Δ9-THC modulation of fatty acid 2-hydroxylase (FA2H) gene expression: Possible involvement of induced levels of PPARα in MDA-MB-231 breast cancer cells

Abstract

We recently reported that Δ9-tetrahydrocannabinol (Δ9-THC), a major cannabinoid component in Cannabis Sativa (marijuana), significantly stimulated the expression of fatty acid 2 hydroxylase (FA2H) in human breast cancer MDA-MB-231 cells. Peroxisome proliferator-activated receptor α (PPARα) was previously implicated in this induction. However, the mechanisms mediating this induction have not been elucidated in detail. We performed a DNA microarray analysis of Δ9-THC treated samples and showed the selective up-regulation of the PPARα isoform coupled with the induction of FA2H over the other isoforms (β and γ). Δ-THC itself had no binding/activation potential to/on PPARα, and palmitic acid (PA), a PPARα ligand, exhibited no stimulatory effects on FA2H in MDA MB 231 cells; thus, we hypothesized that the levels of PPARα induced were involved in the Δ9-THC mediated increase in FA2H. In support of this hypothesis, we herein demonstrated that; (i) Δ9 THC activated the basal transcriptional activity of PPARα in a concentration-dependent manner, (ii) the concomitant up-regulation of PPARα/FA2H was caused by Δ9-THC, (iii) PA could activate PPARα after the PPARα expression plasmid was introduced, and (iv) the Δ9-THC induced up regulation of FA2H was further stimulated by the co-treatment with L-663,536 (a known PPARα inducer). Taken together, these results support the concept that the induced levels of PPARα may be involved in the Δ9 THC up-regulation of FA2H in MDA-MB-231 cells.

Bongkrekic acid as a selective activator of the peroxisome proliferator-activated receptor γ (PPARγ) isoform

“…As expected, LA stimulated the transcriptional activities of PPARβ (1.19 ± 0.01, p < 0.05) (possibly via 13-(S)-HODE), in addition to PPARα/γ (1.65 ± 0.03 and 1.43 ± 0.06, p < 0.05); MCF-7 cells are known to express a functional 15-LOX enzyme (Kerjaschki et al, 2011). Although some compounds such as ∆ 9 -tetrahydrocannabinol, a major component of marijuana, can activate the PPARα isoform by up-regulating the receptor (Takeda et al, 2014), no observable basal transcriptional activities were detected even at 25 μM BKA (Fig. 3A).…”

“…All plasmid concentrations were equalized with the pcD-NA3.1 vector. Expression plasmids of human PPARα/β/γ, RXRα, and the PPRE reporter construct were gifts from Dr. Curtis J. Omiecinski (Pennsylvania State University, PA, USA) (Takeda et al, 2014). At 24-hr post-transfection, cells were washed with phosphate-buffered saline and changed to MEMα without phenol red supplemented with 5% serum, followed by a treatment with BKA and its derivatives (BKA-1, BKA-2, and BKA-3) or LA for 24 hr.…”

“…The primers used were adiponectin (sense) 5'-GGT CTC GAA CTC CTG GCC TA-3' and adiponectin (antisense) 5'-TGA GAT ATC GAC TGG GCA TGG T-3' (Favre et al, 2011). Primers for the PCR of FA2H and β-actin were taken from previous studies (Takeda et al, 2014). The reaction conditions for all mRNA were 95°C for 10 min, followed by 45 cycles at 95°C for 10 sec, at 56°C for 10 sec, and 72°C for 15 sec.…”

Bisphenol AF as an Inducer of Estrogen Receptor β (ERβ): Evidence for Anti-estrogenic Effects at Higher Concentrations in Human Breast Cancer Cells

“…[17][18][19] Briefly, the human breast cancer cell lines, MCF-7 and SK-BR-3 (obtained from the American Type Culture Collection, Rockville, MD, U.S.A.) were routinely grown in phenol red-containing minimum essential medium (MEMα) (Invitrogen, Carlsbad, CA, U.S.A.) supplemented with 10 mM HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid), 5% fetal bovine serum, 100 U/mL of penicillin, and 100 µg/mL of streptomycin in a humidified incubator within an atmosphere of 5% CO 2 at 37°C. Prior to the chemical treatments, medium was changed to phenol red-free MEMα supplemented with 10 mM HEPES, 5% dextran-coated charcoal-treated serum, 100 U/mL of penicillin, and 100 µg/mL of streptomycin.…”

A Novel Bongkrekic Acid Analog-Mediated Modulation of the Size of Lipid Droplets: Evidence for the Appearance of Smaller Adipocytes

“…The PPRE reporter construct was a gift from Dr. Curtis J. Omiecinski (Pennsylvania State University, PA, U.S.A.). 25) Twenty-four hours after transfection, culture medium was replaced by phenol red-free DMEM supplemented with 10% charcoal-stripped FBS, followed by a treatment with BKA, its analogs, and PIO. Cell extracts were prepared with 100 µL passive lysis buffer (Promega) 24 h after the treatments with the compounds, and 20 µL of the extracts was used in dual-luciferase assays by the GloMAX-Multi Detection System (Promega).…”