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Objective:To investigate the molecular pathogenesis of a newly discovered gene mutation in a family with hereditary coagulation factor Ⅺ(FⅪ) deficiency.Methods:The proband was admitted to the First Affiliated Hospital of Wenzhou Medical University in September 2021 due to "calculus of intrahepatic duct". The patient had no symptoms of spontaneous bleeding.The clinical data and blood samples of the proband and her family members (10 persons in 3 generations) were collected.The activated partial thromboplastin time (APTT) and FⅪ activity (FⅪ:C) were performed by the one-stage clotting assay. FⅪ antigen (FⅪ:Ag) were detected by enzyme linked immunosorbent assay (ELISA). Genomic DNA extracted from peripheral blood cells of subjects was used as template to analyze F11 gene mutation by DNA direct sequencing. Bioinformatics software was used to analyze the effects of mutations on protein structure and function. Wild-type and mutant FⅪ protein expression vectors were constructed and transient transfected into HEK293T cells. The total RNA was extracted from positive transfected cells and then reversely transcribed into cDNA. The mRNA expression level of F11 gene in transfected cells was detected by real-time fluorescence quantitative PCR (qRT-PCR). The content of FⅪ:Ag and the expression of FⅪ protein in transfected cell lysates and culture supernatant were detected by ELISA and western blot.Results:The APTT of the proband was significantly prolonged to 107.9s (reference range 29.0-43.0s), while FⅪ:C and FⅪ:Ag were significantly decreased to 2% (reference range 84%-122%) and 5% (reference range 76%-127%), respectively. Gene sequencing analysis indicated that the proband had c.536C>T (p.Thr161Met) heterozygous missense mutation and c.1556G>A (p.Trp501Ter) heterozygous nonsense mutation in exon 6 and 13 of the F11 gene, respectively. Bioinformatics analysis showed that the amino acids at site 161 of FⅪ protein were threonine (Thr) in the matrix composed of five different species, indicating that Thr161 site was highly conserved among homologous genes in different species. p.Thr161Met heterozygous mutation affected the stability of local intermolecular structure of FⅪ protein. In vitro expression experiments of p.Thr161Met mutation showed that FⅪ protein had a normal synthesis in the cells but secretion dysfunction.Conclusions:c.536C>T (p.Thr161Met) heterozygous missense mutation and c.1556G>A (p.Trp501Ter) heterozygous nonsense mutation were mainly responsible for the decrease of FⅪ in this family. p.Thr161Met mutation was first reported in the world and did not affect the normal synthesis of FⅪ protein, but caused secretion dysfunction.
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A 66-year-old female patient presented with blisters on the scalp and neck for 1 month. She had a history of type 2 diabetes for 6 years, and started taking the dipeptidyl peptidase-4 inhibitor linagliptin at a dose of 5 mg once a day 6 months before the onset of eruption. Skin examination showed scattered mung bean- to peanut-sized blisters on the scalp, and some blisters broke with exudation and crusts. There was a pigeon egg-sized bulla and two mung bean-sized blisters on the left neck, with tense blister walls, clear blister fluids, non-erythematous base, and Nikolsky′s sign was negative. Enzyme-linked immunosorbent assay revealed that the serum levels of anti-BP180 NC16A and anti-BP230 antibodies were 5.81 and 139.76 kU/L respectively. Histopathological examination of the blister on the neck showed subepidermal blister formation, and infiltration with neutrophils and a few eosinophils in the blister. The patient was finally diagnosed with localized anti-BP230-type bullous pemphigoid. This case of anti-BP230-type bullous pemphigoid associated with the dipeptidyl peptidase-4 inhibitor linagliptin was firstly reported in China.
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OBJECTIVE@#To explore the molecular basis for a Chinese pedigree affected with hereditary coagulation factor VII (FVII) deficiency.@*METHODS@#The coding regions of F7 gene were amplified by PCR and sequenced. Suspected variants were confirmed by reverse sequencing and validated in other members from the pedigree. Pathogenicity of the variants was analyzed with multiple bioinformatic tools.@*RESULTS@#Genetic analysis revealed that the proband has carried compound heterozygous c.985T>C (p.Ser329Pro) and c.1091G>A (p.Arg364Gln) variants in exon 8 of the F7 gene. Her mother, brother and son were heterozygous for c.985T>C (p.Ser329Pro), while her father was heterozygous for c.1091G>A (p.Arg364Gln). Phylogenetic analysis suggested that both p.Ser329 and p.Arg364 are highly conserved among homologous species. Online bioinformatic software predicted both variants to be deleterious. Protein model analysis suggested that the Pro329 side chain may form a new hydrogen bond with Leu333. The Pro benzene ring may clash with Glu325 in the p.Ser329Pro variant model. The p.Arg364Gln variant have two additional hydrogen bonds compared with wild type Arg364. Both variants may lead to alteration of the protein structure.@*CONCLUSION@#The p.Ser329Pro and p.Arg364Gln variants in exon 8 of the F7 gene probably account for the reduced FVII in this pedigree.
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OBJECTIVE@#To identify potential mutations of F11 gene in a pedigree affected with hereditary coagulation factor XI (FXI) deficiency and explore its molecular pathogenesis.@*METHODS@#Prothrombin time (PT), activated partial thromboplastin time (APTT), fibrinogen (FIB), coagulation factor VIII activity (FVIIIC), coagulation factor IX activity (FIXC), coagulation factor XI activity (FXIC), coagulation factor XII activity (FXIIC) and lupus anticoagulation (LA) of the proband and eight family members were determined. FXI antigen (FXIAg) was determined by enzyme-linked immunosorbent assay (ELISA). For the proband, potential mutations in the exons, flanking introns and 5'-, 3'-untranslated regions of the F11 gene were screened by direct DNA sequencing. The results were confirmed by reverse sequencing. Suspected mutations were detected in other family members. ClustalX-2.1-win and four online bioinformatic tools (PolyPhen-2, PROVEAN, SIFT, and Mutation Taster) were used to study the conservation and possible impact of the mutations. The structure of the mutational sites was processed with Swiss-PdbViewer.@*RESULTS@#The propositus had prolonged APTT (69.6 s), whose FXIC and FXIAg were reduced to 6.0% and 10.7%, respectively. Her mother, elder sister, one younger sister, little brother, daughter and son showed slightly prolonged APTT and moderate FXIC and FXIAg levels. Gene sequencing revealed that the propositus carried a heterozygous nonsense mutation c.738G>A (p.Trp228stop) in exon 7 and a heterozygous mutation c.1556G>C (p.Trp501Ser) in exon 13. Her mother, elder sister and daughter were heterozygous for the p.Trp228stop mutation, while one younger sister and little brother and son were heterozygous for p.Trp501Ser. Her husband and the youngest sister were of the wild type. Phylogenetic analysis suggested that Trp501 was highly conserved among all homologous species. The p.Trp501Ser was predicted to be "probably damaging","deleterious", "affect protein function" and "disease causing" corresponding to PolyPhen-2, PROVEAN, SIFT and Mutation Taster. Model analysis demonstrated that the non-polar Trp501 has two benzene rings, forming a hydrogen bond with Gln512 in the wild type. Once substituted by Ser501, the side chain may form another hydrogen bond with the benzene of His396. This may affect the normal space conformation and stability of FXI protein.@*CONCLUSION@#The compound heterozygous mutations of the F11 gene probably accounted for the low FXI concentration in this pedigree.
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Female , Humans , Male , Factor XI , Genetics , Factor XI Deficiency , Genetics , Heterozygote , Mutation , Pedigree , PhylogenyABSTRACT
Objective@#To detect potential mutations of the coagulation factor Ⅶ (F7) gene in a pedigree affected with hereditary FⅦ deficiency and explore its molecular pathogenesis.@*Methods@#The FⅦ antigen (FⅦ∶Ag) was analyzed by an enzyme-linked immunosorbent assay (ELISA) method. Prothrombin time (PT), FⅦ activity (FⅦ∶C) and other coagulant parameters were quantified with an one-stage clotting assay. The F7 gene was amplified by PCR and sequenced. Mutational sites were confirmed by reverse sequencing. Impact of amino acid substitution was assessed using SIFT and PolyPhen-2 software. Structure of the mutant protein was analyzed using Swiss-pdb Viewer software based on the three-dimensional structure in the Protein Data Bank.@*Results@#The propositus had prolonged PT (36.3 s), with FⅦ∶C and FⅦ∶Ag significantly reduced to 2% and 44%, respectively. Her father, mother, younger sister and daughter had slightly prolonged PT and reduced FⅦ∶C (86%-120%). The FⅦ∶Ag of her father and younger sister were also reduced. DNA sequencing revealed that the propositus has carried compound heterozygous mutations (Lys341Glu and IVS6-1G>A) of the F7 gene. Her father and younger sister were heterozygous for the IVS6-1G>A mutation, while her mother and daughter were heterozygous for the Lys341Glu mutation. Bioinformatics analysis indicated that Lys341Glu mutation may affect the stability and function of the FⅦ protein.@*Conclusion@#The Lys341Glu and IVS6-1G>A mutations probably underlie the reduced activity of FⅦ in this pedigree.
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OBJECTIVE@#To detect potential mutations of the coagulation factor Ⅶ (F7) gene in a pedigree affected with hereditary FⅦ deficiency and explore its molecular pathogenesis.@*METHODS@#The FⅦ antigen (FⅦ:Ag) was analyzed by an enzyme-linked immunosorbent assay (ELISA) method. Prothrombin time (PT), FⅦ activity (FⅦ:C) and other coagulant parameters were quantified with an one-stage clotting assay. The F7 gene was amplified by PCR and sequenced. Mutational sites were confirmed by reverse sequencing. Impact of amino acid substitution was assessed using SIFT and PolyPhen-2 software. Structure of the mutant protein was analyzed using Swiss-pdb Viewer software based on the three-dimensional structure in the Protein Data Bank.@*RESULTS@#The propositus had prolonged PT (36.3 s), with FⅦ:C and FⅦ:Ag significantly reduced to 2% and 44%, respectively. Her father, mother, younger sister and daughter had slightly prolonged PT and reduced FⅦ:C (86%-120%). The FⅦ:Ag of her father and younger sister were also reduced. DNA sequencing revealed that the propositus has carried compound heterozygous mutations (Lys341Glu and IVS6-1G>A) of the F7 gene. Her father and younger sister were heterozygous for the IVS6-1G>A mutation, while her mother and daughter were heterozygous for the Lys341Glu mutation. Bioinformatics analysis indicated that Lys341Glu mutation may affect the stability and function of the FⅦ protein.@*CONCLUSION@#The Lys341Glu and IVS6-1G>A mutations probably underlie the reduced activity of FⅦ in this pedigree.
Subject(s)
Female , Humans , Male , Factor VII , Genetics , Factor VII Deficiency , Genetics , Genetic Testing , Heterozygote , Mutation , PedigreeABSTRACT
Objective The study aimed to construct a composite score method based on multidimensional quality indi-cators and conduct simulation trials to validate the method. Besides,the quality of breast cancer care for both hospitals and sur-geons was evaluated by the method. Methods The two-parameter logistic latent variable model was constructed as measure-ment model;the latent variables in the measurement model were further incorporated into multilevel structural model as depend-ent variables and one pseudo level was designed for representing multiple latent variables. MCMC method was used to estimate model parameters. Three level and two-dimensional latent variable model was used to analyze the actual data. Results The sim-ulation study showed that the number of quality indicators and surgeons should not be less than 20 to obtain efficient estimate of quality of care;the multilevel and multidimensional latent variable model was applied to analyze the data;surgeons and hospitals who provided superior quality of breast cancer diagnosis and operative procedure were obtained. Conclusion The newly con-structed multilevel and multidimensional latent variable model could effectively address the hieratical structure in quality of care data as well as the multidimensional nature of quality of care,thus,the model can be used to comprehensively and rationally as-sess the quality of care;comprehensive evaluation of quality of care provided ground for linking the ranking of hospitals and per-formance appraisal of surgeons to the quality of care.
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Objective To investigate the expression and clinical significance of mammalian sterile 20-like kinase 1 (MST1) in cervical cancer. Methods Immunohistochemical method was applied to detect the expression level of MST1 protein in specimens of cervical cancer tissues (n=139) and pericarcinomatous tissues (n=20, with≥4 cm distance from the primary tumor's edge). Western blot assay and qPCR were used to detect the protein and mRNA transcription expression levels of MST1 in 20 pairs of cervical cancer tissues and pericarcinomatous tissues, respectively. The correlation between MST1 expression, clinic pathological features and the prognosis were analyzed. Results MST1 was mainly expressed in cytoplasm. The positive expression rate of MST1 was significantly lower in cervical cancer tissues (27%, 38/139) than that in pericarcinomatous tissues (80%, 16/20,χ2=21.62, P<0.01). The expressions levels of MST1 protein and mRNA were both lower in the cervical cancer tissues (P<0.01). In cervical cancer, the positive expression rate of MST1 inⅠb+Ⅱa stage was higher than that ofⅡb+Ⅳstage (P<0.05), the positive expression rate of MST1 in lymph node metastasis was lower than that of without lymph node metastasis (P < 0.05). Values of age, tumor size, histological type and differentiation degree showed no significant difference to positive expression rate of MST1. Moreover, the negative expression of MST1 displayed a significantly poorer overall survival time than that of positive expression of MST1 (Log-rank χ2=28.35, P < 0.01). Conclusion MST1 shows a lower expression in cervical cancer, which may be a new target for clinical treatment and prognosis of cervical cancer.
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Objective The study aimed to construct a composite score method based on multidimensional quality indi-cators and conduct simulation trials to validate the method. Besides,the quality of breast cancer care for both hospitals and sur-geons was evaluated by the method. Methods The two-parameter logistic latent variable model was constructed as measure-ment model;the latent variables in the measurement model were further incorporated into multilevel structural model as depend-ent variables and one pseudo level was designed for representing multiple latent variables. MCMC method was used to estimate model parameters. Three level and two-dimensional latent variable model was used to analyze the actual data. Results The sim-ulation study showed that the number of quality indicators and surgeons should not be less than 20 to obtain efficient estimate of quality of care;the multilevel and multidimensional latent variable model was applied to analyze the data;surgeons and hospitals who provided superior quality of breast cancer diagnosis and operative procedure were obtained. Conclusion The newly con-structed multilevel and multidimensional latent variable model could effectively address the hieratical structure in quality of care data as well as the multidimensional nature of quality of care,thus,the model can be used to comprehensively and rationally as-sess the quality of care;comprehensive evaluation of quality of care provided ground for linking the ranking of hospitals and per-formance appraisal of surgeons to the quality of care.
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Objective To explore the association between the C46T polymorphism of coagulation factor Ⅻ (FⅫ) gene and the involvement of FⅫ activity (FⅫ:C) in patients with unexplained recurrent spontaneous abortion (URSA), and to elucidate its role in the pathogenesis of URSA. Methods This study included 203 patients with URSA (URSA group) and 171 healthy women with at least one child and no history of infertility or miscarriage (control group) in the southern area of Zhejiang Province. The C 46T polymorphism of the FⅫ gene was analyzed with matrix-assisted laser desorption/ionization time of flight-mass spectrometry (MALDI-TOF-MS) in all subjects. The values of prothrombin time, activated partial thromboplastin time (APTT), fibrinogen, FⅫ:C and other coagulant parameters were determined. The frequency distribution of the wild-type (CC), heterozygote (CT), homozygote (TT) genotypes and C and T alleles were compared between the patients and controls. A comprehensive analysis of association was conducted between C46T genotypes and the FⅫ:C levels in URSA patients. Results The CC, CT, TT genotypes of the FⅫgene were observed in 7 (3.4%, 7/203), 83 (40.9%, 83/203) and 113 (55.7%, 113/203) patients with URSA versus 7 (4.1%, 7/171), 46 (26.9%, 46/171) and 118 (69.0%, 118/171) controls. The frequency of CT in the patients with URSA was significantly higher than that in controls, but the frequency of TT in the patients was lower than that in controls (χ2=7.939, OR=1.884, 95%CI:1.210-2.935, P<0.05). The frequencies of allele C and allele T were observed in 97 (23.9%, 97/406) and 309 (76.1%, 309/406) patients with URSA versus 60 (17.5%, 60/342) and 282 (82.5%, 282/342) controls. The distribution frequency of allele T in URSA group was lower than that in control group (χ2=4.510, OR=1.475, 95%CI:1.029-2.115, P<0.05). The FⅫ:C levels in the patients were (102±13)%in CC genotype, (78±11)%in CT genotype and (59± 9)%in TT genotype, respectively. The differences of the FⅫ:C levels between the CC and CT, CT and TT, CC and TT genotypes in the patients were significant (all P<0.05). Conclusions The low level of FⅫ:C maybe result from the T allele of the FⅫgene in URSA patients. The CT genotype might be relative to the pathogenesis of URSA in a Chinese Han female population from the southern area of Zhejiang province.
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Objective To investigate the current situation of thyroid disease by analyzing retrospectively the color poppler ultrasonography data of neck discomfort in Heilongjiang Province.Methods A total of 13 020 cases (18-93 years old) of thyroid ultrasound examination data collected from September 2009 to October 2012 at the Department of Ultrasound,Fourth Affiliated Hospital of Harbin Medical University,were retrospectively analyzed.According to the sonographic features of thyroid (thyroid morphology,size,echo characteristics,blood flow,nodular location,number,calcification,etc),combined with ultrasound diagnosis,the relationships between gender and disease,age and disease,ultrasound diagnosis and disease classification,gender,number of nodules and benign and malignant were analyzed.Results Among the 13 020 cases,524 cases was excluded.There were 2 291 male cases; abnormalities were detected in 1 679 cases,and the abnormal ratio was 73.29％; there were 10 205 female cases; 7 946 cases were abnormal,and the abnormal ratio was 77.86％.The proportion of abnormal thyroid cases of the total number of female was higher than that of male,and the ratio of male and female was 1.00 ∶ 4.73 (1 679∶7 946); gender differences were statistically significant(x2 =18.476,P ＜ 0.01).There were nodular type 5 018 cases,diffuse type 2 012 cases and mixed type 2 603 cases in sonographic findings.The nodular type on ultrasound images showed mainly nodular goiter(79.21％,3 975/5 018).The diffuse type on ultrasonic images showed mainly Hashimoto Thyroiditis(59.24％,1 192/2 012).The mixed cases on ultrasound images showed mainly nodular goiter (89.83％,2 331/2 595).Ultrasound diagnostic results showed that nodular goiter accounted for 65.52％ (6 306/9 625) and Hashimoto Thyroiditis accounted for 17.97％ (1 730/9 625).Proportion of thyroid disease increased gradually with age,reached a peak [29.93％(2 557/8 544) and 28.84％ (2 464/8 544)] between the ages of 41-50 and 51-60 years old.The proportion gradually decreased into[14.17％(1 211/8 544),7.26％(620/8 544)] between the age of 61-71 and ＞ 71 years old.The incidence differences of malignant nodules between different age groups were statistically significant (x2 =407.796,P ＜ 0.01).Among malignant nodules,solitary nodule accounted for 95.76％ (113/118) ; multiple nodules accounted for 4.24％ (5/118),and there were more solitary nodule than multiple nodules(x2 =15.286,P ＜ 0.01).About malignant solitary nodules,women accounted for 87.61％(99/113); men accounted for 12.39％ (14/113),and women's were significantly higher than man's(x2 =360.960,P ＜ 0.01).Correlation analysis showed that the results of ultrasounddiagnosisofvarious thyroid diseases were highly correlated with the ultrasonic types of diseases(r =0.139 99,P ＜ 0.01).Conclusions Nodular goiter and Hashimoto Thyroiditis are the most common adult neck discomfort diagnosed by ultrasound in Heilongjiang Province.The high-risk age of thyroid disease is between 41-60 years old.Thyroid malignant is more common in single nodular,and the number of female patients is significantly higher than that of men.
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The results of RT-PCR and immunohistochemistry showed that the expressions of vascular endothelial growth factor (VEGF) and its receptor ( flk-1 ) in the renal cortex of insulin-resistant rats during the phase of normal blood glucose were significantly increased, which were decreased by telmisartan. The result suggests that telmisartan may ease kidney damage via decreasing VEGF and flk-1 expressions.
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Objective To investigate mupiroein resistance in Staphylococcus aureus (SAU) and the resistance to commonly used antibiotics in mupirocin-resistant strains. Methods Four hundred and ninety clinically isolated SAU strains froin January 2005 to May 2007 in the First Affiliated Hospital,Wenzhou Medical College were screened by mupirocin(5μg)disc diffusion method.Minimum inhibition concentration(MIC)and the amplification of mupA gene were performed to determine the resistance to mupirocin.Resistance to cefoxitin,gentamycin, levofloxacin, trimethoprim/sulfamethoxazole, rifampin, erythromycin, clindamycin, tetracycline and vancomycin in mupirocin-resistant strains was detected by disc diffusion method, and the amplification of mecA gene was performed to confirm the methieillin resistance among mupiroein-resistant strains.Results Twenty-seven mupirocin-resistant strains were obtained,in which 22(81.5%)were hish-level mupirocin resistant(MuH)and the rest were low-level mupirocin resistant(MuL).Among 27 mupirocin-resistant strains,24 were methicillin-resistant Staphylococcus aureus (MRSA)in which 21 were MuH and 3 were MuL strains.Drug sensitivity tests showed that the resistance to gentamycin,levofloxacin,trimethoprim/sulfamethoxazole,rifampin,erythromycin,elindamycin and tetracycline were hish among MuH and MuL strains,and most of these strains were multi-drug resistant.All strains were susceptible to vaneomycin.Conclusions Most of the clinical emerged mupirocin-resistant SAU strains are MuH and show hish resistance to commonly used antibiotics.Therefore,detection and drug sensitivity test of mupirocin-resistant strains should be strengthened in clinic practice in order to prevent it from dissemination.
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OBJECTIVE To review and analyze the change in the MICs of vancomycin,teicoplanin and linezolid in meticillin-resistant Staphylococcus aureus(MRSA) strains isolated in our hospital from 2003 to 2007. METHODS The MICs of vancomycin,teicoplanin and linezolid were tested by Etest method on a sample of randomly selected MRSA strains. RESULTS The incidences of MRSA increased from 52.2% in 2003 to 74.5% in 2007.MIC of vancomycin increased from 1.85 ?g/ml in 2003 to 2.15 ?g/ml in 2007,and teicoplanin MIC geometric mean increased even more markedly from 1.28 ?g/ml in 2003 to 2.07 ?g/ml in 2007.The linezolid MIC remained almost unchanged. CONCLUSIONS The incidences of MRSA were increasing from 2003 to 2007.There is a upward trend in MIC of glycopeptide over the years,in which the increase for teicoplanin is higher than others two.
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Objective: To study the relationship between inteiligence and ADHD (attention deficiency and hyperactivity disorder) . Method: 129 children with ADHD and 87 normal control were evaluated with C- WISC. Results: The IQ of some children with ADHD was in borderline range, their performance of balance between VIQ and PIQ was poor. While the controls had normal IQ. Conclusion: The IQ of children with ADHD is lower than that of normal children, especially the balance between VIQ and PIQ.
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Objective:To evaluate levels of intelligence in children with ADHD. Methods:A total of 129 children with ADHD and 87 normal children were evaluated with C-WISC. Results:Full-scale IQs of most children with ADHD varied between the normal and borderline range, showing incompatible scores on VIQ and PIQ. The overall levels of intelligence of children with ADHD were found be lower than normal controls. Conclusion:There was significant difference in intellectual abilities between ADHD children and normal children.