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.

BackgroundSchizophrenia is a common severe mental disorder with complex pathogenesis. There are few studies on the correlation between kynurenine metabolites in peripheral serum and urine in schizophrenia. ObjectiveTo investigate the concentration of tryptophan-kynurenine metabolites and interleukin-6 （IL-6） in serum and urine in patients with schizophrenia， and their correlation with clinical symptoms， so as to explore potential biological characteristics related to schizophrenia. MethodsA total of 38 patients with schizophrenia who met the criteria of the Diagnostic and Statistical Manual of Mental Disorders， fifth edition （DSM-5）， and were hospitalized or attended outpatient clinic at Hangzhou Seventh People's Hospital from December 2021 to December 2022 were included in the study. Additionally， 26 healthy individuals were concurrently recruited from the community of Hangzhou to serve as a control group. All participants were requested to complete the Positive and Negative Symptom Scale （PANSS）. The levels of tryptophan （TRP）， kynurenine （KYN）， kynurenic acid （KYNA）， quinolinic acid （QUIN）， picolinic acid （PIC）， xanthurenate and 5-hydroxytryptamine （5-HT） in both serum and urine were measured using ultra-high-performance liquid chromatography-triple quadrupole linear ion trap mass spectrometry. Serum and urine IL-6 were measured using enzyme-linked immunosorbent assay. Pearson correlation analysis was conducted to examine the correlation between serum and urinary KYN metabolites， as well as the correlation between metabolite levels and clinical symptoms in the patient group. ResultsPatients with schizophrenia had significantly higher level of IL-6 in serum （U=798.500， P<0.01） and lower level of PIC in urine （U=253.000， P=0.013） compared with the control group. Additionally， level of serum KYN was positively correlated with QUIN/KYNA ratio and QUIN/PIC ratio （r=0.562， 0.438， P<0.05） in patients with schizophrenia. 5-HT/KYN ratio in serum was positively correlated with PANSS total score and negative symptom subscale score （r=0.458， 0.455， P<0.01） in patients with schizophrenia. ConclusionSerum TRP-KYN pathway metabolite levels in patients with schizophrenia were associated with neurotoxic metabolite ratios in urine and the severity of negative symptoms. ［Funded by Zhejiang Medical and Health Science and Technology Program Exploratory （number， 2022KY990）］