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Biomedical subjects

E Persson

Publications and source records attributed to E Persson.

At least 91 records · Page 5Linked to original sources
PubMed1996

Effect of Ca2+ on the structure and function of factor VIIa.

Binding of Ca2+ to factor VIIa is required for its binding to phospholipids, tissue factor, and for obtaining full amidolytic activity. This presentation describes various binding studies performed with recombinant factor VIIa and fragments hereof in order to localize Ca2+ sites and obtain a better understanding of the functional implication of Ca2+ binding to these sites. Results obtained by means of Ca2+-dependent monoclonal antibodies, immunosorbent assays, intrinsic protein fluorescence and terbium luminescence technique are described and the results are interpreted in terms of a putative model.

Calcium-Binding Proteins↗
PubMed1996

HPV-types, cytological and histopathological findings in three groups of women with possible HPV-related disease.

OBJECTIVE: The aim of this investigation was to study three groups of women presenting with possible HPV-infection with regard to HPV-types and cervical dysplasia. METHODS: Eighty women were included. Eighteen of them were present partners to men with condylomas, 20 had clinical vulvar HPV-lesions and 42 were referred due to an abnormal PAP-smear. Samples for HPV-analysis by PCR-technique were taken from the vulva, the portio and the cervical canal. A universal HPV-primer as well as specific primers for HPV 6/11, 16, 18, 31, and 33 were utilized. PAP-smears were taken as well as biopsies from cervix/portio. RESULTS: Seventy-eight percent had HPV-DNA identified. Sixty-seven percent of those with HPV 16 and/or 18 had dysplasia verified by histopathology and 50% of those with 31 and/or 33. Twenty of 21 women with dysplasia had HPV 16, 18, 31 and/or 33 identified. One woman with dysplasia was HPV-negative. Histopathologically verified CIN were diagnosed in all groups investigated. Women referred for suspicion of CIN significantly more often had HPV detected at the cervix/portio. HPV 6/11 was mostly found in women with condylomas. Apart from this the occurrence of the different HPV types were alike in the three groups. CONCLUSION: Infection with HPV is a process and the usefulness of different diagnostic methods seems to depend on when during the course of the disease they are used. HPV-findings in women with dysplasia were all associated with oncogenic virus-types. High-risk virus was often found simultaneously with low-risk virus indicating a covariation in the acquisition of the different HPV-types.

Adolescent↗
PubMed1996

Infections after hysterectomy. A prospective nation-wide Swedish study. The Study Group on Infectious Diseases in Obstetrics and Gynecology within the Swedish Society of Obstetrics and Gynecology.

BACKGROUND: An increased use of prophylactic antibiotics to avoid postoperative infections in women undergoing hysterectomy has been observed in Sweden. This investigation was performed a) to study the infection rate to enable future evaluation of the effect of prophylactic antibiotic regimens and b) to identify subgroups suitable for intervention. METHOD: A prospective study comprising all women undergoing pelvic surgery with hysterectomy during a two month period at forty-two Departments of Obstetrics and Gynecology in Sweden. Relevant information regarding the surgical procedure and the postoperative course was included in a standardized form at discharge from hospital and at a follow-up visit 4 to 6 weeks after surgery. RESULTS: Of the 1060 women included in the study, 23% developed postoperative infections: 9.4% had wound-, cuff-, and/or deep infections, 13% urinary tract infections, and 4% other infections unrelated to the surgical site. Only half of the wound/cuff-/deep infections were diagnosed before discharge from the hospital. Wertheim-Meigs procedures, bleedings per-operatively exceeding 1000 ml, and presence of bacterial vaginosis (BV) were associated with an increased risk of postoperative infections. Among women undergoing abdominal hysterectomy for benign reasons, (n = 159), wound-cuff/deep infection was significantly associated with preoperative BV, i.e. 7 of 28 (25%) vs. 11 of 131 (8%), respectively (relative risk = 3.0, p = 0.01). Infection was associated with prolonged postoperative hospital stay. Antibiotics were given pre- or postoperatively to 236 (22%) of the 1060 women. Reduction in the postoperative infection rate was seen among women undergoing vaginal hysterectomy who were given pre- or postoperative antibiotics. CONCLUSION: The postoperative infection rate after hysterectomy was clinically significant in this population. Wertheim-Meigs procedures, peroperative bleeding > 1000 ml and BV were identified as risk factors for postoperative infections after hysterectomy.

Anti-Bacterial Agents↗
PubMed1996

Young women with symptoms of urinary tract infection. Prevalence and diagnosis of chlamydial infection and evaluation of rapid screening of bacteriuria.

OBJECTIVE: To estimate the prevalence of chlamydial infection among young women with UTI symptoms. To evaluate chlamydia diagnostics with the aid of enzyme immuno assay (EIA) on first-void urine. To evaluate rapid screening of bacteriuria, including low concentrations of common pathogens. DESIGN: EIA for detection of Chlamydia trachomatis antigen and confirmation with immunofluorescence test (DFA) in urine, cervical and urethral chlamydia culture, nitrite and granulocyte esterase test, urine sediment, chamber count, dipslide and conventional urine culture were performed. SETTING: Primary health care (PHC). PATIENTS: 217 women aged 15-35 years attending PHC for dysuria or urgency-frequency. MAIN OUTCOME MEASURES: Frequency of chlamydial infections. Sensitivity, specificity, predictive values of EIA and bacteriuria screening tests, respectively. RESULTS: The frequency of chlamydial infection was 3.7%. In spite of a high specificity of the EIA test (0.94 without DFA) the number of false positives exceeded the number of true positives. No single bacteriuria test showed sufficient diagnostic efficiency. CONCLUSIONS: Routine chlamydia testing in young women with UTI symptoms is recommended. EIA test on urine is of little use. Assessing diagnosis of UTI symptoms requires insight into the use of several rapid tests and a dialogue with the patient.

Adolescent↗
PubMed1995

Amino acid residues 721-729 are required for full factor VIII activity.

Recombinant two-chain factor VIII, from which the B domain had been deleted, was expressed in Chinese hamster ovary cells. In addition to the major product, three minor factor VIII forms were isolated. The A2 domains generated by thrombin cleavage showed different electrophoretic mobilities. Peptide mapping of the A2 domains showed that two of the factor VIII forms had the expected C-terminus of the heavy chain at Arg740 [FVIII-(1-740)] and that the other factor VIII forms had C-termini at Tyr729 [FVIII-(1-729)] or Glu720 [FVIII-(1-720)]. The major FVIII-(1-740) form, FVIII-(1-729), and FVIII-(1-720) contained sulfated tyrosine residues at Tyr718, Tyr719 and Tyr723. The minor FVIII-(1-740) form was shown to lack sulfation at these positions. The specific clotting activity was approximately 1 x 10(4) U/mg for FVIII-(1-740) (both forms) and FVIII-(1-729), but twofold lower for FVIII-(1-720). A time study of thrombin activation showed that FVIII-(1-720) was activated slower than FVIII-(1-740), FVIII-(1-729) and plasma-derived factor VIII. Partially sulfated FVIII-(1-740) was activated at the same rate as the fully sulfated FVIII-(1-740). The equilibrium dissociation constant for binding of factor VIII to inactivated immobilized thrombin was the same for all factor VIII forms, showing that the slower activation of FVIII-(1-720) was not due to a lower affinity for the anion-binding exosite in thrombin.

Amino Acid Sequence↗
PubMed1995

Structurally and functionally distinct Ca2+ binding sites in the gamma-carboxyglutamic acid-containing domain of factor VIIa.

The structural and functional effects of Ca2+ binding to vitamin-K-dependent coagulation factor VIIa were investigated. Conformational changes with a midpoint around 0.7 mM Ca2+ quenched the intrinsic protein fluorescence of a fragment of factor VIIa comprising only the light chain and this coincided with an increase in factor VIIa amidolytic activity in the absence of tissue factor. Ca2+ binding to sites in factor VIIa and in the fragment with an apparent dissociation constant of 1.3-1.4 mM induced binding to phospholipids. A similar Ca2+ dependency was not observed with factor VIIa lacking the N-terminal 38 or 44 residues of the light chain and the observed effects could thus be attributed to gamma-carboxyglutamic-acid-dependent Ca2+ binding. Mg2+ appeared to bind to the site(s) of relatively higher affinity since, although it was less efficient than Ca2+, it stimulated the amidolytic activity and induced quenching of the intrinsic fluorescence. In contrast, Mg2+ did not induce expression of the phospholipid-interactive structure. The binding properties of two monoclonal antibodies that recognized epitopes in the gamma-carboxy-glutamic-acid-rich domain of factor VIIa corroborated the occurrence of two Ca(2+)-induced, sequential structural changes and only one of the antibodies recognized the Mg(2+)-induced structure. Thus Ca2+ binding to the gamma-carboxyglutamic-acid-containing domain appeared to result in at least two distinct structural transitions with different functional consequences. The two (sets of) sites responsible for the observed effects could be distinguished based upon differences in Ca2+ affinity and metal ion selectivity. The interaction between factor VIIa and tissue factor was monitored by means of a direct binding assay and an amidolytic assay. In both systems, half-maximal Ca2+ enhancement was observed at 0.25 mM. This coincided with a Ca(2+)-induced conformational change in factor VIIa associated with fluorescence quenching. The same effect on amidolytic activity was observed with the two N-terminally truncated forms of factor VIIa and it is presumably mediated by Ca2+ binding to a site located in the serine protease part.

1-Carboxyglutamic Acid↗
PubMed1995

Kinetics of the interaction between the human factor VIIIa subunits: effects of pH, ionic strength, Ca2+ concentration, heparin, and activated protein C-catalyzed proteolysis.

Coagulation factor VIIIa consists of a heterotrimer in which the A2 subunit is bound to the A1/A3C1C2 dimer. The dissociation of this complex causes the spontaneous and reversible decay of factor VIIIa activity. In order to characterize the kinetics and affinity of the interaction between A2 and A1/A3C1C2, as well as the influence of different parameters on the interaction, the subunits were chromatographically separated and reassembled in a BIAcore instrument (Pharmacia Biosensor). In the binding experiments, A2 was free in solution, whereas A1/A3C1C2 was immobilized on the dextran surface by direct coupling or captured on an immobilized monoclonal anti-C2 antibody. At our chosen standard condition (pH = 6.0, I = 0.12, and [Ca2+] = 2 mM), the association rate constant, dissociation rate constant, and resulting equilibrium dissociation constant were ca. 1.4 x 10(4) M-1s-1, 2.1 x 10(-4)s-1, and 16 nM, respectively. Increasing the ionic strength or Ca2+ concentration resulted in both slower association and faster dissociation. At 0.3 M NaCl or 25 mM Ca2+, the dissociation constant was > 1 microM. This implies that electrostatic forces involved in the interaction contribute at least one-fourth of the total binding energy. Increasing pH caused a similar effect, yielding a dissociation constant of ca. 0.9 microM at pH 7.5. In those cases where the equilibrium dissociation constants had been determined from solution phase experiments [Fay, P. J., & Smudzin, T. M. (1992) J. Biol. Chem. 267, 13246-13250; Lollar, P., Parker, E. T., & Fay, P. J. (1992) J. Biol. Chem. 267, 23652-23657], these constants agreed well with our results.(ABSTRACT TRUNCATED AT 250 WORDS)

Calcium↗
PubMed1994

Characterization of chemically crosslinked human factor VIIIa.

The activity of factor VIIIa was enhanced and stabilized by treatment of factor VIII with the crosslinker disuccinimidyl suberate. The activity was > 200-fold higher compared with that of native factor VIIIa and was stable for at least 15 days at 4 degrees C and pH 7.2. The crosslinked factor VIIIa was purified by immunoaffinity chromatography and gel filtration. Electrophoretic analysis revealed high-molecular-mass (approximately 150 kDa) molecules as well as the three bands characteristic of native factor VIIIa. Thus crosslinking appeared to yield molecules stabilized by intra- and/or inter-subunit crosslinks. The material was further fractionated using immobilized von Willebrand factor and the factor VIIIa activity could be ascribed to trimers containing only intra-subunit crosslinks. Moreover, reduction of crosslinked factor VIIIa produced using dithiobis(succinimidylpropionate) suggested that the molecules containing intersubunit crosslinks had not been cleaved by thrombin at arginine 372.

Chromatography, Affinity↗
PubMed1994

Alcohol habits among teenagers in Sweden: factors of importance.

This study of 689 secondary school students (13-16 years of age) in Sweden investigates the association between alcohol habits, the availability of alcohol and age, gender, socioeconomic status, ethnic origin and family structure. Data were obtained by anonymous questionnaires in 1991. The availability of alcohol increased from the seventh grade to the ninth as did the proportion of alcohol consumers and students with regular alcohol consumption. Boys in the seventh and eighth grades showed somewhat more advanced alcohol habits than girls, but in the ninth grade the opposite was seen. Students with more advanced alcohol habits and a higher availability of alcohol more often belonged to a lower socioeconomic strata and they lived more often with a single parent. Students of foreign background drank alcohol (especially wine) more regularly. An association was also found between the parents' liberal attitude toward offering alcohol at home and frequent intoxication and the students' experience of illicitly produced liquor, especially among the youngest students. In spite of the Swedish alcohol policy the availability of alcohol is rather high among young people. Special attention in alcohol preventive work should be paid to girls, young people living with one parent, young people in lower socioeconomic groups and young people of foreign origin.

Adolescent↗
PubMed1993

The effect of aspartate hydroxylation on calcium binding to epidermal growth factor-like modules in coagulation factors IX and X.

Hydroxylation of aspartic acid to erythro-beta-aspartic acid (Hya) occurs in epidermal growth factor (EGF)-like modules in numerous extracellular proteins with diverse functions. Several EGF-like modules with the consensus sequence for hydroxylation bind Ca2+, and it has therefore been suggested that the hydroxyl group is essential for Ca2+ binding. To determine directly the influence of beta-hydroxylation on calcium binding in the EGF-like modules from coagulation factors IX and X, we have now measured calcium binding to both the fully beta-hydroxylated and the non-beta-hydroxylated modules of the two proteins. At low ionic strength, the Hya-containing module of factor X has a slightly higher Ca2+ affinity, but at physiological salt concentrations this difference is no longer significant for either factor IX or X. Analysis of the 1H NMR chemical shift differences between the hydroxylated and nonhydroxylated factor X modules show that hydroxylation has no effect on the domain fold. Furthermore, measurements on factor IX show that hydroxylation has no effect on the Ca2+/Mg2+ specificity of the ion binding site. We conclude that the hydroxyl group is not a direct ligand for the calcium ion in these EGF-like modules, nor is it essential for high-affinity Ca2+ binding.

Amino Acid Sequence↗
PubMed1993

Effects of Ca2+ binding on the protease module of factor Xa and its interaction with factor Va. Evidence for two Gla-independent Ca(2+)-binding sites in factor Xa.

The assembly of macromolecular complexes containing factors Xa and Va on suitable phospholipid surfaces is crucial for rapid activation of prothrombin. We have used quantitative affinity chromatography to characterize the interaction between factor Va and intact factor Xa on the one hand and between factor Va and factor Xa lacking the gamma-carboxyglutamic acid (Gla)-containing module on the other. The dissociation constants were found to be 1.0 +/- 0.1 and 9.5 +/- 1.8 microM, respectively. There was good agreement between these dissociation constants and the concentrations of active site-inhibited factor Xa and Gla-domainless factor Xa that caused half-maximal inhibition of prothrombin activation. To investigate whether the noncatalytic modules of factor Xa interacted directly with factor Va, intact modules were isolated from proteolytic digests of factor X and used as inhibitors of prothrombin activation. The inhibitory effect observed with the isolated Gla module in the absence of phospholipid was due to inhibition of the amidolytic activity of factor Xa rather than to an interaction with factor Va. The epidermal growth factor-like modules did not inhibit prothrombin activation. Using antibodies specific for calcium-dependent epitopes in the serine protease module of factor Xa we demonstrated that Ca2+ binding to the Gla module alters the conformation of the catalytic module. Half-maximal binding was observed at approximately 0.8 mM Ca2+. Evidence was also obtained for the presence of two Gla-independent Ca(2+)-binding sites in factor Xa. One of these sites, located in the NH2-terminal epidermal growth factor-like module, was half-saturated at approximately 60 microM Ca2+ in intact factor Xa and at approximately 1.2 mM Ca2+ in Gla-domainless factor Xa. This site appeared not to influence the conformation of the protease module. The second site, which was half-saturated at approximately 0.16 mM Ca2+, appeared to reside in the serine protease module and to alter its conformation as judged by binding of antibodies specific for calcium-dependent epitopes.

1-Carboxyglutamic Acid↗
PubMed1993

Antibodies against papillomavirus antigens in cervical secretions from condyloma patients.

Samples of cervical secretions and serum from 30 women with genital condylomas and 30 age-matched controls were tested for the presence of immunoglobulin A (IgA) and IgG antibodies against a panel of papillomavirus-derived antigens. The same cervical samples were also analyzed for presence of human papillomavirus (HPV) DNA by Southern blotting and polymerase chain reaction. By Southern blotting HPV DNA was detected in 8 of 30 patients with condylomas and 2 of 30 controls, and by the polymerase chain reaction HPV DNA was detected in 14 of 30 patients with condylomas and 5 of 30 controls. A total of 18 of 29 patients with condylomas and 8 of 28 controls had IgA antibodies in cervical secretions to an E2 synthetic peptide, and 17 of 29 patients with condylomas and 5 of 28 controls had local IgA antibodies to an E7 peptide (P < 0.025 and P < 0.005, respectively). The results suggest that measurement of local antibody production against selected HPV antigens may be useful in the study of HPV immunology and, possibly, for the diagnosis of HPV infection.

Adolescent↗