Lymphatic growth factors are expressed in human gingiva and upregulated in gingival fibroblasts after stimulation

BACKGROUND: The lymphatic growth factors vascular endothelial growth factor (VEGF)-C and -D are important for maintenance and growth of lymphatic vessels (lymphangiogenesis), but their localization in human gingiva is unknown. This study investigated the expression of VEGF-C and -D in human gingiva and isolated human gingival fibroblasts (HGFs). In addition, the localization of their main receptor VEGFR-3 was explored. METHODS: Non-inflamed gingiva from six donors was used for immunohistochemistry or isolation of HGFs. HGFs were stimulated with either E.coli lipopolysaccharide (LPS) or IL-6/soluble IL-6 receptor (sIL-6R) complex for 1, 6, and 24 hours. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to quantify the relative changes in gene expression of VEGF-A, -C, and -D and enzyme-linked immunosorbent assay (ELISA) for quantification of protein levels. RESULTS: VEGF-C, -D and VEGFR-3 were seen in keratinocytes, blood vessels and in scattered single cells in gingiva. VEGFR-3 was also found in lymphatic vessels and VEGF-C in cells with fibroblastic appearance. Gene analysis showed no expression of VEGF-D in the HGFs, but showed constitutive expression of VEGF-C and -A. Stimulation of HGFs with LPS or IL-6/sIL-6R complex was followed by gene upregulation of VEGF-C and -A and increased protein levels in cell culture supernatant (P ≤0.05). CONCLUSIONS: The localization of VEGF-C, -D, and VEGFR-3 expression imply that signaling via VEGFR-3 is linked to vascular homeostasis and keratinocyte function under normal conditions in gingiva. Inflammatory stimulation of HGFs upregulates VEGF-C and -A expression and may contribute to angiogenesis and lymphangiogenesis.