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Cytogenetic findings in 250 schizophrenics: evidence confirming an excess of the X chromosome aneuploidies and pericentric inversion of chromosome 9
Schizophrenia Research 40 (1999) 43–47 www.elsevier.com/locate/schres
Cytogenetic findings in 250 schizophrenics: evidence confirming an excess of the X chromosome aneuploidies and pericentric inversion of chromosome 9 H. Kunugi *, K.B. Lee, S. Nanko Department of Psychiatry, Teikyo University School of Medicine, 11-1, Kaga 2 Chome, Itabashi-ku, Tokyo 173-8605, Japan Received 12 October 1998; accepted for publication 11 February 1999
Abstract Chromosomal abnormalities may be of help in identifying disease genes. To search for susceptibility loci for schizophrenia, we have performed chromosomal examinations by using the GTG banding technique for 250 schizophrenics. We found five cases with an aneuploidy of the X chromosome and ten cases with pericentric inversion of chromosome 9 [inv (9)]. These results confirmed an excess of the X chromosome aneuploidies in schizophrenia, indicating a possible involvement of the X chromosome in the pathogenesis of the illness. The observed incidence (4.0%) of inv (9) in our schizophrenic sample was significantly higher ( p=0.013) than that reported in the general population in Japan (1.7%). Although inv (9) has been considered to be a normal variant, our observation implies a possible association between inv (9) and schizophrenia, suggesting that a susceptibility locus for the disease may be located at a breakpoint of the inversion on chromosome 9. © 1999 Elsevier Science B.V. All rights reserved. Keywords: Chromosome 9; Cytogenetics; Pericentric inversion; Schizophrenia; X chromosome
1. Introduction Although many linkage and association studies have been performed, no gene has yet been established as giving major susceptibility to schizophrenia (Crow and DeLisi, 1998). The use of chromosomal examinations could be an alternative or complementary approach to identify a susceptibility locus for schizophrenia (Bassett, 1992). Chromosomal abnormalities may be helpful to find not only candidate regions for linkage studies, but also valuable materials for positional cloning. In this regard, we previously performed a cytogenetic screening on 120 psychotic subjects (116 * Corresponding author. Tel./fax: +81-3-3964-2447. E-mail address: [email protected] (H. Kunugi)
patients with schizophrenia and four with delusional disorder) (Nanko et al., 1993). In this preliminary report, we found two schizophrenics with 45,X/46,XX mosaic and four with pericentric inversion of chromosome 9 [inv (9)]. Subsequently we have examined a second sample of 134 schizophrenics. Here we report chromosomal findings for the combined sample of 250 schizophrenics. Although several studies have conducted a systematic cytogenetic screening of psychotic subjects, many of these date back to more than two decades ago; thereafter, cytogenetic techniques have been progressing, and diagnostic criteria for psychiatric disorders have been refined. To our knowledge, this is the largest study to examine chromosomal abnormalities for schizophrenics diagnosed by DSM criteria.
0920-9964/99/$ – see front matter © 1999 Elsevier Science B.V. All rights reserved. PII: S0 9 2 0 -9 9 6 4 ( 9 9 ) 0 0 03 5 - 3
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H. Kunugi et al. / Schizophrenia Research 40 (1999) 43–47
2. Methods
3. Results
2.1. Subjects
In our previous sample ( Table 1a), there were two cases with 45,X/46,XX mosaic and four with inv (9) [inv (9) (p11q13)] among 116 schizophrenics (Nanko et al., 1993). Among the present sample of 134 schizophrenics, we found nine cases with a chromosomal aberration ( Table 1b: one with a karyotype of 45,X/46,XX, two with 47,XXY, and six with inv (9) (p11q13). We could not find any other abnormalities in autosomes except for inv (9). When the previous and the present samples were combined, there were five cases (2.0%) with an aneuploidy of the X chromosome and ten cases (4.0%) with inv (9) among the 250 schizophrenics. When the incidence of inv (9) was compared with that reported by Yamada (1992), who examined 1513 Japanese normal newborns and found 25 cases with inv (9) (1.65%), the difference was statistically significant (x2=6.1, df=1, p=0.013, odds ratio 2.48, 95% confidence interval: 1.18– 5.23). Among the ten schizophrenic probands with inv (9), we could reach some family members of four probands for karyotyping. Case 3 had a son who developed schizophrenia with a karyotype of inv (9); a detailed description of this family is available elsewhere (Lee et al., 1998). Case 13 had a sister and a brother, both of whom were found to have inv (9); the sister was diagnosed with schizophrenia, whereas the brother had no psychiatric problems. The inversions for Cases 11 and 15 were transmitted from his father and her mother, respectively, neither of whom had any psychotic disease.
The first (previous) sample consisting of 120 psychotic subjects [116 schizophrenics according to the DSM-III-R criteria (American Psychiatric Association, 1987) and four patients with delusional disorder] was described elsewhere (Nanko et al., 1993). For the 116 schizophrenic cases (59 men and 57 women), the mean age (standard deviation; SD) was 38.7 (12.2) years and the mean age of onset (first appearance of florid psychotic symptoms) was 25.2 (8.1) years. Seventy-three patients (62.9%) had a history of at least one admission to a psychiatric hospital. The second (present) sample consisted of 134 patients with schizophrenia [63 men and 71 women; mean age (SD): 44.7 years (13.1)] who were crosssectionally selected from the psychiatric clinic of Teikyo University Hospital and psychiatric wards of two associated psychiatric hospitals located in or around the Tokyo metropolitan area during the period from July 1997 to June 1998. Consensus diagnosis according to the DSM-IV criteria for schizophrenia (American Psychiatric Association, 1994) was made for each patient by at least two psychiatrists by using unstructured clinical interviews and all the available information from the medical records. The mean age of onset was 27.9 (SD 10.0) years. All the subjects were unrelated Japanese. One-hundred twenty-four patients (93%) had a history of admission to a psychiatric hospital. 2.2. Cytogenetic examination With written informed consent, a blood sample was drawn from each patient. Peripheral lymphocyte culture and karyotyping were performed by using G-banding by trypsin Giemsa (GTG). Initially, 20 metaphases were examined for all the subjects; then an additional 10 cells were examined when at least one abnormal cell was observed. Mosaicism was determined if the proportion of the minor cell population was more than 15% (i.e., 5 or more in 30 cells), since low-frequency mosaicism has less significance in affecting phenotype, and it may also be due to artifact.
4. Discussion 4.1. Limitations The patients were not diagnosed with structured interviews, although consensus diagnosis by at least two psychiatrists was made based on clinical interviews and information from medical records. The information on family history was obtained from case records when we could not reach family members. In the combined sample, 197 (79%) had
45
H. Kunugi et al. / Schizophrenia Research 40 (1999) 43–47 Table 1 Chromosomal abnormalities in 250 schizophrenics Case (a) First sample (n=116) 1 2 3 4 5 6 (b) Second sample (n=134) 7 8 9 10 11 12 13 14 15
Sex
Age (years)
Age at onset (years)
Family historya
Karyotype
K.O. T.G. Y.K. Y.H. Y.M. M.A.
F F F M M F
59 57 51 20 37 22
35 46 22 20 27 15
+ − + − − −
mos 45,X/46,XX mos 45,X/46,XX 46,XX,Inv(9)(p11q13) 46,XY,Inv(9)(p11q13) 46,XY,Inv(9)(p11q13) 46,XX,Inv(9)(p11q13)
R.H. T.I. H.T. O.T. H.T. M.I. Y.O. R.T. T.K.
M M F M M M F F F
35 46 48 26 33 50 34 28 22
24 15 17 21 29 23 34 15 21
− − + − − − + − −
47,XXY 47,XXY mos 45,X/46,XX 46,XY,Inv(9)(p11q13) 46,XY,Inv(9)(p11q13) 46,XY,Inv(9)(p11q13) 46,XX,Inv(9)(p11q13) 46,XX,Inv(9)(p11q13) 46,XX,Inv(9)(p11q13)
a At least one individual with schizophrenia within the first degree relatives.
a history of admission to a psychiatric hospital; it is likely that severe forms of schizophrenia were overrepresented in our sample. 4.2. The X chromosome aneuploidies in schizophrenia There were five cases with an aneuploidy of the X chromosome: three cases with a karyotype of 45,X/46,XX mosaics and two with 47,XXY. The proportions of the cell populations of 45,X and 46,XX were 15:85, 15:85, and 5:25. In the first sample of 116 schizophrenics, 100 cells were examined for each subject and mosaicism was determined if the proportion of the minor cell population was more than 5%. In the second 134 subjects, only 30 cells were examined; we did not count cases as mosaicism when the minor cell population was less than 15%. Thus we may have underestimated the incidence of 45,X/46,XX, particularly in the second sample. On the other hand, the X chromosome will be lost with normal aging for a proportion of individuals; we cannot entirely exclude this effect of normal aging. Notably, none of these mosaic cases had ever been diagnosed as Turner’s syndrome, probably because the proportions of 45,X were small (15–17%), which had
only a minor effect on phenotype. Nevertheless, the incidence of 45,X/46,XX in our sample (three cases out of 128 female schizophrenics: 2.3%) was unusually high, compared to that reported in the general population [3.3/100 000 according to Hook and Warburton (1983); 1/16 858 according to Ratcliffe et al. (1986)]. Kaplan and Cotton (1968) and Kaplan (1970) also noted that mosaicism of the X chromosome was substantially increased in the schizophrenic females in their studies. For the two cases with 47,XXY, 100% of the cell lines were abnormal. However, here again, these cases had never been diagnosed as Klinefelter’s syndrome until our karyotyping was done. This may be due to the absence of gross anomalies in physical appearance in the syndrome. Furthermore, the cases had never been married, and therefore had no opportunity to suspect their infertility. The incidence of XXY in our sample (2 among 122 males: 1.6%) was unusually high, compared to that in the general population (approx. 1 in 500 males; Smyth and Bremner, 1998). This gives additional evidence for the increased incidence of Klinefelter’s syndrome in schizophrenics [reviewed by Crow (1988) and DeLisi et al. (1994)]. Although Crow (1988) and DeLisi et al. (1994)
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H. Kunugi et al. / Schizophrenia Research 40 (1999) 43–47
pointed out that the incidence of the XXX syndrome in female and that of XYY in male schizophrenics may also be increased, we found no individual with either karyotype. However, given our sample size, the possibility of increased frequency of XXX or XYY cannot be excluded. 4.3. Pericentric inversion of chromosome 9 and schizophrenia Inv (9) is the most common pericentric inversion in humans. Serra et al. (1990) estimated that the prevalence of inv (9) in European or Europeanderived populations was 0.85% among 7613 newborns. In Japanese, the incidence was 1.65% among 1513 normal newborns ( Yamada, 1992) and 1.3% (7/549) among fetuses whose mothers were aged more than 35 years ( Uehara et al, 1992). Ko et al. (1992) reported 1.2% in 1350 Taiwanese fetuses. It could be estimated that the incidence in Asian populations is approx. 1.5%. In addition to the common occurrence of inv (9), no specific phenotype has been demonstrated to be associated with this chromosomal rearrangement. Thus, inv (9) has generally been considered to be a normal variant rather than an abnormal karyotype (Gardner and Sutherland, 1989). However, there are many studies reporting an association of inv (9) with subfertility, recurrent abortions (e.g., Uehara et al., 1992), and abnormal phenotypes. Axelsson and Wahlstro¨m (1984) reported an unusually increased prevalence (9.7%) of inv (9) among male patients with ‘paranoid psychosis’, a proportion of whom might be schizophrenic. In our study, the incidence of inv (9) was consistently increased for both the first (4/116; 3.4%) and the second (6/134; 4.5%) samples relative to those reported in the Japanese general population. Thus, it is unlikely that our observation of increased frequency of inv (9) in schizophrenics has arisen by chance. Although the study of Gorwood et al. (1991) which investigated 25 familial schizophrenics and that of DeLisi et al. (1988) on 46 male schizophrenics did not report any individual with inv (9), these sample sizes were small. If there is an etiological relationship between
inv (9) and schizophrenia, then it could be hypothesized that one of the two breakpoints of inv (9) may disrupt a gene which is causal to schizophrenia. Based on this hypothesis, we previously performed a linkage study on three Japanese schizophrenic families (Nanko et al., 1994). Although we did not obtain a positive linkage between schizophrenia and DNA markers from the pericentric region of chromosome 9, this might be due to the locus heterogeneity or to the small number of families. A more recent study of Moises et al. (1995), which performed an international two-stage genome scan for linkage with schizophrenia, obtained a positive result ( p=0.009) for a marker (D9S175) on the pericentric region of chromosome 9. Furthermore, Levinson et al. (1998) conducted a genome scan and obtained suggestive evidence for a linkage ( p<0.05) between schizophrenia and a marker (D9S257) located 20 cM from the centromere of chromosome 9. A drawback to the possibility of the causal link between inv (9) and schizophrenia is that many family members of our probands who carried the inv (9) had not developed schizophrenia, although there were some familial cases (Cases 3 and 13). It is clear that the effect of inv (9) on the development of schizophrenia would not be major one, but it may be a risk-increasing factor. Recently, inv (9) has been characterized by using fluorescence in situ hybridization (FISH ), showing that there are several different forms of inv (9) which have differential breakpoints (Samonte et al., 1996). This may indicate that phenotypes of inv (9) may vary depending on the location of breakpoints. FISH analyses for the inverted chromosomes found in our subjects are currently underway.
References American Psychiatric Association, 1987. Diagnostic and Statistical Manual of Mental Disorders (DSM-III-R). 3rd ed., revised, American Psychiatric Association, Washington, DC. American Psychiatric Association, 1994. Diagnostic and Statistical Manual of Mental Disorders (DSM-IV ). 4th ed., American Psychiatric Association, Washington, DC.
H. Kunugi et al. / Schizophrenia Research 40 (1999) 43–47 Axelsson, R., Wahlstro¨m, J., 1984. Chromosome aberrations in patients with paranoid psychosis. Hereditas 100, 29–31. Bassett, A.S., 1992. Chromosomal aberrations and schizophrenia. Br. J. Psychiatry 161, 323–334. Crow, T.J., 1988. Sex chromosome and psychosis: the case for a pseudoautosomal locus. Br. J. Psychiatry 153, 675–683. Crow, T.J., DeLisi, L.E., 1998. The chromosome workshops at the 5th International Congress of Psychiatric Genetics—the weight of the evidence from genome scans. Psychiatr. Genet. 8, 59–61. DeLisi, L.E., Friedrich, U., Wahlstrom, J., Boccio-Smith, A., Forsman, A., Eklund, K., Crow, T.J., 1994. Schizophrenia and sex chromosome anomalies. Schizophr. Bull. 20, 495–505. DeLisi, L.E., Reiss, A.L., White, B.J., Gershon, E.S., 1988. Cytogenetic studies of males with schizophrenia: screening for the fragile X chromosome and other chromosomal abnormalities. Schizophr. Res. 1, 277–281. Gardner, R.J.M., Sutherland, G.R., 1989. Chromosome abnormalities and genetic counselling. In: Motulsky, A.G., Bobrow, M., Harper, P.S., Scriver, C. ( Eds.), Oxford Monographs on Medical Genetics. Oxford University Press, Oxford, pp. 68–79. Gorwood, P., Leboyer, M., Hillaire, D., Jay, M., Carteault, F., Dugain, A.-M., Berg, S., Bois, E., Feingold, J., 1991. Cytogenetic studies of familial schizophrenics. Biol. Psychiatry 29, 618–625. Hook, E.B., Warburton, D., 1983. The distribution of chromosomal genotypes associated with Turner’s syndrome: livebirth prevalence rates and evidence for diminished fetal mortality and severity in genotypes associated with structural X abnormalities or mosaicism. Hum. Genet. 64, 24–27. Kaplan, A.R., 1970. Chromosomal mosaicisms and occasional acentric chromosomal fragments in schizophrenic patients. Biol. Psychiatry 2, 89–94. Kaplan, A.R., Cotton, J.E., 1968. Chromosomal abnormalities in female schizophrenics. J. Nerv. Ment. Dis. 147, 402–417. Ko, T.M., Hsieh, F.J., Chang, L.S., Pan, M.F., Lee, T.Y., 1992. Pericentric inversions of chromosome 9 in Taiwanese fetuses. J. Formos. Med. Assoc 91, 473–474. Lee, K.B., Kunugi, H., Nanko, S., 1998. Familial schizophrenia with pericentric inversion of chromosome 9: a case report. Schizophr. Res. 32, 123–126. Levinson, D.F., Mahtani, M.M., Nancarrow, D.J., Brown,
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D.M., Kruglyak, L., Kirby, A., Hayward, N.K., Crowe, R.R., Andreasen, N.C., Black, D.W., Silverman, J.M., Endicott, J., Sharpe, L., Mohs, R.C., Siever, L.J., Walters, M.K., Lennon, D.P., Jones, H.L., Nertney, D.A., Daly, M.J., Gladis, M., Mowry, B.J., 1998. Genome scan of schizophrenia. Am. J. Psychiatry 155, 741–750. Moises, H.W., Yang, L., Kristbjarnarson, H., Wiese, C., Byerley, W., Macciardi, F., Arolt, V., Blackwood, D., Liu, X., Sjo¨gren, B., Aschauer, H.N., Hwu, H.-G., Jang, K., Livesly, W.J., Kennedy, J.L., Zoega, T., Ivarsson, O., Bui, M.-T., Yu, M.-H., Havsteen, B., Commenges, D., Weissenbach, J., Schwinger, E., Gottesman, I.I., Pakstis, A.J., Wetterberg, L., Kidd, K.K., Helgason, T., 1995. An international twostage genome-wide search for schizophrenia susceptibility genes. Nat. Genet. 11, 321–324. Nanko, S., Fukuda, R., Hattori, M., Sasaki, T., Dai, X.Y., Gill, M., Kuwata, S., Shibata, Y., Kazamatsuri, H., 1994. No evidence of linkage or allelic association of schizophrenia with DNA markers at pericentric region of chromosome 9. Biol. Psychiatry 36, 589–594. Nanko, S., Kunugi, H., Sasaki, T., Fukuda, R., Kawate, T., Kazamatsuri, H., 1993. Pericentric region of chromosome 9 is a possible candidate region for linkage study of schizophrenia. Biol. Psychiatry 33, 655–658. Ratcliffe, S.G., Murray, L., Teague, P., 1986. Edinburgh study of growth and development of children with sex chromosome abnormalities III. Birth Defects 22, 73–118. Samonte, R.V., Conte, R.A., Ramesh, K.H., Verma, R.S., 1996. Molecular cytogenetic characterization of breakpoints involving pericentric inversions of human chromosome 9. Hum. Genet. 98, 576–580. Serra, A., Brahe, C., Millington-Ward, A., Neri, G., Tedeschi, B., Tassone, F., Bova, R., 1990. Pericentric inversion of chromosome 9: prevalence in 300 Down syndrome families and molecular studies of nondisjunction. Am. J. Med. Genet. 7, 162–168. Smyth, C.M., Bremner, W.J., 1998. Klinefelter syndrome. Arch. Intern. Med. 158, 1309–1314. Uehara, S., Akai, Y., Takeyama, Y., Takabayashi, T., Okamura, K., Yajima, A., 1992. Pericentric inversion of chromosome 9 in prenatal diagnosis and infertility. Tohoku J. Exp. Med. 166, 417–427. Yamada, K., 1992. Population studies of Inv (9) chromosomes in 4,300 Japanese: incidence, sex difference and clinical significance. Jpn. J. Hum. Genet. 37, 293–301.
Cytogenetic findings in 250 schizophrenics: evidence confirming an excess of the X chromosome aneuploidies and pericentric inversion of chromosome 9 H. Kunugi *, K.B. Lee, S. Nanko Department of Psychiatry, Teikyo University School of Medicine, 11-1, Kaga 2 Chome, Itabashi-ku, Tokyo 173-8605, Japan Received 12 October 1998; accepted for publication 11 February 1999
Abstract Chromosomal abnormalities may be of help in identifying disease genes. To search for susceptibility loci for schizophrenia, we have performed chromosomal examinations by using the GTG banding technique for 250 schizophrenics. We found five cases with an aneuploidy of the X chromosome and ten cases with pericentric inversion of chromosome 9 [inv (9)]. These results confirmed an excess of the X chromosome aneuploidies in schizophrenia, indicating a possible involvement of the X chromosome in the pathogenesis of the illness. The observed incidence (4.0%) of inv (9) in our schizophrenic sample was significantly higher ( p=0.013) than that reported in the general population in Japan (1.7%). Although inv (9) has been considered to be a normal variant, our observation implies a possible association between inv (9) and schizophrenia, suggesting that a susceptibility locus for the disease may be located at a breakpoint of the inversion on chromosome 9. © 1999 Elsevier Science B.V. All rights reserved. Keywords: Chromosome 9; Cytogenetics; Pericentric inversion; Schizophrenia; X chromosome
1. Introduction Although many linkage and association studies have been performed, no gene has yet been established as giving major susceptibility to schizophrenia (Crow and DeLisi, 1998). The use of chromosomal examinations could be an alternative or complementary approach to identify a susceptibility locus for schizophrenia (Bassett, 1992). Chromosomal abnormalities may be helpful to find not only candidate regions for linkage studies, but also valuable materials for positional cloning. In this regard, we previously performed a cytogenetic screening on 120 psychotic subjects (116 * Corresponding author. Tel./fax: +81-3-3964-2447. E-mail address: [email protected] (H. Kunugi)
patients with schizophrenia and four with delusional disorder) (Nanko et al., 1993). In this preliminary report, we found two schizophrenics with 45,X/46,XX mosaic and four with pericentric inversion of chromosome 9 [inv (9)]. Subsequently we have examined a second sample of 134 schizophrenics. Here we report chromosomal findings for the combined sample of 250 schizophrenics. Although several studies have conducted a systematic cytogenetic screening of psychotic subjects, many of these date back to more than two decades ago; thereafter, cytogenetic techniques have been progressing, and diagnostic criteria for psychiatric disorders have been refined. To our knowledge, this is the largest study to examine chromosomal abnormalities for schizophrenics diagnosed by DSM criteria.
0920-9964/99/$ – see front matter © 1999 Elsevier Science B.V. All rights reserved. PII: S0 9 2 0 -9 9 6 4 ( 9 9 ) 0 0 03 5 - 3
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H. Kunugi et al. / Schizophrenia Research 40 (1999) 43–47
2. Methods
3. Results
2.1. Subjects
In our previous sample ( Table 1a), there were two cases with 45,X/46,XX mosaic and four with inv (9) [inv (9) (p11q13)] among 116 schizophrenics (Nanko et al., 1993). Among the present sample of 134 schizophrenics, we found nine cases with a chromosomal aberration ( Table 1b: one with a karyotype of 45,X/46,XX, two with 47,XXY, and six with inv (9) (p11q13). We could not find any other abnormalities in autosomes except for inv (9). When the previous and the present samples were combined, there were five cases (2.0%) with an aneuploidy of the X chromosome and ten cases (4.0%) with inv (9) among the 250 schizophrenics. When the incidence of inv (9) was compared with that reported by Yamada (1992), who examined 1513 Japanese normal newborns and found 25 cases with inv (9) (1.65%), the difference was statistically significant (x2=6.1, df=1, p=0.013, odds ratio 2.48, 95% confidence interval: 1.18– 5.23). Among the ten schizophrenic probands with inv (9), we could reach some family members of four probands for karyotyping. Case 3 had a son who developed schizophrenia with a karyotype of inv (9); a detailed description of this family is available elsewhere (Lee et al., 1998). Case 13 had a sister and a brother, both of whom were found to have inv (9); the sister was diagnosed with schizophrenia, whereas the brother had no psychiatric problems. The inversions for Cases 11 and 15 were transmitted from his father and her mother, respectively, neither of whom had any psychotic disease.
The first (previous) sample consisting of 120 psychotic subjects [116 schizophrenics according to the DSM-III-R criteria (American Psychiatric Association, 1987) and four patients with delusional disorder] was described elsewhere (Nanko et al., 1993). For the 116 schizophrenic cases (59 men and 57 women), the mean age (standard deviation; SD) was 38.7 (12.2) years and the mean age of onset (first appearance of florid psychotic symptoms) was 25.2 (8.1) years. Seventy-three patients (62.9%) had a history of at least one admission to a psychiatric hospital. The second (present) sample consisted of 134 patients with schizophrenia [63 men and 71 women; mean age (SD): 44.7 years (13.1)] who were crosssectionally selected from the psychiatric clinic of Teikyo University Hospital and psychiatric wards of two associated psychiatric hospitals located in or around the Tokyo metropolitan area during the period from July 1997 to June 1998. Consensus diagnosis according to the DSM-IV criteria for schizophrenia (American Psychiatric Association, 1994) was made for each patient by at least two psychiatrists by using unstructured clinical interviews and all the available information from the medical records. The mean age of onset was 27.9 (SD 10.0) years. All the subjects were unrelated Japanese. One-hundred twenty-four patients (93%) had a history of admission to a psychiatric hospital. 2.2. Cytogenetic examination With written informed consent, a blood sample was drawn from each patient. Peripheral lymphocyte culture and karyotyping were performed by using G-banding by trypsin Giemsa (GTG). Initially, 20 metaphases were examined for all the subjects; then an additional 10 cells were examined when at least one abnormal cell was observed. Mosaicism was determined if the proportion of the minor cell population was more than 15% (i.e., 5 or more in 30 cells), since low-frequency mosaicism has less significance in affecting phenotype, and it may also be due to artifact.
4. Discussion 4.1. Limitations The patients were not diagnosed with structured interviews, although consensus diagnosis by at least two psychiatrists was made based on clinical interviews and information from medical records. The information on family history was obtained from case records when we could not reach family members. In the combined sample, 197 (79%) had
45
H. Kunugi et al. / Schizophrenia Research 40 (1999) 43–47 Table 1 Chromosomal abnormalities in 250 schizophrenics Case (a) First sample (n=116) 1 2 3 4 5 6 (b) Second sample (n=134) 7 8 9 10 11 12 13 14 15
Sex
Age (years)
Age at onset (years)
Family historya
Karyotype
K.O. T.G. Y.K. Y.H. Y.M. M.A.
F F F M M F
59 57 51 20 37 22
35 46 22 20 27 15
+ − + − − −
mos 45,X/46,XX mos 45,X/46,XX 46,XX,Inv(9)(p11q13) 46,XY,Inv(9)(p11q13) 46,XY,Inv(9)(p11q13) 46,XX,Inv(9)(p11q13)
R.H. T.I. H.T. O.T. H.T. M.I. Y.O. R.T. T.K.
M M F M M M F F F
35 46 48 26 33 50 34 28 22
24 15 17 21 29 23 34 15 21
− − + − − − + − −
47,XXY 47,XXY mos 45,X/46,XX 46,XY,Inv(9)(p11q13) 46,XY,Inv(9)(p11q13) 46,XY,Inv(9)(p11q13) 46,XX,Inv(9)(p11q13) 46,XX,Inv(9)(p11q13) 46,XX,Inv(9)(p11q13)
a At least one individual with schizophrenia within the first degree relatives.
a history of admission to a psychiatric hospital; it is likely that severe forms of schizophrenia were overrepresented in our sample. 4.2. The X chromosome aneuploidies in schizophrenia There were five cases with an aneuploidy of the X chromosome: three cases with a karyotype of 45,X/46,XX mosaics and two with 47,XXY. The proportions of the cell populations of 45,X and 46,XX were 15:85, 15:85, and 5:25. In the first sample of 116 schizophrenics, 100 cells were examined for each subject and mosaicism was determined if the proportion of the minor cell population was more than 5%. In the second 134 subjects, only 30 cells were examined; we did not count cases as mosaicism when the minor cell population was less than 15%. Thus we may have underestimated the incidence of 45,X/46,XX, particularly in the second sample. On the other hand, the X chromosome will be lost with normal aging for a proportion of individuals; we cannot entirely exclude this effect of normal aging. Notably, none of these mosaic cases had ever been diagnosed as Turner’s syndrome, probably because the proportions of 45,X were small (15–17%), which had
only a minor effect on phenotype. Nevertheless, the incidence of 45,X/46,XX in our sample (three cases out of 128 female schizophrenics: 2.3%) was unusually high, compared to that reported in the general population [3.3/100 000 according to Hook and Warburton (1983); 1/16 858 according to Ratcliffe et al. (1986)]. Kaplan and Cotton (1968) and Kaplan (1970) also noted that mosaicism of the X chromosome was substantially increased in the schizophrenic females in their studies. For the two cases with 47,XXY, 100% of the cell lines were abnormal. However, here again, these cases had never been diagnosed as Klinefelter’s syndrome until our karyotyping was done. This may be due to the absence of gross anomalies in physical appearance in the syndrome. Furthermore, the cases had never been married, and therefore had no opportunity to suspect their infertility. The incidence of XXY in our sample (2 among 122 males: 1.6%) was unusually high, compared to that in the general population (approx. 1 in 500 males; Smyth and Bremner, 1998). This gives additional evidence for the increased incidence of Klinefelter’s syndrome in schizophrenics [reviewed by Crow (1988) and DeLisi et al. (1994)]. Although Crow (1988) and DeLisi et al. (1994)
46
H. Kunugi et al. / Schizophrenia Research 40 (1999) 43–47
pointed out that the incidence of the XXX syndrome in female and that of XYY in male schizophrenics may also be increased, we found no individual with either karyotype. However, given our sample size, the possibility of increased frequency of XXX or XYY cannot be excluded. 4.3. Pericentric inversion of chromosome 9 and schizophrenia Inv (9) is the most common pericentric inversion in humans. Serra et al. (1990) estimated that the prevalence of inv (9) in European or Europeanderived populations was 0.85% among 7613 newborns. In Japanese, the incidence was 1.65% among 1513 normal newborns ( Yamada, 1992) and 1.3% (7/549) among fetuses whose mothers were aged more than 35 years ( Uehara et al, 1992). Ko et al. (1992) reported 1.2% in 1350 Taiwanese fetuses. It could be estimated that the incidence in Asian populations is approx. 1.5%. In addition to the common occurrence of inv (9), no specific phenotype has been demonstrated to be associated with this chromosomal rearrangement. Thus, inv (9) has generally been considered to be a normal variant rather than an abnormal karyotype (Gardner and Sutherland, 1989). However, there are many studies reporting an association of inv (9) with subfertility, recurrent abortions (e.g., Uehara et al., 1992), and abnormal phenotypes. Axelsson and Wahlstro¨m (1984) reported an unusually increased prevalence (9.7%) of inv (9) among male patients with ‘paranoid psychosis’, a proportion of whom might be schizophrenic. In our study, the incidence of inv (9) was consistently increased for both the first (4/116; 3.4%) and the second (6/134; 4.5%) samples relative to those reported in the Japanese general population. Thus, it is unlikely that our observation of increased frequency of inv (9) in schizophrenics has arisen by chance. Although the study of Gorwood et al. (1991) which investigated 25 familial schizophrenics and that of DeLisi et al. (1988) on 46 male schizophrenics did not report any individual with inv (9), these sample sizes were small. If there is an etiological relationship between
inv (9) and schizophrenia, then it could be hypothesized that one of the two breakpoints of inv (9) may disrupt a gene which is causal to schizophrenia. Based on this hypothesis, we previously performed a linkage study on three Japanese schizophrenic families (Nanko et al., 1994). Although we did not obtain a positive linkage between schizophrenia and DNA markers from the pericentric region of chromosome 9, this might be due to the locus heterogeneity or to the small number of families. A more recent study of Moises et al. (1995), which performed an international two-stage genome scan for linkage with schizophrenia, obtained a positive result ( p=0.009) for a marker (D9S175) on the pericentric region of chromosome 9. Furthermore, Levinson et al. (1998) conducted a genome scan and obtained suggestive evidence for a linkage ( p<0.05) between schizophrenia and a marker (D9S257) located 20 cM from the centromere of chromosome 9. A drawback to the possibility of the causal link between inv (9) and schizophrenia is that many family members of our probands who carried the inv (9) had not developed schizophrenia, although there were some familial cases (Cases 3 and 13). It is clear that the effect of inv (9) on the development of schizophrenia would not be major one, but it may be a risk-increasing factor. Recently, inv (9) has been characterized by using fluorescence in situ hybridization (FISH ), showing that there are several different forms of inv (9) which have differential breakpoints (Samonte et al., 1996). This may indicate that phenotypes of inv (9) may vary depending on the location of breakpoints. FISH analyses for the inverted chromosomes found in our subjects are currently underway.
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