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Hypermethylation of NLRP3 Promoter Region Could be Responsible for Decreased Gene Expression, Inflammasome Malfunction and Gut Dysbiosis in Juvenile Spondyloarthritis Patients

Background/Purpose: Juvenile spondyloarthritis (jSpA) is a complex disease with both genetic and environmental factors contributing to the etiology. Recently obtained gene signatures in jSpA patients revealed TLR4 and CXCR4 gene had increased, while NLRP3 and PTPN12 had decreased expression, though the mechanism(s) responsible for those alterations remained unknown. To elucidate the possible role of epigenetic modifications in the regulation of those genes, DNA methylation analysis was performed. Methods: DNA was isolated from PBMCs of 19 patients diagnosed with jSpA according to ILAR classification criteria for enthesitis related arthritis (ErA) and seven matched healthy children. None of the jSpA participant had symptoms for more than six months and all were untreated. Methylated DNA Immunoprecipitation (MeDIP) was performed in promotor region of differentially expressed genes (TLR4, CXCR4, NLRP3, PTPN12) using the Magmedip kit. Enrichment in MeDIP fraction was determined by quantitative RT-PCR using the AriaMx. MeDIP results were expressed as fold enrichment of immunoprecipitated DNA for each site. Results: Statistical analysis revealed significant hypermethylation of promoter sites in NLRP3 gene (p=0, 0220). No significant alterations in methylation status were observed in promotor regions of other genes (Table 1). Conclusion: Our study indicates the hypermethylation of NLRP3 gene promotor is probably responsible for expression alterations in jSpA patients in the initial phase of the disease. The NLRP3 gene has a crucial role in assembly of NLRP3 inflammasome, innate immune sensor that regulates “danger” response upon various signals. While increased expression of this gene has been found in many autoinflammatory diseases, decreased expression has been associated with IBD in which the microbiota is believed to contribute to the intestinal inflammation. Therefore, it is not entirely surprising decreased expression has also been observed in jSpA, the disease that has clinical and genetic similarities with IBD and is often characterized by subclinical gut inflammation. The growing number of evidence shows any modification of gut microbiota can lead to dysbiosis with long- term consequences for the whole organism. Specifically, in jSpA this could result in increased influx of TLR4 ligands and increased expression of TLR4 gene, as well as in reduction of commensal bacteria with anti-inflammatory properties, namely bacteria Faecalibacterium prausnitzii that inhibits NF-kB signaling, leading to TNF-a abundancy characteristic for jSpA. Since inflammasomes have been shown to shape the microbiota, it is reasonable to assume these processes can at least partially be explained by reduced NLRP3 expression due to hypermethylation. Therefore, our findings could have important implications in understanding of the disease mechanisms and possible therapeutic options.

juvenile spondyloarthritis ; epigenetics ; jSpA ; microRNAs ; methylation ; pondyloarthritis ; epigenetics ; jSpA ; microRNAs ; methylation ;

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