Strong evidence has highlighted the contribution of both genetic and epigenetic factors, which interact with environmental ones, in the pathogenesis of endometriosis and the formulation of the complex phenotype of the disease. This topic has been extensively analyzed and reviewed in various studies conducted by our group and others (11,18,41-44). Briefly, twin studies demonstrated that the sibling genetic relative risk is 2.3 and the estimated overall heritability has a rate of 51% (7,45). Despite the endometriosis-associated loci detected by independent gene association studies, the hypothesis-free approach of Genome Wide Association Studies (GWAS) as well as the high-throughput next generation sequencing (NGS) techniques offer valuable assistance in the efforts concentrated on identifying numerous novel disease susceptibility loci, although the exact functional relevance to endometriosis of most of these single nucleotide polymorphisms (SNPs) remains elusive. Notably, the largest trans-racial GWAS and replication meta-analysis of endometriosis performed to date, managed to identify 31 novel loci, in addition to the 11 previously known, which are involved in progesterone resistance, inflammatory processes, cell cycle regulation and angiogenesis (44). It has been reported that common genetic variation accounts for ~26% of the risk of the disease (46). Notably, the majority of these disease-associated SNPs, revealed by an increasing rate over the last years, confer a moderate effect in endometriosis and, according to the function of the respective proteins, were found to be involved in the immune system and autoimmunity, estrogen-induced cell growth, cell adhesion, differentiation, proliferation, apoptosis, matrix remodeling and oxidative stress (43,47-50). The importance of the epigenetic modifications, which play a pivotal role in pathogenesis of endometriosis, has been described in detail in recent studies from our group (5,51,52).

PS is a multifaceted, complex genetic disease, resulting from an interplay between genetic and environmental factors, as aforementioned. This has been demonstrated by studies involving monozygotic and dizygotic twins, where it was shown that monozygotic twins appear to have a 3-fold increased risk for the disease when compared to the dizygotic twins, while the concordance rate among monozygotic twins was never 100%, thus suggesting a role for environmental factors (53,54). Notably, the overall genetic variation accounts for 50-70% of PS susceptibility (55). Furthermore, a familial recurrence has been well established as well, considering that the incidence of the disease is greater among first- and second-degree relatives of patients than among the general population (56). Currently, advances in large-scale analysis of the human genome provide valuable insights into the genetic dissection of PS, with over 40 genomic regions of the genome and >400 SNPs associated with the disease identified through GWAS and/or NGS approaches (57-60). Most of the identified PS susceptibility genes are skin-specific, mediate epidermal differentiation, are involved in neo-vascularization or are related with adaptive immunity (61). Moreover, many of these genes are involved in antigen presentation, type 1 IFN pathways, T-cell polarization axis, negative regulation of immune response, inflammatory processes and innate immunity (57,62,63). Recently, Patel et al (64) classified the PS-associated genes into categories reflecting the pathway that they participate in and influence. These categories include interleukins, interleukin receptors, human leukocyte antigen (HLA) genes, nuclear factor-κB (NF-κB)-pathway related genes, IFNs, genes participating in the Janus κinase (JAK)-signal transducer and activator of transcription (STAT) pathway. Additional categories include genes linked to hypercholesterolemia and hypertriglyceridemia, negative regulators of T-cell activation, inflammatory markers, and nuclear hormone receptors (64).

PS and PsA are interrelated disorders given that most patients with PsA also have PS. However, PsA represents a distinct entity having its own clinical, genetic and epidemiological features. Notably, PsA exhibits a greater heritability among first-degree relatives in comparison with PS (40), thus suggesting a marked difference regarding the genetic architecture of the two disorders and indicating a larger genetic component for PsA (65). A stronger association with PsA than PS has been observed for various loci, including HLA-B*27 (66), CSF2 (67), PTPN22 (68), KIR (69), IL-13 (70), IL-12B (71), ZNF816A (71), IL23R (57), TNFAIP3 (57), IFNLR1 (57), IFIH1 (57), NFKBIA (57), MIF (72), IL-1A (40) and some variants in LCE3A (39).

The TNF-α gene, consisting of four exons and mapped to 6p21, is a member of the class III MHC genes (97). It codes for the TNF-α cytokine, a key regulator of both autoimmunity and inflammation (98), and it has been correlated with the pathogenesis of various autoimmune, inflammatory and infectious diseases (99). The TNF-α-308 G/A rs1800629 SNP, located in the promoter region of the TNF-α gene, has been associated with an increased susceptibility to severe-stage endometriosis (74) as well as PS with PsA (100,101). It is a well-studied polymorphism from the functional viewpoint, and the presence of minor allele ‘A’ was reported to enhance the transcription levels of TNF-α, compared with allele ‘G’, thus leading to the procuction of higher protein levels in serum (102,103). Notably, this SNP has also been shown to modify the consensus sequence of the AP-2 transcription factor-binding site (104). The role of this polymorphism in endometriosis has been highlighted by the increased levels of TNF-α detected in peritoneal fluid as well as the upregulation of TNF-α gene in peritoneal macrophages and peripheral blood monocytes (105,106). Moreover, high levels of TNF-α have been correlated with the severity of endometriosis (107), while the implantation of ectopic endometrial tissue is facilitated by elevated levels of TNF-α (108). All these findings underscore the important role of this SNP in the development of endometriosis. With regard to PS, the presence of ‘A’ at position-308 in the TNF-a gene has been associated with elevated cytokine production in both psoriatic lesional skin (109,110) and the serum of patients with PS (111). Notably, the GA genotype has been associated with a later age of onset for PsA (101), while an increased frequency of the ‘A’ allele has been observed in female patients with PS (100).

The FOXP3 gene is positioned at the Xp11.23 locus of chromosome X (112). It encodes the FOXP3 transcription factor, a major regulatory factor involved in the T-cell activation and the development and function of Treg cells in the peripheral blood, which are considered to be inhibitors of autoimmune responses (113,114). The FOXP3 rs3761549 (-2383 C/T) SNP, located in an intronic region, has been reported to be associated with endometriosis and PS (76,77). Particularly, it was found that increased risk of PS was associated with the CC genotype and the ‘C’ allele of this SNP (77). This SNP was found to alter the FOXP3 expression pathway and affect Treg regulation in PS (115,116). Notably, an association of the rs3761549 SNP with an increased risk for endometriosis, but not with the severity or stage of the disease has been previously reported (76).

HLA molecules, which play an important role by restricting the antigenic peptides by T cells, are encoded by genes of the MHCs on chromosome 6 and help the body to discriminate between self and non-self proteins (117). These antigens exhibit the highest level of variability within the human genome and function by presenting identifying proteins to cytotoxic T cells, thereby triggering subsequent immune responses (118). The HLA-DRB1 gene belongs to the group of MHC class II genes, which are expressed on the surface of antigen-presenting cells and display peptides to Th CD4⁺ cells, thus inducing their activation (119). The HLA-DRB1 (A/G) rs660895 SNP, has been associated with an increased risk for moderate to severe endometriosis (78). Τhe functional significance of this SNP in endometriosis remains unclear. Patients with endometriosis exhibit reduced levels of circulating HLA class II molecules, while the proportion of HLA-DR-positive stromal and glandular epithelial cells is higher in ectopic endometrium compared with eutopic tiisue (120). Therefore, it has been hypothesized that the HLA-DR antigen may play an important role in the immunological dysregulation observed in endometriosis (78). Notably, the rs660895 SNP of the HLA-DRB1 gene has been shown to be associated with increased suseptibility to PS, particularly in early-onset cases (64,121,122). Additionally, hypomethylation of HLA-DRB1 was also found to play a role in PS pathogenesis by affecting the mRNA expression of HLA-DRB1 (123).

The IL-10 gene is located on 1q31-32, spaning a genomic region of 4.7 kb and containing 5 exons (124). It encodes IL-10, a potent anti-inflammatory cytokine that is mainly produced by T-cell subsets, including Th2 and Treg cells, but it is also produced by macrophages, monocytes, natural killer (NK) cells and B cells (125,126). The IL-10 protein modulates Th2-mediated inflammatory processes and functions as an inhibitor of the synthesis of pro-inflammatory mediators, including, cytokines and chemokines as well as adhesion and accessory molecules, thus suppressing T-cell stimulation (127,128). IL-10 rs1800871 (-819C/T), a functional promoter SNP located in a regulatory region of the promoter, has been reported to be associated with an increased risk for endometriosis and PS (79,80). It has been reported that women with the TT genotype or carrying the ‘T’ allele of rs1800871 appear to have a 2-fold increased risk for endometriosis compared with the carriers of the ‘C’ allele (79,129). Notably, the ‘C’ allele has been associated with higher levels of both IL-10 mRNA and protein compared with the ‘T’ allele in patients (130,131), suggesting that this allele may contribute to the downregulation of cell inflammation in the peritoneal cavity (79). Considering that the aforementioned SNP is located in the promoter area of IL-10 gene, it has been suggested that rs1800871 may change the binding site of a transcription factor, thus decreasing the production of IL-10, altering the immunological homeostasis and resulting in development of PS (80).

The human MTHFR gene is located on chromosome 1p36.3, a region with numerous identifiable SNPs (132) and encodes the MTHFR protein. MTHFR represents a key regulatory enzyme in the folate metabolic pathway, with its main function being the conversion of 5,10-methylenetetrahydrofolate into 5-methyltetrahydrofolate (133). The MTHFR rs1801133 (Ala222Val) SNP, located in exon 4 and substituting nucleotide C with T, has been found to be significantly associated with endometriosis (81,83) and PS (134). This SNP results in the reduced activity of the enzyme in TT homozygotes, thus resulting in a decreased production of the antioxidant glutathione, due to alterations of both gene expression levels and protein function, and increased homocysteine levels (135-138). Notably, the 677T variant, which is suggested to be unstable, may influence the thermostability and activity of the MHTFR protein (139). In endometriosis, individuals homozygous for the TT genotype have been associated with a higher susceptibility to endometriosis (81). Furthermore, the TT genotype and ‘T’ allele were found to increase the risk of PS compared with controls (82,134,140,141). Of note, it was found that rs1801133 also affects the severity of PS according to a cross-sectional study conducted by Karabacak et al (142) and a meta-analysis performed by Wu et al (143).

The STAT4 gene, which is located at 2q32.2-q32.3 and consists of 24 exons (144), codes for the STAT4 protein that is a key transcriptional factor expressed in various immune-related cells, such as macrophages, peripheral blood monocytes and dendritic cells (145). This protein, which is a member of the JAK/STAT pathway, is involved in transmission of signals from IL-12, IL-23 and type 1 IFN (146) and is necessary for the induction of IFN-γ production (147). As a consequence, STAT4 has a crucial role in the establishment of chronic inflammation (148). The intronic STAT4 rs7574865 G/T SNP has been associated with an increased risk of both endometriosis and PS (84,85). Thus, the TT genotype rs7574865 SNP was shown to be associated primarily to cases of endometriosis at minimal or mild stages of endometriosis compared with controls (84). However, the functional role of this SNP in the development of endometriosis remains unclear. A previous bioinformatics analysis was unsuccesful in identifying any activator or transcription factor whose binding sites are disrupted by the presence of the risk allele ‘T’ (149). Thus, considering the role of the STAT4 protein in IFN signaling, it can be inferred that rs7574865 SNP contributes to disease development, especially given the strong association between the Th1 response pattern and deep infiltrating endometriosis (150). Furthermore, the ‘T’ allele was previously found to be associated with the development of PS (85). Notably, STAT4 affects the optimal differentiation of Th17 cells, which are involved in the pathogenesis of both PS and RA (151,152).

The VEGF gene is located on chromosome 6p21.3, consists of 8 exons and gives rise to a family of proteins through alternative splicing events (153). It encodes the VEGF protein, which plays an important role in the regulation of angiogenesis (154) as well as endothelial cell dysfunction (155). Several transcription-factor binding sites have been detected in the 5'-UTR of this gene. Moreover, polymorphisms within this region result in an increase of the transcriptional activity of the gene (156) and may be significant in the etiology of the associated pathological conditions (157). The rs1570360 (-1154G/A) SNP, located within a transcription factor binding site in the VEGF promoter, has been associated with both endometriosis (90,158) and PS (88,89,91). Particularly, the ‘G’ allele and GG genotype have been associated with an increased risk of developing endometriosis in comparison with controls, while it has also been associated with deep infiltrating endometriosis as well as moderate to severe forms of the disease (stages III and IV) (63,90). Notably, four potential transcriptional factor binding sites were found to be created by the ‘G’ allele for EGR1, KLF4, MZF1 and SP2 factors (159). In previous studies, elevated levels of VEGF mRNA and protein have been observed in serum and peritoneal fluid of women with endometriosis (90,160,161). Angiogenic activity has been found to be dysregulated in the eutopic endometrium of patients with endometriosis (162,163). Considering that VEGF is a regulator of various processes contributing to the ectopic implantation and growth of the endometrial tissue (164), including neoangiogenesis and tissue remodeling (153), it has been suggested that any alteration in the levels of VEGF may promote the initiation of endometriosis (63,90). Of note, rs1570360 has been associated with the development of PS and the ‘G’ allele was observed significantly more frequently among patients with PS (88). Furthermore, significantly increased VEGF serum levels were found in patients with PS compared with healthy controls, while a positive correlation between the serum levels of VEGF and the severity of the disease was also detected (88). Collectively, this SNP may alter the function of the immune system and increase the susceptibility to PS.

The IL-1 gene cluster, located on the chromosomal 2q12-13 locus (165), consists of nine inflammation-related genes. The encoded protein, IL-1A, is a pro-inflammatory cytokine that is primarily produced by activated macrophages, and it promotes the activation of macrophages as well as T- and B-lymphocytes (166). Both IL-1A rs17561 (p.A114S) and rs3783553 SNPs, located in exon 5 and the 3'-UTR of the gene, respectively, have been shown to be associated with susceptibility to endometriosis and PsA (40,92,93,167). With respect to endometriosis, it has been suggested that the impairment of the IL-1 family network may result in a disrupted activation of the immune system in the peritoneal cavity of patients with endometriosis compared with the serum of affected women, thus leading to the establishment of chronic inflammation (168). Moreover, the increased expression levels of IL-1A in the peritoneal fluid and serum of women with endometriosis (169,170) may be involved in the implantation of menstrual endometrium on peritoneal surfaces (9). Thus, functional alterations of IL-1A probably play a role in the pathogenesis of endometriosis. In this context, it has been suggested that rs3783553, located in the 3'-UTR, may impair mRNA stability and lead to an alteration of the expression levels of IL-1A (171). A strong association of rs17561, which results in an A114S substitution, and endometriosis has been reported, especially among women with moderate to severe forms of the disease (92). Although, a three-dimensional model of the mature IL-1A protein has been constructed (172), the biological significance of the p.114S variant remains unclear given that no functional analyses have been performed in endometriotic cells thus far. However, by using the bioinformatics tool, PolyPhen-2 (http://genetics.bwh.harvard.edu/pph2/) (173), it was suggested that the risk variant might affect processing efficiency at the protein level (92). An increased expression of IL-1A has been detected in the serum synovial fluid and skin of patients with PsA, but no extensive studies have been performed to date, clarifying the protein function in patients with PsA (174,175). Genetic variants of IL-1A may induce inflammation in PsA through intracellular effects, thus contributing to the development of the disease.

The MIF gene, located on chromosome 22q11.2 region and consisting of three exons (176), encodes the MIF protein, a key pro-inflammatory cytokine produced by T lymphocytes and activated macrophages (177). Furthermore, MIF is involved in the upregulation of IL-6, IFN-γ and TNF-α genes (178). Notably, MIF has been implicated in the pathogenesis of various autoimmune diseases, such as RA, multiple sclerosis, inflammatory bowel disease, PsA and juvenile rheumatoid arthritis (94,179). This protein has also been found to activate several molecular pathways in ectopic endometrial tissue, including immune-inflammatory processes, angiogenesis, and the growth, development and implantation of endometriotic tissue (180). It has been reported that the rs755622-173 G>C SNP, located in the promoter region of the MIF gene, has been associated with an increased risk of endometriosis and PsA (72,94). The increased transcriptional activity of the MIF gene in women with endometriosis, due to the presence of ‘C’ allele, leads to an increased synthesis of local estrogen in ectopic tissues, which has been suggested to be involved in both maintenance and progression of endometriosis (181). A significant increase of MIF levels was found in patients with PsA, while the-173C allele and the CG genotype were revealed to be associated with both an increased susceptibility to PsA and elevated levels of MIF in circulation, in several populations (72). It is worth noting that the increase in serum levels of MIF was observed in early stages of PsA, thus suggesting a crucial role of MIF in PsA onset by elevating its levels at the beginning of the inflammatory process (72). Moreover, a previous study by Donn et al (176) demonstrated that the presence of ‘C’ allele of rs755622 SNP creates a binding site for the activating enhancer binding protein 4 (AP-4) transcription factor, and as a consequence, this allele significantly increases the affinity of the MIF promoter for AP-4 binding.

The protein tyrosine phosphatase non-receptor type 22 (PTPN22) gene, located on the 1p13.3-13.1 region, encodes the lymphoid-specific phosphatase Lyp that prevents spontaneous T-cell activation (182). The Lyp protein physically binds to the Csk protein, thus functioning as a downregulator of T-cell activation (183). The PTPN22 functional polymorphism rs2476601 (R620W, C1885T), leads to the production of a gain-of-function Lyp protein that is unable to bind to the Csk efficiently and, therefore, T-cell activation cannot be suppressed (182). This SNP has been reported to be associated with an increased susceptibility to both endometriosis and PsA (68,95,96). Regarding endometriosis, the minor ‘T’ allele has been associated with an increased risk of the disease. Moreover, this allele appears to be associated with moderate/severe (III/IV) stages of endometriosis, rather than with minimal/mild (I/II) forms (95). In the same context, the minor ‘T’ allele has been found at an increased frequency in patients with PsA compared with the control individuals (68,96). The finding that wild-type Lyp is among the most potent inhibitors of T-cell activation provides a potential explanation for the association between the presence of ‘T’ allele and increased risk of PsA (96). Furthermore, patients with PsA carrying the ‘T’ allele appear to have a significantly higher number of joints with deformities, suggesting that this variant may be associated with a more aggressive disease phenotype (184).