Effects of low-density lipoprotein docosahexaenoic acid nanoparticles on cancer stem cells isolated from human hepatoma cell lines

Abstract

Docosahexaenoic acid (DHA) is an omega-3 polyunsaturated fatty acid with anti-cancer properties. Recently, DHA packaged within low-density lipoprotein (LDL) nanoparticles (LDL-DHA) was demonstrated to be effective in a murine model of hepatocellular carcinoma (HCC). Cancer stem cells (CSCs) are a subpopulation of tumor cells that are resistant to most cancer therapies and thereby, contribute to tumor recurrences. To determine whether LDL-DHA is effective against CSCs derived from human HCC cell lines and tumor bearing rats. Epithelial cellular adhesion molecule positive and CD133 negative (EpCAMCD133) CSCs were isolated from HuH-7 and HepG2 human HCC lines and exposed to varying concentrations (1-60 µM) of LDL-DHA nanoparticles for 0-72 h. HCC tumor bearing rats were treated with 2 mg/kg LDL-DHA nanoparticles for 3 days. Regardless of the cell line employed, LDL-DHA nanoparticles achieved 70-100% killing of EpCAMCD133 CSCs at a concentration of 40 µM after 48 h of exposure while DHA and LDL alone had minimal or no cytotoxic effects. Similar results were obtained with LDL-DHA nanoparticle treatment of EpCAMCD133 adult cancer cells (ACCs). In keeping with these findings were similar levels of low density lipoprotein receptor expression and LDL-DHA nanoparticle induced lipid peroxidation activity and reactive oxygen species in the CSC and ACC populations. However, differences in sensitivity to LDL-DHA treatment were observed in vivo experiments where LDL-DHA nanoparticle treated tumors had a higher percent of surviving CSCs relative to ACCs. Further research on LDL-DHA nanoparticle therapy for human HCC is warranted.

Comparisons of Growth Features and Cancer Stem Cell Prevalences in Intrahepatic and Extrahepatic Cholangiocarcinoma Cell Lines

“…Cell proliferation and doubling times were determined by seeding CCA cells at a density of 2000 cells per well in 96well plates. The number of cells in each well were counted on days 0, 1, 3 and 6 after plating by adding a cell proliferation indicator: premixed WST-1 reagent (Takara Bro, USA) and incubating at 37ºC for 3 hours as described 12 .…”

“…Cells (0.5×10 6 /mL, 2 mL per well) were seeded in 6-well plates and cultured with culture medium with 10% FBS until con uent. Cell monolayers were scratched with a 200 μL pipette tip to generate a wound according to the method described previously 12…”

“…Cells were trypsinized and resuspended in DMEM/F12 medium, supplemented with 1×B-27 serum-free supplement, 4 μg/mL heparin, 100 U/mL penicillin, 100 μg/mL streptomycin, 10 ng/mL EGF, and 10 ng/mL bFGF 12 . Cells were subsequently seeded at densities of 200 cells per well in ultra-low attachment 96-well plates.…”

“…and epithelial cell adhesion molecule (EpCAM) 11,12 . CSCs are thought to be responsible for tumor initiation, growth, migration, invasion and metastasis and each SCSM has been associated with speci c tumor cell features 13,14 .…”

“…CSCs are a subpopulation of tumor cells that have the ability to self-replicate and differentiate into different cell lineages 6 . They can be identi ed by the expression of certain SCSMs including CD13 7 , CD24 8 , CD44 9,10 , CD90, CD133 and epithelial cell adhesion molecule (EpCAM) 11,12 . CSCs are thought to be responsible for tumor initiation, growth, migration, invasion and metastasis and each SCSM has been associated with speci c tumor cell features 13,14 .…”