Curcuminoids Modulate TGF-β1-induced PTHrP in Breast Cancer

Sub-confluent human breast cancer MDA-MB-231 cells were plated at 5 × 10⁴ cells/cm² and incubated overnight in fresh DMEM medium containing 10% fetal bovine serum (FBS) and antibiotics at 37°C and 5% CO₂ in a humidified atmosphere. Cells were pre-incubated in DMEM medium containing 2% FBS with or without curcuminoids (0.1 – 100 μM) for 4 h, and the medium was then refreshed with the addition of recombinant human TGF-β1 (5 ng/mL; R&D Systems) for 24 h. PTHrP expression was measured in conditioned media by commercial radioimmunoassay (Diagnostic Systems Laboratories) and cell number and cytotoxicity were assessed by mitochondrial reduction of MTT (ATCC) and conditioned media content of lactate dehydrogenase (LDH; Promega), respectively, as per manufacturer’s protocol.