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Biomedical subjects

W C Hooper

Publications and source records attributed to W C Hooper.

At least 37 records · Page 2Linked to original sources
PubMed2000

Genetic factors associated with thrombosis in pregnancy in a United States population.

OBJECTIVE: Polymorphisms in the genes for factor V (factor V Leiden), prothrombin, methylenetetrahydrofolate reductase, and angiotensin-converting enzyme have been associated with the occurrence of venous thrombosis. The objective of this study was to determine the relationships of these polymorphisms to thrombosis during pregnancy. STUDY DESIGN: This case-control study included 41 case patients with venous thrombosis during pregnancy and 76 control subjects matched for hospital and for race (white vs black) who had a normal pregnancy. RESULTS: Among white subjects, mutations in the genes for factor V and prothrombin were associated with increased risks of venous thrombosis during pregnancy (factor V: odds ratio, 18.3; 95% confidence interval, 2.7-432; P =.001; prothrombin: odds ratio infinity; 95% lower confidence limit, 1.7; P =.01). No black subject had either of these two mutations. For both black and white subjects the D/D genotype of the gene for angiotensin-converting enzyme entailed increased risk compared with the other genotypes (odds ratio, 2.7; 95% confidence interval, 1.2-6.3; P =.02). The polymorphism in the gene for methylenetetrahydrofolate reductase was unrelated to thrombosis during pregnancy among both blacks and whites. CONCLUSION: Women who had thrombotic complications during pregnancy demonstrated an increased prevalence of genetic mutations related to coagulation. The additional risk of thrombosis during pregnancy associated with such genetic mutations can be substantial.

Black People↗
PubMed1999

Multiplex analysis of mutations in four genes using fluorescence scanning technology.

Multiplex analysis of genetic mutations using fluorescence scanning methodology is an accurate, efficient, and cost-effective approach to genotypic characterization. Fluorescence labeling during the synthesis of polymerase chain reaction primers allows the application of this technology to well-established protocols. We have simultaneously analyzed the four polymorphisms of factor V Leiden (G1691A), prothrombin G20210A, 5,10-methylenetetrahydrofolate reductase C677T, and cystathionine beta-synthase 844ins68. Three of these mutations have been associated with an increased risk of thrombosis. Following polymerase chain reaction with fluorescence-labeled primers, the polymerase chain reaction products were digested with an appropriate restriction enzyme (if necessary for detection of the mutation), diluted into one tube per sample for co-loading (multiplex loading), and analyzed with GeneScan software for fragment analysis following capillary electrophoresis on an ABI PRISM 310 Genetic Analyzer (Foster City, CA, USA). Multiplex loading increased throughput without compromising precision.

Cystathionine beta-Synthase↗
PubMed1999

The relationship between polymorphisms in the endothelial cell nitric oxide synthase gene and the platelet GPIIIa gene with myocardial infarction and venous thromboembolism in African Americans.

STUDY OBJECTIVES: To determine whether the polymorphic dinucleotide repeats found in intron 4 of the endothelial cell nitric oxide synthase (ecNOS) gene and the platelet GPIIIa PLA(1)/A(2) polymorphism are associated with myocardial infarction (MI) and venous thromboembolism (VTE) in African Americans. Because these two genes may interact physiologically, the third objective was to determine if there was a relationship between the polymorphisms with respect to MI and VTE. DESIGN: A hospital-based case-control study. After informed consent was obtained, blood used for DNA extraction was drawn from the subjects. SETTING: The study was conducted in the Anticoagulant Clinic and the Cardiology Clinic at Grady Memorial Hospital in Atlanta Georgia. PATIENTS: Subjects were recruited from African-American patients with a reported history of MI (n = 110) or VTE (n = 91). Control subjects (n = 185) without a history of cardiovascular or venous disease were recruited from an outpatient clinic. MEASUREMENTS AND RESULTS: The 393 ecNOS allele was more common among MI cases (36%; p = 0.01) and VTE cases (35%; p = 0.04) than among control subjects (26%). There was no association between the GPIIIa genotypes and either MI or VTE. However, among the MI subjects, there was a strong association between the ecNOS 393/393 genotype and the Pl(A2) allele. It was also found that the frequency of the 393 allele was higher in African-American persons (0.26) compared with what has been reported for Australian Caucasians (0. 14) and Japanese (0.10). CONCLUSIONS: The 393 allele but not the Pl(A2) allele was significantly associated with both MI and VTE in African Americans. Homozygosity for the 393 allele was significantly associated to the diagnosis of MI prior to the age of 45. The combination of the 393 allele and a Pl(A2) allele was also highly associated with MI. The frequency of the 393 allele was significantly higher in African Americans than what has been reported for other populations. This study furthers not only extends the association of the 393 allele to VTE but has demonstrated an interaction with the Pl(A2) allele with respect to MI.

Adult↗
PubMed1998

The up-regulation of IL-6 and IL-8 in human endothelial cells by activated protein C.

The protein C/protein S anticoagulant pathway has been proposed to be a common link between coagulation and inflammation. Studies have suggested that a component of the anticoagulant pathway, activated protein C (APC), may play a role in the inflammatory response by modulating the effects of cytokines such as TNF and by blocking neutrophil activation. Cytokines are known to be intimately involved in the inflammatory response and to function in part to restore hemostatic balance. To begin to delineate what role APC may have in the inflammatory response, we have investigated the effect of APC on the production of the proinflammatory cytokines IL-6 and IL-8 in primary HUVEC, human microvascular endothelial cells, and human coronary artery endothelial cells. Our results have demonstrated that physiologic concentrations of APC significantly up-regulated the production of both IL-6 and IL-8. This increase, which was seen at both the RNA and protein level, was not due to either thrombin or LPS contamination of the APC preparation. Additional studies also showed that the APC-mediated up-regulation of IL-6 and IL-8 was IL-1 independent. Although neither purified protein C nor protein S alone had an effect on cytokine production, protein S, the cofactor for APC, significantly enhanced the ability of APC to up-regulate IL-6/IL-8 production. These results provide further evidence for a role for APC in the inflammatory response.

Aprotinin↗
PubMed1998

Relation of three genetic traits to venous thrombosis in an African-American population.

A mutation in the Factor V gene (Factor V Leiden), a variant in the 5,10-methylenetetrahydrofolate reductase gene (MTHFR), and an insertion/deletion polymorphism of the angiotensin I-converting enzyme gene (ACE) may be related to abnormal blood clotting. The authors examined the associations between these genetic traits and venous thrombosis among African Americans. This study comprised 93 patients with venous thrombosis and 185 control subjects attending clinics at an urban, public hospital in Atlanta, Georgia, in 1995-1996. Subjects' DNA was extracted from blood and assayed for these genetic traits. Odds ratios were obtained from logistic regression and used as a measure of association between each genetic trait and venous thrombosis. Factor V Leiden was unrelated to venous thrombosis, but the mutation ws too rare among our African-American subjects to evaluate adequately its relation to venous thrombosis. The homozygous and heterozygous genotypes for the V allele of the MTHFR gene were unrelated to venous thrombosis (odds ratio = 0.9, 95% confidence interval 0.5-1.8). Subjects with the deletion/deletion ACE polymorphism experienced a moderate increase in venous thrombosis risk compared with persons with the other genotypes (odds ratio = 1.5, 95% confidence interval 0.9-2.6). However, women with this ACE genotype experienced no increased risk (odds ratio = 0.9, 95% confidence interval 0.5-1.9), whereas men with this genotype had nearly three times the risk (odds ratio = 2.8, 95% confidence interval 1.2-6.2; p value for interaction = 0.06). These data indicate that the prevalence of Factor V Leiden and the V allele of the MTHFR gene is low among African Americans. The D allele of the ACE gene is equally prevalent among African Americans and whites and may be related to venous thrombosis among African-American men.

Adolescent↗
PubMed1998

Prevalence of the prothrombin 20210 G-to-A variant in blacks: infants, patients with venous thrombosis, patients with myocardial infarction, and control subjects.

A genetic variation in the prothrombin gene is located in the 3-untranslated region at position 20210 where a G-->A transition occurs. The prevalence of the mutation is 1% to 2% in white populations, and the mutation is associated with an increased risk of venous thrombosis and myocardial infarction. We report the prevalence of the A allele in blacks at birth; in black control subjects with no history of heart attack, stroke, or blood clots; in black patients with venous thrombosis; and in black patients with myocardial infarction. Among 318 infants, the prevalence of the A allele was 0.2% (1 heterozygote), with an exact, one-sided upper 95% confidence limit of 0.7%. Among 185 control subjects the variant was absent, yielding an exact, one-sided upper 95% confidence limit of 0.8% for the A allele. The heterozygous genotype was found in 2 of 91 subjects with deep vein thrombosis and in none of 123 subjects with myocardial infarction. The very low prevalence of the A allele indicates that the prothrombin variant is not a major cause of venous thrombosis or myocardial infarction in blacks.

3' Untranslated Regions↗
PubMed1998

The role of activated protein C resistance in the pathogenesis of venous thrombosis.

Venous thromboembolism (VTE) is the third most common cardiovascular disease in the United States. VTE is usually a consequence of either acquired or inherited alterations in hemostatic regulatory proteins. These regulatory proteins are predominantly those of the protein C/protein S natural anticoagulant pathway. Acquired deficiencies in this pathway are frequently a consequence of other clinical entities (eg, cancer, AIDS, and diabetes), while inherited deficiencies can be responsible for venous thrombosis in an otherwise healthy individual. The purpose of this article is to briefly describe the pathobiology of the anticoagulant protein system and to review the clinical implications of activated protein C resistance.

Aging↗
PubMed1998

Deletion polymorphism in the angiotensin-converting enzyme gene as a thrombophilic risk factor after hip arthroplasty.

Despite thromboprophylaxis, deep vein thrombosis is a common complication of major orthopedic surgery. Predisposing genetic risk factors are unknown. In this case-control study, we investigated the association of the insertion (I)/deletion (D) angiotensin converting enzyme (ACE) gene polymorphism, Factor V Leiden (R506Q) mutation, and 5,10 methylenetetrahydrofolate reductase (MTHFR) gene polymorphism with post-operative venous thrombosis in 85 patients who underwent elective total hip arthroplasty. The odds of a thrombotic event following hip surgery among subjects with the DD genotype of the ACE gene was increased more than 10-fold compared to subjects with the II genotype (odds ratio 11.7 [95% confidence interval 2.3-84.5]); it was increased 5-fold in subjects with the ID genotype compared to the II genotype (odds ratio 5.0 [95% confidence interval 1.1-34.9]). Mean plasma ACE level in control subjects not on ACE inhibitors at the time of study (n=43) was lowest in persons homozygous for the I allele (18.9+/-7.95 U/l), intermediate in patients with the ID genotype (31.6+/-10.8 U/l) and highest in subjects homozygous for the D allele (44.0+/-7.14 U/l). Mean plasma ACE level among cases was higher (33.0 U/l, n=25) than among controls (29.4 U/l, n=43) but this difference was not statistically significant. Neither the Factor V Leiden mutation nor MTHFR gene polymorphism increased the risk of thrombosis following hip replacement. These results demonstrate that the I/D ACE gene polymorphism is a potent risk factor for thrombosis in subjects undergoing total hip arthroplasty.

Activated Protein C Resistance↗
PubMed1997

Endothelial cell protein S synthesis is upregulated by the complex of IL-6 and soluble IL-6 receptor.

We have recently demonstrated that the proinflammatory cytokine, interleukin-6 (IL-6), could upregulate the production of protein S in the human hepatoma cell line, HepG-2, but not in endothelial cells. In this study, we have demonstrated that the combination of exogenous IL-6 and soluble IL-6 receptor (sIL-6R) could significantly upregulate protein S production in both primary human umbilical vein endothelial cells (HUVEC) and in the immortalized human microvascular endothelial cell line, HMEC-1. The IL-6/sIL-6R complex was also able to rapidly induce tyrosine phosphorylation of the IL-6 transducer, gp130. Neutralizing antibodies directed against either IL-6 or gp130 blocked protein S upregulation by the IL-6/sIL-6R complex. It was also observed that exogenous sIL-6R could also upregulate protein S by forming a complex with IL-6 constitutively produced by the endothelial cell. Two other cytokines which also utilize the gp130 receptor, oncostatin M (OSM) and leukemia inhibitory factor (LIF), were also able to upregulate endothelial cell protein S. This study demonstrates a mechanism that allows endothelial cells to respond to IL-6 and also illustrates the potential importance of circulating soluble receptors in the regulation of the anticoagulation pathway.

Antigens, CD↗
PubMed1996

Oligonucleotide ligation assay for detection of the factor V mutation (Arg506-->Gln) causing protein C resistance.

A point mutation in the Factor V (FV) gene at the activated protein C cleavage site, FV Arg (R)506-->FV Gln (Q)506, is the reported molecular basis for resistance to activated protein C (APC-R). This mutation has been reported in approximately 20-50% of individuals with previously unexplained thrombophilia and 3-5% of the general population. We have adapted an oligonucleotide ligation assay (OLA) for nonisotopic detection of the FV:Q506 mutation which permits rapid screening for this mutation. First, the polymerase chain reaction (PCR) was used for target DNA amplification, thus permitting nonisotopic reporters in the DNA analysis. Then thermostable ligase was used for ligation or covalent coupling of adjacent wild-type and mutant oligonucleotide probes which occurs only when the probes are annealed to a matched amplicon. A colorimetric ELISA-based detection assay was then used to capture 5' biotinylated probes in 96-well streptavidin-coated plates and by virtue of ligation, detection of a 3' digoxigenin reporter probe. Following the addition of anti-digoxigenin conjugate and enhanced alkaline phosphatase signal amplification, colorimetric substrate change was measured in an ELISA plate reader. This assay correctly identified FV genotypes of 290 samples.

Base Sequence↗
PubMed1996

A racial difference in the prevalence of the Arg506-->Gln mutation.

Several recent studies have reported that the factor V Arg506-->Gln mutation is present in 3-10% of adults of European descent. To determine if the prevalence is comparable among Blacks, we have initiated a case-control study in a large urban hospital in Atlanta which serves a substantial black population. We have evaluated 131 black subjects with confirmed venous or arterial thrombosis and 61 black subjects without a history of thrombosis. Only one case and one control were positive for the Arg506-->Gln mutation. We conclude that the mutation is more common among Whites than Blacks.

Arginine↗
PubMed1996

TNF-alpha suppresses IL-1 alpha and IL-6 upregulation of C4b-binding protein in HepG-2 hepatoma cells.

C4b-binding protein (C4BP) is a regulatory protein involved in the regulation of the classical pathway of complement and the natural anticoagulant pathway. C4BP is synthesized by the liver, a target organ of the IL-6 proinflammatory cytokine. C4BP is an acute-phase protein and its basal levels may increase by as much as 4-fold during an inflammatory response. IL-6 which plays a major role in the modulation of the acute-phase proteins, including C4BP, also has been shown by our group to significantly increase hepatocyte production of the anticoagulant protein, protein S. In this study, we have examined the role of cytokine combinations on the production of the C4BP regulatory protein in the HepG-2 hepatoma cell line and report that IL-1 alpha and IL-6 in combination as well as IL-1 alpha and Oncostatin M (OSM) were approximately additive in the upregulation of C4BP while IL-6 and OSM were not and that TNF-alpha blocked both IL-1 alpha and IL-6 but not OSM upregulation of C4BP.

Carcinoma, Hepatocellular↗
PubMed1996

TNF-alpha suppresses IL-6 upregulation of protein S in HepG-2 hepatoma cells.

The pathogenesis of disseminated intravascular coagulation (DIC) has, in part, been attributed to the impairment of the natural anticoagulant protein C/protein S pathway. DIC, which frequently occurs during sepsis, has been linked to cytokines that can induce or modulate procoagulant activity. Three of these cytokines, IL-1 alpha, IL-6, and TNF-alpha have been reported to be increased in the early stages of sepsis. In the present study, we have stimulated HepG-2 hepatoma cell cultures with recombinant human IL-1 alpha, IL-6, TNF-alpha, and oncostatin M (OSM). The results demonstrated that TNF-alpha, and to a lesser degree, IL-1 alpha, could significantly suppress IL-6 upregulation of protein S, whereas the effects of OSM was only suppressed by the combination of IL-1 alpha and TNF-alpha. The combination of IL-1 alpha and TNF-alpha also suppressed protein S production below that of control or basal levels. These results indicate that IL-1 alpha and TNF-alpha may play important regulatory roles in coagulation.

Antineoplastic Agents↗
PubMed1995

Hemostatic properties of the SV-40 transfected human microvascular endothelial cell line (HMEC-1). A representative in vitro model for microvascular endothelium.

HMEC-1 is a SV-40T transfected human microvascular endothelial cell line that constitutively expresses RNA transcripts for plasminogen activator inhibitor 1 (PAI-1), tissue-type plasminogen activator (t-PA), protein S (PS), von Willebrand factor (vWF), and thrombomodulin. Tissue factor (TF) can be induced in response to stimulation with tumor necrosis factor alpha (TNF-alpha), interleukin-1 alpha (IL-1alpha) and phorbol 12-myristate 13-acetate (PMA). Proteins corresponding to PAI-1, t-PA, protein S and vWF genes were constitutively released in the culture supernatant. This cell line is a model that will be useful to investigate coagulation/fibrinolytic properties of microvascular endothelium.

Basal Metabolism↗
PubMed1995

IL-6 upregulates protein S expression in the HepG-2 hepatoma cells.

Several pro-inflammatory cytokines have been shown to be important in the modulation of the procoagulant response. However, what role these cytokines may have in the regulation of coagulation inhibitors is poorly understood. While the hepatocyte is a primary site of synthesis for the anticoagulant protein C and S, it is also a major target cell for the proinflammatory cytokine, IL-6. We have found that stimulation of HepG-2 hepatoma cells with IL-6 (5 ng/ ml) significantly increased the production of the anticoagulant cofactor, protein S, in both a time and dose dependent fashion. This increase was seen at both the RNA and protein level. A mouse monoclonal neutralizing antibody to human IL-6 suppressed the IL-6 effect in a concentration dependent fashion. IL-6 also increased the release of the C4b-binding protein but had no effect on protein C production. When combined with either dexamethasone or soluble IL-6 receptor, the IL-6 response was significantly enhanced. Oncostatin M, a functionally related cytokine, had a similar effect while other related cytokines, IL-11 and leukemia inhibitory factor, only had a marginal effect. IL-1, TGF-beta, and TNF-alpha had no significant effect on protein S production. These results indicate that IL-6 may play an important regulatory role in the anti-coagulant pathway.

Animals↗
PubMed1995

MEG-01s cells have receptors for and respond to IL-3, IL-6, and SCF.

An established megakaryoblastic cell line, MEG-01s, was used to study receptor expression and receptor-mediated responses to factors known to affect megakaryocytopoiesis. In addition, the antigenic characteristics of this cell line were further defined. MEG-01s cells were CD34+CD33+CD38 +/- HLA-DR- and expressed erythroid and granulocytic differentiation antigens as well as many megakaryocytic lineage-restricted antigens. These cells also expressed receptors for interleukin-3 (IL-3), IL-6, and stem cell factor (SCF), as measured by flow cytometry and/or RNA expression. MEG-01s cell proliferation or survival was only marginally influenced by these factors and their combinations. c-kit, the receptor for SCF, was downmodulated by its ligand. This modulation was time-dependent, appeared to involve receptor conformational changes, and became concentration-dependent by day 3. Northern blot analysis indicated that amounts of c-kit RNA increased as downmodulation proceeded. IL-3 induced IL-6 secretion in these cells, which was augmented by a protein kinase-C (PKC) inhibitor, H7, and reduced by a tyrosine kinase inhibitor, genistein. Evidence for autocrine regulation of this cell line by IL-6 was demonstrated by the inhibitory effects of an antisense oligonucleotide on 3H-thymidine (3H-TdR) incorporation. These cells should prove useful for studies of the early signal transduction mechanisms involved in cytokine function.

Antigens, CD↗