[Multidrug resistance increase of tumor cells at the effect of radiation and phorbol ether depends on protein kinase C and reactive oxygen species].

The effects of phorbol ether (PMA) and ionizing radiation on multidrug resistance (MDR) of human larynx cancer cells HEp-2 and the dependences of these effects on protein kinase C (PKC) activity and reactive oxygen species (ROS) production were studied. MDR was determined by transport rate of rhodamine 123 off cells and production of ROS in cells was measured by means of 2'7'-dichlorodigidrofuorescein oxidation to fluorescent 2',7'-dichlorofluorescein. ROS production was increased in cells at PMA treatment. This increase was caused by PKC dependent activation of NADPH-oxidase because the ROS increase suppressed completely with PKC and oxidase inhibitors. It was shown that tumor cell MDR was increased 16 h after PMA (100 nM) and radiation (1 Gy) treatment. The MDR increase depends on PKC activity and ROS increase in cells at both influences since PKC inhibitor and antioxidant, lipoic acid suppressed MDR increase. The obtained data are in accordance with hypothesis about the necessity of activation both signalization and stress reaction for initiation of transcription of transport protein genes which responsible for MDR of tumor cells.

[On the biophysical mechanism of the circum-hour cell rhythm and its role in metabolism].

The universal biophysical mechanism of "circum-hour" oscillations of parameters and properties of various cells and their organelles was considered. The mechanism is the result of nonspecific responses of cells to any damaging agents. At the basis of the "circum-hour" mechanism is the earlier reported inverse dependence of the activity of enzymes on the concentration of low-molecular weight organic substances in medium. The universal character of "circum-hour" oscillations and their probable relationship with the acceleration of metabolism was shown using automatic scanning analyzers of microobjects on numerous objects: cells, cell organelles and compartments.

[On the mechanism of the nonspecific cell response to the action of damaging agents and the nature of hormesis].

A conception of the molecular mechanism of the universal nonspecific cell response (NCR) to the action of all damaging factors of physical and chemical nature is described. The mechanism is shown to be closely related to the general principles of the structural-functional organization of cells and to the properties of biological macromolecules. Among them are the phenomenon of nonspecific regulation of the enzyme activity by low-molecular substances and the compartmentation of the latter within the cell, which is provided by the system of intracellular membranes. Special attention is paid to the role of disturbance of the barrier function, of membranes in the development of the nonspecific cell response, and the activation of repair of cell injuries. Based on the theory of nonspecific cell response, the stimulating effect of weak action of agents that damage the cells at high doses (the phenomenon named as hormesis) is explained.

Dependence of thymocyte apoptosis on protein kinase C and phospholipase A2.

The effect of inhibitors and activators of protein kinase C and phospholipase A2 on radiation-induced apoptosis of rat and mouse thymocytes has been studied. It is shown that the apoptosis is prevented by the protein kinase C inhibitor 1-(5-isoquinolinylsulfonyl)-2-methylpiperasine dihydrochloride and is potentiated by protein kinase C activator phorbol 12-myristate 13-acetate, calcium ionophore A23187 and concanavalin A. The protein kinase C activators initiate apoptosis in mouse but not in rat thymocytes. The inhibitor of phospholipase A2 prevents apoptosis induced by all the factors. The results obtained indicate that both protein kinase C and phospholipase A2 are involved in the thymocyte apoptosis.

Role of arachidonic acid metabolism in thymocyte apoptosis after irradiation.

The present study demonstrates that DNA fragmentation, nuclear pycnosis and trypan blue staining of irradiated thymocytes is prevented by inhibition of the lipoxygenase pathway of arachidonic acid metabolism and is not affected by cyclooxygenase inhibition. Exposed to irradiation [3H]arachidonic acid-labeled thymocytes release radioactive products to the external medium. The process is blocked by the lipoxygenase inhibitor, nordihydroguaiaretic acid. Thus, it can be concluded that irradiation activates arachidonic acid metabolism and that lipoxygenase metabolites play an important role in thymocyte apoptosis.

Intercellular interactions in the interphase death of irradiated thymocytes.

The effect of the interaction of different types of cells on the interphase death and pycnosis of thymocytes irradiated in vitro was studied. When removed from the thymus suspension of cells with natural killer activity, medullary thymocytes and macrophages did not change the radiation-induced death of cortical thymocytes. On the other hand, postirradiation incubation of cortical thymocytes together with unirradiated thymocytes or with cells of certain other cell lines diminished thymocyte death. Mixing the cell suspensions and changing the incubation medium decreased thymocyte death. All of these results indicate that these cells produce soluble mediators that are toxic to the cells that secrete them. The possible nature of these autotoxic mediators has been studied using inhibitors of arachidonic acid metabolism. Inhibitors of phospholipase A2 or lipoxygenase reduced interphase death markedly, while an inhibitor of cyclooxygenase did not. These data suggest that some lipoxygenase products may serve as autotoxic mediators in the interphase death of thymocytes.

Endothelial cell population dynamics in rat brain after local irradiation.

The dynamics of the endothelial cell population was investigated in the rat brain after local irradiation with different doses of X rays. A fluorescent-histochemical technique was used for the visualization of the cells. A decrease in endothelial cell number was observed within 1 day of irradiation with doses of 5-200 Gy. At this time the endothelial cell number had decreased by up to 15% compared with the pre-treatment values. This early dose-independent loss in cell number was maintained for up to 1 month after irradiation. This was then followed by a slow dose-independent decrease in cell density up to 6 months after exposure. Subsequently the depletion of the endothelial cell population exposed to 40 and 60 Gy continued. After a dose of 25 Gy an abortive recovery of cell numbers occurred followed by an abrupt depletion of the endothelial cell population. The possible mechanisms of such changes are discussed.

[Effect of low-molecular amines on DNA conformation and stability of the double helix]. / Vliianie nizhomolekuliarnykh aminov na konformatsiiu i stabil'nost' dvoinoi spirali DNK

Interaction of low-molecular amines (cystamine, cysteamine, cystaphose, asparagine, beta-alanine) with DNA was studied. The amines change the positive circular dichroism (CD) band of DNA as well as temperature and range width of melting. Effect of amines on DNA depends on ionic strength of the solvent, concentration and structure of the ligand. Monamines cause destabilization of DNA double helix followed by stabilization as ligand concentration increases. At concentrations stabilizing the double helix DNA conformation undergoes transition from the B- to C-form. The results obtained enable to relate the stabilizing effect of low-molecular amines and conformational B leads to C-transition to the non-specific interaction of ligand amino groups with DNA phosphates, and the destabilizing effect of monoamines of low concentrations to their interaction with bases, mainly in the denaturated sites of DNA. It is proposed that a stronger effectiveness of amines as compared to monovalent metals in the conformational shift of DNA towards the C-form is due to the additional effect of disturbance of hydrophobic interactions in DNA double helix.

[Mechanism of DNA repair induction by action of ionizing radiation on cells]. / O mekhanizme induktsii reparatsii povrezhdeniia DNK pri deistvii ioniziruiushchego izlucheniia na kletki.

A mechanism is presumed regarding the induction of DNA damage repair at crossing from low to high ionisation doses. Radiation even in low doses disturbs the permeability of membranes and increases the intracellular pH. Alkalation of cells induces changes in chromatin conformation and DNA packing in them. This leads to transcription of different earlier "silent" genes, among them the genes of DNA repair enzymes. This mechanism of induction of DNA damage repair may be common for other physical and chemical agents due to evolutionary-developed unspecific cell reaction vocated for maintenance of intact genome conformation.

[The mechanism of the initiation of low-dose effects]. / O mekhanizme initsiatsii éffektov malykh doz.

A mechanism for initiation effects is proposed. This is based on the theory of non-specific response of cells to damaging influences, which has been developed by the author previously. The effects (adaptive response, stimulation of proliferation, etc.) are believed to be initiated by disturbance of the functions of plasmatic and intracellular membranes which decreases the concentration of low-molecular cellular metabolites and affects their compartmentation. The presence of a lower threshold of the effects, the maximum in the dose dependence as well as dose rate and dose value effects in the range of low doses are explained.

[Is apoptosis a "programmed cell death"?]. / Iavliaetsia li apoptoz "programmirovannoi gibel'iu kletok"?

Many authors consider apoptosis as a programmed cell death (PCD). Their opinion is based usually on the dependence of cell death on the protein-opinion and RNA synthesis, on the intranucleosomal fragmentation of DNA and on the expression of genes, which can induce apoptosis. However our analysis of numerous literature data on the apoptotic death (judging by their morphological criteria) of different cell kinds induced by various agents shows: internucleosomal fragmentation of DNA is neither a trigger, nor a reliable proof for apoptotic death; protein- and RNA synthesis, as well as expression of P53 genes is also not a necessary condition for that and is important only in some combinations of the cell type and state and the nature of the damaging agent. Thus, the existence of immanent cell death program requires some more proofs and for present the term PCD may be undoubtedly attributed only to the genetically determined processes intended to remove such cells and organs which become unnecessary for the following development, in short, PCD is a term on the level of organism. We propose that character and sequence of destructive processes at apoptotic cell death are conditioned by the conformational changes in genome as result of the cellular environment changes caused by the action of damaging agents on the cellular membranes.

[Extrapolation of dose-effect relationships for cytogenetic aberrations from high to low doses]. / O probleme ékstrapoliatsii dozovoi zavisimosti tsitogeneticheskikh povrezhdenii ot bol'shikh doz k malym.

It is known, that extrapolation of dose response curve for cytogenic lesions, based on the linear non-threshold concept is not correct due to anomalies in the low dose region, among them the dose intervals of the so-called "low dose effects" (LDE) and "plato"-effect. As a rule they are attributed to assumed heterogeneity of cell populations or to existence of several repair systems. Analysing the data on modification the cytogenetic lesions we concluded, that in the whole low-dose range functions one on the same repair system, but it "switches on" only at sufficient reorganization of chromatin (accompanied by expression of before "silent" genes), caused by radiation-induced increase of the membranes permeability. In the dose interval of LDE this occurs only in vary narrow diapason of dose rate (approximately(1-30) cGy/min for many cell kinds); in the "plato" dose interval--in its beginning. By the assessment of cancerogenic risk the linear non threshold concept can be used only for dose dependence, obtained when repair is absent or inhibited.

[Interphase death of irradiated thymocytes--result of the "bystander effect"]. / Interfaznaia gibel' obluchennykh timotsitov--rezul'tat "éffekta svidetelia".

It was shown that interphase death of thymocytes, which mechanism was established by author's laboratory already over 10 years ago belongs to phenomena which recently was named as caused by "bystander effect".

[A single mechanism for the initiation of different effects of low doses of ionizing radiation]. / O edinom mekhanizme initsiatsii razlichnykh éffektov malykh doz ioniziruiushchikh izluchenii.

Main regularities of different endpoints of low dose effects (adaptive response, stimulation of proliferation, special radiosensitivity of lymphoid cells, and others) have been examined. It has been shown that these endpoints have a commonness for the dose interval, the shape of the dose-effect curve, the reverse effect of dose rate, non-specificity toward initiating agents, and others. An explanation is suggested for the common mechanism of the initiation of all the studied low dose effects, basing on the theory of the non-specific reaction of cell to external influences. It is concluded that initiation of the low dose effects is conditioned by radiation induced damage of functions of plasmic and internal membranes.

[Problems of the mechanism of radiation and chemical hormesis]. / Problemy mekhanizma radiatsionnogo i khimicheskogo gormezisa.

Based on the "membrane" mechanism of biological action of low radiation doses, described earlier, a possible common explanation of radiation and chemical hormesis initiation is proposed.

[The cause-effect relationships between different indices of radiation lesions of the thymocytes]. / K voprosu o prichinno-sledstvennykh sviazakh mezhdu raznymi pokazateliami radiatsionnogo porazheniia timotsitov.

The dependence of the kinetics of different thymocyte injuries of the experimental conditions was investigated. It was shown that the rate of cell death detecting by cell staining decreases with the increase of the dye molecular size. Different organic substances decrease the staining rate of the dead cells with trypan blue but increase the rate of DNA fragmentation. They change the kinetics of cell injuries development only after but not during irradiation. Development of pycnoses forestalls the DNA fragmentation. Basing on these findings the conclusion was made that the time-course of different thymocyte injuries development cannot be used for a statement what type of injuries--of plasmic membrane or cell nucleus--is initial.

[The role of cell interaction in initiation of radiation-induced death of thymocytes and apoptosis]. / Rol' mezhkletochnogo vzaimodeistviia v initsiatsii radiatsionnoi gibeli timotsitov i problema apoptoza.

The experimental evidence of crucial importance of cell interactions for initiation of radiation-induced thymocyte death was obtained. It has been proposed that ionizing radiation, in contrast to many other cytotoxic agents, initiates thymocyte death simultaneously by different independent pathways, affecting all cellular components. The concept of apoptosis as "programmed cell death" has been critically discussed.