Objective:To observe and analyze the pathogenic gene types and clinical phenotypes of Leber congenital amaurosis (LCA).Methods:A retrospective clinical study. Six patients with LCA confirmed by genetic testing and 18 family members were included in the study. The patients came from six unrelated families. The family was investigated with a specific hereditary eye disease enrichment panel which contained 463 known pathogenic genes and based on targeted exome capture technology first to indentify the potential pathogenic genes and mutations. Then the TULP1 , RPGRIP1 , GUCY2D pathogenic mutations were conformed by Sanger sequencing. The pathogenicity of the gene variation was searched through relevant databases and PubMed literature, and its function was explained by protein prediction software. Results:Of the 6 patients, 3 were males and 3 were females; the age was from 3 to 33 years. Nystagmus, finger pressing eyes, photophobia, and night blindness were seen in 5 cases; electroretinogram showed 3 cases of extinction or near extinction; and 4 cases of retinopathy. The results showed patients with compound heterozygous mutation of c.1318C> T and c.1142T> G, homozygous mutation ofc.1318C> T and compound heterozygous mutation of c.1153G> A and c.1561C> T of TULP1 in Family 1, Family 2 and Family 5, respectively. There were compound heterozygous mutations of RPGRIP1 c.391delG and c.1468-2A> G in Family 3 and c.715delA and c.1765C> T in Family 6, respectively. Homozygous mutation of c.3177_3178delAC of GUCY2D was found in Family 4.The parents of all six patients were carriers of corresponding heterozygous mutations. TULP1 gene c.1142T> G, RPGRIP1 gene c.391delG, c.715delA and c.1765C> T and GUCY2D gene c.3177_3178delAC mutations were novel mutations and unreported. The 381th amino acid locus of product protein of TULP1 gene was highly conserved among species. The protein prediction software predicted that the mutation pathogenic. The c.391delG, c.715delA and c.1765C> T mutations of RPGRIP1 gene and c.3177_3178delAC mutation of GUCY2D gene can lead to early translation termination of their product proteins, which are pathogenic variants. Conclusion:The pathogenic mutations of TULP1, RPGRIP1 and GUCY2D genes led to LCA 15, LCA 6 and LCA 1 in six families.

Objectives: To study and prepare monoclonal antibody against glycophorin A of human erythrocyte(GPA McAb).This antibody is a key reagent in preparation of bispecific antibodies for rapid whole-blood immunoassay.　Methods: BALB/c mice received GPA antigen injection. Hybridoma was produced by traditional techniques. Hybridoma was determined with ELISA and indirect agglutination(IA) method.　Results: All 3 GPA McAb reacted with GPA, and they did not autoagglutenate with four types of red cells(type A,B,O,AB).Of the 3 McAb-GPA, two antibodies were IgG1,one IgG2 subtypes. Two IgG1 McAb-GPA can be used in making bispecific antibody in preparation of rapid whole-blood immunoassay.　Conclusions: Immunogenity of GPA was enhanced after coupling with the native serum albumin of the immunized animal, and high titer GPA McAb could be easily obtained. This result is important in making bispecific antibodies in preparation of rapid whole-blood immunoassay.

Objective To analyze the associations of the interleukin-10 (IL-10) promoter-1082G/A and-819C/T single nucleotide polymorphism (SNP) and serum level of IL-10 with intracranial aneurysm (IAs). Methods The polymerase chain reaction (PCR) and DNA direct sequencing methods were used to detect IL-10 gene promoter district two SNP site,-1082G/A and-819C/T genotype frequency and allele frequency in 206 patients with IAs and 187 controls. Chi-square test was used to analyze differences between two groups. The serum level of IL-10 was analyzed by ELISA, and t-test was used to analyze significant differences between two groups. Results There were significant differences in genotypes of GG and GA+AA, as well as the alleles G and A, in-1082G/A locus between IAs group and control group (P<0.01). There were higher frequencies of genotype GA+AA and the allele A in IAs group than those in control group (P<0.01). There was higher risk of suffering IAs in patients with genotype GA+AA (OR=4.137, 95%CI:2.476-6.914) and the allele A (OR=3.368, 95%CI:2.476-4.583). There were higher frequencies of the genotype CT+TT and the allele T in-819C/T locus in IAs group than those of control group (P<0.01). There was higher risk of suffering IAs in patients with genotype CT+TT (OR=3.393, 95%CI:1.952-5.900) and the allele T (OR=3.764, 95%CI:2.730-5.192). The serum level of IL-10 was significantly lower in IAs group than that of control group (P<0.01). Conclusion The IL-10 promoter SNP influences the expression of IL-10. IL-10 promoter-1082G/A and-819C/T polymorphisms are correlated with the formation of IAs.

OBJECTIVETo study the new method of monitoring and clearing organophosphate in blood during single or mixed organophosphate(OP) poisoning.

METHOD(1) Mixed equal volumes of blood of OP poisoned rat and healthy rat, then determine whole blood cholinesterase (ChE) activity. The descending range of ChE activity represents the level of residual OP in blood. (2) Poisoned rats by single or mixed OP pesticides were injected with 5% NaHCO3 15 ml/kg intraperitoneally, then the level of OP in blood was detected.

RESULTS(1) The monitoring results of blood residual OP by gas chromatography were similar to that by "Mixes blood method", which showed significant difference(P < 0.05) from that before OP administration. (2) NaHCO3 injection could not improve the toxic symptoms and whole blood or brain ChE inhibition in 10 CP poisoned rats, blood residual OP level was also not affected, but lung pathological changes by OP such as interstitial inflammation and oedema showed some relief.

CONCLUSIONThe monitoring of blood ChE by "mixed blood method" may reflect the general level of the blood residual OP within the range of exposure dose. The effect of NaHCO3 was not satisfactory, but it may improve OP-induced lung pathological changes.

Animals ; Cholinesterase Inhibitors ; poisoning ; Cholinesterases ; blood ; Chromatography, Gas ; Insecticides ; poisoning ; Lung ; pathology ; Organophosphate Poisoning ; Organophosphorus Compounds ; blood ; Rats ; Sodium Bicarbonate ; pharmacology

OBJECTIVETo detect potential mutations of PHEX gene in four pedigrees affected with hypophosphatemic rickets (HR) and provide prenatal diagnosis for a fetus at 13th gestational week.

METHODSThe coding regions and exon/intron boundaries of PHEX, FGF23, DMP1, ENPP1, CLCN5 and SLC34A3 genes of the probands were analyzed by targeted next-generation sequencing (NGS). Suspected mutations were verified by Sanger sequencing among unaffected relatives and 200 unrelated healthy individuals. Deletions were confirmed by multiplex ligation-dependent probe amplification (MLPA) detection of probands, unaffected relatives and 20 unrelated healthy individuals. Prenatal diagnosis for a fetus with high risk was carried out through MLPA analysis.

RESULTSFour PHEX mutations were respectively detected in the pedigrees, which included c.850-3C>G, exon 11 deletion, exon 13 deletion and c.1753G>A (p.G585R). Among these, exon 11 deletion, exon 13 deletion and c.1753G>A (p.G585R) were novel mutations and not found among unaffected relatives and healthy controls. In pedigree 3, the same mutation was not found in the fetus.

CONCLUSIONMutations of the PHEX gene probably underlies the disease among the four pedigrees. NGS combined with Sanger sequencing and/or MLPA detection can ensure accurate diagnosis for this disease.

Peptide and protein biologics possess high specificity and high biological activity, but their poor stability and short plasma half-life have limited clinical application. One established strategy to increase half-life of therapeutic proteins is chemical conjugation of the biologic with PEG. Nevertheless, PEGylation technology has some drawbacks, so recombinant polypeptide mimetics of PEG have gradually developed in recent years. Pharmaceutically active protein can be fused with specific amino acid sequences using recombinant DNA technology, and then increase hydrodynamic volume or produce charge effect, which retards kidney filtration and eventually prolongs the half-life. This article mainly reviews kinds of polypeptides and the research progress in half-life extension of therapeutic proteins.

OBJECTIVE: To explore the genetic etiology of a child featuring recurrent oral ulcer. METHODS: Clinical data of the child was collected. Whole exome sequencing was carried out for her. Candidate variant was verified by low-coverage massive parallel copy number variation sequencing (CNV-seq) of the family trio. RESULTS: The child, a 6-year-old girl, has featured recurrent fever and ulcers of the oral mucosa, vulvar and perianal regions. No pathogenic variant was found by whole exome sequencing. However, analysis of chromosome copy number variation using the whole exome sequencing data has revealed mosaicism of trisomy 8. CNV-seq assay has verified the variant in the child, with the percentage of mosaicism being 73%. No abnormality was found in neither of her parents. CONCLUSION A case of mosaicism trisomy 8 with recurrent oral ulcer as the first symptom was diagnosed, which has enriched the phenotypic data of trisomy 8 syndrome.
Humans ; Child ; Female ; Trisomy/genetics* ; Chromosomes, Human, Pair 8/genetics* ; DNA Copy Number Variations ; Oral Ulcer ; Mosaicism

Objective:To study the epidemic characteristics of human brucellosis in Yunnan Province, and to provide a reliable scientific basis for formulating accurate prevention and control strategies of brucellosis.Methods:The epidemic data of human brucellosis in Yunnan Province from January 2019 to December 2021 were collected from the information system of the China Center for Disease Control and Prevention, as well as annual monitoring data on brucellosis reported by various states (municipalities) in Yunnan Province. Descriptive epidemiological methods were adopted to analyze the epidemic situation, distribution characteristics (time, region, population), and serological and pathogenic monitoring results of brucellosis.Results:From 2019 to 2021, 1 408 cases of brucellosis were reported in Yunnan Province, with an average annual incidence of 1.00/100 000. The number of cases increased from 321 in 2019 to 701 in 2021, and the incidence increased from 0.68/100 000 in 2019 to 1.50/100 000 in 2021. The onset time was mainly from April to September (857 cases). The top 3 regions with the highest number of reported cases were Kunming City (483 cases), Qujing City (379 cases), and Honghe Hani and Yi Autonomous Prefecture (281 cases), accounting for 81.18% (1 143/1 408) of the total number of cases. The age of onset was mainly 20 - < 70 years old, accounting for 89.70% (1 263/1 408). There were 958 males and 450 females, with a sex ratio of 2.13 ∶ 1.00. Farmers were the main occupation, accounting for 84.02% (1 183/1 408). From 2019 to 2021, a total of 26 280 serum samples from key populations of brucellosis were monitored in Yunnan Province, with 572 positive serological tests and a positive rate of 2.18% (572/26 280). A total of 169 strains of Brucella were isolated from blood samples from hospitals throughout the province, including 155 strains of sheep type 3 and 14 strains of sheep type 1. Conclusions:From 2019 to 2021, the incidence of human brucellosis in Yunnan Province has been on the rise, with a high incidence in summer and autumn. The main population affected is young and middle-aged male farmers. Therefore, it is necessary to strengthen disease monitoring and health education for key populations during the high incidence season.

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.

Several programmed cell death protein 1 (PD-1) or its ligand (PD-L1) immune checkpoint blocking antibodies are available for clinical treatment, but only some patients show clinical response, so an alternative strategy for tumor immunotherapy is needed.A therapeutic tumor vaccine targeting PD-L1 is a meaningful attempt.In this study, we designed an epitope peptide vaccine targeting PD-L1, and then screened the immunogenic PD-L1 epitope peptide based on the humanized immune system (HIS) mouse model and further investigated its anti-tumor activity.The results show that the designed and screened PD-L1-B1 epitope peptide vaccine not only successfully induced PD-L1-specific humoral and cellular immunity, but also exhibit anti-tumor activity.In addition, immunotherapy increased T-lymphocyte infiltration of tumors and reshaped the tumor immunosuppressive microenvironment.In conclusion, PD-L1-B1 epitope peptide vaccine exhibits potent anti-tumor activity and may be an effective alternative immunotherapeutic strategy for patients insensitive to PD-1/PD-L1 blockade.