Background: Hemophilia is the most common clotting disorder in the hereditary blood clotting disorders causing harm to health and psychology. The disease can lead to disability and leave the burden on families and society as well as the development of race\r\n', u"Objectives: To study the familial characteristics of haemophiliacs treated at Regional Hematology and Blood Transfusion Center of Hue Central Hospital. Subject and method: This was a prospective study. It included 48 patients diagnosed and treated Haemophilia A and B at Regional Hematology and Blood Transfusion Center of Hue Central Hospital from 7/2005 - 8/2007. Results: In 48 patients, there were 23 patients who had obviously familial history (included 12 families). They were siblings, cousins, maternal grandfathers or mother's brothers. Among 67 haemophiliacs, 23 haemophiliacs had been studied (34.32%), 30 haemophiliacs died of the disease (44.77%). Most of them died at childhood, below age of 15 years (80.64%). Conclusion: Numbers of deaths in the family was not related to the severity of the disease. The age of clinical detection, morphology, number, site, characteristics of haemorrhage as well as the level of articuar injures were not completely the same between the haemophiliacs of the same family. \r\n", u'\r\n', u'
Hemophilia A/ history ; pathology

We report a case of hemophilic pseudotumor in the ulna of a 6-year-old boy treated with radiation therapy. A total dose of 900 cGy in 6 fractions was given in 6 consecutive days. Progression of cystic changes was halted within a month. New bone formation and trabeculation were found on the 4th month. Complete healing of the lesion and bony replacement were found on the 12th month. The patient was followed up to 72 months and there was no evidence of recurrence and no bone growth disturbance. Radiation therapy can be an effective alternative modality in treating hemophilic pseudotumor.
Bone Cysts/radiotherapy* ; Bone Cysts/pathology ; Bone Cysts/etiology* ; Case Report ; Child ; Hemophilia A/pathology ; Hemophilia A/complications* ; Human ; Male ; Ulna/pathology*

OBJECTIVE: To identify mutations within the factor VIII gene in Korean patients with severe hemophilia A. DESIGN: A laboratory analysis. METHODS: We systematically sequenced the promoter, all exons and splice junctions of factor VIII gene in 23 unrelated Korean patients with severe hemophilia A. Patients with factor VIII gene inversion were excluded. RESULTS: Twelve patients (52.2%) showed a point mutation, among which 6 were nonsense mutations and the other 6 were missense mutations. A large deletion was found in 6 (26.1%) patients, a small deletion in 2 (8.7%), a small insertion in one patient. Two patients had compound mutations: one patient had two missense mutations, and the other had a missense mutation and 4 bp insertion. Ten (43.5%) out of 23 mutations found are novel. CONCLUSION: Mutations within the factor VIII gene found in Korean patients with moderate to severe hemophilia A are diverse as expected. And we have found 10 novel mutations. Our results can help understanding the molecular pathology of hemophilia A.
Codon, Nonsense ; Exons ; Factor VIII* ; Hemophilia A* ; Humans ; Mutation, Missense ; Pathology, Molecular ; Point Mutation

OBJECTIVETo investigate the molecular mechanism of haemophilia B caused by the novel mutation of Arg327Ile (R327I) in FIX gene.

METHODSThe R327I, R327Ala(A), R327Lys(K), R327Asn(N) and a replacement mutant (FIXβFVII), in which FIX β strand 324-329 was replaced by that of FVII 298-303, expression plasmids were constructed with site-directed mutagenesis method based on the wild-type (WT) FIX expression plasmid. The HEK293 cell was transiently transfected, then the activity of FIX (FIX:C) was assayed by one stage method in the conditioned medium, while the FIX:Ag in both the conditioned media and the cell lysates was measured by ELISA. The molecular weight and the semi-quantity of expressed FIX were analyzed by Western blot. Fluorescent protein expression plasmid was constructed to investigate the synthesis and secretion of the FIX R327I mutation in the viable cells.

RESULTSFIX:C of the R327I mutant protein was 4.49% of the level of the WT in the conditioned medium, and the FIX:Ag of the R327I mutant protein in the conditioned medium and the cell lysates was 31.02% and 129.29% compared to that of WT, respectively. The mutation was characterized as cross-reaction material reduced (CRMR). The viable cell fluorescent assays showed that the R327I protein was more in both the viable cells and in lysosome than that of WT. The FIX:C of the R327A, R327K, R327N and FIXβFVII mutants was reduced compared to that of WT, the reduction of FIX:C of FIXβFVII was the most significantly amount among all the mutants in medium. FIX:Ag of all the mutants in the medium, except that the R327K increased, was reduced. The result of Western blot showed that the molecular weight of R327I protein was the same as that of WT, but the amount of the protein was much less compared with WT in the conditioned medium.

CONCLUSIONThe abnormal synthesis and secretion as well as the abnormal function of the R327I mutant protein causes haemophilia B. The residue of R327 as well as the β strand domain of R327 located play important roles of the specific function of FIX.

Factor IX ; genetics ; HEK293 Cells ; Hemophilia B ; genetics ; pathology ; Humans ; Mutagenesis, Site-Directed ; Mutation ; Transfection

To explore the diagnoses and treatment of hemophilic pseudotumor, one case with hemophilic pseudotumor misdiagnosed and treated by operation, was observed and analyzed. The result showed that the final diagnosis of this case was following: hemophilia A (mild type) and hemophilic pseudotumor with injury of femoral nerve. The final diagnosis was given from inquiring case history and family history additionally, and drawing assistance from laboratory examination and computed tomography. After operation, the patient's wound healed very well through supplying coagulation factors positively. In conclusion, it was important for inquiring case history and family history particularly and thinking highly of laboratory examination to reduce the misdiagnosis and error of therapy for this case. If paying attention to preoperative preparation, the danger of hemorrhage during operation can be reduced and wound after operation can heal more rapidly.
Adult ; Diagnostic Errors ; Femoral Nerve ; pathology ; Hematoma ; diagnosis ; etiology ; Hemophilia A ; complications ; Humans ; Male

OBJECTIVE To generate hemophilia A (HA) patient-specific inducible pluripotent stem cells (iPSCs) and induce endothelial differentiation. METHODS Tubular epithelial cells were isolated and cultured from the urine of HA patients. The iPSCs were generated by forced expression of Yamanaka factors (Oct4, Sox2, c-Myc and Klf4) using retroviruses and characterized by cell morphology, pluripotent marker staining and in vivo differentiation through teratoma formation. Induced endothelial differentiation of the iPSCs was achieved with the OP9 cell co-culture method. RESULTS Patient-specific iPSCs were generated from urine cells of the HA patients, which could be identified by cell morphology, pluripotent stem cell surface marker staining and in vivo differentiation of three germ layers. The teratoma experiment has confirmed that such cells could differentiate into endothelial cells expressing the endothelial-specific markers CD144, CD31 and vWF. CONCLUSION HA patient-specific iPSCs could be generated from urine cells and can differentiate into endothelial cells. This has provided a new HA disease modeling approach and may serve as an applicable autologous cell source for gene correction and cell therapy studies for HA.
Cell Differentiation ; Hemophilia A ; pathology ; therapy ; urine ; Humans ; Induced Pluripotent Stem Cells ; cytology ; transplantation ; Urine ; cytology

Aged, 80 and over ; Dyspnea ; etiology ; Glottis ; pathology ; Hematoma ; complications ; Hemophilia A ; complications ; Humans ; Laryngeal Diseases ; complications ; Male

Adolescent ; Bone Neoplasms ; diagnosis ; diagnostic imaging ; etiology ; Hemophilia A ; complications ; diagnostic imaging ; pathology ; Humans ; Male ; Tomography, X-Ray Computed

The objective of this study was to explore the feasibility of intestinal epithelial cell and human source vector used in gene therapy for hemophilia B. The intestinal epithelial sw480 cells were transfected with human source vector plasmid pHrnF9 which contained human coagulation factor IX gene. Transcription of its mRNA were measured by RT-PCR. The transfection efficiency were observed under fluorescence microscope. The expression of its protein and coagulant activities in the transfected sw480 cells were measured by ELISA and one-stage method. The results showed that the expression of hFIX mRNA could be detected after transfection. The transfection efficiency reached to the maximum at 48 hour. The hFIX protein amount was 11.3 +/- 0.23 ng/(10(6) cells.24 h) at 24 hours after transfection and reached to 29.34 +/- 1.00 ng/(10(6) cells.24 h) at 48 hours and decreased to 12.45 +/- 0.15 ng/(10(6) cells.24 hr) at 72 hours. Sw480 cells transfected with pHrnF9 were capable of producing hFIX with coagulant activity. The coagulant activity reached to (6.07 +/- 0.17)%/10(6) cells at 48 hours and decreased to 1.81 +/- 0.06%/10(6) cells at 72 hours. It is concluded that the sw480 cells transfected with pHrnF9 plasmid can express hFIX with coagulant activity, the intestinal epithelial cells may become target cells in the gene therapy for hemophilia B.
Cell Line, Tumor ; Epithelial Cells ; metabolism ; Factor IX ; genetics ; metabolism ; Genetic Vectors ; genetics ; Hemophilia B ; therapy ; Humans ; Intestines ; pathology ; RNA, Messenger ; genetics ; metabolism ; Transfection

OBJECTIVE: To explore the effect of lenalidomide on human fibroblast-like synovial cells (HFLS) and the therapeutic efficacy on hemophilic arthropathy in hemophilia A mice model. METHODS: In vitro, to remodel the inflammatory environment of synovial tissue after hemorrhage, ferric citrate and recombinant TNF-α were added into the cell culture medium of HFLS. Cell Counting Kit-8 (CCK-8), Enzyme-linked immunosorbent assay (ELISA), Quantitative Real-time PCR (RT-qPCR) and flow cytometry were employed for detection of the effects of lenalidomide on the proliferation ability, pro-inflammatory cytokines release and apoptosis of HFLS cells. In vivo, hemophilia arthropathy was remodeled in hemophilia A mice by induction of hemarthrosis. A series of doses of lenalidomide (0.1, 0.3 and 1.0 g/kg) was administrated intra-articularly. Tissues of knee joints were collected on the 14th day after administration, and the protective effect of lenalidomide on arthritis in hemophilia A mice were evaluated by RT-qPCR and histological grading. RESULTS: In vitro, compared with the untreated control group, lenalidomide could significantly inhibit the proliferation of HFLS cells (P<0.05), and the effect was the most significant when the concentration was 0.01 μmol/L (P<0.001). Compared with the control group, lenalidomide could significantly inhibit the expression levels of TNF-α, IL-1β, IL-6 and IFN-γ in HFLS cells (P<0.05). The flow cytometry results showed that lenalidomide could enhance the apoptotis of HFLS cells (P<0.05). The results of RT-qPCR showed that lenalidomide could significantly reduce the mRNA expression levels of TNF-α, IL-1β, IL-6,MCP-1 and VEGF in the joint tissues (P<0.05). Histological results showed that compared with the injured group, lenalidomide could significantly reduce the pathological sequela after hemarthrosis induction, e.g. synovial thickening and neo-angiogenesis in the synovium. The protection displayed a dose-response pattern roughly. CONCLUSION In vitro, lenalidomide can inhibit the proliferation of HFLS cells, promote their apoptosis, and inhibit the expression of pro-inflammatory cytokines. In vivo, lenalidomide can significantly decrease the expression of pro-inflammatory cytokines in the joints of mice, and prevent the development of inflammation and neo-angiogenesis. The results provide a theoretical and experimental basis for the clinical application of lenalidomide in the treatment of hemophilic arthropathy.
Animals ; Arthritis ; Cytokines/metabolism* ; Hemarthrosis/pathology* ; Hemophilia A/genetics* ; Humans ; Interleukin-6 ; Lenalidomide ; Mice ; Neovascularization, Pathologic ; RNA, Messenger ; Sincalide ; Tumor Necrosis Factor-alpha ; Vascular Endothelial Growth Factor A