Good god of course it was tested on human cells.... Please don't just make things up and then go around acting like some kind of authority when you haven't even bothered to do even a minimum level of research.
From page 8 of the study.
CXXC5 attenuates ALP activity and proliferation in human hair follicle dermal
papilla cells via interaction with Dvl-1
The role of CXXC5 in modulating ALP activity and cell proliferation was investigated using
human hair follicle dermal papilla cells (HFDPCs). Transfection with CXXC5 significantly
inhibited expression of β-catenin, ALP, and PCNA (Figure 2a). Analyses using
immunocytochemical and ALP staining also showed that the levels of both β-catenin and
were diminished by CXXC5 transfection (Figure 2b). In contrast, CXXC5 knockdown
induced the expression of β-catenin, ALP and PCNA (see Supplementary Figure S3). CXXC5
possesses a Dvl-binding motif (DBM) at its C terminus that is essential for CXXC5 function
as a negative regulator of the Wnt/β-catenin pathway (London et al., 2004). Transfection with
CXXC5∆DBM, which lacks the DBM (London et al., 2004), did not inhibit the expression of
β-catenin and ALP, as well as PCNA in HFDPCs (Figure 2c). Overexpression of CXXC5
markedly reduced the transcriptional activity of a Wnt/β-catenin reporter gene (Figure 2d).
Furthermore, ALPL promoter activity and ALP activity were significantly lowered by
CXXC5 overexpression (Figures 2e and f). Cell proliferation was also decreased in HFDPCs
by 66% at 72 hours after transfection with CXXC5 (Figure 2g). To further clarify the role of
the Wnt/β-catenin pathway in CXXC5 function, we confirmed the effect of Wnt3a in
HFDPCs. Interestingly, β-catenin and CXXC5 were concomitantly increased in human
dermal papilla cells treated with Wnt3a as shown by western blot and immunocytochemical
analyses (see Supplementary Figure S4). Based on in vivo results, we expected Wnt3a-
dependent induction of CXXC5 to be a negative feedback mechanism. We also found that
CXXC5 interacted with Dvl-1 in human hair follicle cells, but only detected this interaction
when HFDPCs were treated with Wnt3a (Figure 2h). However, CXXC5∆DBM failed to
interact with Dvl-1 in HFDPCs, even when the cells were treated with Wnt3a (Figure 2h).
Endogenous CXXC5 also bound to Dvl-1 in Wnt3a-treated HFDPCs as shown by western
blot analyses of immunoprecipitates (see Supplementary Figure S5). To confirm the
possibility of further Wnt/β-catenin pathway activation after disrupting the negative feedback
regulation of CXXC5, we co-treated HFDPCs with both CXXC5 siRNA and Wnt3a.
Expression of β-catenin, ALP, and PCNA were synergistically increased by co-treatment with
CXXC5 siRNA and Wnt3a (Figures 2i and j). In summary, our data indicate that CXXC5
interacts with Dvl in a Wnt3a-dependent manner to negatively regulate the Wnt/β-catenin
pathway in HFDPCs.
