Cellular Pharmacology of Curcumin With and Without Piperine

Prior reports have suggested that piperine enhances curcumin anti-carcinogenesis. We tested the hypothesis that piperine increases the intracellular concentrations of curcumin by improving intracellular uptake or reducing curcumin efflux or metabolism in breast cells. We incubated SUM149, MCF10A, primary normal human breast cells, ALDH+, and ALDH-CD44+24- SUM149 cells with curcumin with or without piperine at concentrations 1 uM to 15 uM for time periods of 15 minutes to 24 hours. We assayed cell viability by MTT assay and proliferation by primary mammosphere assay. Curcumin and its metabolites were assayed using liquid chromatography mass spectroscopy. Curcumin, but not piperine, showed significantly higher effects on the viability of breast cancer SUM149 cells than in non-tumorigenic MCF10A cells. Curcumin + piperine synergistically reduced viability of SUM149 cells but had a concentration dependent effect upon MCF10A cell viability. Cellular uptake of curcumin in SUM149 is significantly higher, while the efflux in SUM149 is significantly lower than in MCF10A, which correlated with cell viability. Piperine did not alter curcumin cellular uptake, efflux, or metabolism in any of the cell models. The observed synergism of piperine+curcumin in reducing breast stem cell self renewal is likely due to independent anti-carcinogenesis effects rather than any effects upon intracellular curcumin concentrations.

Cadmium exposure inhibits branching morphogenesis and causes alterations consistent with HIF-1α inhibition in human primary breast organoids

ABSTRACTBackgroundDevelopmental cadmium exposure in vivo disrupts mammary gland differentiation, while exposure of breast cell lines to cadmium causes invasion consistent with the epithelial-mesenchymal transition (EMT). The effects of cadmium on normal human breast stem cell development have not been measured.ObjectiveThe objective of this study was to quantify the effects of cadmium exposure on normal breast stem cell proliferation and differentiation.MethodsWe tested the effects of two physiologically relevant doses of cadmium: 0.25µM and 2.5μM on reduction mammoplasty patient-derived breast cells using the mammosphere assay, organoid formation in 3D hydrogels, and tested for molecular alterations using RNA-seq. We functionally validated our RNA-seq findings with a HIF-1α transcription factor activity reporter line and pharmaceutical inhibition of HIF-1α in mammosphere and organoid formation assays.Results2.5μM cadmium reduced primary and secondary mammosphere formation and branching structure organoid formation rates by 33%, 40%, and 83%, respectively. Despite no changes in mammosphere formation, 0.25μM cadmium treatment inhibited branching organoid formation in hydrogels by 68%. RNA-seq revealed that cadmium treatment downregulated genes associated with extracellular matrix formation and EMT, while upregulating genes associated with metal response including metallothioneins and zinc transporters. In the RNA-seq data, cadmium treatment also downregulated HIF-1α target genes including LOXL2, ZEB1, and VIM. Cadmium treatment significantly inhibited HIF-1α activity in a luciferase assay, and the HIF-1α inhibitor acriflavine ablated mammosphere and organoid formation.DiscussionThese findings show that cadmium, at doses relevant to human exposure, inhibited human mammary gland development, potentially through disruption of HIF-1α activity. These findings do not support cadmium being a breast cancer initiator via induction of stem cell proliferation, but instead implicate cadmium as an inhibitor of mammary gland morphogenesis.