Celecoxib, a COX-2 inhibitor, synergistically potentiates the anti-inflammatory activity of docosahexaenoic acid in macrophage cell line.

BACKGROUND: The anti-inflammatory properties of eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA) and non-steroidal anti-inflammatory drugs overlap in many ways. The aim of this study was to examine the individual and synergetic anti-inflammatory effects of celecoxib, EPA and DHA in RAW-264.7 cell line. METHODOLOGY: The cells were exposed to EPA, DHA, celecoxib, rosiglitazone, GW9662 alone or their combination, and stimulated with 5 µg/mL lipopolysaccharide (LPS). Nitric oxide (NO), tumor necrosis factor-α (TNF-α), interleukin (IL)-6 and prostaglandin-E2 (PGE2) levels were estimated in the medium using enzyme-linked immunosorbent assays. The cyclooxygenase-2 (COX-2) and inducible Nitric Oxide Synthase (iNOS) expression were analyzed in the cell lysate by immunoblotting. Peroxisome proliferator-activated receptor γ (PPARγ) and nuclear factor-κB (NF-κB) transcription factor activation assays were performed in the nuclear extract. RESULTS: Combined treatment of DHA (50 µM) and celecoxib (20 µM) significantly inhibited LPS induced synthesis of NO, TNF-α, IL-6 and PGE2 levels in the cells, compared to the individual treatments. In addition, DHA and celecoxib diminished the COX-2 and iNOS expression in the cells. This was associated with increased PPARγ activity, supressed NF-κB activity in the nucleus. We determined whether GW9662, a specific PPARγ inhibitor, could abolish the anti-inflammatory effect of DHA and celecoxib. GW9662 has abolished the DHA and celecoxib induced PPARγ activation, but did not alter the NF-κB mediated anti-inflammatory effects induced by celecoxib and DHA. Interestingly, EPA did not exhibit any inhibitory effect on these parameters. CONCLUSION: Our results suggest that DHA and celecoxib exhibit anti-inflammatory effect through inhibition of NF-κB, independent of PPARγ. Co-administration of celecoxib and DHA would be promising approach for the treatment of inflammatory diseases.

Ascorbyl stearate stimulates cell death by oxidative stress-mediated apoptosis and autophagy in HeLa cervical cancer cell line in vitro.

In this study, Asc-s was evaluated for anti-cancer effect using cervical cancer cells (HeLa). Results determine that Asc-s treatment-induced dose-dependent inhibition of proliferation of HeLa cells and induced apoptosis. Flow-cytometry analysis shows Asc-s treatment-induced accumulation of cells at sub-G0/G1 stage of cell cycle and induced apoptosis as confirmed by DAPI, propodium iodide, and acridine staining in HeLa cells. Asc-s entered the cells and metabolized to ascorbate and stearate moieties, increased membrane permeability, and decreased membrane fluidity in HeLa cells. Asc-s treatment-induced dose-dependent increase in autophagy protein LC3-II, mRNA levels and decreased Nrf-2 levels in HeLa cells. It is hypothesized that both ascorbyl radical and stearoyl moieties of Asc-s induced cytotoxicity by generating reactive oxygen species (ROS) and modulating membrane fluidity/permeability leading to apoptosis/autophagy of HeLa cells. Thus, our findings demonstrate that Asc-s as anti-proliferative and apoptosis inducing compound in cervical cancer cells.

Ascorbyl stearate and ionizing radiation potentiate apoptosis through intracellular thiols and oxidative stress in murine T lymphoma cells.

Ascorbyl stearate (Asc-s) is a derivative of ascorbic acid with better anti-tumour efficacy compared to its parent compound ascorbic acid. In this study, we have examined radio-sensitizing effect of Asc-s in murine T cell lymphoma (EL4) cells at 4 Gy. Asc-s and radiation treatment reduced cell proliferation, induced apoptosis in a dose dependent manner by arresting the cells at S/G2-M phase of cell cycle. It also decreased the frequency of cancer stem cells per se, with significantly higher decrease in combination with radiation treatment./Further, Asc-s and radiation treatment increased the level of reactive oxygen species (ROS), drop in mitochondrial membrane potential (MMP) and increased caspase-3 activity resulting in apoptosis of EL4 cells. Further it also significantly decreased GSH/GSSG ratio due to binding of Asc-s with thiols. The increase in oxidative stress induced by Asc-s and radiation treatment was abrogated by thiol antioxidants in EL4 cells. Interestingly, this redox modulation triggered significant increase in protein glutathionylation in a time dependent manner. Asc-s treatment resulted in glutathionylation of IKK, p50-NF-kB and mutated p53, thereby inhibiting cancer progression during oxidative stress. Asc-s quenches GSH ensuing Asc-s + GSH adduct thereby further modulating GSH/GSSG ratio as evident from HPLC and docking studies. The anti-tumour effect of Asc-s along with radiation was studied by injecting EL4 cells in synegenicC57/BL6 male mice. Intraperitoneal injection of Asc-s followed by radiation exposure at 4 Gy to the tumour bearing mice resulted in radio-sensitization which is evident from significant regression of tumour as evident from tumour burden index. The survival study supports the data that Asc-s pre-treatment enhances radio-sensitization in murine lymphoma. Our data, suggest that Asc-s and ionizing radiation induced cell cycle arrest and apoptosis by perturbing redox balance through irreversible complexes of thiols with Asc-s, disturbed mitochondrial membrane permeability and activation of caspase-3 in EL4 cells.