Journal: Cell Proliferation

Article Title: Microphthalmia‐associated transcription factor/T‐box factor‐2 axis acts through Cyclin D1 to regulate melanocyte proliferation

doi: 10.1111/cpr.12227

Figure Lengend Snippet: Knockdown of Tbx2 inhibited the proliferation of melanoblasts in primary neural crest cell cultures. Neural tube explants were isolated from E9.5 C57BL/6J embryos and cultured in melanocyte induction medium. MITF staining labels melanoblasts. (a) Double immunolabelling for MITF (green nuclear staining) and TBX2 (red nuclear staining) at different days culture. MITF expression began at day 2 while TBX2 and MITF were co‐expressed only at day 4. White arrows indicate positive cells. Bar = 20 μm. (b, c) siRNA‐mediated Tbx2 down‐regulation reduced percentage of MITF and Ki67 double‐positive cells. Primary neural crest cells were transfected with a negative control (si‐C) or Tbx2‐specific siRNAs (si‐Tbx2‐1 and si‐Tbx2‐2). MITF and Ki67 double‐positive cells were recorded and counted 4 days after transfection. Bar = 50 μm (b). Note that number of Ki67 and MITF double‐positive cells was dramatically reduced after knockdown of Tbx2 (c). Experiments were performed in triplicate and are represented as mean ± SD. Significance was determined by Student's t‐test whereby **P < 0.01.

Article Snippet: Melan‐a cells were plated in 96‐well plates, 2000 cells per well, and transfected using LipoJet TM in vitro transfection kit (SignaGen Laboratories) with 40 n m siRNA, in each well.

Techniques: Knockdown, Isolation, Cell Culture, Staining, Expressing, Transfection, Negative Control

Journal: Cell Proliferation

Article Title: Microphthalmia‐associated transcription factor/T‐box factor‐2 axis acts through Cyclin D1 to regulate melanocyte proliferation

doi: 10.1111/cpr.12227

Figure Lengend Snippet: Knockdown of Tbx2 inhibited melan‐a cell proliferation. (a) Western blot analysis of TBX2 expression using anti‐TBX2 antibody. Note that expression of TBX2 was significantly reduced in melan‐a cells transfected with Tbx2‐specific siRNAs (si‐Tbx2‐1 and si‐Tbx2‐2). Intensity of bands was measured using ImageJ software and fold‐change was normalized to that of mock cells. (b) Proliferation analysis for melan‐a cells transfected with si‐C, si‐Tbx2‐1 or si‐Tbx2‐2. Cell numbers were recorded from day 1 to day 4 after transfection. Note that knockdown of Tbx2 led to reduced cell growth compared to mock cells. (c) Immunolabelling for Ki67 (red nuclear staining), a marker for proliferating cells, in melan‐a cells transfected with si‐C, si‐Tbx2‐1 or si‐Tbx2‐2. Number of Ki67‐positive cells was reduced in Tbx2 knockdown cells after 3 days transfection by si‐Tbx2. Bar = 50 μm. (d) Percentage of Ki67‐positive cells was determined based on data similar to those shown in Fig. ​Fig.2c.2c. Data are from triplicate experiments and are represented as mean ± SD. **P < 0.01.

Article Snippet: Melan‐a cells were plated in 96‐well plates, 2000 cells per well, and transfected using LipoJet TM in vitro transfection kit (SignaGen Laboratories) with 40 n m siRNA, in each well.

Techniques: Knockdown, Western Blot, Expressing, Transfection, Software, Staining, Marker