[Clinical and genetic characteristics of 9 rare cases with coexistence of dual genetic diagnoses].

Objective: To analyze the clinical and genetic characteristics of pediatric patients with dual genetic diagnoses (DGD). Methods: Clinical and genetic data of pediatric patients with DGD from January 2021 to February 2022 in Peking University First Hospital were collected and analyzed retrospectively. Results: Among the 9 children, 6 were boys and 3 were girls. The age of last visit or follow-up was 5.0 (2.7,6.8) years. The main clinical manifestations included motor retardation, mental retardation, multiple malformations, and skeletal deformity. Cases 1-4 were all all boys, showed myopathic gait, poor running and jumping, and significantly increased level of serum creatine kinase. Disease-causing variations in Duchenne muscular dystrophy (DMD) gene were confirmed by genetic testing. The 4 children were diagnosed with DMD or Becker muscular dystrophy combined with a second genetic disease, including hypertrophic osteoarthropathy, spinal muscular atrophy, fragile X syndrome, and cerebral cavernous malformations type 3, respectively. Cases 5-9 were clinically and genetically diagnosed as COL9A1 gene-related multiple epiphyseal dysplasia type 6 combined with NF1 gene-related neurofibromatosis type 1, COL6A3 gene-related Bethlem myopathy with WNT1 gene-related osteogenesis imperfecta type XV, Turner syndrome (45, X0/46, XX chimera) with TH gene-related Segawa syndrome, Chromosome 22q11.2 microduplication syndrome with DYNC1H1 gene-related autosomal dominant lower extremity-predominant spinal muscular atrophy-1, and ANKRD11 gene-related KBG syndrome combined with IRF2BPL gene-related neurodevelopmental disorder with regression, abnormal movement, language loss and epilepsy. DMD was the most common, and there were 6 autosomal dominant diseases caused by de novo heterozygous pathogenic variations. Conclusions: Pediatric patients with coexistence of double genetic diagnoses show complex phenotypes. When the clinical manifestations and progression are not fully consistent with the diagnosed rare genetic disease, a second rare genetic disease should be considered, and autosomal dominant diseases caused by de novo heterozygous pathogenic variation should be paid attention to. Trio-based whole-exome sequencing combining a variety of molecular genetic tests would be helpful for precise diagnosis.

LncRNA FOXC2-AS1 stimulates proliferation of melanoma via silencing p15 by recruiting EZH2.

OBJECTIVE: The aim of this study was to elucidate the role of FOXC2-AS1 in promoting the proliferative ability and inhibiting apoptosis of melanoma by silencing p15, thereafter regulating the progression of melanoma. PATIENTS AND METHODS: FOXC2-AS1 levels in melanoma patients with or without metastasis and those with the tumor in different stages were detected by quantitative real-time polymerase chain reaction (qRT-PCR). Regulatory effects of FOXC2-AS1 on viability and apoptosis in melanoma cells were assessed, and subcellular distribution of FOXC2-AS1 was analyzed. Subsequently, the interactions of FOXC2-AS1 with EZH2 and SUZ12 were explored by RNA-Binding Protein Immunoprecipitation (RNA-RIP) assay. Through chromatin immunoprecipitation (ChIP) assay, the role of FOXC2-AS1 to regulate p15 transcription by recruiting EZH2 was verified. At last, regulatory effects of FOXC2-AS1/p15 axis on viability and apoptosis in melanoma cells were investigated. RESULTS: It was found that FOXC2-AS1 was upregulated in melanoma tissues, especially those with metastasis or stage II-IV. Melanoma patients expressing high level of FOXC2-AS1 showed worse survival than those with low level. Knockdown of FOXC2-AS1 inhibited viability, and stimulated apoptosis in A375 and sk-mel-110 cells. Besides, P15 level was upregulated in melanoma cells transfected with si-FOXC2-AS1, and FOXC2-AS1 was mainly distributed in cytoplasm. RNA-RIP assay confirmed that FOXC2-AS1 was mainly enriched in anti-EZH2 and aniti-SUZ12. Knockdown of EZH2 could markedly upregulate protein level of p15 in melanoma cells. Furthermore, it was verified that FOXC2-AS1 inhibited p15 transcription via recruiting EZH2, and the knockdown of p15 could partially reverse the regulatory effects of FOXC2-AS1 on viability and apoptosis in melanoma. CONCLUSIONS: FOXC2-AS1 stimulates proliferative ability in melanoma via silencing p15.

[Clinical phenotype and genotype of early-onset facioscapulohumeral muscular dystrophy type 1].

Objective: To explore the clinical, pathological and genetic characteristics of early-onset facioscapulohumeral muscular dystrophy type 1 (FSHD1), in order to increase awareness of the disease. Methods: In this retrospective study, the history of 3 patients, who were diagnosed with early-onset FSHD1 by molecular genetic test in Pediatric Outpatient Department of Peking University First Hospital from 4(th) June 2012 to 4(th) June 2018, were collected. Their clinical data, genotypes, phenotypes and pathological features of muscle biopsy were analyzed. Results: All the three patients were males at the age of 14 years, 11 years and 9 years 11 months, respectively, whose onset age was between infancy and early childhood and they got confirmed diagnosis within 4 to 10 years after the onset of illness. Their molecular genetic testing indicated that the number of D4Z4 repeat arrays located in 4qA were 2, 3 and 4, which was consistent with the characteristics of early-onset FSHD1. Their common clinical manifestations were facial, scapular and proximal lower limb muscle progressively and asymmetrically weakness. All patients had different severity of spine deformity and high-frequency dominant sensorineural hearing loss, however, the phenotype of the third patient with 4 D4Z4 repeats was significantly the most severe. Conclusions: Early-onset FSHD1 usually concealed onset and is difficult to diagnose. Its precise diagnosis depends on molecular genetic techniques, but the genotypes of 3 patients here are not corresponding to phenotypes strictly and it is necessary to accumulate more cases for further analysis in order to provide a more reliable basis for the relationship of genotype-phenotype and prognosis evaluation of the disease.

Sequential releasing of VEGF and BMP-2 in hydroxyapatite collagen scaffolds for bone tissue engineering: Design and characterization.

The property of biomaterials to control the sequential release of growth factors has been widely concerned in the field of bone repair and regeneration. Double-factor sustained-release scaffolds have good biocompatibility and bioactivity. O­Carboxymethyl chitosan microspheres (O-CMCS) has antimicrobial activity, biodegradability, biocompatibility and film formation. This study developed scaffolds materials with microspheres with a purpose of forming a controlled slow release secondary structure. O-CMCS were used as drug carrier to construct the compound sustained-release system with rhBMP-2 and VEGF double factors release. The O-CMCS were loaded with rhBMP-2 and the hydroxyapatite collagen (HC) scaffolds were loaded with VEGF to prepare the scaffolds. These were double factors composite sustained-release system scaffolds. The biocompatibility of the sustained-release system was evaluated by in vitro and in vivo experiments. HC composite scaffolds were characterized by scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FT-IR), X-ray diffraction detection (XRD), in vitro sustained release test, in vitro cell culture and in vivo animal experiments. The results showed that rhBMP-2/VEGF in HC composite scaffolds (HCBV) successfully achieved the sequential release of the double factors, which could benefit bone regeneration.

Expression and clinical significance of glucose transporter 1 mRNA in bronchial brushing liquid-based cytology specimens from patients with and without lung cancer.

OBJECTIVE: The aim of this study was to evaluate the diagnostic utility of glucose transporter 1 (GLUT1) mRNA expression in bronchial brushing specimens from patients with lung cancer. METHODS: GLUT1 mRNA levels were detected by reverse transcription-polymerase chain reaction (RT-PCR) in SurePath(TM) liquid-based cytology bronchial brushing specimens from patients with lung cancer (n=76) and benign lung disease (n=154). RESULTS: Compared with patients with benign disease and compared with cytology, GLUT1 mRNA was found significantly more frequently in patients with all carcinomas, squamous cell carcinomas, adenocarcinomas and small cell carcinomas, as well as central, peripheral and diffuse carcinomas (P<0.01). Minor differences were noted in GLUT1 mRNA and cytology results between histological types and tumour location but were not statistically significant. The diagnostic performance of RT-PCR analysis of GLUT1 mRNA was significantly higher than cytology in terms of sensitivity (97.4 ± 3.6% versus 65.8 ± 10.7) and negative predictive value (98.6 ± 1.9%, versus 85.6 ± 5.1%) but specificity (90.9 ± 4.5%) and positive predictive value (84.1 ± 7.6%) were lower than cytology (100%). CONCLUSIONS: Using liquid-based cytology, RT-PCR can be performed on bronchial brushing specimens to detect GLUT1 mRNA expression, and may be a useful adjunct to cytology diagnosis. It was more sensitive than cytology but its lower specificity should be taken into account.