The 21 Trysomy - Current Stage of Chromosomal Research

Jérôme Lejeune

In Progress in human genetics : 144-177. Bearn et Steinberg Ed.


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History and terminology

In 1844 and 1846, Seguin described a particular type of mental retardation which he called, "furfuraceous idiocy." His description is still considered by such specialists as Benda (1962) to be the "most ingenious description of the physical characteristics of the mongoloid growth deficiency."

The term "mongoloid" was introduced later, following the racial hypothesis advanced by Langdon Down (1866) who suggested that the mental deficiency was related to the resurgence of traits of the mongolian races. Fortunately, other types of mental deficiencies which he classified as negroid and Malaysian, have been discarded from medical nosography; mongolism, however, still persists.

A recent proposal for avoiding the term mongolism was made by Allen et al. (1961), who suggested the names "Langdon Down syndrome" or "21-Trisomy." Unfortunately, "Langdon Down Syndrome" would represent the consecration of both a historical error, since Seguin was the first to describe the condition, and an etiological error, since the additionnal chromosome which causes the disease has no relation to mongolian races. In the present review the disease will be referred to as "21-trisomy" and the patients as "21-trisomics." Regardless of the standards of its recognition, the 21 chromosome is by definition the chromosome which produces "mongolism" when present in triplicate. The use of such a specialized word may be premature, but the discovery of etiological agents has always led to the renaming of diseases. Such accepted terms as "staphylococcemia" and "hypoparathyroidism" seem no longer difficult to understand, although they were undoubtedly considered unprepossessing at the time of their introduction,

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Clinical features

"Furfuraceous cretinism, with its milk white rosy and peeling skin, with its short-comings of all integuments, which give an unfinished aspect to the truncated fingers and nose; with its cracked lips and tongue, with its red, ectopic conjuctive, coming out to supply the curtailed skin at the margins of the lids" (Seguin, 1866 ) . . . is a quite obvious syndrome. The mark of imperfect definition thrown on the bodies of the children, makes them so alike that "il suffit d'avoir vu l'un d'entre eux pour ne plus risquer de les méconnaitre" (Turpin, 1931).

Aside from this particular aspect, the major symptom of the disease is mental deficiency. Although carefully studied by many investigators, the 21-trisomies do not basically differ from other feeble-minded persons, at least from the point of view of intelligence tests. Dunsdon et al. (1960), have recently stated that the I.Q. of these children is rarely greater than 80 per cent. During the first years of life, however, the I.Q. may frequently be as high as 70 per cent.

The progressive regression of the I.Q. of 21-trisomics is exemplified by the large study of Oster (1953) and has been elsewhere discussed (Lejeune, 1960). This deterioration has recently been confirmed by Zeaman and House (1962). The morphological picture of the 21-trisomic children and the associated malformations are too well known to be reiterated. For a clinical review, the reader is referred to the monographs of Øster (1953) and Benda (1960). we will attempt to review the multiple components of the syndrome in so far as they throw light on its occurrence, its nature, and its consequences.

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I. Epidemiology of the Disease

Frequency

The 21-trisomy is a relatively, common disease, possibly the most frequent of all congenital anomalies.

Table 1 summarizes current data on its frequency at birth, which is of the order of 1/600. There seems to be no difference in frequency between mongolian populations (Japan and Hawaïï) and white populations (Europe, America, Australia).

Table I
AuthorsRegionFrequency
Jenldns (1953)Chicago1/636
Malpas (1937)Liverpool1/776
Carter and MacCarthy (1951)London1/666
0ster (1953)Zeeland1/765
Collman and Stoller (1961)Australia1/688
Schull and Neel (1962)Hiroshima and Nagasaki01/785
Wagner (1962)Honolulu1/478
Jaworska (1962)Poland1/574
Maternal Aga Effect

Maternal age was the first factor recognized to be important in the etiology of the disease. Shuttleworth observed in 1895 that nearly half of the patients were the last-born of a large family. In 1909, he suggested that the aging of the mother had probably an unfavorable influence, and considered the 21-trisomics as "exhaustion products." Since these early observations, numerous authors have studied the disease; the affect of maternal age has been the only point of agreement in etiological research on 21-trisomy before the recognition of the chromosomal aberration.

Paternal age, as well as birth rank, have been demonstrated to have no influence (Jenkins,1933; Penrose,1933 and 1934). Recently, paternal age has been implicated in the rare cases of transmission of 21-22 translocations, (Penrose,1962 ); however, it is only the maternal age affect which seems to be of statistical importance. Relevant data on this point have been presented by Penrose (1961) and by Carter and Evans (1961). The graph shown in Figure 1 is based on data of Carter and Evans. It should also be pointed out that a maternal age affect has been observed in two other trisomies (17-trisomy and 13-trisomy) and is of the same order of magnitude (Lejeune, 1962).


Fig. 1. - Number of 21-trisomic children per thousand birth, relation with the age of the mother (drawn after the data of Carter and Evans, 1961).

Clinical Aspects

The characteristic clinicat features of the disease suggest that 21-trisomic children are affected from head to foot, in that every part of their body shows small departures from normal. It is the accumulation of these deviations, each of which is often at the borderline of the normal, which permits the diagnosis of the disease.

Certain peculiarities are especially significant.

Dermatoglyphic Traits.

There is a vast literature on the dermal ridges of the hands and feet of 21-trisomics. Extensive studies by varions authors stress four traits which are thought to be typical of the disease (Crookshank, 1931; Penrose,1954; Turpin and Lejeune, 1953b) :

1 ) The transversality of the ridges of the distal portion of the hand.

2 ) The medio-palmar position of the axial triradius (the t position).

3 ) The lunar loop on the hypothenar region.

4 ) The "simian line" crossing the palet transversely.

It bas been shown (Turpin and Lejeune, 1954a and b) that these four traits have taxonomic significance, since they are typical of monkeys and therefore permit a discrimination between monkeys and the anthropoids. The apparent resurgence of phylogenetic characters in 21-trisomy leads to the conclusion that a severe ge netic imibalance can overcome genic modifications accumulated during species differentiation. Other trisomies, especially 13-trisomy, also show changes in the dermatographs (Uchida, Patau and Smith, 1962), and it appears that the genetic mechanism of these traits is based on a complex interaction between numerous genes. A review of dermatoglyphic changes in various imbalanced karyotypes leads to the same conclusion (Penrose, 1963).

Nuclear Segmentation of Polymorphs

Decreased nuclear segmentation of the polymorphonuclear leukocytes was the, first cellular anomaly recognized in this disease. Described in 1947 by Turpin and Bernyer and widely confirmed since (Shapiro,1949; Lüers and Lüers,1954; Mittwoch, 1957 and 1961; Canevini and Maderna, 1962), this shift to the left of the Arneth count is definitely not related to the infections frequently observed in these children, but to a constitutional anomaly of the nuclei. In addition, the so-called "drumsticks," i. e., the small appendices seen on the nuclei of polymorphs of females, are rarer among 21-trisomic females than among normal women (Mittwoch,1961).

Predisposition to Leukemia

The high incidence of acute leukemia among 21-trisomic children was recognized before the chromosomal cause of the disease was discovered. Many authors (Schunk and Lehman, 1954; Bernard, Mathe, Delorme and Garnoud, 1955; Krivit and Good, 1956; Merrit and Marris, 1956; Stewart, 1961; Wald, Gorges, Li, Turner and Harnois, 1961; Holland, Doll and Carter, 1962 ), have demonstrated the relation between the two diseases, and it can be stated that a 21-trisomic child has roughly a twenty fold greater risk of dying of acute leukemia than a normal child.

The type of leukemia is sometimes lymphoblastic and sometimes myeloblastic, but generally speaking it is an acute process. According to Stewart (1961), most of the cases are of the stem-tell type, regardless of the actual morphology of the pathologie elements. Also, it should be recalled that the risk of congenital leukemia is extraordinarily high among 21-trisomics. This type of disease is extrernely rare; although no final statistics can be given, the excess of 21-trisomies is very great among congenitally affected patients. It seems worth emphasizing that congenital leukemia has also been observed in a case of 13-trisomy (Schade, Schoeller and Schultze, 1962). This association between the 21-trisomy syndrome and the leukemic process and the abnormal segmentation of the polymorph nucleus suggests that these congenital malformations are fundamentally related to the underlying genetic cause of the disease.

Genetic Aspects

A genetic basis for the 21-trisomy disease was suspected long ago on the basis of three sets of data: twin data, reproduction of 21-trisomic mothers, and accumulation of the disease in exceptional families.

Data on Twins

Most of the available information in the literature is summarized in Table 2, which is based on the combined data of ester (1953), Allen and Baroff (1955), and Carter and Evans (1961). The most striking finding is that dizygotic twins are as a rule discordant, and monozygotic twins always concordant. This result excludes a specific effect of the uterine environment. Nevertheless an exceptional case of monozygotic heterokaryotic twins has recently been observed (Lejeune, Lafourcade, Scharer, de Wolff, Salmon, Haines and Turpin, 1962), in which one of the twins was 21-trisomic and the other was normal. This exceptional type of twinning will be discussed subsequently.

Table 2 - Sets of Twins Containing at Least One 21-Trisomic. After the data of Øster (1953), Allen and Baroff (1955), carter and Evans (1961)
One of Them 21-trisomicBoth 21-trisomicsNo. of Pairs
Dizygous (different sexes)59059
Monozygous (same sexes and identity tests)01313
Probably dizygous (same sex)36339
Zygosity not stated (same sex)331447
12830158

Reproduction of 21-Trisomic Mothers

Very rarely 21-trisomie females procreate. Nothing is known about pregnancies of normal women fathered by a 21-trisomic male although there is no reason to suppose that 21-trisomic males are sterile. If unreliable observations are excluded, eleven 21-trisomie mothers have been reported in the literature (Table 3). As shown in Table 3, the affected mothers are relatively young (at birth of their children). Contrarily their own mothers were rather old at the birth of these trisomic mothers. This is in accordance with the maternal age effect previously quoted. Fifteen pregnancies from these mothers have been recorded. If monozygotic twins are counted as one zygote, these pregnancies may be divided into five 21-trisomics (including a still-born), and eight nontrisomics (including six normals, one mental defective and one still-born). Thus, there is fairly good agreement between these data and a 1:1 segregation of a dominant character. The ratio is in accord with the possibility of meiotic reduction in trisomies, who should produce in equal number normal (haplo-21) and abnormal gametes (diplo-21).

Table 3-Progeny of 21-Trisomic Mothers
ReferenceAge of the MotherAge of the Mother's MotherFather of the ChildChildren
Sawyer (1949); Sawyer and Shatter (1957)25?Father of the mother (incest)Normal girl; now a student 18 yr. old
Lelong et al. (1949)3042Feeble minded (not 21-trisomic)Boy, clinical syndrome of 21-tris.; dead 1 month
Rehn and Thomas (1957); Stiles and Goodman (1961)19 (tris.-21)30unknownGirl (tris.-21)
Forssman and Thyssel (1957) Lehmann and Forssman (1960)20 (tris.-21)42Blind, epileptic (46 chromosomes N)Normal boy (46 chromosomes N)
Schlaug (1957)2939Father of the mother (incest)Abnormal girl; syndrome very different from the classical 21-tris.
Hanhart (1960)21 (tris.-21)Brother of the mother (incest)Girl-(tris.-21)
Hanhart et al. (1961)23 (tris.-21)44Feeble-minded (46 chromosomes N)Boy-Typical syndrome of 21-tris.; dead 1 1/2 yr.
Mullins etal. (1960)2222Feeble mindedNormal boy
Levan and Hsu (1960)???Normal boy (46 chromosomes N)
Thuline and Priest (1961)14 (tris.-21)p?Normal monozygous twin boys (46 chromosomes N) dead
Thompson (1961)21?Feeble mindedNormal boy, now 10 yr. old
20'??Female macerated foetus
22??Male dead foetus probably 21-trisomic
Note: The subjects for which chromosomal analyses are available are indicated: (tris.-21) or (46 chromosomes N).
Familial Occurrence

Recurrence of the disease in a given family is a rare event. Most of the cases are isolated occurrences in an otherwise normal family, but the number of sibships including at least two 21-trisomics is significantly higher than would be expected by chance alone (Penrose, 1934; Ester, 1953; Carter and Evans, 1961). One exceptional instance includes a family of seven children, five of whom are affected (Turpin and Lejeune, 1953a). Such cases suggest existence of a particular mechanism for the disease. As will be discussed subsecfuently, the occurrence of a balanced translocation in one of the parents in these sibships explains the recurrence of the disease. Nevertheless, as pointed out by Hamerton, Briggs, Giannelli and Carter (1961), even if translocation cases are excluded, a small excess of recurrence still occurs in some farnilies; these authors raised the problems of gonadal mosaics and the familial tendency to abnormal segregation. The accumulation of minor stigmata of the disease in healthy members of an affected family has also been reported, such as furrowed tongue (Turpin and Caratzali, 1933; Turpin, Bernyer and Teissier,1947) and elevation of axial triradius (Penrose,1954). The existence of hypothetical genes which increase the chance of mis-segregation of chromosomes should also be considered.

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II. The Chromosomal Anomaly

Speculations on the etiology of mongolism from 1846 to 1959 are sa numerous that it is difficult to imagine a factor which has not been incriminated (see warkany, 1960, for review) . From the general data pointing toward a constitutional disease which acts on numerous traits two authors (Waardenburg,1932; Bleyer, 1934) drew the formal conclusion that the disease was determined by a chromosomal aberration and accepted this etiology as the only one possible. These two remarkable papers show that in biology logical deductions may precede physical observation by a quarter of a century. Other authors have also discussed the chromosomal hypothesis to explain exceptional families (Penrose, 1939), or to explain the general data (Turpin and Caratzali,1937; Fanconi, 1939). Nevertheless., the etiology of the disease remained unproved until the observation of the 21-trisomy.

The Typical 21-Trisomy

Prior to 1958 the only published attempt to observe the chromosomes of affected patients was the investigation of Mittwoch (1952); with the limitations of the available techniques, she was unable to arrive at a firm conclusion. She observed 24 chromosomal masses at meiosis and supposed that this was explained by the somatic number of 48, which was the accepted diploid number at that time.

Two years after the establishment of 46 as the correct somatic chromosome number of man (Tjio and Levan, I956; Ford and Hamerton, 1956), an affected child with 47 chromosomes was observed in Paris. A further study of two other cases was completed in 1958 and the existence of an extra chromosome in this syndrome was published in January 1959 (Lejeune, Gautier and Turpin, 1959b) . Additional confirmations were published by Ford, Jones, Miller, Mittwoch, Penrose, Ridler and Shapiro (1959), Jacobs, Baikie, Court Brown and Strong (1959), and Böök, Fraccaro and Lindsten (1959). Subsequently the 21-trisomy was demonstrated as the cause of the disease in all human races (Makino, Tonomura and Matsunaga, 1960; Lee, Schmid and Smith, 1961; Kleisner, 1961; Conen, Bell and Rance, 1962).

As exemplified in Figures 2 and 3, the presence in triplicata of a small satellited acrocentric chromosome can be established beyond any doubt in suitable cells of affected individuals (for technical discussion, see Lejeune, 1960). Many authors have discussed whether the triplicated chromosome is number 21 or 22. In the "Denver agreement" it was stated that chromosomes should be classed in decreasing order of length; if satellites are excluded, chromosome 22 is slightly longer than 21, thus emphasises the difficulty raised by the numerical classification.

A definite position must be taken in this byzantinian quarrel; it can be presented as follows : The chromosome which produces the so-called "mongolism" when in triplicate, is to be called 21. The other small acrocentric pair is then numbered 22. It should also be pointed out, as accepted in Denver, that if new morphological characters were found which would aid in recognizing chromosome 21, they would not warrant a change of the already accepted number.


Fig. 2 - The 47 chromosomes of a 21-trisomic male.


Fig. 3 - Karyotype of the preceding cell. Remark: three small acrocentrics with satellites (21-trisomy). One of the 22 also exhibit satellites but less prominent.

The Trisomic State

The observation of three similar elements is not in itself sufficient reason for accepting the hypothesis that the elements are homologous chromosomes present in true trisomy. Fortunately, meiotic studies have been performed on testicular biopsies of 21-trisomic males. The production of trivalents has been recorded in a pure trisomic (Miller, Mittwoch and Penrose, 1960 ) and also in translocation carriers (Hamerton, Cowie, Giannelli, Briggs and Polani, 1961).

Since the synaptic process is the best indication of homology, it may be concluded that the trisomic nature of the disease is not only logically established, but that it has been confirmed cytologically as far as it is possible to do so in a species which does not possess polytenic giant chromosomes.

Hidden Trisomies

As previously stated, most of the 21-trisomics, perhaps 95 per cent, exhibit three free 21 chromosomes and have a diploid chromosome number of 47. The first exception to this rude was reported by Polani, Briggs, Ford, Clarke and Berg (1960), in a girl exhibiting only 46 chromosomes although suffering from the typical clinical disease. Although the child was born of normal parents (Carter, Hamerton, Polani, Gunalp and weller, 1960), she had only four small acrocentric chromosomes. The extra 21 chromosome was found to be attached to a large acrocentric chromosome of the 13-15 group. Such transfer of chromosomal material was very similar to the first case of translocation reported in man (Turpin, Lejeune, Lafourcade and Gautier, 1959), which also occurred between a small and a large acrocentric chromosome.

The number of instances of translocation in the literature is now significant. Their general importance has been previously discussed (Turpin and Lejeune, 1961), and their role in 21-trisomy has been recently reviewed (Carr,1962 ). For the sake of simplicity they can be classified into two categories, those between a large and a small acrocentric chromosome and those between two small acrocentric chromosomes.

Translocation Between a Large and a Small Acrocentric

In general, the large acrocentric involved in this rearrangement (see Fig. 4) cannot be cytologically recognized, although it certainly belongs to the 13-15 group. For simplicity it will be referred to here as a (13) chromosome; the number in bracket refers to the whole class and does not specify whether the element is in fact a 13, a 14 or a 15 chromosome.

The small acrocentric is by definition chromosome 21, since it produces the 21-trisomy syndrome if in triplicate. Several families have been investigated in which the segregation of the rearranged chromosome has been observed. The data are summarized in Table 4. At a first glance it can be estimated that normal persons (46 chromosomes), normal persons with translocations (45 chromosomes), and 21-trisomics with translocations (46 chromosomes), are observed in approximately equal proportions among the progeny of carrier mothers.

It may be assumed that the reciprocal of the 21-trisomies (i.e., the haplo-21 zygotes) are produced in comparable number but do not survive. In such a case the conclusion would be that once in every two meioses, there is a mis-segregation of the free 21 chromosome. In other words, the translocation completely disturbs the assomment of the 21 chromosome, and the free one and the translocated one segregate independently (see Fig. 5). Table 4 is also useful for the purpose of genetic counselling, as it shows that a translocation carrier mother will as a rule give birth to a 21-trisomic child once in every three pregnancies.

The picture concerning the fathers is quite different, as is seen in the table; only two cases of trisomy-21, one of which is the propositus, are recorded among 49 children. The reason for such a difference between orale and female carriers is not understood.

Table 4 - Progeny of Carriers of a 21-(13) Translocation
Carrier ParentChildren
N.46 N.45 (T)Tris.-21, 46 (T) Tris.-21*Normal* Total
Mothers: 3223 2625 (14)131198
Fathers: 1515 221 ( 1)11049
( ) No. indicates the number of propositi; (T) indicates carrier of translocation. Compiled from the data published by Penrose et al. (1960); Carter et al. (1960); Penrose and Delhanty (1961); Puckton et al. (1961); Hamerton et al. (1961); German et al. (1962); Sergovich et al . (1962); Forssman and Lehmann (1962); Breg et al. (1962); Atkins et al. (1962); Biesele et al. (1962); MacIntyre et al. (1962) Shaw (1962). * Only the phenotype is known.


Fig. 4 - Translecation 21-13. During the time a 21 and a (13) lie close together, a break occurs close to the centromere of each chromosome. The fragments recombine either in the previous way (no detectable affect) or rearrange producing a new hybrid chromosome 21-13 and a centric fragment bearing the satellites, which is secondarily lost. A varions amount of chromosome material can he attached to the centric fragment. The quantity of genes lost determine the eventual phenotypic affect of the translocation. This general process of rearrangement between acrocentrics is commonly referred to by the rather misleading term centris fusion.


Fig. 5 - Progeny of a carrier of a 21-(13) translocation. In the simplified diagram, of meiosis each chromosome is represented for sake of simplicity by one chromatid instead of two. The progeny show the four types of possible children of which only the three first are effectively observed, the type Haplo-21 being not known (probably lethal).

Translocations Between Two Small Acrocentrics

In this case it is difficult to tell whether the translocated chromosome is formed by the fusion of a 21 and a 22, between two 21's or even between two 22's. The recognition of the 21 is difficult in itself and becomes particularly troublesome in cases of translocation. The nucleolar material, divided among the satellites is the object of competition between the acrocentric chromosomes, as shown by Nawaschin (1934). Hence the use of the satellites as markers of 21 becomes misleading under these conditions.

The available data on the progeny of carriers of 21-(21) type translocations are summarized in Table 5.

As illustrated in Figure 6 the progeny of an isochromosome 21-21 carrier should be composed exclusively of 21-trisomics, since the haplo-21 zygotes are probably lethal. The families described by Forssman and Lehmann (1962) and Dallaire et al, (1962), seem to fit this prediction, being composed of 13 21-trisomies among 13 children. Conversely, the family reported. by Shaw (1969), is in favor of a 21-22 translocation since she finds 2 21trisomics amont 18 children with seven normal children and eight translocation carriers. Finally, the examples of mosaicism of Fraccaro et al. (1960) and Hamerton et al. (1961), can be attributed to either type, as the appearance of normal children is an insufficient reason to exclude the 21-21 type of translocation,

Although it has been reported that paternal age may act as a predisposing factor this has net yet been proved in casas of 21-trisomy caused by 21-21 translocation (Penrose, 1962). The only conclusion to be drawn from the available data is that contrarily to the 21-(13) type, the 21-21 translocation induce 21-trisomy as often at least in the progeny of male carriers than in the progeny of carrier mothers.

Before closing this brief discussion of the data it must be stressed that many of the translocation 21-trisomies are not transmitted, but arise de novo in the affected child (Gustavson, 1962; Carter, Hamerton, Polani, Gunalp and weller,1960; Scherz,1962). Both a 2I-trisomic with 47 chromosomes and a 21-trisomic caused by translocation have been observed in a sibship of which both parents were normal (Penrose, 1963).

Table 5 - Progeny of Carriers of a 21-(21) Translocation
Carrier ParentChildren
21-22 (likely)N.46N.45(T)Tris.-21, 46(T)Tris.-21*N*Total
Fathers: 24(2)48
Fathers: (mosaics): 22(1)147
Mothers: 2325
Total9(3)74 20
21-21 (likely)
Fathers: 2 751(1) 13
Mothers: 231(1)4
( ) No. indicates the number of propositi; (T) indicates carrier of translocation. Compiled after the data of Hamerton et al. (1961); Forssman and Lehmann (1962); Fraccaro et al. (1960); Show (1962); Mukherjee et al. (1962); Dallaire et al. (1962).* Only the phenotype is known.


Fig. 6 - Progeny, of a 21-21 Translocation carrier. At meiosis the new chromosome 21-21 has no homologue and moves directly to one pole, the other cell being entirely deprived of 21 chromosome. As seen the progeny can only include 21-trisomies or miscarriages. Note: In case of a 21-22 translocation, the transmission would be analogous to the case of a 21-(13) translocation (Fig. 5).

Mosaic Trisomies

Although 21-trisomies seem to be affected in every part of their economy, careful investigation has shown that sonne individuals, exhibiting the 21-trisomy syndrome in its classical form, or in a less clear cut picture ("mild affection") were, in part, mosaics. Their cell population was not uniformly composed of 21-trisomie cells, but included a varying proportion of normal, diplo-21. cells. The data are summarized in Table 6.

From these few data, it is not yet possible to deduce a relationship between the percentage of the 21-trisomie tells and the mental and physical status of the individual, although the patient with tetra-21 cells showed a particularly severe mental retardation.

Related to this problem is the biological relevance of the percentage observed. We have no good evidence that 21-trisomie cells and 21-diploid cells do not compete against each other under the conditions of observation. If this indeed is the case, the number of mitoses observed is not an unbiased estimate of the cells present in the sample studied. Secondly, the number of biopsies which can possibly be made is limited, and only few regions of the body can be examined. There are very good indications that mosaic individuals can differ from one tissue to another (Fraccaro, Bott, Salzano, Russel and Cranston, 1962). The physiological effect of this "geographical" mosaicism could be very different depending upon whether a high percentage of aneuploid cells was localized in one organ or another. Selection between diplo-21 and tripla-21 karyotypes during embryonic development is another uncertainty.

Table 6 - Mosaics for 21-Trisomy
AuthorsTissueNo. of CellsPourcentage of KaryotvpesRemarks
Diplo-21Triplo-21Tetra-21Other
Clarke et al. (1961) mother 26 yr. father 26 yr.Skin89 53% 47% Femelle 21/2 yr.; morphology and dermatoglyphs are typical; I.Q. 100; no 21-trisomic cells in the blood
4438% 62%
Blood22100%0%
Fitzgerald and Lycette (1961) mother ? father ?Blood10042%53%5%Male 51 yr.; low grade mental deficiency; diagnosis evident, few stigmata lacking (no epicanthus no simian crease, normal ears)
Gustavson and Ek (1961) mother 24 yr. father 29 yr.Skin6030%30%40%Male 12 yr.; had a brother 21-trisomic, dead at 6 months; low grade mental defi-ciency; typical deformities; trisomy-21 plus a small metacentric (21-21) (?)
Nichols et al. (1962)No details available
Warren et al. (1961)Blood9358%27%?15%Femelle. Typical aspect; annular pancreas; two other cases cited, one observed by Lytt, the other by Conen
Hayashi et al. (1962) mother 17 yr. father ?Blood100 55% 45%Male 3 yr.; incomplete picture of 21-trisomy; I.Q. reported in the normal range
1857% 43%
Richard and Stewart (1962)Blood4548%52%Male Typical case; low grade mental defi-ciency
Lindsten et al. (1962)Blood and Skin29362%35%3%Femelle Mild clinical picture although diagno-sis is evident; borderline intelligence
Blank et al. (1962) mother 40 yr.BloodFemelle Mother of a typical 21-trisomic; few stigmata of the disease
Association with Other Chromosomal Changes (21-Trisomy and XXY)

The most frequent association is that of 21-trisomy and the Klinefelter syndrome (XXY), resulting in a karyotype with 48 chromosomes. Several cases have been described; Ford, Jones, Miller, Mittwoch, Penrose, Ridler and Shapiro (1960), described the first case, followed by Lanman, Sklarin, Cooper and Hirschhorn (1960) and Lehmann and Forssman (1960).

A case of monozygotic twins, both 21-trisomics + XXY, was published in 1961 by Hustinx, Eberle, Ceerts, ten Brink and Woltring and a similar case of twins has been recorded in our laboratory (Inst. Prog. obsv. No. 721).

The expected converse association between 21-trisomy and the Turner syndrome has not been reported. We have searched for 21-trisomic females exhibiting pterygium colli and found one such case but her karotype was 21-trisomy, diplo-X. It is possible that the severe genetic imbalance of the XO condition superimposed on the deleterious effect of the 21-trisomy does not allow the embyronic development of such a zygote.

Another association with 48 chromosomes, 21-trisomy and 18-trisomy, has been described (Gagnon, Katyk-Longtin, de Groot and barbeau, 1961) but the malformed infant, exhibiting the physical stigmata of both syndromes, was not viable.

Related Malformations

Atypical cases, relating to chromosomal changes, have also been described. A girl exhibiting a congenital malfor- mation looking somewhat like 21-trisorny but having probably a 21-16 translocation, has been reported by Böök, Gustavson and Santesson (1961). Also a mosaic boy exhibiting in some cells a very small extra chromosome has been recorded by Ilbery, Lee and Winn (1961), and is considered a "partial 21-trisomic."

Other cases showing trisomy for a small acrocentric have been quoted repeatedly. Firm conclusions on the genetic identity of the chromosomes involved cannot be drawn from a consideration of these cases (Lejeune, 1962).

Possible partial trisomies involving very small translocations on the 21 have also been reported: Chu, Rubinstein and Warkany (1961) and Gray, Mutton and Ashby (1962). A few perplexing cases have been reported: a borderline case with normal chromosomes by Schmid, Lee and Smith (1961); a typical syndrome with normal karyotype by Hall (1962) (who could not exclude mosacicism) ; and two trisomies for a small acrocentric with so-called "non-mongoloid" mental deficiency by Zellweger, Mikano and Abbo (1962). Further investigation of these cases seems highly desirable to define the clinical disease as well as the chromosomal status.

This survey of exceptional cases must be evaluated in the light of the knowledge that probably more than 1,000 individuals with 21-trisomy have been studied in varions laboratories and that even an apparently homogeneous disease may show clinical variation. Every 21-trisomic has a unique complement of genes on the 21 chromosome and many genic differences exist between the 21 chromosomes present in human population.

In addition, as in the case of mental retardation observed by Ellis, Marshall and Penrose (1962), the extra chromosome can be rearranged, resulting in trisomy for varions chromosomal segments.

Pathogenesis of the Chromosomal Aberrations

General factors predisposing to abnormal segregation of chromosomes have been recently reviewed (Lejeune, 1962) and only the data directly relevant to 21 trisomy will be discussed here.

Characteristics of Chromosome 21

The presence of satellites on this chromosome and the relation between these bodies and the nucleolus has led to the view that this type of acrocentric could be at greater risk of abnormal segregation than others. Mitotic association of acrocentric chromosomes is well substantiated (Ferguson-Smith and Handmaker, 1961). Thus, chromosome 21 could be at greater risk of translocation with another satellited acrocentric (Ohno, Trujillo, Kaplan and Kinosita, 1961) . The existence of the (13)-trisomy (Patau, Smith, Therman, Inhorn and Wagner, 1960) and of the numerous 21-(13) and 21-21 translocations previously reported strongly supports this argument. Also the identical tune of replication of these two types of chromosomes points in the same direction (Gilbert, Muldal, Lajtha and Rowley, 1962) .

The Interchromosomal Effect

The evidence that structural changes of chromosomes increase the risk of abnormal segregation of the X chromosome has been amply demonstrated in Drosophila by the work of Sturtevant (1944). The same affect seems to be detectable in man.

The structural affect is clearly demonstrated in the progeny of carriers of a translocation involving the 21. Moreover, the translocation can induce abnormal segregation of the 21, even if this chromosome is not transmitted to the trisomic child. This is well exemplified in the family described by Moorhead, Mellman and wenar (1961), in which a mother carrying a probable 22-(13) translocation gave birth to a child with a free 21-trisomy, and without having received the translocated chromosome.

In a family described by Shaw (1962), the mother had an apparent deletion of a small arm and satellite of a 21 (or 22). She had five children; two were normal (both showing the "marker" chromosome), and three were 21-trisomies with 47 chromosomes (one of them exhibited the marker chromosome and the other did not). In a second family the mother of two typical 21-trisomies with 47 chromosomes had a normal set of 46 chromosomes but exhibited an excess of genetic material in one of the big acrocentrics. This excess, possibly including part of the 21, is thought to be the cause of the mild "stigmata" found in her phenotype. It seems likely that these chromosomal rearrangements make the meiotic pairing unstable or irregular and thus increase the risk of mis-segregation of homologues, or even of chromosomes which are not homologous with the rearranged one.

A less likely possibility is that chromosome rearrangements increase the risk of mis-segregation in the somatic cells. This is well known in Drosophila melanogaster in which gynandromorph mosaics result from the elimination of a ring-X chromosome (Brown and Hannah, 1952). Some data in man also point in this direction. Segmentary heterochromia of the iris, supposed to be due to chromosomal change when produced by irradiation (Lejeune, Turpin, Rethore and Mayer, 1960), has been recorded in a normal child belonging to one of the sibships described by Shaw and listed above (the mother with a possible 21-(13) translocation). Also this effect on meiotic and mitotic stability would possibly explain the accumulation of chromosomal diseases in certain sibships (Mosier, Scott and botter, 1961; Böök, Santesson and Zetterqvist, 1961; Hauschka, Hasson, Goldstein, Koepf and Sandberg, 1962; Cooper and Hirschhorn, 1961; Benirschke, Brownhill, Hoefnagel and Allen, 1962; Johnston, 1961; Zellweger and Mikano,1961) and it seems at least an attractive hypothesis to suggest that familial recurrence of 21-trisomy could be explained by some undetected structural rearrangements in the chromosomes of parents.

Such an instability of the karyotype could be related to the occurrence in these families of a neoplastic process, such as leukemia (see below) (Buckton, Harnden, Baikie and Woods, 1961; Miner, Breg, Schmickel and Tretter, 1961; Hungerf ord, 1961).

Aging of the Mother

This predisposing factor, as previously mentioned, has no clear cut counterpart in the experimental animal; at least the data are not available. Nevertheless in the case of somatic loss of the ring X-chromosome quoted above, the aging of the eggs has a very striking effect (Hannah,1955). The same effect was found by Patterson, Brewster and Winchester (1932) on production of XXY, but only if the eggs were irradiated with x-rays.

The maternal effect, as far as experimental data are concerned, appears to affect somatic cells more than the meiotic process. This possible restriction to the blastomeric stage could be compatible with 21-trisomy, since there are instances in which the occurrence of the chromosomal disease is clearly postzygotic.

The Time of Occurrence of the Aberration

As discussed in the problem of translocations, there are instances in which we can assume fairly safely that a diplo-21 gamete was produced by a carrier parent. The same is true in the rare cases of reproduction of a 21-trisomie mother. Only in these instances which are obviously very rare (less than a few per cent of the total frequency of the disease) can we assume confidently that the error occurred at the meiotic process.

In contrast, two types of data show that the error can be postzygotic. The first concerns those cases of mosaicism in which obviously the "variant cells" were produced after gametic fusion. The probability of a double fertilization proposed by Gartler, Waxman and Giblett (1962) to explain their case of hermaphroditism can certainly not apply to all the mosaics with 21-trisomy. The second evidence is furnished by the exceptional type of twinning, the "heterokaryotic monozygotism" described by Lejeune, Lafourcade, Scharer, de Wolff, Salmon, Haines and Turpin (1962). A pair of monochorionic twins was observed who were identical for all the blood groups tested, but who differed in that 21-trisomy occurred in one while a normal karyotype (and phenotype) occurred in the other. A case of monozygotic twins differing in one chromosome had been previously observed (Turpin, Lejeune, Lafourcade, Chigot and Salmon, 1961; Lejeune and Turpin, 1961). This instance, an X0 Turner and her XY normal monozygotic co-twin, is no longer unique, since a similar set of twins has been recently observed by Edwards, Dent and Crooke (1963, cited by Lindsten et al., 1963).

Figure 7 shows one possible way of producing these twins. Alternatively the twins could arise from a 21-trisomie egg and a secondary loss of the extra chromosome at the blasteromeric stage. In this second hypothesis the normal twin would have been naturally "cured" of his congenital disease.

If spontaneous mosaicism for the sex chromosome, as well as its production in animals by irradiating the fertilized egg (Russel and Saylors, 1961), is taken into consideration, the general picture emerges that the most sensitive stages of development are the time of the fusion of the gametes or the first few cleavages of the blastomeres. If this hypothesis is true, it could very well be that most of the 21-trisomies are not of gametic origin, as generally accepted, but blastomeric, with the eventual death of the recripocal clone (haplo-21).


Fig. 7 - Abnormal segregation of the blastomeric stage. Only the 21 chromosomes are represented. First mitosis of the zygote is normal. Then, one of the blastomeres divides normally, the other one give rise to two abnormal cells: a 21-trisomic and a Haplo-21 (which probably disappears). lf cleavage in two embryos occurs at this four cell stage, the result is monozygotic heterokaryotic twins (see text). If only one embryo develops, the result will be a diplo 21/triplo 21 mosaic (see text).

The Leukemic Process

The strong association between 21-trisomy, free or by translocation (German, DeMayo and Bearn, 1962), and acute leukemia is well known. New karyotypic studies have led to precise results. Tough, Court Brown, Baikie, Buckton, Harnden, Jacobs, King and McBride (1961), reported five cases of apparently unchanged karyotype in the peripheral blood of 21-trisomic children affected by an acute leukemia. These authors raise the possibility that the true leukemic cells were not, in fact, recorded in the mitoses and that this negative finding is not definite.

Recently, leukoblastic abnormality was observed in a 21-trisomie girl in whom a chromosomal variation leading to an abnormal karyotype of 54 chromosomes was observed in the bone marrow and in the blood (Lejeune, Berger, Haines, Lafourcade, Vialatte, Satge and Turpin, 1963). The hemorrhagic disorder, which was characterized by purpura and bleeding, necessitated many blood transfusions, was diagnosed at birth and was still present at 2 years and 2 months. Analysis of cells revealed a chromosomal complement which varied from 47 to 55 and showed that each number was correlated with a specific karyotype. The additional chromosomes were (in increasing number) small acrocentrics (v), big acrocentrics (T), medium sized metacentrics (M) and small metacentrics (c).

Two facts emerged from this study : (1) Every karyotype of an immediately greater number contained all the supernumeraries found in the karyotype of an immediately inferior number, plus one; (29) every new supernumerary occurred twice before the acquisition of a new one. The general data are summarized in Table 7.

Although this apparent evolution of a clone is not yet substantiated by other observations, it is worth noting that a previous observation of a 47/49 mosaic was reported in the bone marrow of a 21-trisomie girl by Piazzi and Rondinini (1961) (it is not known if this child was suffering from leukemia). The observation by boss and Atkins (1962) of another 21-trisomie with leukemia also showed 47/49 mosaicism in the blood, In both these instances the 49 chromosome karyotype was due to a typical 21-trisomy with two additional small acrocentrics (21-22).

The availability of only three cases for study prevents any generalization, but it is tempting to suppose that is was not by chance in the three instances now known that the karyotypic variations started with the same type of chromosome and with the appearance of two additional identical ones.

Other examples of the presence in duplicata of additional chromosomes in neoplastic disease have been reported. One case concerns apparently normal big acrocentrics (13-15) (Sasaki, 1961), and another obviously rearranged giant acrocentrics in cells of exudate of cancerous origin (de Grouchy, Vallee and Lamy, 1963).

The possibility of chromosomal interaction resulting from the trisomy in the causation of leukemic process in the 21-trisomie children is for the moment an open question, as also are the possible laws controlling the karyotypic changes in these neoplastic growths.

Table 7 - Association Between Karyotype and Chromosome Number in the Cells of a Trisomy-21 Patient with Leukemia
No. of ChromosomesKaryotvpeNo. of Cells
47tris.-2122
48tris.-21+1v3
49tris.-21+2v1
50tris.-21+2v+1T1
51tris.-21+2v+2T1
52tris.-21+2v+2T+1M3
53tris.-21+2v+2T+2M7
54tris.-21+2v+2T+2M+1c36
55tris.-21 +2v+2T+2M+2c3
62tris.-21+2v+2T+2M+2c+18+5M+21
Total78
Exceptions : One cell with 51 chromosomes : tris.-21 + 2v + 1T + 1M; one cell with 53 chromosomes : tris.-21 + 2v + 2T + 1M + 1c. - the symbols used, described in Lejeune (1960), represent: v = group) (21-22); T = group (13-15); M = group (6-12-X); c = group (19-20).
The Genic Confient of the 21 Chromosome

The study of chromosomal duplication (or deletion) as a means of detecting genes which show a dosage affect has been a very powerful tool in experimental cytogenetics. Its application to the mapping of human chromosomes has been attempted, especially in the 21-trisomy syndrome. In such research very great care has to be exercised in the selection of phenotypic characteristics as "markers." Fine and immutable morphalogical traits, like dermatoglyphs, have been proposed, but their alteration in all three different types of trisomies (see above) prevent any conclusion about the precise location of the hypothetical genes controlling them.

Comparisons between members of a monozygotic heterokaryotic set of twins for somatic traits are also possible, since these individuals are genetically identical except for one 21 chromosome. In the case quoted above, the 21-trisomic differed from his normal co-twin by having straight hair, whereas the normal boy has curly hair. It connot be concluded that the "straight hair gene" is on chromosome 21, but only that some genes on this chromosome interfere with the manifestation of this character.

The difficulty of genetic analysis with morphological traits is well known. Biochemical characteristics seem more satisfactory as they are probably nearer to the primary products of gene activity.

Alkaline Phosphatase Activity of Polymorphic Leukocytes

The estimation of alakaline phosphatase activity in polymorphic leukocytes provides an apparently simple biochemical test applicable at the cellular level. Since the first repart of Alter, Lee, Pourfar and Dobkin (1962), numerous publications have confirmed the fact that alkaline phosphatase activity is higher in 21-trisomics than in normal children. The ratio of activities is of the order of 3 to 2, and is strikingly similar to the ratio of 21 chromosome numbers (Trubowitz, Kirman and Masek, 1962; King, Gillis and Baikie, 1962). No correlation his found with the Arneth count (Lennox, White and Campbell, 1962), or with the neutrophil count (King, Gillis and Baikie, 1962). Thus the relation between alkaline phosphatase activity and the karyotype seems independent of the tendency of 21-trisomies to incur infections. The tentative localization of the structural gene on the 21 chromosome is nevertheless rendered insecure by the negative correlation between this enzymatic activity and prepubertal age, which suggests that a correlation may exist with the level of sex hormones (Alter, Dobkin, Pourfar and Lee, 1963). Nevertheless the conservative hypothesis that genes affecting alkaline phosphatase activity are located on the 21 can reasonably be made, with the reservation that other genes, possibly many, can also influence this trait.

Another indication that alkaline phosphatase activity may be related to the genes on chromosome 21 arises from the observation of a deleted small acrocentric (Ph1 chromosome) in leukocytes of patients suffering from chronic granulocytic leukemia (Nowell and Hungerford, 1960; Tough, Court Brown, Baikie, Buckton, Harnden, Jacobs and Williams, 1962). If the deleted chromosome is indeed the 21, the low phosphatase activity found in this disease (Valentine and Beck, 1951) could be attributed to the loss of the locus involved.

In addition the observation of haplo-(21 or 22) clones in acute myeloblastic leukemia (Ruffie and Lejeune, 1962) supports the supposition, drawn from the Ph1 chromosome, that genes exist on the 21 which are concerned with the regulation of polymorphonuclear production.

It must be stressed that this second type of inference is based on the assumption that the Ph1 chromosome is really a deleted 21. This tempting assumption, although not proved, could also account for the shift to the left of the low Arneth count in 21-trisomics. By this hypothesis a simple mechanism could be postulated: a balanced karyotype gives normal counts, a triplo-21 depresses the lobulation of the nuclei of the polymorphs and a partial or total deletion of the 21 leads to an excessive proliferation of this type of cell.

As has been shown in the paragraph concerning leukemias in 21-trisomics, an extra supernumerary of this same acrocentric type can also be encountered in the leucoblastic reaction. The significance of this last effect is not immediately apparent, but we can at least imagine that hyperrepression of the granulocytic differentiation could result in blastic proliferation.

Biochemical Changes

Biochemical changes may be more sensitive indicators of gene overdosage than cellular effects. Many biochemical traits have been reported to be peculiar in 21-trisomies. Thus blood calcium level (Stern and Lewis, 1958) and serum esterase level (Stern and Lewis, 1962) are reported to be lower in 21-trisomics than in normal individuels, whereas the serum uric levels (Fuller, Luce and Mertz, 1962) are reported to be higher. In a systematic search for biochemical changes related to 21-trisorny, tryptophan metabolism has been investigated, and a decreased urinary excretion of xanthurenic acid and of 5-hydroxyindolacetic acid has been reported (Gershoff, Hegsted and Trulson, 1958; Gershoff, Mayer and Kulcyzcki, 1959; O'Brien, Groshek and Streamer, 1960). As observed by Jerome, Lejeune, and Turpin (1960) and Jerome (1962), a decreased excretion of 5-hyclroxyindolacetic acid, xanthurenic acid, kynurenic acid, and indolacetic acid seems to be a biochemical symptom of the disease. No direct relation between these metabolic changes and chromosomal constitution has yet been established, although acceleration of some steps by enzymatic overdosage has been discussed. This hypothesis of over production of enzymes by trisomics seems to be confirmed by the alkaline phosphatase data, but cannot yet be related to experiments in other species. In bacteria, however, there are indications that genetic overdosage induces an excess of enzymatic activity.

Haut

Conclusions

Although chromosomal research has greatly improved our knowledge of the disease, the pathogenesis of 21-trisomy has still to be discovered.

Two main questions need to be answered:

1. How can normal genetic information be deleterious if in excess ?

2. What biochemical changes are produced by this imbaIance of the karyotype ?

The cytogenetic epoch of 21-trisomy has obviously not closed, but it is hoped that a biochemical era will soon open. The discovery of specific metabolic changes in 21-trisomies would be an outstanding advance, and the successful control of these changes, if realizable, would receive a most precious reward: the first reasonable hope of correcting the catastrophic consequences of an inborn chromosomal essor.


Haut

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