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Biomedical subjects

C Heilmann

Publications and source records attributed to C Heilmann.

At least 73 records · Page 4Linked to original sources

Characterization of the antibody response to a Haemophilus influenzae type b conjugate vaccine in children with recurrent lower respiratory tract infection.

Children with recurrent lower respiratory tract infection (RLRI) may respond poorly to polysaccharide antigens. To examine how such children respond to a polysaccharide coupled to a protein carrier, we immunized 15 children with RLRI aged 8-69 months and 15 carefully age-matched healthy controls once with a Haemophilus influenzae type b (Hib) conjugate vaccine. Total IgG subclasses, total antipolysaccharide Hib antibodies, and antipolysaccharide Hib antibodies of IgM, IgG, IgA, and IgG1-4 specificity were determined by ELISA. There were no significant differences between the two groups in any single total IgG subclass, but total IgG measured as the sum of all four subclasses was significantly lower in the children with RLRI than in the controls (P = 0.036). Before vaccination, the children with RLRI had significantly less IgG antipolysaccharide Hib antibody than the controls (P = 0.005), whereas 1 month later they had significantly more IgM antibody (P = 0.038). No other significant differences were found between the groups before or after immunization with respect to antipolysaccharide Hib antibodies. Since naturally occurring IgG antibodies are thought to be acquired partly as a consequence of antigenic stimulation on mucosal surfaces, we hypothesize that the low level of specific IgG found before immunization, as well as the low total IgG in the children with RLRI, may reflect an impaired ability to prime through mucosal surfaces. This is supported by our finding of an increased IgM response to Hib conjugate vaccine in these children, since this isotype predominates in the primary immune response, i.e., in the absence of immunologic memory.(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies↗

Transplantation of a lobe of lung from mother to child following previous transplantation with maternal bone marrow.

A left lower lobe of the lung was transplanted from a mother to her child, who had previously received a maternal bone marrow transplant for an immune defect. Following the bone marrow transplantation, the child had developed severe pulmonary fibrosis. Surgery and the early postoperative course have been uncomplicated. Immunosuppression with corticosteroids was administered for a short period, after which all immunosuppressive treatment was discontinued. The operation and the outcome are described both in the donor and recipient. Rehabilitation was slow, but one year later the patient is doing well.

Adult↗

A new type of Ca(2+)-dependent, Mg(2+)-stimulated ATPase of rat liver plasma membrane.

Incubation of a glycoprotein fraction obtained from rat liver plasma membrane which has been previously well characterized using [gamma-32P]ATP results in the phosphorylation of a 230-kDa glycoprotein (pgp230). It is composed of a 120-kDa subunit (pgp120) and a 110-kDa subunit (pgp110) linked by interchain disulfide bonds. Peptide maps of pgp120 and pgp110 suggest extensive similarity in their polypeptide chains. Glycan analysis reveals between four and six hybrid-type oligosaccharide chains for both phosphoproteins. Immunoblotting using monoclonal antibodies and endoglycosidase digestion exclude an identity of pgp120 or pgp110 with the hepatocyte plasma membrane glycoproteins dipeptidylpeptidase IV or the taurocholate transport protein, which co-purify and co-migrate in SDS/PAGE. Protein phosphorylation is Ca(2+)-dependent (K0.5(Ca2+) = 0.35 microM, in the absence of Mg2+). In the presence of Mg2+, the glycoprotein undergoes rapid cycles of phosphorylation and dephosphorylation, resulting in ATPase activity. Analysis of phosphorylated amino acids identifies phosphothreonine as the major one. Photoaffinity labeling with 8-azido-[alpha-32P]ATP demonstrates the presence of one or more ATP binding site(s). Preincubation of pgp230 with various purine or pyrimidine nucleotides (ATP, UTP, TTP, ADP, GDP, AMP, CMP) or known P2-purinoceptor agonists or antagonists (adenosine 5'-[alpha,beta-methylene]triphosphate, 2-methyl-thio-adenosine 5'-triphosphate, suramin) inhibits its phosphorylation by [gamma-32P]ATP. The biological function of pgp230 is unknown at present. Several findings of the present study are compatible with the idea that pgp230 may be involved in a P2-purinoceptor function of the hepatocyte. Following this concept, a mechanism is discussed where a cytosolically exposed high-affinity Ca(2+)-binding site of pgp230 would allow for receptor feedback control, via phosphorylation and dephosphorylation, by sensing changes in cytosolic Ca2+ concentration.

Adenosine Triphosphate↗

Comparative analysis of a biofilm-forming Staphylococcus epidermidis strain and its adhesion-positive, accumulation-negative mutant M7.

We have isolated a stable slime-negative mutant, M7, from the wild-type Staphylococcus epidermidis RP62A by mitomycin mutagenesis. Besides its inability to produce slime in the test tube this mutant differed also in two other properties from its parent strain: it lacked the ability to accumulate on a surface, and it did not produce a 115 kDa and a 18 kDa extracellular protein. In all other tested properties such as initial adherence, growth rate, cell-wall composition, surface characteristics, DNA restriction profile, the presence of a 29 kb antibiotic resistance plasmid, and antimicrobial susceptibility profile, M7 was indistinguishable from its wild-type. The mutant is an important basis for further study of the pathogenesis of polymer-associated S. epidermidis infections.

Bacterial Adhesion↗

Recent results of the joint ESA-DARA/IBMP experiments Biokosmos "Seeds".

Comparison of experimental data obtained from short (SDEF) and long duration exposure flights (LDEF) have recently led to results which will be significant for longer and/or repeated sojourn of man in space. Under orbital conditions biological stress and damage are induced in test subjects by cosmic radiation, especially the high energetic, densely ionizing component of heavy ions. Plant seeds were successful model systems for a biotest in studying the physiological damages and mutagenic effect caused by ionizing cosmic radiation in particular stem cells. Dosimetrically, the subdivision into charge- and Let-groups reveals the contribution of the intermediate group (LET = 350-1000 MeV/cm) due to the medium heavy ions (Z = 6-10). Their relative contribution increases with the lower inclination of the orbit of LDEF-1; on the other hand, the total fluence becomes higher with longer duration of the flight. The observed endpoints of the biological radiation damage hint at a correlation with particle dose rate rather than with the dose; additionally, data on shielding effects inside and outside the space craft and its exposure were gained from the different SDEF- and LDEF-missions.

Arabidopsis↗

Influence of a long duration exposure, 69 months, to the space flight factors in Artemia cysts, tobacco and rice seeds.

Three french laboratories have participated in the Free Flyer Biostack experiment. Artemia cysts, tobacco seeds and rice caryopsis and embryos were used. Biological objects in monolayers were dead. In opposite, a large fraction of samples used in bulk survived. A stimulatory effect occurred in the first steps of development in Artemia cysts. In fact, the larval survival was unchanged or slightly reduced. In tobacco a drastic decrease in germination and survival rate was observed. Space flight did not induce genetic changes. In rice, results depend on the variety which was investigated; the growth rate stimulation in flight samples is discussed with respect to controls.

Animals↗

Immuno-gold electron microscopical detection of heat shock protein 60 (hsp60) in mitochondria of rat hepatocytes and myocardiocytes.

We characterize the specificity of a polyclonal antibody against heat shock protein 60 (hsp60) and present an application for ultrastructural localization studies of this protein. The antibody was obtained from an IgG fraction (AB 121) originally raised against the calcium binding protein calsequestrin by immunoabsorption on isolated rat liver hsp60. As shown by partial N-terminal amino acid sequence analysis of immunoprecipitated proteins AB 121 contained reactivities against hsp60, calsequestrin and the glycoprotein fetuin. In rat heart AB 121 recognized calsequestrin and hsp60. In human and rat liver the only reacting protein was hsp60. In rat erythrocytes the antibody bound to 61 kDa and 58 kDa isoforms of fetuin. According to published data no amino acid sequence homologies nor common motifs are found between calsequestrin, hsp60 and fetuin. As the first application the anti-hsp60 antibody was used for immuno-gold electron microscopical localization of hsp60: in myocardiocytes and hepatocytes of the rat strong labelling was obtained exclusively in mitochondria. No extramitochondrial structures were labelled. The specificity of the antibody and its ability to be visualized by immuno-gold electron microscopy offers the possibility to study the expression of this protein in the liver and in other organs. Possible clinical applications of these studies are discussed, since hsp60 could be a target antigen of autoantibodies in diseases such as autoimmune hepatitis, primary sclerosing cholangitis or primary biliary cirrhosis.

Amino Acid Sequence↗

Opposite effects of actively and passively acquired immunity to the carrier on responses of human infants to a Haemophilus influenzae type b conjugate vaccine.

Vaccination of infants with Haemophilus influenzae type b (Hib) capsular polysaccharide (HibCP) coupled to carrier proteins has proven protective against invasive Hib diseases in several trials. However, insufficient immunogenicity has been noted in certain populations. Therefore, studies analyzing factors influencing the antibody response to conjugate vaccines are needed. In this study, the response to HibCP coupled to tetanus toxoid (TT) was examined in relation to (i) priming with or coadministration of the carrier protein and (ii) the levels of passively acquired maternal TT antibodies. One hundred forty-four infants were vaccinated with HibCP-TT at 5 and 6 months. They were randomized into three groups that received TT as part of a diphtheria-tetanus-polio vaccine at either 6 and 7 months (group A), 5 and 6 months (group B), or 4 and 5 months (group C). Maternally acquired TT antibodies inhibited the anti-HibCP response to the first HibCP-TT dose in groups A and B (r = -0.5 and -0.4, respectively; P < 0.005). In these groups, infants with prevaccination anti-TT levels above the median failed to reach the defined long-term protective level of HibCP antibodies (1 microgram/ml) more often than infants with low prevaccination levels after the first (P = 0.0001) and the second (P = 0.01) doses of HibCP-TT. In contrast, active priming with TT at 4 months resulted in a threefold-higher median level of anti-HibCP (group C; 1.34 micrograms/ml) than in the unprimed group (group A; 0.40 microgram/ml) after the first dose of HibCP-TT (P = 0.01). Coadministration of TT had no enhancing effect (group B; 0.58 microgram/ml). No significant differences between the median anti-HibCP levels were seen after the second HibCP-TT dose (6.72, 9.63, and 11.44 micrograms/ml in groups A, B, and C, respectively; P = 0.25).

Antibodies, Bacterial↗

Heavy-chain isotype patterns of human antibody-secreting cells induced by Haemophilus influenzae type b conjugate vaccines in relation to age and preimmunity.

The influence of preexisting immunity on the heavy-chain isotypes of circulating antibody-secreting cells (AbSC) induced by vaccination with Haemophilus influenzae type b (Hib) capsular polysaccharide (HibCP) coupled to tetanus toxoid (TT) or diphtheria toxoid (DT) and by vaccination with TT or DT alone in 51 healthy adults and 9 infants was studied. In adults, the isotypes of TT and DT AbSC were dominated by immunoglobulin G1 (IgG1) followed by IgG4 and IgA1. HibCP AbSC were dominated by the isotype IgA1 followed by (in decreasing order) IgG2, IgA2, IgM, and IgG1. The isotype distributions of TT and DT AbSC were independent of whether the toxoids were coupled to HibCP, and the isotypes of HibCP AbSC were not influenced by the nature of the carrier (TT or DT). Furthermore, the isotype distributions were unaffected by recent immunization with components of the conjugates, although this reduced the numbers of AbSC. The heavy-chain gene usage of HibCP AbSC in adults differed clearly from that in infants, which was restricted largely to the genes mu, gamma 1, and alpha 1, all lying upstream in the heavy-chain constant-region gene locus, while the usage in adults also, to different extents, involved the downstream genes gamma 2 and alpha 2. The ratio between the numbers of HibCP AbSC using heavy-chain genes from the downstream duplication unit (gamma 2, gamma 4, and alpha 2) and those using genes from the upstream duplication unit (gamma 3, gamma 1, and alpha 1) correlated with the preimmunization level of natural HibCP antibodies (r = 0.59; P = 0.00002). A possible role of natural exposure for Hib or cross-reactive bacteria on the mucosal surfaces in the shaping of the isotype response to HibCP conjugate vaccines is discussed.

Adult↗

Human liver calreticulin: characterization and Zn(2+)-dependent interaction with phenyl-sepharose.

A 60-kDa human calreticulin was isolated from liver homogenates. The protein was identified as calreticulin by its NH2-terminal amino acid sequence, by its mobility in SDS-PAGE, by its immunoreactivity with anti-calreticulin antibodies, by its Ca2+ binding, and by its localization to isolated ER membranes. In this study we show that Ca2+ binding to calreticulin results in Ca(2+)-dependent aggregation and precipitation of the protein. We also show that calreticulin and calsequestrin bind Zn2+ in 65Zn2+ overlay. In addition we have discovered that calreticulin exhibits a Zn(2+)-dependent interaction with hydrophobic matrix of phenyl-Sepharose that can be utilized in the purification of the protein.

Animals↗

Non-epitope-specific suppression of the antibody response to Haemophilus influenzae type b conjugate vaccines by preimmunization with vaccine components.

Recently, conjugate vaccines containing Haemophilus influenzae type b capsular polysaccharide (HibCP) coupled to protein carriers were introduced for use in infants and certain adult risk groups. Similar conjugate vaccines against other capsulated bacteria are currently under development for both children and adults. Despite its potential importance, the possible influence of preexisting immunity to the components of such conjugates on the vaccination response in humans has been addressed by few studies. To study this issue, we randomized 82 healthy adult volunteers into six groups and vaccinated them twice, with a 4-week interval between immunizations. Four groups received tetanus toxoid (TT) or diphtheria toxoid (DT) and then HibCP coupled to TT (HibCP-TT) or DT (HibCP-DT). Two groups received HibCP-TT followed by HibCP-DT or vice versa. The total antibody levels to HibCP, TT, and DT and the anti-HibCP immunoglobulin G1 (IgG1) and IgG2 levels were measured before and 4 weeks after the immunizations. For some of the vaccinees, the number of circulating antibody-secreting cells was evaluated 7 days after immunization. Surprisingly, preimmunization with the relevant carrier protein reduced the subsequent increase in the total HibCP antibody levels (P < 0.05), affecting the IgG1 and the IgG2 subclasses equally. Also, the responses to the carrier portions of the conjugates were suppressed, as demonstrated by the lack of significant increases in the antibody levels (P > or = 0.29) and, for HibCP-TT, by reduced numbers of anticarrier antibody-secreting cells (P = 0.009). Similar non-epitope-specific suppression was seen in the groups receiving both conjugates. Thus, preimmunization with one conjugate reduced the subsequent response to the carrier portion of the other conjugate (HibCP-DT and then HibCP-TT, P = 0.00002; HibCP-TT and then HibCP-DT, P = 0.06) as well as to HibCP itself. Possible mechanisms behind this non-epitope-specific suppression and its relevance for vaccine development are discussed.

Adult↗

[The bactericidal effect of oxybuprocaine. A possible cause of false negative results in bronchoalveolar lavage].

New approaches in the diagnosis of pneumonia, especially in intensive care units, are quantitative cultures of bronchoalveolar (BAL) fluid or the protected specimen brush. The sensitivity of these methods, however, has often been found to be as low as 50-60%. One possible explanation for the low sensitivity of these diagnostic tools is the antimicrobial activity of local anaesthetics used in bronchoscopy. Therefore, we investigated the bactericidal properties of oxybuprocaine, a topical anaesthetic used for bronchoscopy in our clinic, in order to test the reliability of specimens obtained from BAL. METHODS. The bactericidal activity of oxybuprocaine in concentrations of 1%, 0.1%, 0.05% and 0.01% was tested by constructing time-kill curves for Streptococcus pneumoniae, Hemophilus influenzae, Pseudomonas aeruginosa and Escherichia coli. Five stains of each bacterial species were tested. The inoculum size was 10(4) ml, and bacteria were counted after 10, 20, 30, 60 and 120 min. RESULTS. The resulting time-kill curves are demonstrated in Figs. 1-4. The most sensitive bacteria were S. pneumoniae and H. influenzae, in which significant bactericidal activity could be shown even with a 0.01% solution of oxybuprocaine. E. coli and P. aeruginosa were also inhibited, but only at the highest concentration of 1%. CONCLUSIONS. The use of local anaesthetics before material is taken for culture, e.g. from BAL, may give rise to false-negative results and should therefore be avoided or reduced. For each local anaesthetic used in bronchoscopy, the concentrations that can be used without the risk of false-negative results should be determined.

Anesthetics, Local↗

A simplification of the enzyme-linked immunospot technique. Increased sensitivity for cells secreting IgG antibodies to Haemophilus influenzae type b capsular polysaccharide.

A simplified enzyme-linked immunospot (ELISPOT) technique is described for the detection of cells secreting antibodies to tetanus toxoid (TT), diphtheria toxoid (DT) or Haemophilus influenzae type b capsular polysaccharide (PRP). By combining the cell suspension with the enzyme-linked secondary antibodies in one incubation, the second incubation and washing procedure could be omitted from the original technique. The simplified assay had the same sensitivity for anti-TT and anti-DT spot-forming cells as the ordinary ELISPOT assay. The IgG anti-PRP spots were, however, improved both in quality and in quantity (median: 40% more spots), while the detection of IgM and IgA anti-PRP spot-forming cells was the same in the two techniques. This simplified technique can probably also be used to save time in other antigen systems and should be considered when designing ELISPOT assays for the detection of polysaccharide-specific antibody-secreting cells.

Antibodies, Bacterial↗

An improved haemolytic plaque assay for the detection of cells secreting antibody to bacterial antigens.

Recent advances in the development of conjugate polysaccharide vaccines for human use have stimulated interest in the use of assays detecting antibody-secreting cells (AbSC) with specificity for bacterial antigens. Here we present improved haemolytic plaque-forming cell (PFC) assays detecting AbSC with specificity for tetanus and diphtheria toxoid as well as for Haemophilus influenzae type b and pneumococcal capsular polysaccharides. These assays were found to be less time consuming, more economical and yielded 1.9-3.4-fold higher plaque numbers than traditional Jerne-type PFC assays. In the case of anti-polysaccharide AbSC of the IgG isotype, the increase was as high as 7.4-11.8 times. Evidence is presented that the pronounced improvement in the detection of the latter is due to the presence of aggregating anti-IgG antibody from the beginning of the assay. It is proposed that in the case of low affinity of anti-polysaccharide antibodies aggregation of secreted monomeric antibody (IgG) is critical for plaque formation and increases the avidity of binding to target cells.

Antibodies↗

Combined deficiencies of the pyruvate dehydrogenase complex and enzymes of the respiratory chain in mitochondrial myopathies.

In six patients with mitochondrial (encephalo-) myopathy investigations of skeletal muscle revealed a defect of pyruvate dehydrogenase complex (PDHC) in combination with one or more respiratory chain complex deficiencies. A combination of defects of this kind has not been reported previously. Five of the six patients presented within the 1st year of life and had a severe clinical course. Intrafamilial variability of the clinical course in dizygotic twins both suffering from a cytochrome c oxidase deficiency and one of them also from a PDHC deficiency suggests an additional effect of PDHC deficiency on the clinical symptoms. Immunoblot studies of PDHC in five of the patients revealed no abnormalities in their subunit pattern, rendering a defect of mitochondrial protein import or assembly unlikely. The finding of a combined PDHC and respiratory chain deficiency has implications for the diagnostic approach, for therapy and genetic counselling. The exact pathogenetic mechanism of this combination of defects remains to be elucidated.

Female↗

Restriction fragment length polymorphism of two HLA-B-associated transcripts genes in five autoimmune diseases.

The restriction fragment length polymorphism of the two human HLA-B-associated transcripts (BATs) genes, BAT1 and BAT2, identifying polymorphic bands of 12, 8, 2.5, and 1.1 kb, and at 3.3, 2.7, 2.3, and 0.9 kb, respectively, was investigated in patients with primary biliary cirrhosis (PBC), systemic lupus erythematosus (SLE), pauciarticular juvenile rheumatoid arthritis (P-JRA), rheumatoid arthritis (RA), and primary Sjögren's syndrome (pSS), and in healthy Danes. The BAT2/RsaI 2.7-kb band fragment was more frequent in PBC, pSS, and SLE than in controls, but the p values did not reach significance when corrected for multiple comparisons. For pSS and SLE, the associations may be secondary to primary associations with HLA-B8 because the BAT2/RsaI 2.3-kb band, which is allelic to the BAT2/RsaI 2.7-kb band, is strongly negatively associated with HLA-B8 and HLA-DR3. The only significance obtained shows that the HLA-B8 frequency is increased in BAT2/RsaI 2.7-kb positive pSS patients as compared to the corresponding controls indicating that the HLA-B8 association may be strongest. No missing or extra DNA fragments were observed in the disease groups when compared with controls indicating that gross deletions or duplications of the BAT1 and BAT2 genes in the patients are unlikely. In conclusions, it cannot be excluded that the BAT2/RsaI 2.7-kb band may contribute to the susceptibility to PBC, pSS, and SLE.

Autoimmune Diseases↗