Radiation Protection. Anti-Radiation Vaccine. Isolation

of Radiation Toxins from Irradiated Plants .

Dmitri Popov MD (Russia), PhD (Russia-Canada)Advanced Medical Technology and Systems Inc.

intervaccine@gmail.comProf. Jeffrey Jones, Baylor Medical School, USA.

Radiation Effects. Toxicity of Plants after irradiation.• Research Proposal:

Radiation Protection. Anti-Radiation Vaccine. Isolation of Radiation Toxins from Irradiated Plants .• Dmitri Popov• Full-text · Research Proposal · Jan 2016

• File name: RadiationProtectionAntiRadiationVaccine.pptxDOI: 10.13140/RG.2.1.3094.4403

Radiation Effects. Toxicity of Plants after irradiation.

Keywords: Radiation; Apoptosis; Necrosis; programmed cell death; intrinsic/extrinsic pathway; granzyme A/B; perforin.

Radiation Protection. Technology of Anti Radiation vaccine.• Programmed cell death (PCD) occurs in animals and plants under

various stresses include different types of radiation.• The ability to modulate the life and prevent death of an irradiated

cells is recognized as a great therapeutic potential.• Apoptosis also occurs as a defense mechanism such as in immune

reactions or when cells are damaged by disease or physical or chemical agents.

Radiation Protection. Technology of Anti Radiation vaccine.• Irradiation with any type of radiation or viruses, group of drugs used

for cancer chemotherapy results in DNA damage in some cells, which can lead to apoptotic death through a p53-dependent pathway.

Radiation Protection. Technology of Anti Radiation vaccine.After irradiation is also the issue of distinguishing apoptosis from necrosis, two processes that can occur independently, sequentially, as well as simultaneously (Hirsch, 1997; Zeiss, 2003).Can immunological mechanisms induce apoptosis or necrosis?Yes. Can inhibition of pathological, immunological mechanisms prevent apoptosis or even necrosis?Inhibition of pathological, immunological could prevent massive apoptosis. Inhibition of pathological, immunological could prevent massive necrosis.Possible yes.

Radiation Protection. Technology of Anti Radiation vaccine.• At low doses, a variety of injurious stimuli such as heat, radiation,

hypoxia and cytotoxic anticancer drugs can induce apoptosis but these same stimuli can result in necrosis at higher doses. • Toxicologic Pathology, 35:495–516, 2007 Copyright C by the Society of Toxicologic

Pathology ISSN: 0192-6233 print / 1533-1601 online DOI: 10.1080/01926230701320337

• Apoptosis: A Review of Programmed Cell Death SUSAN ELMORE NIEHS, Laboratory of Experimental Pathology, Research Triangle Park, North Carolina 27709, USA

Radiation Protection. Technology of Anti Radiation vaccine.• Although the mechanisms and morphologies of apoptosis and

necrosis differ, there is overlap between these two processes. Evidence indicates that necrosis and apoptosis represent morphologic expressions of a shared biochemical network described as the “apoptosis-necrosis continuum” (Zeiss, 2003).

Radiation Protection. Technology of Anti Radiation vaccine.• Whether a cell dies by necrosis or apoptosis depends in part on the

nature of the cell death signal, the tissue type, the developmental stage of the tissue and the physiologic milieu (Fiers et al., 1999; Zeiss, 2003).• Interesting.

Radiation Protection. Technology of Anti Radiation vaccine.• Necrosis is an uncontrolled and passive process that usually affects

large fields of cells whereas apoptosis is controlled and energy-dependent and can affect individual or clusters of cells. Necrotic cell injury is mediated by two main mechanisms; interference with the energy supply of the cell and direct damage to cell membranes.• Apoptosis: A Review of Programmed Cell Death SUSAN ELMORE NIEHS,

Laboratory of Experimental Pathology, Research Triangle Park, North Carolina 27709, USA• http://tpx.sagepub.com/content/35/4/495.full.pdf+html

Radiation Protection. Technology of Anti Radiation vaccine.• Some of the major morphological changes that occur with necrosis

include cell swelling; formation of cytoplasmic vacuoles; distended endoplasmic reticulum; formation of cytoplasmic blebs; condensed, swollen or ruptured mitochondria; disaggregation and detachment of ribosomes; disrupted organelle membranes; swollen and ruptured lysosomes; and eventually disruption of the cell membrane (Kerr et al., 1972; Majno and Joris, 1995; Trump et al., 1997)• http://tpx.sagepub.com/content/35/4/495.full.pdf+html

Radiation Protection. Technology of Anti Radiation vaccine.• This loss of cell membrane integrity results in the release of the

cytoplasmic contents into the surrounding tissue, sending chemotatic signals with eventual recruitment of inflammatory cells. Because apoptotic cells do not release their cellular constituents into the surrounding interstitial tissue and are quickly phagocytosed by macrophages or adjacent normal cells, there is essentially no inflammatory reaction (Savill and Fadok, 2000; Kurosaka et al., 2003).• http://tpx.sagepub.com/content/35/4/495.full.pdf+html

Radiation Protection. Technology of Anti Radiation vaccine.• Until recently, apoptosis has traditionally been considered an

irreversible process with caspase activation committing a cell to death and the engulfment genes serving the purpose of dead cell removal. However, the uptake and clearance of apoptotic cells by macrophages may involve more than just the removal of cell debris. Hoeppner et al. have shown that blocking engulfment genes in C. elegans embryos enhances cell survival when cells are subjected to weak pro-apoptotic signals (Hoeppner et al., 2001).• http://tpx.sagepub.com/content/35/4/495.full.pdf+html

Radiation Protection. Technology of Anti Radiation vaccine.• Some research suggests that DNA repair is activated early in the p53-

induced apoptotic process and that this DNA repair may be involved in reversing the cell death pathway in some circumstances.• http://tpx.sagepub.com/content/35/4/495.full.pdf+html

Radiation Protection. Technology of Anti Radiation vaccine.• The mechanisms of apoptosis are highly complex and sophisticated,

involving an energy-dependent cascade of molecular events . To date, research indicates that there are two main apoptotic pathways: the extrinsic or death receptor pathway and the intrinsic or mitochondrial pathway. However, there is now evidence that the two pathways are linked and that molecules in one pathway can influence the other (Igney and Krammer, 2002).• http://tpx.sagepub.com/content/35/4/495.full.pdf+html•

Radiation Protection. Technology of Anti Radiation vaccine.• There is an additional pathway that involves T-cell mediated cytotoxicity

and perforin-granzyme-dependent killing of the cell. The perforin/granzyme pathway can induce apoptosis via either granzyme B or granzyme A. The extrinsic, intrinsic, and granzyme B pathways converge on the same terminal, or execution pathway. This pathway is initiated by the cleavage of caspase-3 and results in DNA fragmentation, degradation of cytoskeletal and nuclear proteins, cross-linking of proteins, formation of apoptotic bodies, expression of ligands for phagocytic cell receptors and finally uptake by phagocytic cells. The granzyme A pathway activates a parallel, caspase-independent cell death pathway via single stranded DNA damage (Martinvalet et al., 2005).

Radiation Protection. Technology of Anti Radiation vaccine.• Apoptotic cells exhibit several biochemical modifications such as

protein cleavage, protein cross-linking, DNA breakdown, and phagocytic recognition that together result in the distinctive structural pathology described previously (Hengartner, 2000).

Radiation Protection. Technology of Anti Radiation vaccine.• Caspases are widely expressed in an inactive proenzyme form in most

cells of plants and mammals, and once activated can often activate other pro-caspases, allowing initiation of a protease cascade. • This proteolytic cascade, in which one caspase can activate other

caspases, amplifies the apoptotic signaling pathway and thus leads to rapid cell death.• http://tpx.sagepub.com/content/35/4/495.full.pdf+html

Radiation Effects. Toxicity of Plants after irradiation.• Programmed cell death (PCD) occurs in animals and plants under

various stresses and during development. Recently, vacuolar processing enzyme (VPE) was identified as an executioner of plant PCD.• “A cellular suicide strategy of plants: vacuole-mediated cell death”• Apoptosis 2006; 11: 905–911 C 2006 Springer Science + Business

Media, LLC. Manufactured in The United States. DOI: 10.1007/s10495-006-6601-1. N. Hatsugai et al.

Radiation Effects. Toxicity of Plants after irradiation.• Recently, vacuolar processing enzyme (VPE) was identified as an

executioner of plant PCD. VPE is a cysteine protease that cleaves a peptide bond at the C-terminal side of asparagine and aspartic acid.• “A cellular suicide strategy of plants: vacuole-mediated cell death”• Apoptosis 2006; 11: 905–911 C 2006 Springer Science + Business

Media, LLC. Manufactured in The United States. DOI: 10.1007/s10495-006-6601-1. N. Hatsugai et al.

Radiation Effects. Toxicity of Plants after irradiation.• Programmed cell death (PCD) is an active, genetically controlled• process leading to selective elimination of unwanted or damaged cells

in eukaryotes. PCD is essential for growth and development of multicellular organisms as well as for proper response to environment (Gechev et al., 2006; Lam, 2004).

Radiation Effects. Toxicity of Plants after irradiation.• Plant PCD is associated with a number of developmental processes

including embryo formation, degeneration of the aleurone layer during monocot seed germination, differentiation of tracheary elements in water-conducting xylem tissues, formation of root aerenchyma and epidermal trichomes, anther tapetum degeneration, floral organ abscission, pollen self-incompatibility, remodeling of some types of leaf shape, and leaf senescence. (Gechev et al., 2006; Thomas and Franklin-Tong, 2004).• Programmed Cell Death in Plants: New Insights into Redox Regulation• and the Role of Hydrogen Peroxide• Ilya Gadjev,1,* Julie M. Stone,† and Tsanko S. Gechev*

Radiation Effects. Toxicity of Plants after irradiation.• Hydrogen peroxide (H2O2) and other reactive oxygen species (ROS)

have become recognized to be key modulators of PCD of as well as many other biological processes such as growth, development, and stress adaptation (Gechev et al., 2006).

Radiation Effects. Toxicity of Plants after irradiation.• Although specific ROS receptors/sensors remain largely elusive,

downstream components of H2O2 and ROS signal transduction networks controlling plant PCD have been identified, including protein kinases, protein phosphatases, and transcription factors. The majority of these are restricted to plants, with only a few genes having close homologues in animals. (Gechev et al., 2006).

Radiation Effects. Toxicity of Plants after irradiation.• Hydrogen peroxide is produced in all cellular compartments as a

result of reactions of energy transfer, electron leakage from saturated electron transport chains, and the activities of various oxidases and peroxidases (Apel and Hirt, 2004).

Radiation Effects. Toxicity of Plants after irradiation.• Programmed cell death (PCD) is a process by which cells in many

organisms die. The basic morphological and biochemical features of PCD are conserved between the animal and plant kingdoms. Cysteine proteases have emerged as key enzymes in the regulation of animal PCD.

Radiation Effects. Toxicity of Plants after irradiation.• The discovery that cell death is a tightly regulated (programmed) process

has stirred a great deal of interest in its mechanisms. Studies of animal systems have shown that the execution of programmed cell death (PCD) or apoptosis is controlled by a multistep signaling pathway (McConkey and Orrenius, 1994; Stewart, 1994).• In plants, PCD has been implicated in xylogenesis (Fukuda, 1996; Groover et al., 1997), in some forms of senescence, and in the hypersensitive response to pathogens and environmental stresses (Greenberg, 1996; Mittler and Lam, 1996; Lamb and Dixon, 1997).

Radiation Effects. Toxicity of Plants after irradiation.• Although a detailed understanding of how plant cells die is still largely

unknown, recent studies have shown that the apoptotic pathways of the animal and plant kingdoms are morphologically and biochemically similar (Greenberg, 1996; Levine et al., 1996; Wang et al., 1996).

Radiation Effects. Toxicity of Plants after irradiation.• Specifically, the morphological hallmarks of apoptosis include

cytoplasmic shrinkage, nuclear condensation, and membrane blebbing (Earnshaw, 1995; Martins and Earnshaw, 1997); the biochemical events involve calcium influx, exposure of phosphatidylserine and activation of specific proteases and DNA

fragmentation, first to large 50-kb fragments and then to nucleosomal ladders (McConkey and Orrenius, 1994; Stewart, 1994; Wang et al., 1996; O’Brien et al., 1998).

Radiation Effects. Toxicity of Plants after irradiation.• The Involvement of Cysteine Proteases and Protease Inhibitor Genes in the

Regulation of Programmed Cell Death in Plants. • The Plant Cell, Vol. 11, 431–443, March 1999, www.plantcell.org © 1999

American Society of Plant Physiologists.

Radiation Effects. Toxicity of Plants after irradiation.• VPE processing system mediates a cellular suicide strategy in plants. In animals, dying cells are

packaged into apoptotic bodies• and then engulfed by phagocytes. In contrast, because plants do not have phagocytes and the

cells are surrounded by rigid cell walls, plant• cells must degrade their materials by themselves. VPE, which has caspase-1-like activity, is

accumulated after perception of death signals such• as pathogen infection. VPE is involved in activation of the target proteins to provoke

disintegration of the vacuolar membranes. Consequently,• the vacuolar hydrolytic enzymes leave the vacuole for the cytosol and degrade cellular

components. Plants have evolved a death strategy that• is mediated by the VPE processing system, which is not seen in animals. A cellular

suicide strategy of plants: vacuole-mediated cell death. N. Hatsugai et al. DOI: 10.1007/s10495-006-6601-1

Radiation Effects. Toxicity of Plants after irradiation.• The genome of an organism is under constant attack from endogenous and exogenous

DNA damaging factors, such as reactive radicals, radiation, and genotoxins. Therefore, DNA damage response systems to sense DNA damage, arrest cell cycle, repair DNA lesions, and/or induce programmed cell death are crucial for maintenance of genomic integrity and survival of the organism. Genome sequences revealed that, although plants possess many of the DNA damage response factors that are present in the animal systems, they are missing some of the important regulators, such as the p53 tumor suppressor. These observations suggest differences in the DNA damage response mechanisms between plants and animals. In this review the DNA damage responses in plants and animals are compared and contrasted. In addition, the function of SUPPRESSOR OF GAMMA RESPONSE 1 (SOG1), a plant-specific transcription factor that governs the robust response to DNA damage, is discussed. Biology 2013, 2, 1338-1356; doi:10.3390/biology2041338

Radiation Effects. Toxicity of Plants after irradiation.• Can irradiated plant’s cells used for feeding induce radiation disease

of mammals?• Yes.

Radiation Effects. Toxicity of Plants after irradiation.• A careful analysis by FDA of all Army data present (including 31 loose-

leaf notebooks of animal feeding test results) showed significant adverse effects produced in animals fed irradiated food...• http://www.mercola.com/article/irradiated/irradiated_research.htm• In the course of legalizing the irradiation of beef, chicken, pork, fruit,

vegetables, eggs, juice, spices and sprouting seeds -- a process that has spanned nearly 20 years -- the U.S. Food and Drug Administration has dismissed or ignored a substantial body of evidence suggesting that irradiated food may not be safe for human consumption.• http://www.mercola.com/article/irradiated/irradiated_research.htm

Radiation Effects. Toxicity of Plants after irradiation.• What were these adverse effects?• A decrease of 20.7 percent in surviving weaned rats.• A 32.3 percent decrease in surviving progeny of dogs.• Dogs weighing 11.3 percent less than animals on the control diets...

Carcinomas of the pituitary gland, a particularly disturbing finding since this is an extremely rare type of malignant tumor."• Food irradiation: An FDA report. FDA Papers, Oct. 1968.

Radiation Effects. Toxicity of Plants after irradiation.• Fatal Internal Bleeding in Rats (I)• "A significant number of rats consuming irradiated beef died from

internal hemorrhage within 46 days, the first death of a male rat coming on the 11th day of feeding. This rat became sluggish on the 8th day of the regimen and started refusing food. He continued to be morbid during the next two days, did not eat any food, lost weight and appeared anemic. He was found dead on the 11th day.• Vitamin K deficiency in rats induced by feeding of irradiated beef.• Journal of Nutrition, 69:18-21, 1959. (Cosponsored by the Surgeon

General of the US Army)

Radiation Effects. Toxicity of Plants after irradiation.• Fatal Internal Bleeding in Rats (II)• "Hemorrhagic death had occurred in all males fed irradiated diets by

day 34... There is evidence to suggest that inefficient absorption of vitamins, i.e. vitamin K, from the intestinal tract may contribute to a deficiency state." [Note: Vitamin K plays a major role in blood clotting.]• Influence of age, sex, strain of rat and fat soluble vitamins on

hemorrhagic syndromes in rats fed irradiated beef.• Federation Proceedings, 19:1045-1048, 1960. (Cosponsored by the

Surgeon General of the US Army)

Radiation Effects. Toxicity of Plants after irradiation.• Fetal Deaths in Mice• "Freshly irradiated diets produced elevated levels of early deaths in

[mice fetuses]... The increase in early deaths would suggest that the diet when irradiated has some mutagenic potential."• Irradiated laboratory animal diets: Dominant lethal studies in the

mouse.• Mutation Research, 80:333-345, 1981.• http://www.mercola.com/article/irradiated/irradiated_research.htm

Radiation Effects. Toxicity of Plants after irradiation.• Toxic effects of irradiated foods. Nature, 211:302, 1966.• A Thalidomide Warning (II)• "Irradiating can bring about chemical transformations in food and food components

resulting in the formation of potential mutagens, particularly hydrogen peroxide and various organic peroxides.• It is now realized, especially since the thalidomide episode, that older testing protocols

do not detect the more subtle population hazards such as mutagens and teratogens. In view of the serious consequences to the human population which could arise from a high level of induced mutations, it is desirable that protocols for irradiated food should include in vivo tests on mammals for possible mutagenicity."• Mutagenicity and cytotoxicity of irradiated foods and food components.• http://www.mercola.com/article/irradiated/irradiated_research.htm

Radiation Effects. Toxicity of Plants after irradiation.• Bulletin of the World Health Organization, 41:873-904, 1969. (Cosponsored by

the US Atomic Energy Commission and Food and Drug Administration)• A Host of Problems• "Numerous studies have been carried out to ascertain whether cytotoxic effects

occur when un irradiated biological test systems are cultured or fed with irradiated media or food. In such studies, adverse physiological growth retardation and inhibition, cytological cell division inhibition and chromosome aberrations and genetical effects have been observed in a wide range of test systems, ranging from bacteriophages to human cells... The available data suggest that a variety of free radicals may act as the toxic and mutagenic agents.“• http://www.mercola.com/article/irradiated/irradiated_research.htm

Radiation Effects. Toxicity of Plants after irradiation.• Cytotoxic and mutagenic effects of irradiated substrates and food

material. Radiation Botany, 11:253-281, 1971.• A Cancer Warning• "An increase in concentration of a mutagen in food by irradiation

will increase the incidence of cancer. It will take four to six decades to demonstrate a statistically significant increase in cancer due to mutagens introduced into food by irradiation. When food irradiation is finally prohibited, several decades worth of people with increased cancer incidence will be in the pipeline.“• http://www.mercola.com/article/irradiated/irradiated_research.htm

Radiation Effects. Toxicity of Plants after irradiation.• Growth, reproduction, survival and histopathology of rats fed beef

irradiated with electrons. Food Research, 20:193-214, 1955.• Chromosomal Damage to Human Cells (I)• "Irradiated sucrose solutions were extremely toxic to human white blood

cells. Cell divisions were inhibited. Degenerated cell divisions were observed and the chromosomes were grossly damaged. The DNA was clumped or the chromosomes appeared shattered or pulverized. In contrast, treatment with un irradiated sucrose at the same concentration had no apparent effect on the mitotic rate and the chromosomes were not visibly damaged.“• http://www.mercola.com/article/irradiated/irradiated_research.htm

Radiation Effects. Toxicity of Plants after irradiation.• Cytotoxic and radiomimetic activity of irradiated culture medium on

human leukocytes. Current Science, 16:403-404, 1966.• Toxic Chemical Formed in Food Containing Fat (I)• "When food containing fat is treated by ionizing radiation, a group of

2-alkylcyclobutanones [toxic chemicals] is formed. To date, there is no evidence that the cyclobutanones occur in unirradiated food. In vitro experiments using rat and human colon cells indicate that 2-dodecylcyclobutanone (2-DCB)... is clearly cytotoxic and genotoxic.“• http://www.mercola.com/article/irradiated/irradiated_research.htm

Radiation Effects. Toxicity of Plants after irradiation.• Radiation Toxins – Effects of Radiation Toxicity, Molecular

Mechanisms of Action, Radiomimetic Properties and Possible Countermeasures for Radiation Injury.• http://www.intechopen.com/books/current-topics-in-ionizing-

radiation-research/radiation-toxins-molecular-mechanisms-of-toxicity-and-radiomimetic-properties-

Radiation Effects. Toxicity of Plants after irradiation.• Some results under way.

Radiation Effects. Toxicity of Plants after irradiation.