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. 2020 May 29;15(5):e0233737.
doi: 10.1371/journal.pone.0233737. eCollection 2020.

Hypomethylation of IL1RN and NFKB1 genes is linked to the dysbalance in IL1β/IL-1Ra axis in female patients with type 2 diabetes mellitus

Sona Margaryan  1   2   3 , Eva Kriegova  1 , Regina Fillerova  1 , Veronika Smotkova Kraiczova  1 , Gayane Manukyan  1   2   3
Affiliations

Hypomethylation of IL1RN and NFKB1 genes is linked to the dysbalance in IL1β/IL-1Ra axis in female patients with type 2 diabetes mellitus

Sona Margaryan et al. PLoS One. .

Abstract

Inflammation has received considerable attention in the pathogenesis of type 2 diabetes mellitus (T2DM). Supporting this concept, enhanced expression of interleukin (IL)-1β and increased infiltration of macrophages are observed in pancreatic islets of patients with T2DM. Although IL-1 receptor antagonist (IL-1Ra) plays a major role in controlling of IL-1β-mediated inflammation, its counteraction effects and epigenetic alterations in T2DM are less studied. Thus, we aimed to analyze the DNA methylation status in IL1RN, RELA (p65) and NFKB1 (p50) genes in peripheral blood mononuclear cells (PBMCs) from treated T2DM patients (n = 35) and age-/sex- matched healthy controls (n = 31). Production of IL-1β and IL-1Ra was analyzed in plasma and supernatants from LPS-induced PBMCs. Immunomodulatory effects of IL-1β and IL-1Ra were studied on THP-1 cells. Average DNA methylation level of IL1RN and NFKB1 gene promoters was significantly decreased in T2DM patients in comparison with healthy controls (P< 0.05), which was associated with the increased IL-1Ra (P< 0.001) and IL-1β (P = 0.039) plasma levels in T2DM patients. Negative association between average methylation of IL1RN gene and IL-1Ra plasma levels were observed in female T2DM patients. Methylation of NFKB1 gene was negatively correlated with IL-1Ra levels in the patients and positively with IL-1β levels in female patients. LPS-stimulated PBMCs from female patients failed to raise IL-1β production, while the cells from healthy females increased IL-1β production in comparison with unstimulated cells (P< 0.001). Taken together, the findings suggest that hypomethylation of IL1RN and NFKB1 gene promoters may promote the increased IL-1β/IL-1Ra production and regulate chronic inflammation in T2DM. Further studies are necessary to elucidate the causal direction of these associations and potential role of IL-1Ra in anti-inflammatory processes in treated patients with T2DM.

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Conflict of interest statement

The authors have declared that no competing interests exist.

Figures

Fig 1
Fig 1. Average DNA methylation levels of target genes in PBMCs from type 2 diabetes mellitus (T2DM, n = 35) patients and healthy controls (Healthy, n = 31).
(A) Mean methylation levels in IL1RN gene promoter (across 16 CpG loci) in all subjects and males/females; (B) Mean methylation levels in NFKB1 gene promoter (across 65 CpG loci) in all subjects and males/females. Results are expressed as mean ± SEM. *P< 0.05.
Fig 2
Fig 2. Plasma levels of cytokines in type 2 diabetes mellitus (T2DM, n = 35) patients and healthy controls (Healthy, n = 30).
(A) Levels of IL-1Ra in all subjects and males/females; (B) Levels of IL-1β in all subjects and males/females. Results are expressed as mean ± SEM. *P< 0.05, ***P< 0.001.
Fig 3
Fig 3. An association between mean DNA methylation and cytokine plasma levels in type 2 diabetes mellitus (T2DM) patients and healthy controls (Healthy).
(A) Correlation between IL1RN promoter mean methylation level and IL-1Ra plasma levels in males (green) and females (violet); (B) Correlation between NFKB1 promoter mean methylation level and IL-1Ra plasma levels in all studied subjects (black); (C) Correlation between NFKB1 promoter mean methylation level and IL-1β plasma levels in males/females.
Fig 4
Fig 4. LPS-stimulated production of IL-1Ra and IL-1β by PBMCs from type 2 diabetes mellitus patients (T2DM, n = 32) and healthy controls (Healthy, n = 27).
PBMCs isolated from males (green) and females (violet) were treated with LPS (100ng/ml) and left unstimulated as a control (Ctl) for 24h. Data are presented as mean ± SEM. *P< 0.05, ***P< 0.001.
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Internal grant agency of Palacky University (IGA UP_2020_016), Ministry of Health Czech Republic MH CZ-DRO (FNOL, 00098892) as well as Russian-Armenian University within the framework of the grant of the Ministry of Education and Science of the Russian Federation.