OBJECTIVE: To develop monoclonal antibodies that can specifically recognize human von Willebrand factor (VWF) propeptide (VWFpp) in plasma, and establish a rapid and reliable method for the detection of VWFpp antigen in plasma by using the double-antibody sandwich ELISA with the obtained anti-VWFpp monoclonal antibody. METHODS: The recombinant human VWFpp (D1 and D2 regions) protein expressed in eukaryotic cells was used as immunogen to immunize BALB/c mice with routine method, so as to obtain clones of fusion cells. After screening and identification, hybridoma cell lines secreting monoclonal antibodies against VWFpp were selected, and then double-antibody sandwich ELISA assay was used to construct VWFpp antigen detection kit for the determination of VWFpp in human plasma. The levels of VWFpp antigen in plasma of 12 leukemia patients who underwent bone marrow transplantation were dynamically detected. RESULTS: Two hybridoma cell lines that can be subcultured continuously and secrete monoclonal antibodies against VWFpp were obtained and named SZ175 and SZ176 respectively. Identified by ELISA and Western blot, the antibodies could both specifically recognize VWFpp but couldn't recognize mature VWF (without propeptide). Based on the principle of double-antibody sandwich ELISA, monoclonal antibodies SZ175 and SZ176 were successfully made into a kit for detecting VWFpp antigen. The plasma VWFpp levels of leukemia patients before and after bone marrow transplantation were dynamically detected. The results showed that the plasma VWFpp levels of the patients after transplantation were significantly higher than those before transplantation. CONCLUSION Two monoclonal antibodies against VWFpp were successfully prepared, and a double-antibody sandwich ELISA detection kit for VWFpp antigen was constructed, which provides a powerful tool for further study on the biological function of VWFpp, the clinical diagnosis and classification of von Willebrand disease (VWD), and the prognostic monitoring of endothelial injury-related diseases.
Animals ; Mice ; Humans ; von Willebrand Factor ; Antibodies, Monoclonal ; Protein Precursors/metabolism* ; von Willebrand Diseases/diagnosis* ; Prognosis

BACKGROUND: Definition of possible von Willebrand disease (VWD) included levels of von Willebrand factor antigen (VWF:Ag) or von Willebrand factor ristocetin cofactor activity (VWF:RCo) below 30 U/dL. The purpose of this study was to determine the prevalence of possible VWD in Ulsan, Korea. We also analyzed the influence of demographic factors such as age, gender, and blood group on subject levels of VWF.METHODS: Between March 2011 and September 2011 we prospectively investigated 1,039 subjects (271 children, 768 adults). Blood samples were collected for the determination of VWF:Ag, VWF:RCo, and factor VIII coagulation assay (FVIII:C). A standardized questionnaire was administered to evaluate hemorrhagic symptoms at the time of first examination, using a bleeding score ranging from 0 to 3.RESULTS: Forty five subjects met the criteria for possible VWD, for a prevalence of 4.3%. Subjects of Group O had a significantly lower mean FVIII:C, VWF:Ag, and VWF:RCo value than subjects of group A,B, or AB (P<0.001).CONCLUSION: Our results suggest that the prevalence of VWD may be much higher than previously reported. Efforts to increase the awareness and diagnosis of VWD may help improve identification of patients with bleeding disorders and lead to early, appropriate management with safe and efficacious therapies.
Child ; Demography ; Diagnosis ; Factor VIII ; Hemorrhage ; Humans ; Korea ; Prevalence ; Prospective Studies ; Ulsan ; von Willebrand Diseases ; von Willebrand Factor ; Surveys and Questionnaires

Adolescent ; Diagnostic Errors ; Hemophilia A ; diagnosis ; Humans ; Male ; von Willebrand Diseases ; diagnosis

Bleeding Time ; Child, Preschool ; Female ; Hemorrhage ; diagnosis ; Humans ; von Willebrand Diseases ; diagnosis

OBJECTIVETo analyze phenotype and genotype of three Chinese pedigrees with von Willebrand disease (vWD), and explore the molecular mechanism.

METHODSBleeding time (BT), activated partial thromboplastin time (APTT), ristocetin-induced platelet aggregation (RIPA), von Willebrand factor (vWF): ristocetin cofactor (RCof) (vWF:RCof), vWF antigen (vWF:Ag), vWF activity (vWF:A) test, vWF collagen binding assay (vWF:CB), vWF and Factor VIII (FVIII) binding assay (vWF:FVIII:B) and multimer analysis were used for phenotype diagnosis. Genomic DNA was extracted from the peripheral blood (PB). All the 52 exons and flanking sequences of the probands' vWF gene were amplified by PCR and analyzed by direct sequencing.

RESULTSAPTT were prolonged in all three probands, while BT were normal excepting for proband 3. Plasma RIPA, vWF:RCo, vWF:Ag, vWF:A and vWF:CB were decreased in different extents. In multimer analysis, proband 3 lost the large and intermediate molecular weight multimers, while proband 1 and 2 were normal. Gene analysis in the three probands revealed three heterozygous missense mutations of 144067 G→A (R2287Q) in exon 39, 110374G→A (R1374H) and 110770C→T (S1506L) in exon 28 and heterozygous polymorphism 110667G→A (D1472H) in exon 28, respectively.

CONCLUSIONThe three heterozygous mutations (R2287Q, R1374H and S1506L) and an heterozygous polymorphism (D1472H) are genetic defects of the hereditary vWD of the three pedigrees respectively. R2287Q is a novel mutation reported for the first time in the literature.

Adult ; Child ; DNA Mutational Analysis ; Female ; Genotype ; Heterozygote ; Humans ; Male ; Pedigree ; Phenotype ; von Willebrand Diseases ; diagnosis ; genetics ; von Willebrand Factor ; genetics ; metabolism

OBJECTIVETo analyse the epidemiological feature, clinical characters and therapeutic regimens for von Willebrand disease(VWD).

METHODSThe clinical data and laboratory tests results of 162 VWD patients in our center were analyzed.

RESULTSThere were 76 males and 86 female among these patients with the mean age of 7.2(2.0-41.0) and 20.7(5.0-48.0) years, respectively. 86 patients (53.1%) were identified to be type 1 VWD, 34 patients (21.0%) type 3 VWD and 42 patients (25.9%) type 2 VWD. Among type 2 VWD patients, 33 patients were type 2A, 4 patients type 2M, 5 patients type 2B. Eighty-seven patients (53.7%) had a definite family history of bleeding tendency. The most common and specific bleeding symptoms were easy bruising (61.7%), epistaxis (53.7%), prolonged bleeding after surgery or minor injury (53.1%). Menorrhagia (66.3%) was common in female patients. The analysis of Vicenza bleeding scores in all patients showed that only 56(34.6%) patients had abnormal bleeding scores. FVIII/VWF concentrates and cryoprecipitate were applied to 45 patients (27.8% ), Desmopressin (DDAVP) to 8 patients. Eight female patients need oral contraceptives jointly to control menorrhagia. Hysterectomy had to be performed in 2 female patients with VWD.

CONCLUSIONVWD was a common congenital bleeding disorder with heterogeneous characters, it was necessary to screen, identify, classify accurately this disease in order to supply to effectively individualized treatment.

Adolescent ; Adult ; Child ; Child, Preschool ; Female ; Humans ; Male ; Middle Aged ; Retrospective Studies ; Young Adult ; von Willebrand Diseases ; diagnosis ; therapy ; von Willebrand Factor

PURPOSE: von Willebrand disease is a common inherited bleeding disorder characterized by high degree of variable clinical presentation due to either quantitative or qualitative defects in von Willebrand factor. Its incidence in Korea is not well studied while that in western countries is extensively studied. METHODS: We classified 16 cases of vWD from 14 unrelated families based on vWF antigen, ristocetin cofactor activity, factor VIII activity and vWF multimeric patterns analysed by agarose gel electrophoresis, according to a revised classification by ISTH. RESULTS: There were 12 cases (75%) of type 1 vWD or 2M/2N with normal multimeric pattern, 3 cases (18.75%) of type 2 vWD lacking high molecular weight multimers and only 1 case of type 3 vWD with no multimers. CONCLUSION: The proportion of each vWD subtype in Korea is similar to that in western countries, however, accurate diagnosis based on ristocetin induced platelet aggregation test, factor VIII binding assay and molecular genetic diagnosis seems to be necessary for a more complete classification of vWD.
Classification ; Diagnosis ; Electrophoresis, Agar Gel ; Factor VIII ; Hemorrhage ; Humans ; Incidence ; Korea* ; Molecular Biology ; Molecular Weight ; Platelet Aggregation ; Ristocetin ; von Willebrand Disease, Type 3 ; von Willebrand Diseases* ; von Willebrand Factor

BACKGROUND: von Willebrand disease (VWD) is one of the most common inherited bleeding disorders. However, the number of patients who register to the Korea Hemophilia Foundation (KHF) is much lower than the expected prevalence rate and only few hospitals perform tests for diagnosis autonomously. Thus, we surveyed practical realities of VWD in Yeungnam region. METHODS: Patients with VWD (N=267) who were diagnosed at eleven university hospitals from March 1995 to March 2018 were enrolled in this study. We evaluated the medical records from each hospital retrospectively. RESULTS: Two hundred and twenty-eight children and 39 adults met the diagnostic criteria for VWD. Seventy-eight (57.4%) patients had the blood type O. Fifty-eight patients were definite type 1 (21.7%), 151 were possible type 1 (56.6%), and the others were type 2. Abnormal laboratory findings were the most common factor for the diagnosis in children. VWF mutations were detected in 17 patients. Patients with a family history showed age of diagnosis of 9 y, which is higher than in those with no family history (6 yr), and also showed a higher rate of significant bleeding (32.1% vs. 14.2%). VWF:RCo and VWF:Ag tests were performed in-hospital at only 1 of 11 hospitals. Twelve of 267 patients were enrolled at the KHF (4.5%). CONCLUSION: A high rate of out-sourcing studies may result in inaccurate diagnosis. The registration rate to the KHF is still lower than the prevalence rate. A comprehensive nationwide registration system is necessary in order to identify the actual prevalence rate and promote the diagnosis of VWD in Korea.
Adult ; Child ; Diagnosis ; Hemophilia A ; Hemorrhage ; Hospitals, University ; Humans ; Korea ; Medical Records ; Prevalence ; Retrospective Studies ; von Willebrand Diseases

BACKGROUND: von Willebrand disease (VWD) is the most common inherited bleeding disorder with a prevalence of up to 1%. However in Korea, only 126 VWD patients were registered in Korea Hemophilia Foundation (KHF). The aim of this study was to determine the status of VWD patients in Korea. We analyzed VWD patients by age, gender, blood group, family history and bleeding history.METHODS: One hundred twenty-six VWD patients registered in the KHF by December 2016, and 74 patients diagnosed at six university hospitals were enrolled in this study. We evaluated the medical records from the KHF and the questionnaires from six university hospitals retrospectively.RESULTS: Seventeen patients misdiagnosed and ten patients duplicated were excluded. One hundred nine patients registered in the KHF and 64 patients diagnosed at six university hospitals met the criteria for VWD. The blood type O accounts for 72 (51.8%). VWF mutation was detected in 30 patients (17.3%). Median age at diagnosis was 10.5 yr. The bleeding score of adults was higher than that of children (P < 0.001). The most common bleeding symptom was epistaxis (48.5%). The distribution of VWD types was: 67% of type 1, 30.1% of type 2, and 2.9% of type 3.CONCLUSION: Even though only six hospitals responded to the survey, 64 patients not registered in the KHF were diagnosed with VWD. Our results suggest the prevalence of Korean VWD might be higher than previously reported. A nationwide registration system is warranted in order to accurately identify the national prevalence of VWD.
Adult ; Child ; Diagnosis ; Epistaxis ; Hemophilia A ; Hemorrhage ; Hospitals, University ; Humans ; Korea ; Medical Records ; Prevalence ; Retrospective Studies ; von Willebrand Diseases

A 68-year-old woman with known severe aortic stenosis was admitted to the hospital because of hematochezia and dizziness. She had received several blood transfusions over the preceding 3 years and undergone right hemicolectomy 2 years ago for severe lower gastrointestinal bleeding. Postoperative histology revealed angiodysplasia involving the ascending colon. After the hemicolectomy, she continued to have hematochezia and anemia and required additional blood transfusions for anemia. During this admission, platelet count, activated partial-thromboplastin time, von Willebrand factor antigen, and von Willebrand factor ristocetin cofactor were normal. She had a severe deficiency of high-molecular-weight multimers of von Willebrand factor. Colonoscopy showed angiodysplasia in the transverse colon at this time. Successful coagulation of the bleeding angiodysplasia was achieved by argon plasma coagulator. No additional bleeding was observed thereafter. We report a case of Heyde's syndrome with abnormal von Willebrand factor in a patient who presented with intestinal angiodysplasia and aortic stenosis.
Aged ; Anemia/etiology/therapy ; Angiodysplasia/*complications ; Aortic Valve Stenosis/*complications ; Colonic Diseases/*complications ; English Abstract ; Female ; Gastrointestinal Hemorrhage/*etiology ; Humans ; Syndrome ; von Willebrand Disease/*complications/diagnosis