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Biomedical subjects

C Arnold Spek

Publications and source records attributed to C Arnold Spek.

At least 19 recordsLinked to original sources
PubMed2006

Hedgehog morphogen in cardiovascular disease.

In this review, we focus on the basic biology of the important developmental Hedgehog (Hh) protein family, its general function in development, pathway mechanisms, and gene discovery and nomenclature. Hh function in cardiovascular development and recent findings concerning Hh signaling in ischemia models are discussed in more detail, and future perspectives are proposed. In light of the recent discovery of Hh transport by insect lipophorin, we also hypothesize a role for low-density lipoprotein (LDL) in mammalian Hh transport, creating a surprising role for LDL in cardiovascular disease.

Animals↗
PubMed2006

Low dose endotoxin priming is accountable for coagulation abnormalities and organ damage observed in the Shwartzman reaction. A comparison between a single-dose endotoxemia model and a double-hit endotoxin-induced Shwartzman reaction.

The clinical response of sepsis to a systemic inflammatory infection may be complicated by disseminated intravascular coagulation or DIC. In order to experimentally study the syndrome of DIC, we aimed for a severe sepsis model complicated by disseminated coagulation. Most-simplified-experimental models describing coagulation abnormalities as a consequence of sepsis are based on single dose endotoxemia. The so called-Shwartzman reaction contrarily, is elicited by a low dose endotoxin priming followed by an LPS challenge and is characterized by pathological manifestations that represent the syndrome of DIC. In order to investigate whether the Shwartzman reaction is superior to a single endotoxin challenge as a model for sepsis-induced DIC and to determine what the pathological effect is of an encounter of low endotoxin prior to an LPS challenge, we undertook the present study. In this study we demonstrate that low-dose endotoxin priming prior to an LPS challenge in the Shwartzman reaction is accountable for micro-vascular thrombosis in lung and liver and subsequent (multi-) organ failure, not observed after a single-dose endotoxin challenge, which indicates that the Shwartzman reaction is well suited-model to study sepsis-induced DIC adversities. Remarkably, only minor differences in the innate immune response were established between the single-dose endotoxin challenge and the Shwartzman reaction.

Journal Article↗
PubMed2006

Nuclear receptors Nur77, Nurr1, and NOR-1 expressed in atherosclerotic lesion macrophages reduce lipid loading and inflammatory responses.

OBJECTIVE: Atherosclerosis is an inflammatory disease in which macrophage activation and lipid loading play a crucial role. In this study, we investigated expression and function of the NR4A nuclear receptor family, comprising Nur77 (NR4A1, TR3), Nurr1 (NR4A2), and NOR-1 (NR4A3) in human macrophages. METHODS AND RESULTS: Nur77, Nurr1, and NOR-1 are expressed in early and advanced human atherosclerotic lesion macrophages primarily in areas of plaque activation/progression as detected by in situ-hybridization and immunohistochemistry. Protein expression localizes to the nucleus. Primary and THP-1 macrophages transiently express NR4A-factors in response to lipopolysaccharide and tumor necrosis factor alpha. Lentiviral overexpression of Nur77, Nurr1, or NOR-1 reduces expression and production of interleukin (IL)-1beta and IL-6 proinflammatory cytokines and IL-8, macrophage inflammatory protein-1alpha and -1beta and monocyte chemoattractant protein-1 chemokines. In addition, NR4A-factors reduce oxidized-low-density lipoprotein uptake, consistent with downregulation of scavenger receptor-A, CD36, and CD11b macrophage marker genes. Knockdown of Nur77 or NOR-1 with gene-specific lentiviral short-hairpin RNAs resulted in enhanced cytokine and chemokine synthesis, increased lipid loading, and augmented CD11b expression, demonstrating endogenous NR4A-factors to inhibit macrophage activation, foam-cell formation, and differentiation. CONCLUSIONS: NR4A-factors are expressed in human atherosclerotic lesion macrophages and reduce human macrophage lipid loading and inflammatory responses, providing further evidence for a protective role of NR4A-factors in atherogenesis.

Atherosclerosis↗
PubMed2006

Signal transduction induced by activated protein C: no role in protection against sepsis?

The anticoagulant activated protein C (APC) is historically known as a risk factor for venous thrombosis. However, after the positive results of the protein C worldwide evaluation in severe sepsis (PROWESS) trial, which showed that APC was the first drug that considerably reduced sepsis-related mortality, APC is considered a pleiotropic protein with both anticoagulant and anti-inflammatory properties. In addition, in vitro studies have suggested that APC-induced intracellular signal transduction is a potential mechanism by which APC might be protective against sepsis. Recently, however, the efficacy of APC in sepsis has been argued, and also the extent to which the signal transduction capacity of APC contributes to its pro-survival effects is debated. Here, we review the role of APC in the body natural defense against sepsis and discuss the mechanism by which APC might act at a cellular level.

Animals↗
PubMed2006

Hedgehog turns lipoproteins into Janus-faced particles.

Hedgehog is an important morphogenetic signal during embryonic development. The molecule contains several hydrophobic moieties, including cholesterol and palmitoyl groups, apparently incompatible with long-range functioning. Very recent research, however, performed in the fruitfly Drosophila melanogaster, demonstrated that hedgehog can be carried by insect lipoproteins which strongly resemble mammalian low-density lipoprotein (LDL), enabling the morphogen to act at long distances from its source. Such long-range signaling of hedgehog is in perfect agreement with current insight that in mammals hedgehog signaling is not terminated after gestation has been completed but remains active throughout life, mediating among others revascularization of adult ischemic cardiac tissue. In agreement, hedgehog gene therapy is highly effective in experimental rodents for limiting the damage after coronary occlusion. If indeed hedgehog is transported on lipoproteins in mammals, an interesting change of paradigm might emerge for LDL. No longer would LDL merely be bad news for the progression of atherosclerotic plaques, as it also conveys a revascularization and tissue salvage signal in the form of hedgehog.

Animals↗
PubMed2006

Local activation of the tissue factor-factor VIIa pathway in patients with pneumonia and the effect of inhibition of this pathway in murine pneumococcal pneumonia.

OBJECTIVE: The tissue factor (TF)-factor VIIa (FVIIa) complex not only is essential for activation of blood coagulation but also affect the inflammatory response during sepsis. The objective of this study was to determine the role of TF-FVIIa in pneumonia caused by Streptococcus pneumoniae, the most important causative organism in community-acquired pneumonia and a major cause of sepsis. DESIGN: A controlled, in vivo laboratory study. SETTING: Research laboratory of a health sciences university. PATIENTS AND SUBJECTS: Patients with unilateral community-acquired pneumonia and female BALB/c mice. INTERVENTIONS: Bilateral bronchoalveolar lavage was performed in patients with community-acquired pneumonia. In mice, pneumonia was induced by intranasal inoculation with S. pneumoniae with or without concurrent inhibition of TF-FVIIa by subcutaneous injections of recombinant nematode anticoagulant protein (rNAPc2). MEASUREMENTS AND MAIN RESULTS: Patients with unilateral community-acquired pneumonia demonstrated elevated concentrations of FVIIa, soluble TF, and thrombin-antithrombin complexes in bronchoalveolar lavage fluid obtained from the infected site compared with the uninfected site. Mice with S. pneumoniae pneumonia displayed increased TF expression and fibrin deposits in lungs together with elevated thrombin-antithrombin complex levels in bronchoalveolar lavage fluid; inhibition of TF-FVIIa by rNAPc2 attenuated the procoagulant response in the lung but did not affect host defense, as reflected by an unaltered outgrowth of pneumococci and an unchanged survival. CONCLUSIONS: These data suggest that TF-FVIIa activity contributes to activation of coagulation in the lung during pneumococcal pneumonia but does not play an important role in the antibacterial host defense in this murine model.

Adult↗
PubMed2006

Role of the factor V Leiden mutation in septic peritonitis assessed in factor V Leiden transgenic mice.

OBJECTIVE: The factor V Leiden (FVL) mutation (Arg506Glu) results in the production of an FV protein that when activated is relatively resistant to inactivation by activated protein C and thereby leads to predisposition to thrombosis. The rather high prevalence of the FVL mutation in the general population prompted speculation about a potential survival benefit for individuals carrying the FVL allele. Indeed, both clinical and experimental animal data suggest that a heterozygous FVL genotype might protect against the lethal consequences of sepsis. We sought to confirm the survival advantage of heterozygous FVL mice in septic disease. DESIGN: Controlled animal experiment. SETTING: Academic research laboratory. SUBJECTS: Wild-type, heterozygous, and homozygous FVL mice subjected to 1 x 10 live bacteria as model for septic peritonitis. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: The intraperitoneal injection of E. coli led to growth and dissemination of bacteria and provoked an inflammatory response as evident from elevated cytokine levels (interleukin-6, interleukin-10, and tumor necrosis factor-alpha), induced thrombin-antithrombin complex levels, increased granulocyte influx into the peritoneal cavity, liver necrosis, and adhesion of leukocytes to the vessel wall, resulting in approximately 50% mortality after 72 hrs. The FVL genotype had no significant effect on bacterial outgrowth, markers of inflammation (i.e., tumor necrosis factor-alpha levels of 152 [96.2-200], 152 [99.7-1745], and 110 [99.7-177] pg/mL in peritoneal lavage fluid at t = 20 hrs for wild-type, heterozygous, and homozygous FVL mice, respectively), thrombin generation (i.e., thrombin-antithrombin complex levels of 19.9 [9.31-37.4], 10.4 [6.55-15.8], and 12.6 [8.24-29.0] ng/mL in peritoneal lavage fluid at t = 6 hrs for wild-type, heterozygous, and homozygous FVL mice, respectively), and/or survival (50%, 36%, and 50% for wild-type, heterozygous, and homozygous FVL mice, respectively). CONCLUSIONS: The FVL allele has no beneficial effect in mouse septic peritonitis, and the general protective effect of FVL in sepsis needs further investigation.

Animals↗
PubMed2006

Repression of smoothened by patched-dependent (pro-)vitamin D3 secretion.

The developmentally important hedgehog (Hh) pathway is activated by binding of Hh to patched (Ptch1), releasing smoothened (Smo) and the downstream transcription factor glioma associated (Gli) from inhibition. The mechanism behind Ptch1-dependent Smo inhibition remains unresolved. We now show that by mixing Ptch1-transfected and Ptch1 small interfering RNA-transfected cells with Gli reporter cells, Ptch1 is capable of non-cell autonomous repression of Smo. The magnitude of this non-cell autonomous repression of Smo activity was comparable to the fusion of Ptch1-transfected cell lines and Gli reporter cell lines, suggesting that it is the predominant mode of action. CHOD-PAP analysis of medium conditioned by Ptch1-transfected cells showed an elevated 3beta-hydroxysteroid content, which we hypothesized to mediate the Smo inhibition. Indeed, the inhibition of 3beta-hydroxysteroid synthesis impaired Ptch1 action on Smo, whereas adding the 3beta-hydroxysteroid (pro-)vitamin D3 to the medium effectively inhibited Gli activity. Vitamin D3 bound to Smo with high affinity in a cyclopamine-sensitive manner. Treating zebrafish embryos with vitamin D3 mimicked the smo(-/-) phenotype, confirming the inhibitory action in vivo. Hh activates its signalling cascade by inhibiting Ptch1-dependent secretion of the 3beta-hydroxysteroid (pro-)vitamin D3. This action not only explains the seemingly contradictory cause of Smith-Lemli-Opitz syndrome (SLOS), but also establishes Hh as a unique morphogen, because binding of Hh on one cell is capable of activating Hh-dependent signalling cascades on other cells.

Animals↗
PubMed2006

Ethyl pyruvate exerts combined anti-inflammatory and anticoagulant effects on human monocytic cells.

Sepsis is characterized by a concurrent activation of inflammation and coagulation. Recently, recombinant human activated protein C was shown to decrease mortality in patients with severe sepsis presumably due to a combined anti-inflammatory and anticoagulant effect. These promising findings led to a search for other products that influence both the inflammatory and the procoagulant response to severe infection. Ethyl pyruvate (EP) was recently identified as an experimental anti-inflammatory agent during endotoxemia and sepsis. The aim of the present study was to investigate whether EP influences coagulation besides its anti-inflammatory effects. For this we investigated the effects of EP on the expression and function of tissue factor (TF), the principal initiator of coagulation activation in sepsis, in human monocytic (THP-1) cell cultures. EP dose-dependently inhibited the production of tumor necrosis factor (TNF)-alpha, macrophage inflammatory protein (MIP)-1alpha and MIP-1beta by lipopolysaccharide (LPS)-stimulated THP-1 cells at mRNA and protein level, thereby confirming its anti-inflammatory properties in this in-vitro system. In addition, EP dose-dependently attenuated the increases in TF mRNA levels, TF-protein-surface expression and cell-surface-associated TF activity in LPS-stimulated THP-1 cells. These results demonstrate for the first time that EP is a compound with combined anti-inflammatory and anticoagulant effects.

Anti-Inflammatory Agents↗
PubMed2005

Antiinflammatory effects of salmeterol after inhalation of lipopolysaccharide by healthy volunteers.

RATIONALE: Salmeterol is a beta2-adrenoreceptor agonist used in the treatment of obstructive pulmonary disease. Salmeterol inhibits inflammatory responses by neutrophils and mononuclear cells in vitro and in mouse models of lung inflammation in vivo. OBJECTIVE: To determine the effect of salmeterol on LPS-induced lung inflammation in humans. METHODS: Thirty-two healthy subjects were enrolled in a single-blinded, placebo-controlled study. Subjects inhaled 100 microg salmeterol or placebo (t=-0.5 h) followed by 100 microg LPS or normal saline (t=0 h; n=8/group). Measurements were performed in bronchoalveolar lavage fluid and purified alveolar macrophages obtained 6 h post-challenge. MEASUREMENTS AND MAIN RESULTS: Inhalation of LPS was associated with neutrophil influx, neutrophil degranulation (myeloperoxidase, bactericidal/permeability-increasing protein and elastase), release of cytokines (tumor necrosis factor alpha and interleukin 6) and chemokines (interleukin 8, epithelial cell-derived neutrophil attractant 78, macrophage inflammatory proteins 1alpha and 1beta), activation of alveolar macrophages (upregulation of HLA-DR and CD71; enhanced expression of mRNAs for 13 different mediators of inflammation), and protein leakage (all p<0.05 vs. placebo/saline). Pretreatment with salmeterol inhibited LPS-induced neutrophil influx, neutrophil degranulation (myeloperoxidase), tumor necrosis factor alpha release, and HLA-DR expression (all p<0.05 vs. placebo/LPS), while not significantly influencing other responses. CONCLUSION: Salmeterol exerts antiinflammatory effects in the pulmonary compartment of humans exposed to LPS.

Adrenergic beta-Agonists↗
PubMed2005

Low molecular weight heparin attenuates multiple organ failure in a murine model of disseminated intravascular coagulation.

OBJECTIVE: Bacterial sepsis causes widespread vascular inflammation that frequently leads to disseminated intravascular coagulation (DIC). Although intravascular coagulation contributes to organ failure, it is often debated whether anticoagulant therapy produces any beneficial effects in patients with DIC. The aim of this study was to document potential beneficial effects of low molecular weight heparin (LMWH) in a lipopolysaccharide-induced DIC model. DESIGN: Controlled animal experiment combined with an in vitro laboratory study. SETTING: Academic research laboratory. SUBJECTS: C57BL/6 mice subjected to two injections of Serratia Marcescens lipopolysaccharide (LPS) resulting in the generalized Shwartzman's reaction as a model for DIC. INTERVENTIONS: LMWH (5 IU of anti-Xa activity) or saline was administered before both LPS injections and 10 hrs after the first exposure to LPS. To test the effect of LMWH on LPS-driven monocyte inflammatory responses, a human monocyte-human umbilical vein endothelial cell co-culture was used to determine E-selectin expression as a marker of monocyte adherence. MEASUREMENTS AND MAIN RESULTS: In our murine DIC model, LMWH had no effect on markers of inflammation. In addition, no effect of LMWH was detected on monocyte adherence in the human monocyte-human umbilical vein endothelial cell co-culture. Organ damage, contrarily, was significantly reduced as determined by hepatic necrosis (p < .05), lung epithelial protein leakage (p < .05), and creatinine release from kidneys into plasma (p < .01). LMWH protection from organ failure resulted in an increase in survival (p = .06) in this model for DIC. CONCLUSIONS: These results demonstrate the significance of blood coagulation in the progression of DIC and hint at a beneficial role for LMWH anticoagulation in the management of DIC.

Animals↗
PubMed2004

High throughput mRNA profiling highlights associations between myocardial infarction and aberrant expression of inflammatory molecules in blood cells.

Studies on the role of inflammation in cardiovascular disease focus on surrogate markers like plasma levels of C-reactive protein or interleukins that are affected by several factors. In this study we employ an approach in which the inflammatory mRNA profile of leucocytes is measured directly in a multigene system. We investigated the mRNA profile for 35 inflammatory markers in blood samples in a case-control study including 524 men with a history of myocardial infarction and 628 control subjects. Compared with controls, patients showed mRNA profiles with increased levels of most inflammatory mRNAs. The 2 most prominent mRNA risk indicators encoded the secreted protein macrophage migration inhibitory factor (crude odds ratio [OR], 3.4 for the highest quartile versus the lowest quartile (95% confidence interval [CI95], 2.3-4.9), and the intracellular regulator proteinase inhibitor 9 (OR, 2.5 for the highest versus the lowest quartile (CI95, 1.8-3.5), both showing an increase in odds ratio with increasing quartiles. Leucocytes in the blood of patients with myocardial infarction are more active in transcription of inflammatory genes, as evidenced by mRNA profiling. These data support the hypothesis that an inflammatory response involving leucocytes plays a role in the pathogenesis of myocardial infarction.

Adult↗
PubMed2004

Inhibition of the tissue factor/factor VIIa pathway does not influence the inflammatory or antibacterial response to abdominal sepsis induced by Escherichia coli in mice.

BACKGROUND: Anticoagulants have gained increasing attention for the treatment of sepsis. Inhibition of the tissue factor (TF)/factor (F) VIIa pathway has been shown to attenuate the activation of coagulation and to prevent death in a primate model of sepsis caused by gram-negative bacteria. METHODS: To determine the role of the TF/FVIIa complex in the host response to peritonitis, mice received an intraperitoneal injection of live Escherichia coli with or without concurrent treatment with recombinant nematode anticoagulant protein c2 (rNAPc2), a selective inhibitor of the TF/FVIIa pathway. RESULTS: Peritonitis was associated with an increase in the expression of TF at the tissue level and activation of coagulation, as reflected by elevated levels of thrombin-antithrombin complexes and by increased fibrin(ogen) deposition in the liver and lungs. rNAPc2 strongly attenuated this procoagulant response but did not influence the inflammatory response (histopathology, leukocyte recruitment to the peritoneal cavity, and cytokine and chemokine levels). Moreover, rNAPc2 did not alter bacterial outgrowth locally or dissemination of the infection, and survival was not different between rNAPc2-treated mice and control mice. CONCLUSIONS: These data suggest that TF/FVIIa activity contributes to the activation of coagulation during E. coli peritonitis but does not play a role in the inflammatory response or antibacterial host defense.

Animals↗
PubMed2004

FVIIa:TF induces cell survival via G12/G13-dependent Jak/STAT activation and BclXL production.

Tissue factor (TF), apart from activating the extrinsic pathway of the blood coagulation, is a principal regulator of embryonic and oncogenic angiogenesis, inflammation, leukocyte reverse transmigration, and tumor progression. It has become clear that these events are mediated by intracellular signal transduction elicited by TF/factor VIIa (FVIIa) interaction, but the details of this signaling remain largely obscure. In this study, we show that FVIIa/TF-interaction produces STAT5 phosphorylation, STAT5 nuclear translocation and transactivation of a STAT5 reporter construct. FVIIa-dependent STAT5 activation was dependent on FVIIa proteolytic activity but not on generation of the downstream coagulation factors Xa and thrombin, nor on the TF cytoplasmic domain. FVIIa-induced STAT5 phosphorylation was dependent on functional G12/G13 class G proteins and Jak2 activity, but not Jak1 or Tyk2. Finally, we show that FVIIa leads to cell survival through a Jak2/STAT5-dependent production of the antiapoptotic STAT5 target Bcl(XL) as well as Jak2-dependent activation of the antiapoptotic protein PKB. In conclusion, our results show that FVIIa induces cell survival through STAT5-dependent Bcl(XL) production and Jak2-dependent activation of PKB. Finally, we demonstrated for the first time that TF/FVIIa-signal transduction is dependent on G12/G13 class G proteins.

Active Transport, Cell Nucleus↗
PubMed2004

Coagulation factors VIIa and Xa inhibit apoptosis and anoikis.

The molecular mechanisms enabling cancer cells to survive loss-of-adhesion-induced apoptosis in the early phases of metastasis remain largely obscure. Interestingly, the overexpression of tissue factor (TF) on tumor cells is associated with successful metastasis and it has now become clear that coagulation factor VIIa (FVIIa), the natural binding partner of TF induces signal transduction in TF-expressing cells. Hence, we investigated the effects of FVIIa-TF interaction on cell survival. We observed that FVIIa, at physiologically relevant concentrations, inhibits cell death and caspase-3 activation induced by serum deprivation and loss of adhesion (lack of integrin signaling) in TF-overexpressing cells, but not in non-TF-expressing cells. This FVIIa effect was not dependent on the formation of the downstream coagulation products FXa or thrombin and was inhibited using an active site-blocked form of FVIIa (FVIIai). FVIIa incubation resulted in the prolonged activation of both the phosphatidylinositide-3-(OH) kinase and p42/p44 MAP kinase pathways, and studies employing pharmacological inhibitors revealed that both the pathways are required for FVIIa-induced cell survival and inhibition of caspase-3 activity. Finally, TF:FVIIa-induced FXa generation dramatically increased cell survival. We propose that FVIIa-induced cell survival may explain why overexpression of TF is associated with successful metastasis.

Animals↗
PubMed2004

Hedgehog: an unusual signal transducer.

Hedgehog proteins are of pivotal importance for development and maintenance of tissue patterns in adult organisms. Despite the role of Hedgehogs in differentiation and tumorigenesis, signal transduction of Hedgehog remains a relatively uncharted area of signalling biochemistry. For proper Hedgehog distribution into tissues, two highly unusual covalent modifications are necessary, palmitoylation of a secreted protein and the attachment of a cholesterol group, making Hedgehog the only established sterolated protein in nature. Hedgehog exerts its function via two membrane-bound receptors, Patched and Smoothened; presumably, Patched transports a cholesterol derivate out of cells which inhibits Smoothened. Binding of Hedgehog to Patched impedes this proposed pump function and thus Inhibition of Smoothened, which leads to expression of genetic Hedgehog targets via relief of transcriptional repression. These atypical features make Hedgehog physiology unique in biology and may explain why this field has attracted such significant attention.

Animals↗
PubMed2004

Tissue factor: from 'just one of the coagulation factors' to a major player in physiology.

As early as the nineteenth century, it was recognized that tissues exhibit clot-promoting activity. The realization that a single protein was responsible for this procoagulant effect led to the discovery of tissue factor (TF), but for many years it was thought that TF activity was restricted to the activation of an auxiliary pathway that had little biological relevance. Research over recent decades has greatly improved our understanding of TF function, and today the protein is recognized to be the primary biological initiator of the coagulation cascade. Furthermore, it is now clear that TF is a major player in several other physiological processes. We present recent data suggesting that TF is vital not only for effective haemostasis, but also for angiogenesis and tumour metastasis, for embryonic development, and for induction of a pro-inflammatory response. These advances in our knowledge of TF function will improve our understanding of several pathophysiological conditions, and may expand the therapeutic options available for their treatment.

Animals↗