis [71].
A group of researchers led by Ovejero-Benito MC found that among psoriasis patients, H3K4 methylation is increased in PBMCs (Peripheral Blood Mononuclear Cells) compared to the control group, which may contribute to differential gene expression in PBMCs. Moreover, the researchers found differences in H3K4 and H3K27 methylation levels among patients on biological drug therapy (ustekinumab, secukinumab, adalimumab, ixekizumab), depending on the effectiveness of the drugs used. This raises the possibility of using markers of histone methylation as biomarkers of treatment response [72].
The results of a study by Zhang T. showed an increase in the activity of histone H3K27me3, enhancer of zeste homolog 2 (EZH2), and histone H3K27 methylase in psoriatic epidermis [73].
In another study, researchers from China [74] described hypoacetylation of global histone H4 in PBMC cells of psoriasis patients compared to healthy control groups. Additionally, a negative correlation was observed between histone H4 acetylation and disease activity as measured by the PASI score. The mRNA levels of P300, CBP and SIRT1 were significantly reduced in PBMC of patients with psoriasis compared to healthy controls, whereas the mRNA levels of HDAC1, SUV39H1 and EZH2 were significantly elevated [74].  ET-1 is made by psoriatic keratinocytes and inhibits apoptosis.
Inflammation increases the production of ET-1, which in turn results in the chronic induction of keratinocyte proliferation

Conclusions
Despite significant progress in the diagnosis and treatment of psoriasis, many questions about the pathogenesis and prevention of the disease remain unanswered. In recent years, more than 60 psoriasis susceptibility loci have been discovered [104][105][106][107]. However, the identified genes explain only 28% of the heritability of psoriasis, suggesting the existence of additional, as yet unidentified, sources of heritability [32]. Epigenetic modifica-

Conclusions
Despite significant progress in the diagnosis and treatment of psoriasis, many questions about the pathogenesis and prevention of the disease remain unanswered. In recent years, more than 60 psoriasis susceptibility loci have been discovered [104][105][106][107]. However, the identified genes explain only 28% of the heritability of psoriasis, suggesting the existence of additional, as yet unidentified, sources of heritability [32]. Epigenetic modifications passed from generation to generation may be partly responsible for this discrepancy. In addition, different environmental exposures can induce somatic epigenetic modifications, altering an individual's susceptibility to the disease.
Psoriasis-specific epigenetic regulation may represent novel therapeutic targets and serve as a potential biomarker for diagnosis and treatment monitoring. However, this requires a better understanding of the epigenetic changes associated with psoriasis. There is still a lack of studies based on multicellular models, in vivo studies, and combination therapies using epigenetic targeting combined with standard treatment of psoriasis. A better understanding of the epigenetic mechanisms underlying the pathogenesis of psoriasis will accelerate and facilitate the future use of epigenetic modifiers in the treatment of psoriasis and may also contribute to the development of effective methods for prevention of this disease.