Sphingosine kinase inhibitor, SKI-II confers protection against the ionizing radiation by maintaining redox homeostasis most likely through Nrf2 signaling.

Exposure to ionizing radiation (IR) set a series of deleterious events causing acute radiation syndrome and mortality, posing the need for a potent and safe radio-protective drug. IR induces cell death predominantly by causing oxidative stress and macromolecular damage. The pre-existing antioxidant defence machinery of the cellular system plays a crucial role in protecting the cells against oxidative stress by activation of Nrf2. The current study was undertaken to investigate the radio-protective potential of sphingosine kinase inhibitor (SKI-II), which was demonstrated to activate Nrf2 signaling. The safety and efficacy of SKI-II were evaluated with cell cytotoxicity, proliferation index, and clonogenic survival assays in different cell lines, namely Raw 264.7, INT-407, IEC-6 and NIH/3T3 cell lines. A safe dose of SKI-II was found radio-protective in all the cell lines linked with the activated antioxidant defence system, thereby resulting in the amelioration of IR induced oxidative stress. SKI-II pretreatment also significantly reduced DNA damage, micronuclei expression, and accelerated DNA repair kinetics as compared to IR exposed cells. Reduced oxidative stress and enhanced DNA repair significantly reduced apoptosis and suppressed the pro-death signaling associated with IR exposure. Furthermore, the in-vitro observation was verified in the in-vivo model (C57 BL/6). The Intra-peritoneal (IP) administration of SKI-II, 2 h before a lethal dose of IR exposure (7.5 Gy) resulted in 75% survival. These results imply that SKI-II ameliorates IR-induced oxidative stress and cell death by inducing anti-oxidant defence system and DNA repair pathways, thus strengthening its potential to be used as radiation countermeasure.

3,4-Dimethoxyphenyl bis-benzimidazole derivative, mitigates radiation-induced DNA damage.

Radiation-induced DNA damage initiates a series of overlapping responses that include DNA damage recognition and repair, induction of cell cycle checkpoints, senescence and/or apoptosis. This study assessed the DNA damage response and whole genome expression profile in two mammalian cell lines (HEK and U87) in response to (5-{4-methylpiperazin-1-yl}-2-[2'-(3,4-dimethoxyphenyl)-5'-benzimidazolyl] benzimidazole) DMA and ionizing radiation. DMA has been shown to act as a potent radiation protector, yielding significant levels of protection, i.e., 20.9% in HEK cells and 21.2% in U87 cells. Our findings revealed treatment with DMA significantly reduced γ-H2AX, 53BP1 and Rad51 foci formation after irradiation. MAP kinase, WNT signaling and p53 pathways were found to be activated in DMA-treated cells. In addition, the DNA damage response genes, HSP70, HSPD1, PRDX1, PRX, CALR, NPM, UBC, and SET showed differential regulation in DMA, DMA + radiation and radiation-treated cells. The data suggest that DMA-influenced repertoire of repair proteins, which are an indispensable part of the cell, interplay with each other to reduce DNA damage and maintain the genomic integrity of the cell.

Role of tumor necrosis factor-α in Gram-negative bacterial lipopolysaccharides induced implantation failure.

Background and aims: Gram-negative bacterial lipopolysaccharides (LPS) are known causative agents for pregnancy loss in mothers with genital tract infections. In this study, we attempt to test the role of tumor necrosis factor-α (TNF-α) in the normal physiological processes of preimplantation embryonic development and LPS induced pregnancy loss in mice. Since the preimplantation mouse embryos grow in an unattached state for a considerable period (day 1-4.5) of its development in the maternal environment, it is possible that a critical level of soluble and biologically active TNF-α is maintained in the maternal environment, and that any alteration in this could lead to implantation failure. Here we determine the pattern and level of expression of TNF-α gene in preimplantation stage embryos and uterus collected from control and LPS treated pregnant animals during different stages of preimplantation period of pregnancy by reverse transcriptase-polymerase chain reaction. Methods: The concentrations and biological activity of soluble TNF-α protein present in oviductal fluid (OF) and uterine fluid (UF), in the normal and LPS treated animals, were determined by enzyme-linked immunosorbent assay and 3-(4,5-dimethylthiazol-2-yl) 2,5-diphenyltetrazolium bromide assay on L929 cells, respectively. TNF-α was also given i.p. to study its effect on implantation. Results: An early expression of TNF-α messenger ribonucleic acid in the preimplantation stage embryos collected from LPS treated animals was observed along with a significant rise in the level of biologically active soluble TNF-α in the OF. Similarly, the level of bioactive and soluble TNF-α present in the UF from LPS treated animals was significantly higher as compared to the control on day 4.42 of pregnancy. Conclusions: TNF-α given i.p. exerted similar effects on pregnancy as that of LPS. An incessant exposure of the preimplantation stage embryos to significantly high levels of maternal bioactive free/soluble TNF-α, and an alteration in the normal pattern of its expression in the preimplantation stage embryos may be one of the causes of failure of implantation leading to poor pregnancy outcome in LPS treated mouse. (Reprod Med Biol 2005; 4: 79- 89).