Objective:To investigate the DMD genetic variants of the Chinese population with Duchenne (DMD) and Becker muscular dystrophies (BMD).Methods:A cross-sectional study was conducted on 2 690 unrelated patients with DMD and BMD aged 0-18 who visited the Genetic and Prenatal Diagnosis Center of the First Affiliated Hospital of Zhengzhou University from January 2005 to February 2022. The clinical data, such as gender, age, clinical manifestations, and address, were collected. Multiplex ligation-dependent probe amplification, next generation sequencing panel, Sanger sequencing, and PCR amplification were used to detect the variants of the DMD gene in the patients, whose clinical information and gene detection results were descriptively analyzed.Results:The 2 690 patients included 2 648 males and 42 females, with an age of 6.0 (4.0, 9.0) years. The serum creatine kinase increased in all patients. Pathogenic DMD gene variants were detected in the 2 618 patients, including 1 875 cases (71.6%) large deletions, 231 cases (8.8%) duplications, and 512 cases (19.6%) small variants. Among the deletion variants, the deletion of 3 exons was the most common, accounting for 15.4% (288/1 875); and hotspot deletion involved exons 45 to 50, accounting for 6.3% (119/1 875). Exon 2 was the most common type duplication region, accounting for 13.0% (30/231). Small variants were distributed in all 79 exons of the DMD gene, with no hotspots. In addition, the 46 small variants were previously unreported.Conclusion:Exon deletion is the most common type of DMD gene variant, followed by small variants and exon duplication.

Objective:To explore the significance and interpretation of low-depth whole-genome copy number variation sequencing (CNV-seq) in prenatal diagnosis in detecting DMD gene variations in fetuses without a family history of genetic diseases, and to investigate the results of family testing. Methods:Retrospectively collected case data of 16 fetuses with DMD gene deletions or duplications detected by low-depth whole-genome CNV-seq from December 2019 to August 2023 at the First Affiliated Hospital of Zhengzhou University. Amniotic fluid or chorionic villus samples and peripheral blood from family members were collected for all 16 cases, and genomic DNA was extracted. The fetal chromosomal copy number variations were detected using CNV-seq technology and the DMD gene deletions or duplications were verified by multiplex ligation-dependent probe amplification (MLPA), followed by family validation to trace the source of variation. The pathogenicity of the DMD gene deletion or duplication fragments was analyzed based on online Mendelian genetics databases and family validation results. Results:All 16 cases denied a family history of monogenic diseases. The indications for CNV-seq prenatal diagnosis were high-risk Down syndrome screening in nine cases, advanced maternal age in two cases, abnormal fetal ultrasound in three cases, and non-invasive prenatal DNA testing suggesting X chromosome abnormalities in two cases. CNV-seq results indicated nine cases of DMD gene duplication variations and seven cases of DMD gene deletion variations. MLPA validation confirmed results consistent with CNV-seq detection. Family analysis showed that three cases were de novo variations, 12 cases were inherited from the mother, one case had a mother with normal peripheral blood testing but a sister carrying the same variation, suggesting a high possibility of the mother being a carrier of gonadal mosaic. The likelihood of pathogenic variation was high in seven cases of deletion; nine cases were duplication variations, four of which were located within the DMD gene and could potentially disrupt the gene, leading to disease, while the other five variations were located in the 5' untranslated region or 3' untranslated region, considered benign variations. Conclusions:Low-depth whole-genome CNV-seq can effectively detect large deletion and duplication variations of the DMD gene in fetuses without a family history, preventing the birth of children with de novo variations. However, the pathogenicity of fetuses with large DMD gene duplications should be assessed based on family validation. When the duplication region includes the 5' untranslated region or 3' untranslated region of the DMD gene, it is more likely to be a polymorphic variation.

OBJECTIVE: To assess the value of re-sampling for patients who had failed non-invasive prenatal testing (NIPT) due to low cell-free fetal DNA (cffDNA) fraction. METHODS: Clinical data of 20 387 patients undergoing NIPT test was reviewed. The patients were re-sampled when initial blood test did not yield a result due to cffDNA fraction. The results were analyzed, and the outcome of pregnancy was followed up. RESULTS: Among all samples, 17 (0.08%) had failed to yield a result due to low cffDNA fraction, all of which accepted re-sampling. A result was attained in 16 cases, with a success rate of 94.12%. Only one sample had failed the re-test. CONCLUSION For patients who had failed the initial NIPT due to low cffDNA fraction, re-sampling should be considered with gestational week and ultrasound results taken into consideration.
Aneuploidy ; Cell-Free Nucleic Acids/genetics* ; DNA/genetics* ; Female ; Fetus ; Humans ; Pregnancy ; Prenatal Diagnosis

Objective:To analyze the clinical manifestations of congenital cataract in 12 families and gene variants causing the disease.Methods:The method of pedigree investigation was adopted.Clinical data of 27 patients from 12 Chinese Han families with congenital cataract were collected, and genomic DNA was extracted from peripheral blood samples of patients and family members.Candidate variants were screened by next generation sequencing and were verified by Sanger sequencing.Population frequency of the variants were obtained through the Genome Arrgregation Database (gnomAD).Pathogenicity of variants was analyzed through the Human Gene Mutation Database (HGMD), Database of Single Nucleotide Polymorphism (dbSNP) and PubMed, and the mutation effect was interpreted by protein prediction softwares including SIFT, PolyPhen_2 and MutationTaster.The conservation analysis of amino acid sequences of variants was performed by GERP+ + software.Diagnosis was confirmed by clinical ophthalmic phenotype, medical history and mutation analysis.This study adhered to the Declaration of Helsinki.The study protocol was approved by the Ethics Committee of the First Affiliated Hospital of Zhengzhou University (No.KS-2018-KY-36).Written informed consent was obtained from all subjects and their guardians.Results:Pathogenic genetic variants were found in all the 12 families, 9 of which had known pathogenic variants including MIP c.97C>T, GJA8 c.593G>A, CRYBA4 c.277T>C, CRYBB2 c.563G>A and c.436G>C, CRYGC c.470G>A, CRYGD c.70C>A, PAX2 c.70dupG as well as OCRL E5-E16dup, and 3 novel potential pathogenic variants including CRYGD c.422delG, ELP4 c.886C>A and CRYBB2 c.434G>C. CRYGD c.422delG could lead to the early termination of translation of protein products, which was pathogenic.The nucleotide and amino acid sites of ELP4 c.886C>A and CRYBB2 c.434G>C were highly conserved among species, and were predicted as harmful.The 12 families were consistent with co-segregation. Conclusions:CRYGD c.422delG, ELP4 c.886C>A and CRYBB2 c.434G>C may be novel pathogenic variants of congenital cataract.

Objective:To explore the application effect of flipped classroom model based on Simodont dental training system in the standardized training teaching of prosthodontics.Methods:The control experiment was used in this study. Seventy two students from Batch 2018 and Batch 2019 of Stomatology Hospital of Air Force Medical University were selected and randomly divided into experimental group (flipped classroom model based on Simodont dental training system) and control group (Simodont dental training system training mode after traditional teaching), with 18 students every academic year in each group. Questionnaire survey was conducted to evaluate the teaching effect, and the results of after-class theory test and practical computer test were compared between the two groups. SPSS 20.0 was used for chi-square test and t test. Results:The experimental group was better than the control group in enhancing classroom interest, improving the ability of independent analysis and problem-solving, and cultivating the ability of cooperation and expression ( P<0.05). The scores of after-class theory test and practical computer test in the experimental group [(23.36±0.21) points and (90.56±0.52) points] were significantly better than those in the control group[(21.81±0.25) points and (88.31±0.48) points] ( P<0.01). Conclusion:The flipped classroom model based on Simodont dental training system can effectively improve the effect of standardized training and teaching of professional skills in prosthodontics. At the same time, the students' ability of independent analysis and problem solving, cooperation and communication and expression are effectively improved.

OBJECTIVE: To explore the genetic basis for a Chinese patient suspected for Canavan disease. METHODS: Whole exome sequencing (WES) was carried out for the proband, and candidate variants were verified by Sanger sequencing of the proband, her parents and brother. Prenatal diagnosis was provided to her mother by chorionic villi sampling (CVS) upon her subsequent pregnancy. RESULTS: The proband, a 4-month-old female infant, had manifested drowsiness, hypotonia and apathy. Urine metabolism screening showed elevated N-acetylaspartic acid. Cranial magnetic resonance imaging revealed abnormal myelination and multiple abnormal signals in large brain areas. WES revealed that the proband has harbored compound heterozygous variants of the ASPA gene, namely c.187A>G (p.Arg63Gly) in exon 1 and c.634+1G>A (P.?) in exon 4. Sanger sequencing confirmed that the c.187A>G (p.Arg63Gly) and c.634+1G>A (p.?) variants were respectively inherited from her mother and father. Her phenotypically normal brother has carried a heterozygous c.634+1G>A (p.?) variant. Prenatal diagnosis by CVS indicated that the fetus was a heterozygous carrier of the c.187A>G variant. CONCLUSION WES can facilitate the diagnosis of Canavan disease, particularly for those lacking specific phenotypes of the disease. The compound heterozygous variants of the ASPA gene probably underlay the Canavan disease in this patient, and the result has enabled prenatal diagnosis for this family.
Canavan Disease/genetics* ; China ; Female ; Humans ; Male ; Mutation ; Pedigree ; Pregnancy ; Prenatal Diagnosis

OBJECTIVE: To explore the genetic etiology of a patient with mucopolysaccharidosis type II (MPSII). METHODS: The IDS gene of the proband and his mother was detected by Sanger sequencing, agarose gel electrophoresis, real-time PCR and multiple ligation-dependent probe amplification (MLPA). Prenatal diagnosis was performed on amniotic fluid sample. RESULTS: Agarose gel electrophoresis, real-time PCR, and MLPA all showed that exon 2 of IDS gene of the proband was deleted, for which his mother was normal. Prenatal diagnosis showed that the fetus was a normal male. CONCLUSION The de novo deletion of exon 2 of the IDS gene probably underlay the MPSII in this patient. Above finding has broadened the mutation spectrum of the IDS gene. The combined methods for the detection of IDS gene mutations could make accurate prenatal diagnosis for MPSII.
China ; Exons ; Female ; Humans ; Male ; Mucopolysaccharidosis II/genetics* ; Mutation ; Pedigree ; Pregnancy ; Prenatal Diagnosis/methods*

Objective:To explore the genetic etiology of a child with suspected propionic acidemia.Methods:Genomic DNA was extracted from peripheral blood sample of the child and subjected to high-throughput sequencing to screen pathogenic variants of genes associated with methylmalonic acidemia and propionic acidemia, including MUT, MMACHC, MMAA, MMAB, MMADHC, LMBRD1, PCCA, PCCB and SLC22A5. Candidate variants were verified by Sanger sequencing of the proband, her parents and sister. Results:The proband was found to harbor two pathogenic variants of the MUT gene, namely c. 1560+ 2T>C and c. 729_730insTT (p.Asp244fs), but not in genes associated with propionic acidemia. Her sister and father had carried c. 1560+ 2T>C, and her mother had carried c. 729_730insTT (p.Asp244fs). Conclusion:The proband was diagnosed as methylmalonic acidemia due to compound heterozygous variants of c. 1560+ 2T>C and c. 729_730insTT (p.Asp244fs) of the MUT gene. Her elder sister and parents were all carriers. Genetic testing has facilitated differential diagnosis of methylmalonic acidemia and propionic acidemia in this pedigree.

The clinical data of 380 patients undergoing loop electrosurgical excision procedure(LEEP) conization for cervical intraepithelial neoplasia grade 3(CIN3) at the Affiliated Hospital of Xuzhou Medical University from January 2017 to December 2019 were retrospectively analyzed. Univariate and multivariate logistic regression analyses were used to determine the risk factors of positive margin and residual lesions following conization. One hundred and twelve cases (29.5%) had positive margins, and the risk factors for positive margins were positive high-risk human papillomavirus ( OR=4.92, 95% CI: 1.81-13.36), high-grade lesions on thin cytologic test (TCT)screening ( OR=3.95, 95% CI: 2.42-6.42), and glandular involvement ( OR=3.58, 95% CI: 1.93-6.63). The hysteretomy was performed following conization in 169 hundred and sixty-nine patients who had difficulty in follow-up or no fertility intension, among whom 51 (30.2%) had residual lesions; logistic regression analysis showed that positive margins on LEEP were a risk factor for residual lesions at the time of hysterectomy ( OR: 2.83, 95% CI: 1.44-5.59). The study indicates that positive HR-HPV, high-grade TCT lesions, and lesions involving glands are risk factors for positive margins in CIN3 patients undergoing LEEP conization; and positive margins is risk factor for residual lesions following conization,to which more attention should be paid in clinical practice.

OBJECTIVE: To identify the etiology of a patient with severe symptoms of DMD and to trace its pathogenic gene, so as to provide a basis for genetic counseling and clinical intervention. METHODS: Multiple ligation-dependent probe amplification (MLPA) technique was used to analyze exon deletion/repetitive variant of DMD gene, and further analysis was performed by chromosome G-banding, fluorescence in situ hybridization (FISH) and SNP array analysis. RESULTS: The MLPA results of the proband showed that the exon 1-79 of DMD gene were deleted, the G-banding karyotype of blood sample was 46, XY, and the deletion of the short arm of X chromosome was found by FISH. SNP array results showed that 5.8Mb (29 628 158-35 434 714) deletion occurred in the Xp21.2p21.1 region of X chromosome, and the patient was diagnosed as the contiguous deletion syndrome involving the genes of IL1RAPL, MAGEB1-4, ROB, CXorf2, GM, AP3K7IP, FTHL1, DMD, FAM47A, TMEM47, and FAM47B. CONCLUSION The exact pathogenic site of this family is the deletion of 5.8 Mb (29 628 158-35 434 714) in the Xp21.2p21.1 region of X chromosome, which can be used for prenatal diagnosis. High resolution SNP array technique plays an important role in detecting potential chromosome abnormalities in patients.
Dystrophin/genetics* ; Exons ; Female ; Gene Deletion ; Humans ; In Situ Hybridization, Fluorescence ; Muscular Dystrophy, Duchenne/genetics* ; Pregnancy ; Prenatal Diagnosis