Microbeam Studies of the Sensitivity of Structures Within Living Cells

Scanning Microscopy Volume 6 Number 1 Article 13 12-30-1991 Microbeam Studies of the Sensitivity of Structures within Living Cells L. A. Braby Pacific Northwest Laboratory Follow this and additional works at: https://digitalcommons.usu.edu/microscopy Part of the Biology Commons Recommended Citation Braby, L. A. (1991) "Microbeam Studies of the Sensitivity of Structures within Living Cells," Scanning Microscopy: Vol. 6 : No. 1 , Article 13. Available at: https://digitalcommons.usu.edu/microscopy/vol6/iss1/13 This Article is brought to you for free and open access by the Western Dairy Center at DigitalCommons@USU. It has been accepted for inclusion in Scanning Microscopy by an authorized administrator of DigitalCommons@USU. For more information, please contact [email protected]. Scanning Microscopy, Vol. 6, No. 1, 1992 (Pages 167-175) 0891-7035/92$5 .00+ .00 Scanning Microscopy International, Chicago (AMF O'Hare), IL 60666 USA MICROBEAMSTUDIES OF THESENSITIVITY OF STRUCTURESWITHIN LIVING CELLS L.A. Braby* Battelle, Pacific Northwest Laboratory Richland WA.99352 (Received for publication May 4, 1991, and in revised form December 30, 1991) ABSTRACT INTRODUCTION Determining the biological effects of low doses A new generation of charged-particle microbeam of radiation with high linear energy transfer (LET) irradiation systems is being installed at several is complicated by the stochastic nature of charged laboratories around the world. These systems have particle interactions. Populations of cells been designed to answer some fundamental questions exposed to very low radiation doses contain a few about the hazards of low doses of ionizing radia cells which have been hit by a charged particle, tion. The key to answering these 1ong standing while the majority of the cells receive no radia questions is the ability to detect each charged tion damage. At somewhat higher doses, a few cells particle as it interacts with a cell, and limit the receive two or more events. Because the effects of exposure of each cell to a predetermined value. damage produced by separate events can interact in The ability of ionizing radiation to damage the cell, we have had to make assumptions about the living systems was recognized soon after the nature of these interactions in order to interpret discovery of x-rays. Si nee then, the nature of the results of the experiments. Many of those this damage has been studied extensiv~ly in order assumptions can be tested if we can be sure of the both to optimize the benefit in medical appl ica number of charged- particle events which occur in tions such as cancer therapy, and to minimize the individual cells, and correlate this number with effects of envi ronmenta1 exposures such as those the biological effect. produced by radon progeny captured in energy Wehave developed a special irradiation facili efficient houses. We probably know more about the ty at Pacific Northwest Laboratory (PNL) to control effects of radiation than any other environmental the actual number of charged particle tracks that carcinogen, and yet we still lack answers to basic pass through cell nuclei . The beam from a 2 MeV questions such as the shape of the dose response tandem accelerator is collimated to approximately relationship at low doses. The major factor 5 µm. Cells, grown in special dishes with l.5µm limiting the investigation of low dose effects has thick plastic bottoms, are positioned so that the been the stochastic nature of the physical interac desired portion of the cell aligns with the colli tion of ionizing radiation with individual cells, mator. A shutter in the beam line is opened and but this limitation can be overcome by irradiating closed after the desired number of particle tracks individual cells with specific numbers of parti has been counted. cles. This approach can be used to investigate the For a very long time microbeam irradiation has effects of the interaction between irradiated and been used to study the function of living cells and unirradiated cells in an organized system, as well to investigate the response to radiation. Initial as to study the effects of spatial and temporal ly UVlight was used, but gradually techniques for distribution of radiation damage within single charged-particle microbeams were added (Zirkle cells. We expect that this approach will lead to 1957). The goals of these experiments have evolved a better understanding of the mechanisms of high as has our understanding of the structure of living LET radiation effects. cells and the effects of radiation have developed. The earliest experiments used a microbeam as a tool for microsurgery to investigate the function of KEYWORDS: microbeam, single particle, high linear subcellular structures within individual cells. energy transfer, cell survival. Later, the emphasis changed to determining the portion of the cell which is sensitive to the effects of radiation, and now the emphasis is on understanding the mechanisms which allow very small chemical changes in individual cells to produce major health effects such as cancer. *Address for correspondence: L. A. Braby P8-47 Battelle Northwest Richland WA 99352 Phone No. (509) 376-5915 FAX(509) 376-9944 167 L.A. Braby Radiation Effects on Living Systems It is well known that ionizing radiations damage living material . Both beneficial appl i ca tions as we17 as potential risks such as cancer induction were identified soon after X-rays were discovered. In order to estimate the magnitude of the effect which would be produced by a specific radiation exposure, it was necessary to define a unit for measuring the radiation. Radiations from different sources, for example alpha particles from radon and gammarays from potassium 40, all deposit energy by producing ionizations and excitations in matter. It was observed that all types of ionizing radiation produced the same type of initial ioniza tions, and that the number of i oni zat ions was proportional to the energy deposited, so the radiation dose was defined in terms of the energy deposited per unit mass. The definition of this quantity has evolved with changes in measurement systems and with improving understanding of the physical processes involved, but the current definition (ICRU 1980) is still based on the assumption that the biological effect is related to the energy deposited by the radiation. However, the relationship is not a simple one. The inacti vation of dry enzymes and some other simple systems is a linear function of dose, but most plant and animal systems display a distinctly nonlinear behavior (Elkind and Whitmore 1967). At the lowest doses that can be used in experiments the effect per unit dose is relatively small, but as the dose increases, the effect per unit dose also increases. At still higher doses, the effect begins to de crease again (see high dose rate gammaray curve in Figure 1). This curve indicates that products of successive energy transfers to the cell interact with each other to increase the effectiveness of the later events, but if the dose is high enough, Figure 2. Photograph of a mammaliancell culture with the doses (in cGy) to the cell nuclei calcu lated by computer simulation. The mean single event dose to a cell nucleus was assumed to be 40 Neutrons cGy and the dose to the sample was 40 cGy. High Dose Rate additional factors which prevent expression of the effect become dominant. Pl ant and animal ce 11s al so show an increase in biol ogi cal effect with increasing linear energy transfer {LET), i.e. the ti stopping power of the radiation, in spite of the wl fact that the same type of ionizations are produced by all radiations. However, the spatial separation of the ionizations along the track and the range of delta rays extending from the track do differ with / ~ma LET and charged particle velocity, and it is now ,_____-- ~~~- Dose assumed that clusters of ionizations in volumes a Rate few nanometers in diameter are responsible for much 1////////////// 1/////////////////////,1///,½1////, of the observable effect (Goodhead 1987). Finally, it is observed that the magnitude of the response Dose of most biological systems depends on the dose rate at which the damage is delivered. For all end Figure 1. Schematic dose response relationships points which have been tested, decreasing the dose for a typical biological system. Experimental data rate of low LET radiation reduces the effect. are usually available only for the relatively high However, for malignant transformation by high LET doses indicated by the curves, while concerns about particles, the dose rate effect is still unsettled. hea 7th risks deal primarily with sma11 effects In some experiments the transformation seems to indicated by the shaded area. increase with decreasing dose rate, while in other 168 Microbeam Studies of Living Cells experiments the effect decreases with decreasing dose rate (the dashed curves in Figure 1). Figure l also demonstrates a major limitation of the data currently available for determining the risks associated with radiation exposure. All of the experimental data are for doses which give a relatively high probability of producing an effect, while the concern in health protection is over the low rates of effects which occur at lower radiation doses. Pr act i cal matters such as the number of cells which must be counted and the effects of biological variability between experimental cul tures are generally responsible for the lack of low dose experiments with low LET radiation. However, the interpretation of low dose experiments with high LET radiations is limited by the physics of the charged-particle interactions which deposit the Figure 3. Small beams can be produced by simple energy. Dose is an average quantity, the energy collimation (left) or by focusing (right) an image deposited divided by the mass of the irradiated of a distant source with a short focal length lens.

Load more