Lysophosphatidic acid regulates SPHK1 and GPR68/OGR1 in human gingival fibroblasts

Abstract

Our laboratory has established that lysophosphatidic acid (LPA) regulates the biology of human oral fibroblasts, and that gingival fibroblasts (GF) express the LPA1–5 receptor subtypes. We have also shown that LPA controls immediate‐early gene transcription of inflammatory cytokines, and their receptors and regulators in GF; therefore, we hypothesized that LPA would modulate transcription of the isozymes SPHK1 and/or SPHK2, which make sphingosine‐1‐phosphate (S1P). S1P is under much investigation as a major regulator in immunity and inflammation. Critically, S1P is a primary controller of human neutrophil responses (such as chemotaxis, degranulation, and oxidative burst), making it highly relevant to the inflammation seen in periodontal disease. After GF treatment with 1 × 10−5 M 18:1 LPA for 2h or 8h, RNA was extracted. Agilent Whole Human Genome Oligonucleotide Microarray analysis revealed that LPA consistently and significantly upregulated SPHK1 versus control (3.9 ± 0.5 and 4.7± 0.7 fold at 2h and 8h, respectively). Of great interest, the proton‐sensing receptor GPR68/OGR1 was highly induced: 7.3 ± 4.0 fold (2h) and 18.8 ± 12.8 fold (8h). LPA1–3 can all dimerize with GPR68 /OGR1, which is linked to COX‐2 and inflammatory cytokine induction in other systems. QRT‐PCR was performed to validate the microarray results, using 3 biologic replicates for four periodontal disease‐relevant up‐regulated genes at the 8h time point. These results (level of significance, p < 0.01) provide preliminary evidence that LPA likely exerts complex actions during periodontal inflammation by inducing S1P production and through LPA1–3 hetero‐dimerizing with GPR68. Support: Health Future Foundation (D.R.C).