Prebiotic Mannan Oligosaccharide Pretreatment Improves Mice Survival Against Lethal Effects of Gamma Radiation by Protecting GI Tract and Hematopoietic Systems.

Total body irradiation (TBI) results in critical injuries in a dose dependent manner that primarily damages highly proliferating tissues including hematopoietic stem cells (HSCs) and intestinal crypt stem cells etc. This may result in hematopoietic syndrome leading to bone marrow failure and gastrointestinal syndrome leading to chronic intestinal functional alterations. Death results from the gastrointestinal syndrome due to sepsis, bleeding, dehydration, and multi-system organ failure. We demonstrate that the prebiotic mannan oligosaccharide (MOS) pretreatment substantially prolongs survival in both male and female mice when administered 2 h prior to radiation either through oral or intraperitoneal route. The radioprotective efficacy of MOS was found to be age dependent and improves survival even in aged mice (12-13 months old). MOS pretreatment effectively abrogates radiation-induced hematopoietic injury and accelerates recovery of lymphocytes and WBCs and alleviates depletion of circulatory blood cells. Results also illustrate that MOS pretreatment abolishes crypt cell death and denudation of villi in comparison to the respective irradiated animals and ameliorates the overall radiation-induced damage to the GI system. MOS pretreatment facilitates intestinal recovery leading to enhanced animal survival demonstrating its protection efficacy against TBI induced mortality. Moreover, MOS pretreated animals show signs of accelerated recovery in terms of severity of radiation sickness symptoms including weight loss and completely abolish TBI associated mortality.

Role of Nrf2-antioxidant in radioprotection by root extract of Inula racemosa.

Purpose:Inula racemosa, a Trans-Himalayan plant is an important medicinal herb. In this study, the radio-modulatory efficacy of aqueous root extract of I. racemosa was investigated. Materials and methods: Normal Kidney Epithelium cells were treated with extract (50-200 µg/ml) and exposed to 3 Gy of γ-radiation, while C57BL/6 mice were administered with extract (300-600 mg/kg BW) intraperitoneally prior to exposure to 7.5 Gy of γ-radiation to assess radiation modulatory efficacy. Results: The administration of extract (30 min and 1 h) prior to radiation exposure improved the survival of NKE cells (as measured by proliferation), restored MMP and ROS levels as compared to radiation-exposed alone cells. These cells showed up-regulated Nrf2 protein levels at 7 h and increased expression of HO-1 and NOQ1 protein at 24 h In mice, the 30 days whole body survival study demonstrated that extract pre-treatment increases survival or delays the onset of radiation-induced mortality as against 70% mortality of 7.5 Gy of γ-radiation. Conclusions: The aqueous extract of roots of I. racemosa enhanced the survival of irradiated NKE cells and rescued C57BL/6 mice against WBI-induced mortality. The radiation modulation efficacy was mediated through cumulative activation of HO-1 and NQO1 downstream of Nrf2 translocation in NKE cells. Abbreviations: ARE: Antioxidant Response Element; FITC: Fluorescein isothiocyanate; GCS: Glutamylcysteine synthase; HO-1: Heme oxygenase-1; LPS: Lipopolysacharide; MRP: Multidrug Resistance-Associated Proteins; NQO1: NAD(P)H Quinone Dehydrogenase 1; NRH: Quinone Oxidoreductase 2 (NQO2); PBS: Phosphate Buffer Saline; PKA: Protein Kinase A; PKC: Protein Kinase C; PI3-kinase: Phosphatidylinositol 3-Kinase; SRB: Sulforhodamine B; UV: Ultra-Violet radiation.

Crosstalk of toll-like receptors signaling and Nrf2 pathway for regulation of inflammation.

Inflammation as a second line of defense of innate immunity plays a crucial role in eliminating invading pathogens (bacteria, viruses, fungi as well as other parasites). The inflammatory response may also activate adaptive immune system involving lymphocytes to mount either antibody dependent or cell-mediated immune responses to clear pathogenic insult. However, if continued, the inflammatory processes may become uncontrolled culminating in cellular injury and tissue destruction, thereby manifesting itself in chronic form. The chronic inflammation has been associated with numerous human pathological conditions like allergies and autoimmune diseases, atherosclerosis, arthritis, Alzheimer's disease, cancer, obesity, type 2 diabetes, schizophrenia, neuro-degenerative diseases and numerous others. The dysregulated inflammatory process is associated with overproduction of free radicals leading to oxidative stress and activation of different cell signaling pathways. The regulation of inflammation by TLR signaling as well as Nrf2 pathways separately is widely documented. Since both these major signaling pathways modulate inflammation, they may crosstalk to bring about coordinated inflammatory responses. The linkage between TLR signaling and Nrf2-Keap1 pathway may serve as a bridge between immune regulation and oxidative stress responses through regulation of inflammation. Also, inflammation is reportedly responsible for the plethora of diseased conditions; a study of its regulation by targeting the TLR-Nrf2 cross-talks may also be beneficial for the development of therapeutic therapies or prophylactic treatments. Hence, present review focuses on the crosstalk between TLR signaling and Nrf2 pathway with respect to their role in modulation of inflammation in normal as well as pathologic conditions.

Mannan oligosaccharide requires functional ETC and TLR for biological radiation protection to normal cells.

BACKGROUND: Low LET Ionizing radiation is known to alter intracellular redox balance by inducing free radical generation, which may cause oxidative modification of various cellular biomolecules. The extent of biomolecule-modifications/ damages and changes in vital processes (viz. cellular homeostasis, inter-/intra-cellular signaling, mitochondrial physiology/dynamics antioxidant defence systems) are crucial which in turn determine fate of cells. RESULTS: In the present study, we expended TLR expressing (normal/ transformed) and TLR null cells; and we have shown that mannan pretreatment in TLR expressing normal cells offers survival advantage against lethal doses of ionizing radiation. On the contrary, mannan pretreatment does not offer any protection against radiation to TLR null cells, NKE ρ° cells and transformed cells. In normal cells, abrupt decrease in mitochondrial membrane potential and endogenous ROS levels occurs following treatment with mannan. We intend to irradiate mannan-pretreated cells at a specific stage of perturbed mitochondrial functioning and ROS levels to comprehend if mannan pretreatment offers any survival advantage against radiation exposure to cells. Interestingly, pre-irradiation treatment of cells with mannan activates NFκB, p38 and JNK, alters mitochondrial physiology, increases expression of Cu/ZnSOD and MnSOD, minimizes oxidation of mitochondrial phospholipids and offers survival advantage in comparison to irradiated group, in TLR expressing normal cells. CONCLUSION: The study demonstrates that TLR and mitochondrial ETC functions are inevitable in radio-protective efficacy exhibited by mannan.

Phytochemical analysis and differential in vitro cytotoxicity assessment of root extracts of Inula racemosa.

The root of Inula racemosa is known for its antifungal, hypolipdemic and antimicrobial properties in traditional Indian Ayurvedic and Chinese system of medicine. The biological efficacy of Inula species is mainly due to the presence sesquiterpene lactone (Isoalantolactone and Alantolactone), which are reported to be inducers of Nrf2 antioxidant pathway. The investigation of properties and efficacy of root extracts of I. racemosa and their comparison was done with a view to find most efficacious extract for use at cellular level (both normal and transformed). In the present study different extracts of root of I. racemosa (aqueous, ethanolic, and 50% aqueous-ethanolic) were prepared and compared for their antioxidant potential, reducing capacity, polyphenol content and flavonoid content. Our investigations suggested that the aqueous extract possess highest antioxidant capacity and reducing potential. The polyphenol content was found to be highest in aqueous extract in comparison with other two extracts. However, all the three extracts showed less flavonoid content. Further, the preliminary phytochemical screening of all the extracts revealed the presence of terpenoids, phytosterols and glycosides. The TLC profile of ethanolic and 50% aqueous-ethanolic extracts showed the presence of alantolactone while aqueous extracts did not exhibit its strong presence. This warrants the need of more stringent techniques for characterization of aqueous extract in future. The in vitro cell based toxicity assays revealed that the aqueous extract was less toxic to kidneys cells while ethanolic extract was toxic to cells even at low concentrations. Hence, the current investigations showed better efficacy of the aqueous extract with respect to other extracts and found to be promising for its future application at in vitro levels.

Studies on radiation sensitization efficacy by silymarin in colon carcinoma cells.

Recent reports demonstrated the role of silymarin as a cytoprotective agent for normal cells against ionizing or non-ionizing (UV) radiation, and in inhibiting the chemically initiated or promoted carcinogenesis in several malignancies, such as skin or prostate cancers. Silymarin is a plant flavonoid obtained from milk thistle; the main active principles in milk thistle are silybin (silibinin), sylichrisitin and silydianin, commonly referred as silymarin. In the present study, we aimed to investigate the radiation modulatory effects of silymarin on cancer cells. For this, we used the HCT-15 and RKO colon cancer cell lines as a model. Pre-irradiation treatment of cells with silymarin (20 mg/ml) followed by radiation exposure inhibits colon cancer cell proliferation and enhances cell death in a time-dependent manner. We have also examined the changes in p53 phosphorylation at Ser15, phosphorylation of p38 and their association with DNA damage. Silymarin was found to reduce proliferation of the human colon carcinoma cells in a concentration and time-dependent manner. Moreover, percentage of cell death was also increased in combined treatment (20µg/ml of silymarin + radiation). Our studies indicate that the combination increases the arrest of cells in G2/M phase of cell cycle, DNA damage-induced decrease in mitochondrial membrane potential (MMP) and a decrease of the reactive oxygen species (ROS) levels, which are associated with an increase in cell death. Altogether, these results suggest that silymarin sensitizes colon cancer cells to radiation, strategy with potential for colon cancer treatment. Noteworthy, since silymarin was previously shown to confer protection against radiation in at least some types of normal tissues, additional studies are needed to further investigate the potential of silymarin in colon cancer therapy when combined with radiation, its potential protective effects on normal tissues and its mechanisms of action.

Biodistribution and scintigraphic evaluation of 99mTc-Mannan complex.

Technetium-99m (99mTc) is extensively used in nuclear medicine, mostly used to label radiopharmaceuticals and in radio diagnostics. In the present study, we directly radiolabeled mannan with 99mTc by using Tin(II) Chloride Dihydrate (SnCl2·2H2O) as a reducing agent. Mannan, a TLR agonist is a complex carbohydrate identified as a potential modulator of biological effects of ionizing radiation, both in vitro and in vivo, in our laboratory. Under in vivo conditions mannan modulates radiation response when administered through either oral or parenteral routes. The present study aims to understand the pharmacologic biodistribution of the 99mTc-mannan complex in mice (via oral, i.p. and i.v. routes) using non-invasive scintigraphic imaging and invasive radiometry. Qualitative and quantitative analysis of 99mTc-mannan complex was performed by ITLC-SG, ascending paper chromatography. Radio-complexation efficiency of >98% was consistently achieved with hydrolyzed reduced Tc-99m being 1-2%. We confirmed stability of complex in saline and serum up to 24 h at room temperature. Biodistribution studies were performed using the above radiocomplex in BALB/c mice and 99mTc-mannan complex was administered though oral, i.p. and i.v. routes. To our expectations, most of the radioactivity accumulated in stomach and small intestine in mice with oral administration, along with insignificant activity in the remaining studied organs. It suggests that 99mTc-mannan complex did not get absorbed from the gut and was removed as such in the fecal material. On the contrary, i.p. and i.v administration of mannan resulted in significant accumulation of the 99mTc-mannan complex in kidney, liver, intestine, lungs, spleen, bone marrow, blood and heart, at both 1 h and 4 h after i.v. administration. The remaining organs (stomach, testis and muscles) showed lower accumulation of the 99mTc-mannan complex. 99mTc-mannan complex was adminstered (i.v.) in New Zealand white rabbits and it was evident from the scintigraphic images that mannan cleared very rapidly from the administration site and reached into systemic circulation. No activity in the thyroid, salivary gland, or gastric mucosa suggests an insignificant amount of free pertechnetate in the 99mTc-complex preparation, further confirming the in vivo stability of the radiolabeled mannan complex. Significant amount of radioactivity in liver, intestine and kidneys suggests hepatobiliary as well as renal routes of clearance. The bio-availability of the complex varies with the route of administration. An entirely different biodistribution pattern exists when the same molecule is administered through oral or parenteral route. Our study is the first step towards a better understanding of the mechanisms involved in radiation modulation offered by mannan administration, in vivo.

Chemical composition and biological activities of trans-Himalayan alga Spirogyra porticalis (Muell.) Cleve.

The freshwater alga Spirogyra porticalis (Muell.) Cleve, a filamentous charophyte, collected from the Indian trans-Himalayan cold desert, was identified on the basis of morpho-anatomical characters. Extracts of this alga were made using solvents of varying polarity viz. n-hexane, acetonitrile, methanol and water. The antioxidant capacities and phenolic profile of the extracts were estimated. The methanol extract showing highest antioxidant capacity and rich phenolic attributes was further investigated and phytochemical profiling was conducted by gas chromatography-mass spectrometry (GC/MS) hyphenated technique. The cytotoxic activity of methanol extract was evaluated on human hepatocellular carcinoma HepG2 and colon carcinoma RKO cell lines. The anti-hypoxic effect of methanol extract of the alga was tested on in vivo animal system to confirm its potential to ameliorate oxidative stress. The antioxidant assays viz. ferric reducing antioxidant power (FRAP), 2,2'-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS), 1,1-diphenyl-2-picrylhydrazyl (DPPH) and nitric oxide (NO) radical scavenging capacities, ß-carotene-linoleic acid bleaching property and lipid peroxidation exhibited analogous results, wherein the algal extracts showed significantly high antioxidant potential. The extracts were also found to possess high content of total proanthocyanidin, flavonoid and polyphenol. GC/MS analysis revealed the presence of thirteen chemotypes in the methanol extract representing different phytochemical groups like fatty acid esters, sterols, unsaturated alcohols, alkynes etc. with substantial phyto-pharmaceutical importance. The methanol extract was observed to possess anticancer activity as revealed from studies on HepG2 and RKO cell lines. In the present study, S. porticalis methanol extract also provided protection from hypoxia-induced oxidative stress and accelerated the onset of adaptative changes in rats during exposure to hypobaric hypoxia. The bioactive phytochemicals present in this trans-Himalayan alga are of enormous interest and can be utilized sustainably for discovery of novel drugs against oxidative stress.

Development of Novel Anti-Cd20 Monoclonal Antibodies and Modulation in Cd20 Levels on Cell Surface: Looking to Improve Immunotherapy Response.

Rituximab has been revolutionized and validated CD20 targeting monoclonal antibody. Although, it is widely used for lymphoma therapy and many patients have been benefited. However significant numbers of patients are refractory or developed resistance to current therapies due to low level of CD20 expression and/or availability on cells surface. Thus development of novel anti-CD20 mAbs with great cell killing ability and enhance CD20 levels on cell surface can potentially exploit lymphoma therapy. In this scenario, we are summarizing the recently developed mAbs against CD20 and compounds that have ability to induce CD20 expression at significant level. We also are providing information regarding combination strategy for use of radiation and anti-CD20 mAbs in vitro. However, it will need to be determined by rigorous at pre-clinical and clinic testing. We hope this review will be beneficial for current research in the area of immunotherapy or radio-immunotherapy.

NF-kB as a key player in regulation of cellular radiation responses and identification of radiation countermeasures.

Nuclear factor (NF)-κB is a transcription factor that plays significant role in immunity, cellular survival and inhibition of apoptosis, through the induction of genetic networks. Depending on the stimulus and the cell type, the members of NF-κB related family (RelA, c-Rel, RelB, p50, and p52), forms different combinations of homo and hetero-dimers. The activated complexes (Es) translocate into the nucleus and bind to the 10bp κB site of promoter region of target genes in stimulus specific manner. In response to radiation, NF-κB is known to reduce cell death by promoting the expression of anti-apoptotic proteins and activation of cellular antioxidant defense system. Constitutive activation of NF-κB associated genes in tumour cells are known to enhance radiation resistance, whereas deletion in mice results in hypersensitivity to IR-induced GI damage. NF-κB is also known to regulate the production of a wide variety of cytokines and chemokines, which contribute in enhancing cell proliferation and tissue regeneration in various organs, such as the GI crypts stem cells, bone marrow etc., following exposure to IR. Several other cytokines are also known to exert potent pro-inflammatory effects that may contribute to the increase of tissue damage following exposure to ionizing radiation. Till date there are a series of molecules or group of compounds that have been evaluated for their radio-protective potential, and very few have reached clinical trials. The failure or less success of identified agents in humans could be due to their reduced radiation protection efficacy. In this review we have considered activation of NF-κB as a potential marker in screening of radiation countermeasure agents (RCAs) and cellular radiation responses. Moreover, we have also focused on associated mechanisms of activation of NF-κB signaling and their specified family member activation with respect to stimuli. Furthermore, we have categorized their regulated gene expressions and their function in radiation response or modulation. In addition, we have discussed some recently developed radiation countermeasures in relation to NF-κB activation.

Surface levels of CD20 determine anti-CD20 antibodies mediated cell death in vitro.

BACKGROUND: The sensitivity of human Burkitt's lymphoma cells to rituximab (Rtx) and tositumomab (Tst) was assessed on cells expressing different levels of CD20 on surface. Cells that harbor low CD20 levels may resists against therapeutics response to CD20-specific antibodies. We postulated that, radiation-induced modulation of CD20 surface levels may play a crucial and central role in determining the relative efficacy of rituximab and tositumomab in treating Burkitt's lymphoma disease. Here, we examined the γ-radiation-induced CD20 expression in the Burkitt lymphoma cell line 'Daudi' and the relation of differential levels of CD20 with anti-CD20 mAbs mediated cell death. METHODOLOGY: In this study we examined kinetics of CD20 expression following sub lethal doses ofγ-radiation to Daudi cells and thereafter anti-CD20 mAbs (rituximab and tositumomab) were added in cell suspensions. The correlation of kinetics of CD20 expression and cells treated with anti-CD20 mAbs/or corresponding isotype Abs with special reference to changes in mitochondrial membrane potential and reactive oxygen species generation was also examined. Further, we also investigated the efficacy of anti-CD20 mAbs and possible induction of cell death in relation to levels of CD20 cell surface expression. CONCLUSION: This report provides evidence that CD20 expression can be induced by exposure of cells to γ-radiation. In addition, these findings demonstrated that the efficacy of anti-CD20 mAbs is dependent on the surface levels of CD20. Based on these findings, we hypothesized (i) irradiation just prior to immunotherapy may provide new treatment options even in aggressive B cell tumors, which are resistant to current therapies in vivo (ii) The efficacy of induction of apoptosis varies with type of monoclonal antibodies in vitro.

Antioxidant capacities, phenolic profile and cytotoxic effects of saxicolous lichens from trans-Himalayan cold desert of Ladakh.

Fourteen saxicolous lichens from trans-Himalayan Ladakh region were identified by morpho-anatomical and chemical characteristics. The n-hexane, methanol and water extracts of the lichens were evaluated for their antioxidant capacities. The lichen extracts showing high antioxidant capacities and rich phenolic content were further investigated to determine their cytotoxic activity on human HepG2 and RKO carcinoma cell lines. The ferric reducing antioxidant power (FRAP), 2,2'-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt (ABTS), 1,1-diphenyl-2-picrylhydrazyl (DPPH) and nitric oxide (NO) radical scavenging capacities and ß-carotene-linoleic acid bleaching property exhibited analogous results where the lichen extracts showed high antioxidant action. The lichen extracts were also found to possess good amount of total proanthocyanidin, flavonoid and polyphenol. The methanolic extract of Lobothallia alphoplaca exhibited highest FRAP value. Methanolic extract of Xanthoparmelia stenophylla showed the highest ABTS radical scavenging capacity. The n-hexane extract of Rhizoplaca chrysoleuca exhibited highest DPPH radical scavenging capacity. Highest antioxidant capacity in terms of ß-carotene linoleic acid bleaching property was observed in the water extract of Xanthoria elegans. Similarly, Melanelia disjuncta water extract showed highest NO scavenging capacity. Among n-hexane, methanol and water extracts of all lichens, the methanolic extract of Xanthoparmelia mexicana showed highest total proanthocyanidin, flavonoid and polyphenol content. From cytotoxic assay, it was observed that the methanolic extracts of L. alphoplaca and M. disjuncta were exhibiting high cytotoxic effects against cancer cell growth. Similarly, the water extract of Dermatocarpon vellereum, Umbilicaria vellea, X. elegans and M. disjuncta and the methanolic extract of M. disjuncta and X. stenophylla were found to possess high antioxidant capacities and were non-toxic and may be used as natural antioxidants for stress related problems. Our studies go on to prove that the unique trans-Himalayan lichens are a hitherto untapped bioresource with immense potential for discovery of new chemical entities, and this biodiversity needs to be tapped sustainably.

In vitro evaluation of antioxidant and radioprotective properties of a novel extremophile from mud volcano: implications for management of radiation emergencies.

A thermophilic bacterium, designated as RH 127, was isolated from mud volcano (Baratang Islands) of Andaman region, India (12°07'N 92°47'E/12.117°N 92.783°E) for the first time. Biochemical tests and 16S rRNA gene sequencing indicate that it belongs to the genus Geobacillus. The strain showed 98% confirmed 16S rRNA gene sequence homology with Geobacillus toebii. The bacteria was extracted in various solvent systems and three different fractions prepared. In the present study, antioxidant and radioprotective activity of extracts (INM-7860, INM-7861, and INM-7862) of bacterium G. toebii (strain RH 127) were evaluated. The fractions were evaluated for their introspective comparison of the relative antioxidant efficiency. The antioxidative activities, DPPH radical scavenging effects, hydroxyl radical scavenging effects, membrane protection, antihemolytic activity, and linoleic acid degradation efficacies were assayed. INM-7861 and INM-7862 activated NF-κB expression, as evidenced by reporter assay studies, and thereby contributed to overall radioprotective effect. INM-7862 exhibited best results. This study explicitly shows that the extracts of G. toebii have immense potential as a radiation countermeasure agent.

Radical scavenging and radiomodulatory effects of Psoralea corylifolia Linn. substantiated by in vitro assays and EPR spectroscopy.

The present study is the first report of the radiomodulatory effects of Psoralea corylifolia Linn. The extract (IBG-RA-26) prepared from P. corylifolia was chemically analysed by HPLC, LC-MS/MS and NMR. The total polyphenolic content of IBG-RA-26 was 0.287 mg/ml of quercetin equivalents. IBG-RA-26 exhibited a dose-dependent increase in 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity. It exhibited comparable (> 50%) site-specific and non-site-specific hydroxyl radical scavenging activity in higher concentration ranges (500-1000 microg/ml), while at lower concentrations (5-50 microg/ml) it exhibited significantly (p < 0.05) higher non-site-specific scavenging ability compared to site-specific activity. Nitric oxide scavenging activity of IBG-RA-26 (5-1000 microg/ml) increased in a concentration-dependent manner, while maximum superoxide ion scavenging ability (79%) was observed at 50 microg/ml. The electron donation potential of IBG-RA-26 was found to be higher than that of ascorbic acid at lower concentrations (up to 5 microg/ml). Analysis of the ability of IBG-RA-26 to protect membranes against gamma-radiation, utilizing an artificial membrane system (liposome), revealed a significant (p < 0.05) decrease in the formation of malondialdehyde (MDA) as a function of the concentration of IBG-RA-26. Radiation-induced lysis of human erythrocytes was monitored and efficacy of IBG-RA-26 was tested in the concentration range 25-1000 microg/ml, with significant protective efficacy observed in the range 25-50 microg/ml. IBG-RA-26 rendered significant (p < 0.05) protection against radiation (0.25 kGy)-induced DNA damage. EPR spectroscopy was used to investigate the DPPH radical scavenging capacity of IBG-RA-26. IBG-RA-26 exhibited a good DPPH radical scavenging capacity in a concentration-dependent manner. By direct EPR spectroscopy we have also demonstrated the possible formation of free radical species in a solution of IBG-RA-26. The wide spectrum of radioprotective and antioxidant properties exhibited by IBG-RA-26 indicate that P. corylifolia has potential as a radiomodulatory agent.

Evaluation of silymarin as a promising radioprotector.

Silymarin, a purified extract of seeds of Silybum marianum L. and well known for its hepatoprotective abilities, has been evaluated for inherent utility as a radioprotective agent. A fraction (INM-7035) was authenticated by characterizing the percentage composition of silybin A and B (39.9% and 57.4%). Free radical scavenging activities of INM-7035 against superoxide radicals (>68%), hydroxyl radicals (>33.75%), DPPH (67.2%), and ABTS (32.4%) were also evaluated. The fraction chelated (>30%) ferrous ions, thereby able to restrict amplification. INM-7035 exhibited >50% peroxyl radical scavenging activity in the lipid phase along with dose-dependent (R2 = 0.990) reducing power in the aqueous phase. Radiation-induced free radical flux can lead to disruption of biomolecules like membrane lipids. INM-7035 completely inhibited lipid peroxidative stress in case of membranes against supralethal radiation stress in the liposomal system. The ability of INM-7035 to modulate the levels of NF-kappaB, indicated its inherent potential as a radioprotective bioactive constituent.

Podophyllum hexandrum as a potential botanical supplement for the medical management of nuclear and radiological emergencies (NREs) and free radical-mediated ailments: leads from in vitro/in vivo radioprotective efficacy evaluation.

Management of radiation-induced reactive oxygen/nitrogen species requires a holistic approach to mitigate the deleterious effects of free radicals. Flora of the Himalayas, which prevails under extreme climatic conditions, has been explored for its potential utility to develop radioprotective drugs. The Himalayan high altitude medicinal plant, Podophyllum hexandrum Royle, was selected on the basis of its unique properties, and a novel fractionated nonpolar extract (REC-2003) was prepared and evaluated for radioprotective efficacy, in vitro as well as in vivo. The free radical scavenging activity of REC-2003 was found to be > 75% (20 µg/ml) with maximum superoxide scavenging activity (57.56 ± 1.38%) recorded at 1 mg/ml concentration (tetrazolium-based estimation). More than 30% inhibition of nitric oxide radicals was observed at concentrations > 0.5 mg/ml, while hydroxyl radical scavenging activity (deoxy-D-ribose assay) exhibited a dose-dependent (100-600 µg/ml) increase. Significantly high (90%) protection to human erythrocytes was observed at 75 µg/ml, which was found to be the most optimized dose. Similarly, more than 90% inhibition was observed against lipid peroxidation (evaluated by estimating levels of malondialdehyde). The significant antihemolytic potential of REC-2003 could be attributed to its ability to scavenge free radicals, reduce peroxidative stress on lipid membranes, and render protection to DNA (evaluated using plasmid relaxation assay). All these activities holistically contributed toward the radioprotective ability. REC-2003 (8 mg/kg BW; intraperitoneal (i.p.), -30 min) rendered > 80% total-body protection in Swiss Albino Strain 'A' mice [against lethal radiation (10 Gy)] in a 30-day survival assay. Phytochemical characterization of the constituents of REC-2003 revealed the presence of polyphenolics (flavonoids). The characterized constituents also included the aryl-tetralin lignans like podophyllotoxin, its glycoside, 4'-demethyl derivative, and epi-podophyllotoxin. The optimized requisite single dose (8 mg/KgBW; i.p., -30 min) for obtaining significant radioprotection is reasonably low and establishes its future utility as a dietary supplement in the medical management of free radical-mediated diseases and specifically for rescue missions during nuclear and radiological emergencies (NREs).

An agonist of toll-like receptor 5 has radioprotective activity in mouse and primate models.

The toxicity of ionizing radiation is associated with massive apoptosis in radiosensitive organs. Here, we investigate whether a drug that activates a signaling mechanism used by tumor cells to suppress apoptosis can protect healthy cells from the harmful effects of radiation. We studied CBLB502, a polypeptide drug derived from Salmonella flagellin that binds to Toll-like receptor 5 (TLR5) and activates nuclear factor-kappaB signaling. A single injection of CBLB502 before lethal total-body irradiation protected mice from both gastrointestinal and hematopoietic acute radiation syndromes and resulted in improved survival. CBLB502 injected after irradiation also enhanced survival, but at lower radiation doses. It is noteworthy that the drug did not decrease tumor radiosensitivity in mouse models. CBLB502 also showed radioprotective activity in lethally irradiated rhesus monkeys. Thus, TLR5 agonists could potentially improve the therapeutic index of cancer radiotherapy and serve as biological protectants in radiation emergencies.

Regulation of CD20 expression by radiation-induced changes in intracellular redox status.

Increasing the levels of CD20 expression in cells that harbor low CD20 levels may enhance their responsiveness to CD20-specific antibody therapies. Here, we examined the regulation of CD20 expression after treatment with 0.5-2.0 Gy X-irradiation and hydrogen peroxide (H(2)O(2)), in the presence or absence of known antioxidants, in the Burkitt lymphoma cell lines Daudi and Raji. Irradiation of cells enhanced cell-surface CD20 expression; the kinetics and extent of this change were cell-type specific and time-dependent. The kinetics of reactive oxygen species generation and changes in mitochondrial membrane potential after irradiation were also correlated with changes in CD20 expression. Raji and Daudi cells treated with H(2)O(2) showed a 2-to 2.5-fold increase in CD20 expression at 12 and 20 h, respectively. Buthionine sulfoximine, which depletes glutathione, also increased surface CD20, whereas antioxidants, such as PEG-catalase, PEG-SOD, vitamin C, and amifostine, decreased CD20 expression induced by radiation or H(2)O(2). The antioxidant-mediated decrease in CD20 expression induced by radiation or H(2)O(2) suggests a mechanism involving redox regulation. These results demonstrate the critical role of radiation-induced oxidative stress in CD20 expression and may have implications for defining and improving the efficacy of CD20-targeted antibody therapy and radioimmunotherapy.

E2F4 function in G2: maintaining G2-arrest to prevent mitotic entry with damaged DNA.

Mammalian cells undergo cell cycle arrest in response to DNA damage through multiple checkpoint mechanisms. One such checkpoint pathway maintains genomic integrity by delaying mitotic progression in response to genotoxic stress. Transition though the G2 phase and entry into mitosis is considered to be regulated primarily by cyclin B1 and its associated catalytically active partner Cdk1. While not necessary for its initiation, the p130 and Rb-dependent target genes have emerged as being important for stable maintenance of a G2 arrest. It was recently demonstrated that by interacting with p130, E2F4 is present in the nuclei and plays a key role in the maintenance of this stable G2 arrest. Increased E2F4 levels and its translocation to the nucleus following genotoxic stress result in downregulation of many mitotic genes and as a result promote a G0-like state. Irradiation of E2F4-depleted cells leads to enhanced cellular DNA double-strand breaks that may be measured by comet assays. It also results in cell death that is characterized by caspase activation, sub-G1 and sub-G2 DNA content, and decreased clonogenic cell survival. Here we review these recent findings and discuss the mechanisms of G2 phase checkpoint activation and maintenance with a particular focus on E2F4.

Radioprotection by plant products: present status and future prospects.

The development of radioprotective agents has been the subject of intense research in view of their potential for use within a radiation environment, such as space exploration, radiotherapy and even nuclear war. However, no ideal, safe synthetic radioprotectors are available to date, so the search for alternative sources, including plants, has been on going for several decades. In Ayurveda, the traditional Indian system of medicine, several plants have been used to treat free radical-mediated ailments and, therefore, it is logical to expect that such plants may also render some protection against radiation damage. A systematic screening approach can provide leads to identifying potential new candidate drugs from plant sources, for mitigation of radiation injury. This article reviews some of the most promising plants, and their bioactive principles, that are widely used in traditional systems of medicine, and which have rendered significant radioprotection in both in vitro and in vivo model systems. Plants and their constituents with pharmacological activities that may be relevant to amelioration of radiation-mediated damage, including antiemetic, antiinflammatory, antioxidant, cell proliferative, wound healing and haemopoietic stimulatories are also discussed.