Subtelomeric rearrangements in idiopathic mental retardation
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《美国医学杂志》
1 Departments of Pediatrics & Pathology, University of Texas Medical Branch, Galveston, TX, USA
2 Departments of Pediatrics, University of Texas Medical Branch, Galveston, TX, USA
3 Pediatrics, Texas Tech Health Sciences Center, Lubbock, TX, USA
Abstract
Objective: To estimate the frequency of subtelomeric rearrangements in patients with sporadic and non-syndromic idiopathic mental retardation (IMR). Methods: A total of 18 IMR patients were taken for the study. Selection criteria included no known syndromes or chromosomes abnormalities and known causes of IMR. All patients signed an informed consent to participate. Chromosome analysis was carried out on all patients to rule out gross chromosome abnormalities. Lymphocyte cultures were initiated and harvested using standard protocols. For fluorescence in situ hybridization (FISH), Chromoprobe Multiprobe-T system was used. This system consists of 24 embossed areas with each area having one reversibly bound subtelomere probe for a specific chromosome. The subtelomere probes were differentially labeled with green fluorescence for short arm and orange for the long arm. Hybridization, washing and staining are done using standard protocols. A minimum of 5 metaphases were analyzed per chromosome per patient. Results: A total of 2 subtelomeric rearrangements were detected (11.1%). Case 1 involved a 17-year-old with severe MR, profound deafness and dysmorphic features with reciprocal translocation t(3;7)(q26.2; p15.1). The second case involved a 4.6-year-old with mild developmental delay and a terminal deletion of the long arm of chromosome 2, del(2) (q37.3). The frequency of abnormalities detected in our study is in agreement with published reports. Conclusion: Subtelomeric screening with FISH is a useful tool for investigation of IMR, however, it is not cost effective in all cases. Conventional chromosome analysis coupled with targeted FISH testing might be the optimal strategy for investigation of IMR.
Keywords: Idiopathic mental retardation; Subtelomeric rearrangements; FISH
Chromosome abnormalities are the single most common cause of mental retardation (MR) in humans. Approximately 40% of cases with severe idiopathic MR (IMR) have chromosome abnormalities, while the frequency is about 10% for mild IMR.[1] Because of the unique structural and functional characteristics of telomeres such as their enrichment for pseudogenes and genes leading to mispairing during meiosis[2],[3],[4],[5],[6],[7],[8] increased recombination rates at telomeres.[9], [10] It has been suggested that imbalances involving these regions might be a significant contributor to IMR.[1]
Recent evidence suggests that approximately 4-35% of cases with IMR have subtelomeric rearrangements. [11],[12],[13],[14],[15],[16],[17],[18],[19],[20],[21],[22],[23],[24],[25],[26] Most of the subtelomere studies are partially retrospective, thus can be biased in favor of familial cases. There was only one prospective study of consecutive group of children with MR of unknown etiology[27] (van Karnebeek et al., 2002). Contrary to the published reports, this study showed that the frequency of subtelomeric rearrangements detected using fluorescence in situ hybridization (FISH) is very low (0.5%). Because of the conflicting reports and wide variations in the frequency of such rearrangements, a study has been initiated to screen IMR patients with or without dysmorphic features and no family history of MR using subtelomeric FISH panels.
Materials and Methods
Selection of patients
For the study patients were recruited from the child development and genetics clinics. The cohort consisted of 20 patients with MR of unknown etiology, all of whom were younger than 18 years of age. All the patients are clinically evaluated by either developmental pediatricians and/or clinical geneticist. As such those patients that fit any of the distinct clinical syndromes such as Down, Prader-Willi, DiGeorge, fragile (X), etc., are excluded from this study. Selection criteria included no known syndromes or chromosome abnormalities and known causes of their MR. Standard assessment of all patients included routine diagnostic tests including chromosome analysis to rule out major abnormalities; prenatal, perinatal and postnatal history, family history to rule out mental retardation; psychiatric disorders; neurological examination and behavioral assessment. All patients signed an informed consent to participate.
Cytogenetic and molecular cytogenetic methods
To rule out gross abnormalities, chromosome analysis was carried out on all patients either in authors' laboratory or in an outside laboratory. Lymphocyte cultures were initiated and harvested using standard protocols. Cell suspension with chromosomes was dropped onto template slides with 24 chambers. For FISH studies, Chromoprobe Multiprobe-T system (Cytocell Inc., Banbury, UK) was used. This system consists of 24 embossed areas with each area having one reversibly bound subtelomeric probe for a specific chromosome. The subtelomeric probes are differentially labeled with green fluorescence for short arm and orange for long arm. Hybridization, washing and staining are done using standard protocols. A minimum of 5 metaphases were analyzed per chromosome per patient. Only those metaphases where both p arm and q arm signals can be seen were considered for analysis. Results were considered conclusive if the first 5 metaphases showed 100% concordant results. If a probe on the multiprobe device could not be scored, FISH was performed separately using the specific subtelomeric probe from the same manufacturer. Similarly all the abnormal results were reconfirmed using chromosome specific telomere probes and paint probes. Parental chromosome and FISH analyses were carried out when ever possible in those cases where an abnormality was detected. Photographic documentation is maintained for all abnormal results.
Results
A summary of the cytogenetic, subtelomere FISH results, demographic data and brief description of major clinical features of all the patients is presented in table1. In the majority of the cases (16/18) a normal karyotype and normal subtelomere FISH results were observed. The remaining 2 cases (2/18 = 11.1%) showed abnormal subtelomere FISH results. Of the 2 cases with chromosome rearrangements, case GC was previously described. [27] In brief, this patient is 17 year old with severe developmental delay, seizures, mental retardation and several dysmorphic features. He also had bilateral hearing loss and severe visual impairment. Subtelomere FISH and subsequent chromosome analysis showed that he has a balanced translocation between the long arm of chromosome 3 and the short arm of chromosome 7 Figure1. The karyotype was interpreted as 46,XY, t(3;7) (q27; p21.2). Subsequent FISH studies with whole chromosome paint probes confirmed the rearrangement. The second case (case TM) involved a 5-year-old with isolated developmental delay. Subtelomere FISH analysis showed a terminal deletion of the long arm of chromosome 2 Figure2. Subsequent repeat analysis with Vysis probe for 2q telomere confirmed the deletion.
Of the 18 cases studied with subtelomere probe panels, 8 (44.4%) patients were reported to have severe developmental delay, while 9 (50%) have moderate to mild developmental delay. Five (27.8%) patients had dysmorphic features including epicanthal folds, broad nasal bridge, anteverted nares, high arched palate, cleft palate and posteriorly rotated ears. Most of the patients had no family history of mental retardation, though some have family members affected with Down syndrome. One mother and son in the present cohort were developmentally delayed with speech difficulties. In this family, the mother is described as slow and the son has moderate developmental delay. Both of them showed normal results with subtelomere probes. Some of the patients had several investigations including targeted FISH studies to rule out microdeletions.
Discussion
The frequency of subtelomeric rearrangements varies considerably from 4 to 35% in reported studies, but the general consensus is that segmental aneusomy of the subtelomeric regions accounts for 6% of patients with mental retardation/developmental delay and/or non-specific dysmorphic features.[15],[23],[25],[28] The studies' wide ranging results have been attributed to differences in one or more of the following: inclusion criteria, recruitment methodology, clinical evaluation of study subjects, quality of chromosome preparations and different laboratory methods used to identify the subtelomeric rearrangements.[26] Most of the studies with high frequency of subtelomeric rearrangements were retrospective, and as such there is likelihood for selection bias towards patients who are likely to have chromosome abnormalities and inclusion of patients with classical microdeletion syndromes and or known chromosomal abnormalities. For these reasons, the data regarding the frequency of subtelomeric rearrangements in IMR are not considered a true reflection of the population frequency. Thus, investigators have not come to agreement about the true prevalence of subtelomeric rearrangements.
The two abnormal cases observed in the present cohort were referred to us from an outside laboratory with normal chromosome analysis reports. Only after detection of abnormalities with subtelomeric probes, was chromosomal analysis performed again, and then high resolution GTG banding analysis showed the abnormality even at the chromosome level. Thus if we exclude the two cases the actual frequency of subtelomere rearrangements detected in the present would be 0%. These results are in agreement with published reports[1] indicating that subtelomeric imbalance is not a significant cause of IMR. Joyce et al[1] further suggested that such subtelomeric rearrangements might be a common finding in the general population. These authors have suggested that the presence of subtelomeric rearrangements in normal individuals might be confined to those regions of the genome with few functional genes. There has been increasing evidence of non-pathogenic euchromatic rearrangements that are visible at cytogenetic levels[29],[30],[31] and such rearrangements represent perhaps normal population variants without clinical significance. Several studies have shown that such polymorphic variants are present within the subtelomeric regions. [32],[33],[34]
One of the best-known polymorphic variants is the 2q subtelomeric deletion.[33],[34] The subtelomeric region on distal 2q has been known to exhibit a chromosome length polymorphism.[35] Detection of a deletion depends on the subtelomeric probe used. Some probes may detect a deletion while other probes may not.[36] One of the abnormalities seen in the present cohort was the same distal 2q deletion. Using Vysis 2q subtelomeric probe, the authors confirmed the presence of deletion, but were unable to perform parental studies due to lost contact with this family. Recent subtelomeric studies in children with autism have suggested that there might be gene(s) present on the distal region of chromosome 2q that predispose to autism.[36] Buxbaum et al also suggested such a possible linkage to distal 2q in autism.[37] In the present cohort of 18 patients, one patient was detected with deletion of distal 2q, and this patient is also thought to have autistic behavior. However, if we consider the clinical phenotype of the 18 patients in the present study, 8 patients had autistic behavior (44.4%) but only one showed 2q deletion. Thus the frequency of 2q deletion in autistic children in the present cohort was 12.5%(1/8) and this is similar to the results reported by Wolff et al.[36] The second subtelomeric rearrangement seen in the present cohort involved a balanced reciprocal translocation. Since most reported subtelomeric studies have shown segmental anueploidy to be the cause of IMR, this may indicate that the present case is unique.
The frequency of subtelomeric rearrangements seen in our cohort is very low compared to other published reports. Ours is only the second report of such low frequency of subtelomeric rearrangements. The difference can be explained by differences of methodology: (1) None of the patients in our cohort have a family history of MR. Studies have shown that significant differences exists in the frequency of abnormalities detected when patients with and without family history are compared and (2) In the present cohort, only 5 of the 18 patients (28%) of the study had dysmorphic features.
When patients with dysmorphic features are included in studies, the yield of subtelomeric rearrangements is expected to be higher.[23],[38] Indeed, the fact that in the present study one of the two subtelomeric rearrangements is seen in a patient with several dysmorphic features further strengthens this hypothesis. (3) The authors have not included known chromosomal abnormalities or syndromes in the present cohort, and thus in the present study frequency is lower than other studies with less strict inclusion criteria.
In summary, in the present limited study of 18 patients with non-syndromic developmental delay/mental retardation/and/or non-specific dysmorphic features, the authors observed a very low frequency of subtelomeric rearrangements. This is in accordance with other published reports with strict inclusion criteria.[1], [26] Based on the authors experience and the literature, it is felt that the best approach to diagnosis of segmental aneusomy in IMR/developmental delay/non-specific dysmorphic features is high resolution chromosome analysis followed by targeted FISH studies. Subtelomeric FISH studies might still be warranted and are useful in those cases where there is family history or considerable dysmorphic features associated with severe MR or developmental delay.
Acknowledgements
The authors express their gratitude to the families of children included in this study. The authors are also grateful to Shuliu Zhang, Jill Northup, and Judy Hawkins for the excellent technical assistance and discussions. Some part of this study is supported by funds from Texas Department of Health to Drs. LLH and GVNV.
References
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10. Laurie DA, Hulten MA. Further studies on bivalent chiasma frequency in human males with normal karyotypes. Ann Hum Genet 1985; 49: 189-201.
11. Flint J, Wilkie AO, Buckle VJ, Winter RM, Holland AJ, McDermid HE. The detection of subtelomeric chromosomal rearrangements in idiopathic mental retardation. Nat Genet 1995; 9: 132-140.
12. Hoovers JM, Bijlsma EK, Sluijter S, Hennekam RCM. Subtelomeric rearrangements as a cause for mental retardation. Eur J Hum Genet 1998; 6: 41-42.
13. Viot G, Gosset P, Fert S, Prieur M, Turleau C, Raoul O, De Blois MC, Lyonnet S, Munnich A, Vekemans M. Cryptic subtelomeric rearrangements detected by FISH in mentally retarted and dysmorphic patients. Am J Hum Genet 1998; 63: A44.
14. Vorsanova SG, Yurov YB, Demidova IA, Kolotii AD, Soloviev IV. FISH analysis of microaberrations near telomeric chromosomal regions in children with mental retardation. Am J Med Genet 1998; 96: A345.
15. Knight SJL, Regan R, Nicod A, Horsley SW, Kearney L, Homfray T, Winter RM, Bolton P, Flynt J. Subtle chromo-somal rearrangements in children with unexplained mental retardation. Lancet 1999; 354: 1676-1681.
16. Lamb AN, Lytle CH, Aylsworth AS, Powel CM, Rao KW, Hendrickson M, Carey JC, Opitz JM, Viskochil DH, Leonard CO, Brothman AR, Stephan M, Bartley JA, Hackbarth M, McCarthy D, Proffitt J. Low proportion of subtelomeric rearrangements in a population of patients with mental retardation and dysmorphic features. Am J Hum Genet 1999;65: A924.
17. Slavotinek A, Rosenberg M, Knight S, Gaunt L, Fergusson W, Killoran C, Clayton-Smith J, Kingston H, Campbell RHA, Flint J, Donnai D, Biesecker L. Screening for submicroscopic chromosome rearrangements in children with idiopathic mental retardation using microsatellite markers for the chromosome telomeres. J Med Genet 1999; 36: 405-411.
18. Colleaux L, Rio M, Heuertz S, Moindrault S, Turleau C, Ozilou C, Gosset P, Raoult O, Lyonnet S, Cormier-Daire V, Amiel J, Le Merrer M, Picq M, de Blois MC, Prieur M, Romana S, Cornelis F, Vekemans M, Munnich A. A novel automated strategy for screening cryptic telomeric rearrangements in children with idiopathic mental retardation. Eur J Hum Genet 2001; 9: 319-327.
19. Kirchhoff M, Rose H, Lundsteen C. High resolution comparative genomic hybridization in clinical cytogenetics. J Med Genet 2001; 38: 740-744.
20. Riegel M, Baumer A, Jamar M, Delbecque K, Herens C, Verloes A, Schinzel A. Submicroscopic terminal deletions and duplications in retarded patients with unclassified malformation syndromes. Hum Genet 2001; 109: 286-294.
21. Rosenberg MJ, Killoran C, Dziadzio L, Chang S, Stone DL, Meck J, Aughton D, Bird LM, Bodurtha J, Cassidy SB, Graham JM Jr., Grix A, Guttmarcher AE, Hudgins L, Kozma C, Michaelis RC, Pauli R, Peters KF, Rosenbaum KN, Tifft CJ, Wargowski D, Williams MS, Biesecker LG. Scanning for telomeric deletions and duplications and uniparental disomy using genetic markers in 120 children with malformations. Hum Genet 2001; 109: 311-318.
22. Rossi E, Piccini F, Zollino M, Neri G, Caselli D, Tenconi R, Castellan C, Carrozzo R, Danesino C, Zuffardi O, Ragusa A, Castiglia L, Galesi O, Greco D, Romano C, Pierluigi M, Perfume C, Di Rocco M, Faravelli F, Dagna BF, Bonaglia M, Bedeschi M, Borgatti R. Cryptic telomeric rearrangements in subjects with mental retardation associated with dysmorphism and congenital malformations. J Med Genet 2001; 38: 417-420.
23. Anderlid B-M, Schoumans J, Annerιn G, Sahlιn S, Kyllerman M, Vujic M, Hagberg B, Blennow E, Nordenskj φld M. Subtelomeric rearrangements detected in patients with idiopathic mental retardation. Am J Med Genet 2002; 107: 275-284.
24. Baker E, Hinton L, Callen DF, Altree M, Dobbie A, Eyre HJ, Sutherland GR, Thompson E, Thompson P, Woollatt E, Haan E. Study of 250 children with idiopathic mental retardation reveals nine cryptic and diverse subtelomeric chromosome anomalies. Am J Med Genet 2002; 107: 285-293.
25. Clarkson B, Pavenski K, Dupuis L, Kennedy S, Meyn S, Nezrati MM, Nie G, Weksberg R, Withers S, Quercia N, Teebi AS, Teshima I. Detecting rearrangements in children using subtelomeric FISH and SKY. Am J Med Genet 2002; 107: 267-274.
26. van Karnebeek CDM, Koevoets C, Sluijter S, Bijlsma EK, Smeets DFMC, Redeker EJ, Hennekam RCM, Hoovers JMN. Prospective screening for subtelomeric rearrangements in children with mental retardation of unknown aetiology: the Amsterdam experience. J Med Genet 2002; 39: 546-553.
27. Tonk VS, Wyandt HE, Huang X, Patel N, Morgan DL, Kukolich M, Lockhart LH, Velgaleti GVN. Disease associated balanced chromosome rearrangements (DBCR): report of two new cases. Ann Genet 2003; 46: 37-43.
28. Jalal SM, Harwood AR, Sekhon GS, Lorentz CP, Ketterling RP, Babovic-Vuksanovic D, Meyer RG, Ensenauer R, Anderson MH, Michels VV. Utility of subtelomeric fluorescent DNA probes for detection of chromosome anomalies in 425 patients. Genet Med 2003; 5: 28-34.
29. Barber JCK, Joyce CA, Collinson MN, Nicholson JC, Willatt LR, Dyson HM, Bateman MS, Green AJ, Yates JRW, Dennis NR. Duplication of 8p23.1: a cytogenetic anomaly with no established clinical significance. J Med Genet 1998; 35: 491-496.
30. Barber JCK, Reed CJ, Dahoun SP, Joyce CA. Amplification of a pseudogene cassette underlies euchromatic variation of 16p at the cytogenetic level. Hum Genet 1999;104:211-218.
31. Gardner RJM, Sutherland GR. Chromosome abnormalities and genetic counseling. Oxford University Press, Oxford. 1996.
32. Harwood A, Anderson M, Jalal SM. Technical comparison of two commercially available telomere specific probe set. J Assoc Genet Tech 2001; 27: 132-134.
33. Jalal SM, Harwood AR, Anderson MH, Sekhon GS, Ketterling RP, Michels VV. Analysis by telomere specific FISH probes of 191 karyotypically normal patients with nonspecific dysmorphic features or developmental delay, or history of multiple miscarriages. Am J Hum Genet 2001; 69: 333.
34. Fan Y-S, Zhang Y, Speevak M, Farrell S, Jung JH, Siu VM. Detection of submicroscopic aberrations in patients with unexplained mental retardation by fluorescence in situ hybridization using multiple subtelomeric probes. Genet Med 2001; 3: 416-421.
35. Macina RA, Negorev DG, Spais C, Ruthig LA, Hu X-L, Riethman HC. Sequence organization of the human chromosome 2q telomere. Hum Mol Genet 1994; 3: 1847-1853.
36. Wolff DJ, Clifton K, Karr C, Charles J. Pilot assessment of the subtelomeric regions of children with autism: Detection of a 2q deletion. Genet Med 2002; 4: 10-14.
37. Buxbaum JD, Silverman JM, Smith CJ, Kilifarski M, Reichert J, Hollander E, Lawlor BA, Fitzgerald M, Greenberg DA, Davis KL. Evidence for a susceptibility gene for autism on chromosome 2 and for genetic heterogeneity. Am J Hum Genet 2001; 68: 1514-1520.
38. de Vries BB, White SM, Knight SJ, Regan R, Homfray T, Young ID, Super M, McKeown, Splitt M, Quarrell OWJ, Trainer AH, Niermeijer MF, Malcolm S, Flint J, Hurst JA, Winter RM. Clinical studies on submicroscopic subtelomeric rearrangements: a check list. J Med Genet 2001; 38: 145-150.(Velagaleti Gopalrao V N, )
2 Departments of Pediatrics, University of Texas Medical Branch, Galveston, TX, USA
3 Pediatrics, Texas Tech Health Sciences Center, Lubbock, TX, USA
Abstract
Objective: To estimate the frequency of subtelomeric rearrangements in patients with sporadic and non-syndromic idiopathic mental retardation (IMR). Methods: A total of 18 IMR patients were taken for the study. Selection criteria included no known syndromes or chromosomes abnormalities and known causes of IMR. All patients signed an informed consent to participate. Chromosome analysis was carried out on all patients to rule out gross chromosome abnormalities. Lymphocyte cultures were initiated and harvested using standard protocols. For fluorescence in situ hybridization (FISH), Chromoprobe Multiprobe-T system was used. This system consists of 24 embossed areas with each area having one reversibly bound subtelomere probe for a specific chromosome. The subtelomere probes were differentially labeled with green fluorescence for short arm and orange for the long arm. Hybridization, washing and staining are done using standard protocols. A minimum of 5 metaphases were analyzed per chromosome per patient. Results: A total of 2 subtelomeric rearrangements were detected (11.1%). Case 1 involved a 17-year-old with severe MR, profound deafness and dysmorphic features with reciprocal translocation t(3;7)(q26.2; p15.1). The second case involved a 4.6-year-old with mild developmental delay and a terminal deletion of the long arm of chromosome 2, del(2) (q37.3). The frequency of abnormalities detected in our study is in agreement with published reports. Conclusion: Subtelomeric screening with FISH is a useful tool for investigation of IMR, however, it is not cost effective in all cases. Conventional chromosome analysis coupled with targeted FISH testing might be the optimal strategy for investigation of IMR.
Keywords: Idiopathic mental retardation; Subtelomeric rearrangements; FISH
Chromosome abnormalities are the single most common cause of mental retardation (MR) in humans. Approximately 40% of cases with severe idiopathic MR (IMR) have chromosome abnormalities, while the frequency is about 10% for mild IMR.[1] Because of the unique structural and functional characteristics of telomeres such as their enrichment for pseudogenes and genes leading to mispairing during meiosis[2],[3],[4],[5],[6],[7],[8] increased recombination rates at telomeres.[9], [10] It has been suggested that imbalances involving these regions might be a significant contributor to IMR.[1]
Recent evidence suggests that approximately 4-35% of cases with IMR have subtelomeric rearrangements. [11],[12],[13],[14],[15],[16],[17],[18],[19],[20],[21],[22],[23],[24],[25],[26] Most of the subtelomere studies are partially retrospective, thus can be biased in favor of familial cases. There was only one prospective study of consecutive group of children with MR of unknown etiology[27] (van Karnebeek et al., 2002). Contrary to the published reports, this study showed that the frequency of subtelomeric rearrangements detected using fluorescence in situ hybridization (FISH) is very low (0.5%). Because of the conflicting reports and wide variations in the frequency of such rearrangements, a study has been initiated to screen IMR patients with or without dysmorphic features and no family history of MR using subtelomeric FISH panels.
Materials and Methods
Selection of patients
For the study patients were recruited from the child development and genetics clinics. The cohort consisted of 20 patients with MR of unknown etiology, all of whom were younger than 18 years of age. All the patients are clinically evaluated by either developmental pediatricians and/or clinical geneticist. As such those patients that fit any of the distinct clinical syndromes such as Down, Prader-Willi, DiGeorge, fragile (X), etc., are excluded from this study. Selection criteria included no known syndromes or chromosome abnormalities and known causes of their MR. Standard assessment of all patients included routine diagnostic tests including chromosome analysis to rule out major abnormalities; prenatal, perinatal and postnatal history, family history to rule out mental retardation; psychiatric disorders; neurological examination and behavioral assessment. All patients signed an informed consent to participate.
Cytogenetic and molecular cytogenetic methods
To rule out gross abnormalities, chromosome analysis was carried out on all patients either in authors' laboratory or in an outside laboratory. Lymphocyte cultures were initiated and harvested using standard protocols. Cell suspension with chromosomes was dropped onto template slides with 24 chambers. For FISH studies, Chromoprobe Multiprobe-T system (Cytocell Inc., Banbury, UK) was used. This system consists of 24 embossed areas with each area having one reversibly bound subtelomeric probe for a specific chromosome. The subtelomeric probes are differentially labeled with green fluorescence for short arm and orange for long arm. Hybridization, washing and staining are done using standard protocols. A minimum of 5 metaphases were analyzed per chromosome per patient. Only those metaphases where both p arm and q arm signals can be seen were considered for analysis. Results were considered conclusive if the first 5 metaphases showed 100% concordant results. If a probe on the multiprobe device could not be scored, FISH was performed separately using the specific subtelomeric probe from the same manufacturer. Similarly all the abnormal results were reconfirmed using chromosome specific telomere probes and paint probes. Parental chromosome and FISH analyses were carried out when ever possible in those cases where an abnormality was detected. Photographic documentation is maintained for all abnormal results.
Results
A summary of the cytogenetic, subtelomere FISH results, demographic data and brief description of major clinical features of all the patients is presented in table1. In the majority of the cases (16/18) a normal karyotype and normal subtelomere FISH results were observed. The remaining 2 cases (2/18 = 11.1%) showed abnormal subtelomere FISH results. Of the 2 cases with chromosome rearrangements, case GC was previously described. [27] In brief, this patient is 17 year old with severe developmental delay, seizures, mental retardation and several dysmorphic features. He also had bilateral hearing loss and severe visual impairment. Subtelomere FISH and subsequent chromosome analysis showed that he has a balanced translocation between the long arm of chromosome 3 and the short arm of chromosome 7 Figure1. The karyotype was interpreted as 46,XY, t(3;7) (q27; p21.2). Subsequent FISH studies with whole chromosome paint probes confirmed the rearrangement. The second case (case TM) involved a 5-year-old with isolated developmental delay. Subtelomere FISH analysis showed a terminal deletion of the long arm of chromosome 2 Figure2. Subsequent repeat analysis with Vysis probe for 2q telomere confirmed the deletion.
Of the 18 cases studied with subtelomere probe panels, 8 (44.4%) patients were reported to have severe developmental delay, while 9 (50%) have moderate to mild developmental delay. Five (27.8%) patients had dysmorphic features including epicanthal folds, broad nasal bridge, anteverted nares, high arched palate, cleft palate and posteriorly rotated ears. Most of the patients had no family history of mental retardation, though some have family members affected with Down syndrome. One mother and son in the present cohort were developmentally delayed with speech difficulties. In this family, the mother is described as slow and the son has moderate developmental delay. Both of them showed normal results with subtelomere probes. Some of the patients had several investigations including targeted FISH studies to rule out microdeletions.
Discussion
The frequency of subtelomeric rearrangements varies considerably from 4 to 35% in reported studies, but the general consensus is that segmental aneusomy of the subtelomeric regions accounts for 6% of patients with mental retardation/developmental delay and/or non-specific dysmorphic features.[15],[23],[25],[28] The studies' wide ranging results have been attributed to differences in one or more of the following: inclusion criteria, recruitment methodology, clinical evaluation of study subjects, quality of chromosome preparations and different laboratory methods used to identify the subtelomeric rearrangements.[26] Most of the studies with high frequency of subtelomeric rearrangements were retrospective, and as such there is likelihood for selection bias towards patients who are likely to have chromosome abnormalities and inclusion of patients with classical microdeletion syndromes and or known chromosomal abnormalities. For these reasons, the data regarding the frequency of subtelomeric rearrangements in IMR are not considered a true reflection of the population frequency. Thus, investigators have not come to agreement about the true prevalence of subtelomeric rearrangements.
The two abnormal cases observed in the present cohort were referred to us from an outside laboratory with normal chromosome analysis reports. Only after detection of abnormalities with subtelomeric probes, was chromosomal analysis performed again, and then high resolution GTG banding analysis showed the abnormality even at the chromosome level. Thus if we exclude the two cases the actual frequency of subtelomere rearrangements detected in the present would be 0%. These results are in agreement with published reports[1] indicating that subtelomeric imbalance is not a significant cause of IMR. Joyce et al[1] further suggested that such subtelomeric rearrangements might be a common finding in the general population. These authors have suggested that the presence of subtelomeric rearrangements in normal individuals might be confined to those regions of the genome with few functional genes. There has been increasing evidence of non-pathogenic euchromatic rearrangements that are visible at cytogenetic levels[29],[30],[31] and such rearrangements represent perhaps normal population variants without clinical significance. Several studies have shown that such polymorphic variants are present within the subtelomeric regions. [32],[33],[34]
One of the best-known polymorphic variants is the 2q subtelomeric deletion.[33],[34] The subtelomeric region on distal 2q has been known to exhibit a chromosome length polymorphism.[35] Detection of a deletion depends on the subtelomeric probe used. Some probes may detect a deletion while other probes may not.[36] One of the abnormalities seen in the present cohort was the same distal 2q deletion. Using Vysis 2q subtelomeric probe, the authors confirmed the presence of deletion, but were unable to perform parental studies due to lost contact with this family. Recent subtelomeric studies in children with autism have suggested that there might be gene(s) present on the distal region of chromosome 2q that predispose to autism.[36] Buxbaum et al also suggested such a possible linkage to distal 2q in autism.[37] In the present cohort of 18 patients, one patient was detected with deletion of distal 2q, and this patient is also thought to have autistic behavior. However, if we consider the clinical phenotype of the 18 patients in the present study, 8 patients had autistic behavior (44.4%) but only one showed 2q deletion. Thus the frequency of 2q deletion in autistic children in the present cohort was 12.5%(1/8) and this is similar to the results reported by Wolff et al.[36] The second subtelomeric rearrangement seen in the present cohort involved a balanced reciprocal translocation. Since most reported subtelomeric studies have shown segmental anueploidy to be the cause of IMR, this may indicate that the present case is unique.
The frequency of subtelomeric rearrangements seen in our cohort is very low compared to other published reports. Ours is only the second report of such low frequency of subtelomeric rearrangements. The difference can be explained by differences of methodology: (1) None of the patients in our cohort have a family history of MR. Studies have shown that significant differences exists in the frequency of abnormalities detected when patients with and without family history are compared and (2) In the present cohort, only 5 of the 18 patients (28%) of the study had dysmorphic features.
When patients with dysmorphic features are included in studies, the yield of subtelomeric rearrangements is expected to be higher.[23],[38] Indeed, the fact that in the present study one of the two subtelomeric rearrangements is seen in a patient with several dysmorphic features further strengthens this hypothesis. (3) The authors have not included known chromosomal abnormalities or syndromes in the present cohort, and thus in the present study frequency is lower than other studies with less strict inclusion criteria.
In summary, in the present limited study of 18 patients with non-syndromic developmental delay/mental retardation/and/or non-specific dysmorphic features, the authors observed a very low frequency of subtelomeric rearrangements. This is in accordance with other published reports with strict inclusion criteria.[1], [26] Based on the authors experience and the literature, it is felt that the best approach to diagnosis of segmental aneusomy in IMR/developmental delay/non-specific dysmorphic features is high resolution chromosome analysis followed by targeted FISH studies. Subtelomeric FISH studies might still be warranted and are useful in those cases where there is family history or considerable dysmorphic features associated with severe MR or developmental delay.
Acknowledgements
The authors express their gratitude to the families of children included in this study. The authors are also grateful to Shuliu Zhang, Jill Northup, and Judy Hawkins for the excellent technical assistance and discussions. Some part of this study is supported by funds from Texas Department of Health to Drs. LLH and GVNV.
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