PMC

Effects of Wnt5a on the Wnt/Ror2 signalling pathway in melan-a cells. (A) Melan-a cells were infected with AdWnt5a. After 0h, 16h, 24h and 48h, RT-PCR analyses were performed with primers specific for Ror2, JUN, JNK1, JNK2 and GADPH. (B) The relative mRNA expression levels. These data were representative results of three independent experiments. *p< 0.05.

Effects of Wnt5a on the proliferation of melan-a cells. (A) Melan-a cells infected by Ad-Wnt5a. The infection efficiency was observed by green fluorescence. (Left, phase contrast images; Right, fluorescent images) Scale bars represent 100 µm. (B) RT-PCR and Western-blot analysis of Wnt5a mRNA and protein after infection with AdWnt5a or AdGFP. (C) Proliferation of melan-a cells infected with different doses of AdWnt5a or AdGFP as measured by the MTT assay. (D) Proliferation of melan-a cells infected with AdGFP or AdWnt5a as measured by BrdU incorporation. BrdU incorporation was detected by immunofluorescent staining (Left), and the percentage of BrdU positive cells were quantitatively measured (Right). Red, BrdU; Blue, DAPI. Scale bars represent 50 µm. *p< 0.05, **p< 0.01. n =3.

Effects of Wnt5a on the melanogenesis of melan-a cells. (A) Tyrosinase activity of melan-a cells infected with different doses of AdWnt5a or AdGFP analyzed by tyrosinase activity assay. (B) Melanin synthesis of melan-a cells infected with AdWnt5a or AdGFP. (C-D) Effect of Wnt5a on the expression of TRP1 and tyrosinase in melan-a cells. (C) Melan-a cells were infected with AdWnt5a or AdGFP. RT-PCR analyses were performed with primers specific for TRP1, tyrosinase and GADPH after 0h, 16h, 24h and 48h. (D) Melan-a cells were infected with AdWnt5a or AdGFP. After 48 h, Western blot analyses were performed with antibodies specific for TRP1, tyrosinase and GADPH. (E) Melan-a cells infected with AdWnt5a or AdGFP were implanted subcutaneously into nude mice. Visual pigmentation were exhibited in Day 1 and 3. Implanted sites were retrieved and subjected to Fontana-Masson staining. Melanin-positive cells were indicated with arrows. Bar=50 μm. These data were representative results of three independent experiments. *p< 0.05, **p< 0.01.

Effects of Wnt5a on the canonical Wnt signalling pathway in melan-a cells. (A) Effect of Wnt5a on the expression of β-catenin mRNA and protein as measured by RT-PCR and Western blot assays. (B) Effect of Wnt5a on the tyrosinase activity in AdWnt3a-infected melan-a cells. The cells were infected with AdGFP, AdWnt3a, or co-infected with AdWnt3a and AdWnt5a. After 48 h, tyrosinase activity was analyzed by tyrosinase activity assay. (C-D) Effect of Wnt5a on the expression of β-catenin, TRP1 and tyrosinase in AdWnt3a-infected melan-a cells. The cells were infected with AdGFP, AdWnt3a, or co-infected with AdWnt3a and AdWnt5a for 48 h. (C) RT-PCR analyses were performed with primers specific for β-catenin, TRP1, tyrosinase and GADPH (Left) and the relative mRNA expression levels were quantitatively measured (Right). (D) Western blot analyses were performed with antibodies specific for β-catenin, TRP1, tyrosinase and GADPH (Left) and the relative protein expression levels were quantitatively measured (Right). These data were representative results of three independent experiments. *p< 0.05.