PubMed Health⌕ Search

Biomedical subjects

Michael Braun

Publications and source records attributed to Michael Braun.

13 recordsLinked to original sources
PubMed2026

UGT1A1 genotype testing for irinotecan: A guideline developed by the UK Centre of Excellence in Regulatory Science and Innovation in Pharmacogenomics (CERSI-PGx).

Irinotecan, a topoisomerase I inhibitor, is available as both non-pegylated and pegylated formulations. The non-pegylated formulation is licensed for use in advanced colorectal cancer either in combination with other agents or as monotherapy. However, it is also used off-label across a range of gastrointestinal malignancies and in rare malignancies such as glioblastoma and sarcomas. The pegylated formulation is licensed for use as combination therapy in adult patients with metastatic pancreatic adenocarcinoma. Irinotecan is hydrolysed to its active metabolite, SN-38, which is predominantly inactivated by the enzyme uridine diphosphate glucuronosyltransferase UGT1A1. UGT1A1 is encoded by the gene UGT1A1, which is polymorphically expressed, with allele frequencies varying across populations. Poor metabolizers carry two variants that reduce UGT1A1 enzyme expression or activity, leading to increased risk of irinotecan toxicity. Any patient who is about to be prescribed irinotecan for an epithelial malignancy should have pharmacogenetic testing, to identify clinically relevant UGT1A1 variants, where testing is available. Irinotecan dose should be reduced by 30% at Cycle 1 treatment in poor metabolizers for all indications, with doses titrated thereafter based on tolerability and neutrophil counts. The lack of evidence precludes us from making any recommendation for rare malignancies such as sarcomas. Our guideline is consistent with other international pharmacogenetics prescribing guidelines. This guideline is grounded in the latest evidence but cannot account for all individual factors relevant to patient care. Therefore, prescribers must conduct a thorough assessment of each patient's risk-benefit profile, ensuring that therapy is optimized to maximize benefits while minimizing potential harms.

Humans↗
PubMed2003

Flow cytometric analysis of T cell proliferation in a mixed lymphocyte reaction with dendritic cells.

BACKGROUND: Dendritic cells (DCs) are the most potent antigen-presenting cells. They can be generated in vitro from CD14+ cells, and also from CD34+ progenitor cells. Although T cell proliferation using [3H] thymidine incorporation assay has been used widely to check DC function, this technique only provides limited information about the T cell proliferation. Here, we describe a novel method for quantitative analysis of T cell proliferation using flow cytometry. MATERIALS AND METHODS: DCs were generated from CD14+ cells from six healthy blood donors. Monocytes were isolated using positive selection with magnetic cell sorting (MACS) and then cultured with IL-4, GM-CSF, IL-1beta, IL-6, TNF-alpha and PGE(2) to yield fully mature DCs. Allogeneic naive T lymphocytes with known mismatches in HLA classes I and II were cocultured with DCs. Naive T cells without DC stimulation served as negative controls. T cells were harvested on days 0, 3, 5, 7, 9, 11 and analysed by flow cytometry. CD3-ECD and CD4-fluorescein isothiocyanate (FITC) or CD8-FITC antibodies were used to distinguish T cell subsets, whereas T cell activation was measured by assessment of HLA-DR, CD45RO, CD25 and CD71 expression. For T cell quantification, fluorescent microparticles were used. Dead cells were excluded with 7-AAD. The bromdeoxyridine (BrdU)-incorporation ELISA procedure was also performed in order to compare with the T cell proliferation assay with regard to absolute cell counts and CD71 expression. RESULTS: The initial T cell concentration on day 1 was 203.9+/-39.7 (173-265) CD3+/CD4+ cells/micro l and 184.5+/-41.6 (148-260) CD3+/CD8+ cells/micro l. The maximal T cell proliferation was recorded on day 7 with a five- to tenfold T cell expansion which resulted in 1994.9+/-383 (1446-2404) CD3+/CD4+ cells/micro l and 944+/-303.7 (560-1483) CD3+/CD8+ cells/micro l. Furthermore, activation markers of both cell lineages were upregulated and reached maxima on days 7 (CD71) and 9 (CD25, HLA-DR). T cell count/micro l as well as CD71 expression both correlated significantly with BrdU incorporation. CONCLUSION: Flow cytometric analysis permits simple, precise and rapid quantification of T cell proliferation in a mixed lymphocyte reaction with DCs. Activation, proliferation and cell viability can be simultaneously determined. CD71 is particularly well suited as an activation marker for the simultaneous measurement of T cell proliferation. Thus, specific T cell subsets involved in antigen-specific proliferation can be evaluated in detail.

Bromodeoxyuridine↗
PubMed2003

Smoothness-based forces for deformable models: a long-range force and a corner fitting force.

Deformable models, originally proposed by Terzopoulos et al. (Artif. Intell. 36 (1988) 91) and Kass et al. (Int. J. Comput. Vision 1 (1988) 321) in 1988, have been widely used in medical image segmentation. However, they manifest two well-known limitations: the lack of an appropriate long-range force to drive the model surface towards the object boundary and poor performance at high curvature boundaries (such as corners) due to the models' intrinsic smoothness constraint. In this paper, a new smoothness force with local control is proposed. The local control is used to devise a long-range force, referred to as the self-zoom force, and a corner fitting force. The self-zoom force enables the model surface to expand and shrink without a limit in range. The corner fitting force propels the model surface to fit high-curvature boundaries. Experiments demonstrate that the model surface is driven to the object boundary by the new forces even if the initial estimate is not close and the object is nonconvex or has a high local curvature.

Computer Simulation↗
PubMed2003

Software for image registration: algorithms, accuracy, efficacy.

Image registration is finding increased clinical use both in aiding diagnosis and guiding therapy. There are numerous algorithms for registration, which all involve maximizing a measure of similarity between a transformed floating image and a fixed reference image. The choice of the similarity measure depends, to some extent, on the application. Methods based on the use of the joint intensity histogram have become popular because of their flexibility and robustness. A distinction is made between rigid-body and non-rigid transformations. The latter are needed for inter-subject registration or intra-subject registration in cases where the region of the body of interest is not considered rigid. Non-rigid transformation is normally achieved using a global model of the deformation but can also be defined by a set of locally rigid transformations, each constrained to a small block in the image. There is scope for further research on the incorporation of appropriate constraints, especially for the application of non-rigid transformations to nuclear medicine studies. Most of the initial practical concerns regarding image registration have been overcome and there is increasing availability of commercial software. There are several approaches to the validation of registration software, with validation of non-rigid algorithms being particularly difficult. Studies have demonstrated the accuracy on the order of half a pixel for both intra- and inter-modality registration (typically 2 to 3 mm). Although hardware-based registration has now become possible by using dual-modality instruments, software-based registration will continue to play an important role in nuclear medicine.

Algorithms↗
PubMed2003

Mutant analysis of the Escherichia coli FhuA protein reveals sites of FhuA activity.

The FhuA outer membrane protein of Escherichia coli actively transports ferrichrome, albomycin, and rifamycin CGP 4832, and confers sensitivity to microcin J25, colicin M, and the phages T1, T5, and phi80. Guided by the FhuA crystal structure and derived predictions on how FhuA might function, mutants were isolated in the cork domain (residues 1 to 160) and in the beta-barrel domain (residues 161 to 714). Deletion of the TonB box (residues 7 to 11) completely inactivated all TonB-dependent functions of FhuA. Fixation of the cork to turn 7 of the barrel through a disulfide bridge between introduced C27 and C533 residues abolished ferrichrome transport, which was restored by reduction of the disulfide bond. Deletion of residues 24 to 31, including the switch helix (residues 24 to 29), which upon binding of ferrichrome to FhuA undergoes a large structural transition (17 A) and exposes the N terminus of FhuA (TonB box) to the periplasm, reduced FhuA transport activity (79% of the wild-type activity) but conferred full sensitivity to colicin M and the phages. Duplication of residues 23 to 30 or deletion of residues 13 to 20 resulted in FhuA derivatives with properties similar to those of FhuA with a deletion of residues 24 to 31. However, a frameshift mutation that changed QSEA at positions 18 to 21 to KKAP abolished almost completely most of FhuA's activities. The conserved residues R93 and R133 among energy-coupled outer membrane transporters are thought to fix the cork to the beta-barrel by forming salt bridges to the conserved residues E522 and E571 of the beta-barrel. Proteins with the E522R and E571R mutations were inactive, but inactivity was not caused by repulsion of R93 by R522 and R571 and of R133 by R571. Point mutations in the cork at sites that move or do not move upon the binding of ferrichrome had no effect or conferred only slightly reduced activities. It is concluded that the TonB box is essential for FhuA activity. The TonB box region has to be flexible, but its distance from the cork domain can greatly vary. The removal of salt bridges between the cork and the barrel affects the structure but not the function of FhuA.

Amino Acid Sequence↗
PubMed2003

In vivo reconstitution of the FhuA transport protein of Escherichia coli K-12.

The FhuA protein in the outer membrane of Escherichia coli actively transports ferrichrome and the antibiotics albomycin and rifamycin CGP 4832 and serves as a receptor for the phages T1, T5, and phi80 and for colicin M and microcin J25. The crystal structure reveals a beta-barrel with a globular domain, the cork, which closes the channel formed by the barrel. Genetic deletion of the cork resulted in a beta-barrel that displays no FhuA activity. A functional FhuA was obtained by cosynthesis of separately encoded cork and the beta-barrel domain, each endowed with a signal sequence, which showed that complementation occurs after secretion of the fragments across the cytoplasmic membrane. Inactive complete mutant FhuA and an FhuA fragment containing 357 N-proximal amino acid residues complemented the separately synthesized wild-type beta-barrel to form an active FhuA. Previous claims that the beta-barrel is functional as transporter and receptor resulted from complementation by inactive complete FhuA and the 357-residue fragment. No complementation was observed between the wild-type cork and complete but inactive FhuA carrying cork mutations that excluded the exchange of cork domains. The data indicate that active FhuA is reconstituted extracytoplasmically by insertion of separately synthesized cork or cork from complete FhuA into the beta-barrel, and they suggest that in wild-type FhuA the beta-barrel is formed prior to the insertion of the cork.

Bacterial Outer Membrane Proteins↗
PubMed2002

Iron transport and signaling in Escherichia coli.

Bacteria solve the iron supply problem caused by the insolubility of Fe(3+) by synthesizing iron-complexing compounds, called siderophores, and by using iron sources of their hosts, such as heme and iron bound to transferrin and lactoferrin. Escherichia coli, as an example of Gram-negative bacteria, forms sophisticated Fe(3+)-siderophore and heme transport systems across the outer membrane. The crystal structures of three outer membrane transport proteins now allow insights into energy-coupled transport mechanisms. These involve large long-range structural transitions in the transport proteins in response to substrate binding, including substrate gating. Energy is provided by the proton motive force of the cytoplasmic membrane through the activity of a protein complex that is inserted in the cytoplasmic membrane and that contacts the outer membrane transporters. Certain transport proteins also function in siderophore-mediated signaling cascades that start at the cell surface and flow to the cytoplasm to initiate transcription of genes encoding proteins for transport and siderophore biosynthesis.

Anti-Bacterial Agents↗
PubMed2002

Image registration: an essential tool for nuclear medicine.

There is increasing interest in being able to automatically register medical images from either the same or different modalities. Registered images are proving useful in a range of applications, not only providing more correlative information to aid in diagnosis, but also assisting with the planning and monitoring of therapy, both surgery and radiotherapy. The practising nuclear medicine specialist is faced with a dilemma in choosing an appropriate method since the literature in the field is extensive, with conflicting evidence as to what methods are optimal. Although most barriers to implementing registration in routine practice have been removed, there remains a lack of commercial, validated software. The alternative is to install a dual-modality instrument. The objective of this review is to present a general overview of medical image registration with emphasis on the application and issues relevant to nuclear medicine.

Algorithms↗
PubMed2002

Active transport of iron and siderophore antibiotics.

Bacteria solubilize iron (Fe(3+)) with secreted siderophores, which are then taken up as Fe(3+)-siderophore complexes. Some bacteria also use iron in heme, hemoglobin, hemopexin, transferrin and lactoferrin of eukaryotic hosts. Crystal structures of two outer membrane transport proteins, FhuA and FepA, and biochemical data reveal strong long-range conformational changes of the proteins upon binding of Fe(3+)-siderophore complexes and in response to energy transfer from the cytoplasmic membrane into the outer membrane via the TonB-ExbB-ExbD protein complex. The crystal structure of the periplasmic binding protein FhuD strongly deviates from the uniform overall structure of binding proteins hitherto determined. Sideromycins, antibiotics that contain Fe(3+)-siderophore complexes as carriers, are highly effective, as they enter cells via Fe(3+)-siderophore transport systems. In this review, recently published data is discussed to demonstrate the state of understanding of iron transport across the outer membrane and the cytoplasmic membrane.

Amino Acid Sequence↗
PubMed2002

Diffusion through channel derivatives of the Escherichia coli FhuA transport protein.

FhuA is a multifunctional protein in the outer membrane of Escherichia coli that actively transports [Fe3+]ferrichrome, the antibiotics albomycin and rifamycin CGP 4832, and mediates sensitivity of cells to the unrelated phages T5, T1, phi80 and UC-1, and to colicin M and microcin J25. The energy source of active transport is the proton motive force of the cytoplasmic membrane that is required for all FhuA functions except for infection by phage T5. The FhuA crystal structure reveals 22 antiparallel transmembrane beta-strands that form a beta-barrel which is closed by a globular N-terminal domain. FhuA still displays active transport and sensitivity to all ligands except microcin J25 when the globular domain (residues 5-160) is excised and supports weakly unspecific diffusion of substrates across the outer membrane. Here it is shown that isolated FhuADelta5-160 supported diffusion of ions through artificial planar lipid bilayer membranes but did not form stable channels. The double mutant FhuADelta5-160 Delta322-336 lacking in addition to the globular domain most of the large surface loop 4 which partially constricts the channel entrance, displayed an increased single-channel conductance but formed no stable channels. It transported in vivo[Fe3+]ferrichrome with 45% of the rate of wild-type FhuA and did not increase sensitivity of cells to antibiotics. In contrast, a second FhuA double mutant derivative which in addition to the globular domain contained a deletion of residues 335-355 comprising one-third of surface loop 4 and half of the transmembrane beta-strand 8 formed stable channels in lipid bilayers with a large single-channel conductance of 2.5 nS in 1 m KCl. Cells that synthesized FhuADelta5-160 Delta335-355 showed an increased sensitivity to antibiotics and supported diffusion of maltodextrins, SDS and ferrichrome across the outer membrane. FhuADelta5-160 Delta335-355 showed no FhuA specific functions such as active transport of [Fe3+]ferrichrome or sensitivity to the other FhuA ligands. It is concluded that FhuADelta5-160 Delta335-355 assumes a conformation that is incompatible with any of the FhuA functions.

Anti-Bacterial Agents↗
PubMed2002

Leishmania priming of human dendritic cells for CD40 ligand-induced interleukin-12p70 secretion is strain and species dependent.

A major question in the study of leishmaniasis is what dictates clinical disease expression produced by different Leishmania species, i.e., cutaneous versus systemic and healing versus nonhealing. Animal models using a Leishmania species associated with self-limiting cutaneous disease (L. major) have revealed that protective immunity requires CD40/CD40 ligand (CD40L)-dependent, interleukin-12 (IL-12)-driven Th1 responses. We recently showed that L. major can prime human dendritic cells (DCs) for CD40L-triggered IL-12p70 secretion and that these cells can drive a Th1 response in autologous T cells from sensitized individuals. Here we show that in contrast to L. major, Leishmania species responsible for visceral disease (L. donovani), as well as species associated with persistent, cutaneous lesions and occasional systemic disease (L. tropica), did not induce CD40L-dependent IL-12p70 production, despite comparable levels of uptake by DCs. Up-regulated surface expression of CD40 did not correlate with IL-12p70 production, and appreciable CD40L-induced IL-12p40 secretion was observed in uninfected as well as infected DCs, regardless of species. Reverse transcription-PCR analysis confirmed that the production of heterodimeric IL-12 was limited by expression of IL-12p35 mRNA, which was dependent on both a microbial priming signal and CD40 engagement for its high-level induction. The intrinsic differences in the ability of Leishmania species to prime DCs for CD40L-dependent IL-12p70 secretion may account, at least in part, for the evolution of healing and nonhealing forms of leishmanial disease.

Animals↗
PubMed2002

The isolated perfused bovine uterus as a model for mucous membrane irritation and inflammation.

Isolated perfused bovine uteri were used to study the irritation potential of antiseptics frequently used in veterinary practice for the treatment of endometritis. Comparable to the isolated perfused bovine udder (Kietzmann et al., 1993), the viability of uteri obtained directly after slaughtering of cows, was tested with the help of biochemical parameters. These parameters of the viability were nearly unchanged over five hours. Thereafter, different antiseptics were tested for their irritation potential on the mucous membrane with the help of the MTT-assay. Here the irritant effect of Lugol's iodine solution is demonstrated exemplarily. An additional study was performed to establish a hemoperfused uterus. An injection of arachidonic acid into the serosa induces a visible inflammatory reaction and a marked increase of the prostaglandin E2 synthesis. In conclusion, the isolated perfused bovine uterus seems to be a promising in vitro model for mucous membrane irritancy as well as for inflammatory reactions.

Animals↗