Guideline for diagnosis and management of congenital dysfibrinogenemia.

INTRODUCTION: Congenital dysfibrinogenemia (CD) is characterized by dysfunction induced by an abnormal fibrinogen molecule structure that results in blood coagulation dysfunction. The clinical manifestations of CD patients are asymptomatic, bleeding and thrombosis. The majority of patient are asymptomatic. However, the single fibrinogen detection method is easy to cause missed diagnosis or misdiagnosis of CD patients. The treatment strategies of CD patients with different clinical manifestations are also different. METHODS: Combing the existing experimental diagnosis technology, literature and our research results, a simple and practical CD diagnostic criteria was proposed. And based on the relevant literature and existing treatment guidelines, more comprehensive treatment recommendations are summarized. RESULTS: In this new criteria, combination Clauss method and PT derived method was proposed to detect fibrinogen and its ratio was used to diagnose for CD. Diagnosis also needs to be combined the clinical manifestations, family investigation and genetic testing. According to different clinical manifestation (bleeding, thrombosis or asymptomatic), treatment methods and strategies are different. The treatment of CD patients should consider the patient's personal and family history of bleeding or thrombosis. Treatment of thrombosis and pregnancy may be more challenging. The risk of bleeding and thrombosis should be evaluated and balanced at all times during clinical treatment. These detailed treatment recommendations can provide reference for patients with different clinical manifestations of CD. CONCLUSIONS: The new CD diagnosis criteria and comprehensive treatment recommendations can effectively improve the diagnosis and treatment of CD.

Genetic mutation analysis of hereditary spherocytosis in Guangxi Zhuang Autonomous Region.

Hereditary spherocytosis (HS) is a common, hereditary hemolytic anemia (HHA) that is attributed to the disturbance of five erythrocyte membrane proteins. HS is also common in Guangxi, China. Target region capture high-throughput sequencing technology was used to analyze genetic mutations found in HS patients. Pedigree analysis was also performed, in some cases, to provide an optimized approach for the etiological diagnosis of complex, hereditary hemolytic anemia. Blood samples from the probands and their families were assessed by laboratory tests, target region capture high-throughput sequencing technology, and Sanger sequencing. We detected 79 HS patients from 37 unrelated families. The mutations observed in these patients were found mainly in four HS-related genes. These included SLC4A1, which was mutated in 31.65% of patients (25/79), SPTA1 (30.78% (24/79)), EPB42 (6.33% (5/79)), and SPTB (5.06% (4/79)). Composite genotype was observed in 26.58% (21/79) of patients and included mutations in two or more HS-related genes or mutations in HS-related genes combined with thalassemia or G6PD deficiency. No significant differences in clinical symptoms were found among patients of various genotypes except total bilirubin. Mean reticulocyte volume (MRV) and mean sphered cell volume (MSCV) of the composite genotype were significantly different from other groups. A total of 28 mutation types were found in HS-related genes. Using high-throughput sequencing technology, we also found some cases that had been misdiagnosed. MRV and MSCV are more significant in compound mutations as sensitive determinants of HS. High-throughput sequencing technology can be used to provide a more effective etiological diagnostic method for HS, with high efficiency and specificity.

Misdiagnosis of a patient with congenital dysfibrinogenemia: A case report and literature review.

BACKGROUND: We reported a patient with congenital dysfibrinogenemia who was misdiagnosed and reviewed relevant literature, in order to discuss the methods to reduce misdiagnosis. METHODS: A 23-year-old pregnant woman was found to be with low fibrinogen in antenatal examination at another province teaching hospital, who was misdiagnosed to have hypofibrinogenemia. Fibrinogen infusion or cryoprecipitation was recommended if necessary. The patient came to our hospital for further diagnosis and treatment considering the safety of herself and the fetus. We examined the coagulation function and gene sequencing of the pregnant woman and her family members. RESULTS: Fibrinogen (Clauss method) was significantly reduced in the patient and her mother, while the level of fibrinogen (PT-derived method) was normal. Thrombin time was prolonged. Heterozygous mutation site was found in exon 2 of the FGA gene, c.104G > A(p.Arg35His). CONCLUSION: When the fibrinogen (Clauss method) is significantly reduced and the thrombin time is prolonged, PT-derived method and the investigation of family coagulation function should be added, which can be used to diagnose and distinguish congenital dysfibrinogenemia from hypofibrinogenemia.

Congenital dysfibrinogenemia in major surgery: A description of four cases and review of the literature.

BACKGROUND: Congenital dysfibrinogenemia is characterized by qualitatively abnormal fibrinogens with resultant blood coagulation dysfunction. The clinical manifestations are high heterogeneity. Treatment for dysfibrinogenemia should be personalized. Here, we reported four congenital dysfibrinogenemia patients with the major surgery, in order to discuss the treatment and diagnosis of congenital dysfibrinogenemia. METHODS: We reported four asymptomatic congenital dysfibrinogenemia patients with the major surgery (valve replacement, brain surgery, tumorectomy, hysterectomy) in our study. Routine coagulation tests, hepatorenal function and gene analysis, thrombelastogram were performed. RESULTS: Four congenital dysfibrinogenemia patients all showed prolonged TT, low level of activity fibrinogen and normal fibrinogen antigen. Case1 showed a heterozygous mutation in exon 2 of the FGA, c.1223G > C, which turns the codon for residue Aα Gly13 into Arg (p. Gly13Arg). DNA sequencing of case2 showed that a heterozygous mutation in exon 8 of the FGG (c.5877G > A) with being responsible for the Arg â†’ His substitution at position 301 of the γ chain (p. Arg301His). Case3 and case 4 failed to do genetic testing for other reason. Four congenital dysfibrinogenemia patients were asymptomatic in the daily life. Personal and family history revealed no abnormal bleeding or thrombotic events. These four patients did not receive special treatment and management before surgery. They all had a smooth operation. CONCLUSIONS: Misdiagnosis and unnecessary infusion bring huge health risks to patients. Correct diagnosis of congenital dysfibrinogenemia is the key to avoid misdiagnosis or unnecessary infusion. Asymptomatic patients with congenital dysfibrinogenemia do not need cryoprecipitate or fibrinogen input before major surgery.

A Multi-Agent Platform to Explore Strategies for Age-Friendly Community Projects in Urban China.

BACKGROUND AND OBJECTIVES: Although numerous efforts have been made to promote age-friendly communities (AFCs) in urban China, challenges such as the engagement and management of stakeholders, budget constraints, and policy issues remain. This article describes the work of designing a multi-agent platform (MAP) for the briefing stage of AFC projects. RESEARCH DESIGN AND METHODS: The process to design the MAP is first described, and the components and variables are identified. Then, a case study of a stakeholder consensus formation process is conducted using an agent-based simulation. Next, according to the simulation results, strategies to handle the conflicts arising among the stakeholders of AFC projects are proposed. RESULTS: According to the agent-based simulation conducted, both the initial approval rate and the outside connection rate will affect the stakeholder consensus formation process. Although a higher initial approval rate and a lower outside connection rate may reduce the average convergence time, the results show that 3-5 rounds of information exchange are still needed before a consensus or dissent is formed. DISCUSSION AND IMPLICATIONS: Investors are suggested to communicate with residents and alleviate their concerns regarding AFC projects to facilitate the consensus formation process during the briefing stage of AFC projects; they can also organize activities for residents to exchange information and ideas. The simulation conducted, together with the MAP built in this research, will serve as a reference to help researchers and practitioners further understand the briefing stage and explore efficient strategies for the successful implementation of AFC projects in urban China.

The ß-chain mutation p.Trp433Stop impairs fibrinogen secretion: A novel nonsense mutation associated with hypofibrinogenemia.

BACKGROUND: Congenital hypofibrinogenemia is characterized by proportional decreases in fibrinogen activity and immunoreactive fibrinogen levels. Here, we describe a new case with the bleeding risk identified in our hospital. METHODS: The proband was cut and bled for 3 h. Coagulation testing, gene analysis, thrombelastogram, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), in vitro plasmid construction, and functional analyses were performed to explore the pathogenic mechanism. RESULTS: Coagulation testing of the male proband revealed low levels of fibrinogen detected by two methods (the Clauss method and the PT-derived method); his two sons had normal coagulation results. DNA sequencing of the proband revealed a heterozygous point mutation in exon 8 of the FGB gene causing Trp→Stop substitution and a polymorphic site (p.Leu92Phe). Human Trp433 was found to be highly conserved. SDS-PAGE showed that the fibrinogen level of the proband was markedly lower than that of healthy controls. Using high-performance liquid chromatography-mass spectrometry, a mutated Bß chain was not detected in circulation. In vitro expression analyses indicated that the mutation affected the secretion of fibrinogen. The TEG results indicated that the proband had a prolonged K time, a lower CI value, and a lower angle value. CONCLUSION: We report a new case with a novel nonsense mutation that resulted in hypofibrinogenemia. The results indicate that the nonsense mutation may cause misfolding of the D domain, which then affects the secretion of fibrinogen.

Congenital dysfibrinogenemia caused by γAla327Val mutation: structural abnormality of D region.

BACKGROUND: : Congenital dysfibrinogenemia (CD) is a coagulation disorder caused by mutations in the fibrinogen genes, which result in abnormal fibrinogen function. However, the precise pathogenesis underlying it remains unclear. METHODS: : In this study, we identified a novel heterozygous mutation in an asymptomatic patient with CD caused by γ Ala327Val mutation. Aimed to investigate the pathogenesis, functional studies of fibrinogen isolated from the proband and her family members were performed, such as coagulation function, fibrinogen aggregation test, and fibrin clot lysis test. Coagulation was monitored using a thromboelastometer, and the fibrin clot network structure was observed by scanning electron microscopy. The effect of the mutation on fibrinogen structure and function was predicted by molecular modeling. RESULTS: : The fibrinogen activity concentration in patients with CD was significantly lower than that in healthy individuals, indicating that fibrinogen activity was low. Proband's fibrinogen activity concentration was 0.75 g/L(Clauss method) and antigen concentration (immune turbidimetry method) was 1.59 g/L(normal reference range for both parameters: 2.0-4.0 g/L). Thromboelastography showed that the K value of patients with CD was higher than that of healthy individuals and Angle values were decreased, indicating that mutation impaired fibrinogen function. Compared to fibrinogen from healthy individuals, fiber network structure of the proband was loose, pore size was increased, and fiber branch nodes were increased. CONCLUSIONS: : Ala327Val heterozygous missense mutation leads to changes in the structure of fibrinogen D region and impairs the aggregation function of fibrinogen. This mutation is reported here for the first time.

A New Compound Heterozygous Mutation Of BSCL2 In A Chinese Zhuang Ethnic Family With Congenital Generalized Lipodystrophy.

PURPOSE: This study aims to report the clinical features of an infant with CGL in a Chinese Zhuang ethnic family, whose family members were discovered to carry new pathogenic mutations in the BSCL2. PATIENTS AND METHODS: In this study, we report clinical and molecular investigations of CGL disease in a family of 4 members (parents and two sons). We used whole exome sequencing (WES) in the family to examine the genetic cause of the disease. RESULTS: The proband presented with skin pigmentation, hypertriglyceridemia and diabetes. WES identified a previously unreported compound heterozygous mutation in the BSCL2 (c.545_546insCCG heterozygous mutation and exon 3 heterozygous deletion) in the proband. His mother is a heterozygous carrier of the c.545_546insCCG mutation and his father and brother are carriers of the exon 3 heterozygous deletion. CONCLUSION: Compound heterozygous mutation of the BSCL2 (new c.545_546insCCG heterozygous mutation and new exon 3 heterozygous deletion) was detected in the proband with characteristic clinical manifestations of CGL2.

A novel fibrinogen gamma-chain mutation, p.Cys165Arg, causes disruption of the γ165Cys-Bß227Cys disulfide bond and ultimately leads to hypofibrinogenemia.

BACKGROUND: Congenital hypofibrinogenemia is a type of hereditary disease characterized by impaired fibrinogen synthesis and/or secretion induced by mutations in the fibrinogen gene. OBJECTIVES: We investigated the phenotypes, genotypes, and pathogenesis of congenital hypofibrinogenemia in an affected family. PATIENTS/METHODS: The proband had a risk of bleeding; therefore, conventional coagulation screening was performed for the proband and her family members. Mutation sites in all exons and flanking sequences of FGA, FGB, and FGG were identified, with matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) performed to indicate the expression of abnormal chains. The effect of the mutation sites on fibrinogen structure and function was predicted by molecular modeling, and purified plasma fibrinogen from the proband was analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis and scanning electron microscopy. Thromboelastography was applied to assess the risk of bleeding and clotting in the proband. RESULTS: Fibrinogen levels in the proband were 1.21 g/L, 1.31 g/L, and 1.38 g/L according to Clauss assay, the prothrombin time method, and enzyme-linked immunosorbent assay, respectively. A novel heterozygous mutation (γCys165Arg), a heterozygous mutation (AαIle6Val), and two genetic polymorphisms (AαThr331Ala and BßArg478Lys) in fibrinogen were found in the proband, and MALDI-TOF MS indicated absence of the mutated chain in patient plasma. Additionally, the heterozygous mutation (γCys165Arg) displayed substitution of a nonpolar γ165Cys (low mass) with a positively charged Arg (high mass) along with a small fiber diameter and loose network structure. CONCLUSIONS: Fibrinogen γCys165Arg mutations cause damage to the interchain disulfide bonds of fibrinogen and hinder fibrinogen secretion, possibly explaining the pathological mechanism associated with congenital hypofibrinogenemia.

Abnormal fibrinogen with an Aα 16Arg → Cys substitution is associated with multiple cerebral infarctions.

We found a heterozygous dysfibrinogenemia caused by a substitution of AαArg16Cys. The proband suffered multiple cerebral infarctions. Routine coagulation tests revealed a prolonged thrombin time. The fibrinogen levels in the functional assays were considerably lower than the levels in the immunological assays. The polymerization of the purified fibrinogen was strongly impaired in the presence of calcium. As previously observed in other heterozygous Aα R16C variants, the release rate and amount of fibrinopeptide A (FPA) were lower in the proband than those in normal controls. Additionally, the release of fibrinopeptide B (FpB) was delayed. The immunoblotting analysis using antibodies against human serum albumin indicated that albumin is bound to Aα R16C. The mass spectrometry analysis showed that the Aα R16C fibrinogen chains appeared in the patient's circulation. The clot structure analysis using scanning electron microscopy (SEM) revealed that the fibrin network was dense and consisted of thin and highly branched fibres. Using overlaid fibrinolytic enzymes in a clot lysis experiment, clot degradation was observed to be delayed. These results indicated that the thrombotic tendency may be ascribed to a fibrinolytic resistance caused by an abnormal clot structure with thin fibres and fibrinogen-albumin complexes.

Combined use of Clauss and prothrombin time-derived methods for determining fibrinogen concentrations: Screening for congenital dysfibrinogenemia.

BACKGROUND: In this study, the significance of fibrinogen concentration assessed by a combination of Clauss and prothrombin time (PT)-derived methods for screening for congenital dysfibrinogenemia were investigated, and the screening efficiency of fibrinogen PT-derived/Clauss ratio on congenital dysfibrinogenemia was analyzed. METHODS: We compared fibrinogen concentrations determined by the Clauss, PT-derived, and enzyme-linked immunosorbent assay (ELISA) methods in 73 patients with congenital dysfibrinogenemia and 81 normal controls. Receiver operating characteristic (ROC) curves were utilized to evaluate the efficacy of fibrinogen PT-derived/Clauss ratio in screening for congenital dysfibrinogenemia. RESULTS: Fibrinogen concentrations determined by the Clauss method were dramatically lower than by the PT-derived method and ELISA, and correlated poorly with the latter two methods in patients with congenital dysfibrinogenemia. Fibrinogen concentrations in normal controls were slightly lower according to the Clauss method than to the PT-derived method and ELISA; however, each method yielded results within the normal range and the correlation was good. The area under the ROC curve of fibrinogen PT-derived/Clauss ratio for diagnosis of congenital dysfibrinogenemia was 1 with a standard error of 0, 95% confidence interval of 0.976-1.00, and optimal critical diagnosis point of 1.43. When fibrinogen PT-derived/Clauss ratio was >1.43, the sensitivity and specificity for diagnosis of congenital dysfibrinogenemia were both 100%. CONCLUSIONS: The combined use of Clauss and PT-derived methods for determining fibrinogen concentrations improves the efficiency of screening for congenital dysfibrinogenemia, as the fibrinogen PT-derived/Clauss ratio has high sensitivity and specificity in diagnosis of congenital dysfibrinogenemia. This ratio could serve an important screening tool for this disease.

Three cases of congenital dysfibrinogenemia in unrelated Chinese families: heterozygous missense mutation in fibrinogen alpha chain Argl6His.

Congenital dysfibrinogenemia (CD) is a qualitative fibrinogen disorder caused by an abnormal fibrinogen molecule structure, leading to dysfunctional blood coagulation. This study describes 3 cases of dysfibrinogenemia identified in the unrelated Chinese pedigrees.Routine coagulation screening tests were performed on the probands and their families. The antigens and functionality of fibrinogen was measured using an immunoturbidimetry assay and the Clauss method, respectively. To identify the genetic mutation responsible for these dysfibrinogens, genomic DNA extracted from the blood was analyzed using PCR amplification and direct sequencing. The presence of the mutant chains was determined using matrix-assisted laser desorption/ionization time of flight (MALDI-TOF) mass spectroscopy. Purified plasma fibrinogen of 3 probands was analyzed using SDS-PAGE, fibrinogen clottability, fibrin polymerization, fibrinopeptide release, and scanning electron microscopy (SEM).The 3 probands had a long thrombin time. Levels of functional fibrinogen were found to be very low, while the fibrinogen antigen was within the normal range. DNA sequencing revealed a heterozygous Arg16His substitution in the fibrinogen Aα chain (FGA). The mutant chains were found to be expressed using MALDI-TOF mass spectroscopy. SDS-PAGE did not reveal any difference in the molecular weights of 3 polypeptide chains between normal and abnormal fibrinogens. Fibrinogen clottability showed a slower fibrin clot formation than the healthy control. Fibrin polymerization, after addition of thrombin, showed a prolonged lag phase and decreased final turbidity. The kinetics of fibrinopeptides release revealed a decreased amount of the released fibrinopeptide A. SEM of the patient's fibrin clot was found to be abnormal.Results indicate that the 3 probands with dysfibrinogenemia were caused by mutations of Aα chain Arg16His. Mutation of this fibrinogen induced dysfunction of plasma fibrinogen.

Using concentric circles and wedge grating for camera calibration.

This paper presents a method for camera calibration based on the orthogonal vanishing point calibration using concentric circles grating and wedge grating. This method, which we believe is new, uses the high-precision characteristics of phase extraction to obtain the feature points, thus decreasing the calibration errors caused by the traditional marker extraction errors of gray pattern. According to the simulation experiment analysis results, the concentric circles grating was designed with seven periods and the wedge grating was designed with four periods. In the real measuring experiment, the grating target and the similar gray concentric circles target were used to calibrate the camera, respectively. Through comparing the reprojective errors of the two methods, the method proposed is proven to improve the calibration accuracy and robustness for the vanishing point calibration algorithm.

Analysis and identification of phase error in phase measuring profilometry.

Both the analysis of phase errors which occur at the abrupt discontinuities in phase measuring profilometry (PMP) and the identification method are presented in this paper. The sampling effect of CCD will cause a dilution of accuracy in PMP, especially at abrupt discontinuities on the object surface. The existing methods cannot efficiently identify the abrupt discontinuities. We analyze the relationship between the phase, the height and the equivalent wavelength. By viewing the phase as the argument of a vector we find out that CCD sampling introduces errors into the measurement and the phase is nonlinear to the equivalent wavelength at the abrupt discontinuities. Therefore temporal phase unwrapping (TPU) is introduced into the measurement to identify the abrupt discontinuities. Computer simulations and practical experiment validate the feasibility of this method.

Eliminating the zero spectrum in Fourier transform profilometry using empirical mode decomposition.

Empirical mode decomposition is introduced into Fourier transform profilometry to extract the zero spectrum included in the deformed fringe pattern without the need for capturing two fringe patterns with pi phase difference. The fringe pattern is subsequently demodulated using a standard Fourier transform profilometry algorithm. With this method, the deformed fringe pattern is adaptively decomposed into a finite number of intrinsic mode functions that vary from high frequency to low frequency by means of an algorithm referred to as a sifting process. Then the zero spectrum is separated from the high-frequency components effectively. Experiments validate the feasibility of this method.