Biomedical subjects

Matthias Müller

Publications and source records attributed to Matthias Müller.

At least 19 recordsLinked to original sources

PubMed2007

Continuous SvO2 measurement in infants undergoing congenital heart surgery--first clinical experiences with a new fiberoptic probe.

BACKGROUND: SvO2-guided therapy, using fiberoptic oximetric catheters can help to improve the outcome after complex congenital heart surgery especially in infants undergoing the Stage 1-Norwood-Procedure. So far, fiberoptic catheters have to be placed transthoracically by the surgeon into the vena cava or the pulmonary artery putting the infant at an additional risk of bleeding at the time of catheter removal. METHODS: We used a new percutaneously applicable fiberoptic probe for continuous monitoring of central venous saturation in three infants undergoing modifications of the Stage 1-Norwood-Procedure (reconstruction of the aortic arch), two in combination with a bidirectional cavopulmonary connection (Glenn shunt), the third with reconstruction of the pulmonary arteries (biventricular repair). The approved clinical monitoring system consisted of a small (2 F) fiberoptic probe and a bed-side-monitor. The probe was inserted via the routine central venous access for such a case. RESULTS: Continuous SvO2 measurement is feasible in infants using the CeVOX system in combination with routine central venous access, and the advantages of continuous monitoring are discussed. CONCLUSIONS: We believe that compared with transthoracically inserted oximetric catheter, the presented percutaneous technique avoids additional risks at the time of catheter removal (i.e. bleeding) and has become part of our standard management.

Cardiopulmonary Bypass↗

PubMed2006

Matrix metalloproteinase-19 expression in keratinocytes is repressed by transcription factors Tst-1 and Skn-1a: implications for keratinocyte differentiation.

Matrix metalloproteinase-19 (MMP-19), unlike other members of the MMP family, is expressed in basal keratinocytes of intact epidermis whereas keratinocytes in suprabasal and higher epidermal layers express this enzyme only during cutaneous disorders. As the activity of MMP-19 effects proliferation, migration, and adhesion of keratinocytes we examined whether transcription factors involved in keratinocyte differentiation repress the expression of MMP-19. Using luciferase reporter assays, POU transcription factors Tst-1 (Oct-6) and Skn-1a (Oct-11) markedly downregulated the activity of MMP-19 promoter in COS-7 cells and HaCaT keratinocytes. Tst-1 alone was able to inhibit 85% of the promoter activity. Skn-1a exhibited a weak inhibitory effect although it synergistically increased effects of Tst-1. HaCaT cells stably transfected with Tst-1 showed a strong decrease of activity of MMP-19 promoter that correlated with suppression of MMP-19, cytokeratin 14 and 5, decreased cell proliferation, and altered expression of involucrin and loricrin. The expression of MMP-9 was also significantly reduced in Tst-1 expressing keratinocytes. MMP-2 was substantially affected during its activation whereas the expression of MMP-28 was unchanged. Our results suggest that Tst-1 and Skn-1a regulate expression of MMPs in keratinocytes and effect both the expression and activation of these proteolytic enzymes.

Animals↗

PubMed2006

Min-map bias Monte Carlo for chain molecules: biased Monte Carlo sampling based on bijective minimum-to-minimum mapping.

A novel Monte Carlo (MC) simulation scheme based on Theodorou's bijective mapping strategy [D. N. Theodorou, J. Chem. Phys. 124, 034109 (2006)] is introduced. This min-map bias Monte Carlo acts in combination with any other proper, bare MC. It carries over the bare MC move from the original configuration space Omega(0), where trial move acceptance may be low, to a different configuration space, Omega(1), where acceptance is higher. The bare MC move is then performed in Omega(1) and the resulting configuration is finally mapped back to Omega(0). Mappings between Omega(0) and Omega(1) entail weighted selection of trial configurations, the bias of which is subsequently removed in the overall acceptance criterion. The new method is applied, in conjunction with continuum configurational bias as bare MC scheme, to the simulation of explicit hydrogen linear alkanes in the canonical ensemble. Min-map bias MC is found to alleviate the pervasive problem of very low acceptance rates encountered when using an explicit molecular description.

Journal Article↗

PubMed2006

A derivative of lipid A is involved in signal recognition particle/SecYEG-dependent and -independent membrane integrations.

A cell-free system was developed that allows the correct integration of single and multispanning membrane proteins of Escherichia coli into proteoliposomes. We found that physiological levels of diacylglycerol were required to prevent spontaneous integration into liposomes even of the polytopic mannitol permease. Using diacylglycerol-containing proteoliposomes, we identified a novel integration-stimulating factor. Integration of mannitol permease was dependent on both the SecYEG translocon and this factor and was mediated by signal recognition particle and signal recognition particle receptor. Integration of M13 procoat, which is independent of both signal recognition particle/signal recognition particle receptor and SecYEG, was also promoted by this factor. Furthermore, the factor stimulated the post-translational translocation of presecretory proteins, suggesting that it also mediates integration of a signal sequence. This factor was found to be a lipid A-derived membrane component possessing a peptide moiety.

Bacterial Outer Membrane Proteins↗

PubMed2006

The suitability of different DHB isomers as matrices for the MALDI-TOF MS analysis of phospholipids: which isomer for what purpose?

Although the analysis of large biomolecules is the prime application of matrix-assisted laser desorption and ionization time-of-flight mass spectrometry (MALDI-TOF MS), there is also increasing interest in lipid analysis. Since lipids possess relatively small molecular weights, matrix signals should be as small as possible to avoid overlap with lipid peaks. Although 2,5-dihydroxybenzoic acid (DHB) is an established MALDI matrix, the question whether just this isomer is ideal for lipid analysis was not yet addressed. UV absorptions of all six DHB isomers were determined and their laser desorption spectra recorded. In addition, all isomers were used as matrices to record positive and negative ion mass spectra of selected phospholipids (phosphatidylcholine and -serine): In the order 2,5-, 2,6-, 2,3- and 2,4-DHB, the quality of the positive ion lipid spectra decreases. This correlates well with the decreasing acidity of the applied DHB isomers. The 3,4- and 3,5- isomers give only very weak positive ion signals especially of acidic lipids. In contrast, the most suitable matrices in the negative ion mode are 2,5-, 2,4- and 3,5-DHB. 2,6-DHB does not provide any signal in the negative ion mode due to its marked acidity. Finally, differences in the crystallization behavior of the pure matrix and the matrix/lipid co-crystals were also monitored by atomic force microscopy (AFM): 2,5-DHB gave the smallest crystals and the skinniest layer. It is concluded that basically all DHB isomers can be used as MALDI matrices but the 2,5-isomer represents the most versatile compound.

Gentisates↗

PubMed2006

Identification of single-point mutations in mycobacterial 16S rRNA sequences by confocal single-molecule fluorescence spectroscopy.

We demonstrate the specific identification of single nucleotide polymorphism (SNP) responsible for rifampicin resistance of Mycobacterium tuberculosis applying fluorescently labeled DNA-hairpin structures (smart probes) in combination with single-molecule fluorescence spectroscopy. Smart probes are singly labeled hairpin-shaped oligonucleotides bearing a fluorescent dye at the 5' end that is quenched by guanosine residues in the complementary stem. Upon hybridization to target sequences, a conformational change occurs, reflected in a strong increase in fluorescence intensity. An excess of unlabeled ('cold') oligonucleotides was used to prevent the formation of secondary structures in the target sequence and thus facilitates hybridization of smart probes. Applying standard ensemble fluorescence spectroscopy we demonstrate the identification of SNPs in PCR amplicons of mycobacterial rpoB gene fragments with a detection sensitivity of 10(-8) M. To increase the detection sensitivity, confocal fluorescence microscopy was used to observe fluorescence bursts of individual smart probes freely diffusing through the detection volume. By measuring burst size, burst duration and fluorescence lifetime for each fluorescence burst the discrimination accuracy between closed and open (hybridized) smart probes could be substantially increased. The developed technique enables the identification of SNPs in 10(-11) M solutions of PCR amplicons from M.tuberculosis in only 100 s.

Bacterial Proteins↗

PubMed2006

Secreted modular calcium-binding protein-1 localization during mouse embryogenesis.

BM-40 is an extracellular matrix-associated protein and is characterized by an extracellular calcium-binding domain as well as a follistatin-like domain. Secreted modular calcium-binding protein-1 (SMOC-1) is a new member of the BM-40 family. It consists of two thyroglobulin-like domains, a follistatin-like domain and a new domain without known homologues and is expressed ubiquitously in many adult murine tissues. Immunofluorescence studies, as well as immunogold electron microscopy, have confirmed the localization of SMOC-1 in or around basement membranes of adult murine skin, blood vessels, brain, kidney, skeletal muscle, and the zona pellucida surrounding the oocyte. In the present work, light microscopic immunohistochemistry has revealed that SMOC-1 is localized in the early mouse embryo day 7 throughout the entire endodermal basement membrane zone of the embryo proper. SMOC-1 mRNA is synthesized, even in early stages of mouse development, by mesenchymal as well as epithelial cells deriving from all three germ layers. In embryonic stage day 12, and fetal stages day 14, 16, and 18, the protein is present in the basement membrane zones of brain, blood vessels, skin, skeletal muscle, lung, heart, liver, pancreas, intestine, and kidney. This broad and organ-specific distribution suggests multifunctional roles of SMOC-1 during mouse embryogenesis.

Animals↗

PubMed2006

Differential compartmentalization and distinct functions of GABAB receptor variants.

GABAB receptors are the G protein-coupled receptors for the main inhibitory neurotransmitter in the brain, gamma-aminobutyric acid (GABA). Molecular diversity in the GABAB system arises from the GABAB1a and GABAB1b subunit isoforms that solely differ in their ectodomains by a pair of sushi repeats that is unique to GABAB1a. Using a combined genetic, physiological, and morphological approach, we now demonstrate that GABAB1 isoforms localize to distinct synaptic sites and convey separate functions in vivo. At hippocampal CA3-to-CA1 synapses, GABAB1a assembles heteroreceptors inhibiting glutamate release, while predominantly GABAB1b mediates postsynaptic inhibition. Electron microscopy reveals a synaptic distribution of GABAB1 isoforms that agrees with the observed functional differences. Transfected CA3 neurons selectively express GABAB1a in distal axons, suggesting that the sushi repeats, a conserved protein interaction motif, specify heteroreceptor localization. The constitutive absence of GABAB1a but not GABAB1b results in impaired synaptic plasticity and hippocampus-dependent memory, emphasizing molecular differences in synaptic GABAB functions.

Animals↗

PubMed2006

Effect of isoflurane on echocardiographic left-ventricular relaxation indices in patients with diastolic dysfunction due to concentric hypertrophy and ischemic heart disease.

OBJECTIVE: The purpose of this study was to investigate whether isoflurane, a known negative lusitropic agent, exacerbates diastolic dysfunction in patients with preexisting impaired relaxation. DESIGN: Prospective, experimental study. SETTING: Single-institution, university hospital. PARTICIPANTS: Twenty-five patients with diastolic dysfunction due to concentric hypertrophy and ischemic heart disease undergoing elective coronary artery bypass graft surgery. INTERVENTIONS: After approval of the local ethics committee and informed consent, patients randomly received sufentanil/midazolam anesthesia plus either 0.5 to 1.0 minimum alveolar concentration of isoflurane (n = 15) or weight-adjusted boli of urapidil (n = 10) during preparation of the internal mammary artery. Changes in hemodynamic parameters and echocardiographic diastolic indices before and after drug administration were compared. Filling pressures during the study were kept constant within normal range. MEASUREMENTS AND MAIN RESULTS: Hemodynamic changes measured by invasive arterial and pulmonary arterial pressures were comparable between isoflurane and urapidil. Both interventions led to a marked reduction in afterload that was accompanied by a significant increase in thermodilution cardiac output and stroke volume. Transesophageal echocardiographic relaxation indices were also comparable between groups. Transmitral and tissue Doppler E waves increased significantly, leading to larger E/A and Em/Am ratios; whereas the deceleration time and the isovolumetric relaxation time decreased significantly. CONCLUSION: Isoflurane did not exacerbate diastolic dysfunction in patients with concentric hypertrophy and ischemic heart disease. In contrast, isoflurane led to a "normalization" of the relaxation pattern that was attributed to a reduction in left-ventricular loading conditions.

Aged↗

PubMed2006

Alternate recruitment of signal recognition particle and trigger factor to the signal sequence of a growing nascent polypeptide.

Different from cytoplasmic membrane proteins, presecretory proteins of bacteria usually do not require the signal recognition particle for targeting to the Sec translocon. Nevertheless signal sequences of presecretory proteins have been found in close proximity to signal recognition particle immediately after they have emerged from the ribosome. We show here that at the ribosome, the molecular environment of a signal sequence depends on the nature of downstream sequence elements that can cause an alternate recruitment of signal recognition particle and the ribosome-associated chaperone Trigger factor to a growing nascent chain. While signal recognition particle and Trigger factor might remain bound to the same ribosome, both ligands are clearly able to displace each other from a nascent chain. The data also imply that a signal sequence owes its molecular environment to the fact that it remains closely apposed to the ribosomal exit site during growth of a nascent secretory protein.

Adenosine Triphosphatases↗

PubMed2006

Global down-regulation of gene expression in the brain using RNA interference, with emphasis on monoamine transporters and GPCRs: implications for target characterization in psychiatric and neurological disorders.

RNA interference (RNAi) is a natural mechanism for regulating gene expression, which exists in plants, invertebrates, and mammals. We investigated whether non-viral infusion of short interfering RNA (siRNA) by the intracerebroventricular route would enable a sequence-specific gene knockdown in the mouse brain and whether the knockdown translates into disease-relevant behavioral changes. Initially, we targeted enhanced green fluorescent protein (EGFP) in mice overexpressing EGFP. A selective knockdown of both EGFP protein and mRNA was observed throughout the brain, with lesser down-regulation in regions distal to the infusion site. We then targeted endogenous genes, encoding the dopamine (DAT) and serotonin transporters (SERT). DAT-siRNA infusion in adult mice produced a significant down-regulation of DAT mRNA and protein and elicited hyperlocomotion similar, but delayed, to that produced on infusion of GBR-12909, a potent and selective DAT inhibitor. Similarly, SERT-siRNA infusion resulted in significant knockdown of SERT mRNA and protein and elicited reduced immobility in the forced swim test similar to that obtained on infusion of citalopram, a very selective and potent SSRI. Application of this non-viral RNAi approach may accelerate target validation for neuropsychiatric disorders that involve a complex interplay of gene(s) from various brain regions.

Animals↗

PubMed2005

Sphingosine kinase type 2 is essential for lymphopenia induced by the immunomodulatory drug FTY720.

FTY720, a potent immunomodulatory drug in phase 2/3 clinical trials, induces rapid and reversible sequestration of lymphocytes into secondary lymphoid organs, thereby preventing their migration to sites of inflammation. As prerequisite for its function, phosphorylation of FTY720 to yield a potent agonist of the sphingosine-1-phosphate receptor S1P(1) is required in vivo, catalyzed by an as-yet-unknown kinase. Here, we report on the generation of sphingosine kinase 2 (SPHK2) knockout mice and demonstrate that this enzyme is essential for FTY720 phosphate formation in vivo. Consequently, administration of FTY720 does not induce lymphopenia in SPHK2-deficient mice. After direct dosage of FTY720 phosphate, lymphopenia is only transient in this strain, indicating that SPHK2 is constantly required to maintain FTY720 phosphate levels in vivo.

Animals↗

PubMed2005

Twin-arginine-specific protein export in Escherichia coli.

In many prokaryotic organisms, secretory proteins harboring a twin-arginine consensus motif are exported in a fully folded conformation via the twin-arginine translocation (Tat) pathway. In Escherichia coli, Tat involves the three structurally and functionally different membrane proteins TatA, TatB, and TatC. Whereas TatC proteins function in the specific recognition of substrate, TatA might be the major pore-forming subunit.

Escherichia coli↗

PubMed2005

Basement membrane composition in the early mouse embryo day 7.

Basement membranes (BM) are specialized structures of the extracellular matrix known to be involved in various early developmental processes. Despite numerous investigations on the localization of BM components, it remains unknown which molecules are expressed in early developmental stages and by which germ layers these proteins are produced. Therefore, we tested for all known laminin chains, nidogens, collagen type IV, and perlecan by means of light microscopic immunostaining and performed in situ reverse transcriptase-polymerase chain reaction to detect the mRNAs specific for laminin alpha1, laminin beta1, the alpha1 chain of collagen type IV, nidogen-2, and perlecan in the early mouse embryo, day 7, in vivo. Only the laminin chains alpha1, beta1, and gamma1 were detected immunohistochemically throughout the entire endodermal and ectodermal BM zones of the embryo proper. The mRNA of laminin alpha1, laminin beta1, collagen type IV, nidogen-2 and perlecan were expressed in the ectoderm-derived mesoderm, in the endoderm as well as in the ectoderm. In contrast, Reichert's membrane was positive for all laminin chains except for the alpha4, alpha5, beta3, and gamma3 chains. Moreover, maternal epithelial as well as mesenchymal cells expressed laminins, nidogen-1 and nidogen-2, collagen type IV, and perlecan. In conclusion, laminin-1 might be the only laminin isoform in the early mouse embryo that, together with the other main BM components, nidogens, collagen type IV, and perlecan, is synthesized by all three germ layers.

Animals↗

PubMed2005

Localization of Apaf1 gene expression in the early development of the mouse by means of in situ reverse transcriptase-polymerase chain reaction.

Apoptosis is an essential ubiquitous process that controls the duration of the life span of cells, thus playing a crucial role in morphogenetic, histogenetic, and phylogenetic developmental processes. Apaf1 (apoptosis protease activating factor 1) is one of the central mediators of the intrinsic apoptotic pathway and a part of the apoptosome, which activates procaspase-3 and promotes cell death. Gene knockout of Apaf1 in mice leads to late embryonic lethality with malformations such as the persistence of interdigital webs and hyperplasia of brain and retina. Therefore, Apaf1 is generally believed to play a crucial role in developmental apoptosis and have a widespread expression. However, its pattern of expression in early development remains unknown. To specify whether Apaf1 indeed plays this key role, we investigated the pattern of gene expression for Apaf1 in mouse embryos on day 7, 9, and 12 of development. Our results show, that gene expression for Apaf1 first occurs within the embryo between day 7 and 9 of development, becoming more widespread toward day 12 and then includes structures, such as yolk sac, mesenchyme, cartilage, heart anlage, otic vesicle, peridermis, and anlagen of the spinal ganglia and vertebral bodies. Our results also show that gene expression for Apaf1 is not ubiquitous in early mouse development. This finding indicates that cell death processes are independent of or less dependent on Apaf1 during this time. Of interest, an active gene expression for Apaf1 is also present in organ anlagen such as heart or intestine, in which no obvious phenotype is seen after Apaf1 deletion. This finding suggests a possible role for Apaf1 in such anlagen as a putative alternative compensatory pathway, which could be switched on in the case of defects in the mediators that are normally involved in such organs.

Animals↗

PubMed2005

Strong and ubiquitous expression of transgenes targeted into the beta-actin locus by Cre/lox cassette replacement.

Conventional approaches to produce transgenic mice recurrently yield unpredictable patterns and levels of transgene expression, a situation calling for the development of new techniques to overcome these drawbacks in the context of overexpression studies. Here we present an efficient method for rapid and reproducible transgenesis using the recombinase mediated cassette exchange (RMCE) (Bouhassira et al.: Blood 90:3332-3344, 1997) procedure. A lox511-EGFP-TK/neo-loxP cassette was placed under the control of the endogenous mouse beta-actin promoter. Heterozygous mice revealed strong and ubiquitous EGFP expression throughout embryogenesis and adulthood. Reproducibly, the same expression pattern was obtained with RMCE when it was used to replace the EGFP-harboring cassette by ECFP or placental alkaline phosphatase (PLAP) reporter genes (DePrimo et al.: Transgenic Res 5:459-466, 1996). Furthermore, the RMCE procedure proved efficient as well in embryonic stem (ES) cells as directly in zygotes. Our results demonstrate ubiquitous expression of floxed transgenes in the endogenous beta-actin locus and they support the general use of the beta-actin locus for targeted transgenesis.

Actins↗

PubMed2004

Neurochemical and behavioral consequences of widespread gene knockdown in the adult mouse brain by using nonviral RNA interference.

Gene expression analysis implicates an increasing number of novel genes in the brain as potential targets for the treatment of neurological and psychiatric disorders. Frequently, these genes are ubiquitously expressed in the brain and, thus, may contribute to a pathophysiological state through actions in several brain nuclei. Current strategies employing genetically modified animals for in vivo validation of such targets are time-consuming and often limited by developmental adaptations. Somatic gene manipulation using viral-mediated RNA interference (RNAi) has emerged recently, although restricting the target validation to specific brain nuclei. We investigated whether nonviral infusion of short interfering RNA (siRNA) into the ventricular system would enable a sequence-specific gene knockdown. The temporality and extent of siRNA-induced down-regulation were analyzed by targeting a transgene, EGFP, in mice overexpressing EGFP. Extensive knockdown of EGFP was observed, especially in regions adjacent or dorsoventrally and mediolaterally distant to the infusion site (dorsal third ventricle), with lesser knockdown in more distal regions. We challenged our RNAi approach to generate a specific knockdown of an endogenous gene, encoding the dopamine transporter (DAT) in regions (ventral midbrain) far distal to the infusion site. DAT-siRNA infusion in adult mice produced a significant down-regulation of DAT mRNA and protein in the brain and also elicited a temporal hyperlocomotor response similar to that (but delayed) obtained upon infusion of GBR-12909, a pharmacologically selective DAT inhibitor. Application of this nonviral RNAi approach may accelerate target validation for neuropsychiatric disorders that involve a complex interplay of gene(s) from various brain regions.

Animals↗

PubMed2004

Isolation and characterization of bifunctional Escherichia coli TatA mutant proteins that allow efficient tat-dependent protein translocation in the absence of TatB.

In Escherichia coli, the Tat system promotes the membrane translocation of a subset of exported proteins across the cytoplasmic membrane. Four genes (tatA, tatB, tatC, and tatE) have been identified that encode the components of the E. coli Tat translocation apparatus. Whereas TatA and TatE can functionally substitute for each other, the TatB and the TatC proteins have been shown to perform distinct functions. In contrast to Tat systems of the ABC(E) type found in E. coli and many other bacteria, some microorganisms possess a TatAC-type translocase that consists of TatA and TatC only, suggesting that, in these systems, TatB is not required or that one of the remaining components (TatA or TatC) additionally takes over the TatB function. We have addressed the molecular basis for the difference in subunit composition between TatABC(E) and TatAC-type systems by using a genetic approach. A plasmid-encoded E. coli minimal Tat translocase consisting solely of TatA and TatC was shown to mediate a low level translocation of a sensitive Tat-dependent reporter protein. Suppressor mutations in the minimal Tat translocase were isolated that compensate for the absence of TatB and that showed substantial increases in translocation activities. All of the mutations mapped to the extreme amino-terminal domain of TatA. No mutations affecting TatC were identified. These results suggest that in TatAC-type systems, the TatA protein represents a bifunctional component fulfilling both the TatA and TatB functions. Furthermore, our results indicate that the structure of the amino-terminal domain of TatA is decisive for whether or not TatB is required.

Amino Acid Sequence↗