[ABSTRACT]OBJECTIVETo investigate the effect of accurate airway humidification on hemorrhage, pharyngalgia, mucosal edema and sputum viscosity in patients with low-temperature plasma coblation-assisted tonsillectomy.METHODS58 cases were divided into three groups by using random numbers.In accurate airway humidification group, atomizing inhalation was carried out by AIRVOTM series apparatus; in oxygen atomizing group, budesonide suspension was used; in control group, saline was used. We evaluated the hemorrhage, pharyngalgia, mucosal edema and sputum viscosity in 3 consecutive postoperative days.RESULTSPharyngalgia in accurate airway humidification group and in oxygen atomizing group were both significantly reduced than that of the control group (P<0.001). Besides, in accurate airway humidification group, mucosal edema and sputum viscosity were significantly improved than that of the oxygen atomizing group (P<0.05) and control group (P<0.05).CONCLUSIONAccurate airway humidification could reduce the complications such as pharyngalgia, mucosal edema and purulent sputum after low-temperature plasma coblation-assisted tonsillectomy, and could accelerate recovery from surgery.

Objective:To analyze the genotypes and clinical phenotypes of two families with Hermansky-Pudlak syndrome (HPS).Methods:The method of pedigree investigation was adopted.A Han Chinese HPS family and a Hui Chinese HPS family were enrolled in People's Hospital of Ningxia Hui Autonomous Region from June 2020 to May 2021.Clinical data of two probands and their phenotypically normal parents were collected.Relevant ocular and systemic examinations were carried out.Platelet dense granules in the two probands were observed with an electron microscope.DNA was extracted from peripheral venous blood collected from the subjects.The pathogenic genes were screened by whole exome sequencing.The potential disease-causing variations were analyzed by bioinformatics analysis.Validation and family cosegregation analysis of the pathogenic variations were performed by Sanger sequencing.The relationship between HPS-related gene variations and clinical characteristics was explored.This study adhered to the Declaration of Helsinki.The study protocol was approved by the Ethics Committee of Ningxia Eye Hospital, People's Hospital of Ningxia Hui Autonomous Region (No.2016018).Written informed consent was obtained from each subject or custodian before any medical examination.Results:The two families were consistent with autosomal recessive inheritance pattern.In family 1 with a family history of consanguineous marriage, the proband had no obvious hypopigmentation on his facial skin, hair, eyebrows and eyelashes.Horizontal nystagmus, exotropia, mild visual impairment, iris atrophy, positive light transmission, orange fundus, pigment loss, macular hypoplasia, prolonged prothrombin time in laboratory examination, and a significant reduction of platelet dense granules by electron microscopy were observed.The proband in family 2 had pale brown hair and eyebrows, severe visual impairment, normal iris pigment, longer thrombin time in laboratory tests, and characteristics similar to those of the proband in family 1.A novel homozygous variant c. 2887G>T (p.E963X) was detected in the HPS3 gene of the proband in family 1.The parents of the proband from family 1 both carried a heterozygous variant c.2887G>T (p.E963X).Compound heterozygous variants were detected in HPS5 gene of the proband in family 2, c.2952-2A>C splicing variation and heterozygous deletion (a 3 144-bp deletion, located in chr11: 18302108-18305251, exon 22).The parents of the proband from family 2 carried a heterozygous variation.The three novel variations were labeled as pathogenic according to the ACMG standards and guidelines. Conclusions:Family 1 is with HPS-3 and family 2 is with HPS-5.There is a certain genotype-phenotype correspondence in the two types of HPS.

Objective:To analyze the genotype of hereditary eye diseases with early-onset high myopia (eoHM) and its relationship with phenotype.Methods:The families with eoHM were collected in Ningxia Eye Hospital from January 2019 to June 2020.The medical records of the probands and their family members were inquired and recorded in detail, and the relevant ocular examinations were performed.Peripheral venous blood samples were collected from patients and their family members, and whole-genome DNA was extracted.Sequence capture sequencing technology was applied to screen for disease-causing gene mutations in probands.The detected suspected pathogenic variants were verified by Sanger sequencing and were analyzed by family cosegregation analysis.According to ACMG guidelines, the pathogenicity of novel variants was evaluated.The original literature about hereditary eye diseases with eoHM was searched to analyze the relationship between mutated genes and clinical phenotype.This study protocol adhered to the Declaration of Helsinki.All subjects or their guardians were informed of the purpose and procedure of the study and signed the informed consent form.The study protocol was approved by the Ethics Committee of the People's Hospital of Ningxia Hui Autonomous Region (No.2016018).Results:A total of 20 eoHM families were collected, among which pathogenic variants associated with inherited eye diseases were detected in 8 families.Of the 8 probands, two were diagnosed with familial exudative vitreoretinopathy, one with X-linked retinitis pigmentosa, one with congenital stationary nightblindness, one with Stickler syndrome, one with achromatopsia, one with Leber congenital amaurosis, and one with gyrate atrophy of the choroid and retina.The first diagnosis age of the 8 probands was 4-7 years old, and they were all diagnosed as high myopia, with a refractive status ≤-6.00 DS.Genetic tests showed that the 8 probands carried a heterozygous variant c. 313A>G (p.M105Val) in FZD4 gene, a heterozygous variant c. 14_15insAAGA (p.Asp5fs *) in TSPAN12 gene, a heterozygous frameshift variant c. 2234_2237del (p.Arg745fs) in RPGR gene, a compound heterozygous variant of c. 481C>T (p.Gln161Ter *) and c. 355>T (p.Arg119Cys *) in GPR179 gene, a frameshift variant c. 1659_1660insACGGTGACCCTGGCCGTCCTGG (p.Pro554fs *) in COL2A1 gene, a compound heterozygous variant of c. 1811C>T (p.Thr604Ile *) and c. 967G>A (p.Gly323Ser) in PDE6B gene, a compound heterozygous variant of c. 604_619delTCCACGGCACTCAGGG (p.Ser202fs *) and c. 995G>C (p.Arg332Pro) in GUCY2D gene, a homozygous variant c. 772C>T (p.Pro241Leu) in OAT gene.Seven of them were novel variants.Compared with the previous literature, the clinical and gene phenotypes of the 8 families were analyzed in detail in this study, which provided the basis for the diagnosis of hereditary eye diseases with eoHM. Conclusions:EoHM is closely related to some hereditary eye diseases, which may be the reason for the early diagnosis of children and an important clue for clinicians to detect potential hereditary eye diseases.Further clinical evaluations of ocular structure and function as well as genetic screening in children with eoHM are recommended.

In order to develop a simple and efficient site-directed mutagenesis solution, the Gibson assembly technique was used to clone the cyclin dependent kinase 4 gene with single or double site mutations, with the aim to simplify the overlap extension PCR. The gene fragments containing site mutations were amplified using a strategy similar to overlap extension PCR. Meanwhile, an empty plasmid was digested by double restriction endonucleases to generate a linearized vector with a short adaptor overlapping with the targeted gene fragments. The gene fragments were directly spliced with the linearized vector by Gibson assembly in an isothermal, single-reaction, creating a recombinant plasmid. After the recombinant plasmids were transformed into competent Escherichia coli DH5α, several clones were screened from each group. Through restriction analysis and DNA sequencing, it was found that the randomly selected clones were 100% target mutants. Since there was neither tedious multiple-round PCR amplification nor frequent DNA extraction operation, and there was no need to digest the original plasmid, this protocol circumvents many factors that may interfere with the conventional site-directed mutagenesis. Hence, genes with single or multiple mutations could be cloned easily and efficiently. In summary, the major defects associated with overlap extension PCR and rolling circle amplification were circumvented in this protocol, making it a good solution for site-directed mutagenesis.
Clone Cells ; Mutagenesis, Site-Directed ; Mutation ; Plasmids/genetics* ; Polymerase Chain Reaction/methods*

BACKGROUND:Spinal cord injury not only causes serious physical and psychological injuries to patients but also brings a heavy economic burden to society.Spinal cord injury is initially triggered by mechanical trauma,followed by secondary injuries,and as the disease progresses,a glial scar develops. OBJECTIVE:To summarize the pathological process of spinal cord injury and strategies for stem cell transplantation to repair spinal cord injury,aiming to provide the best protocol for treating spinal cord injury. METHODS:Computer search was used to search PubMed and CNKI databases.Chinese search terms were"stem cell transplantation,spinal cord injury".English search terms were"stem cell,spinal cord injury,spinal cord,mesenchymal stem cells,neural stem cells,pathophysiology,clinical trial,primary injury,secondary injury".The literature was screened according to the inclusion and exclusion criteria.Finally,91 articles were included for review analysis. RESULTS AND CONCLUSION:(1)The strategies for repairing spinal cord injury through stem cell transplantation can be divided into exogenous stem cell transplantation and endogenous stem cell transplantation.The exogenous stem cell transplantation strategy for the treatment of spinal cord injury is divided into four kinds:injecting stem cells into the site of injury;transplantation of biomaterials loaded with stem cells;fetal tissue transplantation;transplantation of engineered neural network tissue or spinal cord-like tissue.(2)Compared with a single treatment method,combination therapy can more effectively promote nerve regeneration and spinal cord function recovery.(3)Microenvironment regulating the injury site,magnetic stimulation,electrical stimulation,epidural oscillating electric field stimulation,transcription factor overexpression and rehabilitation therapy can be combined with stem cell transplantation for combination therapy,thereby promoting the recovery of spinal cord function.