Fig 1: Migration of melanoma cells in an APP-rich microenvironment. (A) Treatment with 100 nM of exogenous soluble rhAPP did not affect the migration of C8161 cells compared to untreated control, but significantly reduced migration of WM1552C compared to untreated control (paired t-test, * p = 0.02). (B) Migration of C8161 cells was significantly increased towards brain tissue of AD female (AD-F) (paired t-test, * p = 0.007) and AD male (AD-M) (paired t-test, * p = 0.03) mice compared to brain tissue from age- and sex-matched wildtype (WT) control mice. No significant (ns) difference was observed between C8161 migration towards AD-F and AD-M brain tissue. (C) No significant difference was observed in the migration of WM1552C cells towards AD-F compared to WT-F brain tissue. However, migration of WM1552C cells was significantly reduced towards AD-M compared to WT-M brain tissue (paired t-test, * p = 0.005). Additionally, migration of WM1552C toward AD-M brain tissue was significantly lower than toward AD-F brain tissue (paired t-test, * p = 0.03). (D) Microphotographs of inserts with stained cells representative of the migration results towards the different types of mice brain tissues used are shown. Data are presented as the mean fold OD of control ± SEM of 4 independent experiments.
Fig 2: Effect of DAPT treatment on APP and Aβ expression in C8161 cells. Densiometric analysis of WB bands in (A) shows a significant increase in APP expression in C8161 melanoma cells after 48 h treatment with 30 μM DAPT compared to DMSO control (paired t-test, * p = 0.02) (B). Representative WB in (C) from two independent experiments also shows concomitant expression of low-molecular-weight protein in DAPT-treated C8161, presumably APP degradation product of less than 20 kDa (arrow), in DAPT-treated C8161 cells that is not detected in control cells. C8161 cells treated with 30 μM DAPT ± 100 nM rhAPP show significant increased production of Aβ40 (paired t-test, * p = 0.04) (D) and Aβ42 (paired t-test, * p = 0.04) (E) in conditioned media from DAPT + rhAPP-treated cells compared to DAPT + PBS-treated controls. Data are presented as the mean fold OD of control ± SEM of at least 3 independent experiments.
Fig 3: Endogenous APP expression in melanoma cells. Representative WB showing APP expression in WM1552C and C8161 melanoma cells. A higher expression of a lower-molecular-weight protein was occasionally detected in WM1552C compared to C8161 (arrow). The histogram summarizing the densitometric analysis of the optical density (OD) of the WB bands shows that PS-1-high WM1552C melanoma cells express a significantly lower amount of APP compared to PS-1-low C8161 melanoma cells. OD data are presented as the mean fold change in WM1552C ± SEM, of 3 independent experiments. (unpaired t-test, * p = 0.001).
Supplier Page from BioLegend for Recombinant Human APP751