Objective To construct a uricase-deficient mouse model with stable inheritance using the CRISPR/Cas9 system,and evaluate its ability to simulate the disease characteristics of patients with hyperuricemia.Methods Double single guide RNAs(sgRNAs)were designed on both sides of exon 2～4 of the Uox gene.sgRNA and Cas9 mRNA for gene knockout were microinjected into the fertilized eggs of mice.After culture for 2～4 h,the embryos were transferred to surrogate mother mice to produce an F0 generation.Uox-knockout mice were identified by polymerase chain reaction and sequencing analysis.Positive mice were then mated with wild-type(WT)mice to produce an F1 generation,and heterozygous female and male F1 mice were then selected to obtain homozygous F2 mice.Serum and urine levels of uric acid,creatinine,and urea,and serum alanine aminotransferase(ALT)and aspartate aminotransferase(AST)levels were detected and compared between homozygous and wild-type mice.Pathological changes in kidney and liver tissues were observed by hematoxylin and eosin and Masson staining.Results Urine levels of serum uric acid(male:(4116.8±1928.1)μmol/L,P＜0.001;female:(2998.0±547.7)μmol/L,P＜0.01)and serum levels of uric acid(male:(478.4±114.6)μmol/L,P＜0.001;female:(507.7±129.6)μmol/L,P＜0.001),creatinine((91.8±55.6)μmol/L,P＜0.001),urea((28.6±13.9)mmol/L,P＜0.05),ALT((53.3±23.3)U/L,P＜0.01),and AST((203.3±70.3)U/L,P＜0.001)were significantly increased in Uox-/-mice compared with WT mice.Histopathological examination showed moderate hepatocyte degeneration in the liver,moderate-to-severe tubular cystic dilation,degeneration,and fibrosis in the kidney,glomerular hypertrophy and hyperplasia,small-vessel dilation and congestion,and infiltration of stromal monocytes and lymphocytes in Uox-/-mice.Conclusions We successfully established a homozygous uricase-deficient mouse strain using CRISPR/Cas9 technology,as a suitable animal model for research in the field of hyperuricemia.

Objective To construct a uricase-deficient mouse model with stable inheritance using the CRISPR/Cas9 system,and evaluate its ability to simulate the disease characteristics of patients with hyperuricemia.Methods Double single guide RNAs(sgRNAs)were designed on both sides of exon 2～4 of the Uox gene.sgRNA and Cas9 mRNA for gene knockout were microinjected into the fertilized eggs of mice.After culture for 2～4 h,the embryos were transferred to surrogate mother mice to produce an F0 generation.Uox-knockout mice were identified by polymerase chain reaction and sequencing analysis.Positive mice were then mated with wild-type(WT)mice to produce an F1 generation,and heterozygous female and male F1 mice were then selected to obtain homozygous F2 mice.Serum and urine levels of uric acid,creatinine,and urea,and serum alanine aminotransferase(ALT)and aspartate aminotransferase(AST)levels were detected and compared between homozygous and wild-type mice.Pathological changes in kidney and liver tissues were observed by hematoxylin and eosin and Masson staining.Results Urine levels of serum uric acid(male:(4116.8±1928.1)μmol/L,P＜0.001;female:(2998.0±547.7)μmol/L,P＜0.01)and serum levels of uric acid(male:(478.4±114.6)μmol/L,P＜0.001;female:(507.7±129.6)μmol/L,P＜0.001),creatinine((91.8±55.6)μmol/L,P＜0.001),urea((28.6±13.9)mmol/L,P＜0.05),ALT((53.3±23.3)U/L,P＜0.01),and AST((203.3±70.3)U/L,P＜0.001)were significantly increased in Uox-/-mice compared with WT mice.Histopathological examination showed moderate hepatocyte degeneration in the liver,moderate-to-severe tubular cystic dilation,degeneration,and fibrosis in the kidney,glomerular hypertrophy and hyperplasia,small-vessel dilation and congestion,and infiltration of stromal monocytes and lymphocytes in Uox-/-mice.Conclusions We successfully established a homozygous uricase-deficient mouse strain using CRISPR/Cas9 technology,as a suitable animal model for research in the field of hyperuricemia.

Objective To establish a human α-synuclein nuclear localization signal transgenic mouse model and investigate the effects of α-synuclein nuclear localization on the behavior of mice.Methods Human α-synuclein nuclear localization signal and EGFP lentiviral vectors were constructed.Transgenic mice were created with the microinjection method.Using PCR and Western Blot method to identify the genotypes and protein expression of the transgenic founder mice and their offsprings.The immunofluorescence was used to examine the localization of human α-synuclein in the mouse brain tissue.The behavioral changes of the transgenic mice were evaluated by the open field test,rotarod test,and O maze test.Results The h SNCA-NLS gene was successfully inserted into the mouse genome,the human α-syn was successfully expressed,and the human α-syn has localized with the nuclear.Further studies found that human α-synuclein nuclear localization signal transgenic mice had significant motor dysfunction,astrocyte proliferation and inflammatory response at 2 months of age and exhibited significant anxiety-like symptoms and reduced expression of the γ-aminobutyric acid(GABA)gene at 9 months of age,which persisted until 12 months of age.Conclusions A human α-synuclein nuclear localization signal transgenic mouse model has been successfully established.The mice exhibit significant motor dysfunction and anxiety-like symptoms.The successful establishment of this model provides a foundation for studying the role of α-syn nuclear localization in Parkinson's disease.

Background: Mature oocytes at the metaphase II status (MII-stage oocytes) played an important role in assisted reproductive technology in non-human primates. Objectives: In order to improve the proportion of MII-stage oocytes retrieval, three different superovulation protocols were performed on 24 female cynomolgus monkeys. Methods: All the monkeys received once-daily injection of follicle-stimulating hormone (25 international unit [IU]) on day 3 of the menstruation, 3-day intervals, twice daily for 8–12 days until the time of human chorionic gonadotropin (1,500 IU) injection, on the 14–17th day of menstruation collecting oocytes. The difference between protocol I and protocol II was that 0.1 mg the gonadotropin-releasing hormone agonist was injected on day 1 of the menstruation, while the difference between personalized superovulation protocol and protocol II was that oocytes could be collected on the 14–17th day of menstrual cycle according to the length of each monkey. Results: The total number of oocytes harvested using the personalized superovulation protocol was much higher than that using protocol I (p < 0.05), and the proportion of MII-stage oocytes was significantly greater than that from either superovulation protocol I or II(p < 0.001 and p < 0.01 respectively), while the proportion of immature oocytes at the germinal vesicle was less than that from superovulation protocol I (p < 0.05). Conclusions The personalized superovulation protocol could increase the rate of MII-stage oocytes acquired, and successfully develop into embryos after intracytoplasmic sperm injection, and eventually generated fetus.

ObjectiveTo investigate the association of metabolic syndrome (MS) and its components with the overall survival of pancreatic cancer (PC) patients who do not receive antitumor therapy. MethodsA retrospective analysis was performed for the data of patients who were diagnosed with PC in The Affiliated Hospital of Southwest Medical University from August 2013 to November 2018. Related data were collected, including age, sex, body weight, body height, body mass index (BMI), smoking, drinking, medical history of chronic pancreatitis, medical history of biliary tract diseases and gastritis, medical history of chronic hepatitis B/C, medical history of other tumors, presence or absence of PC in first-grade relatives, blood glucose, blood pressure, high-density lipoprotein cholesterol (HDL-C), triglyceride (TG), MS, and TNM stage. The log-rank test was used for comparison of survival curves between groups, and the univariate and multivariate Cox regression analyses were used to investigate the influencing factors for survival. ResultsA total 269 PC patients were enrolled in this study, with an average survival time of 3 months. The survival analysis showed no significant difference in survival time between the patients with MS and those without MS (P=0.754). There was no significant difference in median survival time between the patients with hypertension, high TG, high HDL-C, or abnormal BMI and those without such abnormality (all P＞0.05). There was a significant difference in median survival time between the patients with hyperglycemia and those without hyperglycemia (hazard ratio ［HR］=1.322, 95% confidence interval ［CI］: 0.985-1.775, P=0.028), and the multivariate Cox regression analysis achieved consistent results (HR=1481, 95% CI: 1.043-2.104, P=0.028). The analysis of the influencing factors for survival time in patients with stage Ⅳ PC showed that the patients with hyperglycemia had a significant reduction in median survival time (HR=1.524, 95%CI: 1.046-2.218, P=0004). ConclusionMS is not an influencing factor for the survival of PC patients, but hyperglycemia is an independent risk factor for poor prognosis in PC patients, especially in those with advanced PC.

ObjectiveTo explore the characteristics and correlation of intelligence development and socioemotional development in 1~3 years old children with expressive language disorder,and provide a basis for early intervention.Methods125 cases with expressive language disorder(language disorder group) and 126 normal children (normal group) are evaluated in two parts (intelligence and social mood) with 0~6 years old neuropsychological development diagnosis scale and Chinese version of urban infant-toddler social and emotional assessment.Results (1)The scores in language disorder group were lower than normal group in the area of free movements (81.60±10.40 vs 89.62±7.94),adaptability (81.48±10.95 vs 91.25±8.89),language (67.46±7.20 vs 89.13±8.24),social behavior (76.61±9.73 vs 90.12±8.13) and total developmental quotient (80.17±6.39 vs 91.15±6.05) (P<0.05).(2) There were significant differences between language disorder group and normal group in the area of externalizing behavior(56.28±10.15 vs 53.57±7.91),deregulation (56.45±10.61 vs 51.11±9.32) and capabilities(46.79±9.08 vs 51.25±7.47) (P<0.05).The boys' scores in internalizing behavior were lower than the girls' (49.19±10.76 vs 54.71±9.90) (P<0.05).(3) The scores of gross motor had positive correlation with externalizing behavior (r=0.220,P<0.05).The scores of language had negative correlation with externalizing behavior and deregulation(r=-0.650,P<0.05;r=-0.470,P<0.05).The scores of social behavior had negative correlation with externalizing behavior (r=-0.208,P<0.05).There was also a negative correlation between total development and deregulation (r=-0.184,P<0.05).(4) Multiple stepwise linear regression results showed that the externalizing behavior domain and mothers' education could predict 44.1% of variances in the language area.ConclusionThe children with expressive language disorder not only show backward in level of language development,but also in the development of other areas,and also bad social behavior and emotional problems.Individualized parenting interventions should be adopted to promote these children's intelligent development in an all-round way.