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Biomedical subjects

Matthias Ernst

Publications and source records attributed to Matthias Ernst.

At least 37 records · Page 2Linked to original sources
PubMed2005

Differential regulation of gastric tumor growth by cytokines that signal exclusively through the coreceptor gp130.

BACKGROUND & AIMS: We have shown that mice with a mutation in gp130 (gp130(757F/F)), the signal transducing receptor for interleukin (IL)-6 family cytokines, have chronic gastric inflammation and develop distal stomach tumors associated with deregulated phosphorylated STAT3 expression. This model recapitulates many characteristics of intestinal-type gastric cancer in humans. METHODS: To evaluate the role of IL-6 and IL-11 as ligands regulating tumor growth and submucosal invasion, we compared tumor characteristics of gp130(757F/F) mice with gp130(757F/F) mice lacking IL-6 or mature T and B cells. RESULTS: As a result of the gp130(757F/F) mutation, expression of IL-6 and IL-11 was greatly up-regulated concomitant with activation of STAT3 and development of tumors. However, the lack of IL-6 or T and B cells did not impact on tumor growth. While IL-6 did not regulate tumor growth or tumor vascularization, gp130(757F/F)/IL-6(-/-) mice showed approximately 10-20-fold more submucosal tumor invasion, reduced mononuclear inflammatory cell infiltrate, and greater IL-11 and matrix metalloproteinase (MMP)-13 and MMP-9 synthesis than gp130(757F/F) mice. Expression of MMP-13 was largely restricted to tumor-associated stroma, but MMP-9 was also expressed in polymorphonuclear cells and a subset of epithelial cells. In addition, treatment with recombinant IL-11 stimulated expression of MMP-13 and MMP-9 in stomachs of wild-type mice. CONCLUSIONS: Increased submucosal invasion in gp130(757F/F)/IL-6(-/-) mice could not be explained by increased vascularization or reduced immunosurveillance but was most likely facilitated by augmented metalloproteinase activity driven by elevated IL-11 levels.

Animals↗
PubMed2004

Mammary gland remodeling depends on gp130 signaling through Stat3 and MAPK.

The interleukin-6 (IL6) family of cytokines signals through the common receptor subunit gp130, and subsequently activates Stat3, MAPK, and PI3K. Stat3 controls cell death and tissue remodeling in the mouse mammary gland during involution, which is partially induced by IL6 and LIF. However, it is not clear whether Stat3 activation is mediated solely through the gp130 pathway or also through other receptors. This question was explored in mice carrying two distinct mutations in the gp130 gene; one that resulted in the complete ablation of gp130 and one that led to the loss of Stat3 binding sites (gp130Delta/Delta). Deletion of gp130 specifically from mammary epithelium resulted in a complete loss of Stat3 activity and resistance to tissue remodeling comparable to that seen in the absence of Stat3. A less profound delay of mammary tissue remodeling was observed in gp130Delta/Delta mice. Stat3 tyrosine and serine phosphorylation was still detected in these mice suggesting that Stat3 activation could be the result of gp130 interfacing with other receptors. Experiments in primary mammary epithelial cells and transfected COS-7 cells revealed a p44/42 MAPK and EGFR-dependent Stat3 activation. Moreover, the gp130-dependent EGFR activation was independent of EGF ligands, suggesting a cytoplasmic interaction and cross-talk between these two receptors. These experiments establish that two distinct Stat3 signaling pathways emanating from gp130 are utilized in mammary tissue.

Animals↗
PubMed2004

Low-power high-resolution solid-state NMR of peptides and proteins.

We demonstrate that two-dimensional solid-state NMR chemical-shift correlation spectra can be recorded under low-power conditions. Except for the cross-polarization period, no rf-field amplitudes above 40 kHz are used. Such experiments require the use of fast (>50 kHz) magic-angle spinning (MAS). A comparison with the high-power version of the experiment shows no general line broadening but some changes in the polarization-transfer dynamics.

Nuclear Magnetic Resonance, Biomolecular↗
PubMed2004

Rotational-resonance distance measurements in multi-spin systems.

It is demonstrated that internuclear distances can be evaluated from rotational-resonance (RR) experiments in uniformly (13)C-labelled compounds. The errors in the obtained distances are less than 10% without the need to know any parameters of the spin system except the isotropic chemical shifts of all spins. We describe the multi-spin system with a simple fictitious spin-1/2 model. The influence of the couplings to the passive spins (J and dipolar coupling) is described by an empirical constant offset from the rotational-resonance condition. Using simulated data for a three-spin system, we show that the two-spin model describes the rotational-resonance transfer curves well as long as none of the passive spins is close to a rotational-resonance condition with one of the active spins. The usability of the two-spin model is demonstrated experimentally using a sample of acetylcholine perchlorate with labelling schemes of various levels of complexity. Doubly-, triply-, and fully labelled compounds lead to strongly varying RR polarization-transfer curves but the evaluated distances using the two-spin model are identical within the expected error limits and coincide with the distance from the X-ray structure. Rotational-resonance distance measurements in fully labelled compounds allow, in particular, the measurement of weak couplings in the presence of strong couplings.

Journal Article↗
PubMed2004

Acquiring signalling specificity from the cytokine receptor gp130.

Repeated use of a relatively small number of intracellular signalling molecules specifies tissue- and cell-type-specific responses to pleiotropic-acting growth factors and cytokines. Currently, gaining a better understanding of these mechanisms is a major challenge. The IL-6 family of cytokines shares a common receptor subunit called gp130. Phenotypic comparisons of mice with amino acid knock-in substitutions that disable individual signalling modules in gp130, with knockout mice lacking ligand-specific gp130 activation or transgenic mice with constitutive gp130 activation, has led to the identification of two molecular mechanisms. One mechanism is based on differential target-gene responsiveness to signalling threshold levels transduced by either the STAT1/3 or the SHP2/ERK cascade, which are under reciprocal negative regulation and together account for the majority of intracellular gp130 signalling. The second mechanism is based on the capacity of certain cell types to integrate the often-conflicting information transduced by these two pathways, and to prevent pathological responses.

Animals↗
PubMed2004

Gastric cancer development in mice lacking the SHP2 binding site on the IL-6 family co-receptor gp130.

BACKGROUND AND AIMS: We have developed a mouse model of gastric cancer that resembles human intestinal-type adenocarcinoma. The aim of this study was to determine the identity and temporal changes in mediators of IL-6 signaling regulating tumor development. METHODS: gp130(757F/F) Mice that lack the SHP2-binding site on the IL-6 family receptor gp130 and have increased STAT 3 activity and wild-type littermates were used. Cohorts were assessed by quantitative histology and immunohistochemistry for gastric cell phenotype and proliferation markers from 4 to 40 weeks of tumor development. Northern blotting and in situ hybridization were used to quantify expression of the tumor suppressor TFF1 and the mitogens gastrin and Reg I. Expression of epidermal growth factor receptor (EGFr) and its ligands was measured by RT-PCR analysis. Age-matched differences in gene expression profiles were tested by ANOVA. RESULTS: Hyperplastic antral tumors with inflammation and ulceration were evident in gp130(757F/F) mice at 4 weeks of age and reached maximum size by 20 weeks. Tumor progression was marked by gastritis, atrophy, intestinal metaplasia, dysplasia, and submucosal invasion after 30 weeks. Both TFF1 and gastrin expression were progressively inhibited during tumorigenesis, whereas Reg I was stimulated. The EGFr and its ligands transforming growth factor (TGF)-alpha and heparin-binding EGF had increased expression corresponding to maximal tumor growth. CONCLUSIONS: gp130(757F/F) Mice rapidly develop distal stomach tumors, with loss of SHP2/Erk/AP-1 transcriptional regulation exemplified by decreased TFF1 expression and increased STAT1/3 regulated genes such as Reg I. Tumor development occurs in a hypogastrinemic environment. Balanced IL-6 signaling is required for maintaining gastric homeostasis.

Adenoma↗
PubMed2004

Imbalanced gp130-dependent signaling in macrophages alters macrophage colony-stimulating factor responsiveness via regulation of c-fms expression.

The mechanisms by which interleukin-6 (IL-6) family cytokines, which utilize the common receptor signaling subunit gp130, influence monocyte/macrophage development remain unclear. Here we have utilized macrophages devoid of either gp130-dependent STAT1/3 (gp130(Delta STAT/Delta STAT)) or extracellular signal-regulated kinases 1 and 2 (ERK1/2) mitogen-activated protein (MAP) kinase (gp130(Y757F/Y757F)) activation to assess the individual contribution of each pathway to macrophage formation. While the inhibition by IL-6 of macrophage colony-stimulating factor (M-CSF)-induced colony formation observed in gp130(wt/wt) mice was abolished in gp130(Delta STAT/Delta STAT) mice, inhibition of macrophage colony formation was enhanced in gp130(Y757F/Y757F) mice. In gp130(Delta STAT/Delta STAT) bone marrow-derived macrophages (BMMs), both IL-6- and M-CSF-induced ERK1/2 tyrosine phosphorylation was enhanced. By contrast, tyrosine phosphorylation of ERK1/2 in response to M-CSF was reduced in gp130(Y757F/Y757F) BMMs, and the pattern of ERK1/2 activation in gp130 mutant BMMs correlated with their opposing responsiveness to M-CSF-induced proliferation. When compared to the level of expression in gp130(wt/wt) BMMs, c-fms expression was elevated in gp130(Delta STAT/Delta STAT) BMMs but reduced in gp130(Y757F/Y757F) BMMs. Finally, an ERK1/2 inhibitor suppressed M-CSF-induced BMM proliferation, and this result corresponded to a reduction in c-fms expression. Collectively, these results provide a functional and causal correlation between gp130-dependent ERK MAP kinase signaling and c-fms gene activation, a finding that provides a potential mechanism underlying the inhibition of M-CSF-dependent macrophage development by IL-6 family cytokines in mice.

Animals↗
PubMed2004

Glycoprotein 130 regulates bone turnover and bone size by distinct downstream signaling pathways.

The gp130-dependent cytokines, which signal through at least two intracellular pathways, regulate osteoclast and osteoblast formation. To define their roles in regulating bone mass, we analyzed mice in which gp130 signaling via either the signal transducer and activator of transcription (STAT) 1/3 (gp130(DeltaSTAT/DeltaSTAT)) or SHP2/ras/MAPK (gp130(Y757F/Y757F)) pathway was attenuated. In gp130(DeltaSTAT/DeltaSTAT) mice, trabecular bone volume (BV/TV) and turnover were normal, but bone length was reduced by premature growth plate closure, indicating an essential role for gp130-STAT1/3 signaling in chondrocyte differentiation. In contrast, while bone size was normal in gp130(Y757F/Y757F) mice, BV/TV was reduced due to high bone turnover, indicated by high osteoclast surface/bone surface (OcS/BS) and osteoblast surface/bone surface (ObS/BS). Furthermore, generation of functional osteoclasts from bone marrow of gp130(Y757F/Y757F) mice was elevated, revealing that while gp130 family cytokines stimulate osteoclastogenesis through the osteoblast lineage, gp130, via SHP2/Ras/MAPK, inhibits osteoclastogenesis in a cell lineage-autonomous manner. Genetic ablation of IL-6 in gp130(Y757F/Y757F) mice exacerbated this osteopenia by reducing ObS/BS without affecting OcS/BS. Thus, while IL-6 is critical for high bone formation in gp130(Y757F/Y757F) mice, it is not involved in the increased osteoclastogenesis. In conclusion, gp130 is essential for normal bone growth and trabecular bone mass, with balanced regulation depending on selective activation of STAT1/3 and SHP2/ras/MAPK, respectively. Furthermore, the latter pathway can directly inhibit osteoclastogenesis in vivo.

Animals↗
PubMed2003

NMR spectra of a microcrystalline protein at 30 kHz MAS.

Proteins are not always available in amounts desirable for solid-state magic-angle spinning (MAS) nuclear-magnetic resonance (NMR) spectroscopy. To maximize the signal-to-noise ratio achievable with small samples, the filling factor must be optimized by using small-diameter MAS rotors. These rotors have the added benefit of allowing higher radio frequency field amplitudes during polarization transfer steps and during decoupling periods as well as allowing higher spinning frequencies. We demonstrate the advantages of relatively fast MAS (30 kHz using a 2.5 mm rotor) compared to MAS at 12 kHz for the 10.4 kDa model protein Crh with 93 residues and show that the signal-to-noise ratio in two-dimensional correlation spectra can be significantly improved by taking advantage of optimized pulse sequences available with rapid MAS.

Bacillus subtilis↗
PubMed2003

Determination of internuclear distances in uniformly labeled molecules by rotational-resonance solid-state NMR.

Rotational-resonance magic-angle spinning NMR experiments are frequently used to measure dipolar couplings and to determine internuclear distances. So far most measurements were performed on samples containing isolated spin pairs. Thus, extensive structure elucidation, for example in biomolecules, requires the preparation of a whole set of doubly labeled samples. Here, we describe the analysis of the rotational-resonance polarization-exchange curves obtained from a single, uniformly labeled sample. It is shown experimentally that, at a magnetic field of 14.09 T, the rotational-resonance conditions in uniformly (13)C-labeled threonine are sufficiently narrow to permit the measurement of five distances between the four carbon spins with an accuracy of better than 10%. The polarization-exchange curves are analyzed using a modified two-spin model consisting of the two active spins. The modified model includes an additional offset in the final polarization, which comes from the coupling to the additional, passive, spins. The validity of this approach is experimentally verified for uniformly (13)C-labeled threonine. The broader applicability of such a model is demonstrated by numerical simulations which quantify the errors as a function of the most relevant parameters in the spin system.

Carbon Isotopes↗
PubMed2003

The role of gp130/IL-6 cytokines in the development of pulmonary fibrosis: critical determinants of disease susceptibility and progression?

Cryptogenic fibrosing alveolitis (CFA), also known as idiopathic pulmonary fibrosis (IPF), is the end stage of a heterogeneous group of disorders in which the deposition of excessive amounts of collagen results in the loss of lung function and premature death. The molecular mechanisms underlying the disease are unknown. Accordingly, there is much debate as to whether pulmonary fibrosis is the end result of (1) a chronic inflammatory process or (2) a disturbance in normal epithelium-fibroblast cross talk, or both. In addition, it appears increasingly likely that there is a genetic component in the development of pulmonary fibrosis. The IL-6 cytokine family is a group of pleiotropic mediators produced by a variety of cells in response to a inflammatory stimuli. These cytokines are grouped together on the basis of weak structural homology, overlapping functions, and shared use of the transmembrane glycoprotein beta-subunit gp130 as part of their multimeric receptor complexes. Activation of these receptor complexes results in the recruitment and phosphorylation of specific transcription factors. In addition, membrane-proximal tyrosine residues act as docking sites for molecules involved in the activation of extracellular signal-related kinase (ERK). However, studies in genetically engineered mice that overexpress members of this family have shown that while overlapping biological activities exist, there are effects specific to individual cytokines. Data from both human and animal studies are now emerging to suggest that members of this cytokine family play an important role in the pathogenesis of fibroproliferative diseases and thus represent a novel group of cytokines implicated in pulmonary fibrosis. Importantly, manipulation of signaling pathways activated by these cytokines may suppress fibrosis but leave innate cellular mechanisms necessary for host defense largely untouched. This may provide guides for the development of novel pharmacological treatment for fibroproliferative diseases.

Animals↗
PubMed2003

Heteronuclear spin decoupling in solid-state NMR under magic-angle sample spinning.

Achieving high spectral resolution is an important prerequisite for the application of solid-state NMR to biological molecules. Higher spectral resolution allows to resolve a larger number of resonances and leads to higher sensitivity. Among other things, heteronuclear spin decoupling is one of the important factors which determine the resolution of a spectrum. The process of heteronuclear spin decoupling under magic-angle sample spinning is analyzed in detail. Continuous-wave RF irradiation leads only in a zeroth-order approximation to a full decoupling of heteronuclear spin systems in solids under magic-angle spinning (MAS). In a higher-order approximation, a cross-term between the dipolar-coupling tensor and the chemical-shielding tensor is reintroduced, providing a scaled coupling term between the heteronuclear spins. In strongly coupled spin systems this second-order recoupling term is partially averaged out by the proton spin-diffusion process, which leads to exchange-type narrowing of the line by proton spin flips. This process can be described by a spin-diffusion type superoperator, allowing the efficient simulation of strongly coupled spin systems under heteronuclear spin decoupling. Low-power continuous-wave decoupling at fast MAS frequencies offers an alternative to high-power irradiation by reversing the order of the averaging processes. At fast MAS frequencies low-power continuous-wave decoupling leads to significantly narrower lines than high-power continuous-wave decoupling while at the same time reducing the power dissipated in the sample by several orders of magnitude. The best decoupling is achieved by multiple-pulse sequences at high RF fields and under fast MAS. Two such sequences, two-pulse phase-modulated decoupling (TPPM) and X-inverse-X decoupling (XiX), are discussed and their properties analyzed and compared.

Ammonium Chloride↗
PubMed2003

Low-power XiX decoupling in MAS NMR experiments.

Low-power XiX proton decoupling under fast magic-angle spinning is introduced. The method is applicable if the MAS frequency exceeds the proton-proton interactions. For rigid organic solids this is the case for MAS frequencies above approximately 40 kHz. It is shown that the quality of the decoupling as well as the sensitivity to frequency offsets can be improved compared to low-power continuous-wave decoupling. The decoupling efficiency is somewhat reduced compared to optimized high-power decoupling: in a peptide sample investigated at an MAS frequency of 50 kHz a loss of about 10% in signal intensity for CH3 and CH groups, and of about 40% for CH2 groups was observed. Taking into consideration, that the rf amplitude in the low-power XiX was about 15 times lower than in high-power XiX decoupling, such a reduction in line intensity is sometimes tolerable.

Carbon Isotopes↗
PubMed2002

Response of Apc(min) and A33 (delta N beta-cat) mutant mice to treatment with tea, sulindac, and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP).

There is growing interest in the potential health benefits of tea, and a recent report described the potent antimutagenic activity of white tea in comparison with green tea against several heterocyclic amines, including 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) [Mutat. Res. 495 (2001) 61]. We compared the inhibitory effects of white and green teas with sulindac, a nonsteroidal anti-inflammatory agent, in two different mouse models of intestinal tumorigenesis. In the Apc(min) mouse, white and green teas given at human-relevant concentrations (1.5% w/v, 2-min brew), and sulindac (80 ppm in the drinking water), each suppressed polyp formation by approximately 50%, and the combination of white tea plus sulindac was more effective than either treatment alone (P=0.05). Mice expressing an N-terminally truncated, oncogenic version of beta-catenin (A 33(delta N beta-cat) mutant mice) developed colonic aberrant crypt foci (ACF) spontaneously, but PhIP treatment increased the incidence and number of ACF per colon. In the normal-looking intestinal mucosa of Apc(min) and A 33(delta N beta-cat) mice, white tea plus sulindac treatment markedly attenuated the expression of beta-catenin protein, and this was recapitulated in vitro in cells transiently transfected with beta-catenin plus Tcf-4 and treated with tea or the major tea polyphenol epigallocatechin-3-gallate (EGCG). Expression of a beta-catenin/Tcf reporter was inhibited by EGCG in the transfected cells, and the beta-catenin/Tcf target genes cyclin D1 and c-jun were downregulated in vivo by tea plus sulindac treatment. Collectively, the data support a chemopreventive role for tea and sulindac against intermediate and late stages of colon cancer, via effects on the beta-catenin/Tcf signaling pathway.

Adenomatous Polyposis Coli Protein↗
PubMed2002

Reciprocal regulation of gastrointestinal homeostasis by SHP2 and STAT-mediated trefoil gene activation in gp130 mutant mice.

The intracellular signaling mechanisms that specify tissue-specific responses to the interleukin-6 (IL-6) family of cytokines are not well understood. Here, we evaluated the functions of the two major signaling pathways, the signal transducers and activators of transcription 1 and 3 (STAT1/3) and the Src-homology tyrosine phosphatase 2 (SHP2)-Ras-ERK, emanating from the common signal transducer, gp130, in the gastrointestinal tract. Gp130(757F) mice, with a 'knock-in' mutation abrogating SHP2-Ras-ERK signaling, developed gastric adenomas by three months of age. In contrast, mice harboring the reciprocal mutation ablating STAT1/3 signaling (gp130(Delta STAT)), or deficient in IL-6-mediated gp130 signaling (IL-6(-/-) mice), showed impaired colonic mucosal wound healing. These gastrointestinal phenotypes are highly similar to the phenotypes exhibited by mice deficient in trefoil factor 1 (pS2/TFF1) and intestinal trefoil factor (ITF)/TFF3, respectively, and corresponded closely with the capacity of the two pathways to stimulate transcription of the genes encoding pS2/TFF1 and ITF/TFF3. We propose a model whereby mucosal wound healing depends solely on activation of STAT1/3, whereas gastric hyperplasia ensues when the coordinated activation of the STAT1/3 and SHP2-Ras-ERK pathways is disrupted.

Adenoma↗
PubMed2002

Constitutive activation of the SRC family kinase Hck results in spontaneous pulmonary inflammation and an enhanced innate immune response.

To identify the physiological role of Hck, a functionally redundant member of the Src family of tyrosine kinases expressed in myelomonocytic cells, we generated Hck(F/F) "knock-in" mice which carry a targeted tyrosine (Y) to phenylalanine (F) substitution of the COOH-terminal, negative regulatory Y(499)-residue in the Hck protein. Unlike their Hck(-/-) "loss-of-function" counterparts, Hck(F/F) "gain-of-function" mice spontaneously acquired a lung pathology characterized by extensive eosinophilic and mononuclear cell infiltration within the lung parenchyma, alveolar airspaces, and around blood vessels, as well as marked epithelial mucus metaplasia in conducting airways. Lungs from Hck(F/F) mice showed areas of mild emphysema and pulmonary fibrosis, which together with inflammation resulted in altered lung function and respiratory distress in aging mice. When challenged transnasally with lipopolysaccharide (LPS), Hck(F/F) mice displayed an exaggerated pulmonary innate immune response, characterized by excessive release of matrix metalloproteinases and tumor necrosis factor (TNF)alpha. Similarly, Hck(F/F) mice were highly sensitive to endotoxemia after systemic administration of LPS, and macrophages and neutrophils derived from Hck(F/F) mice exhibited enhanced effector functions in vitro (e.g., nitric oxide and TNFalpha production, chemotaxis, and degranulation). Based on the demonstrated functional association of Hck with leukocyte integrins, we propose that constitutive activation of Hck may mimic adhesion-dependent priming of leukocytes. Thus, our observations collectively suggest an enhanced innate immune response in Hck(F/F) mice thereby skewing innate immunity from a reversible physiological host defense response to one causing irreversible tissue damage.

Amino Acid Substitution↗