Porphyromonas gingivalis LPS regulates MnSOD in HGFs through TLR4-NF-κB pathway

Abstract

Objectives: Porphyromonas gingivalis is a keystone periodontal pathogen and its lipopolysacharide (LPS) is strongly associated with periodontal disease. P. gingivalis LPS (PgLPS) displays remarkable heterogeneity containing both tetra- (LPS1435/1449) and penta-acylated (LPS1690) lipid A structures. We recently demonstrated that PgLPS differentially modulates the immuno-inflammatory response in human gingival fibroblasts (HGFs). The present study further investigated the expression and regulation of antioxidants in HGFs in response to heterogeneous PgLPS. Methods: Primary HGFs were treated with PgLPS1435/1449 and PgLPS1690 as well as E. coli LPS in appropriate dose- and time-dependent assays. Modulation of oxidative stress and anti-oxidant defense pathways were examined using gene-expression arrays. The identified biomarkers were further validated by real-time qPCR and Western blot. Involvement of TLRs and signaling pathways (MAPK/NF-κB) for PgLPS-induced MnSOD expression was elucidated by Western blot and antibody-mediated blocking assays.

Results: Gene-array results showed that PgLPS1690 upregulated significantly the antioxidants MnSOD (SOD2), SOD3 and PXDNL compared to the PgLPS1435/1449. PgLPS1690 and E. coli LPS significantly upregulates MnSOD, PRDX1 and TXN1 mRNAs in dose- and time-dependent manners. MnSOD mRNA expression was significantly inhibited in PgLPS1690-treated cells through NF-κB and SAPK/JNK pathways. In contrast, blockage of NF-κB and P38 MAPK significantly inhibited MnSOD expression in E. coli LPS-treated cells. Both TLR4 and TLR2 were involved in the regulation of MnSOD expression in PgLPS1690-treated HGFs, whereas only TLR4 was involved in E. coli LPS-treated HGFs.

Conclusions: This study unravels for the first time the oxidative stress and antioxidant expression of HGFs in response to heterogeneous PgLPS. MnSOD could play a key role in antioxidant defense of gingival tissues through TLRs and NF-κB signaling pathway by targeting P. gingivalis to shift its LPS structure, which could be a strategy to restore the antioxidant status of gingival tissues and thereby preventing periodontal diseases. (Supported by the Hong Kong RGC, No. HKU766909M).