Objective:To explore the effects of hyperoxic environments on renal metabolites to understand the potential mechanisms that contribute to pathologic retinal vascular neovascularization and renal injury through metabolomic studies in a mouse model of oxygen-induced retinopathy (OIR) model.Methods:Sixteen C57/B6J mice pups born to day 7 (P7) were randomly and equally divided into an OIR model group and a normal control group using a randomized numerical table of mother mice.Mice were reared standardly from birth until day 7 (P7), then mice and their mother mice in the OIR group were placed in a hyperoxic (75±2)% chamber until day 12 (P12) and then reared normally.Mice in the normal control group were reared normally throughout.Mice in two groups were killed by carbon dioxide euthanasia on postnatal day 17 (P17). The mice retinal wholemount from the two groups were made and stained with isolectin B4 (IB4) to observe the morphology of retinal vessels, central non-perfusion area and pathological neovascularization.The kidney tissue of P17 mice was analyzed by liquid chromatograph mass spectrometer.After anticoagulant treatment, the whole blood of mice was centrifuged and precipitated, and the obtained plasma without cellular components was analyzed by targeted metabonomics.Mass spectral information was interpreted using metabolomics data processing software Progenesis QI v2.3.Overall differences in metabolic profiles were distinguished by unsupervised principal component analysis and orthogonal partial least squares analysis (OPLS-DA). The fold change and P values of metabolites were compared between the two groups.The variable importance of projection value>1 and P value<0.05 was used to screen out differential metabolites.Metabolic pathway enrichment analysis of differential metabolites was performed based on the KEGG database.The feeding and use of animals were strictly in accordance with the requirements of the Ethics Committee of Jinan University, and the research protocol was reviewed and approved by the Ethics Committee of Jinan University (No.20200401-54). Results:The IB4 staining of retinal wholemounts showed that the retinal blood vessels were evenly distributed in the P17 mice from control group.The peripheral retinal vessels were tortuous and disordered with a large non-perfusion area in central region in P17 mice from OIR group, and a large number of neovascularization clusters were formed at the junction of the nonperfusion area and the vascular area of the retina, showing strong fluorescent staining.The relative area of retinal nonperfusion area in OIR group was (25.16±3.50)%, which was significantly larger than (0.63±0.30)% in normal control group ( t=12.07, P<0.001). The OPLS-DA parameter R2X cum (0.578), interpretation rate R2Y cum (0.978) and prediction rate Q2 cum (0.857) values were all greater than 0.5, indicating that the OPLS-DA model had a good predictive ability.A total of 26 main differential metabolites were found, among which 17 were up-regulated and 9 were down-regulated, including glycerophospholipids (PC 20∶4(5Z, 8Z, 11Z, 14Z)/0∶0, PC 22∶6(4Z, 7Z, 10Z, 13Z, 16Z, 19Z)/0∶0, PC 14∶1(9Z)/20∶2(11Z, 14Z), PE P-18∶0/20∶4(6E, 8Z, 11Z, 14Z)(5OH[S]), amino acid metabolites (arginine, ornithine, pipecolic acid, and hydroxylysine), purines (guanine, hypoxanthine, hydroxypurinol), and fatty acids (methyl 15-palmitate, 2, 6, 8, 12-tetramethyl-2, 4-tridecadien-1-ol), and so on.Differential metabolites were mainly enriched in ABC transporters (L-arginine, taurine, inositol, adenosine, N-acetyl-D-glucosamine, L-glutamine), aminoacyl-tRNA biosynthesis (L-isoleucine, L-proline, L-arginine, L-histidine, L-glutamine), arginine biosynthesis (L-arginine, L-ornithine, L-glutamine) metabolic pathways.The plasma targeted metabonomics showed that the differential amino acid metabolites were mainly enriched in metabolic pathways such as aminoacyl-tRNA biosynthesis, arginine biosynthesis and metabolism, and ABC transporters. Conclusions:ABC transporter, aminoacyl-tRNA biosynthesis, and arginine biosynthesis metabolic pathways in OIR mice may participate in the pathological changes of renal injury and neovascularization in retinopathy of prematurity.

[Objective] To evaluate the effects of preoperative ureteroscopy (URS) on the prognosis of patients with upper tract urothelial carcinoma (UTUC) after radical nephroureterectomy (RNU). [Methods] Data of 712 UTUC patients who received RNU in West China Hospital during May 2003 and Jun.2019 were retrospectively analyzed. Patients were divided into URS group (n=187) and non-URS group (n=525) according to whether URS was performed before RNU. Kaplan-Meier curves were used to analyze the overall survival (OS), cancer-specific survival (CSS), and intravesical recurrence-free survival (IVRFS). Cox proportional risk model was used for risk assessment. Subgroup analysis and interaction test were used to further verify the results. [Results] Significant differences were observed between the two groups in terms of body mass index (BMI), diabetes mellitus, surgical method, hydronephrosis, tumor location, tumor grade, lymph node metastasis, lymphovascular invasion and tumor diameter (P<0.05). Kaplan-Meier survival analysis showed that IVRFS was significantly lower in the URS group than in the non-URS group (P<0.001), and the difference was more pronounced in renal pelvis carcinoma (P<0.001); there were no differences in OS and CSS between the two groups (P>0.05). Cox multivariate regression showed that URS was an independent risk factor for intravesical recurrence (HR=2.12, 95%CI: 1.34-3.36, P<0.001). [Conclusion] Preoperative URS can increase the recurrence rate of UTUC, but it has no effect on the OS and CSS.

Objective:To explore the value of whole exome sequencing for the inferential analysis of recessive genetic disease carrier status for couples with a child died of Primary immunodeficiency (PID).Methods:Clinical data was collected from four couples with a childbearing history of PID who had sought genetic counseling and undergone genetic testing at Henan Provincial People′s Hospital from February 2017 to December 2021. Whole exome sequencing (WES) was performed on both partners of each couple, and candidate variants were validated by Sanger sequencing and fluorescent quantitative PCR. Prenatal diagnosis was conducted on fetuses of these couples after confirming the variants.Results:A total of six variants were detected in four genes including IL2RG, BTK, CYBB, and DUOX2. Among these, the c.1265G>A and c.3329G>A variants of the DUOX2 gene and the c. 676C>T variant of the IL2RG gene were previously known as pathogenic variants. On the other hand, the Exon5_8del variant of the IL2RG gene, the c. 184_185delAC variant of the BTK gene, and the c. 472A>T variant of the CYBB gene were unreported previously. Based on the guidelines from the American College of Medical Genetics and Genomics, the IL2RG: Exon5_8del, BTK: c. 184_185delAC and CYBB: c. 472A>T variants were classified as likely pathogenic (PVS1+ PM2_Supporting+ PP4).Prenatal diagnosis was conducted for three couples during their subsequent pregnancies, and the results revealed that the fetuses had the wild-type genotypes at the c. 184_185 position of the BTK gene, the c. 472 position of the CYBB gene, and the c. 676 position of the IL2RG gene. Follow-up examinations one year after birth has found no abnormality in the infants. Conclusion:WES is an important tool to infer and analyze the carryier status for couples who had given births to children died of PID and improve the positive detection rate.

Objective:To explore the genetic etiology of a pedigree with intellectual disability and explore its pathogenesis.Methods:A Chinese pedigree which had presented at the Henan Provincial People′s Hospital in March 2023 was selected as the study subject. Clinical data of the pedigree were collected, along with peripheral venous blood samples from its members. Whole exome sequencing (WES) was carried out, and candidate variants were verified by Sanger sequencing. Amniotic fluid was collected for prenatal diagnosis. This study was approved by the Medical Ethics Committee of the Henan Provincial People′s Hospital (No. 2019-134).Results:Both the proband (a 6-year-old male) and his mother (30 years old) had various degrees of intellectual and motor impairment. WES revealed that the proband has harbored a de novo heterozygous c. 2563_2567dup (p.Lys856fs) variant of the UBE3A gene, while his mother, maternal grandmother and fetus had all harbored a novel heterozygous c. 409+ 1G>A variant of the RNF13 gene. Based on the guidelines from the American College of Medical Genetics and Genomics (ACMG), both variants were predicted to be pathogenic (PVS1+ PS1+ PM2_Supporting; PVS1+ PM2_Supporting+ PP3). Conclusion:Based on the clinical manifestations and the result of genetic testing, the heterozygous c.2563_2567dup (p.Lys856fs) variant of the UBE3A gene probably underlay the intellectual disability and developmental delay in the proband, whilst the heterozygous c. 409+ 1G>A variant of the RNF13 gene may underlie the intellectual disability in the proband′s mother and grandmother. Above results have enabled genetic counseling and prenatal diagnosis for this pedigree.

Objective:To analyze the clinical phenotype and genetic characteristics of probands in 3 pedigrees of complex hereditary spastic paraplegia type 5 (HSP5) who developed symptoms during childhood, and the genetic diagnostic methods of HSP5 to improve the diagnosis and differential diagnosis of this disease.Methods:The clinical data of 3 HSP5 families admitted to Henan Provincial People′s Hospital from June 2020 to January 2023 were collected. Whole exome sequencing (WES) was performed on the patients to analyze phenotype-related single nucleotide variation (SNV) and small fragment insertion/deletion (INDEL) variation. At the same time, the sequencing data were used to analyze the dynamic mutation regions of specific genes.Results:The probands in the 3 families had complex HSP: the proband in family 1 showed weakness of both lower limbs, urgency of urination and ataxia; the proband in family 2 showed slightly lower intelligence, weakness of both lower limbs, dysarthria, and brain magnetic resonance imaging showed white matter lesions; the proband in family 3 showed muscle weakness, spasm, frequent urination and ataxia of both lower limbs. The sequencing results showed that the CYP7B1 gene c.1171G>T (paternal) and c.1249C>T (maternal) compound heterozygous mutations were found in proband 1 and his younger brother. The CYP7B1 gene c.334C>T (paternal) and c.259+2T>C (maternal) compound heterozygous mutations were found in proband 2 and her younger sister. The CYP7B1 gene c.334C>T (paternal) and c.1082G>A (maternal) compound heterozygous mutations were found in proband 3. And c.1171G>T was a new variant that had not been reported before. Dynamic mutation analysis showed that the numbers of CAG repeats of ATXN1/2/3/6/7/8/12, DRPLA, TBP genes were within the normal range. According to the clinical manifestations and genetic examination results of the children in the 3 pedigrees, the diagnosis of HSP5 was clear. Conclusions:The 3 families in the study all had complex HSP5 caused by compound heterozygous mutations of the CYP7B1 gene. WES can analyze SNV, INDEL and dynamic mutations simultaneously to make the maximum clear diagnosis and can be used as an effective detection method for HSP5.

Objective:To investigate the clinical efficacy of botulinum toxin A in the treatment of hand hyperhidrosis.Methods:One hundred patients with hand hyperhidrosis were treated with botulinum toxin A (BTXA, Lanzhou Biotechnology Development Co., Ltd., Botulinum Toxin Type A for Injection Hengli) injection, a total of 200 U. Each hand was injected with 100 U respectively. The curative effect was evaluated by self-made questionnaire. The scores of the two were added. The subjective and objective evaluation were carried out before and after injection, and the patients were rechecked 2 weeks, and 1, 4 and 6 months after injection. Efficacy, patient satisfaction and adverse reactions were evaluated.Results:Compared with before treatment, the effective rate increased 2 weeks after injection, 1 month after injection, 4 months after injection and 6 months after injection, and the difference was statistically significant (Chi-square value was 31.54, 36.33, 28.34, 25.23, respectively, P<0.05). After 6 months of follow-up, the effect gradually decreased, and the curative effect could be maintained for about 10 months. After recurrence, the symptoms of hand sweating were still reduced. Satisfaction 96%; Adverse reactions were mild, subcutaneous blood stasis, 27% hand muscles were slightly weak, and returned to normal after 2-3 weeks. Conclusions:Botulinum toxin A injection has certain curative effect, high safety and less adverse reactions. It is an ideal method for the treatment of hand hyperhidrosis.

Objective:To analyze clinical characteristics of and causative genes in two families with dystrophic epidermolysis bullosa, and to reveal the pathogenesis of the disease and mechanisms underlying phenotypic differences between patients.Methods:DNA was extracted from peripheral blood samples of members from two families with dystrophic epidermolysis bullosa, and subjected to high-throughput sequencing and Sanger sequencing.Results:The clinical manifestations of the 2 probands in the 2 families were consistent with the diagnosis of dystrophic epidermolysis bullosa, and the symptoms of the proband in family 1 were more serious than those of other patients in the family. Genetic testing showed that all patients in family 1 carried a mutation c.6082G>C (p.G2028R) in the COL7A1 gene, and the proband and her phenotypically normal mother and uncle also carried a splice-site mutation c.7068+2 (IVS91) T>G in the COL7A1 gene, both of which were first reported. The proband in family 2 carried the mutations c.6081_6082 ins C (p.G2028Rfs*71) and c.1892G>A (p.W631X, first reported) in the COL7A1 gene, which were inherited from her father and mother, respectively.Conclusion:The two pathogenic mutations may be the molecular mechanism underlying the severe clinical phenotype in the proband in family 1; the first reported mutations enriched the mutation spectrum of the COL7A1 gene.

OBJECTIVE: To explore the clinical characteristics and genetic basis of two Chinese pedigrees affected with Joubert syndrome. METHODS: Clinical data of the two pedigrees was collected. Genomic DNA was extracted from peripheral blood samples and subjected to high-throughput sequencing. Candidate variants were verified by Sanger sequencing. Prenatal diagnosis was carried out for a high-risk fetus from pedigree 2. RESULTS: The proband of pedigree 1 was a fetus at 23⁺⁵ weeks gestation, for which both ultrasound and MRI showed "cerebellar vermis malformation" and "molar tooth sign". No apparent abnormality was noted in the fetus after elected abortion. The fetus was found to harbor c.812+3G>T and c.1828G>C compound heterozygous variants of the INPP5E gene, which have been associated with Joubert syndrome type 1. The proband from pedigree 2 had growth retardation, mental deficiency, peculiar facial features, low muscle tone and postaxial polydactyly of right foot. MRI also revealed "cerebellar dysplasia" and "molar tooth sign". The proband was found to harbor c.485C>G and c.1878+1G>A compound heterozygous variants of the ARMC9 gene, which have been associated with Joubert syndrome type 30. Prenatal diagnosis found that the fetus only carried the c.485C>G variant. A healthy infant was born, and no anomalies was found during the follow-up. CONCLUSION The compound heterozygous variants of the INPP5E and ARMC9 genes probably underlay the disease in the two pedigrees. Above finding has expanded the spectrum of pathogenic variants underlying Joubert syndrome and provided a basis for genetic counseling and prenatal diagnosis.
Female ; Humans ; Pregnancy ; Pedigree ; Cerebellum/abnormalities* ; Abnormalities, Multiple/diagnosis* ; Eye Abnormalities/diagnosis* ; Kidney Diseases, Cystic/diagnosis* ; Phosphoric Monoester Hydrolases/genetics* ; Retina/abnormalities* ; East Asian People ; Mutation

OBJECTIVE: To explore the clinical features and genetic etiology of two children with intellectual developmental disorder and microcephaly with pontine and cerebellar hypoplasia (MICPCH). METHODS: Two children with MICPCH who were presented at the Henan Provincial People's Hospital between April 2019 and December 2021 were selected as the study subjects. Clinical data of the two children were collected, along with peripheral venous blood samples of them and their parents, and amniotic fluid sample of the mother of child 1. Whole exome sequencing (WES), array-comparative genomic hybridization (aCGH) and real-time quantitative PCR (qPCR) were carried out for the children, their parents and the fetus. The pathogenicity of candidate variants were evaluated. RESULTS: Child 1 was a 6-year-old girl featuring motor and language delay, whilst child 2 was a 4.5-year-old girl mainly featuring microcephaly and mental retardation. WES revealed that child 2 has harbored a 158.7 kb duplication in Xp11.4 (chrX: 41446160_41604854), which has encompassed exons 4~14 of the CASK gene. The same duplication was not found in either of her parents. aCGH revealed that child 1 has harbored a 29 kb deletion at Xp11.4 (chrX: 41637892_41666665), which encompassed exon 3 of the CASK gene. The same deletion was not found in either of her parents and the fetus. The above results were confirmed by qPCR assay. Above deletion and duplication were not found in the ExAC, 1000 Genomes and gnomAD databases. Based on the guidelines from the American College of Medical Genetics and Genomics (ACMG), both variants were rated as likely pathogenic (PS2+PM2_Supporting). CONCLUSION The deletion of exon 3 and duplication of exons 4~14 of the CASK gene probably underlay the pathogenesis of MICPCH in these two children, respectively.
Humans ; Child ; Female ; Child, Preschool ; Microcephaly/genetics* ; Developmental Disabilities/genetics* ; Intellectual Disability/complications* ; Comparative Genomic Hybridization ; Mutation

OBJECTIVE: To explore the genetic basis for fetus with bilateral lateral ventriculomegaly. METHODS: Fetus umbilical cord blood and peripheral blood samples of its parents were collected. The fetus was subjected to chromosomal karyotyping, whilst the fetus and its parents were subjected to array comparative genomic hybridization (aCGH). The candidate copy number variation (CNV) were verified by qPCR, Application goldeneye DNA identification system was used to confirm the parental relationship. RESULTS: The fetus was found to have a normal karyotype. aCGH analysis indicated that it has carried a 1.16 Mb deletion at 17p13.3, which partially overlapped with the critical region of Miller-Dieker syndrome (MDS), in addition with a 1.33 Mb deletion at 17p12 region, which is associated with hereditary stress-susceptible peripheral neuropathy (HNPP). Its mother was also found to harbor the 1.33 Mb deletion at 17p12. qPCR analysis confirmed that the expression levels of genes from the 17p13.3 and 17p12 regions were about the half of that in the normal control, as well as the maternal peripheral blood sample. Parental relationship was confirmed between the fetus and its parents. Following genetic counseling, the parents has chosen to continue with the pregnancy. CONCLUSION The fetus was diagnosed with Miller-Dieker syndrome due to the de novo deletion at 17p13.3. Ventriculomegaly may be an important indicator for prenatal ultrasonography in fetuses with MDS.
Pregnancy ; Female ; Humans ; Classical Lissencephalies and Subcortical Band Heterotopias ; Comparative Genomic Hybridization ; DNA Copy Number Variations ; Fetus ; Hydrocephalus ; Prenatal Diagnosis ; Chromosome Deletion