The correlation of age of onset with CTG trinucleotide repeat amplification in myotonic dystrophy.

The gene for myotonic dystrophy (DM) has recently been isolated and amplification of an unstable CTG trinucleotide repeat, located within the DM gene, has been identified in virtually all patients studied to date. A high proportion of DM families who are studied show a progressively earlier age of onset with succeeding generations and, in the few pedigrees reported so far, an increasing degree of amplification of the CTG repeat has been noted to parallel this trend. It has been implicit in several of the original reports on the nature of the changes in the DM gene that knowledge of CTG amplification status at the DM locus of a person will provide useful information concerning prognosis. However, no studies of genotype-phenotype correlation have been reported and there are no specific data on which to base such counsel. In this paper we report the correlation between the degree of CTG amplification and age of onset in 109 DM gene carriers from 17 families. Included are parent-child and sib-sib comparisons which provide a framework in which to incorporate DNA diagnostic studies when counselling subjects and families at risk for DM.     

Somatic instability of the myotonic dystrophy (CTG)n repeat during human fetal development

Myotonic dystrophy is characterised by the striking level of somatic heterogeneity seen between and within tissues of the same patient, which probably accounts for a significant proportion of the pleiotropy associated with this disorder. The congenital form of the disease is associated with the largest (CTG)n repeat expansions. We have investigated the timing of instability of myotonic dystrophy (CTG)n repeats in a series of congenitally affected fetuses and neonates. We find that during the first trimester the repeat is apparently stable and that instability only becomes detectable during the second and third trimesters. In our series repeat instability is apparent only after 13 weeks gestational age and before 16 weeks. The appearance of heterogeneity shows some tissue specificity, with heart most commonly having the largest expansion. The degree of heterogeneity is not correlated with initial expansion size as gauged by chorionic villus and blood (CTG)n repeat sizes.      

Caught in the middle: two genes troubled by trinucleotide expansion in myotonic dystrophy

Trinucleotide repeat expansion at the myotonic dystrophy locus reduces expression of DMAHP Klesert et al. (1997) Nat Genet 16: 402–406
Expansion of the myotonic dystrophy CTG repeat reduces expression of the flanking DMAHP gene Thornton et al. (1997) Nat Genet 16: 407–409     

一个非CTG非CCTG重复扩展型强直性肌营养不良家系

目的　探讨一个强直性肌营养不良 (myotonic dystrophy,DM)家系的分子遗传学基础。方法　采用长模板 PCR扩增、Southern印迹技术和特定染色体区域的基因组扫描技术 ,检测一个强直性肌营养不良家系成员 DMPK基因中 CTG及 ZNF9基因中 CCTG短串重复拷贝数。结果　家系中接受检测的2 6个成员中 ,两种序列的重复拷贝数均在正常范围内 ,即 :(1)该家系中无 DM1DMPK基因 (CTG) n重复数的异常增加 ;(2 )该家系中无 DM2 ZNF9基因 (CCTG) n重复的异常增加 ;(3)受检者相隔 4年的两次血样标本中 ,DMPK基因 (CTG) n重复的拷贝数无明显差异。DMPK及 ZNF9两侧微卫星标志的 L od值均小于 1。结论　除 CTG、CCTG短串联重复序列异常扩展外 ,可能存在新的强直性肌营养不良的致病基因。     

CTG repeat instability in a human embryonic stem cell line carrying the myotonic dystrophy type 1 mutation

Human embryonic stem cells (hESC) are considered to be an indefinite source of self-renewing cells that can differentiate into all types of cells of the human body and could be used in regenerative medicine, drug discovery and as a model for studying early developmental biology. hESC carrying disease-causing mutations hold promise as a tool to investigate mechanisms involved in the pathogenesis of the disease. In this report, we describe the behaviour of an expanded CTG repeat in the 3' untranslated region of the DMPK gene in VUB03_DM1, a hESC line carrying the myotonic dystrophy type 1 (DM1) mutation compared with the normal CTG repeat in two hESC lines VUB01 and VUB04_CF. Expanded CTG repeats were detected by small amount PCR, small pool PCR and Southern blot analysis in consecutive passages of VUB03_DM1. An important instability of the CTG repeat was detected during prolonged in vitro culture, showing stepwise increases of the repeat number in consecutive passages as well as a higher range of variability. This variability was present in cells of different colonies of the same passage and even within single colonies. The high repeat instability is in contrast to the previously observed stability of the repeat in preimplantation embryos and in fetuses during the first trimester of pregnancy. This in vitro culture of affected hESC represents a valuable model for studying the biology of repeat instability.     

Correlations between individual clinical manifestations and CTG repeat amplification in myotonic dystrophy

Myotonic dystrophy (DM) is a multisystemic disease caused by the expansion of a CTG repeat, located in the 3'-untranslated region of the DMPK gene. The number of CTG repeats broadly correlates with the overall severity of the disease. However, correlations between CTG repeat number and presence/absence or severity of individual clinical manifestations in the same patients are yet scarce. In this study the number of CTG repeats detected in blood cells of 24 DM subjects was correlated with the severity of single clinical manifestations. The presence/absence of muscular atrophy, respiratory insufficiency, cardiac abnormalities, diabetes, cataract, sleep disorders, sterility or hypogonadism is not related to the number of CTG repeats. Muscular atrophy and respiratory insufficiency are present with the highest frequency, occurring in 96 and 92% of the cases, respectively. A significant correlation was found with age of onset (r = -0.57, p<0.01), muscular disability (r = 0.46, p<0.05), intellective quotient (r = -0.58, p<0.01) and short-term memory (r= -0.59, p<0.01). Therefore, the CTG repeat number has a predictive value only in the case of some clinical manifestations, this suggesting that pathogenetic mechanisms of DM may differ depending on the tissue.     

Reported relationship between increased CTG repeat lengths in myotonic dystrophy and azoospermia

Dear Sir, The letter of Dean et al. stated that 16 out of 236 alleles(6.8%) had >18 CTG repeats and that seven out of the 15 menwho were found to     

强直性肌营养不良基因CTG重复数与EMG、NCV对比分析

目的：探讨强直性肌营养不良（ＤＭ）患者及家系成员三核苷酸重复数ＣＴＧ（胞嘧啶、胸腺嘧啶、鸟嘌呤）的变化与ＥＭＧ、ＮＣＶ的关系。方法：用聚合酶链反应（ＰＣＲ）扩增及ＤＮＡ杂交法对５例临床诊断ＤＭ患者及三个家系的１６名成员进行ＤＭ基因的ＣＴＧ重复数和ＥＭＧ、ＮＣＶ测定。结果：１０名正常人ＣＴＧ重复数是３０个，ＥＭＧ、ＮＣＶ正常，５例ＤＭ病人ＣＴＧ重复数均在８５个以上，其中２例在１６０５个以上，明显高于正常，１６名家系成员中除４名正常，余１２例ＣＴＧ重复数均超过正常基因，而且，ＣＴＧ重复数与临床症状、ＥＭＧ、ＮＣＶ轻重有关。结论：ＤＭ基因诊断与其临床诊断、ＥＭＧ、ＮＣＶ改变相一致     

Preimplantation genetic diagnosis for myotonic dystrophy type 1: upon request to child

Preimplantation genetic diagnosis (PGD) is an alternative to prenatal diagnosis for patients at risk of transmitting an inherited disease such as myotonic dystrophy type 1(DM1) to their offspring. In this paper, the clinical application of preimplantation diagnosis for DM1 upon request to children born is described in a large cohort of risk couples. PGD could be offered to all 78 couples opting for PGD regardless of the triplet repeat size. The incidence of major complications was minimalised following a careful assessment in affected DM1 females anticipating possible cardiological, obstetrical and anaesthetical problems. A live-birth delivery rate per cycle with oocyte retrieval of 20% was the outcome. Forty-eight of the 49 children born are in good health and have normal psychomotor development.     

Minimal expression of myotonic dystrophy: a clinical and molecular analysis.

A clinical and molecular study is reported of 83 patients considered to be minimally affected with myotonic dystrophy (DM). These had been identified in three ways: 60 subjects were identified on clinical grounds and were divided into those with and those without neuromuscular involvement (groups I and II); nine subjects were at high risk of carrying the DM gene but had a normal phenotype (group III); and 14 were parents of definitely affected patients where neither parent showed clinical abnormalities (group IV). PCR analysis of the CTG repeat in the DM gene showed a range of 70 to 230 repeats for the younger at risk patients in group III, while the asymptomatic gene carriers in group IV had 53 to 60 repeats. The sensitivity of diagnosis by EMG was found to be 39%. For ophthalmic signs this was 97.5%. This suggests that assignment on the basis of minimal clinical features carries a significant error. Molecular analysis, in conjunction with established clinical investigations, should prove valuable in the identification and exclusion of minimal myotonic dystrophy.     

Correlation between the onset age of Huntington's disease and length of the trinucleotide repeat in IT-15

Huntington's disease (HD) is an autosomal dominant disorderwith a variable age of onset that is influenced by the sex ofthe affected parent. The recent recognition that HD is causedby an expanded triplet repeat suggests the possibility thatthe onset age may be predicted by the length of the repeat.This hypothesis was tested by assaying the length of the repeatin 114 individuals who were clinically diagnosed with HD andhad a known onset age. Every individual had an expanded allele.The range was from 36 to 82 repeats (mean = 48.4 ± 9.51)and larger than the normal range (6 to 31). The size of theexpanded allele was correlated with the age of onset (r = –0.65p <.0001). Despite the highly significant correlation, therepeat size explains less than half of the variance in onsetage. Furthermore, the age of onset cannot be predicted fromthe size of the triplet repeat, particularly if the number ofrepeats is in the smaller end of the expanded range. If therepeat Is     

Comparison of the myotonic dystrophy associated CTG repeat in European and Japanese populations.

Gene amplification using polymerase chain reaction (PCR) was carried out on DNA samples from a total of 92 normal subjects and 52 subjects with myotonic dystrophy (DM) from European and Japanese populations, to determine the copy number of the CTG repeat associated with DM for each group. In the two populations, the number of repeats on normal chromosomes only were compared, as CTG copy number on DM chromosomes was difficult to determine by PCR alone. In this study, normal chromosomes were found which had as many as 35 copies of the repeat, which is larger than the normal range reported previously but still does not overlap with the repeat number associated with DM pathology, which is at least 50 copies. Using data from normal chromosomes from unrelated subjects, the frequencies of five, 11, and 13 copies of the CTG repeat were found to be significantly different between the two populations, with five and 11 copies more commonly seen in the European population and 13 copies in the Japanese population. This difference may be the result of natural divergence of the normal chromosomes between the population groups.     

The correlation of CTG repeat length with material and social deprivation in myotonic dystrophy

Socioeconomic deprivation has long been recognized as a prominent feature of myotonic dystrophy type 1 (DM1), but studies performed before the discovery of the mutation causing DM1 may have suffered an ascertainment bias towards the more severe forms of the disease. We have sought to clarify the relationship between CTG repeats, muscular impairment, and socioeconomic characteristics of 200 patients with DM1. Patients with DM1 reported lower educational attainment, lower employment rate, lower family income, and higher reliance on social assistance than the reference population. Logistic regression showed, on one hand, that CTG repeats and marital status were significant predictors of social assistance recipiency and, on the other hand, that CTG repeats and gender were significant predictors of low social support from family, after adjustment for age, gender, degree of muscular impairment, CTG repeats, educational level, and marital status. For example, each additional 100 CTG repeats was found to increase the odds of relying on social assistance by about 35% and having low social support by about 22%. The chances of experiencing socioeconomic deprivation are loaded heavily against patients with DM1. The relationship between increased CTG repeat length and higher risk of material and social deprivation must be acknowledged in the clinical management of DM1.     

DNA mismatch repair protein expression and microsatellite instability in primary mucosal melanomas of the head and neck

AIMS: To examine the expression of DNA mismatch repair (MMR) proteins and the presence of microsatellite instability (MSI) in seven primary mucosal melanomas of the head and neck (MMHN). METHODS AND RESULTS: Haematoxylin and eosin staining and immunohistochemical analysis for routine diagnostic markers and for MMR proteins were performed. Six cases were examined for MSI. Four cases were monomorphous and three cases were pleomorphic type MMHN. Melanocytic markers were positive in all cases. Immunoreactivity for MMR proteins was weak in normal epithelium. The neoplastic tissue in six cases showed positivity for all MMR proteins with different percentages. One case showed weak positivity for hMSH2 and hMSH6 and no immunoreactivity for hMLH1 or hPMS2. Staining intensity was higher in tumour cells than in matched normal mucosa in three cases for hMSH2 and hMLH1 and in two cases for hPMS2. None of the examined cases showed MSI. CONCLUSIONS: Expression of hMSH2 and hMLH1 proteins was up-regulated in three cases, whereas in two cases that of hPMS2 was increased. hMSH6 expression was comparable to that of normal cells in all cases. The percentage of positive neoplastic cells and the intensity of staining seemed to be greater in pleomorphic melanomas. Six cases were MMR-proficient and microsatellite stable.     

Complex patterns of male germline instability and somatic mosaicism in myotonic dystrophy type 1

The genetic basis of myotonic dystrophy type 1 (DM1) is the expansion of a CTG repeat in the 3' untranslated region of DM1PK. Once into the disease range, the repeat becomes highly unstable and is biased toward expansion in both somatic and germline tissues. Intergenerational differences usually reveal an increase in allele length, concordant with the clinical anticipation characteristic of DM1, but there have also been cases with intergenerational contractions of the repeat length, accompanied by apparent anticipation. In order to gain a better understanding of this intergenerational behaviour, we have obtained semen samples from six DM males and used single molecule analyses to compare the allele distributions present in their sperm and blood with those of their offspring. We have confirmed that the male germline mutational pathway is distinct from that of the soma, but the extent of variation is highly variable from one individual to another and not obviously correlated with progenitor allele length. Nonetheless, in all cases the alleles present in the father's sperm overlap with those observed in their offspring. These data also provide further indications that the interpretation of intergenerational transmissions by standard analyses is frequently compromised by the masking of germline differences by age-dependent somatic expansion in the parent.