OBJECTIVE: To explore the clinical characteristics and genetic basis of two brothers featuring X-linked alpha thalassemia mental retardation (ATR-X) syndrome. METHODS: An infant who had presented at the Qilu Children's Hospital in 2020 for unstable upright head and inability to roll over and his family were selected as the study subjects. The clinical features of the child and one of his brothers were summarized, and their genomic DNA was subjected to targeted capture and next generation sequencing (NGS). RESULTS: The brothers had presented with mental retardation and facial dysmorphisms. NGS revealed that they had both harbored a hemizygous c.5275C>A variant of the ATRX gene located on the X chromosome, which was inherited from their mother. CONCLUSION The siblings were diagnosed with ATR-X syndrome. The discovery of the c.5275C>A variant has enriched the mutational spectrum of the ATRX gene.
Humans ; Infant ; Male ; alpha-Thalassemia/diagnosis* ; Ataxia Telangiectasia Mutated Proteins/genetics* ; East Asian People ; Intellectual Disability/genetics* ; Mental Retardation, X-Linked/diagnosis* ; Pedigree ; X-linked Nuclear Protein/genetics*

OBJECTIVE: To analyze the (CGG)n repeats of FMR1 gene among patients with unexplained mental retardation. METHODS: For 201 patients with unexplained mental retardation, the (CGG)n repeats of the FMR1 gene were analyzed by PCR and FragilEase RESULTS: For the 201 patients with unexplained mental retardation, 15 were identified with full mutations of the FMR1 gene. The prevalence of fragile X syndrome (FXS) in patients with unexplained mental retardation was determined as 7.5% (15/201). Prenatal diagnosis was provided for 6 pregnant women with pre- or full mutations. Analysis revealed that women with mental retardation and full FMR1 mutations exhibited a skewed XCI pattern with primary expression of the X chromosome carrying the mutant allele. CONCLUSION FXS has a high incidence among patients with unexplained mental retardation. Analysis of FMR1 gene (CGG)n repeats in patients with unexplained mental retardation can facilitate genetic counseling and prenatal diagnosis for their families. FMR1 gene (CGG)n repeats screening should be recommended for patients with unexplained mental retardation.
Female ; Fragile X Mental Retardation Protein/genetics* ; Fragile X Syndrome/genetics* ; Humans ; Intellectual Disability/genetics* ; Mutation ; Pregnancy ; Prenatal Diagnosis

OBJECTIVE: To determine the CGG repeat number and methylation status of FMR1 gene for fetuses whose mothers have carried a FMR1 mutation. METHODS: For 30 pregnant women, the fetal CGG repeat number was determined with a GC-rich PCR system by using chorionic villus, amniotic fluid or umbilical blood samples. The methylation status of the FMR1 gene was confirmed with Southern blotting. RESULTS: In total 30 prenatal diagnoses were performed for 29 carriers of FMR1 gene mutations and 1 with FMR1 gene deletion mosaicism. Three fetuses were found to carry premutations, 9 were with full mutations and 1 with mosaicism of premutation and full mutations. Eighteen fetuses were normal. CONCLUSION Considering the genetic complexity of Fragile X syndrome (FXS), single method may not suffice accurate determination of their genetic status. The pitfalls and technical limitations of protocols requires adoption of personalized strategy for its prenatal diagnosis.
Female ; Fragile X Mental Retardation Protein ; genetics ; Fragile X Syndrome ; diagnosis ; Heterozygote ; Humans ; Mutation ; Pregnancy ; Prenatal Diagnosis

OBJECTIVETo study the pattern of CGG repeat instability within germline cells derived from two male fetuses affected with Fragile X syndrome (FXS).

METHODSThe length and methylation status of CGG repeats within the testes of a fetus carrying a full FXS mutation and another fetus carrying mosaicism FXS mutation were analyzed with Southern blotting and AmplideX FMR1 PCR. Immunohistochemistry was also applied for the measurement of FMR1 protein (FMRP) expression within the testes.

RESULTSFor the fetus carrying the full mutation, Southern blotting analysis of the PCR product has detected an expected band representing the full mutation in its brain and a premutation band of > 160 CGG repeats in its testis. Whereas the pattern of premutation/full mutation in mosaic testis was similar to that in peripheral blood and no sign of contracted fragment was found other than a band of about 160 CGG repeats. Immunohistochemistry assay with a FMRP-specific antibody demonstrated a number of FMRP-positive germ cells, which suggested a contraction from full mutation to premutation alleles.

CONCLUSIONThis study has clarified the instability pattern of CGG repeat and expression of FMRP protein within the testes of fetuses affected with FXS, confirming that the CGG repeat can contract progressively within the germline. The FMRP expression in the testis is consistent with spermatogonium proliferation, and thus the contraction from full mutation to unmethylated premutations may occur for the requirement of FMRP expression during spermatogenesis. The better understanding of FMRP function during germ cell proliferation may elucidate the mechanism underlying the contraction of full FXS mutation in male germline.

Abortion, Eugenic ; Blotting, Southern ; Brain ; embryology ; metabolism ; DNA Methylation ; Fatal Outcome ; Fetus ; cytology ; metabolism ; Fragile X Mental Retardation Protein ; genetics ; metabolism ; Fragile X Syndrome ; diagnosis ; genetics ; Humans ; Immunohistochemistry ; Male ; Mosaicism ; Mutation ; Polymerase Chain Reaction ; Spermatozoa ; metabolism ; Testis ; cytology ; embryology ; metabolism ; Trinucleotide Repeat Expansion ; genetics

ObjectiveTo investigate the clinical (including reproductive) manifestations and genetic characteristics of familial fragile X syndrome (FXS).

METHODSWe collected the clinical data about a case of familial FXS by inquiry, testicular ultrasonography, semen analysis, determination of sex hormone levels, and examinations of the peripheral blood karyotype and Y chromosome microdeletions. Using Southern blot hybridization, we measured the size of the CGG triple repeat sequence of the fragile X mental retardation-1 (FMR1) gene and determined its mutation type of the pedigree members with a genetic map of the FXS pedigree.

RESULTSAmong the 34 members of 4 generations in the pedigree, 3 males and 1 female (11.76%) carried full mutation and 9 females (26.47%) premutation of the FMR1 gene. Two of the males with full FMR1 mutation, including the proband showed a larger testis volume (>30 ml) and a higher sperm concentration (>250 ×10⁶/ml), with a mean sperm motility of 50.5%, a mean morphologically normal sperm rate of 17.5%, an average sperm nuclear DNA fragmentation index (DFI) of 18.5%, a low level of testosterone, normal karyotype in the peripheral blood, and integrity of the azoospermia factor (AZF) region in the Y chromosome. One of the second-generation females carrying FMR1 premutation was diagnosed with premature ovarian failure and another 3 with uterine myoma.

CONCLUSIONSSome of the FXS males in the pedigree may present macroorchidism and polyzoospermia but with normal semen parameters. In the intergenerational transmission, premutation might extend to full mutation, with even higher risks of transmission and extension of mutation in males, especially in those with >80 CGG triple repeat sequences. Therefore, it is recommended that the couples wishing for childbearing receive genetic testing, clinical guidance, and genetic counseling before pregnancy and, if necessary, prenatal diagnosis and preimplantation genetic diagnosis.

Chromosome Deletion ; Chromosomes, Human, Y ; genetics ; DNA Fragmentation ; Female ; Fragile X Mental Retardation Protein ; genetics ; Fragile X Syndrome ; genetics ; Genetic Testing ; Humans ; Infertility, Male ; genetics ; Karyotyping ; Male ; Mutation ; Organ Size ; Pedigree ; Pregnancy ; Preimplantation Diagnosis ; Risk ; Sex Chromosome Aberrations ; Sex Chromosome Disorders of Sex Development ; genetics ; Sperm Count ; Testis ; diagnostic imaging ; pathology

OBJECTIVETo investigate whether the four boys with delayed motor development and intellectual disability suffer from MECP 2 duplication syndrome.

METHODBlood specimens and clinical data of four patients and mothers of patient 2 and patient 4 were collected. Genomic DNA was extracted from peripheral blood using DNA extraction kit. At first multiplex ligation-dependent probe amplification (MLPA) was employed in 4 patients, two distinct kits SALSA P036 and P070 for sub-telomere screening, and SALSA P245 for the 22 common microdeletion and microduplication syndromes. Then array-CGH analysis was carried out. Two mothers of patients were tested by array- comparative genomic hybridization (CGH) and X chromosome inactivation analysis.

RESULTAll the 4 patients presented with severe hypotonia, delayed motor development, intellectual disability and absent or limited language. Three patients manifested recurrent pneumonia in infancy except patient 2. Four patients had duplication on chromosome Xq28 with MLPA kit SALSA P245. Array-CGH identified the size of each duplication on Xq28. The precise size of each duplication was different in the four patients: patient 1, 14.931 Mb, patient 2, 0.393 Mb, patient 3, 0.482 Mb and patient 4, 0.299 Mb. To compare Xq28 duplications with UCSC database (http://genome.ucsc.edu/) revealed that each duplication harbors the MECP 2 and HCFC 1 gene. Mothers of patient 2 and patient 4 also carried microduplication on Xq28. X chromosome inactivation analysis demonstrated completely skewed inactivation (0: 100) and it is the inactive allele that passed on to the patients.

CONCLUSIONFor patients that present with delayed motor development, intellectual disability, hypotonia, absent or limited language and recurrent infection, combination of MLPA and array- CGH is effective and specific diagnostic methods of MECP 2 duplication syndrome.

Chromosomes, Human, X ; genetics ; Comparative Genomic Hybridization ; Gene Duplication ; Humans ; Male ; Mental Retardation, X-Linked ; diagnosis ; genetics ; Methyl-CpG-Binding Protein 2 ; genetics ; Multiplex Polymerase Chain Reaction ; X Chromosome Inactivation

OBJECTIVETo establish a method of methylation-sensitive restriction enzymes based quantitative PCR (MSRE-qPCR) for analysis of CpG island DNA of FMR1 gene, and to assess its value for molecular diagnosis of fragile X syndrome.

METHODSThirty boys with mental retardation and abnormal repeats of 5'(CGG)n in the FMR1 gene and 20 mothers were analyzed by conventional PCR screening. Eag I was used to digest genomic DNA, and qPCR was performed to amplify CpG island in the FMR1 gene using both undigested and digested templates. Raw Ct values were obtained through quantitative PCR amplification. The degree of CpG island methylation was calculated by 2 - U+0394 U+0394 Ct. The result of MSRE-qPCR was verified by Southern blotting. 30 healthy females and 30 healthy males were used as controls to optimize the established MSRE-qPCR method.

RESULTSThe ranges of 2 - U+0394 U+0394 Ct value for normal methylation, partial methylation and full methylation were determined. Among the 30 patients, 3 were found to have partial methylation of CpG island of the FMR1 gene, and 27 were found to have full methylation (3/30 results were verified by Southern blotting). Only 7 mothers were found abnormal methylation of CpG island of FMR1 gene, whilst the remaining 13 mothers were normal.

CONCLUSIONMSRE-qPCR is a quick and reliable method for quantitative analysis of CpG island methylation status in FMR1 gene, which may provide a new strategy for the diagnosis of fragile X syndrome.

CpG Islands ; DNA Methylation ; Female ; Fragile X Mental Retardation Protein ; genetics ; Fragile X Syndrome ; diagnosis ; genetics ; Humans ; Male ; Sex Factors

Fragile X mental retardation 1 is the gene underlying fragile X syndrome (FXS). Its product, fragile X mental retardation protein, is closely involved with development of brain and neurons. PCR and Southern blotting have been the major methods for laboratory diagnosis of FXS. In this article, the progress in the molecular diagnosis of FXS is reviewed.
Fragile X Mental Retardation Protein ; genetics ; Fragile X Syndrome ; diagnosis ; genetics ; Humans ; Pathology, Molecular ; methods

Fragile X-associated tremor/ataxia syndrome(FXTAS) is a neurodegenerative disease caused by FMR1 gene permutation(PM). The main clinical manifestations are intention tremor and/or ataxia, and the pathogenesis was related to RNA toxicity. In this paper, the research progress of clinical manifestatios, pathological characteristics, epidemiology and molecular mechanisms will be reviewed.
Ataxia ; genetics ; Female ; Fragile X Mental Retardation Protein ; genetics ; Fragile X Syndrome ; complications ; diagnosis ; genetics ; pathology ; Humans ; Male ; Tremor ; genetics

Mutation of the ATRX gene leads to X-linked alpha-thalassemia/mental retardation (ATR-X) syndrome and several other X-linked mental retardation syndromes. We report the first case of ATR-X syndrome documented here in Korea. A 32-month-old boy came in with irritability and fever. He showed dysmorphic features, mental retardation and epilepsy, so ATR-X syndrome was considered. Hemoglobin H inclusions in red blood cells supported the diagnosis and genetic studies confirmed it. Mutation analysis for our patient showed a point mutation of thymine to cytosine on the 9th exon in the ATRX gene, indicating that Trp(C), the 220th amino acid, was replaced by Ser(R). Furthermore, we investigated the same mutation in family members, and his mother and two sisters were found to be carriers.
Amino Acid Substitution ; Body Dysmorphic Disorders/complications ; Child, Preschool ; DNA Mutational Analysis ; Epilepsy/complications ; Exons ; Hemoglobin H/*genetics ; Humans ; Male ; Mental Retardation/complications ; Mental Retardation, X-Linked/complications/diagnosis/genetics ; Point Mutation ; Republic of Korea ; alpha-Thalassemia/complications/diagnosis/genetics