Objective:To investigate the signaling pathway of inhibiting macrophage phagocytosis of TIR domain-containing protein encoded by Escherichia coli (TcpC) N-terminal ubiquitin ligase active fragments of uropathogenic Escherichia coli. Methods:Bioinformatics software was used to analyze the amino acid sequences and the function of TcpC N-terminal ubiquitin ligase active fragments as well as the functional sites. PCR was performed to amplify tcpc-330, tcpc-450 and tcpc-510 genes and a prokaryotic expression system was constructed to express the target proteins. The recombinant proteins rTcpC-N110, rTcpC-N150 and rTcpC-N170 were purified by Ni-NTA affinity chromatography. LPS in the recombinant proteins was removed by Detoxi-gel chromatography. The expression of MyD88 at protein and mRNA levels in macrophages incubated with rTcpC-N110, rTcpC-N150, rTcpC-N170 or rTcpC-TIR was detected by Western blot and qRT-PCR. The activation of NF-κB signal pathway and the levels of proinflammatory factors in macrophages incubated with the above TcpC protein fragments were measured by Western blot and ELISA, respectively. Results:Cys12, Trp104 and Trp106 in the N-terminal fragment of TcpC were crucial amino acids in maintaining its ubiquitin ligase activity. The target recombinant proteins rTcpC-N110, rTcpC-N150 and rTcpC-N170 were successfully expressed and purified. After Detoxi-gel chromatography, rTcpC-N110, rTcpC-N150 and rTcpC-N170 extracts were undetectable for LPS. TcpC ubiquitin ligase fragments inhibited the expression of MyD88 at protein level, but not affect its expression at mRNA level in macrophages. LPS-induced phosphorylation of NF-κB signaling pathway-related proteins p50 and p65 was significantly inhibited in macrophages treated with TcpC ubiquitin ligase fragments. Moreover, LPS-induced production of pro-inflammatory factors was also significantly inhibited.Conclusions:The recombinant proteins rTcpC-N110, rTcpC-N150 and rTcpC-N170 could inhibit the expression of MyD88 at protein level and suppress the activation of NF-κB signaling pathway, suggesting that they were closely related to the inhibition of innate immune activity of macrophages.

Objective:To investigate whether the hypervirulent Klebsiella pneumoniae (hvKP) induces liver abscess through activating NLRP3 inflammasome. Methods:K1-hvKP and K35-non-hvKP bacterial suspensions were intraperitoneally injected into C57BL/6 mice to establish the models of liver abscess. Human peripheral blood neutrophils were sorted by immunomagnetic beads with CD45 + and Gr-1 + , and the purity was detected by flow cytometry. The concentrations of capsular polysaccharide of K1-hvKP and K35-non-hvKP were detected by total carbohydrate assay kit. The expression of IL-18 and IL-33 by neutrophils at mRNA and protein levels was detected by real-time fluorescence quantitative PCR and ELISA, respectively. The activation of NLRP3 inflammasome in neutrophils was detected by Western blot. Neutrophil extracellular trap formation (NETosis) was observed under confocal laser scanning microscope. Results:The C57BL/6 mice with K1-hvKP infection had significantly serious liver abscess as compared with the K35-non-hvKP-infected mice. The purity of human neutrophils was more than 95%. The concentration of capsular polysaccharide in K1-hvKP was significantly higher than that in K35-non-hvKP. Compared with K35-non-hvKP, K1-hvKP significantly promoted the neutrophils to express IL-18 and IL-33 at both mRNA and protein levels, enhanced the activation of NLRP3 and induced NETosis.Conclusions:This study suggested that hvKP could promote NETosis by activating NLRP3 inflammasome to cause liver abscess.

Objective:To investigate the role of TcpC in uropathogenic Escherichia coli (UPEC)-induced cystitis in mice and to preliminarily analyze the pathogenic mechanism. Methods:C57BL/6 mice were injected with 10 9 CFU wild-type UPEC CFT073 (CFT073 wt) or tcpc gene-deleted mutant (CFT073 Δ tcpc) from urethra into bladder to construct the mouse model of cystitis. The mice were sacrificed 3 d after infection and the bladders were taken to observe the gross pathological changes. Histopathological changes in bladder tissues were observed after HE staining. Immunohistochemistry was used to detect TcpC in bladder tissues. Bacterial loads in urine samples of UPEC-infected mice were counted by tenfold dilution method, and the presence of tcpc gene in the genomic DNA of bacteria from the bladder and urine samples of CFT073 wt-infected mice was measured by PCR. Real-time quantitative RT-PCR (qRT-PCR) and Western blot were performed to detect the expression of TcpC at mRNA and protein levels in macrophages after CFT073 wt infection. The influence of UPEC strains on the activation of NF-κB signaling pathway in macrophages were determined by Western blot. The levels of proinflammatory factors and the bacterial and cell activity after infecting macrophages with UPEC strains were detected by ELISA, laser confocal microscope and fluorescence microscope, respectively. Results:Compared with the mice with CFT073 Δ tcpc infection, CFT073 wt-infected mice had significantly enlarged bladder and severe neutrophil infiltration and abundant TcpC in bladder tissues. The number of bacteria in the urine of CFT073 wt-infected mice was significantly greater than that of the CFT073 Δ tcpc group. PCR results showed that the bacteria in bladder or urine were CFT073 wt. The expression of TcpC at both mRNA and protein levels in macrophages increased significantly after CFT073 wt infection. Moreover, in CFT073 wt-infected macrophages, the expression of IκBα was promoted and the phosphorylation of p65 and the production of proinflammatory factors were suppressed. TcpC was instrumental in the survival and invasion of CFT073 wt in macrophages. Conclusions:TcpC expression increased significantly in mice with CFT073 wt-induced cystitis. TcpC inhibited the activation of NF-κB signaling pathway and the production of proinflammatory factors in macrophages to improve the survival rate of CFT073 wt, which was closely related to the pathogenesis and immune evasion of UPEC.

ObjectiveTo explore the related factors of troublemaking behaviors among patients with mental disorders induced by amphetamine-type stimulants （ATS）， and to provide references for the formulation of relevant intervention measures for ATS-induced mental disorders. MethodsA total of 105 patients who met the diagnostic criteria of International Classification of Diseases， tenth edition （ICD-10） for ATS-induced mental disorders were included， and classified into troublemaking group and non-troublemaking group. The general demographic data and clinical data of the selected individuals were collected， and all patients were assessed using Social Support Rating Scale （SSRS）. Then univariate analysis and multivariate Logistic regression model were used to screen the related factors of troublemaking behaviors. ResultsThe scores of SSRS， objective support dimension and social support utilization dimension were significantly lower in troublemaking group than those in non-troublemaking group， with statistical differences ［（24.10±6.59） vs. （28.94±5.59）， t=3.364， P=0.001； （5.50±1.96） vs. （8.20±2.13）， t=5.183， P<0.01； （4.60±2.26） vs. （6.28±1.90）， t=3.435， P=0.001］. Multivariate Logistic regression analysis showed that male （OR=6.061， P=0.014） was a risk factor， while high social support level （OR=0.873， P=0.018） was the protective factor for troublemaking behaviors among patients with ATS-induced mental disorders. ConclusionPatients with ATS-induced mental disorders of the males and with low social support level are at high risk of troublemaking behaviors.

Objective To assess the longitudinal mitral annular plane systolic excursion ( M APSE) of different directions in normal fetuses during mid‐late pregnancy based on two‐dimensional speckle tracking imaging ( ST I) . Methods Seventy‐six normal fetuses during middle and late pregnancy were selected at 26－32 weeks of gestation . T he peak M APSE was measured by free angle M‐mode echocardiography ( FAM ) perpendicular to the lateral annulus in the mitral annular plane . The time‐displacement curves of interventricular septal mitral annulus in three different directions including points A ,B and C through transverse level of apex were recorded by STI . T he peak M APSE of interventricular septal mitral annulus ( SEPT‐M APSE‐A ,SEPT‐M APSE‐B ,SEPT‐M APSE‐C) in three different directions including points A ,B and C and the time to peak ( T T P :SEPT‐T T P‐A ,SEPT‐T T P‐B ,SEPT‐T T P‐C) were recorded respectively . T he time‐displacement curves of lateral mitral annulus in three different directions including points A ,B and C through transverse level of apex were recorded by STI . T he peak M APSE of lateral mitral annulus ( LAT‐M APSE‐A ,LAT‐MAPSE‐B ,LAT‐MAPSE‐C) in three different directions including points A ,B and C ,the time to peak( LA T‐T T P‐A ,LA T‐T T P‐B ,LA T‐T T P‐C) were recorded respectively . Finally ,the data were analyzed statistically . Results T he peak M APSE of the lateral mitral annulus in 3 different directions including points A ,B and C[ LA T‐M APSE‐A ( 3 .62 ± 1 .01) mm ,LA T‐M APSE‐B ( 3 .95 ± 1 .04) mm ,LAT‐M APSE‐C ( 4 .45 ± 1 .05) mm ] were greater than those of the interventricular septum mitral annulus[ SEPT‐MAPSE‐A (3 .41 ± 0 .63)mm ,SEPT‐MAPSE‐B (3 .07 ± 0 .50) mm ,SEPT‐MAPSE‐C (2 .82 ± 0 .51) mm] . LAT‐M APSE‐C and SEPT‐M APSE‐A were the largest longitudinal excursions of mitral annulus . T he differences were statistically significant in points B and C ( P ＜0 .05) . T here was no significant difference in point A ( P ＞0 .05) . LA T‐M APSE‐C was less than FAM‐M APSE [ ( 6 .06 ± 1 .35 ) mm ] . T here was a significant difference between them ( P ＜0 .05 ) . Strong correlation was found between them ( r ＝0 .896 , P＜0 .05) . T here were no significant differences in the time to peak of interventricular septal mitral annulus [ SEPT‐T T P‐A ( 0 .210 ± 0 .008 ) s ,SEPT‐T T P‐B ( 0 .213 ± 0 .008 ) s ,SEPT‐T T P‐C ( 0 .210 ± 0 .005 ) s] in directions including points A ,B ,C ( P＞ 0 .05 ) . T here were no significant differences in time to peak of lateral mitral annulus [ LAT‐T T P‐A ( 0 .210 ± 0 .008 ) s , LAT‐T T P‐B ( 0 .213 ± 0 .006 ) s , LAT‐T T P‐C ( 0 .210 ± 0 .007) s] in directions inclucling points A ,B ,C ( P ＞0 .05) . Conclusions Longitudinal systolic motion of fetal left ventricular wall during mid‐late pregnancy has good synchronization . Longitudinal motion of fetal mitral annulus is a comprehensive movement of multiple directions and different degrees of displacement ,with the movement perpendicular to the annulus as the maximum displacement direction . T he displacement parameters of mitral annulus measured by ST I can reflect the left ventricular longitudinal systolic function and have clinical application value in evaluating the left ventricular longitudinal systolic function of fetuses .

Objective: To assess the longitudinal mitral annular plane systolic excursion (MAPSE) of different directions in normal fetuses during mid-late pregnancy based on two-dimensional speckle tracking imaging (STI). Methods: Seventy-six normal fetuses during middle and late pregnancy were selected at 26-32 weeks of gestation. The peak MAPSE was measured by free angle M-mode echocardiography (FAM) perpendicular to the lateral annulus in the mitral annular plane. The time-displacement curves of interventricular septal mitral annulus in three different directions including points A, B and C through transverse level of apex were recorded by STI. The peak MAPSE of interventricular septal mitral annulus (SEPT-MAPSE-A, SEPT-MAPSE-B, SEPT-MAPSE-C) in three different directions including points A, B and C and the time to peak (TTP: SEPT-TTP-A, SEPT-TTP-B, SEPT-TTP-C) were recorded respectively. The time-displacement curves of lateral mitral annulus in three different directions including points A, B and C through transverse level of apex were recorded by STI. The peak MAPSE of lateral mitral annulus (LAT-MAPSE-A, LAT-MAPSE-B, LAT-MAPSE-C) in three different directions including points A, B and C, the time to peak(LAT-TTP-A, LAT-TTP-B, LAT-TTP-C) were recorded respectively. Finally, the data were analyzed statistically. Results: The peak MAPSE of the lateral mitral annulus in 3 different directions including points A, B and C[LAT-MAPSE-A (3.62±1.01)mm, LAT-MAPSE-B (3.95±1.04)mm, LAT-MAPSE-C (4.45±1.05)mm] were greater than those of the interventricular septum mitral annulus[SEPT-MAPSE-A (3.41±0.63)mm, SEPT-MAPSE-B (3.07±0.50)mm, SEPT-MAPSE-C (2.82±0.51)mm]. LAT-MAPSE-C and SEPT-MAPSE-A were the largest longitudinal excursions of mitral annulus. The differences were statistically significant in points B and C (P<0.05). There was no significant difference in point A (P>0.05). LAT-MAPSE-C was less than FAM-MAPSE[(6.06±1.35)mm]. There was a significant difference between them(P<0.05). Strong correlation was found between them(r=0.896, P<0.05). There were no significant differences in the time to peak of interventricular septal mitral annulus [SEPT-TTP-A (0.210±0.008)s, SEPT-TTP-B (0.213±0.008)s, SEPT-TTP-C (0.210±0.005)s] in directions including points A, B, C(P>0.05). There were no significant differences in time to peak of lateral mitral annulus[LAT-TTP-A(0.210±0.008)s, LAT-TTP-B(0.213±0.006)s, LAT-TTP-C(0.210±0.007)s] in directions inclucling points A, B, C(P>0.05). Conclusions Longitudinal systolic motion of fetal left ventricular wall during mid-late pregnancy has good synchronization. Longitudinal motion of fetal mitral annulus is a comprehensive movement of multiple directions and different degrees of displacement, with the movement perpendicular to the annulus as the maximum displacement direction. The displacement parameters of mitral annulus measured by STI can reflect the left ventricular longitudinal systolic function and have clinical application value in evaluating the left ventricular longitudinal systolic function of fetuses.

Objective: To explore the genetic basis for a pedigree affected with Marfan syndrome (MFS). Methods: Clinical data of the patients was collected.With genomic DNA extracted from peripheral blood samples, potential mutation was detected by targeted exome sequencing.Candidate variants were validated by Sanger sequencing and bioinformatic analysis. Results: Targeted exome sequencing and Sanger sequencing revealed a missense c. 649T>C(p.Trp217Arg) variant in the exon 7 of FBN1 gene, which was unreported previously.Bioinformatics analysis suggested that the variant can cause amino acid replacement and affect the structure and function of fibrillin-1. Conclusion A novel missense variant of the FBN1 gene was identified, which probably underlies the autosomal dominant MFS in this pedigree.

Objective To calculate Z-score for mitral and tricuspid color blood flow widths in normal fetuses and fetuses with dilated coronary sinuses ( CS ) using fetal echocardiography ,and explore the application value of Z-score of the color flow widths of atrioventricular valves in normal fetuses and fetuses with dilated CS . Methods Two hundred and thirty-eight normal fetuses (control group) with a gestational age of 16 to 38 weeks were studied by color Doppler echocardiography . Gestational age ( GA ) ,biparietal diameter (BPD) ,femoral length (FL) ,aortic inner diameter (AOd) ,pulmonary artery diameter (PAd) ,and heart area ( HA) were measured as independent variables ,and mitral and tricuspid valve color flow widths were measured as the dependent variables . Z-score models were established by regression analysis . Thirty fetuses with dilated CS (dilated CS group) from 22 to 33 weeks'gestation were involved . The Z-score of the CS fetus was calculated based on the established Z-score models and were compared with those of the normal fetuses . Results The independent sample t-test showed that there were no significant differences in the Z-scores of the blood flow width of the fetal mitral and tricuspid valves between dilated CS group and control group ( P >0 .05) . Conclusions The simple dilated CS does not affect the mitral valve diastolic blood flow ,so there is no significant effect on the filling of left ventricular blood flow .

OBJECTIVE: To assess the clinical application of single nucleotide polymorphism microarray (SNP array) in patients with intellectual disability/developmental delay(ID/DD). METHODS: SNP array was performed to detect genome-wide DNA copy number variants (CNVs) for 145 patients with ID/DD in Women's Hospital, Zhejiang University School of Medicine from January 2013 to June 2018. The CNVs were analyzed by CHAS software and related databases. RESULTS: Among 145 patients, pathogenic chromosomal abnormalities were detected in 32 cases, including 26 cases of pathogenic CNVs and 6 cases of likely pathogenic CNVs. Meanwhile, 18 cases of uncertain clinical significance and 14 cases of likely benign were identified, no significant abnormalities were found in 81 cases (including benign). CONCLUSIONS SNP array is effective for detecting chromosomal abnormalities in patients with ID/DD with high efficiency and resolution.
Chromosome Aberrations ; DNA Copy Number Variations ; Genome-Wide Association Study ; Humans ; Intellectual Disability ; diagnosis ; genetics ; Oligonucleotide Array Sequence Analysis ; standards ; Polymorphism, Single Nucleotide

OBJECTIVE: To assess the clinical application of single nucleotide polymorphism microarray (SNP array) in prenatal genetic diagnosis for fetuses with absent nasal bone. METHODS: Seventy four fetuses with absent nasal bone detected by prenatal ultrasound scanning were recruited from Women's Hospital, Zhejiang University School of Medicine during June 2015 and October 2018. The chromosome karyotypes analysis and SNP array were performed. The correlation between absent fetal nasal bone and chromosome copy number variants was analyzed. RESULTS: Among 74 fetuses, 19 were detected to have chromosomal abnormalities, including 16 cases of trisomy-21, 1 case of trisomy-18 and two cases of micro-deletion/duplication. Among 46 cases with isolated absence of nasal bone, 3 had trisomy-21, and 1 had a micro-duplication. Absence of nasal bone in association with nuchal translucency thickening had a higher rate of abnormal karyotypes compared with isolated absence of nasal bone (=32.27,<0.01). CONCLUSIONS Fetuses with absent nasal bone and nuchal translucency thickening are likely to have chromosome abnormalities, and SNP array testing is recommended to exclude the chromosome abnormalities.
Chromosome Aberrations ; Female ; Fetus ; Humans ; Nasal Bone ; abnormalities ; Oligonucleotide Array Sequence Analysis ; standards ; Polymorphism, Single Nucleotide ; genetics ; Pregnancy ; Pregnancy Trimester, First ; Prenatal Diagnosis ; methods