Sprouty2 Regulates Cell Proliferation And Migration In Periodontal Ligament Cells

Objectives: The periodontal ligament (PDL) plays an important role as a reservoir of mesenchymal stem cell. The cells in PDL tissue are critical participants during periodontal tissue regeneration. Sprouty (Spry) is an intracellular protein includes four mammalian homologs, which are expressed in most tissues. Mammalian Spry proteins were originally identified as an inhibitor of fibroblast growth factor (FGF) receptor, and it functions as a negative regulator of receptor tyrosine kinases (RTKs) signaling. However, Spry2 does not affect epidermal growth factor (EGF)-induced ERK activation. Since both FGF and EGF have been suggested as possible growth factor for periodontal cytokine therapy, we aimed to investigate whether Spry2 could be new therapeutic targets for enhancing periodontal tissue regeneration.
Methods: We used multipotent clonal human PDL cell line 1-17. The cells were transfected with small interfering RNA (siRNA) specific for Spry2. Using WST-8 assay and Ki-67 staining, growth factor-induced cell proliferation was investigated. In order to evaluate osteoblastic differentiation, ALP staining and Real-time PCR detecting genes associated with calcification were performed. Cell migration was assessed by a scratch wound healing assay and boyden chamber assay, and lamellipodia formation was confirmed by fluorescence microscope.
Results: Transfection of Spry2 siRNA enhanced bFGF+EGF-induced ERK activation in 1-17 cells. Spry2 siRNA 1-17 cells stimulated by bFGF+EGF proliferated faster than control cells. Migration activity and lamellipodia formation of bFGF+EGF-stimulated Spry2 siRNA 1-17 cells were promoted compared with control cells accompanied by increase in PI3K, Akt and Rac1 activity. On the other hand, osteoblastic differentiation of Spry2 siRNA 1-17 cells was inhibited.
Conclusions: bFGF+EGF induced proliferation and migration was up-regulated in Spry2 siRNA 1-17 cells, whereas osteogenic differentiation was inhibited. Since selective proliferation and migration of PDL cells are important for periodontal tissue regeneration, Spry2 could be new therapeutic targets for periodontal regeneration.