Molecular and Cellular Radiobiology

Characteristics of ionizing radiation such as charge, mass, and energy vary among the different types of radiation. These attributes determine the extent to which different radiation modalities transfer energy into biologic tissue. To understand the way ionizing radiation causes injury and how the effects may vary in biologic tissue, three important concepts must be studied: 1.Linear energy transfer (LET) 2.Relative biologic effectiveness (RBE) 3.Oxygen enhancement ratio (OER)

Linear Energy Transfer (LET) When passing through a medium, ionizing radiation may interact with it during its passage and, as a result, deposit energy along its path (called a track). The average energy deposited per unit length of track is called linear energy transfer (LET). The energy average is calculated by dividing the track into equal energy intervals and averaging the lengths of the tracks that contain that specific energy amount. LET is generally described in units of kiloelectron volts (keV) per micron (1 micron [μm] = 10−6 m). Because the amount of ionization produced in an irradiated object corresponds to the amount of energy it absorbs and because both chemical and biologic effects in tissue coincide with the degree of ionization experienced by the tissue, LET is an important factor in assessing potential tissue and organ damage from exposure to ionizing radiation.

Low-LET Radiation High-LET Radiation Gamma rays Alpha particles X-rays Ions of heavy nuclei Charged particles released from interactions between neutrons and atoms Low-energy neutrons

Low-LET Radiation Low-LET radiation is electromagnetic radiation such as x-rays and gamma rays (short-wavelength, high-energy waves emitted by the nuclei of radioactive substances). This penetrating electromagnetic radiation is sparsely ionizing and interacts randomly along the length of its track. It does not relinquish all of its energy quickly. When low-LET radiation interacts with biologic tissue, it causes damage primarily through an indirect action that involves the production of free radicals. Also, but much less likely, the radiation may directly induce single-strand breaks in the ladderlike deoxyribonucleic acid (DNA) structure. Because low-LET radiation generally causes sublethal damage to DNA, repair enzymes can usually reverse the cellular damage. Because of a property known as wave-particle duality, we can also refer to x-rays and gamma rays as streams of particles, each of which has no mass and no charge

High-LET Radiation High-LET radiation includes particles that possess substantial mass and charge. These radiations cause dense ionization along their length of track. Some typical examples of high-LET radiations are alpha particles, ions of heavy nuclei, and charged particles released from interactions between neutrons and atoms. Low-energy neutrons, which carry no electrical charge, also are a form of high-LET radiation. All of these lose energy more rapidly than low-LET radiations because they produce much more ionization per unit of distance traveled. As a result, they exhaust their energy in a shorter length of track and therefore cannot travel or penetrate as far. Because high-LET radiations deposit more energy per unit of biologic tissue traversed, they are more destructive to biologic matter than are low-LET radiations.

For radiation protection, high-LET radiation is of greatest concern when internal contamination is possible—that is, when a radionuclide has been implanted, ingested, injected, or inhaled. Then the potential exists for irreparable damage because, with high-LET radiation, multiple-strand breaks in DNA are possible.

Relative Biologic Effectiveness Biologic damage produced by radiation escalates as the LET of radiation increases, because identical doses of radiations of various LETs do not render the same biologic effect. The relative biologic effectiveness (RBE) describes the relative capabilities of radiation with differing LETs to produce a particular biologic reaction. RBE of the type of radiation being used is the ratio of the dose of a reference radiation (conventionally 250-kVp x-rays) to the dose that is necessary to produce the same biologic reaction in a given experiment. The reaction is produced by a dose of the test radiation delivered under the same conditions

Protraction and Fractionation If a dose of radiation is delivered over a long period of time rather than quickly, the effect of that dose is lessened. Stated differently, if the time of irradiation is lengthened, a higher dose is required to produce the same effect. This lengthening of time can be accomplished in two ways. If the dose is delivered continuously but at a lower dose rate, it is said to be protracted. Six hundred rad (6 Gyt) delivered in 3 min (200 rad/min [2 Gyt/min]) is lethal for a mouse. However, when 600 rad is delivered at the rate of 1 rad/hr (10 mGyt/hr) for a total time of 600 hr, the mouse will survive If the 600 rad dose is delivered at the same dose rate, 200 rad/min, but in 12 equal fractions of 50 rad (500 mGyt), all separated by 24 hr, the mouse will survive. In this situation, the dose is said to be fractionated. Dose protraction and fractionation cause less effect because time is allowed for intracellular repair and tissue recovery

The oxygen enhancement ratio (OER) is the ratio of the radiation dose required to cause a particular biologic response of cells or organisms in an oxygen-deprived environment to the radiation dose required to cause an identical response under normal oxygenated conditions. Because high-LET radiations such as alpha particles produce their biologic effects from direct action—namely, direct ionization and disruption of biomolecules—the presence or absence of oxygen is of no consequence. Therefore, the OER of high-LET radiation is approximately equal to 1. For low-LET radiation a significant fraction of bioeffects are caused by indirect actions in which a chemical species called a free radical is formed. A free radical is a solitary atom or most often a combination of atoms that behaves as an extremely reactive single entity as a result of the presence of an unpaired electron. Free radicals dramatically increase the amount of biologic damage. However, the presence of oxygen in biologic tissues makes the damage produced by these free radicals permanent because oxygen reacts with free radicals to produce organic peroxide compounds, which represent non-restorable changes in the chemical composition of the target material. Without oxygen, damage produced by the indirect action of radiation on a biologic molecule may be repaired, but when damage occurs through an oxygen-mediated process, the end result is permanent, or fixed. This phenomenon has been called the oxygen fixation hypothesis.

MOLECULAR EFFECTS OF IRRADIATION In living systems, biologic damage resulting from exposure to ionizing radiation may be observed on three levels: molecular, cellular, and organic. Any visible radiation-induced injuries of living systems at the cellular or organic level always begin with damage at the molecular level. Molecular damage results in the formation of structurally changed molecules that may impair cellular functioning

Classification of Ionizing Radiation Interaction When ionizing radiation interacts with a cell, ionizations and excitations (the addition of energy to a molecular system, transforming it from a ground state to an excited state) are produced either in vital biologic macromolecules (such as DNA), or in water (H2O), the medium in which the cellular organelles are suspended. Based on the site of the interaction, the action of radiation on the cell is classified as either direct or indirect. In direct action, biologic damage occurs as a result of ionization of atoms on master, or key, molecules (DNA), which can cause these molecules to become inactive or functionally altered. Indirect action refers to the effects produced by reactive free radicals that are created by the interaction of radiation with water (H2O) molecules. These unstable, highly reactive agents have the potential to substantially disrupt master molecules, resulting in cell death. Direct action may occur after exposure to any type of radiation. However, direct action is much more likely to happen after exposure to high-LET radiations such as alpha particles, which produce a very large number of ionizations in a very short distance of travel. This is in contrast to exposure to low-LET radiations such as x-rays, which are only sparsely ionizing.

Direct Action When ionizing particles interact directly with vital biologic macromolecules such as DNA, ribonucleic acid (RNA), proteins, and enzymes, damage to these molecules occurs from absorption of energy through photoelectric and Compton interactions. The ionization or excitation of the atoms of the biologic macromolecules results in breakage of the macromolecules' chemical bonds, causing them to become abnormal structures, which may in turn lead to inappropriate chemical reactions. Thus, when enzyme molecules are damaged by interaction with ionizing particles, essential biochemical processes may not occur in the cell at the appropriate time. For example, if an enzyme is inactivated, it will not be available to facilitate a particular biochemical reaction. Should this occur during the synthesis of a particular protein, the protein will not be manufactured, and if this protein was intended to perform a specific function, its nonexistence will hinder or prevent that function. In the event that other cell operations depend on the suppressed function, these operations sustain some type of damage as well, and so a biologic chain reaction essentially occurs.

Ionization of Water Molecules X-ray photons may interact with and ionize water molecules contained within the human body. Such an interaction between an x-ray photon and a water molecule creates an ion pair consisting of a water molecule with a positive charge (HOH+) and an electron (e−). After the original ionization of the water molecule, several reactions can occur. One is that the positively charged water molecule (HOH+) may recombine with the electron (e−) to re-form a stable water molecule (HOH+ + e− = H2O). If this happens, no damage occurs. Alternatively, the electron (the negative ion) may join with another water molecule, producing a negative water ion (H2O + e− = HOH−).

Production of Free Radicals The positive water molecule (HOH+) and the negative water molecule (HOH−) are basically unstable. Hence they will break apart into smaller molecules. HOH+ becomes a hydrogen ion (H+) and a hydroxyl radical (OH*), whereas HOH− becomes a hydroxyl ion (OH−) and a hydrogen radical (H*). The asterisk symbolizes a free radical. A free radical is a configuration of one or more atoms having an unpaired electron but no net electrical charge. This object is highly reactive because the unpaired electron will pair up with another electron even if it has to break a chemical bond to do this. Hence, the interaction of radiation with water results in the formation of an ion pair, H+ and OH− (hydrogen ion and hydroxyl ion), and two free radicals, H* and OH* (a hydrogen radical and a hydroxyl radical)

Production of Undesirable Chemical Reactions and Biologic Damage Because the hydrogen and hydroxyl ions usually recombine to form a normal water molecule, the existence of these ions as free agents within the human body is insignificant in terms of biologic damage. The presence of hydrogen and hydroxyl free radicals, however, is not insignificant. As molecules containing an unpaired electron in their outer shell, they are chemically unstable and very reactive. They can produce undesirable chemical reactions and cause biologic damage by transferring their excess energy to other molecules, thereby either breaking these molecules' chemical bonds or at the very least causing point lesions (i.e., altered areas caused by the breaking of a single chemical bond) in the molecule. Approximately two thirds of all radiation-induced damage is believed to be ultimately caused by the hydroxyl free radical (OH*). In addition, because free radicals have excess energy and can travel through the cell, they are capable of destructively interacting with other molecules located at some distance from their place of origin.

Hydrogen and hydroxyl radicals are not the only destructive substances that may be produced during the radiolysis of water. A hydroxyl radical (OH*) may bond with another hydroxyl radical (OH*) and form hydrogen peroxide (OH* + OH* = H2O2), a substance that is poisonous to the cell. Also, a hydroperoxyl radical (HO2*) is formed when a hydrogen free radical (H*) combines with molecular oxygen (O2). This radical and hydrogen peroxide are believed to be among the primary substances that produce biologic damage directly after the interaction of radiation with water.

Indirect Action When free radicals previously produced by the interaction of radiation with water molecules act on a molecule such as DNA, the damaging action of ionizing radiation on the vital biologic macromolecule is indirect in the sense that the radiation is not the immediate cause of injury to the macromolecule. It is the by-products of the radiation, the free radicals, that are the immediate cause of this damage. Because the human body is 80% water and less than 1% DNA, essentially all other effects of irradiation in living cells result from indirect action

Recovery In vitro experiments show that human cells can recover from radiation damage. If the radiation dose is not sufficient to kill the cell before its next division (interphase death), then given sufficient time, the cell will recover from the sublethal radiation damage it has sustained. This intracellular recovery is due to a repair mechanism inherent in the biochemistry of the cell. Some types of cells have greater capacity than others for repair of sublethal damage. At the whole-body level, this recovery from radiation damage is assisted through repopulation by surviving cells. If a tissue or organ receives a sufficient radiation dose, it responds by shrinking. This is called atrophy, and it occurs because some cells die and disintegrate and are carried away as waste products. If a sufficient number of cells sustain only sublethal damage and survive, they may proliferate and repopulate the irradiated tissue or organ. The combined processes of intracellular repair and repopulation contribute to recovery from radiation damage.