Differential action of polyunsaturated fatty acids and eicosanoids on bleomycin-induced cytotoxicity to neuroblastoma cells and lymphocytes

Abstract

IntroductionThis study was conducted to examine whether bleomycin-induced growth inhibitory action on human neuroblastoma cells (IMR-32) is influenced by anti-inflammatory metabolites of polyunsaturated fatty acids (PUFAs): lipoxin A4 (LXA4), resolvin D1 and protectin D1 in vitro.Material and methodsThe in vitro study was conducted using monolayer cultures of exponentially growing IMR-32 cells. The effects of various PUFAs and eicosanoids and anti-inflammatory metabolites of PUFAs such as lipoxin A4 (LXA4), resolvin D1 and protectin D1 on the growth of IMR-32 cells and human lymphocytes in vitro were investigated. The potential of PUFAs, eicosanoids and LXA4, resolvin D1 and protectin D1 to modify the growth inhibitory effects of bleomycin was also studied in IMR-32 cells and human lymphocytes.ResultsPUFAs inhibited the growth of IMR-32 cells (EPA > DHA = AA > GLA = ALA > DGLA = LA) significantly (p < 0.001) while prostaglandins were found to be not effective. Bleomycin-induced growth inhibitory action on IMR-32 cells was augmented by PUFAs and its metabolites (p < 0.05). PUFAs and LXA4 did not inhibit the growth of human lymphocytes and bleomycin-induced growth inhibitory action was also not enhanced by these bioactive lipids.ConclusionsBioactive lipids have differential action on normal human lymphocytes and tumor cells in vitro. The apparent lack of effect of PUFAs in combination with bleomycin on the growth of human lymphocytes in comparison to their growth inhibitory action on IMR-32 cells suggests that PUFAs can be used in combination with bleomycin to target tumor cells with little concern over this combination’s effect on the growth of human lymphocytes. Further studies are warranted to evaluate these differential effects under in vivo conditions.

n‐6 and n‐3 Fatty acids and their metabolites augment inhibitory action of doxorubicin on the proliferation of human neuroblastoma (IMR‐32) cells by enhancing lipid peroxidation and suppressing Ras, Myc, and Fos

“…The pellet of cells obtained were counted in a hemocytometer and at least >90% of the cells were found to be viable by trypan blue dye exclusion method. These cells were used for further studies .…”

“…The cells were allowed to attach for 24 h by which time they reached 80% confluence . At the end of 24 h, cells were treated with different doses of doxorubicin (0.044–1.4 µg/mL) and incubated for further 24 and 48 h. At the end of 24 and 48 h, cell viability was assayed by MTT assay (3‐(4,5‐dimethylthiazole‐2‐yl)‐2, 5‐diphenyltetrazolium bromide) as described previously . The percentage of cell proliferation was expressed as percentage of cell growth compared with control .…”

“…IMR‐32 cells were seeded at a density of 1 × 10 4 /100 µL of culture media per well in 96‐well plates. The cells were allowed to attach for 24 h, at the end of which cells were supplemented with different doses of n‐6, n‐3 PUFAs, their metabolites prostaglandins/leukotrienes/lipoxin A4/resolvins/protectins and incubated for an additional 24 h. At the end of the treatment, cell viability was assessed by MTT assay as described previously .…”

“…After attachment period, cells were treated simultaneously with AA/EPA (30 μg/mL), doxorubicin (0.175 μg/mL) for 24 h. After the treatment period, spent was collected and cells were washed with 500 μL of PBS (pH 7.4). Cells were lysed with NET lysis buffer, both cell lysate and supernatant were used for measuring nitric oxides and lipid peroxides production and only the lysate was used for measuring the activities of various antioxidant enzymes as described previously [7,9,[31][32][33][34][35]].…”

“…We also evaluated the effect of AA and EPA and doxorubicin alone and in combination on the expression of various oncogenes/anti‐oncogenes including Ras, Myc and Fos , NF‐kB, IkB, and NF‐kB/IkB ratio; BCL‐2/BAX ratio, cytochrome C, caspases 9 and 3, p53, and 5‐LOX. We selected IMR‐32 cells for this study since, our previous studies showed that PUFAs have selective growth inhibitory action on these cells in vitro and as brain tumors are highly resistant to conventions treatments and have poor prognosis and hence, are in need of newer therapeutic options.…”