Journal: Advanced Functional Materials
Article Title: A Bio‐Functional Wnt3a Gradient Microarray for Cell Niche Studies
doi: 10.1002/adfm.202301941
Figure Lengend Snippet: Figure 8. Maintenance of human hair dermal papilla cells (HHDPCs) on the Wnt3a gradient. A) Study design and workflow of characterizing the fibronectin meshwork deposited by the HHDPCs cultured on Wnt3a gradient. B) Representative immunofluorescence images of fibronectin deposited by the HHDPCs cultured on the NA gradient bound with bWnt3a (LOL = 12, 100 ng of input) i) and on the NA gradient only in the culture medium ii), for 24 h. Scale bar: 100 µm. Linear regression analysis of the relationship between the area of fibronectin matrix iii), the total length of fibronectin matrix iv), the percentage (%) of the high-density fibronectin matrix v) and the lacunarity of the fibronectin matrix vi) on the Wnt3a or NA gradient. Evaluation of β-catenin nuclear accumulation in HHDPCs cultured on the Wnt3a gradient by counting the number of cells positive in the nuclear β-catenin vii). n = 4 in two independent experiments. The differences of means among groups were analyzed by one-way ANOVA with Bonferroni’s post-hoc tests. *p < 0.05, **p < 0.01. C) Representative immunofluorescence images of ALP expression in the HHDPCs maintained on the NA gradient bound with bWnt3a (LOL = 12, 100 ng of input) for 24 h i) and the quantification of the area ii) and the level iii) of ALP expression. Scale bar: 100 µm. The differ- ences of means among groups were analyzed by Welch’s ANOVA with Dunnett’s T3 post-hoc tests. *p < 0.05, ***p = 0.0002, ****p < 0.0001. ns: not significant (p > 0.05). n = 9, 26, 32, and 30 cells in 18, 27, 36, and 45 mW group, respectively. D) Representative immunofluorescence images of BMP2 expression in the HHDPCs maintained on the NA gradient bound with bWnt3a (LOL = 12, 100 ng of input) for 24 h i) and the quantification of the area ii) and the level iii) of BMP2 expression. Scale bar: 100 µm. The differences of means among groups were analyzed by Welch’s ANOVA with Dunnett’s T3 post-hoc tests. ns: not significant (p > 0.05). n = 21, 20, 17, and 24 cells in 18, 27, 36, and 45 mW group, respectively. E) Representative immunofluo- rescence images of versican expression in the HHDPCs maintained on the NA gradient bound with bWnt3a (LOL = 12, 100 ng of input) for 24 h i) and the quantification of the area ii) and the level iii) of versican expression. Scale bar: 100 µm. The differences of means among groups were analyzed by Welch’s ANOVA with Dunnett’s T3 post-hoc tests. ns: not significant (p > 0.05). n = 12, 15, 39, and 38 cells in 18, 27, 36, and 45 mW group, respectively.
Article Snippet: Specifically, cells undergone fixation and permeabilization were incubated with mouse anti-fibronectin primary antibody (sc-8422, Santa Cruz Biotechnology, Dallas, TX, USA) (1:200) and rabbit anti-β-catenin primary antibody (ab16051, Abcam) (1:150) at 4 °C for overnight.
Techniques: Cell Culture, Immunofluorescence, Expressing