Biomedical subjects

B Hubner

Publications and source records attributed to B Hubner.

7 recordsLinked to original sources

PubMed2001

Increase in calcitonin gene related peptide (CGRP) and decrease in mast cells in dihydroepiandrosterone (DHEA)-induced polycystic rat ovaries.

UNLABELLED: The polycystic ovary is reported to correspond with a high density in intraovarian nerve fibers and their sympathetic hyperresponsiveness. Peptidergic nerves may also be involved in this process. An interaction between nerve fibers and mast cells is assumed because of nerve growth-factor production by mast cells. Here we investigated CGRP-positive nerve fibers and mast cells in polycystic ovaries induced in immature rats with dihydroepiandrosterone (DHEA). The DHEA treated ovaries contained less corpora lutea than controls (mean +/- SEM: 4.3 +/- 0.6 versus 11.3 +/- 0.9, P > 0.001) and less intact antral follicles (4.7 +/- 0.7 versus 8.1 +/- 1.1; P < 0.05) according to the histometric approach. By immunolabelling more CGRP-positive nerve fibers were found in the DHEA treated ovaries than in controls (mean +/- SEM per one section: 23.2 +/- 5.8 fibers versus 10.3 +/- 0.9 and 171 +/- 44.7 varicosities versus 84 +/- 9.5). This was confirmed by dot blot analysis, showing a significant higher CGRP signal intensity per microgram homogenized ovaries of the DHEA treated group compared to the untreated (P < 0.05). Toluidine-blue-stained mast cells populated the medulla in both groups, yet had strikingly decreased in the DHEA treated ovaries (23.5 +/- 3.9 versus 89 +/- 5.6, P < 0.005). CONCLUSION: The increase in CGRP-positive nerve fibers and the decrease of toluidine-blue-stained mast cells points to an altered neuroimmune function in DHEA-induced polycystic rat ovaries.

Animals↗

PubMed2000

High incidence of parvovirus B19 DNA in synovial tissue of patients with undifferentiated mono- and oligoarthritis.

A common problem in rheumatological practice is inflammatory joint disease that cannot be classified. The prognosis of such undifferentiated arthritides is uncertain. The synovial tissue of 41 consecutive patients with various forms of arthritis was tested for the presence of viral DNA in a diagnosis-unaware fashion, using the polymerase chain reaction (PCR). Of all tested viruses, cytomegalovirus and parvovirus B19 were positive (each in 10 patients, two double-positives), whereas herpes simplex virus was positive in two patients. Rubella virus RNA was detected in three specimens. When the positivity for viral material was analysed in terms of distribution among the various diagnostic groups, it became evident that five out of 10 parvovirus B 19-positive patients belonged to the undifferentiated arthritis group, whereas cytomegalovirus-positive patients were spread among all diagnostic groups. This indicates the possibility of a new diagnostic category of undifferentiated mono- and oligoarthritis, which can be identified by the presence of parvovirus B19 DNA in synovial tissue.

Adolescent↗

PubMed2000

Detection of multiple viral DNA species in synovial tissue and fluid of patients with early arthritis.

OBJECTIVE: Viruses have a role in the pathogenesis of various forms of arthritis. This study aimed at determining whether viral DNA can be detected in joint samples in the early stages of idiopathic arthritides. METHODS: Synovial fluid (SF) and synovial tissue (ST) samples were obtained from 73 patients, with undifferentiated arthritis (n=22), rheumatoid arthritis (n=13), spondyloarthropathy (n=17), crystal arthropathy (n=8), osteoarthritis (n=7), septic arthritis (n=5), and trauma (n=1). The presence of viral DNA was investigated by polymerase chain reaction analysis. RESULTS: Cytomegalovirus was present in 25 patients, parvovirus B19 in 15 patients, Epstein-Barr virus in 12 patients, and herpes simplex virus in 16 patients (in ST, SF, or both), respectively. The joint samples were negative for viral DNA from adenovirus and varicella-zoster virus. In ST, eight patients were double positive for parvovirus B19 and another viral DNA, with herpes simplex virus being the most prevalent. Seven patients were double positive for other viruses (cytomegalovirus, herpes simplex virus, Epstein-Barr virus). In SF, four patients were double or triple positive for viral DNA. Paired samples were available in 56 patients. In these, viral DNA was detected in 37 patients in ST, as compared with 19 in SF. CONCLUSION: These data show that one or more viruses can be detected in the synovial specimens of patients with early arthritis, irrespective of the clinical diagnosis. This observation might be explained by migration of inflammatory cells harbouring viral DNA into the inflamed joints.

Arthritis↗

PubMed1999

Structural analysis of an RNase T1 variant with an altered guanine binding segment.

The ribonuclease T1 variant 9/5 with a guanine recognition segment, altered from the wild-type amino acid sequence 41-KYNNYE-46 to 41-EFRNWQ-46, has been cocrystallised with the specific inhibitor 2'-GMP. The crystal structure has been refined to a crystallographic R factor of 0.198 at 2.3 A resolution. Despite a size reduction of the binding pocket, pushing the inhibitor outside by 1 A, 2'-GMP is fixed to the primary recognition site due to increased aromatic stacking interactions. The phosphate group of 2'-GMP is located about 4.2 A apart from its position in wild-type ribonuclease T1-2'-GMP complexes, allowing a Ca(2+), coordinating this phosphate group, to enter the binding pocket. The crystallographic data can be aligned with the kinetic characterisation of the variant, showing a reduction of both, guanine affinity and turnover rate. The presence of Ca(2+) was shown to inhibit variant 9/5 and wild-type enzyme to nearly the same extent.

Amino Acid Sequence↗

PubMed1999

Modification of ribonuclease T1 specificity by random mutagenesis of the substrate binding segment.

Attempts to modify the guanine specificity of ribonuclease T1 (RNase T1) by rationally designed amino acid substitutions failed so far. Therefore, we applied a semirational approach by randomizing the guanine binding site. A combinatorial library of approximately 1.6 million RNase T1 variants containing permutations of 6 amino acid positions within the recognition loop was screened on RNase indicator plates. The specificity profiles of 180 individual clones showing RNase activity revealed that variant K41S/N43W/N44H/Y45A/E46D (RNaseT1-8/3) exhibits an altered preference toward purine nucleotides. The ApC/GpC preference in the cleavage reaction of this variant was increased 4000-fold compared to wild-type. Synthesis experiments of dinucleoside monophosphates from cytidine and the corresponding 2'3'-cyclic diesters using the reverse reaction of the transesterification step showed a 7-fold higher ApC synthesis rate of RNase 8/3 than wild-type, whereas the GpC synthesis rates for both enzymes were comparable. This study shows that site-directed random mutagenesis is a powerful additional tool in protein design in order to achieve new enzymatic specificities.

Amino Acid Sequence↗

PubMed1995

Destabilization of a protein helix by electrostatic interactions.

Electrostatic interactions between charged residues and the helix dipole in a protein were investigated by protein engineering methods. In ribonuclease T1, two surface-exposed acidic residues (Glu28 and Asp29) are located near the carboxyl terminus of the alpha-helix between residues 13 and 29. They were replaced, individually and in concert, by the uncharged amides Gln28 and Asn29, and the stabilities of the wild-type protein and its variants were determined as a function of pH. The effects of the two mutations are additive. Either one leads to a marginal destabilization by 0.7 kJ/mol at pH 2 but to a strong stabilization by about 3.2 kJ/mol at pH 7. This suggests that the deprotonations of Glu28 and Asp29 reduce the free energy of stabilization of folded ribonuclease T1 by about 4 kJ/mol each. This destabilization is probably caused by unfavorable electrostatic interactions of Glu28 and Asp29 with the negative end of the helix dipole. The activation energies for the unfolding of the different variants of ribonuclease T1 change in parallel with the differences in the thermodynamic stability when the pH is varied. This indicates that the unfavorable electrostatic interactions of Glu28 and Asp29 are lost very early in unfolding, and are not present in the activated state of unfolding.

Asparagine↗

PubMed1992

Chronic occupational exposure to organic solvents. XV. Glycol ether exposure during the manufacture of brakehoses.

Twenty-two persons (20 men and 2 women) were examined for their external and internal exposure to the glycol ether 1-methoxypropan-2-ol (PGME) during the production, leak testing and mounting of brakehoses. For the measurement of external exposure, personal air monitoring was the method of choice. Average concentrations of PGME of 82.2 mg/m3 (22.3 ppm), 68.6 mg/m3 (18.6 ppm) and 11.3 mg/m3 (3.1 ppm) were found in the air of the brakehose production, leak test and mounting areas, respectively. For the estimation of internal exposure to PGME, this glycol ether was measured in both urine and blood. The biological samples were taken post-shift. The highest internal exposure levels were found in the brakehose production section and in the leak test area. The average post-shift concentrations for PGME in workers in the brakehose production section were 4.6 mg/l in urine and 13.5 mg/l in blood; the corresponding figures for workers in the leak test area were 4.2 mg/l in urine and 11.0 mg/l in blood. In blood and urine samples of workers engaged in the mounting area, PGME levels were below the detection limits. The elimination kinetics of PGME were also studied in three highly exposed persons, and mean excretion half-lives of PGME of approximately 4.4 h were found. On the basis of our results we made a rough calculation of a future biological tolerance value: we would except that concentrations of 38-109 mg per litre of blood and 10-31 mg per litre of urine would correspond to the German MAK value for PGME (375 mg/m3).

Adult↗