Figure 5.

Functional validation of evRSPO1 in vitro

(A) TOPFlash reporter assay in HEK293 cells treated with evWT and evRSPO1 (n = 3).

(B) Western blot analysis of unphosphorylated β-catenin and total β-catenin in evWT- or evRSPO1-treated HEK293 cells.

(C) Representative confocal images of Axin2-mGFP expression in evWT- or evRSPO1-treated intestinal organoids.

(D) Quantification of the Axin2-mGFP signals in intestinal organoids from (C) (n = 3).

(E) Representative bright-field images of evWT- or evRSPO1-treated intestinal organoids.

(F) Quantification of the number of crypt-villus structures in intestinal organoids from (E) (n = 3). Statistical data are presented as mean ± SD. Two-tailed unpaired Student’s t-tests were used to compare the two groups. The p-values are calculated and evaluated for statistical significance (ns p ≥ 0.05 and ∗∗∗∗p < 0.0001).