Lipid Raft Integrity Is Required for Survival of Triple Negative Breast Cancer Cells.
10.4048/jbc.2016.19.4.372
- Author:
Anil BADANA
1
;
Madhuri CHINTALA
;
Gayathri VARIKUTI
;
Nagaseshu PUDI
;
Seema KUMARI
;
Vijaya Rachel KAPPALA
;
Rama Rao MALLA
Author Information
1. Cancer Biology Lab, Department of Biochemistry, GIS, GITAM University, Visakhapatnam, India. dr.rrmalla@gmail.com
- Publication Type:Original Article
- Keywords:
Beta-cyclodextrin;
Cholesterol;
Membrane microdomains;
Triple negative breast neoplasms
- MeSH:
Apoptosis;
Caveolin 1;
Cell Adhesion;
Cell Cycle;
Cell Cycle Checkpoints;
Cell Line;
Cell Proliferation;
Cell Survival;
Cholesterol;
Detergents;
Fibronectins;
Filipin;
Flow Cytometry;
In Vitro Techniques;
Membrane Microdomains;
Membranes;
Methods;
Neoplasm Metastasis;
Nystatin;
Protein-Tyrosine Kinases;
Triple Negative Breast Neoplasms*;
Vitronectin
- From:Journal of Breast Cancer
2016;19(4):372-384
- CountryRepublic of Korea
- Language:English
-
Abstract:
PURPOSE: Lipid rafts are cholesterol enriched microdomains that colocalize signaling pathways involved in cell proliferation, metastasis, and angiogenesis. We examined the effect of methyl-β-cyclodextrin (MβCD)-mediated cholesterol extraction on the proliferation, adhesion, invasion, and angiogenesis of triple negative breast cancer (TNBC) cells. METHODS: We measured cholesterol and estimated cell toxicity. Detergent resistant membrane (DRM) and non-DRM fractions were separated using the OptiPrep gradient method. Cell cycles stages were analyzed by flow cytometry, apoptosis was assessed using the TdT-mediated dUTP nick end-labeling assay, and metastasis was determined using a Matrigel invasion assay. Neo-vessel pattern and levels of angiogenic modulators were determined using an in vitro angiogenesis assay and an angiogenesis array, respectively. RESULTS: The present study found that the cholesterol-depleting agent MβCD, efficiently depleted membrane cholesterol and caused concentration dependent (0.1–0.5 mM) cytotoxicity compared to nystatin and filipin III in TNBC cell lines, MDA-MB 231 and MDA-MB 468. A reduced proportion of caveolin-1 found in DRM fractions indicated a cholesterol extraction-induced disruption of lipid raft integrity. MβCD inhibited 52% of MDA-MB 231 cell adhesion on fibronectin and 56% of MDA-MB 468 cell adhesion on vitronectin, while invasiveness of these cells was decreased by 48% and 52% respectively, following MβCD treatment (48 hours). MβCD also caused cell cycle arrest at the G2M phase and apoptosis in MDA-MB 231 cells (25% and 58% cells, respectively) and in MDA-MB 468 cells (30% and 38% cells, respectively). We found that MβCD treated cells caused a 52% and 58% depletion of neovessel formation in both MDA-MB 231 and MDA-MB 468 cell lines, respectively. This study also demonstrated that MβCD treatment caused a respective 2.6- and 2.5-fold depletion of tyrosine protein kinase receptor (TEK) receptor tyrosine kinase levels in both TNBC cell lines. CONCLUSION: MβCD-induced cholesterol removal enhances alterations in lipid raft integrity, which reduces TNBC cell survival.
