Brain Damage Among Individuals Exposed Prenatally to Ionizing Radiation:

A 1993 Review WILLIAM J. SCHULL

U.T. Health Sciences Center, Medical Genetics Center, Houston, Texas, USA

Key Words. Prenatal radiation exposure . Mental retardation Intelligence quotient . Neuronal migration . Japan

ABSTRACT Mental retardation as a result of prenatal

exposure to ionizing radiation is not a common phenomenon when compared to the incidence of cancer, but it has nevertheless been well- documented. This article describes results from studies of individuals who were exposed prenatally to radiation in Hiroshima and Nagasaki. The critical time of exposure, when the most significant damage was done, was dur- ing the 8th-15th week of gestation, with a lesser

effect at 16-25 weeks. Individuals in the study were assessed by measurement of an intelli- gence quotient and by examination of school performance. Studies show that the period of 8- 15 weeks of gestation coincides with a key time for neuronal cell migration in the developing brain. There is continuing investigation of the mechanism of this migration and how it might be disrupted by ionizing radiation. Stem Cells 1997~15(~~ppl2):129-I33

INTRODUCTION Perhaps it seems odd that a discussion of brain damage should be included in a symposium deal-

ing with the effects of exposure to low doses of ionizing radiation. After all, on the basis of current information and the sorts of doses that we anticipate from an accident such as that which occurred at Chernobyl or might occur in other accidents, organic brain damage would not appear to be a par- ticularly likely risk. Leaving aside the psychosocial damage that may ensue, organic brain damage would not appear to be a major threat.

However, this topic may indeed be appropriate because of all the stories that have emerged from the studies of the survivors of Hiroshima and Nagasaki; the most poignant by far is the story of those individuals who were exposed prenatally. A substantial number of them have been condemned from their birth to a life of social inadequacy. This has been a burden not only for the exposed person, but also to their families. Now, as these individuals reach the ages of 47 and 48 and their parents are dying, there is great concern about who will manage the care of these individuals in the future. This discussion will begin with a few brief remarks about the nature of the studies, a review of the evidence that has emerged from these experiences in Hiroshima and Nagasaki, and conclude with a discussion of some of the unanswered questions predicted by these results.

Radiation Znjuiy and the Chernobyl Catastrophe. STEM CELLS 1997;1S(suppl2):129-133 0 1997 AlphaMed Press. All rights reserved.

130 Brain Damage Among the Prenatally Exposed

CLINICAL AND SOCIAL CONSEQUENCES OF PRENATAL RADIATION EXPOSURE We have followed a group of approximately 1,600 individuals. Aniong those we have seen 30 cases

of severe mental retardation, whereas our expectation would have been something on the order of 9 or 10. For the purposes of this study, an individual with severe mental retardation is defined as one who is unable to do simple arithmetic problems, form coherent sentences or to manage his or her own personal affairs, or who is sufficiently handicapped to require institutionalization. It is difficult to correlate this level of functioning with an equivalent intelligence quotient (IQ), but in most instances it is certainly well below 70. Thirty cases of severe mental retardation would not seem a large number, especially in com- parison to the number of cases of cancer that have occurred. However, the poignancy of those events transcends the smallness of the numbers.

Even before the studies in Hiroshima and Nagasaki began, it was apparent that impairment of mental function might be one of the consequences of prenatal exposure. The basis for that supposition began with work performed in the U.S. in the late 1920s by Goldstein and Murphy, 1929 and Murphy, 1947, both of whom were professors of Obstetrics and Gynecology at the University of Pennsylvania [ 1,2]. They solicit- ed responses from a large number of practicing obstetricians in the U S . to determine how many instances of severe mental retardation they had seen after treatment with radium of pregnant women. The number that they saw was distressing. Prior to their work, which was really the first systematic study, there had been anecdotal accounts and case reports of severely retarded children born to mothers who had received radio- therapy, but it was difficult from that happenstance literature to really know how profound the problem was.

It is surprising, in a sense, that occurrence of retardation was not more frequent because in the 1910s and early 1920s, ionizing radiation was actually being used as a means of therapeutic abortion. The desired interruption did not occur in a substantial number of cases, leading to full-term pregnancies, and often catastrophic consequences for the resulting child.

When we analyzed the data on maternal uterine dose from Hiroshima and Nagasaki combined, we reasoned that ionizing radiation could impinge only upon those embryological events that were actually occurring at the time of exposure. This led us to suppose that mental retardation would not be a risk uniformly distributed across all ages of gestation and that, in fact, the preponderance (over 80% of the cases) occurred in the window of time from 8 to 15 weeks after fertilization [3]. At this time two major events are occurring in the developing brain. It is the period of the most rapid pro- duction of cells destined to populate the cerebral cortex, and it is the period when these cells migrate from their places of origin, in the circumferential proliferative zones, to the cortex of the brain itself. Although to some extent some of these fundamental events extend into the next period, i.e., 16-25 weeks, this is much more a period of rapid synaptogenesis than it is a period of populating the cere- bral cortex. These landmarks refer only to the cortex and not to the cerebellum, where the events are somewhat different in time.

The concentration of cases of mental retardation in this window (8-15 weeks) led us to suppose that there might be less dramatic effects or impairment of cortical function, and that these effects might manifest themselves in other ways that would be amenable to measurement. We learned that, in fact, some of the early investigators with the Atomic Bomb Casualty Commission did have suffi- cient foresight to make those measurements. We therefore turned to information on IQs that had been accumulated largely in the year 1955 [4]. This does not necessarily suggest that an intelligence test actually measures intelligence; nevertheless, whatever it is measuring is the same thing in all age groups and in all dose groups.

We compared two different dosimetries during the period fi-om 0 to 7 weeks, where a priori one might argue that we would not necessarily expect effects because proliferative cells have not yet cast their des- tiny, as it were, and are not yet neuronal cells. Neuronal cells do not multiply. In theory, therefore, in this very early period there was the prospect of repopulation. However, there was a very sharp decline in the group exposed at 8-15 weeks, a somewhat lesser decline in the period of 16-25 weeks, and no evidence of

Schull 131

a decline whatsoever beyond the 26th week [4]. Again, using this measure, totally distinct from the clinical recognition of severe mental retardation, we identified the same two periods of vulnerability (even when one excludes the children who were clinically recognized to be mentally retarded).

In 1955, at the time that these measurements were being made, the school authorities in Hiroshima and Nagasaki were approached to see whether they would permit us to copy school performance records. At this time these children were about 10-1 1 years old, and most of them had completed the first four years of school. In a Japanese school at that time, as today, a student was exposed to seven different sub- jects and was graded independently in each of these subjects. Children were assigned grades of -2 to +2. Minus two implied that they were in the lowest five percentile of the class, -1 represented the next low- est 20 percentile, zero was the middle 50 percentile, and equivalent scores were used on the positive side. Since there were seven different subjects, we elected simply to combine the grades for each child. We pooled all of the individual scores on the supposition that the mean would satisfy some of the assump- tions inherent in the test of significance more than examining the individual scores. When this was done, the school performances of these children revealed the same sort of impairment that we saw with regard to the IQ test which had been conducted by trained psychometrists in the clinical fac Commission [5,6]. In the schools, there were four different teachers grading the children in each of their four years; these teachers were unfamiliar with the prenatal exposure of these children to radiation.

These results led us to suspect that there might be other forms of damage that had not been consid- ered. One of the obvious ones, since by this time we were beginning to understand what we thought might be one of the mechanisms, was evidence of more profound cortical dysfunction manifesting as seizures. Seizures can be precipitated by febrile episodes, by post-vaccinia reactions and by a variety of causes not directly related to exposure to ionizing radiation. Consequently we examined all seizures, but the group that we focused on primarily was the so-called unprovoked group (i.e., with seizures that could not be explained by trauma, febrile episode or other factors) [7]. As far as we could ascertain from talk- ing with their parents, there was no precipitating cause for these seizures other than the exposure of the child to ionizing radiation.

We found that at 1 Gy, severe mental retardation is increased by 50-fold. Intelligence tests among those not described as mentally retarded show a fall of almost 30 IQ points. Thus, an individual who would have been at the 100 value is now diagnosed as having borderline intelligence. The data on school performance essentially moved a child from the middle of his class to the lower five percentile. Seizures were increased 20-fold. In spite of our observations, several very important questions remain unan- swered. For example, the data are so sparse in some instances that we cannot describe the dose-response relationship with great reliability. There has been considerable concern expressed as to whether there might be a threshold in this dose-response relationship. In fact, the late Robin Mole has argued somewhat strongly that, at least with respect to severe mental retardation, there may be a threshold at 0.5 Gy[8-101. Personally, I am uncertain whether this threshold exists and would not accept a threshold of 0.5 Gy. Robin Mole was an outstanding radiation biologist, and it does indicate the range of opinions that can exist given the current limitations of the data. I believe that we will not resolve this question by any epi- demiological study. This leads to the second area of great interest, that is, what are the molecular and cellular events that are impaired by exposure to ionizing radiation?

CELLULAR CONSEQUENCES OF PRENATAL RADIATION EXPOSURE

Until relatively recently we had little information as to what structural impairments might result from prenatal exposure to ionizing radiation. Until 1986, we had information from autopsies of only four individ- uals in this situation. Of those four, only one had received a dose in excess of 1 cGy. The two with normal intelligence had typical brains with normal weight and appearance. Of the two individuals who were diag- nosed as retarded, one was a woman who died in her early 20s who had received about 1 cGy of ionizing radiation in the 31st to 32nd week of her gestation. The brain was small, weighing 1,000 g (about two-thirds

132 Brain Damage Among the Prenatally Exposed

of the weight that one would anticipate), but otherwise structurally normal. The other case was a young man who died when he was 16 years old of a fulminating viral encephalitis. He had been exposed in Nagasaki to a dose of a little less than 1.5 Gy. There were massive areas of ectopia surrounding the ventricles. It was clear that a very substantial number of the neurons that this young man had produced during his gestation had never left the proliferative zones. We know that they had to be functional and that they had established some form of connection, or they would not be visible at all. We now know from an embryological standpoint that neuronal cells failing to establish connections die and are cleared from the brain [ 1 I]. In the case of the 16- year-old, those cells had made connections although they were obviously not appropriate. The child had a severe behavioral problem from the earliest years, with sexual aberrancy being one of the common dysfunc- tional states. He had bilateral radiation cataracts and bilateral adrenal hypoplasia, which may have also con- tributed to the peculiarities of his behavior. One could not, however, make any compelling argument about migration being an important phenomenon on the basis of a single case.

Fortunately, the development of magnetic resonance imaging has allowed us to examine a number of living brains among individuals exposed prenatally. These studies have confirmed the impression that there is an impairment of cell migration [12].

While these studies made it more convincing that cellular migration was involved in radiation- induced brain damage, they did not directly address the question of what was transpiring at low doses. For this we turned to experimental systems, and we are now investigating what occurs in the rat brain when it is exposed to doses of 5, 10, 15 and 20 cGy at the equivalent stage in develop- ment. These experiments have demonstrated that there is a steady dose-dependent diminution in the number of cells that have moved out of the proliferative zone [13]. We can duplicate this through either in vivo experiments or in vitro experiments, i.e., by labeling the animal while still alive, sacrificing the animal, taking pieces of brain tissue that can be cultured, labeling those and allowing them to proceed for approximately two divisions after the labeling has occurred. The lin- ear trend in retardation of migration is significant, implying that even at doses as low as 5 cGy something is happening in these brains.

The next question is why is there an impairment in migration? Perhaps it is because of an alter- ation in the site of skeletal proteins, since we know that migration involves not only the shape of the cell but also its capacity to relate to the pathways over which it migrates. Therefore, not only are surface phenomena important, but also intracellular phenomena involving cytoskeletal pro- teins. At doses as low as 10 cGy we cannot show any impairment to surface proteins or cytoskele- tal proteins that are expressed only in neuronal cells (i.e., those that have cast their fate and will not divide any longer), using any of the known monoclonal antibodies. However, when we look at the neuronal cell adhesion molecule (N-CAM), which is a ubiquitous cell marker that is also extremely important in the early stages of migration, there is an easily recognizable qualitative impairment of expression of N-CAM at doses as low as 10 cGy [13]. In other words, we find inhi- bition of the molecule that is very critical for allowing the neuronal cells to migrate. Thus, one of the proteins that is central to the migratory phenomenon has been shown to be impaired at rela- tively low doses of ionizing radiation. Surprisingly, however, those cells recovered their capacity to express N-CAM approximately 48 h later.

These findings were not entirely unexpected since Ludwig Feinendegen has found very similar changes [14, 151. The transient inhibition of the functions of hematopoietic stem cells at doses as low as 1 cGy does not explain why these cells recover. We are now interested in attempting to deter- mine whether low doses of ionizing radiation cause some impairment of gap junction function which affects calcium flux, an extremely important phenomenon in the brain. If those gap junctions are inhibited only for a period of time, we would like to determine if either new ones are established or the old ones are reactivated. We may be able to explore this question because of the availability of monoclonal antibodies that appear to be specific for gap junctions.

Schull 133

CONCLUSIONS In this review, I have attempted to indicate important future directions, not only for these studies, but

also for virtually all of the issues that are discussed in this symposium. Resolution of these questions will not, in my view, come from epidemiological studies alone. It will be essential to develop better interactions between the experimentalist and the epidemiologist than have occurred in the past, since the only prospects for understanding what has actually transpired is to understand the molecular mechanisms involved.

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