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S R Poort

Publications and source records attributed to S R Poort.

At least 37 records · Page 2Linked to original sources
PubMed1995

Prevalence of protein C deficiency in the healthy population.

Reported prevalence rates for protein C (PC) deficiency in the population at large have varied widely. The differences presumably reflect the existence of an apparently high number of clinically recessive forms of the deficiency. In an attempt to document more precisely the prevalence of PC deficiency in the healthy population we have measured PC activity in just under 10,000 blood donors in the West of Scotland. After repeat testing of donors with low results and then further observation and selection, 32 donors were identified who had individual mean PC activities below the age- and gender-specific study reference range. Assessment of available first degree relatives, and also PC gene analysis in 23 of these donors, allowed identification of at least 14 with an inheritable deficiency (8 by both family study and gene analysis, 3 by family study alone and 3 by gene analysis alone). Two recurring and seven unique point mutations, only one of which has been previously described, were identified. The observed prevalence of inherited PC deficiency was 1.45 per 1000 (95% CI, 0.79/1000 to 2.43/1000). However after correcting for the possibility of missing some genuine inherited deficiencies we estimated the prevalence to be as high as 1 in 500. All cases of hereditary deficiency were asymptomatic with regard to thrombosis and none had a strong family history of thrombosis.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗
PubMed1994

Determination of the allelic and haplotype frequencies of three polymorphisms in the promoter region of the human protein C gene.

The frequencies of three polymorphisms in the promoter region of the human protein C gene (C/T at -1654, A/G at -1641 and A/T at -1476) have been determined. All three polymorphisms show allelic frequencies of about 0.60 and 0.40. The determination of the frequencies of the haplotypes of these polymorphisms showed that three of them (CGT, TAA and CAA) account for 88% of the observed haplotypes.

Alleles↗
PubMed1994

Homozygosity for a novel missense mutation in the prothrombin gene causing a severe bleeding disorder.

A patient with a severe bleeding tendency and hypoprothrombinemia (Factor II activity 2%, Factor II antigen 5%) was screened for the presence of alterations in his prothrombin gene. Direct sequencing of PCR fragments derived from the coding and flanking regions of the prothrombin gene, revealed that the patient was homozygous for an A-->G substitution in exon 3. This substitution predicts the replacement of Tyr44 (TAC) by Cys (TGC) in the prothrombin molecule. Both parents were found to be heterozygous for the same mutation. Further family studies revealed complete cosegregation of the mutation with the prothrombin deficiency. Only the five homozygous brothers and sisters of the propositus were clinically affected (severe hemorrhages including epistaxis, soft tissue, muscle and joint bleedings in all, and severe hemorrhages in the two women). The bleeding tendency therefore seems to inherit as an autosomal recessive trait.

Adenine↗
PubMed1993

Increased risk of venous thrombosis in carriers of hereditary protein C deficiency defect.

The relevance of heterozygosity for hereditary protein C deficiency as a risk factor for venous thrombosis has been disputed because heterozygotes without symptoms have been identified among blood donors and relatives of homozygotes. As a result, clinicians do not know whether to offer prophylaxis or not. We have compared thrombosis-free survival in 161 heterozygous and normal members of the families of 24 heterozygotes for protein C deficiency referred from several centres in the Netherlands and with a history of symptoms. We studied the influence of heterozygosity and of putative additional risk factors on the occurrence of thrombotic events noted when a medical history was taken. Protein C activities were measured but a diagnosis of heterozygosity was based on the presence of the specific mutation in one of the protein C genes identified in the proband of the family. We found a significant difference in the thrombosis-free survival of the 77 heterozygotes and 84 normals: by age 45, 50% of heterozygotes and 10% of normal relatives can be expected to have had a manifestation of venous thromboembolism. The presence of such a mutation was clearly associated with an increased risk of venous thrombotic events. Thrombotic events occurred more often in years in which the patient had been immobile for more than a week or had had surgery. Other putative risk factors showed no significant effect in the incidence of thrombotic events. About 50% of all first episodes and 65% of recurrences of venous thromboembolism in the heterozygotes were spontaneous--ie, there was no predisposing event such as surgery or pregnancy. There was no increased risk for arterial occlusions in heterozygotes. We conclude that members of the family of a symptomatic heterozygote proband who are heterozygous for the mutation in the protein C gene have an increased risk of venous thrombotic events compared with their normal family members. For such individuals prophylactic anticoagulation should be considered; the decision will need to be taken on an individual basis.

Adolescent↗
PubMed1993

Twelve novel and two recurrent mutations in 14 Austrian families with hereditary protein C deficiency.

The molecular basis of hereditary type I and type II protein C deficiency was studied in a panel of 14 unrelated Austrian families. By direct sequencing of the nine exons and their splice junctions sequence alterations were found in one of the protein C alleles in all but one subject. In twelve subjects a single alteration was found whereas in one subject one of the protein C alleles carried two sequence abnormalities. Whenever DNA from family members was available (11 of the 14 cases) cosegregation of the protein C deficiency with the mutation was observed. In contrast to what has been found previously in a panel of Dutch patients with hereditary protein C deficiency, none of the 14 mutations occurred in more than one family. Only two of the genetic defects (157Arg-->Stop and 178Arg-->Gln) have been found previously in other geographic locations. These data confirm the large genetic heterogeneity of protein C deficiency.

Amino Acid Sequence↗
PubMed1991

The spectrum of genetic defects in a panel of 40 Dutch families with symptomatic protein C deficiency type I: heterogeneity and founder effects.

Heterozygosity for protein C deficiency is associated with thromboembolic episodes, but clinical symptoms are nonrandomly distributed among protein C deficient families. This finding has led to the provisional definition of clinically dominant and clinically recessive protein C deficiency. We report here the molecular basis of hereditary, clinically dominant protein C deficiency in a panel of 40 Dutch probands from apparently independent families. All but one subject was a heterozygote for a point mutation in the protein C gene. These 39 subjects shared 15 mutations, six of which occurred in more than one proband (between two and nine). The diversity in the 15 mutations, together with the observation that the most frequent Dutch mutation was also found in a Swedish family with clinically recessive protein C deficiency, makes it unlikely that the molecular basis of protein C deficiency will be different between the clinically dominant and recessive forms. The recurrence of one of the mutations is most likely due to a founder effect, which suggests that when an additional hereditary factor is involved in the clinical severity of protein C deficiency this factor may remain linked to the protein C gene over many generations.

Base Sequence↗
PubMed1990

Two mutations of the factor IX gene including a donor splice consensus deletion and a point mutation in a Dutch patient with severe hemophilia B.

The abnormal factor IX gene of a patient with severe hemophilia B (hemophilia B Ursem) was selected for study. All of the coding and their flanking regions and parts of the 5'- and 3'-untranslated regions of the factor IX gene were amplified from the patient's genomic DNA by using the polymerase chain reaction (PCR). By analyzing the nucleotide sequence of the PCR products we have identified two mutations in the patient's factor IX gene, viz. a tetranucleotide deletion (GAGT, nt 6492 to 6495) or (TGAG, nt 6491 to 6494) in the 5'-donor splice site consensus at the exon 2-intron B boundary, and a point mutation at nucleotide 31103 in the catalytic domain (exon 8) of factor IXa, which changes the codon for valine 328 (GTT) to one for isoleucine (ATT). PCR-amplified exon 8 from 45 normal males and 55 normal females had the codon for valine-328. We propose that the deletion within the donor splice-site consensus is the cause of the disease in this individual, whereas the substitution of valine-328 by isoleucine may be a neutral variant which is, at least, very rare in the normal population. In a family study the DNA sequence of the patient's mother shows both the G to A transition in exon 8 and the 5'-donor splice consensus deletion in intron B in one allele.

Amino Acid Sequence↗
PubMed1990

Application of two neutral MspI DNA polymorphisms in the analysis of hereditary protein C deficiency.

Screening of restriction enzyme digested DNA from normal and protein C deficient individuals with a variety of probes derived from the protein C locus has revealed the existence of two neutral MspI polymorphism. One polymorphism (MI), which is located approximately 7 kb upstream of the protein C gene, has allelic frequencies of 69 and 31%, and was used to exclude extensive gene deletions as a likely cause of type I protein C deficiency in 50% of cases in a panel of 22 families. Furthermore, the same polymorphism has been used in 5 doubly affected individuals establishing compound heterozygosity in 3 of these. The second, intragenic, polymorphism (MII) has allelic frequencies of 99 and 1% in the normal population. The frequency of the rare allele of this RFLP was with 7% much higher in a panel of 22 Dutch families with protein C deficiency. Interestingly, in all three probands that were heterozygous for MII the rare allele of MII coincided with a point mutation that leads to a stop codon in amino acid position 306 of the protein C coding sequence. This mutation may account for 14% of the protein C deficient individuals in The Netherlands.

Alleles↗
PubMed1990

Mutations in hemophilia Bm occur at the Arg180-Val activation site or in the catalytic domain of factor IX.

Hemophilia Bm is characterized by a strikingly prolonged plasma ox brain prothrombin time. In an attempt to find an explanation for this phenomenon we have analyzed various aspects of the Bm variants factor IX Deventer, factor IX Milano, factor IX Novara, and factor IX Bergamo. Proteolytic cleavage by factor XIa was normal in two Bm variants, but absent at the Arg180-Val bond in the other two. In the latter variants Arg180 was replaced by either Trp or Gln, whereas Val181----Phe and Pro368----Thr replacements have occurred in the variants that were normally cleaved by factor XIa. In all four variants the Bm effect could be neutralized with a single monoclonal antibody against factor IX. Also, after treatment with factor XIa, none of the Bm variants reacted with antithrombin III (in contrast to normal factor IXa). Purified factor IX Deventer (one of the variants with a replacement of Arg181), either with or without pretreatment with factor XIa, was found to be a more effective competitive inhibitor of the factor VIIa-tissue factor-induced factor X activation than similarly treated normal factor IX. In addition, this inhibitory effect was much more pronounced when bovine tissue factor was used instead of human tissue factor. We propose that the normal activation of factor IX not only produces a conformational change around the active site serine that allows efficient substrate binding and catalysis, but that the same conformational change is instrumental in effectively dissociating factor IXa from the activating factor VIIa-tissue factor complex. Amino acid replacements that disrupt this conformational transition directly (e.g. Pro368----Thr near the catalytic center) or indirectly (mutations at the Arg180-Val activation site) therefore lead to a combination of 1) the loss of coagulant activity and 2) an inhibitory effect in the ox brain prothrombin time assay.

Amino Acid Sequence↗
PubMed1988

Immunoradiometric assays for the Ca(II)-dependent and Ca(II)-independent conformation of human protein C.

From a rabbit antiserum against human protein C, two subpopulations of antibodies recognizing Ca(II)-dependent and NonCa(II)-dependent epitopes of human protein C respectively were affinity purified for use in solid-phase immunoradiometric assays. The two assays were specific for PC:Ca(II)Ag and PC:NonCa(II)Ag and highly sensitive with a lower limit of detection of 2 ng/ml. The concentration of PC:Ca(II)Ag and PC:NonCa(II)Ag and their ratio in healthy volunteers was 0.91 +/- 0.16 U/ml, 0.95 +/- 0.14 U/ml and 0.97 +/- 0.18. In a group of patients treated with oral anticoagulants the ratio PC:Ca(II)Ag/PC:-NonCa(II)Ag was significantly decreased (to about 0.6) as compared with normal controls suggesting the presence of sub- and noncarboxylated protein C molecules (PIVKA-PC) in plasma of patients on oral anticoagulants. Analysis of plasmas of patients with a hereditary (n = 8) or isolated (n = 4) type II protein C deficiency and a history of thrombo-embolic disease did not reveal patients with a PC:Ca(II)Ag level and a PC:Ca(II)Ag/PC:NonCa(II)Ag ratio below the lower limit of the normal range suggesting that the frequency of genetic protein C variants with defects in the Ca(II)-dependent conformation is low.

Animals↗
PubMed1987

Immunoradiometric assay for the calcium-stabilized conformation of human protein S.

Rabbit polyclonal anti-protein S serum was fractionated with immobilized human protein S to establish solid-phase immunoradiometric assays recognizing Ca(II)-dependent and NonCa(II)-dependent epitopes of human protein S. The two assays were specific for PS:Ca(II)Ag and PS:NonCa(II)Ag and highly sensitive with a lower limit of detection of about 2.5 ng/ml. PS:Ca(II)Ag and PS:NonCa(II)Ag levels were measured in immunopurified protein S, thrombin-modified protein S and chymotrypsin-cleaved protein S. Only in chymotrypsin-cleaved protein S an important discrepancy between the two antigen levels was observed. Ranges for the concentration of PS:Ca(II)Ag and PS:Non-Ca(II)Ag and their ratio were established in plasma of healthy individuals (0.92 +/- 0.13 U/ml, 0.98 +/- 0.21 U/ml, 0.96 +/- 0.17, respectively). In a group of patients using oral anticoagulant therapy the ratio PS:Ca(II)Ag/PS:NonCa(II)Ag decreased at increasing intensity of anticoagulation suggesting the presence of sub- and noncarboxylated protein S molecules. In plasma of patients with a hereditary type I protein S deficiency PS:Ca(II)Ag and PS:NonCa(II)Ag were reduced to the same extent: mean ratio 1.02 +/- 0.12 in the group not on oral anticoagulant treatment and 0.94 +/- 0.10 in the group on oral anticoagulant therapy. Analysis of patients with a history of unexplained thrombo-embolic disease did not reveal individual patients with a PS:Ca(II)Ag/PS:NonCa(II)Ag ration below the lower limit of the normal range (mean ratio 1.05 +/- 0.17), suggesting that the frequency of genetic protein S variants with defects in the Ca(II)-stabilized conformation is very low.

Anticoagulants↗
PubMed1986

Rabbit polyclonal antibodies against the calcium-dependent conformation of factor IX and their application in solid phase immunoradiometric assays.

Two subpopulations of antibodies were isolated from rabbit polyclonal antiserum directed against human factor IX: one against the Ca(II)-dependent conformation of factor IX and one against the Ca(II)-independent conformation of factor IX. The two subpopulations were used for the development of immunoradiometric assays (IRMA's) for factor IX:Ca(II)Ag and factor IX:NonCa(II)Ag respectively. Ranges for the concentration of factor IX:Ca(II)Ag and factor IX:NonCa(II)Ag were established in plasmas of healthy volunteers, patients treated with oral anticoagulants and hemophilia B patients. In the group of patients using oral anticoagulant therapy a progressively reduced ratio of factor IX:Ca(II)Ag to factor IX:NonCa(II)Ag was observed with increasing intensity of oral anticoagulant treatment. Variant factor IX molecules from hemophilia B patients (CRM-, CRM(Red) and CRM+) with a defective Ca(II) binding or defective conformational transition induced by Ca(II) binding, were identified. These defects are absent in variant factor IX molecules from one hemophilia Bm patient and from patients with hemophilia B Leyden.

Administration, Oral↗
PubMed1985

Determination of plasma protein S--the protein cofactor of activated protein C.

Protein S, an important cofactor of activated protein C, and C4b-binding protein were purified from human plasma. Specific antibodies against the purified proteins were raised in rabbits and used for the development of immunologic assays for these proteins in plasma: an immunoradiometric assay for protein S (which measures both free protein S and protein S complexed with C4b-binding protein) and an electroimmunoassay for C4b-binding protein. Ranges for the concentrations of these proteins were established in healthy volunteers and patients using oral anticoagulant therapy. A slight decrease in protein S antigen was observed in patients with liver disease (0.78 +/- 0.25 U/ml); no significant decrease in protein S was observed in patients with DIC (0.95 +/- 0.25 U/ml). Criteria were developed for the laboratory diagnosis of an isolated protein S deficiency.

Anticoagulants↗
PubMed1981

Effect of feeding diets of different composition on the protein synthetic pattern of the rat pancreas.

Rats were fed a protein-rich or carbohydrate-rich diet for 3 weeks. By means of a double-label technique, we compared the protein-synthesizing patterns under these conditions. After 3 weeks of feeding, the relative rates of all the identified secretory enzymes and of some unidentified components had changed. The synthetic rates of the proteolytic proenzymes and of ribonuclease were increased after feeding a protein-rich diet, whereas those of the amylases and of lipase were lowered. The protein synthetic patterns of the rat pancreas under two extreme feeding conditions were compared to that of the pancreas of rats constantly fed a balanced diet of an intermediate composition. The results indicated that changes in relative rate of synthesis are mainly provoked by the protein-rich diet. Only the synthetic rate of the anodic chymotrypsinogen was strongly influenced by the carbohydrate-rich diet.

Amylases↗