Radiation Response Of Mammalian Cells Research Articles

Bryostatin-1 causes radiosensitization of BMG-1 malignant glioma cells through differential activation of protein kinase-Cδ not evident in the non-malignant AA8 fibroblasts.

Bryostatin-1 (bryo-1), a non-phorbol ester, is known to sensitize mammalian cells against certain chemotherapeutic drugs. We assessed its ability to modify radiation response of mammalian cells using Chinese hamster fibroblasts AA8 cells and human malignant glioma BMG-1 cells. In the malignant glioma BMG-1 cell line, bryo-1 pre-treatment significantly enhanced radiation-induced growth inhibition and cytogenetic damage, and further reduced the clonogenic cell survival as compared to cells irradiated at the clinically relevant dose of 2Gy. PKCδ expression increased significantly when bryo-1 pre-treated BMG-1 glioma cells were irradiated at 2Gy and induced prolonged ERK-1/2 activation associated with p21 overexpression. Silencing PKCδ resulted in inhibition of bryo-1-induced radiosensitization. In contrast, bryo-1 failed to alter radiosensitivity (cell survival; growth inhibition; cytogenetic damage) or activate ERK1/2 pathway in the AA8 fibroblasts despite PKCδ phosphorylation at its regulatory (Y155) domain, indicating alternate mechanisms in these non-malignant cells as compared to the glioma cells. This study suggests that bryo-1 may effectively enhance the radiosensitivity of malignant cells and warrants further in-depth investigations to evaluate its radiosensitizing potential in various cell types.

Identification of the human DEAD-box protein p68 as a substrate of Tlk1

The activity of the human protein kinase Tlk1 is down-regulated within minutes after exposure of cells to ionizing radiation. In order to identify signaling pathways which might be relevant in the radiation response of mammalian cells we screened nuclear proteins for substrates of Tlk1. Among several proteins one could be identified as p68 RNA helicase. Furthermore, it could be shown that Tlk1 phosphorylates immunoprecipitated p68. The phosphorylation of the C-terminal fragment of p68 by rTlk1 reduced its affinity to single stranded RNA in a gel shift assay. In addition, it could be demonstrated that increasing the Tlk1 activity in HT1080 cells by forced Tlk1 overexpression leads to an increased phosphorylation of endogenous p68, arguing that p68 might be a physiological substrate of Tlk1. These findings open the possibility that Tlk1 might participate in diverse biologic functions like cell growth and differentiation, pre-mRNA splicing, and transcriptional coactivation.

The effects of 2-deoxyglucose and amino-oxyacetic acid on the radiation response of mammalian cells in vitro

. Cells use substrates such as glucose and glutamine to provide energy for repair of radiation damage. Glutaminolysis and glycolysis were inhibited by aminooxyacetic acid (AOAA) and 2-deoxyglucose (2DG), respectively, to inhibit metabolism of these substrates in order to determine the effect on radiation response of CHO-KI cells in vitro. Exposure to treatments which inhibit energy metabolism resulted in alterations in radiosensitivity and, in general, a reduction in cellular recovery rate after gamma -irradiation but varied with regard to the extent of recovery. The greatest inhibition of recovery relative to that in normal culture medium was found with medium which lacked glucose and glutamine and contained 2DG and AOAA. In contrast, medium lacking glucose and glutamine without the addition of inhibitors resulted in an increase in recovery. It is proposed that the efficiency of energy pathways such as glycolysis and glutaminolysis and their interaction are determinants of both radiosensitivity and recovery.

Effects of strong magnetic fields on cell growth and radiation response of human T-lymphocytes in culture.

Experiments were undertaken in order to verify whether or not a strong magnetic field would have any biological effects on the cell growth, viability and radiation response of mammalian cells. Magnetic field exposures were conducted using a superconducting magnet with freshly-isolated human peripheral blood T-lymphocytes maintained at their normal growing temperature of 37 degrees C. The static magnetic fields with intensities up to 6.3-tesla (T) exerted little influence on the cell growth and viability of actively-growing T-lymphocytes under normal cell-culture conditions. On the other hand, the T cells exposed to the magnetic fields (4 T-6.3 T) during PHA stimulation were inhibited in their cell growth when compared to controls. The effects of the magnetic fields with intensities up to 2 T on cell growth properties, however, were minimal in this system. Also, the radiosensitivity of T-lymphocytes previously exposed to the strong magnetic fields was more sensitive than that of control cells. These results suggest that exposure to a static magnetic field of 4 T or stronger might lead to physiological and growth abnormalities at the cellular level.

Involvement of poly (ADP-ribose) in the radiation response of mammalian cells.

The evidence implicating poly (ADP-ribose) in the radiation response of mammalian cells is reviewed. It is concluded that the apparently conflicting results using inhibitors of ADP-ribosyl transferase (ADPRT) can be explained by a working hypothesis. This hypothesis maintains that poly (ADP-ribose) is required for repair of radiation damage (presumably to facilitate ligation). In most cells the synthesis of poly (ADP-ribose) is not rate limiting for repair and therefore, an almost complete inhibition of ADPRT activity is required to potentiate the radiation response. In radiation-sensitive cells (e.g. resting lymphocytes, L5178Y-S cells) with a deficient poly (ADP-ribose) metabolism, its synthesis can become rate limiting for repair. In such cells even a partial inhibition of ADPRT activity may enhance radiation-induced cell killing. It is suggested that if such differences exist between normal and cancer cells, they can be utilized to improve the therapeutic ratio of radiotherapy.

Effect of membrane fatty acid changes on the radiation sensitivity of human lymphoid cells.

To test the influence of changes in membrane fatty acid composition on the radiation response of mammalian cells, human LDV cells were cultured in medium containing delipidated serum supplemented with either oleic or linoleic acid. Analysis of lipid extracts of the cells by gas liquid chromatography showed that, after 3 or more days growth in oleic or linoleic acid supplemented media, there were substantial overall increases in the proportion of oleate and linoleate, respectively, in the cellular lipid. Smaller absolute changes were measured in nuclear phospholipid, which was found to be low in unsaturated phospholipid compared to the rest of the cell. Fluorescence polarization measurements using diphenylhexatriene indicated an increased membrane fluidity in cells grown in the presence of excess linoleic acid and to a lesser extent for oleic acid. The clonogenic capacity of the cells after irradiation in air or nitrogen was not altered by any of these membrane compositional changes. The lack of effect on radiation sensitivity, in contrast to that reported for bacteria (E. coli K1060), is consistent with the fact that little or no change was brought about in the nuclear membrane composition, since evidence from partial cell irradiation experiments indicates that the cell nucleus is the sensitive target for cell killing by ionizing radiation. The ability of the cell to maintain a low level of unsaturated phospholipids in its nuclear membrane may be an important general defence mechanism against free radical damage to chromatin mediated by lipid peroxidation.

Use of solubilizing agents to study radiosensitization in vitro by compounds of low water solubility.

A major disadvantage in studying the effects of chemicals on the radiation response of mammalian cells in vitro can be the poor water solubility of the chemicals. Using Chinese hamster cells we have overcome this problem by using dimethyl sulphoxide and ethanol as co-solvents. From studies using these co-solvents and misonidazole as a standard we have demonstrated that these co-solvents have no effect on sensitizing efficiency and cytotoxicity. A number of poorly water soluble 2-nitroimidazoles of promising structure have been studied. The results, together with those from studies on misonidazole and the co-solvents are presented.

Radioprotection of mammalian cells by procaine.

The cellular membrane has been recognized for some time as an important target for radiation injury (Myers, 1970; Alper, 1971). In addition to this, it is also believed to play an important role in repair processes, thus influencing the ultimate survival of the irradiated cells. In an attempt to elucidate these mechanisms further, we have examined the ability of certain “membrane-active” compounds to modify the radiation response of mammalian cells under various experimental conditions. In this report we present the radioprotective effect afforded by a local anaesthetic, procaine hydrochloride (PH), on L5178Y, a murine leukaemic cell line. Asynchronous L5178Y cells were grown in Fischer's Medium plus 10% horse serum. Cultures were harvested by centrifugation and resuspended in serum-free medium to final concentration of 2 × 105 cells/ml. Cells were irradiated at room temperature with a Picker 250 kV X-ray generator at a dose rate of 60 rad/min. An appropriate amount of 1 M PH solution, made up in saline, ...

The effect of hypoxia on the growth and radiation response of mammalian cells in culture.

Abstract During prolonged hypoxia, a progressive change occurs in the radiosensitivity of cultured mammalian cells. An attempt was made to determine whether this could be explained on the basis of a progressive change in the life cycle or age distribution of cells during hypoxia. Cell population growth kinetics and the cell life cycle were studied by two conventional methods of analysis. Both indicated an appreciable heterogeneity in the cell populations with regard to the ability of individual cells to negotiate their life cycle during hypoxia. This heterogeneity was confirmed by studying the development of individual microcolonies under hypoxia. From this study, the following conclusions were reached regarding changes which occurred during hypoxia: (1) the growth rate was decreased and the life cycle lengthened; (2) the overall proportion of cells in DNA synthesis decreased with time; (3) on average, a disproportionate lengthening of the G1 phase of the life cycle was implicated; and (4) these changes w...

RADIOBIOLOGICAL SUPPORT FOR RADIOTHERAPY: WHAT KIND OF RESEARCH?

The radiation response of mammalian cells varies not only with their origin but also with conditions of growth; so laboratory observations on human or other mammalian cells and tissues are inadequate as a guide to the mode of their response in situ. Thus there is no basis at present, other than clinical experience, for prescribing the most effective radiotherapeutic regimen for treatment of a given tumour. The problem is exemplified by results with a pair of rat-tumour lines of common origin and similar histology. Neutrons would be more successful than conventional radiotherapy for one member of the pair but less so for the other; whereas the positions would be reversed if hyperbaric-oxygen therapy were under test. Thus radiobiological research should be directed towards uncovering biological characteristics associated with variations in radiation response. To this end, the most suitable methods are not necessarily those claimed to be "related to the clinical situation ", although these may be regarded as more worthy of funding. On the other hand, much more quantitative information is needed from clinicians on the relationship of dose to effect on both human tumours and normal tissues.

Variation in Radiation Response of Mammalian Cells as a Function of Oxygen Tension

Complete radiation survival curves for mammalian cells as a function of partial pressure of oxygen have been obtained with considerable medium present. The results show an apparent decrease in extrapolation number (n) when irradiation is carried on during conditions of moderate hypoxia (5000 ppm oxygen) with the n value returning to the air level when the hypoxia becomes extreme. This phenomenon can be explained by radiochemical depletion of oxygen. At less than 25 ppm oxygen, the cells are not capable of sublethal repair, while at 200 ppm oxygen repair proceeds at almost the aerobic rate.

Radiation response of mammalian cells grown in culture: III. Modification of X-ray survival of Chinese hamster cells by 5-bromodeoxyuridine

Chinese hamster cells (strain V79-1) cultured in vitro show an increased response to the lethal effects of X-rays when they are grown prior to irradiation in 5-bromodeoxyuridine (BUDR). These cells appear to be more sensitive than the human cell line, D98/AG, studied by Erickson and Szybalski to the radiation modifying as well as the cytotoxic effects of BUDR. BUDR produces a striking change in the extrapolation number of X-ray survival curves of V79-1 as well as the change in slope reported in D98/AG. Both of these changes appear to be the result of alterations in cellular radiation response and not the result of population heterogeneity. When cells having an increased X-ray response induced by BUDR are removed from contact with BUDR, they reassume X-ray survival parameters similar to normal cells within one or two generations.

Influence of Temperature on the Radiation Response of Mammalian Cells in Tissue Culture

Mammalian cells in tissue culture were irradiated at 5 un. Concent 85% C, 37 un. Concent 85% C, or 40 un. Concent 85% C. Cells irradiated at 5 un. Concent 85% C demonstrate radioresistance compared to cells irradiated at 40 un. Concent 85% C by a factor of 1.59. It is suggested that low temperature results in protection rather than enhances recovery. The nature of this protection is not known but it is thought to be associated closely with slowed rate processes and recombination of molecules in the metionic state before deieterious reactions can take place. (auth)