Chromosomes were first seen and named in the nineteenth century. Chromosome is a combination of Greek words meaning colored body. It was early appreciated that these brightly staining objects appearing in the cell nucleus must be the “stuff of heredity,” the very vessels of our genetic inheritance. But not until 1956, following a serendipitous laboratory error (cells washed in an overly hypotonic solution) that had a fortunate technical consequence, was it possible to get a clear look at human chromosomes, and Hsu (1979) provides a personal perspective of these events. Three years later came the first demonstration of a medical application, with Dr. Jérôme Lejeune's discovery of the extra chromosome in Down syndrome (Lejeune et al., 1959), and this was followed shortly by the recognition of the other major aneuploidy syndromes. Thereafter, medical cytogenetics evolved into a practice in its own right, and it is now a well-established and mainstream medical and laboratory discipline. These brief facts are well rounded out in Gersen (1999).
“Colored bodies” has become an especially apt derivation with the development of newer staining techniques. We can now see different parts of chromosomes in many different colors, whether true or computer-generated false colors. The images produced by this kaleidoscopic karyotyping can be rather beautiful. Black and white photographs are less splendid but usually suffice (Fig. 1-1). If microarray technology takes over, “disembodied colors” might be a more accurate expression, but the word chromosome will surely last forever.
Chromosomal Morphology
Chromosomes have a linear appearance: two arms are continuous at the centromere. Reflecting the French influence in the establishment of the cytogenetic nomenclature, the shorter arm is designated p (for petit) and the longer is q (next letter in the alphabet). In the early part of the cell cycle, each chromosome is present as a single structure, a chromatid, a single DNA molecule. During the cell cycle (Fig. 1-2) the chromosomes replicate, and two sister chromatidsform. Now the chromosome exists as a double-chromatid entity. Each chromatid contains exactly the same genetic material. This replication is in preparation for cell division so that, after the chromosome has separated into its two component chromatids, each daughter cell receives the full amount of genetic material. It is during mitosis that the chromosomes contract and become readily distinguishable. (At other times in the cell cycle, chromosomes are attenuated and not visible.)
Routine cytogenetic analysis is done on mitotic cells, usually obtained from blood. Blood lymphocytes have two convenient properties for the cytogeneticist: they are easily obtained, and they are easily stimulated to go into mitosis. The chromosomes of the small number of lymphocytes studied are taken as representative of the chromosomal constitution of (essentially) every other cell of the body. In the case of prenatal diagnosis, the cells from amniotic fluid or chorionic villi are likewise assumed (with certain caveats) to represent the fetal chromosomal constitution.
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Figure 1-1. Chromosomes as they appear viewed through the microscope. |
The 46 chromosomes come in 23 matching pairs, and constitute the genome. One of each pair came from the mother, and one from the father. For 22 of the chromosome pairs, each member (each homolog) has the same morphology in each sex: these are the autosomes. The sex chromosome (gonosome) constitution differs: the female has a pair of X chromosomes, and the male has an X and a Y chromosome. The single set of homologs—one of each autosome plus one sex chromosome—is the haploid set. The haploid number (n) is 23. The haploid complement exists, as such, only in the gametocytes (ovum and sperm). All other cells in the body—the soma—have a double set: the diploid complement (2n) of 46. If there is a difference between a pair of homologs, in the sense of one being structurally rearranged, the person is described as a heterozygote.
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Figure 1-2. Outline of chromosome activity during the mitotic cycle. |
The chromosomes are distinguishable on the basis of their size, centromere position, and banding pattern. The centromere may be in the middle, off-center, or close to one end—meta-centric, submetacentric, and acrocentric, respectively. Originally the chromosomes were assigned to groups A through G according to their general size and the position of the centromere. With banding, each chromosome is individually distinguishable. The diagrammatic representation of the banding pattern is the ideogram (Appendix A). The autosomes are numbered from largest to smallest, no. 1 through 22 (to split hairs, this order is not exact; for example, chromosomes 10 and 11 are shorter than chromosome 12, and chromosome 21 is smaller than 22). Certain parts of some chromosomes may show variation in the population. Increasing precision in banding (highresolution banding) permits progressively more subtle definition of the chromosome (Fig. 1-3). Chromosomes are conventionally displayed “cut and paste” and arranged with p arms upwards, in their matching pairs. This is a karyotype (Fig. 1-4). The word karyotype is also used in the general sense of chromosomal constitution. Cytogeneticists describe karyotypes with a shorthand notation, the International System for Human Cytogenetic Nomenclature (ISCN, 1995); an outline is given in Appendix B.
Chromosomal Structure and Function
The two chemical components of chromatin are DNA and protein. Proteins provide the scaffolding of the chromosome, and are divided into histone and non-histone proteins. Histones are strongly conserved DNA-associated proteins; the fact that they differ little between species such as ourselves and the sweet pea (for example) indicates how fundamentally important their role is in maintaining the integrity of the chromosome. Chromatin exists in differently condensed forms: the less condensed euchromatin and the more condensed heterochromatin. Euchromatin contains the coding DNA—the genes—while heterochromatin comprises non-coding DNA. Chromosomes are capped at the terminal extremities of their long and short arms by telomeres, specialized DNA sequences comprising many repeats of the sequence TTAGGG, that can be thought of as sealing the chromatin and preventing its fusion with the chromatin of other chromosomes. The centromere is a specialized region of DNA that, at mitosis, provides the site at which the spindle apparatus can be anchored and draw each separated chromatid to opposite poles of the dividing cell. Centromeric heterochromatin contains “satellite DNA,” so-called because these DNA species have different buoyant densities and produce distinct humps on a density gradient distribution. (These are not to be confused with the satellites on acrocentric chromosomes.) A separate issue is the “packaging question”: how the centimeters of DNA are compacted into micron-length chromosomes. The presently preferred model is that the chromatin fibers are thrown into loops extending outward from a backbone, this backbone being formed as adhesive sites dotted along the fibers come together and construct a continuous linear stack.
Miller and Therman (2001) treat this question in detail.
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Figure 1-3. Increasing resolution of banding (chromosome 11). (Courtesy D. R. Romain.) |
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Figure 1-4. Chromosomes arranged in formal karyotype. |
CHROMOSOME ABNORMALITY
Chromosomes are distributed to each daughter cell during cell division in a very precise process. Yet it is prone to error. From our perspective, the two cell divisions of meiosis, during which the gametes are formed, are of central importance. Most of the discipline of medical cytogenetics focuses on the consequences of disordered meiosis that produce a chromosomally abnormal gamete, causing a chromosomal abnormality in the conceptus. A chromosome abnormality that is present from conception and involves the entire body is a constitutional abnormality. If an additional cell line with a different chromosomal complement arises before the basis of the body structure is formed (i.e., in embryonic or pre-embryonic life) and becomes an integral part of the organism, constitutional mosaicism results. In this book we are concerned almost solely with constitutional abnormalities. Acquired chromosomal abnormality, of course, exists, and indeed it is a major initiating and sustaining cause in most cancers, a fact voluminously attested by the work of Mitelman et al. (2002), but this is more the field of study of the cytomolecular pathologist than the genetic counselor.
An incorrect amount of genetic material carried by the conceptus disturbs and distorts its normal growth pattern (from zygote → blastocyst → embryo → fetus). In trisomy there are three of a particular chromosome, instead of the normal two. In monosomy only one member of the pair is present. Two of each is the only combination that works properly! It is scarcely surprising that a process as exquisitely complex as the development of the human form should be vulnerable to a confused outflow of genetic instruction from a nucleus with a redundant or incomplete database.
Trisomy and monosomy for a whole chromosome were the first cytogenetic mechanisms leading to an abnormal phenotype to be identified. More fully, we can list the following pathogenetic mechanisms that arise from chromosomal abnormalities:
1. A dosage effect, with a lack (deletion [del]) or excess (duplication [dup]) of chromosomal material, whether for a whole chromosome or a part of a chromosome
2. A direct damaging effect, with disruption of a gene at the breakpoint of a rearrangement
3. An effect due to the incongruent parental origin of a chromosome or chromosomal segment (genomic imprinting)
4. A position effect, whereby a gene in a new chromosomal environment functions inappropriately.
We discuss these mechanisms in more detail in following chapters.
Autosomal Imbalance
Structural Imbalance
As noted above, imbalance may involve the gain or loss of a whole chromosome—full ane-uploidy—or of part of a chromosome—partial aneuploidy. The abnormality may occur in the nonmosaic or mosaic state. Loss (i.e., monosomy) of chromosomal material generally has a more devastating effect on growth of the conceptus than does an excess of material (i.e., trisomy). Certain imbalances lead to certain abnormal phenotypes. The spectrum is listed in outline form in Table 1-1 and in more detail in Table 1-2. Most full autosomal trisomies and virtually all full autosomal monosomies set development of the conceptus so awry that, sooner or later, abortion occurs—the embryo “self-destructs” and is expelled from the uterus. This issue is further explored in Chapter 21. A few full trisomies are not necessarily lethal in utero, and many partial chromosomal aneuploidies are associated with survival through to the birth of an infant.
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Table 1.1. Spectrum of Effects Resulting from Constitutional Chromosomal Abnormality |
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Characteristically, “survivable imbalances” produce a phenotype of widespread dysmorphogenesis, and there may be malformation of internal organs and limbs. It is often in the facial appearance (facies) that the most specific physical abnormality is seen. The most complex organ of all, the brain, is the most vulnerable to a less than optimal genetic constitution; and some compromise of mental and intellectual functioning, usually to the extent of an obvious deficit, is nearly invariable. Advances in cytogenetic technology have allowed the detection of subtle deletions and duplications, and while the physical phenotype in many of these cases may be rather bland, compromise of neurological functioning is typical (Curry et al., 1997). Thus, the central concern of most people seeking genetic counseling for a chromosomal condition is the fear of having a physically and mentally handicapped child.
Functional Imbalance
A correct amount of chromatin does not necessarily mean that the phenotype will be normal. Inappropriate inactivation, or activation, of a segment of the genome can distort the genetic message. Some segments of the genome require only monosomic expression, and the homologous segment on the other chromosome is inactivated. If this control fails, both segments can become activated, or both inactivated; the over- or underexpression of the contained loci can cause phenotypic abnormality. The classic example of this is genomic imprinting according to parent of origin, which is discussed in Chapters 2 and 20. A rather specialized example arises with the X-autosome translocation. A segment of X chromosome can fail to be inactivated, or X-inactivation can spread into an autosomal segment (Chapter 5).
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Table 1.2. Impact of Constitutional Chromosomal Abnormality on Human Mortality and Morbidity |
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Sex Chromosomal Abnormality
Sex chromosome (gonosome) imbalance has a much less deleterious effect on the phenotype than does autosomal aneuploidy. In both male and female one, and only one, completely active X chromosome is needed. X chromosomes in excess of one are almost wholly inactivated, as the normal 46,XX female exemplifies. With X chromosome excess or deficiency, a partially successful buffering mechanism exists in which the imbalance is counteracted in an attempt to achieve the effect of a single active X. Excess whole X chromosomes are inactivated; abnormal X chromosomes are selectively inactivated to leave the normal X as the active one; and if in the female one X is missing, the single X remaining is not subject to inactivation. In X imbalance, the reproductive tract and brain are the organs predominantly affected. The effect may be minimal. As for Y chromosome excess, there is a rather limited phenotypic consequence, because this chromosome is composed largely of constitutively inactive material.
The fragile X syndromes concern, obviously enough, a sex chromosome. They began life, as their names attest, as cytogenetic abnormalities, but are now seen largely as single gene disorders, testable by molecular methodology. Nevertheless, their cytogenetic patrimony is necessary knowledge for the counselor, and it remains perfectly appropriate that we continue to include the fragile X syndromes in this book.
Frequency and Impact of Cytogenetic Pathology
According to the window of observation, chromosomal disorders make a greater or lesser contribution to human mortality and morbidity. Looking at prenatal existence, chromosomal mortality is very high, and aneuploidy is the major single cause of spontaneous abortion. Perinatal and early infant death has a significant chromosomal component, of which trisomies 18 and 21 are major elements. As for morbidity, chromosomal defects are the basis of a substantial fraction of all intellectual deficit, and many of these retarded individuals will also have structural malformations that cause functional physical disability. Among a mentally retarded population, Down syndrome is the predominant contributor in the fraction who have a chromosome abnormality, while the increasing ability to pick up subtle deletions puts this category in second place (Fig. 1-5). During adolescence, many sex chromosome defects come to light, when pubertal change fails to occur; and in young adulthood, chromosomal causes of infertility are recognized. Few new cytogenetic defects come to attention later in adult life, but many retarded children survive well into adulthood and some into old age, and some require lifelong care from their families or from the state. This latter group imposes a considerable emotional and financial burden. While some parents and caregivers declare the emotional return they have from looking after these individuals, for others this responsibility is a source of continuing, unresolved grief.
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Figure 1-5. The relative proportions of different cytogenetic categories in 835 karyotypically abnormal individuals of a mentally retarded population in South Carolina studied in 1989–1994 (Phelan et al., 1996). If the exercise were to be repeated now, a lesser fraction due to Down syndrome might possibly be expected, and a new category would be needed for subtelomeric and other small deletions not detectable on classical cytogenetics. (Courtesy M. C. Phelan, and with the permission of the Greenwood Genetic Center.) |
Hook (1992) has summarized the categories of cytogenetic pathology and their impact, and we have reproduced his synopsis in Table 1-2. In Table 1-3 we set out the birth incidence of the various categories of chromosomal abnormality. Overall, around 1 in 120 liveborn babies has a chromosomal abnormality, and about half of these are phenotypically abnormal due to the chromosome defect. If we were to look at 5-day blastocysts, the fraction might be close to a half. If we studied a population of 70-year-olds, we could expect to see very few individuals with an unbalanced autosomal karyotype. The finer the cytogenetic focus, the greater the incidence: with the highest resolution banding and the application of molecular methodologies, some previously unrecognized defects would be included.
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Table 1.3. Frequency of Chromosome Abnormality in Newborns |
P.10
Research Application of Cytogenetic Pathology. The phenotypes that result from chromosome abnormalities can point the way to discovery of the causative genes. An early example of deletion mapping was the recognition that the gene for retinoblastoma is on chromosome 13, given the association of this cancer with the 13q syndrome. Another cancer gene to be similarly mapped was APC (adenomatous polyposis coli), following the observation of polyposis in a retarded del(5)(q22q23) individual. The triple dose of chromosome 21 in Down syndrome was a signpost on the way to finding the β-amyloid precursor protein (APP) gene as one of the loci for Alzheimer's disease. A translocation with one breakpoint at a particular spot on the long arm of chromosome 17 in an individual with neurofibromatosis-1 provided an entrée to cloning the NF1 gene (Ledbetter et al., 1989). Some of these translocations may turn out to be false leads, such as Brett et al.'s (1996) report of a 3;8 translocation in a family with Gilles de la Tourette syndrome, a neuropsychological disorder of multiple tics and obsessive-compulsive behavior. Initially, there seemed suggestive evidence that a gene for this disorder might have mapped to 3p21.1; but with further analysis and the discovery that some affected family members did not have the translocation, the case collapsed. We have conducted a review of chromosomal conditions in which epilepsy is a feature, with the aim of providing leads to epilepsy genes (Singh et al., 2002a).
It is a general principle that many important scientific discoveries are made serendipitously; or, stated differently, “opportunity comes to the prepared mind.” Voullaire et al. (1993) identified an extra structurally abnormal chromosome (ESAC), in a child with a nonspecific picture of physical abnormality and intellectual deficit, which had no C-band positive centromere (only a constriction). Conventional wisdom has it that a chromosome cannot be stably transmitted at cell division if it has no centromere. These workers studied this ESAC, and discovered that it did have a simple, but functional, centromere. This observation led the way to the delineation of the neocentromere (p. 273). This elemental structure could beused as the basis for designing an artificial centromere, a necessary component of a human artificial chromosome (HAC). HACs have a potential medical role as vectors of therapeutic genes.
LOOKING AT CHROMOSOMES
Chromosomes are analyzed in the cytogenetics laboratory under the light microscope at a magnification of about 1000×. The chromosomes must be stained to be visible, and a great many staining techniques are available to demonstrate different features of the chromosome. We list some of these, in particular those with a more immediate practical application to the clinical issues we discuss in this book.
1. Plain staining (“solid staining”). Many histologic dyes, including Giemsa, orcein, and Leishman, stain chromosomes uniformly. Until the early 1970s these were the only stains available.
2. Giemsa or G-banding. This is the main staining method in use in routine clinical cytogenetics. It allows for precise identification of every chromosome and for the detection and delineation of structural abnormalities. At the 400–550 band level, rearrangements down to about 5 mega-bases (Mb) in length can be discerned, at least in regions where the banding pattern is distinctive. Its precision is increased by manipulations designed to arrest the chromosome in its more elongated state at early metaphase or prometaphase—high-resolution banding. Alternative methods to demonstrate essentially the same morphology are quinacrine or Q-banding and reverse or R-banding. In R-banded chromosomes the pale staining regions seen in G-banding stain darkly, and vice versa.
3. Centromere or C-banding. This technique stains constitutive heterochroma-tin—the centromeric heterochromatin, some of the material on the short arms of the acrocentric chromosomes, and the distal part of the long arm of the Y chromosome. Constitutive heterochromatin, by definition, has no direct phenotypic effect.
4. Replication banding. This technique is used primarily to identify inactive X chromatin. A nucleotide analog (BrdU) is added either as a pulse at the beginning or toward the end of the cell cycle, to allow the cytogenetic distinction of chromatin that replicates early from that which replicates late. It produces a banding pattern similar to that of R-banding.
5. NOR (silver) staining. The nucleolar organizing regions (NOR) contain multiple copies of genes coding for rRNA and are sited on the satellite stalks of the acrocentric chromosomes. Those that actively produce rRNA stain black with silver nitrate. Silver (Ag) staining can be used in characterization of small marker chromosomes and in translocations involving the short arms of the acrocentric chromosomes. This technique has now been largely been replaced by fluorescence in situ hybridization (FISH) using a probe that hybridizes to short arm sequences on all the acrocentric chromosomes.
6. Distamycin A/DAPI staining. This fluorescent stain identifies the heterochromatin of chromosomes 1, 9, 15, 16, and Y. A particular use is in distinguishing theinverted duplication 15 chromosome (p. 273) from other small marker chromosomes.
7. Fluorescence in situ hybridization (FISH), and variations. The major cytogenetic advance of the 1990s was the ability to identify specific chromosomes and parts of chromosomes by in situ hybridization with labeled probes. This technique is widely used to detect submicroscopic deletions and to characterize more obvious chromosome anomalies. The hybridization method may be direct or indirect. Direct attachment of a detectable molecule (e.g., a fluorophore) to the probe DNA enables its microscopic visualization immediately after its hybridization to the target DNA in the chromosome. The more sensitive indirect procedure requires special modification of the probe with a hapten detectable by affinity cytochemistry. The most popular systems are the biotin-avidin and digoxigenin systems. By using combinations of biotin-, digoxigenin-, and fluorophorlabeled probes, multiple simultaneous hybridizations can be done to locate different chromosomal regions in one preparation (multicolor FISH).
8. Submicroscopic telomeric analysis. The subtelomeric regions are, in general, gene-rich, and very small rearrangements in these regions can be of profound effect. Probes have been developed for targeted subtelomeric FISH that may identify subtle rearrangements not detectable on routine banded analysis, and commercial kits are available (Knight and Flint, 2000).
9. Comparative genomic hybridization (CGH). Here, fluorophore-tagged DNA from the patient is applied to a metaphase slide prepared from a “standard” normal person. It has been described as a sort of “FISH in reverse.” Relative excesses and deficiencies of patient DNA bind differentially to the reference chromosomes, and yield different colors on exciting the fluorophores. This procedure has even been applied successfully to archival pathology material. It may have application in preimplantation genetic diagnosis, where it offers the advantage of testing the whole karyotype. High-resolution CGH refers not to a more stretched chromosome preparation but to a further level of sophistication of the computer software that is used to analyze the images by adjusting for the idiosyncratic patterns that each homolog may have. Very small imbalances may be identifiable by this approach, and the nature of uncertain rearrangements clarified (Knight and Flint, 2000; Kirchhoff et al., 2001; Ness et al., 2002).
Chromosomes examined by various techniques are illustrated in Figure 1-6. Full detail of these techniques can be found in Keagle and Gersen (1999), Blancato (1999), Mark (2000), and Miller and Therman (2001), while Trask (2002) provides a historical span of the cytogeneticist's skill.
Non-cytogenetic Methods
While these new techniques are largely in the research sphere, as at the present writing, cytogeneticists will certainly want to maintain a watching brief. A chromosome report of the future may resemble the graph in Figure 1-7, or at any rate be based on this sort of raw data. Whatever techniques come to be used, the fundamental purpose of the cytogenetic report will, of course, remain the same. Descriptions about the technologies used will be important addenda to reports, as they may inform the clinician about the intensity of chromosome analysis undertaken and the need for further possible analysis.
Microarray Analysis. Of all the new procedures for examining chromosomes, microarray analysis is the new technique most likely to supplant classical cytogenetics. The fundamental principle is essentially the same as in CGH, noted above; indeed, this process is a comparative genomic hybridization, using the array, rather than the metaphase spread, as substrate. The actual microarray comprises thousands of spots of reference DNA sequences, applied in a precisely grided manner on a slide (or “chip”). Patient and control DNA are applied to the slide, and hybridization takes place. Excess chromatin conventionally gives a red color, deficient hybridization gives green, and an even amount of hybridization is seen as yellow (produced from an overlapping of equal amounts of red and green). Red therefore signals duplication, green signals deletion, and yellow denotes normal segments. The pattern of red, yellow, and green spots on the microarray is captured and subject to computer analysis. An array with 3000 spots could detect unbalanced rearrangements at a 1 Mb resolution across the entire genome (Snijders et al., 2000); and, as a start, a chromosome 22 array has been prepared (Buckley et al., 2002). The balanced state of rearranged chromosomes is not detected.
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Figure 1-6. Chromosome pairs 1, 6, 15, 16, and Y and X stained by various techniques: plain stain (a), G-banding (b), replication banding (c), C-banding (d), Ag-NOR stain (e), and Q-banding (f). |
Molecular Analysis of Telomeric Sequences. Since telomeric imbalances may be relatively frequent, and often undetectable even by high-resolution cytogenetics, approaches have been devised to assess these regions by DNA-based methodologies. They have the particular advantages that all subtelomeric regions are assessed in a single test and several samples can be run together as a batch. Two techniques are mentioned here, one quantitative, the other qualitative; no doubt further variations on these themes will be developed. In multiplex amplifiable probe hybridization (MAPH), genomic DNA is analyzed with a set of small (140–600 bp) probes for subtelomeric sequences from every chromosome, and the degree of amplification is quantified (Hollox et al., 2002). Greater or lesser amplification indicates a duplication or deletion on that particular chromosome arm. Microsatellite transmission analysis might be a name to apply to the method reported in Colleaux et al. (2001). Polymorphic sequences in the subtelomeric regions are genotyped, and aberrant inheritance—one parent transmitting other than one allele—is directly observed on an electropherogram plot.
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Figure 1-7. The style of a cytogenetic study of the future? In microarray-based comparative genomic hybridization (CGH), probes are used for subtelomeric regions, here on a child with a del(18)(q21.2). T/R, test/reference ratio. The two points below the lower threshold (a T/R of 0.8) in the no. 18 column reflect a reduced amount of DNA from that part of the genome which is recognized by the two 18q-derived probes. (From J. A.Veltman et al., Am. J. Hum. Genet., 70, 1269–1276, 2002, courtesy J. A. Veltman, © The American Society of Human Genetics, and with the permission of the University of Chicago Press.) |
ETHICAL AND COUNSELING ISSUES
Our focus in this book is on the biology of chromosomal defects and the reproductive risks they may entail. Certain bioethical issues, coming to be more formally defined in the late twentieth century do, however, demand attention. Counselors must hold fast to the following requirements: (1) that they act beneficently toward their clients,1 and (2) that they strive to make their services accessible to those who may need them.
Guilt in a Carrier
Sometimes a chromosomal diagnosis is made in an older child or even an adult, when the parents have held for years to the notion that obstetric misadventure, or a virus, or some other blameable event was the cause of the child's condition. Some people find it upsetting to have to readjust, and to know that they may have been the source of the abnormality. They are likely to use words like “guilt,” “blame,” and “fault.” Helping these people to adjust to the new knowledge is a challenge for the counselor.
Nondirective Counseling
In a Western ethos, the counselor is required to respect the autonomy of the client, and this largely translates into the principle that counseling be nondirective. Counseling may in fact never be truly nondirective, and we need to have an awareness of our own biases so that our advice will be valid, as seen by those to whom we give it. Rentmeester (2001) comments that, since it is “impossible for human language to convey facts purely, without any spoor of values”, and since “risk cannot be appreciated without consideration of values”, it is neither helpful nor possible to try to be value-neutral. There is a fine line between directive and detached counseling, a point nicely illustrated in Karp's (1983) deft essay, “The Terrible Question” (required reading for every counselor). Ingelfinger (1980) comments, admittedly in a somewhat different context: “A physician who merely spreads an array of vendibles in front of the patient and then says, ‘Go ahead and choose, it's your life,’ is guilty of shirking his duty, if not of malpractice.” Rentmeester offers the refreshing advice that it is not necessarily unprofessional to answer a patient's question: ‘What would you do?’. It is the skill of the counselor that helps clients reach the decision that is right for them and feel satisfied that they have done so. The subtleties and complexities of attempting to be nondirective in the setting of a prenatal diagnosis clinic are discussed by Anderson (1999), who analyzes responses of couples who did or who did not choose to have testing. She emphasizes the wide range of beliefs and values that people can have, and the likelihood for failed communication if these differences are not appreciated.
In some other societies, the perceived good of the group may carry more weight than the professed wishes of the individual. The degree to which one society can seek to influence practice in another is a matter of some controversy, well illustrated by the response in the West to the “eugenic” Chinese Maternal and Infant Health Care Law of 1994 (Lancet editorial, 1995). The subtleties of the issue have led to keenly pointed argument (correspondence in the American Journal of Human Genetics, 65, 1197–1201, 1999). Knoppers (1998) comments on the subtle boundary between the need to respect cultural, religious, and social diversity, and the imperative to adhere to tenets of generally accepted rights and ethics. More provocatively, she points to a “political and moral one-upmanship” which has colored the argument, and which may confuse deciding between what is “immoral state policy or just plain common sense.”
Mental Retardation and Genetic Abortion
Intellectual deficiency is a condition for which many parents are unwilling to accept a significant recurrence risk; this is hardly remarkable, since intellectual function is such an obvious attribute of humanness. The great majority of those who chose to have prenatal diagnosis opt for pregnancy termination if a chromosomal condition implying major mental defect is identified. In a French series of trisomy 21 diagnosed prenatally and postnatally over the period 1984–1990 in the Marseille district, all of the 76 cases of prenatal diagnosis were followed by abortion (Julian-Reynier et al., 1995). Some patients for whom abortion is not acceptable may nevertheless choose prenatal diagnosis for reassurance, or for the preparedness that certain knowledge can allow. Community views on mental handicap are changing and the late twentieth century has seen something of an exodus from institutions and from special schools, as the mentally and psychologically disabled join the mainstream, some more successfully than others. Counselors need to handle the tension inherent in these views and the views of parents who want to avoid having a handicapped child; and the separate conflict that parents experience when a decision is taken to terminate an otherwise wanted pregnancy. As we discuss above, the doctrine of nondirective counseling is a central tenet of modern practice; it is a test of counselors' professionalism that their own views not unduly influence the advice and counsel that they give. De Crespigny et al. (1998) document the experiences and comments of a number of couples in their book Prenatal Testing Making Choices in Pregnancy, intended for the lay public. Walters (1995) and Tillisch (2001) offer personal perspectives. First, Walters:
Defending the right of women who are carrying babies with Down's syndrome to have abortions is not pleasant. Anyone who does so is likely to sound heartless, especially if they have no first-hand experience. It is even harder for me. I am the father of a Down's syndrome baby…. It is the most painfulthing I will ever say but my wife Karen and I wish she had had a test. If she had, we would have terminated the pregnancy. I must be a callous swine, mustn't I? … Her birth was a tragedy, but not so different to any tragedy that can strike out of the blue, such as a crippling accident. Just as we work to avoid other tragedies, I see nothing wrong in using Down's tests to avoid the tragedy of human handicap…. I know that I would rather not haveexisted at all than to be, like her, sentenced to a life of confusion, frustration, pain and possibly loneliness when Karen and I are gone. If I feel guilt, it is that I was responsible for her birth. To me that guilt is far worse than anything I would have felt had I prevented it.
Tillisch is the mother of a child with the del(1)(p36) syndrome (p. 277). Anomalies had been detected on ultrasonography during the pregnancy, but an amniocentesis returned a normal cytogenetic result. The child had a stormy neonatal course, and in due course the chromosomal defect was identified. Tillisch writes:
I'm so thankful that the amniocentesis results were inaccurate. Since we didn't learn of Kasey's diagnosis until she was 9 months old, we were able to get to know, love, and admire Kasey as an individual, as our daughter. We didn't allow doctors to define her for us…. From a mother's perspective, Kasey's future is bright. She receives treatment and will soon go to a public school. We will allow Kasey to show us her potential, rather than labeling her “severely mentally retarded” and casting her off to be locked away from society…. My father once asked, if Icould ever make Kasey “whole”, would I? Without any hesitation, I answered: absolutely not. Adding the missing genes would make Kasey a different person, a stranger.
These differing, one could say polar, views of parents find some parallels in the positions of those whom we could consider as the philosophers of our profession. In a provocative address to the American Society of Human Genetics in 1970, Lejeune deplored the application of his original cytogenetic discovery to the prenatal diagnosis of Down syndrome. Epstein (2002) reflected, some three decades later, on Lejeune's influence, and while not stepping back from the standpoint that prenatal diagnosis is a proper and valid medical procedure, he does acknowledge (as must we) that a plurality of views exists, and that the genetics community must be sensitive to, and must respect, the range of views in the community.
Brock (1995) discusses the philosophy of “wrongful handicap,” addressing the esoteric issue of whether not producing a child who would suffer has harmed that potential child, and enunciates a principle that “individuals are morally required not to let any possible child for whose welfare they are responsible experience serious suffering or limited opportunity if they can act so that, without imposing substantial burdens or costs on themselves or others, any alternative possible child for whose welfare they would be responsible will not experience serious suffering or limited opportunity.” This position could be seen as providing an ethically based framework for making a decision to terminate an abnormal pregnancy and to conceive again.
Pregnancy and the Mentally Retarded
One issue that scarcely was an issue at the outset of the discipline, but which now looms quite significantly, is that of the rights of the intellectually handicapped to have children (Elkins et al., 1986a). What of the person with Down syndrome or some partial trisomy compatible with fertility in whom a question of procreation arises? Zühlke et al. (1994) give an example in describing a man with Down syndrome who developed a relationship with a mentally retarded girl living in the same house. She requested removal of an intrauterine contraceptive device, became pregnant, and the normal baby is being brought up by the maternal grandmother. On the one hand, the right of the handicapped person to experience parenthood is debated; and the American Academy of Pediatrics (1990) expressed reservation about the sterilization of intellectually handicapped women on the basis of anticipated hardship to others. On the other hand, Gillon (1987) notes that normal people have the option of being sterilized, and the mentally handicapped should have the same right. The Law Lords in Great Britain concur that sterilization may be in the best interest of the handicapped person herself (Brahams, 1987). Many parents or guardians, not wishing to become “parental grandparents,” favor sterilization. Some regard hysterectomy as having the double benefit of ensuring sterility and facilitating personal hygiene; others consider only reversible contraception to be acceptable. The High Court of Australia decided in 1992 that the parents of a handicapped child cannot themselves lawfully allow sterilization, but that a court authorization is required, and noted that this requirement “ensures a hearing from those experienced in different ways in the care of those with intellectual disability and from those with experience of the long term social and psychological effects of sterilization” (Monahan, 1992). Ten years later, it appeared that very few unlawful sterilizations of minors were being performed in the state of Victoria (Grover et al., 2002).
When a retarded woman with a chromosomal defect is pregnant, or is pregnant by a retarded man, one or other of the couple having an unbalanced karyotype, and the pregnancy is recognized in time, the grounds for termination are substantial. The ethical issue arises over the difficulty (or impossibility) of securing the woman's informed consent versus the expressed wishes of her guardians. Martínez et al. (1993) report from Alabama a mother with cri du chat syndrome who was severely retarded and had no speech, and was pregnant by an unknown male, and “although pregnancy termination had been desired by the patient's grandmother, social and legal limitations prevented access to this procedure.” Some less severely affected persons (if they are able to grasp the issue) may not regard it as undesirable to have a child like themselves; they may nonetheless have the insight to recognize their own deficiency and not wish to pass it on. We may perhaps read this into the brief report of Bobrow et al. (1992) of a man with Down syndrome fathering a child, and the mother seeking a first-trimester prenatal diagnosis (the baby was normal). There is the concept of imagining what a retarded person would want, were they intellectually competent to make a decision, a concept that some would regard as paternalistic (and infringing personal autonomy) and others see as valid. The sociology rather than the biology will exercise the coun-selor's mettle in this uncommonly encountered situation.
The other party involved is the child. Is having good parenting a right? What of a normal child born, say, to a man carrying a dup(10) (p13p14) chromosome and a mother with idiopathic mental defect? How can the interests of the child and those of the parents be resolved? This is an actual case that we have seen (Voullaire et al., 2000a): it was quite poignant as this mildly retarded man, who had some insight into his own handicap, struggled to understand how he might best be a father to his 46,XX baby, and expressed sadness at the abnormal behavior displayed by his older 46,XY,dup(10) child. The capable and willing grandmother stepped into the breach; but when the daughter is older, and assuming she is of normal intelligence, how will the realization of her parents' abnormality affect her? Whether a normal child in this sort of setting has a legal claim for “dissatisfied life” is an intriguing and as yet (to our knowledge) untested notion (Shaw, 1984).
Testing Children
To state the obvious, familial rearrangements are familial. It is very natural that parents would be concerned whether children they already have might be carriers, once an abnormality has been identified in one of them. Children certainly need to know their carrier status, sooner or later. It would be very unfortunate (and possibly create an exposure to legal redress) if a failure to transmit information led to another affected child unknowingly being born elsewhere in the family, as occurred in Burn et al.'s (1983) report of a family with a translocation that was the cause of cri du chat syndrome in two generations. Genetic counselors are attuned, however, to the principle of not taking away a child's right to make their own informed decision, in the fullness of time, to learn about genetic risks they may face, the principle being that the child's future autonomy is to be respected. The American Society of Human Genetics, and the American College of Medical Genetics (1995) has determined that “timely medical benefit to the child should be the primary justification for genetic testing in children and adolescents,” and it is true that a balanced chromosomal rearrangement will have no influence on a person's physical health, other than, in due course, their reproductive health (and the issue is thus to be seen in a different light than testing for adult-onset disease). Questions have been raised whether testing could damage a child's self-esteem, distort the family's perceptions of the child, and have adverse effects upon the child's capacity to form future relationships (Clarke et al., 1994).
Parents' views are also not without validity. Clayton (1995) comments that there is the possibility of conflict with parents, as physicians increasingly act as advocates for the child's interests, but notes further that “children are generally ill-served if their parents feel they have not been listened to”; she also draws the conclusion that this is a medicoethical rather than a medicolegal issue. McConkie-Rosell et al. (1999) sought opinions from a group of 65 parents of fragile X children attending a national conference in Portland, Oregon, in 1996. They noted a “strong belief in a parental right to make the decision regarding carrier status in their children,” with about half considering that they should have the right to decide when their child should be tested and informed of the result. The Genetic Interest Group in the United Kingdom gently chides the profession in commenting that “the vast majority of people are better able to understand the implications than they are often given credit for,” and has enunciated the following principle: “After suitable counseling, parents have the right to make an informed choice about whether or not to have their children tested for carrier status. Ideally, children should only be tested when of an age to be involved in the decision” (Dalby, 1995). It may be that earlier concerns overstated the potential for harm: at least with respect to the Mendelian cancer-predisposing syndrome familial adenomatous polyposis, children having undergone predictive testing and receiving a positive gene test result experienced no increase in anxiety, or depression or lack of self-esteem (Michie et al., 2001). Indeed, Robertson and Savulescu (2001) see potential benefit to the child, and support the view that, as a general rule, the parents' views should prevail, and a request for predictive testing be respected. There is also the practical point that many parents will have had a prenatal karyotype from amniocentesis or chorionic villus sampling; and it may not seem entirely logical to decline to test a postnatal child.
From this discussion, we conclude that a conservative stance, but not an immovable one, is appropriate. In debating the issue with them, many parents will see the wisdom of the declared position of the profession and be well satisfied (and possibly relieved) with the advice to leave testing until the child can decide. Equally, there will be occasions when acquiescence to a parental request may be reasonable. Either the parent's mind is set at rest, or they know of the need to raise the issue with the child at a suitable age, which should be with the assistance of the genetic counseling clinic. The task for the counselor is to assist parents in deciding what age would be suitable for their child, as “there is no universal ‘right’ age,” as McConkie-Rosell et al. (2002) comment with respect to fragile X carrier testing, and to convey the information in such a way that concern for the future is kept in perspective and the child's self-confidence is kept intact.
Family Studies
More widely, the parents' siblings and cousins could be carriers. Grandparental karyotypes may be useful in knowing which branch of a family to follow. The rights of individuals could potentially clash with the obligation that comes with belonging to a family: “no man is an island, entire unto himself,” and some may see altruism as a duty. Austad (1996) proposes that the family's right to know about “sensitive genetic information” should take precedence over the individual's right not to know. He considers it “alarming to use the principle of autonomy to renounce the co-responsibility for others, in this case, relatives,” and goes on to state that “we cannot exclude ourselves from the genetic fellowship of fate into which we are born.” If counselors take pains to provide clear information and to do so sensitively, such studies should proceed without unfortunate consequence. A suitable approach, in most families, will be to ask the person coming to the clinic to take the responsibility of bringing the matter to the attention of relatives, with appropriate support from the counselor. A letter couched in terms that it could be shown to other family members and provide contact points for further information is often useful.
Access to Prenatal Diagnostic Services
It would not be economically feasible or sensible to make definitive prenatal diagnosis (chorionic villus sampling or amniocentesis) available to every pregnant woman. Even among those for whom testing is, in principle, freely available, a proportion will not present, either because they are opposed to abortion or because they have not been informed about or have not understood the issues involved (Halliday et al., 2001). Those who can afford it and who do not meet criteria (essentially maternal age or other particular indicators of risk) for acceptance in the public system may have the privilege of access to private testing. Mass screening methodologies (Chapter 22) may bypass the inequity inherent in the public–private dichotomy.
Legal barriers may arise in some jurisdictions. In the United States, as Miller et al. (2000) comment, “there is perhaps no more divisive subject than abortion.” A possible ban on second-trimester (14–27 weeks) abortions would considerably affect couples having prenatal cytogenetic diagnosis, since many chromosomal abnormalities are discovered in the second trimester, and particularly in the period 14–18 weeks. Miller et al. calculate that a second-trimester ban would have a net annual cost of $74 million in the state of Michigan and $2 billion in the United States, based on the estimated lifetime costs of individuals with various congenital defects (including those other than chromosomal).
If prenatal testing is not made available, or if an abnormal result is reported but this has not been passed on to the parents, the option of pregnancy termination is denied them. Here, the legal concept of the “right not to be born” may be invoked (Weber, 2001). This issue is controversial. French courts made landmark decisions in 2000 and 2001 in which substantial financial compensation was granted to parents of children with Down syndrome. Whatever the legalities, the lesson for counselors is that testing should be offered to those for whom it may be appropriate, and they should be diligent and careful in ensuring that prenatal testing results are safely conveyed to the right person.
The Status of Embryos at in Vitro Fertilization
Lejeune has commented, indeed provided extensive testimony, on the ethical position respecting abortion and discarding an unwanted embryo. At a famous court case dealing with a dispute about in vitro fertilization (IVF) embryos in Blount County, Tennessee, in 1989, he insisted on the point that human life commences at conception and therefore that disposing of a zygote is, in essence, no different from the induced abortion of an established pregnancy. This argument is not necessarily seen as convincing to those pragmatic couples who choose to have preimplantation diagnosis to avoid the predicament of having to decide on a course of action following prenatal diagnosis of a chromosomal abnormality at chorionic villus sampling or amniocentesis. One Catholic thinker is of the opinion that “human personhood” of the embryo does not inhere until the stage at which embryonic cells have differentiated and the primitive streak has appeared (at about the end of the second week post-conception) (Ford, 1988). Prior to that time, when the “pro-embryo,” as he prefers to call it, is only a personne en devenir, “we should resist the conceptual and linguistic temptation to attribute an unwarranted ontological unity to an actual multiplicity of developing human blastomeres” (see also footnote 1, p. 391).
More liberally, Isaacs (2002) discusses the concept of a continuum, in which the “moral status” of the fetus increases in value through pregnancy (and indeed after birth); and some couples seem intuitively to follow this line. These issues underlie arguments about the validity or not of the term “pre-embryo” (Jones and Veeck, 2002; Tacheva and Vladimirov, 2002, et seq.)
Predictive Genetic Testing
Counselors are very familiar with the concept of predictive genetic testing, that is to say, offering genetic testing to people who are presently well but who are at risk for having inherited a particular genotype that may, at some stage in adult life, be the basis of the onset of disease. Its widest application is in the fields of cancer genetics and neurogenetics. With respect to translocations in the balanced state that may confer a predisposition to cancer, mention is made on p. 95. Concerning a neurogenetic focus, the delineation in recent times of the neurodegenerative disorder associated with the fragile X premutation in males (p. 223) is adding a layer of complexity to counseling issues with fragile X families. “Cascade screening” in a family may now need to take into account the fact that identifying the premutation in a male could have implications for that man's health in later life; and the detection of the pre-mutation at prenatal diagnosis will now raise a new question. The degree of risk (that is, the penetrance with respect to the neurological phenotype) has yet to be precisely defined, and the counselor will need to keep abreast of evolving knowledge. Fragile X premutation testing may become a part of the routine assessment of adult-onset ataxia, a common presenting diagnosis at the neurogenetic clinic (Macpherson et al., 2003), and such testing should, if feasible, be done with the individual's awareness of potential wider implications for the family.
