Characterization of triglyceride rich lipoproteins with very light density by ultracentrifugation and agarose gel electrophoresis using triglyceride- and cholesterol-staining

Hypertriglyceridemia is an independent risk factor for atherosclerosis. This risk is most likely due to accumulation of circulating triglyceride rich lipoproteins with heterogeneous particles. The identification and characterization of these triglyceride rich lipoproteins is important to detect abnormality of triglyceride metabolism. In the present study, we developed a new method that combines ultracentrifugation and agarose gel electrophoresis with triglyceride- and cholesterol-staining. We investigated 40 subjects with hypertriglyceridemia. Triglyceride rich lipoproteins with very light density were recovered in the aqueous fraction after ultracentrifugation (17,000 x g, 15 min). The lipoproteins recovered in the aqueous fraction contained chylomicrons, if present, their remnants, and light-VLDL (d <1.000 g/ml) containing apoB-100, but not normal VLDL (d <1.006 g/ml) and IDL. Triglyceride rich lipoproteins in the aqueous fraction were characterized by electrophoresis patterns of triglyceride- and cholesterol-staining. Forty patients with hypertriglyceridemia were separated into 8 groups according to their electrophoretic patterns. In lipoproteins recovered in the aqueous fraction from each group, the triglyceride level was correlated with the respective cholesterol level. In summary, a system using ultracentrifugation and agarose gel electrophoresis with triglyceride- and cholesterol-staining is useful for characterization of triglyceride rich lipoproteins and their remnants.     

Association between AMPD1 gene polymorphism and coagulation factors in patients with coronary heart disease

The aim of this study was to investigate whether the C34T and G468T variations in the adenosine monophosphate deaminase-1 (AMPD1) gene were associated with intima-media thickness of the carotid and brachial artery, endothelial function of the brachial artery, glucose metabolism, haemostatic variables and cardiac hypertrophy in patients (n = 109) with coronary heart disease. The plasminogen activator inhibitor-1 activity and the von Willebrand factor were higher in the CC homozygote group compared to the CT/TT group (p < 0.05). There were no differences between the groups regarding intima-media complex of the carotid and brachial artery, presence of plaque in the carotid region, flow-mediated dilatation, ejection fraction or dimensions of the heart. In conclusion, there were no differences between the mutant AMPD1 allele carriers and CC homozygotes regarding surrogate values for atherosclerosis, endothelial function, dimensions and ejection fraction of the heart, glucose tolerance and other well-known cardiovascular risk factors, whereas plasminogen activator inhibitor-1 activity and von Willebrand levels were lower in the mutant AMPD1 allele carriers.     

Removal of hepatitis B virus from a concentrate of coagulation factors II,VII, IX and X by hydrophobic interaction chromatography

Deliberately added hepatitis B surface antigen (HBsAg) and hepatitis B virus (HBV) could be removed by hydrophobic interaction chromatography from a concentrate of coagulation factors II, VII, IX and X. Chromatography in high concentrations of salt, preferably on octanoic acid hydrazide-Sepharose 4B, resulted in a 10⁴–10⁵-fold reduction of HBsAg. The binding properties of HBsAg and HBV to the hydrophobic gels were similar and the Chromatographie method seems particularly useful in removing low concentrations of virus-related material from protein solutions. The yield of protein from the coagulation factor concentrate was about 85% and the material showed the same stability and biological activities as before the chromatography.     

Polymorphisms in the genes for coagulation factors II, V, and VII in patients with ischemic heart disease

BACKGROUND: Cardiovascular disease remains the leading cause of mortality in the United States, accounting for approximately 33% of all deaths in this country. Of these deaths, most are due to acute myocardial infarctions (AMIs), which are associated with thrombotic coronary artery obstruction and/or occlusion. These events could potentially be due to alterations in genes coding for coagulation factors. Several polymorphisms have been described in the factor II, V, and VII genes, which may predispose one to increased risk for ischemic heart disease (IHD). OBJECTIVE: To determine if mutations in 3 coagulation factor genes could predispose an individual to increased risk for arterial thrombosis as a mechanism for developing unstable angina (UA) or AMI. METHODS: We examined 125 hospitalized patients (mean age, 53 +/- 6 years, 79 men and 46 women), including 32 with AMI, 68 with UA, and 25 noncardiac controls, for a genetic predisposition for increased risk of IHD. EDTA-anticoagulated whole blood was collected at the time of hospital admission. DNA was extracted, and the polymorphisms were detected by polymerase chain reaction amplification of these genes with subsequent restriction enzyme digestion and gel electrophoresis. RESULTS: Our results showed that 3 (9.4%), 3 (4.4%), and 1 (4%) individuals were heterozygous for prothrombin G20210A and 3 (9.4%), 5 (7.4%), and 1 (4%) individuals were heterozygous for factor V Leiden in the AMI, UA, and control groups, respectively. The following genotype frequencies for the factor VII R353Q polymorphism were identified: 25 (78.1%), 56 (82.4%), and 18 (72%) with RR and 7 (21.9%), 12 (17. 6%), and 7 (28%) with RQ in the AMI, UA, and control groups, respectively. No QQ homozygotes were identified. For the HVR4 size polymorphism, the following genotypes were identified: 3 (9.4%), 4 (5.9%), and 5 (20%) individuals with H7H7; 11 (34.4%), 33 (48.5%), and 12 (48%) with H6H7; and 18 (56.2%), 31 (45.6%), and 8 (32%) with H6H6 genotypes in the AMI, UA, and control groups, respectively. There were no H7H5 and H6H5 genotypes found in this study. CONCLUSIONS: Although the frequency differences of these polymorphisms in patients with AMI and UA were not statistically significant from those in controls, several trends are consistent with what has been reported in the literature. Although any of these or other undefined genetic abnormalities may result in IHD, it is possible that phenotypic predisposition to IHD initially presents as UA. A larger population study addressing the significance of these polymorphisms in the sequence of events that lead to IHD, including cases of UA, is warranted.     

Association between necropsy evidence of disseminated intravascular coagulation and coagulation variables before death in patients in intensive care units.

The necropsy findings in 21 patients on an intensive care unit, on whom coagulation studies had been performed immediately before death, were assessed. Eleven of the patients were retrospectively studied and 10 were reviewed consecutively in a prospective study. Fifteen patients (eight retrospective and seven prospective) had evidence of disseminated intravascular coagulation. Microthrombi were most often found in the lungs and kidneys. The most common abnormal coagulation tests in patients with necropsy evidence of disseminated intravascular coagulation were raised serum concentrations of fibrinogen and fibrin degradation products, prolonged prothrombin time, and reduced platelet counts. Reduced fibrinogen concentrations and a prolonged thrombin time were the least commonly observed abnormalities. There was no difference in either the prevalence or magnitude of abnormality of any particular coagulation variable test result between those patients with evidence of disseminated intravascular coagulation at necropsy and those without.     

Effect of coenzyme A on triglyceride and very-low-density lipoproteins secretion in cultured rat hepatocytes.

Exogenous coenzyme A (CoA) decreases plasma triglycerides, cholesterol, and Apo B in man. CoA regulates lipid metabolism favouring beta-oxidation in hepatic peroxisomes of very-long-chain fatty acids. Furthermore recent studies show that CoA participates in the transport processes which occur in the Golgi apparatus. The aim of this study was to establish whether exogenous CoA is able to modify the lipid composition of very-low-density lipoproteins (VLDL) and the VLDL secretion in rat hepatocyte culture. The presence of 5mM CoA produces a significant decrease of VLDL triacylglycerol, of VLDL total cholesterol and of VLDL esterified cholesterol by 32%, 39% and 41% respectively. This decrease is observed in all the three days of hepatocyte culture. On the third day a significant decrease of cytosolic triacylglycerols is also observed. The decrease in VLDL secretion depends on the concentration of CoA added to the culture medium. Our study shows that exogenous CoA decreases the plasma VLDL concentration because it reduces VLDL secretion by the hepatocytes.     

Association between idiopathic premature ovarian failure and fragile X premutation

A total of 106 women affected by premature ovarian failure (POF) were evaluated for fragile X (FRAXA) premutation. The POF patients were classified as having a familial condition (33 women), at least one relative with early menopause (12 women), or a sporadic condition (61 women). The FRAXA premutation was only detected in patients with familial (four out of 33) or sporadic POF (two out of 61). In general, the results obtained indicated that the prevalence [six out of 106, 6%, 95% confidence interval (CI) 3-11%] of FRAXA premutation is significantly higher in women affected by POF than expected (P = 1.24x10(-3)), suggesting a phenotype consequence of the premutation alleles. This relationship is more convincingly derived from the observation in two analysed pedigrees of a co-segregation between FRAXA and POF. These findings suggest a possible involvement of premutated alleles in ovarian failure, and indicate the utility of POF families screening for FRAXA premutation in order to prevent the transmission of mental retardation syndrome.     

早期自然流产与环境因素的相关性分析

目的探讨与早期自然流产相关的环境危险因素,为降低自然流产率、减少不良妊娠结局的发生提供参考。方法采用问卷调查的方式,分析比较了97例在厦门市妇幼保健院诊断为早期自然流产的妇女和76例同期正常分娩的妇女的个人史资料和早孕期环境接触情况。结果单因素分析显示,自然流产组妇女的妊娠次数、既往自然流产次数、早孕期被动吸烟、每日手机通话次数以及使用染发剂的比例显著高于对照组(P均<0.05)。多因素分析显示,以上提到的三个环境接触因素均可增加早期自然流产的危险。结论早孕期被动吸烟、每日手机通话次数以及使用染发剂均与自然流产有一定关联,因此孕妇在怀孕期间应尽量避免被动吸烟、长时间使用电磁辐射大的设备以及接触染发剂等化学物质。     

Effects of low-density lipoproteins on blood coagulation and fibrinolytic activity

In vitro experiments showed that copper-oxidized low-density lipoproteins activate factors of the prothrombin complex in the whole blood and inhibit fibrin generation in both blood and plasma. Moreover, oxidized low-density lipoproteins inhibit fibrinolysis and impair the structure of fibrin clot, which results in hypercoagulation. Translated fromByulleten' Eksperimental'noi Biologii i Meditsiny, Vol. 129, No. 6, pp. 637–639, June, 2000     

Association of the Robin sequence with the fragile X syndrome.

We report on 4 individuals with the fragile X [fra(X)] syndrome and the Robin sequence (or elements of that sequence). To our knowledge, this association has been described in only one other boy. However, males with the fra(X) syndrome have been reported to have an increased incidence of cleft palate. We recommend that children with a cleft palate or the Robin sequence be assessed for developmental delays and a family history of mental retardation. The fra(X) syndrome may be one of the genetic causes of the Robin sequence and, when indicated, children with the sequence should be tested for fra(X).     

Platelet and coagulation factors in proliferative diabetic retinopathy.

Plasma beta-thromboglobulin, platelet factor 4, fibrinogen, fibrinopeptide A, antithrombin III, factor VIII related antigen, alpha 2-macroglobulin, platelet count, and total glycosylated haemoglobin were measured in three well matched groups of subjects: non-diabetic controls, diabetics without retinopathy, and diabetics with proliferative retinopathy. beta-thromboglobulin and platelet factor 4 concentrations were significantly higher in the diabetics with retinopathy than in the controls and platelet factor 4 was also increased in the diabetics without retinopathy compared with controls. Fibrinogen concentration was raised in diabetics without retinopathy compared with controls, diabetics with retinopathy compared with controls, and diabetics with retinopathy compared with those without. Fibrinopeptide A concentration did not differ significantly between groups. Antithrombin III levels were increased in diabetics with retinopathy compared with controls, and in diabetics with retinopathy compared with those without. Factor VIII related antigen values were higher in both the diabetic groups when compared with the controls. Fibrinopeptide A concentration correlated with both beta-thromboglobulin and platelet factor 4 in each of the three groups. Haemostatic abnormalities in diabetes have been shown, although a hypercoagulable state has not been confirmed. These changes in platelet and coagulation function may be secondary to the development of microvascular disease and their role in the pathogenesis of retinopathy remains uncertain.