Differential mRNA and tissue expression of lymphangiogenic growth factors (VEGF-C and -D) and their receptor (VEGFR-3) during tail regeneration in a gecko

Lymphangiogenesis, the growth of new lymph vessels, has important roles in both normal and pathological lymphatic function. Despite recent advances, the precise molecular mechanisms behind the lymphangiogenic process remain unclear. The Australian marbled gecko, Christinus marmoratus, voluntarily drops its tail (autotomy) as a predator avoidance strategy. Following autotomy a new tail is regenerated including lymphatic drainage pathways. We examined the molecular control of lymphangiogenesis within the unique model of the regenerating gecko tail. Partial sequences were obtained of the gecko lymphangiogenic growth factors, vascular endothelial growth factor C (VEGF-C) and VEGF-D along with their receptor VEGFR-3. These were highly homologous to other vertebrates. Quantitative real-time polymerase chain reaction (PCR) demonstrated up-regulation of VEGF-C, VEGF-D and VEGFR-3 mRNA expression during the early and middle stages of tail regeneration (between 4 and 9 weeks following autotomy), in late regeneration (12 weeks) and during mid-regeneration (7 and 9 weeks), respectively. VEGF-C and VEGF-D immunostaining was observed lining some lymphatic-like and blood vessels in early-mid tail regeneration, indicating possible associations of the proteins with VEGFRs on endothelia. Keratinocytes and fibroblasts also showed positive staining of VEGF-C and VEGF-D in early-mid tail regeneration. Additionally, VEGF-C was localised in adipose tissue in all tail states examined. This work suggests that specific timings exist for the expression of the lymphangiogenic growth factors, VEGF-C and VEGF-D, and their receptor, VEGF-R3, throughout the regeneration of a functional lymphatic network. Along with localisation data, this suggests potential functions for the growth factors in lymphangiogenesis and angiogenesis throughout tail regeneration.