Nicotinamide N-Methyltransferase in the Inflammatory Pathogenesis of Graves' Orbitopathy

Abstract

Purpose: Nicotinamide N-methyltransferase (NNMT) has been implicated in inflammatory autoimmune disease pathogenesis, although its pro-inflammatory role in Graves' orbitopathy (GO) is unclear. Therefore, we investigated the influence and mechanisms of NNMT in GO inflammation.

Methods: We evaluated NNMT mRNA expression in GO and non-GO orbital tissues via reverse transcription-quantitative PCR analysis. A pro-inflammatory process was induced in primary cultured orbital fibroblasts via interleukin (IL)-1β treatment, and NNMT expression was assessed by Western blotting. To further investigate the role of NNMT in GO inflammation, we inhibited NNMT expression and activity using small interfering RNA (siRNA) and pharmacologic antagonists, respectively. The production of inflammatory cytokines and intracellular signaling molecules were analyzed via Western blotting and enzyme-linked immunosorbent assay analysis.

Results: NNMT mRNA expression levels were higher in GO orbital tissues than in healthy orbital tissues. Tissues from patients with type Ⅱ GO showed higher NNMT expression than those with type Ⅰ GO. Pro-inflammatory stimulation induced NNMT expression in dose- and time-dependent manners. NNMT siRNA and antagonists attenuated the expression of pro-inflammatory cytokines (IL-6, IL-8, and monocyte chemotactic protein-1), cyclooxygenase-2, and prostaglandin E2 in orbital fibroblasts. NNMT silencing downregulated the active forms of intracellular signaling molecules (extracellular signal-regulated kinase, c-Jun-terminal kinase, and p38).

Conclusions: Our results demonstrate that NNMT was associated with the inflammatory mechanisms of GO. Inhibiting NNMT, either through mRNA silencing or pharmacologic antagonism, markedly reduced pro-inflammatory reactions. These findings suggest that targeting NNMT is a promising therapeutic strategy for managing inflammation in GO.