Biomedical subjects

Markus Hecker

Publications and source records attributed to Markus Hecker.

33 records · Page 2Linked to original sources

PubMed2004

CD154/CD40-mediated expression of CD154 in endothelial cells: consequences for endothelial cell-monocyte interaction.

OBJECTIVE: CD40 ligand (CD154) expressed on activated T helper cells is a key costimulatory molecule for antigen-presenting cells expressing the corresponding receptor CD40. Moreover, CD40 stimulation in nonimmune cells, such as endothelial cells, may play an important role in atherogenesis. One gene product that is induced in endothelial cells on exposure to CD154 is CD154 itself. METHODS AND RESULTS: In human primary cultured endothelial cells, constitutive CD154 expression was virtually absent and insensitive to proinflammatory cytokines such as tumor necrosis factor alpha and/or interferon-gamma. However, CD154 expression was markedly induced, both on the mRNA and protein level, after CD40 stimulation. Moreover, CD40-positive human monocytes (THP-1 cell line) transmigrating through CD154-expressing endothelial cells responded with an increased expression of interleukin-1beta (IL-1beta) mRNA, indicative of their activation. This increase in IL-1beta expression was confirmed on the protein level and could be abrogated by prior treatment of the endothelial cells with a neutralizing anti-CD154 antibody. CONCLUSIONS: By way of CD154-induced CD154 expression, human endothelial cells thus seem capable of influencing the progression of proinflammatory reactions, including atherogenesis through activation of extravasating monocytes.

Animals↗

PubMed2004

Focal adhesion protein zyxin is a mechanosensitive modulator of gene expression in vascular smooth muscle cells.

Excessive deformation of vascular smooth muscle cells (SMCs) caused by a prolonged increase in blood pressure (eg, in hypertension) results in an adaptive remodeling of the vessel wall that is characterized by SMC hypertrophy or hyperplasia and contributes to fixation of the increase in blood pressure. The onset of this process is characterized by a unique change in gene expression in the SMC. However, thus far, no transcription factor has been identified that specifically mediates mechanosensitive gene expression in these cells. Therefore, the role of a putative mechanotransducer, the cytoskeletal protein zyxin, was investigated in rat aortic cultured SMCs. Immunofluorescence and Western blot analysis revealed that on exposure to cyclic stretch, but not to osmotic stress or treatment with proinflammatory cytokines, zyxin dissociates from focal adhesions and accumulates in the nucleus. Unlike zyxin, vinculin, another focal adhesion-associated protein, did not translocate. Moreover, antisense oligonucleotide downregulation of zyxin protein abundance suggested that zyxin accumulation in the nucleus is a prerequisite for mechanosensitive gene expression in these cells. Thus, stretch-induced endothelin B receptor expression, for example, was attenuated, whereas that of tenascin-C was augmented after zyxin suppression. The data are consistent with a role for zyxin in transducing mechanical stimuli from the cell membrane to the nucleus in vascular SMCs and in controlling the expression of mechanosensitive genes that have been implicated in hypertension-induced arterial remodeling.

Adaptation, Physiological↗

PubMed2004

Inhibition of signal transducer and activator of transcription 1 attenuates allergen-induced airway inflammation and hyperreactivity.

BACKGROUND: Transcriptional factors of the signal transducer and activator of transcription (STAT) family play an important role in orchestrating immune reactions. OBJECTIVE: The aim of the current study was to investigate the role of STAT-1 in murine allergen-induced sensitization and development of airway inflammation (AI) and airway hyperreactivity (AHR), cardinal features of bronchial asthma. METHODS: BALB/c mice were systemically sensitized to ovalbumin and challenged with ovalbumin through the airways. Decoy oligonucleotide (ODN) specific for STAT-1 was applied once locally to the airways of sensitized animals before allergen airway challenges. RESULTS: Single application of decoy ODN markedly and significantly reduced numbers of eosinophils and lymphocytes in bronchoalveolar lavage fluids compared with those seen in sensitized and challenged animals receiving mutant control ODN. Associated with this decrease in eosinophilic AI were significantly reduced levels of IL-5 in BAL fluid, of CD40 expression in peribronchial infiltrates, and of vascular cell adhesion molecule 1 expression in vascular endothelial cells, respectively. In addition, development of AHR after allergen sensitization and airway challenges was effectively abolished after local STAT-1 decoy ODN treatment. CONCLUSION: The data indicate that a decoy ODN neutralizing STAT-1 effectively inhibits allergen-induced AI and AHR, probably by attenuating upregulation of costimulatory and adhesion molecules, and suggest a significant role of STAT-1 in asthma pathology.

Allergens↗

PubMed2004

Plasma sex steroid concentrations and gonadal aromatase activities in African clawed frogs (Xenopus laevis) from South Africa.

Adult African clawed frogs (Xenopus laevis) were collected from a corn-growing region (CGR) and a non-corn-growing region (NCGR) with different exposure profiles for atrazine and related triazines. Physical, chemical, and biological parameters from the catchment areas were also measured. Frogs were surveyed for possible effects of exposure to triazine herbicides on plasma testosterone (T) and estradiol (E2) titers, gonadal aromatase activity, and gonad growth (GSI). Concentrations of both T and E2 varied among locations and were correlated to some accessory factors, such as pH, several ions, and metals. Greatest median plasma T concentrations (males: 19 ng/ml; females: 16 ng/ml) occurred in frogs inhabiting NCGR as compared to those from the CGR (males: 4 ng/ml; females: 1 ng/ml). Median E2 concentrations were also greater in frogs collected from the NCGR (males: 3 ng/ml; females: 28 ng/ml) than those in frogs from the CGR (males: 2 ng/ml; females: 5 ng/ml). Because some exposure to agricultural chemicals at both regions occurred, as did simultaneous exposures to multiple chemicals, a regression analysis was employed. Negative correlations were observed between plasma T concentrations and concentrations of atrazine, deisopropylatrazine, deethylatrazine, and tertbuthylazine in females and between T and diaminochlorotriazine in males. Estradiol in females exhibited a significant negative correlation with atrazine and deethylatrazine. No correlations were observed between gonadal aromatase activity or GSI and any of the agricultural chemicals measured. Median aromatase activities in ovaries varied among sampling sites ranging from 7 to >3000 times greater than those in males when measurable. Testicular aromatase activity was below the detection limit of the assay in male frogs at most of the sites. Although exposure to agricultural inputs did not affect aromatase activities, effects of atrazine or coapplied pesticides on sex steroid homeostasis cannot be excluded at this point.

Agriculture↗

PubMed2003

Interleukin-10 induction of nitric-oxide synthase expression attenuates CD40-mediated interleukin-12 synthesis in human endothelial cells.

Interleukin-10 (IL-10) is a potent anti-inflammatory cytokine in Th1 cell-mediated chronic inflammatory diseases such as, e.g. Crohn's disease. Moreover, IL-10 has been shown to limit the progression of atherosclerosis, presumably by influencing endothelial cell function. Here we demonstrate that under pro-inflammatory conditions expression of the human IL-10 receptor gene is enhanced in endothelial cells in vitro and in vivo. Subsequent exposure to IL-10 results in an up-regulation of both endothelial nitric-oxide synthase (NOS-3) expression and activity. Gel mobility shift analyses and decoy oligonucleotide experiments suggest that this effect of IL-10 is mediated through activation of the transcription factor STAT-3 (signal transducer and activator of transcription-3). One functional consequence of IL-10 up-regulation of NOS-3 abundance in cultured endothelial cells is the attenuation of CD154-induced IL-12 p40 expression. Moreover, CD154-induced IL-12 p40 expression is enhanced after blockade of NOS-3 activity but attenuated in the presence of exogenous nitric oxide. Increased NOS-3 expression may, thus, be one mechanism by which IL-10 exerts its anti-inflammatory effects in Th1 cell-mediated chronic inflammatory diseases.

CD40 Antigens↗

PubMed2002

3-hydroxy-3-methylglutaryl coenzyme A reductase-independent inhibition of CD40 expression by atorvastatin in human endothelial cells.

OBJECTIVE: 3-Hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors (statins) exert potent anti-inflammatory effects that are independent of their cholesterol-lowering action. We have investigated the effects of these drugs on cytokine-stimulated CD40 expression in human cultured endothelial cells and monocytes. METHODS AND RESULTS: Reverse transcription-polymerase chain reaction and Western blot analysis revealed that treatment of either cell type with atorvastatin, cerivastatin, or pravastatin (1 to 10 micromol/L) inhibited interferon-gamma plus tumor necrosis factor-alpha-stimulated CD40 expression by approximately 50%, an effect that was not reversed by the HMG-CoA reductase product mevalonic acid (400 micromol/L). In contrast, mevalonic acid prevented the inhibitory effect of atorvastatin on cytokine-stimulated vascular cell adhesion molecule-1 expression and subsequent adhesion of THP-1 monocytes to the cultured endothelial cells. Transcription factor analysis revealed an inhibition by atorvastatin of nuclear factor-kappaB plus signal transducer and activator of transcription-1-dependent de novo synthesis of interferon regulatory factor-1, governing cytokine-stimulated CD40 expression in these cells. One consequence of this statin-dependent downregulation of CD40 expression was a decrease in CD40 ligand-induced endothelial interleukin-12 expression. CONCLUSIONS: By interfering with cytokine-stimulated CD40 expression in vascular cells, statins thus seem capable of attenuating CD40 ligand-induced proinflammatory responses, including atherosclerosis. In addition, they point to the coexistence of HMG-CoA reductase-dependent and -independent effects of statins in the same cell type.

Atorvastatin↗

PubMed2002

Plasma biomarkers in fish provide evidence for endocrine modulation in the Elbe River, Germany.

Blood plasma samples were collected from wild bream (Abramis brama L.) in the Elbe River, Germany, and analyzed for the yolk protein precursor vitellogenin (VTG), a biomarker for estrogen exposure, and the sex steroids 11-ketotestosterone (11KT), testosterone (T), and 17beta-estradiol (E2) to investigate for evidence of endocrine modulation. In addition, the gonadal status and the prominence of spawning tubercles were investigated. Nine riverine sites were investigated on the Elbe that were influenced by different sources of endocrine-active substances. Bream were collected from a lake that received no domestic or industrial discharges as a control. Plasma VTG concentrations were significantly higher in male bream from the Czech border to the middle Elbe, with the highest concentrations in fish sampled at the locations near Magdeburg and downstream of Dresden (between 20 and 100 times higher than in the controls), regions that are characterized by high levels of effluent discharges into the river. Following the Elbe from this site to the sea, the concentrations of plasma VTG in males were lower than at Meissen but were still elevated above the controls. 11KT and E2 titers showed suppressions in their normal concentrations at some locations (those receiving the greatest industrial discharges). There were reciprocal relationships between inhibitory effects on gonadal growth, maturation, and plasma sex steroids and exposure to pollutants, such as organotins, pesticides, or metals. However, there was no single chemical that alone could explain the observed inhibitory effects on sexual development. The results indicate that the endocrine system in wild bream is disrupted in stretches of the Elbe River.

Animals↗

PubMed2002

CCAAT/enhancer-binding protein decoy oligodeoxynucleotide inhibition of macrophage-rich vascular lesion formation in hypercholesterolemic rabbits.

Many cytokine genes, including those encoding acute-phase proteins and immunoglobulins, share binding sites for the CCAAT/enhancer-binding protein (C/EBP) in their 5'-flanking regions, and C/EBP-related transcription factors regulate cell proliferation during terminal differentiation. Therefore, C/EBP represents an attractive target for inhibiting restenosis after balloon angioplasty. In a rabbit model of restenosis that combines balloon injury of the carotid artery with cholesterol-mediated chronic inflammation, a decoy oligodeoxynucleotide (ODN) capable of neutralizing C/EBP was administered to the site of injury for 30 minutes. Electrophoretic mobility shift analysis confirmed that C/EBP activity in decoy ODN-treated segments was virtually absent after 2 days. Morphometric analysis after 28 days revealed significant reduction (up to 50%) of neointimal formation and intravascular inflammation in decoy ODN-treated segments compared with mutant control ODN or vehicle-treated segments. In addition, de novo synthesis of endothelin-1 and the number of proliferating cell nuclear antigen-positive smooth muscle cells in the vessel wall were markedly attenuated at day 3. These findings suggest that decoy ODN-based neutralization of C/EBP may be a feasible and effective method to limit restenosis after angioplasty brought about, at least in part, by inhibiting the de novo synthesis of endothelin-1.

Animals↗

PubMed2002

Decoy oligodeoxynucleotide against activator protein-1 reduces neointimal proliferation after coronary angioplasty in hypercholesterolemic minipigs.

OBJECTIVES: We sought to demonstrate, in an appropriate animal model, that co-medication with a transcription factor-blocking agent limits restenosis after percutaneous transluminal coronary angioplasty (PTCA). BACKGROUND: Enhanced synthesis in the vessel wall of endothelin-1 (ET-1), a powerful co-mitogen for vascular smooth muscle cells, appears to be one mechanism that promotes restenosis after PTCA. Deformation-induced expression of prepro-ET-1 is governed by the transcription factor, activator protein-1 (AP-1). METHODS: An anti-AP-1 decoy oligodeoxynucleotide (dODN) strategy was devised in which the dODN-containing solution (20 nmol) was administered locally through a Dispatch catheter into the coronary arteries of hypercholesterolemic minipigs at the time of PTCA (AVE-GFX stent). RESULTS: Treatment with an AP-1 dODN, mimicking the consensus binding site of the transcription factor, significantly reduced neointimal formation in the coronary arteries of hypercholesterolemic minipigs (n = 10 to 12), compared with vehicle-treated coronary arteries, after four weeks of follow-up (neointimal area 2.64 +/- 0.33 vs. 4.81 +/- 1.04 mm(2) [mean +/- SEM]; p < 0.05). This effect was maintained after eight weeks (neointimal area 2.04 +/- 0.22 mm(2); n = 3) and correlated with a reduction in both nuclear translocation of AP-1 and ET-1 synthesis in the vessel wall 48 h after PTCA (n = 4). In contrast, an AP-1 mutant dODN, to which the transcription factor does not bind, showed no effect on neointimal formation at either time point (n = 3 to 7). Moreover, a consensus dODN directed against CCAAT/enhancer binding protein (C/EBP), another deformation-sensitive transcription factor, did not significantly affect neointimal formation after four weeks (n = 3). CONCLUSIONS: These findings demonstrate the feasibility, efficacy and specificity of the anti-AP-1 dODN approach to the treatment of restenosis, which principally but not exclusively targets deformation-induced ET-1 synthesis in the vessel wall. Provided that these findings can be extrapolated to the situation of patients with coronary artery disease, the observed extent of the inhibitory effect of the AP-1 dODN treatment suggests that this co-medication may greatly reduce the incidence of in-stent restenosis.

Angioplasty, Balloon, Coronary↗

PubMed2002

Cytokine-inducible CD40 expression in human endothelial cells is mediated by interferon regulatory factor-1.

Given the significance of CD40-CD40 ligand interactions in chronic inflammatory diseases including atherosclerosis, the transcriptional regulation of CD40 expression as a potential therapeutic target was investigated in human umbilical vein cultured endothelial cells. Exposure to interferon-gamma (IFN-gamma) plus tumor necrosis factor-alpha resulted in a marked synergistic de novo expression of CD40, which, according to electrophoretic mobility shift analysis, was attributable to activation of the transcription factors nuclear factor-kappaB (NF-kappaB), signal transducer and activator of transcription-1 (STAT-1), and interferon regulatory factor-1 (IRF-1). Subsequent time-course studies revealed that de novo synthesis of IRF-1 preceded that of CD40. Decoy oligodeoxynucleotide (ODN) neutralization of STAT-1 or IRF-1, but not of NF-kappaB, inhibited cytokine-stimulated CD40 expression by 60% at both the mRNA and protein levels, and this effect was mimicked by antisense ODN blockade of IRF-1 synthesis. In contrast, CD40 expression in response to IFN-gamma stimulation was sensitive to neutralization of STAT-1 only. These findings suggest that depending on the cytokine composition, CD40 expression in human endothelial cells under proinflammatory conditions is governed by STAT-1 either directly or indirectly through de novo synthesis of IRF-1. Moreover, decoy ODN neutralization of these transcription factors may provide a novel therapeutic option for interfering with CD40-CD40 ligand-mediated inflammatory responses in vivo.

Animals↗

PubMed2002

Atorvastatin inhibition of cytokine-inducible nitric oxide synthase expression in native endothelial cells in situ.

Animal experimental studies have demonstrated that inducible nitric oxide synthase (iNOS) expression correlates with neointima formation and is prevented by HMG-CoA reductase inhibitors (statins). In the present study we have investigated the underlying mechanism of action of these drugs in isolated segments of the rat aorta. Western blot analysis and immunohistochemistry revealed that tumour necrosis factor alpha (TNFalpha) plus interferon-gamma (IFNgamma) synergistically induce iNOS gene expression in the endothelium but not in the smooth muscle of these segments while constitutive endothelial NO synthase (eNOS) abundance was markedly reduced. Pre-treatment with 1 - 10 microM atorvastatin, cerivastatin or pravastatin decreased TNFalpha plus IFNgamma stimulated iNOS expression in the endothelium irrespective of the presence of the HMG-CoA reductase product mevalonate (400 microM). Electrophoretic mobility shift assay experiments confirmed that the combination of TNFalpha plus IFNgamma causes activation of the transcription factors STAT-1 and NF-kappaB in native endothelial cells. Neutralization of these transcription factors by employing the corresponding decoy oligonucleotides confirmed their involvement in TNFalpha plus IFNgamma stimulated iNOS expression. Translocation of both transcription factors was attenuated by atorvastatin, and this effect was insensitive to exogenous mevalonate. The present findings thus demonstrate a specific HMG-CoA reductase-independent inhibitory effect of statins, namely atorvastatin, on cytokine-stimulated transcription factor activation in native endothelial cells in situ and the subsequent expression of a gene product implicated in vascular inflammation. This effect may be therapeutically relevant and in addition provide an explanation for the reported rapid onset of action of these drugs in humans.

Animals↗

PubMed2002

L-arginine deficiency and supplementation in experimental acute renal failure and in human kidney transplantation.

BACKGROUND: The "L-arginine paradox" refers to situations where L-arginine (L-Arg) supplementation stimulates nitric oxide (NO) synthesis, despite saturating intracellular concentrations. This paradox is frequently observed in acute renal failure (ARF). First, the effects of L-Arg on renal function of rats with ARF were studied. Based on the promising results from these initial studies, the second part of our study searched for a form of ARF in humans that could be studied easily under conditions with little variance and yet was linked with endothelial dysfunction. Thus, we investigated the effects of L-Arg supplementation immediately after kidney transplantation in 54 patients. METHODS: In uranyl nitrate-induced ARF in rats the effects of L-Arg and L-NNA (inhibitor of nitric oxide synthase; NOS) on glomerular filtration rate (GFR), renal plasma flow (RPF), blood pressure (BP) and NOx (NO2- +NO3-) excretion were examined. Tissue L-Arg levels, NOS activities, immunodetection of NOS and superoxide dismutase (SOD), activities of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase and xanthine oxidase, and nitrotyrosine immunoreactive protein (NT-IR) were determined and compared to sham operated animals. Secondly, in a randomized, double-blind study, the effects of L-Arg on GFR and RPF were investigated in 54 kidney transplant recipients, receiving IV L-Arg for three days. GFR and RPF were measured on days 1, 3, 5 and 10 by scintigraphy. RESULTS: In experimental ARF, decreased RPF and GFR were associated with reduced tissue L-Arg levels, endothelial NOS-III expression, NO formation and NOx excretion. Reduction in GFR, RPF and NOx excretion were reversed upon administration of exogenous L-Arg. There also was a loss of Cu,Zn-SOD, a key enzyme against oxidative stress, and an elevation of NT-IR, an indicator of nitrosative stress and suggested marker for pathological actions of NO. However, NT-IR was not dependent on de novo NO synthesis and not related to the functional effects of l-Arg administration. In kidney transplant recipients receiving organs with a short cold ischemia time (CIT) and from young donors, that is, those with a higher likelihood of a functional endothelium, early administration of L-Arg improved renal function. CONCLUSION: Both experimental and clinical data show that \L-Arg deficiency and endothelial dysfunction are pathomechanistically relevant in ARF. The data suggest a therapeutic potential for the administration of L-Arg in ARF and kidney transplantation, at least in patients receiving kidneys with shorter CIT and from younger donors.

Acute Kidney Injury↗

PubMed2001

Cytokine-induced down-regulation of zfm1/splicing factor-1 promotes smooth muscle cell proliferation.

One hallmark of inflammation is the proliferation of bystander cells such as vascular smooth muscle cells (SMC), a process governed by growth factors and cytokines. Whereas cytokine induction of gene products promoting inflammation and proliferation is well characterized, little is known about the concomitant down-regulation of potentially counter-regulatory gene products in these cells. By employing the suppression subtractive hybridization-PCR technique, RNA isolated from rat aortic SMC treated with the cytokines interleukin-1 beta (IL-1 beta) and tumor necrosis factor alpha (TNF alpha) was subtracted from RNA of control cells. Eleven genes were identified, the expression of which fell by 44-77%. One, the transcriptional repressor splicing factor-1 or zfm1, was characterized further. Antisense oligonucleotide suppression of zfm1 protein synthesis mimicked the stimulatory effects of IL-1 beta and TNF alpha on SMC proliferation and expression of the chemokine MCP-1 and the vascular cell adhesion molecule-1. Moreover, in an in vivo mouse model of atherosclerosis, zfm1 abundance was decreased in proliferating arterial SMC. These findings suggest a role for zfm1 in controlling both proliferation and expression of pro-inflammatory gene products in SMC. Therefore, cytokine-induced down-regulation of zfm1 expression may contribute to the pathogenesis of hyperproliferative inflammatory diseases.

Animals↗

PubMed2000

Endothelium-Derived Hyperpolarizing Factor-Fact or Fiction?

The vascular endothelium releases a diffusible factor that hyperpolarizes and hence relaxes vascular smooth muscle cells predominantly through activation of Ca(2+)-dependent K(+) channels. In the coronary circulation, this endothelium-derived hyperpolarizing factor appears to be a cytochrome P(450)-derived arachidonic acid epoxide, the release of which may play a crucial role in the maintenance of coronary blood flow in arteriosclerosis.

Journal Article↗

PubMedDate not supplied

Met-RANTES inhibition of mucosal perfusion failure in acute intestinal transplant rejection - role of endothelial cell-leukocyte interaction.

Acute rejection-induced microvascular injury results in graft dysfunction, ultimately leading to graft loss. Infiltration of T cells and monocytes as a consequence of an enhanced endothelial cell-leukocyte interaction appears to play an important role in this deleterious process. Recruitment of these pro-inflammatory cells to the vessel wall is mediated by chemokines such as RANTES. Heterotopic small bowel transplantation was performed in rats with the fully allogeneic Brown Norway-Lewis strain combination and, as a control, the syngeneic Lewis-Lewis strain combination. Intravital microscopy was performed from postoperative day 1-7 in both groups. The percentages of perfused villi and villus stasis, mucosal and muscular functional capillary densities, red blood cell velocities, and finally, firm adherence of leukocytes in postcapillary submucosal venules were assessed. Syngeneic small bowel transplantation revealed homogeneous perfusion of villi and muscle layers over the whole study period. Allogeneic small bowel transplantation showed a decline in perfusion from postoperative day 1 until complete failure on postoperative day 7. This was accompanied by a continuous increase in endothelial cell-leukocyte interaction which reached a plateau on postoperative day 5. Met-RANTES treatment at 200 microg/day for 5 days markedly attenuated both the decrease in functional capillary density and the increased endothelial cell-leukocyte interaction in rats following allogeneic small bowel transplantation. We conclude that blocking chemokine receptors, thereby limiting endothelial cell-leukocyte interaction, may constitute a useful therapeutic approach to the prevention of microcirculatory perfusion failure in acute transplant rejection.

Animals↗