Biomedical subjects

S Köhler

Publications and source records attributed to S Köhler.

At least 73 records · Page 4Linked to original sources

PubMed1994

Live Brucella spp. fail to induce tumor necrosis factor alpha excretion upon infection of U937-derived phagocytes.

Tumor necrosis factor alpha (TNF-alpha) plays a central role in activation of first-line defenses of a host against foreign organisms. To determine whether Brucella infection modulated TNF-alpha production, we measured the biological activity of this cytokine in supernatants of U937 cell-derived macrophages and of fresh human monocytes infected with Brucella spp. Neither the smooth nor rough Brucella strains used induced any measurable TNF-alpha excretion upon infection. On the contrary, as reported before for other gram-negative bacteria, phagocytosis of nonpathogenic Escherichia coli was followed by a rapid and transient induction of TNF-alpha release, suggesting an involvement of this cytokine in some autocrine process. As expected, the Brucella strains tested survived and/or multiplied within U937-derived macrophages, whereas E. coli was rapidly eliminated after phagocytosis. Immunoglobulin G opsonization of E. coli strains enhanced their intracellular killing and strongly potentiated TNF-alpha secretion. Immunoglobulin G opsonization of Brucella strains, in contrast, did not lead to TNF-alpha production, although their rate of intracellular multiplication was reduced. Killed brucellae, however, promoted a significant excretion of TNF-alpha from U937-derived macrophages into cell culture supernatants. We finally demonstrated that pretreatment of U937-derived macrophages with exogenous TNF-alpha significantly inhibited intracellular multiplication of Brucella spp. These results and experiments performed on fresh human monocytes or with isolated lipopolysaccharide (LPS) showed that (i) differences in TNF-alpha production observed during macrophage infection by Brucella spp. and E. coli were not due to differences in LPS structure but resulted from active inhibition of TNF-alpha production by a specific process linked to Brucella spp. and (ii) the capacity of Brucella spp. to use pathways avoiding TNF-alpha production during infection may be considered a major attribute of virulence.

Brucella↗

PubMed1994

[Psychosocial disintegration].

Among the patients referred for rehabilitation in the latter half of their working life, many are notable due to considerable discrepancies between their objectively ascertainable performance and its subjectively perceived decline. In these cases, the "substantial threat to earning capacity" cannot be explained by measurable organ deficiencies. Similarly, treatment efforts focussed solely at improved somatic functioning remain inefficient in terms of stabilization of earning capacity, because they do not bring about changes in the cause of subjective performance deterioration. The author in these circumstances assumes the presence of an independent syndrome, called "psychosocial disintegration". He describes the full picture of this disease entity, and suggests causal mechanisms as well as potential for remedial intervention. On account of the considerable social dimension of the disorder outlined, early identification of these gradually developing changes as well as qualified care of the insurants are indispensable. All those involved in treatment and care of the patients or working in some branch of the social security system should be familiar with this psychosocial disintegration syndrome in order to avoid the guidance and counselling mistakes that are frequently the case. As rehabilitation is impossible in case of inhibiting personal attitudes of an insurant, it is advisable to verify the individual's readiness for rehabilitation and/or to strengthen it by appropriate measures before engaging in costly in-patient service provision. If the needed motivation is to be achieved during participation in a rehabilitation measure, extended service provision will invariably be required.

Disability Evaluation↗

PubMed1994

[Development of wisdom teeth as a criterion of age determination].

The study is based on the orthopantomographic evaluation of the formation and the development of the permanent third molar (n = 938 patients with known age between 15 and 25 years). We used a modified method according to Gleiser and Hunt (1955) with 3 stages of crown formation and 7 stages of root formation. The correlation between age and the formation of the third molar is not very close. There is a deviation of +/- 4 years (97% statistical safety). The root is completed at the age of about 23 years (upper jaw: 23 years in males and females, mandible: males 23.2 years, females 23.6 years). There is a slight sexual dimorphism with an accelerated development of the third molar in males. The dentition of the molars runs parallel on both sides of the jaw. Only a small proportion (14%) of all persons has completely formed roots at the age of 21. Retination of the third molars (produced by the second molar and by the ascending branch of the mandible) causes a delay of its development.

Adolescent↗

PubMed1994

[STECO titanium magnets. A new magnetic attachment].

A newly developed telescopic magnet (the Steco titanium magnet) is presented and can guarantee a stable position of the epithesis, as well as still allow a vertical displacement up to 3 mm and at the same time permit a controllable rotation. In combination with other flat magnets (with spherical surfaces) even in unfavorable disparallel positions, a stable, base-adapted epithesial position is attainable and a permanent shaft secured. The Steco magnet is a valuable alternative to other retention mechanisms now available.

Ear, External↗

PubMed1993

A family of transcripts (K2) of Entamoeba histolytica contains polymorphic repetitive regions with highly conserved elements.

Sera from patients with invasive amebiasis were used to identify a cDNA clone (K2p-1) encoding a commonly recognized, repeat-containing antigen of the pathogenic Entamoeba histolytica HM-1:IMSS. K2p-1 was used to isolate 3 cDNA clones (K2 clones); one K2p-1 related clone from the same pathogenic E. histolytica strain and 2 from the nonpathogenic E. histolytica strain SAW-142. The nucleotide sequence and predicted amino acid sequence revealed a closely related family of transcripts differing mainly in the extent and arrangement of an internal region consisting of tandemly arranged repetitive elements. The repetitive units encoding either 12 or 8 amino acids were found to be highly conserved in all the K2 clones analyzed so far, suggesting that the repeat motifs perform functions common to both pathogenic and nonpathogenic E. histolytica. The genomic organization of the K2 genes was different when compared in pathogenic and nonpathogenic E. histolytica and may therefore be used to discriminate between pathogenic and nonpathogenic E. histolytica strains.

Amino Acid Sequence↗

PubMed1993

Listeria monocytogenes--a model system for studying the pathomechanisms of an intracellular microorganism.

Virulence of Listeria monocytogenes is determined by a cluster of five genes in the order plcA, hly, mpl, actA and plcB, which are coordinately regulated by a transcriptional activator, termed PrfA. The gene for PrfA is located in front of plcA. Mutations within each of these genes reduce the virulence considerably and render the mutants unable to properly multiply and/or spread within the infected host cells. Under growth-limiting conditions PrfA-dependent proteins are preferentially synthesised. These studies indicate the existence of additional PrfA-regulated proteins in L. monocytogenes. The synthesis of catalase, superoxide dismutase, LmaA and p60 is not under the control of PrfA. These proteins seem to be also associated with virulence of L. monocytogenes. P60-related proteins are found as major extracellular proteins in all Listeria species but only p60 of L. monocytogenes is able to restore the failure of R-mutants (exhibiting a drastically reduced synthesis of p60) to adhere to 3T6 mouse fibroblasts. Adherence of L. monocytogenes to the epithelial Caco-2 cells seem to be independent of p60. The p60 protein of L. monocytogenes differs characteristically from the p60-related proteins of the nonvirulent Listeria species.

Amino Acid Sequence↗

PubMed1993

The iap gene of Listeria monocytogenes is essential for cell viability, and its gene product, p60, has bacteriolytic activity.

Expression of the iap gene of Listeria monocytogenes in the L. monocytogenes rough mutant RIII and in Bacillus subtilis DB104 caused the disruption of the cell chains which these two strains normally form under exponential growth conditions. The p60 protein produced by L. monocytogenes and B. subtilis DB104 also exhibited bacteriolytic activity detected in denaturing polyacrylamide gels containing heat-killed Micrococcus lysodeikticus. Purification of the p60 protein led to aggregation of p60 and loss of the cell chain disruption and bacteriolytic activities. A cysteine residue in the C-terminal part of p60 which is conserved in all p60-like proteins from the other Listeria species seems to be essential for both activities. The iap gene could not be inactivated without a loss of cell viability, indicating that p60 is an essential housekeeping protein for L. monocytogenes and probably also for other Listeria species. These data suggest that p60 possesses a murein hydrolase activity required for a late step in cell division.

Amino Acid Sequence↗

PubMed1992

The homologous and heterologous regions within the iap gene allow genus- and species-specific identification of Listeria spp. by polymerase chain reaction.

The iap gene of Listeria species encodes protein p60. The comparison of iap-related genes from different Listeria species indicated common and variable regions within these genes which appeared to be specific for each Listeria species. On the basis of the iap gene sequences, pairs of polymerase chain reaction (PCR) primers which allowed the unambiguous identification of all members of the genus Listeria, of groups of related Listeria species, and of L. monocytogenes, exclusively, were selected. The PCR primers specific for L. monocytogenes yielded PCR products which represented essentially the repeat region of the iap gene. The size of these PCR products allowed an estimate of the number of the TN repeat units within the repeat region of the p60 protein of an L. monocytogenes strain. The data indicated that the number of repeat units differed among L. monocytogenes isolates.

Bacterial Proteins↗

PubMed1992

Identification of p60 antibodies in human sera and presentation of this listerial antigen on the surface of attenuated salmonellae by the HlyB-HlyD secretion system.

Antibodies directed against the major secreted protein of Listeria monocytogenes, termed p60, were found more frequently than antilisteriolysin antibodies in sera of listeriosis patients. Anti-p60 antibodies were also identified in all tested sera from healthy individuals. To test whether p60 provides protection against L. monocytogenes, we constructed an attenuated Salmonella typhimurium aroA strain which secretes p60 via the Escherichia coli hemolysin secretion pathway. Application of this Salmonella strain to BALB/c mice prior to an L. monocytogenes infection induced p60 antibodies in these mice and led to a significantly reduced number of viable bacteria in the spleen compared with that in control animals which were primed with the S. typhimurium aroA strain alone.

Animals↗

PubMed1992

Structural and functional properties of the p60 proteins from different Listeria species.

The major extracellular protein p60 of Listeria monocytogenes seems to be required for this microorganism's adherence to and invasion of 3T6 mouse fibroblasts but not for adherence to human epithelial Caco-2 cells. Western blot analysis with polyclonal antibodies against p60 of L. monocytogenes indicated the presence of cross-reacting proteins in the culture supernatants of all Listeria species. Protein p60 of L. monocytogenes could restore adhesion of the L. monocytogenes mutant RIII (impaired in the synthesis of p60) to mouse fibroblasts more efficiently than that of Listeria grayi. The amino acid sequences of the p60-related proteins of L. innocua, L. ivanovii, L. seeligeri, L. welshimeri, and L. grayi indicated highly conserved regions of about 120 amino acids at both the N-terminal and the C-terminal ends. The middle portions of these proteins, consisting of about 240 amino acids, varied considerably. These parts include the repeat domain consisting of repetitions of Thr (T) and Asn (N) which was present only, albeit in different arrangements, in the p60 proteins of L. monocytogenes and L. innocua. The p60-related proteins of L. grayi, L. ivanovii, L. seeligeri, and L. welshimeri each contained an insertion of 54 amino acids which was absent in the p60 proteins of L. monocytogenes and L. innocua.

Amino Acid Sequence↗

PubMed1991

Gene disruption by plasmid integration in Listeria monocytogenes: insertional inactivation of the listeriolysin determinant lisA.

A plasmid integration technique was developed for insertional inactivation of chromosomal Listeria monocytogenes genes. A Listeria-Escherichia coli shuttle vector (pLSV1) was constructed which carried the temperature-sensitive gram-positive replication origin from plasmid pTV32(Ts). An internal fragment of the listeriolysin gene (lisA) was cloned into pLSV1 to create pLSV2. In L. monocytogenes pLSV2 transformants, plasmid pLSV2 integrated into the L. monocytogenes chromosome at a frequency of 2 x 10(-3) via lisA homology and these cells could be selected at 42 degrees C using a plasmid-encoded erythromycin resistance. Plasmid integration resulted in disruption of the lisA gene, production of a truncated, immunologically cross-reactive listeriolysin protein and loss of the hemolytic phenotype. An improved Listeria protoplast transformation method is also described which facilitates genetic manipulation of Listeria species.

Bacterial Toxins↗

PubMed1991

Studies on the pathogenicity of Listeria monocytogenes.

The characterization of mutants of Listeria monocytogenes with reduced virulence properties is described. Reduction in the amount of the extracellular protein p60 (encoded by the ipa gene) leads to cell filaments with impaired invasiveness. Mutants which cannot synthesize listeriolysin are still invasive but unable to survive within phagocytic cells. One type of listeriolysin-negative mutants is defective in the synthesis of a positive regulatory element PrfA which coordinately regulates the listeriolysin gene (lisA) together with several other genes, including those for a phosphatidylinositol-specific phospholipase and a metalloprotease.

Antigens, Bacterial↗

PubMed1991

Expression of the iap gene coding for protein p60 of Listeria monocytogenes is controlled on the posttranscriptional level.

Expression of the iap gene of Listeria monocytogenes encoding a major extracellular protein (p60) was analyzed. Different start sites for transcription of the iap gene were identified by primer extension analysis in L. monocytogenes and in a recombinant Escherichia coli clone. The mutant RIII of L. monocytogenes represents a member of the frequently occurring L. monocytogenes R mutants, which form cell chains and produce greatly reduced amounts of p60. However, the concentrations of iap-specific mRNA were similar in mutant RIII and the wild-type strain. The introduction of additional copies of the iap gene from wild-type L. monocytogenes led to an equal increase of iap mRNA in both strains, but overexpression of protein p60 was only observed in the wild-type strain. The nucleotide sequences of both iap genes and their 5' noncoding regions were identical in all parts that are essential for efficient transcription of the iap gene, translation of the iap-specific mRNA, and transport of the p60 protein. These data suggest that the expression of the iap gene in L. monocytogenes is controlled on the posttranscriptional level by a specific factor that is defective in mutant RIII.

Amino Acid Sequence↗

PubMed1991

[The dynamics of bone apposition in different animal species--a comparative study].

The time of the bone apposition was compared in rats, rabbits and guinea pigs with the aid of enzyme-activity measurements, determination of the proportion of minerals and the judgment of the cell generation. For this intention a defect was made in the distal femur epiphyses at the three species. The animals were observed for a term of 15 weeks. The results showed little differences with regard to the speed of the bone apposition in the three called species. The bone metabolism of rats was higher than those of the other ones. A clear reference value for the enzyme activity to a define point of time of the mineralization could not be ascertained.

Alkaline Phosphatase↗

PubMed1990

Distinct binding sites of Ala48-hirudin1-47 and Ala48-hirudin48-65 on alpha-thrombin.

The interaction of alpha-thrombin with Ala48-hirudin, Ala48-hirudin1-47, and Ala48-hirudin48-65 was analyzed. Mutations at Pro48 were found to cause only slight changes in the kon (human: 3.1 +/- 0.3 x 10(8) M-1 s-1; bovine: 1.03 +/- 0.3 x 10(8) M-1 s-1) and koff (human: 0.4 +/- 0.2 x 10(-3) s-1; bovine: 2.9 +/- 0.4 x 10(-3) s-1) rate constants for the formation of the thrombin-hirudin complex. The amino-terminal fragment Ala48-hirudin1-47 containing the three disulfide bridges and the carboxyl-terminal fragment Ala48-hirudin48-65 were derived from the Ala48 mutant by proteolysis with endoproteinase Lys-C. These fragments inhibit bovine alpha-thrombin clotting activity with IC50 values of 0.6 and 4.9 microM, respectively (2.4 nM for r-hirudin). By mapping the interaction of Ala48-hirudin-derived fragments with bovine alpha-thrombin by limited proteolysis with trypsin and pancreatic elastase distinct binding sites for each fragment were determined. The carboxyl-terminal fragment was found to bind to the proposed anion-binding exosite in the region B62-74, whereas the amino-terminal fragment binds to a region around the elastase cleavage site at residues 150-151 of the alpha-thrombin B-chain.

Alanine↗

PubMed1990

The gene coding for protein p60 of Listeria monocytogenes and its use as a specific probe for Listeria monocytogenes.

The gene of Listeria monocytogenes that encodes a major extracellular protein (p60) was cloned in Escherichia coli. The gene was designated iap, as p60 was previously shown to represent an invasion-associated protein (M. Kuhn and W. Goebel, Infect. Immun. 57:55-61, 1989). The recombinant E. coli clone expressed p60, as shown by immunoblotting. The complete nucleotide sequence of iap was determined. The deduced amino acid sequence of p60 (484 amino acids) contains a putative N-terminal signal sequence of 27 amino acids and an extended repeat region consisting of 19 threonine-asparagine units. Hybridization with the entire iap gene revealed the presence of homologous sequences in most other Listeria species. In contrast, a 400-base-pair internal iap probe which contained the whole repeat region hybridized only with genomic DNA from L. monocytogenes. Four oligonucleotides previously described as specific probes for the detection of L. monocytogenes (A. R. Datta, B. A. Wentz, D. Shook, and M. W. Trucksess, Appl. Environ. Microbiol. 54:2933-2937, 1988) were shown to be part of the iap gene.

Amino Acid Sequence↗

PubMed1990

Ovarian response and pregnancy rates in in vitro fertilization, gamete intrafallopian transfer, and in vivo fertilization therapies after combined gonadotropin-releasing hormone agonist/human menopausal gonadotropin stimulation.

Ovarian stimulation cycles were initiated using human menopausal gonadotropin (HMG) in 318 women (643 cycles) without pretreatment and in 341 women (525 cycles) after pituitary desensitization by pretreating with the gonadotropin-releasing hormone agonists (GnRH-A) buserelin acetate or triptorelin acetate. Significantly higher pregnancy rates were observed in the GnRH-A/HMG group (15%) as compared to the HMG group (7%) following in vitro (p less than 0.05) but not in vivo fertilization therapy (14 vs. 9%, respectively). After in vitro fertilization, the rates increased with increasing length of the active phase of follicular maturation. Gamete intrafallopian transfer, performed only in the GnRH-A/HMG group, led to a pregnancy rate of 34%. Overall, there was a clear trend to higher pregnancy rates in the GnRH-A/HMG group (16%) as compared to the HMG group (8%). Abortion rates were comparable in both groups (24 vs. 19%). The higher pregnancy rates in the GnRH-A/HMG group were attributable to enhanced follicular maturation and optimized ovarian stimulation produced by the hypogonadotropic state. However, an increased risk of the ovarian hyperstimulation syndrome was observed in these patients.

Adult↗

PubMed1989

Solution structure of recombinant hirudin and the Lys-47----Glu mutant: a nuclear magnetic resonance and hybrid distance geometry-dynamical simulated annealing study.

The solution structure of recombinant wild-type hirudin and of the putative active site mutant Lys-47----Glu has been investigated by nuclear magnetic resonance (NMR) spectroscopy at 600 MHz. The 1H NMR spectra of the two hirudin variants are assigned in a sequential manner with a combination of two-dimensional NMR techniques. Some assignments made in our previous paper [Sukumaran, D. K., Clore, G. M., Preuss, A., Zarbock, J., & Gronenborn, A. M. (1987) Biochemistry 26, 333-338] were found to be incorrect and are now corrected. Analysis of the NOE data indicates that hirudin consists of an N-terminal compact domain (residues 1-49) held together by three disulfide linkages and a disordered C-terminal tail (residues 50-65) which does not fold back on the rest of the protein. This last observation corrects conclusions drawn by us previously on hirudin extracted from its natural source, the leech Hirudo medicinalis. The improved sensitivity of the 600-MHz spectrometer relative to that of our old 500-MHz spectrometer, the availability of two variants with slightly different chemical shifts, and the additional information arising from stereospecific assignments of methylene beta-protons and methyl protons of valine have permitted the determination of the solution structure of hirudin with much greater precision than before. Structure calculations on the N-terminal domain using the hybrid distance geometry-dynamical simulated annealing method were based on 685 and 661 approximate interproton distance restraints derived from nuclear Overhauser enhancement (NOE) data for the wild-type and mutant hirudin, respectively, together with 16 distance restraints for 8 backbone hydrogen bonds identified on the basis of NOE and amide NH exchange data and 26 phi backbone and 18 chi 1 side-chain torsion angle restraints derived from NOE and three-bond coupling constant data. A total of 32 structures were computed for both the wild-type and mutant hirudin. The structure of residues 2-30 and 37-48 which form the core of the N-terminal domain is well determined in both cases with an average atomic rms difference between the individual structures and the respective mean structures of approximately 0.7 A for the backbone atoms and approximately 1 A for all atoms. As found previously, the orientation of the exposed finger of antiparallel beta-sheet (residues 31-36) with respect to the core could not be determined on the basis of the present data due to the absence of any long-range NOEs between the exposed finger and the core.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗