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Biomedical subjects

Wolfgang Weber

Publications and source records attributed to Wolfgang Weber.

10 recordsLinked to original sources
PubMed2006

Monitoring tumor glucose utilization by positron emission tomography for the prediction of treatment response to epidermal growth factor receptor kinase inhibitors.

PURPOSE: The mechanisms underlying the sensitivity of non-small cell lung cancer to epidermal growth factor receptor (EGFR) kinase inhibitors are complex, and there are no established markers to accurately predict treatment outcome in individual patients. EXPERIMENTAL DESIGN: We investigated whether tumors responding to EGFR inhibitors can be identified by measuring treatment-induced changes in glucose utilization by positron emission tomography with the glucose analogue fluorodeoxyglucose (FDG-PET). We studied a panel of cell lines with a spectrum of sensitivity to EGFR kinase inhibitors. After incubation with the EGFR kinase inhibitor gefitinib for various time points, FDG uptake, glucose transport rates, and hexokinase activity were determined. FDG uptake in vivo was assessed by microPET imaging of tumor xenografts in mice. RESULTS: In gefitinib-sensitive cell lines, there was a dramatic decrease in FDG uptake as early as 2 hours after treatment. Immunoblots showed the translocation of glucose transporters (GLUT3) from the plasma membrane to the cytosol; glucose transport rates were reduced 2.6-fold at this time. There was also a modest reduction of hexokinase activity. These metabolic alterations preceded changes in cell cycle distribution, thymidine uptake, and apoptosis. MicroPET studies showed an up to 55% decrease of tumor FDG uptake in sensitive xenografts within 48 hours. In contrast, gefitinib-resistant cells exhibited no measurable changes in FDG uptake, either in cell culture or in vivo. CONCLUSION: Glucose metabolic activity closely reflects response to gefitinib therapy. FDG-PET may be a valuable clinical predictor, early in the course of treatment, for therapeutic responses to EGFR kinase inhibitors.

Animals↗
PubMed2006

Lysine as a critical amino acid for IgE binding in Phl p 5b C terminus.

BACKGROUND: Allergens induce the formation of specific immunoglobulin (Ig)E and harbor at least two IgE-binding regions (epitopes) to facilitate crosslinking of basophilic or mast-cell-bound specific IgE antibodies. Studies mapping linear epitopes have shown that these regions often contain charged or hydrophobic amino acids. Nevertheless, these studies are hampered by limited significance due to the often conformational nature of IgE epitopes. This prompted us to study the role of lysines in the context of an intact 3-dimensional model. METHODS: Major allergen Phl p 5b from timothy grass bears 12 lysines in its C-terminal half. Using site-directed mutagenesis, we substituted all 10 surface-exposed lysines by alanines. RESULTS: Although structural integrity of the lysine-deficient mutant was not altered, IgE-binding capacity measured by ELISA inhibition tests and crosslinking activity in ex vivo basophil stimulation and in vivo skin prick tests were significantly diminished. Interestingly, binding of specific IgG antibodies was considerably less reduced by loss of lysines. CONCLUSION: Lysine is an important amino acid for IgE binding in more than one epitope of major grass pollen allergen Phl p 5b C terminus. Allergenicity, but not IgG binding of the molecule, is substantially diminished by single amino acid substitutions without structural integrity being hampered.

Allergens↗
PubMed2005

Offspring from non-stimulated calves at an age younger than two months: a preliminary report.

This study investigated the viability of embryos from non-stimulated 2-3-month-old calves generated in vitro using oocytes from follicles of defined size in terms of their ability to produce full-term pregnancies. Ablation of follicles>or=4 mm was used to induce the emergence of a new follicular wave, and calves (n=3) were laparoscopically punctured three times at 7-day-intervals to recover cumulus-oocyte-complexes (COCs) from follicles>8 (group A) and between 4-8 mm (group B). Calves were aged 49, 56, and 80 days, respectively, at first recovery. Morphologically intact COCs were subjected to in vitro maturation, fertilization, and embryo culture, and compact morulae/blastocysts were transferred on day 7 post-insemination to synchronized virgin heifers. Blood typing was used for maternity analysis. A total of 29 COCs were recovered, 21 cultured, yielding 11 cleaved embryos (52.4%) and 6 compact morulae/blastocysts (28.6%). No differences were observed between groups. Transfer of the 6 embryos to 5 recipients resulted in three pregnancies (one from group A and two from group B). Two normal male offspring (both from group B), with birth weights of 44 and 51 kg, were born, and two donor calves, aged 56 and 59 days, were identified as the dams. In conclusion, the results demonstrate that embryos generated in vitro from oocytes from non-stimulated calves at an age younger than two months are viable in terms of their ability to produce full-term pregnancies, and suggest that the viability of calf embryos is not related to follicle size.

Animals↗
PubMed2005

The in vitro developmental competence of oocytes from juvenile calves is related to follicular diameter.

This study investigated the relationship between follicle size (FS) and developmental competence of calf oocytes. Cumulus-oocyte-complexes (COCs) from follicles>8 (L-COCs; n=19), 4-8 (M-COCs; n=54), and 2-3 mm (S-COCs; n=155) were recovered from non-stimulated 1-4 months old dairy calves post mortem and ex vivo (laparoscopy), and in parallel from slaughtered adult cows from follicles of identical size categories [>8 (n=91); 4-8 (n=138); 2-3 mm (n=193)]. Morphologically intact COCs were subjected to in vitro maturation, fertilization, and embryo culture. Cleavage rate (CR; 46 h post-insemination=p.i.), rate of morulae/blastocysts (M/Bl; day 7 p.i.), and blastocysts (Bl; day 9 p.i.) were recorded. FS had no effect on the CR in calves. However, calf L-COCs yielded the highest rates of M/Bl and Bl compared with the two other size categories (P<0.05). In contrast, calf S- and M-COCs gave similar rates of M/Bl, whereas the proportion of Bl was lowest for S-COCs (P<0.05). This was almost identical to findings in cows, except that the CR was highest for L-COCs and M/Bl yields were lowest for S-COCs (P<0.05). There were no differences between calf and cows with regard to CR for the respective FS categories. L-COCs from calves and cows yielded similar rates of M/Bl and Bl, whereas calf S- and M-COCs yielded lower rates of Bl than S- and M-COCs from cows and a lower rate of M/Bl when S-and M-COCs were analyzed as one group (P<0.05). Whereas the CR was similar in calves and cows, calf COCs yielded lower rates of M/Bl and Bl (P<0.05). In conclusion, the results show that the developmental competence of calf oocytes is higher in those derived from follicles larger than 8 mm, and thus are almost equally as competent as cow oocytes derived from follicles of identical size. This suggests that calf oocytes acquire developmental competence within the large follicle, potentially due to a process similar to prematuration of the oocyte in the adult cow. It is proposed that procedures that facilitate prematuration, such as "coasting" following a preceding superstimulation, might increase the developmental competence of calf oocytes.

Age Factors↗
PubMed2002

Structure of the functional domain of the major grass-pollen allergen Phlp 5b.

The major allergen Phlp 5b from timothy grass pollen induces allergic rhinitis and bronchial asthma in millions of allergic patients worldwide. As an important step towards understanding the interactions between the pollen protein and components of the human immune system, the structure of the C-terminal key domain of Phlp 5b has been determined at 2.0 A resolution and refined to an R value of 19.7%. This is the first known allergen composed entirely of alpha-helices. The protein forms a dimer stabilized by one intermolecular disulfide bridge. Sequence homology suggests that at least all group V and group VI grass-pollen allergens belong to this new class of 'four-helix-bundle allergens'.

Allergens↗
PubMed2002

Biosynthetic precursors of the lipase inhibitor lipstatin.

Three putative intermediates in the biosynthesis of the lipase inhibitor lipstatin were synthesized in stable isotope-labeled form and were added to fermentation cultures of Streptomyces toxytricini. Biosynthetic lipstatin was isolated and analyzed by NMR spectroscopy. [3,10,11,12-(2)H]-(3S,5Z,8Z)-3-hydroxytetradeca-5,8-dienoic acid (9) was shown to serve as a direct biosynthetic precursor of lipstatin. [7,8-(2)H(2)]Hexylmalonate (11) was also incorporated into lipstatin, albeit at a relatively low rate. The leucine moiety of [(13)C-formyl,(15)N]-N-formylleucine (10) was diverted to lipstatin under loss of the (13)C-labeled formyl residue.

Animals↗
PubMed2002

The major extracellular protease of the nosocomial pathogen Stenotrophomonas maltophilia: characterization of the protein and molecular cloning of the gene.

Stenotrophomonas maltophilia is increasingly emerging as a multiresistant pathogen in the hospital environment. In immunosuppressed patients, these bacteria may cause severe infections associated with tissue lesions such as pulmonary hemorrhage. This suggests proteolysis as a possible pathogenic mechanism in these infections. This study describes a protease with broad specificity secreted by S. maltophilia. The gene, termed StmPr1, codes for a 63-kDa precursor that is processed to the mature protein of 47 kDa. The enzyme is an alkaline serine protease that, by sequence homology and enzymic properties, can be further classified as a new member of the family of subtilases. It differs from the classic subtilisins in molecular size, in substrate specificity, and probably in the architecture of the active site. The StmPr1 protease is able to degrade several human proteins from serum and connective tissue. Furthermore, pan-protease inhibitors such as alpha(1)-antitrypsin and alpha(2)-macroglobulin were unable to abolish the activity of the bacterial protease. The data support the interpretation that the extracellular protease of S. maltophilia functions as a pathogenic factor and thus could serve as a target for the development of therapeutic agents.

Amino Acid Sequence↗
PubMedDate not supplied

Phosphoenolpyruvate availability and the biosynthesis of shikimic acid.

The impact of increased availability of phosphoenolpyruvate during shikimic acid biosynthesis has been examined in Escherichia coli K-12 constructs carrying plasmid-localized aroF(FBR) and tktA inserts encoding, respectively, feedback-insensitive 3-deoxy-d-arabino-heptulosonic acid 7-phosphate synthase and transketolase. Strategies for increasing the availability of phosphoenolpyruvate were based on amplified expression of E. coli ppsA-encoded phosphoenolpyruvate synthase or heterologous expression of the Zymomonas mobilis glf-encoded glucose facilitator. The highest titers and yields of shikimic acid biosynthesized from glucose in 1 L fermentor runs were achieved using E. coli SP1.lpts/pSC6.090B, which expressed both Z. mobilis glf-encoded glucose facilitator protein and Z. mobilis glk-encoded glucose kinase in a host deficient in the phosphoenolpyruvate:carbohydrate phosphotransferase system. At 10 L scale with yeast extract supplementation, E. coli SP1.lpts/pSC6.090B synthesized 87 g/L of shikimic acid in 36% (mol/mol) yield with a maximum productivity of 5.2 g/L/h for shikimic acid synthesized during the exponential phase of growth.

3-Deoxy-7-Phosphoheptulonate Synthase↗