ABSTRACT
BACKGROUND: Cardiogenic shock (CS) is the leading cause of death in patients admitted for acute myocardial infarction (MI). Despite the recent advances in reperfusion and medical treatment mortality remains unacceptably high. Whether cells of the blood compartment in CS-patients are activated and release microparticles (cMPs) that may be both messengers and biomarkers of cell damage is not known. We aimed to investigate the cMP subtypes and parental activated cells of ST-elevation MI (STEMI)-patients complicated by CS and that of non-CS STEMI-patients (non-CS) in order to identify a cMP signature that could aid CS patient's risk stratification. METHODS: Clinically-characterized STEMI-patients with and without CS (36/group) were included. Treatment was delivered according to guidelines and included primary percutaneous coronary intervention. cMPs were characterized by triple-labeling flow cytometry using Annexin V and cell surface-specific monoclonal antibodies. RESULTS: Increased levels of leukocyte-derived (neutrophil and granulocyte origin) and platelet-derived cMPs were detected in CS compared to non-CS patients. A signature of cMPs derived from platelets, leukocytes, and endothelium discriminated CS-patients (AUC of 0.743±0.059 [95% CI: 0.628-0.859], P<0.0001) and predicted mortality in CS (AUC of 0.869±0.06 [95% CI: 0.750-0.988], P<0.0001). In CS-patients, a higher number of platelet- and monocyte-cMPs and of tissue factor-rich cMPs associated to worse myocardial blush grade and thrombolysis in myocardial infarction flow. CONCLUSIONS: cMPs derived from proinflammatory and prothrombotic cells were found to be elevated in CS-patients. In treated as per guidelines CS patients, granulocytes and neutrophils remained activated and actively shed cMPs. These cMPs were biomarkers of adverse prognosis in CS. TRANSLATIONAL ASPECT: Increased levels of leukocyte and platelet-derived circulating microparticles (cMPs) are found in cardiogenic shock (CS) patients as compared to non-CS patients. In CS-patients, a higher number of platelet- and monocyte-cMPs and a higher number of tissue factor-rich cMPs were associated to worse myocardial reperfusion. A specific prothrombotic and proinflammatory cMPs signature in cardiogenic shock (CS) patients is a potential discriminator and survival prognostic biomarker for CS, which could aid management and improve clinical outcomes.
Subject(s)
Cell-Derived Microparticles/metabolism , ST Elevation Myocardial Infarction/blood , ST Elevation Myocardial Infarction/epidemiology , Severity of Illness Index , Shock, Cardiogenic/blood , Shock, Cardiogenic/epidemiology , Aged , Biomarkers/blood , Cohort Studies , Female , Humans , Male , Middle Aged , Prospective Studies , Retrospective Studies , ST Elevation Myocardial Infarction/diagnosis , Shock, Cardiogenic/diagnosisABSTRACT
Atherothrombosis is a leading cause of cardiovascular mortality and long-term morbidity. Platelets and coagulation proteases, interacting with circulating cells and in different vascular beds, modify several complex pathologies including atherosclerosis. In the second Maastricht Consensus Conference on Thrombosis, this theme was addressed by diverse scientists from bench to bedside. All presentations were discussed with audience members and the results of these discussions were incorporated in the final document that presents a state-of-the-art reflection of expert opinions and consensus recommendations regarding the following five topics: 1. Risk factors, biomarkers and plaque instability: In atherothrombosis research, more focus on the contribution of specific risk factors like ectopic fat needs to be considered; definitions of atherothrombosis are important distinguishing different phases of disease, including plaque (in)stability; proteomic and metabolomics data are to be added to genetic information. 2. Circulating cells including platelets and atherothrombosis: Mechanisms of leukocyte and macrophage plasticity, migration, and transformation in murine atherosclerosis need to be considered; disease mechanism-based biomarkers need to be identified; experimental systems are needed that incorporate whole-blood flow to understand how red blood cells influence thrombus formation and stability; knowledge on platelet heterogeneity and priming conditions needs to be translated toward the in vivo situation. 3. Coagulation proteases, fibrin(ogen) and thrombus formation: The role of factor (F) XI in thrombosis including the lower margins of this factor related to safe and effective antithrombotic therapy needs to be established; FXI is a key regulator in linking platelets, thrombin generation, and inflammatory mechanisms in a renin-angiotensin dependent manner; however, the impact on thrombin-dependent PAR signaling needs further study; the fundamental mechanisms in FXIII biology and biochemistry and its impact on thrombus biophysical characteristics need to be explored; the interactions of red cells and fibrin formation and its consequences for thrombus formation and lysis need to be addressed. Platelet-fibrin interactions are pivotal determinants of clot formation and stability with potential therapeutic consequences. 4. Preventive and acute treatment of atherothrombosis and arterial embolism; novel ways and tailoring? The role of protease-activated receptor (PAR)-4 vis à vis PAR-1 as target for antithrombotic therapy merits study; ongoing trials on platelet function test-based antiplatelet therapy adjustment support development of practically feasible tests; risk scores for patients with atrial fibrillation need refinement, taking new biomarkers including coagulation into account; risk scores that consider organ system differences in bleeding may have added value; all forms of oral anticoagulant treatment require better organization, including education and emergency access; laboratory testing still needs rapidly available sensitive tests with short turnaround time. 5. Pleiotropy of coagulation proteases, thrombus resolution and ischaemia-reperfusion: Biobanks specifically for thrombus storage and analysis are needed; further studies on novel modified activated protein C-based agents are required including its cytoprotective properties; new avenues for optimizing treatment of patients with ischaemic stroke are needed, also including novel agents that modify fibrinolytic activity (aimed at plasminogen activator inhibitor-1 and thrombin activatable fibrinolysis inhibitor.
Subject(s)
Thromboembolism/therapy , Thrombosis/blood , Thrombosis/therapy , Anticoagulants/therapeutic use , Biomarkers/blood , Blood Coagulation , Erythrocytes/metabolism , Factor VIII/metabolism , Factor XII/metabolism , Factor XIII/metabolism , Humans , Macrophages/metabolism , Netherlands , Phenotype , Plaque, Atherosclerotic/blood , Plaque, Atherosclerotic/diagnosis , Plaque, Atherosclerotic/therapy , Polyphosphates/metabolism , Risk Factors , Signal Transduction , Thromboembolism/blood , Thromboembolism/diagnosis , Thrombosis/diagnosisABSTRACT
BACKGROUND AND AIMS: High LDL triggers dyslipidemia and atherosclerosis, a chronic inflammatory disease with participation of the innate immunity system. Monocytes are recruited to areas of LDL-induced endothelial damage and initiate differentiation. This study was aimed to investigate the effects of LDL on the early transitional stages of monocyte differentiation into macrophages. METHODS: Blood monocytes, isolated from healthy donors by their adhesion properties, were exposed to native-LDL (1.80 mg/mL) for 48-h. Monocyte phenotype was assessed at transcript and miRNA levels by real-time PCR. Protein-expression was determined by western-blot and flow-cytometry. RESULTS: CD14 time-dependently decreased in adhered monocytes, reaching a >4 fold decrease at transcript- and protein-levels after 7-days in culture when cells were already differentiated into macrophages. At 4-days differentiation, monocytes exposed to LDL reduced CD14-transcrition >1.5 fold in mRNA (p = 0.002) and 34% CD14-protein (p = 0.039), whereas increased in CD16-expression (p = 0.019). Besides, LDL induced a significant increase in integrin CD49c (α3-subunit) at mRNA (>2 fold, p = 0.008) and protein (>3 fold, p = 0.045) level and a decrease in the apoptosis-effectors CASP8 and CASP3 (p = 0.002 and p = 0.035, respectively) as well as in the precursor form of the death-receptor DR5 (p = 0.045) without affecting its mRNA-expression level, suggesting a LDL-dependent post-transcriptional regulation of DR5. In silico prediction analysis indicated miR-126-3p as a candidate to regulate DR5-expression and miR-126-3p was shown affected by LDL reaching a significant increase (p = 0.033). CONCLUSIONS: In differentiating human monocytes, LDL stimulates expression of cell-adhesion molecules and downregulates apoptosis-effectors, regulating anoikis and survival programs in the early stage macrophages.
Subject(s)
Anoikis/drug effects , Cell Adhesion/drug effects , Cell Transdifferentiation/drug effects , Lipoproteins, LDL/pharmacology , Macrophages/drug effects , Monocytes/drug effects , Cell Adhesion Molecules/genetics , Cell Adhesion Molecules/metabolism , Cells, Cultured , GPI-Linked Proteins/genetics , GPI-Linked Proteins/metabolism , Humans , Integrin alpha3/genetics , Integrin alpha3/metabolism , Lipopolysaccharide Receptors/genetics , Lipopolysaccharide Receptors/metabolism , Macrophages/metabolism , Macrophages/pathology , MicroRNAs/genetics , MicroRNAs/metabolism , Monocytes/metabolism , Monocytes/pathology , Phenotype , RNA, Messenger/genetics , RNA, Messenger/metabolism , Receptors, IgG/genetics , Receptors, IgG/metabolism , Receptors, TNF-Related Apoptosis-Inducing Ligand/genetics , Receptors, TNF-Related Apoptosis-Inducing Ligand/metabolism , Time Factors , Transcription, Genetic/drug effectsABSTRACT
BACKGROUND: Circulating microparticle (cMP) levels are increased in the acute phase of ST-elevation myocardial infarction (STEMI) and associate with microvascular obstruction; however, the precise cMP-parental cell signature and activation level are not elucidated. Here, we aimed to study the cMP signature in STEMI-patients and whether cMP phenotype changes in relation to onset of pain-to-PCI [ischemic time (IT)]-elapsed time. METHODS: Blood was taken at PCI from the culprit coronary and the peripheral circulation in STEMI-patients (N=40). Two control groups were included: peripheral blood of age-matched patients recovering from STEMI [after 72 h] and of control individuals (N=20/group). cMP-parental origin and activation level were characterized by triple-labeling flow cytometry. RESULTS: Procoagulant annexin V-positive cMPs bearing parental cell markers as well as markers of activated cells displayed a significantly different profile in STEMI-patients, in control individuals and in patients recovering from STEMI. cMPs derived from monocytes, endothelium, and activated vascular cells were higher in the culprit coronary artery than in peripheral blood in STEMI-patients, especially in patients intervened at short IT. Indeed, cMP levels in coronary blood were inversely related to IT duration (more abundant in thrombi with pain-to-PCI time<180 min). CONCLUSIONS: A characteristic [CD66b+/CD62E+/CD142+] cMP signature in the systemic circulation reflects the formation of coronary thrombotic occlusions in STEMI-patients. Changes in the cMP signature in the culprit coronary artery blood reveal the sensitivity of MPs to detect the ischemia-elapsed time. Interestingly, cMPs in peripheral blood may be sensitive markers of the thrombo-occlusive vascular process developing in the coronary arteries of STEMI-patients.
Subject(s)
Cell-Derived Microparticles/metabolism , Coronary Occlusion/blood , Myocardial Infarction/blood , Pain/metabolism , Aged , Angioplasty, Balloon, Coronary/methods , Biomarkers/blood , Biomarkers/metabolism , Coronary Occlusion/metabolism , Coronary Vessels/metabolism , Electrocardiography , Female , Flow Cytometry/methods , Humans , Male , Middle Aged , Myocardial Infarction/metabolism , Myocardial Infarction/surgery , Myocardial Revascularization/methods , Pain/diagnosis , Pain/surgery , Percutaneous Coronary Intervention/methods , Phenotype , Predictive Value of Tests , Severity of Illness Index , Thrombectomy/methods , Thrombosis/metabolismABSTRACT
BACKGROUND: Local fluid dynamics and exposed atherosclerotic lesions regulate thrombus formation. Activated cells in the attached thrombi release microparticles to the circulation (circulating microparticles [cMPs]); however, their phenotype is unknown. OBJECTIVES: To investigate the specific phenotype of the cMPs released by growing thrombi. METHODS/PATIENTS: cMPs released by thrombi growing in different well-characterized thrombogenic conditions were investigated. cMP contents just before and immediately after perfusion of the thrombogenic surfaces were analyzed by triple-labeling flow cytometry. cMPs were tested for their thrombin-generating capacity. The cMPs identified in the ex vivo perfusion experiments were validated in blood of ST-elevation myocardial infarction (STEMI) patients undergoing thrombectomy and percutaneous coronary intervention. Culprit coronary blood (STEMI-CCB) and peripheral artery blood (STEMI-PAB) were simultaneously analyzed and compared with peripheral artery blood from age-matched controls (C-PAB) and peripheral artery blood from patients who had recovered from acute coronary syndrome (ACS) (pSTEMI-PAB). RESULTS: The levels of annexin V(+) cMPs significantly increased in blood collected after perfusion of the exposed thrombogenic surfaces. cMP release was directly related to the formed thrombus mass and the plasma procoagulant activity. Post-thrombus blood showed higher thrombin generation potential and contained higher levels of cMPs carrying glycophorin-A (CD235a(+) ; erythrocyte-derived microparticles [ErMPs]) than preperfusion blood (P < 0.05), whereas the levels of cMPs carrying activated and adhesion platelet markers were decreased. STEMI-CCB and STEMI-PAB had significantly higher ErMP levels than control blood (P < 0.005). ErMP levels were also significantly higher in STEMI-PAB than in pSTEMI-PAB, validating the experimental mechanistic studies and suggesting that ErMPs are markers of ongoing coronary thrombosis (C-statistics: 0.950; 95% confidence interval 0.889-1.000; P < 0.001). CONCLUSION: Glycophorin-A-rich microparticles are released from evolving growing thrombi into the distal perfusing blood, and can be measured in peripheral blood. CD235a(+) cMPs may constitute a novel systemic biomarker of ongoing thrombosis.
Subject(s)
Blood Platelets/metabolism , Cell-Derived Microparticles/metabolism , Coronary Thrombosis/blood , Glycophorins/metabolism , Myocardial Infarction/blood , Platelet Activation , Aged , Biomarkers/blood , Case-Control Studies , Coronary Thrombosis/diagnosis , Coronary Thrombosis/therapy , Female , Flow Cytometry , Humans , Male , Middle Aged , Myocardial Infarction/diagnosis , Myocardial Infarction/therapy , Percutaneous Coronary Intervention , Phenotype , Predictive Value of Tests , Reproducibility of Results , Thrombectomy , Thrombin/metabolism , Time Factors , Treatment OutcomeABSTRACT
Platelet activation is relevant to a variety of acute thrombotic events. We sought to examine adenosine 5'-monophosphate (AMP) mechanisms of action in preventing platelet activation, thrombus formation and platelet-related inflammatory response. We assessed the effect of AMP on 1) P-selectin expression and GPIIb/IIIa activation by flow cytometry; 2) Platelet aggregation and ATP secretion induced by ADP, collagen, TRAP-6, convulxin and thrombin; 3) Platelet rolling and firm adhesion, and platelet-leukocyte interactions under flow-controlled conditions; and, 4) Platelet cAMP levels, sP-selectin, sCD40L, IL-1ß, TGF-ß1 and CCL5 release, PDE3A activity and PKA phosphorylation. The effect of AMP on in vivo thrombus formation was also evaluated in a murine model. The AMP docking with respect to A2 adenosine receptor was determined by homology. AMP concentration-dependently (0.1 to 3 mmol/l) inhibited P-selectin expression and GPIIb/IIIa activation, platelet secretion and aggregation induced by ADP, collagen, TRAP-6 and convulxin, and diminished platelet rolling and firm adhesion. Furthermore, AMP induced a marked increase in the rolling speed of leukocytes retained on the platelet surface. At these concentrations AMP significantly decreased inflammatory mediator from platelet, increased intraplatelet cAMP levels and inhibited PDE3A activity. Interestingly, SQ22536, ZM241385 and SCH58261 attenuated the antiplatelet effect of AMP. Docking experiments revealed that AMP had the same orientation that adenosine inside the A2 adenosine receptor binding pocket. These in vitro antithrombotic properties were further supported in an in vivo model of thrombosis. Considering the successful use of combined antiplatelet therapy, AMP may be further developed as a novel antiplatelet agent.
Subject(s)
Adenosine Monophosphate/metabolism , Blood Platelets/physiology , Mesenteric Veins/physiology , Receptor, Adenosine A2A/metabolism , Thrombosis/blood , Adenosine A2 Receptor Agonists/pharmacology , Adenosine Monophosphate/analogs & derivatives , Animals , Blood Platelets/drug effects , CD40 Ligand/metabolism , Cells, Cultured , Crotalid Venoms/metabolism , Cyclic Nucleotide Phosphodiesterases, Type 3/metabolism , Disease Models, Animal , Humans , Lasers/statistics & numerical data , Lectins, C-Type/metabolism , Mesenteric Veins/drug effects , Mesenteric Veins/radiation effects , Mice , Molecular Targeted Therapy , P-Selectin/genetics , P-Selectin/metabolism , Peptide Fragments/metabolism , Photochemical Processes , Platelet Activation/drug effects , Platelet Glycoprotein GPIIb-IIIa Complex/genetics , Platelet Glycoprotein GPIIb-IIIa Complex/metabolism , Rose Bengal/administration & dosage , Stereoisomerism , Thrombosis/chemically induced , Thrombosis/drug therapy , Triazines/pharmacology , Triazoles/pharmacologyABSTRACT
BACKGROUND: Urokinase-type plasminogen activator (UPA) regulates vascular smooth muscle cell (VSMC) functions relevant in vascular remodeling by facilitating proteolysis at the cell surface and inducing cell signaling pathways. Our previous results demonstrated that aggregated low-density lipoprotein (agLDL) impair cytoskeleton dynamics, a key event contributing to VSMC behavior during progression of atherosclerotic plaques. OBJECTIVES: To investigate whether mechanisms underlying inhibition of cytoskeleton dynamics in lipid-loaded VSMC occurs through a UPA-mediated process. METHODS: Adhesion assay was performed in lipid-loaded human VSMC after 16-h exposition to agLDL (100 µg mL(-1)). Protein subcellular localization and actin-fiber formation were assessed by confocal microscopy. For analysis of protein expression western blots were carried out. Co-immunoprecipitates of UPAR were examined by one-dimensional- or two-dimensional electrophoresis (1-DE or 2-DE), mass spectrometry MALDI-TOF and western blot. RESULTS: agLDL induced UPA subcellular delocalization and significantly decreased UPA levels during attachment of VSMC. UPA (enhanced endogenous-expression or exogenous added) acting as a urokinase-type plasminogen activator receptor (UPAR)-ligand restored actin-cytoskeleton organization and adhesion capacity of lipid-loaded cells to control levels. UPAR co-immunoprecipitated with the unphosphorylated form of myosin regulatory light chain (MRLC) in lipid-loaded cells. The detrimental effects of agLDL on MRLC phosphorylation were reversed by high levels of UPA. The UPA effects on VSMC exposed to agLDL involved FAK phosphorylation. CONCLUSIONS: The detrimental effects of atherogenic LDL on VSMC are mediated by a decrease and delocalization of the UPA-UPAR interaction that result in an impairment of cytoskeleton dynamics and adhesion capacity affecting cell phenotype and function.
Subject(s)
Atherosclerosis/metabolism , Cytoskeleton/metabolism , Lipoproteins, LDL/metabolism , Muscle, Smooth, Vascular/metabolism , Myocytes, Smooth Muscle/metabolism , Receptors, Urokinase Plasminogen Activator/metabolism , Urokinase-Type Plasminogen Activator/metabolism , Atherosclerosis/genetics , Blotting, Western , Cell Adhesion , Cells, Cultured , Electrophoresis, Gel, Two-Dimensional , Focal Adhesion Kinase 1/metabolism , Humans , Immunoprecipitation , Microscopy, Confocal , Myosin Light Chains/metabolism , Phenotype , Phosphorylation , Protein Binding , Protein Transport , RNA Interference , Signal Transduction , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization , Time Factors , Transfection , Urokinase-Type Plasminogen Activator/geneticsABSTRACT
BACKGROUND: Prompt coronary thrombus resolution, reducing time of ischemia, improves cardiac recovery. The factors triggered by ischemia that contribute to the clinical outcome are not fully known. We hypothesize that unabated inflammation due to cardiac ischemia may be a contributing factor. AIMS: As a proof-of-concept, we evaluated the effect of short-term myocardial ischemia on the local and systemic inflammatory response. METHODS: Pigs underwent either 90-min mid-left anterior descending (LAD) coronary artery balloon occlusion (infarct size 25% +/- 1% left ventricle; 29% heart function deterioration) or a sham-operation procedure. Peri-infarcted and non-ischemic cardiac tissue was obtained for histopathologic, molecular and immunohistochemical analysis of inflammatory markers [interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), modified C-reactive protein (mCRP), and human alveolar macrophage-56 (HAM-56)]. Blood (femoral vein) was withdrawn prior to myocardial infarction (MI) induction (t = 0) and at 30 and 90 min to evaluate: (i) systemic cytokine levels (IL-6, TNF-alpha, CRP); (ii) proinflammatory gene and protein expression in peripheral blood mononuclear cells (PBMCs) of tissue factor (TF), cyclo-oxygenase-2 (Cox-2), monocyte chemoattractant protein-1 (MCP-1), and CRP; and (iii) platelet activation (assessed by perfusion studies and RhoA activation). RESULTS: Short-term ischemia triggered cardiac IL-6 and TNF-alpha expression, recruitment of inflammatory cells, and mCRP expression in infiltrated macrophages (P < 0.05 vs. t = 0 and sham). PBMC mRNA and protein expression of MCP-1, Cox-2 and TF was significantly increased by ischemia, whereas no differences were detected in CRP. Ischemia increased cardiac troponin-I, IL-6 and TNF-alpha systemic levels, and was associated with higher platelet deposition and RhoA activation (P < 0.001 vs. t = 0 and sham). CONCLUSION: Short-term myocardial ischemia, even without atherosclerosis, induces an inflammatory phenotype by inducing local recruitment of macrophages and systemic activation of mononuclear cells, and renders platelets more susceptible to activation.
Subject(s)
C-Reactive Protein/genetics , Immunologic Factors/genetics , Myocardial Ischemia/metabolism , Up-Regulation/genetics , Animals , Biomarkers , C-Reactive Protein/metabolism , Chemokine CCL2/genetics , Chemokine CCL2/metabolism , Cyclooxygenase 2/genetics , Cyclooxygenase 2/metabolism , Cytokines , Immune System/cytology , Immunologic Factors/metabolism , Inflammation , Myocardial Ischemia/immunology , Myocardial Ischemia/pathology , Platelet Activation , Swine , Thromboplastin/genetics , Thromboplastin/metabolismABSTRACT
Atherosclerosis is an inflammatory process, triggered by the presence of lipids in the vascular wall, and encompasses a complex interaction among inflammatory cells, vascular elements, and lipoproteins through expression of several adhesion molecules and cytokines. Subendothelial retention of lipoproteins is the key initiating event in atherosclerosis, provoking a cascade of events to pathogenic response. High levels of plasma lipids, particularly low-density (LDL) and very-low-density lipoproteins (VLDL) are among the pathophysiologic stimuli that induce endothelial dysfunction. Endothelial cells regulate coagulation, thrombosis and the fibrinolytic system; the endothelium modulates the activity of smooth muscle cells (vascular tone/proliferation) and controls the traffic of macromolecules and inflammatory cells to the vessel wall. Furthermore, LDLs have been implicated in the induction of changes in permeability, cell adhesion and secretion of vasoactive molecules (nitric oxide [NO]), while VLDLs seem to modulate the fibrinolytic system [tissue plasminogen activator (TPA) and plasminogen activator inhibitor-1 (PAI-1)]. In this review, we will focus on the pathophysiologic functions of lipoproteins in the vascular wall.
Subject(s)
Atherosclerosis/metabolism , Lipoproteins, LDL/metabolism , Animals , Apoptosis , Endothelium, Vascular/cytology , Endothelium, Vascular/metabolism , Humans , Receptors, Lipoprotein/metabolism , Tunica Intima/pathologyABSTRACT
La inflamación juega un papel importante en el inicio, progresión y complicación de la enfermedad arterosclerótica. Estudios epidemiológicos y clínicos han demostrado una estrecha relación entre marcadores de inflamación y el riesgo de sufrir futuros eventos cardiovasculares, tanto en sujetos aparentemente sanos como en pacientes con síndromes crónicos y agudos. Diferentes factores de riesgo cardiovascular, tales como hiperlipidemia o hipertensión y quizás infecciones crónicas (por ejemplo, infecciones peridontales), desencadenan una reacción inflamatoria crónica en la pared vascular arterial, resultado de la cual se desarrollan placas vulnerables, con predisposición a la ruptura y trombosis. De hecho, las lesiones ateroscleróticas presentan un elevado grado de infiltración de componentes celulares inflamatorios (monocitos/macrófagos, T-linfocitos) y la ruptura aguda de las placas está asociada a reacciones inflamatorias. El mejor conocimiento de los mecanismos celulares y moleculares que relacionan inflamación y arteriosclerosis puede no sólo llevar al mejor conocimiento de esta enfermedad, sino que además puede aportar una mejor estratificación pronóstica de riesgo y la identificación de nuevas dianas terapéuticas
Inflammation plays an important role in the onset, progression and complication of arterosclerotic disease. Epidemiologic and clinical studies have demonstrated a close relationship between inflammation markers and the risk of suffering future cardiovascular events, both in apparently healthy subjects as in patients with chronic and acute syndromes. Different cardiovascular risk factors such as hyperlipidemia or hypertension and perhaps chronic infections (for example peridontal infections) precipitate a chronic inflammatory reaction in the arterial vascular wall. Due to this, vulnerable plaques are developed with predisposition to rupture and thrombosis. In fact, the arteriosclerotic lesions have an elevated degree of infiltration of inflammatory cell components (monocytes/macrophages, T-cells) and acute rupture of the plaques is associated to inflammatory reactions. Better knowledge of the cell and molecular mechanisms that relate inflammation and atherosclerosis will not only lead to better knowledge of this disease but also may improve prognostic risk stratification and identification of new therapeutic targetsInflammation plays an important role in the onset, progression and complication of arterosclerotic disease. Epidemiologic and clinical studies have demonstrated a close relationship between inflammation markers and the risk of suffering future cardiovascular events, both in apparently healthy subjects as in patients with chronic and acute syndromes. Different cardiovascular risk factors such as hyperlipidemia or hypertension and perhaps chronic infections (for example peridontal infections) precipitate a chronic inflammatory reaction in the arterial vascular wall. Due to this, vulnerable plaques are developed with predisposition to rupture and thrombosis. In fact, the arteriosclerotic lesions have an elevated degree of infiltration of inflammatory cell components (monocytes/macrophages, T-cells) and acute rupture of the plaques is associated to inflammatory reactions. Better knowledge of the cell and molecular mechanisms that relate inflammation and atherosclerosis will not only lead to better knowledge of this disease but also may improve prognostic risk stratification and identification of new therapeutic targets
Subject(s)
Humans , Intracranial Arteriosclerosis/physiopathology , Inflammation Mediators/analysis , Inflammation/physiopathology , Biomarkers/analysis , Risk Factors , C-Reactive Protein/physiology , Neovascularization, Pathologic/physiopathology , Coronary Disease/physiopathology , Periodontal Diseases/complicationsABSTRACT
La inflamación juega un papel importante en el inicio, progresión y complicación de la enfermedad arterosclerótica. Estudios epidemiológicos y clínicos han demostrado una estrecha relación entre marcadores de inflamación y el riesgo de sufrir futuros eventos cardiovasculares, tanto en sujetos aparentemente sanos como en pacientes con síndromes crónicos y agudos. Diferentes factores de riesgo cardiovascular, tales como hiperlipidemia o hipertensión y quizás infecciones crónicas (por ejemplo, infecciones peridontales), desencadenan una reacción inflamatoria crónica en la pared vascular arterial, resultado de la cual se desarrollan placas vulnerables, con predisposición a la ruptura y trombosis. De hecho, las lesiones ateroscleróticas presentan un elevado grado de infiltración de componentes celulares inflamatorios (monocitos/macrófagos, T-linfocitos) y la ruptura aguda de las placas está asociada a reacciones inflamatorias. El mejor conocimiento de los mecanismos celulares y moleculares que relacionan inflamación y arteriosclerosis puede no sólo llevar al mejor conocimiento de esta enfermedad, sino que además puede aportar una mejor estratificación pronóstica de riesgo y la identificación de nuevas dianas terapéuticas
Inflammation plays an important role in the onset, progression and complication of arterosclerotic disease. Epidemiologic and clinical studies have demonstrated a close relationship between inflammation markers and the risk of suffering future cardiovascular events, both in apparently healthy subjects as in patients with chronic and acute syndromes. Different cardiovascular risk factors such as hyperlipidemia or hypertension and perhaps chronic infections (for example peridontal infections) precipitate a chronic inflammatory reaction in the arterial vascular wall. Due to this, vulnerable plaques are developed with predisposition to rupture and thrombosis. In fact, the arteriosclerotic lesions have an elevated degree of infiltration of inflammatory cell components (monocytes/macrophages, T-cells) and acute rupture of the plaques is associated to inflammatory reactions. Better knowledge of the cell and molecular mechanisms that relate inflammation and atherosclerosis will not only lead to better knowledge of this disease but also may improve prognostic risk stratification and identification of new therapeutic targets
Subject(s)
Humans , Inflammation/physiopathology , Arteriosclerosis/physiopathology , Biomarkers/analysis , Inflammation/epidemiology , Inflammation/complications , Arteriosclerosis/etiology , C-Reactive Protein/adverse effects , Risk Factors , Neovascularization, Pathologic/physiopathology , Periodontitis/physiopathologyABSTRACT
In analogy to solid neoplasms, accumulating data suggest the requirement of angiogenesis also for the development and progression of hematopoietic malignancies including acute myeloid leukemia (AML). Inhibition of increased microvessel density in bone marrow (BM) might be a promising target for pharmacological interventions aimed at reducing disease activity. Among the putative inhibitors of angiogenesis, thalidomide has demonstrated a considerable efficacy in myelodysplastic syndromes (MDS) and AML with overall response rates up to 56% and 25%, respectively. Responders experienced hematologic improvements with increased hemoglobin and platelet counts resulting in temporary transfusion independence. In AML, partial responses--defined as reduction of the leukemic blast cell infiltration of at least 50% in BM--occurred in four of 20 patients after one month of thalidomide administration in a previous phase I/II study. Additionally, we observed a long-term response in one AML patient of more than 20 months, meanwhile fulfilling the criteria of complete remission. The decrease in leukemic blast infiltration in BM of responders was accompanied by a significant reduction of the microvessel density. Overall adverse events caused by the drug consisted mainly of fatigue, constipation, skin rash and polyneuropathy with a tolerable dose of 200-400 mg p.o. per day. In conclusion, thalidomide as a single agent has significant anti-leukemic activity with some evidence for anti-angiogenic effects in BM, although the precise mechanism of action remains to be elucidated.
Subject(s)
Angiogenesis Inhibitors/therapeutic use , Antineoplastic Agents/therapeutic use , Leukemia, Myeloid/drug therapy , Thalidomide/therapeutic use , Acute Disease , Aged , Aged, 80 and over , Angiogenesis Inhibitors/adverse effects , Antineoplastic Agents/adverse effects , Bone Marrow/drug effects , Bone Marrow/pathology , Clinical Trials, Phase I as Topic , Clinical Trials, Phase II as Topic , Female , Humans , Leukemia, Myeloid/pathology , Male , Middle Aged , Myelodysplastic Syndromes/drug therapy , Neovascularization, Pathologic/drug therapy , Pilot Projects , Prospective Studies , Safety , Thalidomide/adverse effects , Treatment OutcomeABSTRACT
Vascular endothelial growth factor (VEGF) and its cellular receptor VEGFR-2 have been implicated as the main endothelial pathway required for tumor neovascularization. However, the importance of the VEGF/VEGFR-2 system for angiogenesis in hematologic malignancies such as AML remains to be elucidated. In 32 patients with newly diagnosed untreated AML, we observed by immunohistochemical analysis of bone marrow biopsies significantly higher levels of VEGF and VEGFR-2 expression than in 10 control patients (P <0.001). In contrast, VEGFR-1 staining levels in AML patients were in the same range as in the controls. Expression of VEGF and VEGFR-2 was significantly higher in patients with a high degree of microvessel density compared to those with a low degree (VEGF: P =0.024; VEGFR-2: P =0.040) and correlated well with bone marrow microvessel density (r(s)=0.566 and 0.609, respectively; P <0.001). Furthermore, in patients who achieved a complete remission following induction chemotherapy VEGFR-2 staining levels decreased into the normal range. In conclusion, our results provide evidence for increased expression of VEGF/VEGFR-2 of leukemic blasts and correlation with angiogenesis in the bone marrow of AML patients. Thus, VEGF/VEGFR-2 might constitute promising targets for antiangiogenic and antileukemic treatment strategies in AML.
Subject(s)
Bone Marrow Cells/metabolism , Endothelial Growth Factors/biosynthesis , Leukemia, Myeloid/metabolism , Lymphokines/biosynthesis , Receptor Protein-Tyrosine Kinases/biosynthesis , Receptors, Growth Factor/biosynthesis , Acute Disease , Adult , Aged , Antineoplastic Combined Chemotherapy Protocols/therapeutic use , Bone Marrow Cells/pathology , Endothelial Growth Factors/genetics , Gene Expression Regulation, Neoplastic , Humans , Leukemia, Myeloid/drug therapy , Leukemia, Myeloid/genetics , Leukemia, Myeloid/pathology , Lymphokines/genetics , Middle Aged , Neovascularization, Pathologic , Receptor Protein-Tyrosine Kinases/genetics , Receptors, Growth Factor/genetics , Receptors, Mitogen/biosynthesis , Receptors, Mitogen/genetics , Receptors, Vascular Endothelial Growth Factor , Remission Induction , Vascular Endothelial Growth Factor A , Vascular Endothelial Growth FactorsABSTRACT
Angiogenesis in Patients with Hematologic Malignancies The importance of angiogenesis for the progressive growth and viability of solid tumors is well established. Emerging data suggest an involvement of angiogenesis in the pathophysiology of hematologic malignancies as well. Recently, we and others have reported increased angiogenesis in the bone marrow of patients with acute myeloid leukemia (AML) and normalization of bone marrow microvessel density when patients achieved a complete remission (CR) after induction chemotherapy. Tumor angiogenesis depends on the expression of specific mediators that initiate a cascade of events leading to the formation of new microvessels. Among these, VEGF (vascular endothelial growth factor), FGF (fibroblast growth factor) and angiopoietins play a pivotal role in the induction of neovascularization in solid tumors. These cytokines stimulate migration and proliferation of endothelial cells and induce angiogenesis in vivo. Recent data suggest an important role for these mediators in hematologic malignancies as well. Isolated AML blasts overexpress VEGF and VEGF receptor 2. Thus, the VEGF/VEGFR-2 pathway can promote the growth of leukemic blasts in an autocrine and paracrine manner. Therefore, neovascularization and angiogenic mediators/receptors may be promising targets for anti-angiogenic and anti-leukemic treatment strategies. The immunomodulatory drug thalidomide inhibits angiogenesis in animal models. Moreover, it has significant activity in refractory multiple myeloma. In a current phase II study for patients with primary refractory or relapsed multiple myeloma using a combination of thalidomide with hyperfractionated cyclophosphamide and dexamethasone (Hyper-CDT), we observed a partial remission in 12 of 14 evaluable patients (86%). Thus, this combination seems to be very potent. Furthermore, we evaluated the safety and efficacy of thalidomide in patients with AML not qualifying for intensive cytotoxic chemotherapy. 20 patients aged 58-85 (median 69) years were recruited to this phase I/II study and were treated with a dose of 200-400 mg per os daily for a duration of 1-40 (median 6) weeks, dependent on the individual tolerability of the drug. In 4 patients we observed a partial response (PR - defined as more than 50% reduction in leukemic blast infiltration in the bone marrow). This was accompanied by an increase in platelet counts and hemoglobin values. One additional patient showed a significant improvement of peripheral blood counts without fulfilling the criteria of a PR. In parallel, we observed a significant decrease in microvessel density in these 5 patients during treatment with thalidomide. In conclusion, thalidomide seems to have anti-angiogenic as well as anti-leukemic activity in AML. The VEGF/VEGFR-2 pathway seems to play an important role in AML. Therefore, receptor tyrosine kinase inhibitors like SU5416 or SU6668 are currently evaluated in the context of phase II studies in AML. We could recently induce a stable remission in a patient with second relapse of her AML refractory towards chemotherapy by administration of SU5416 (compassionate use), a tyrosine kinase inhibitor of VEGFR-2 and ckit. Current and future studies will clarify the role of anti-angiogenic treatment strategies in AML and other hematologic malignancies.
Subject(s)
Bone Marrow/blood supply , Hematologic Neoplasms/pathology , Neovascularization, Pathologic/pathology , Aged , Aged, 80 and over , Bone Marrow/drug effects , Clinical Trials as Topic , Female , Hematologic Neoplasms/drug therapy , Humans , Leukemia, Myeloid, Acute/drug therapy , Leukemia, Myeloid, Acute/pathology , Male , Microcirculation/drug effects , Microcirculation/pathology , Middle Aged , Neovascularization, Pathologic/drug therapy , Thalidomide/adverse effects , Thalidomide/therapeutic use , Treatment OutcomeABSTRACT
The small molecule receptor tyrosine kinase (RTK) inhibitor SU5416 targets the vascular endothelial growth factor receptor 2 and the stem cell factor receptor c-kit. Herein is described the successful treatment of a 65-year-old woman with SU5416, in second relapse of acute myeloid leukemia (AML) and refractory toward standard chemotherapy regimens. After 12 weeks of treatment with SU5416, the blast cell counts (blood and bone marrow) decreased to undetectable levels and the peripheral blood cell counts normalized with the exception of the platelet count (50-80 x 10(9)/L [50-80 x 10(3)/microL]). The duration of the remission is longer than 4 months during maintenance therapy with SU5416. Microvessel density in the patient's bone marrow dropped from 33.4 to 12.3 microvessels/x500-field 8 weeks after SU5416 administration and remains in the normal range. This is the first report of a stable remission achieved after administration of the RTK inhibitor SU5416 in a patient with AML relapse.
Subject(s)
Indoles/administration & dosage , Leukemia, Myeloid/drug therapy , Pyrroles/administration & dosage , Acute Disease , Angiogenesis Inhibitors/administration & dosage , Antineoplastic Agents/administration & dosage , Bone Marrow Cells/chemistry , Endothelium, Vascular/chemistry , Endothelium, Vascular/cytology , Female , Humans , Immunohistochemistry , Middle Aged , Protein-Tyrosine Kinases/antagonists & inhibitors , Receptor Protein-Tyrosine Kinases/drug effects , Receptor Protein-Tyrosine Kinases/metabolism , Receptors, Growth Factor/drug effects , Receptors, Growth Factor/metabolism , Receptors, Vascular Endothelial Growth Factor , Remission Induction/methodsABSTRACT
Vascular endothelial growth factor (VEGF), a multifunctional cytokine, potently stimulates angiogenesis including tumor neovascularization. Although well established in solid tumors, the role of VEGF in bone marrow neoangiogenesis and paracrine tumor-stromal cell interactions in lymphohematopoietic malignancies has not been fully elucidated. In multiple myeloma (MM), marrow neovascularization parallels disease progression. This parallel prompted us to investigate the expression and secretion of VEGF by myeloma cells and its potential effects in myeloma-marrow stroma interactions. The biologically active splice variants VEGF165 and VEGF121 were expressed and secreted by myeloma cell lines and plasma cells isolated from the marrow of patients with MM. As shown by immunocytochemistry or RT-PCR, myeloma cells did not express or weakly expressed the VEGF receptors FLT-1 and FLK-1/KDR, indicating that autocrine stimulation is unlikely. In contrast, FLK-1/KDR was abundantly expressed by marrow stromal cells. Therefore, we studied the effects of VEGF on marrow stroma, focusing on the secretion of interleukin-6 (IL-6), a potent growth factor for myeloma cells and an inhibitor of plasma cell apoptosis. Exposure of stromal and microvascular endothelial cells to recombinant human (rh) VEGF165 or VEGF121 induced a time- and dose-dependent increase in IL-6 secretion (14- to 27-fold at 50 ng/mL after 24 hours, P <.001). Conversely, rhIL-6 stimulated VEGF expression and secretion in myeloma cell lines (40%-60%; P <.05) and to a variable degree (up to 5.3-fold; P <.005) in plasma cells purified from the marrow of patients with MM. This mutual stimulation suggests paracrine interactions between myeloma and marrow stromal cells triggered by VEGF and IL-6. (Blood. 2000;95:2630-2636)
Subject(s)
Cell Communication , Endothelial Growth Factors/physiology , Interleukin-6/physiology , Lymphokines/physiology , Multiple Myeloma/physiopathology , Paracrine Communication , Stromal Cells/pathology , Adult , Aged , Humans , Middle Aged , Multiple Myeloma/metabolism , Multiple Myeloma/pathology , Tumor Cells, Cultured , Vascular Endothelial Growth Factor A , Vascular Endothelial Growth FactorsABSTRACT
The importance of angiogenesis for the progressive growth and viability of solid tumors is well established. In contrast, only few data are available for hematologic neoplasms. To investigate the role of angiogenesis in acute myeloid leukemia (AML), bone marrow biopsies from 62 adults with newly diagnosed, untreated AML (day 0) were evaluated. Further studies were done after the completion of remission induction chemotherapy (day 16 of induction chemotherapy, n = 21; complete remission, n = 20). Microvessels were scored in at least 3 areas (x500 field, 0.126 mm(2)) of the highest microvessel density in representative sections of each bone marrow specimen using immunohistochemistry for von Willebrand factor and thrombomodulin. Microvessel counts were significantly higher in patients with AML (n = 62) compared with control patients (n = 22): median (interquartile range) 24.0 (21.0-27.8)/x500 field vs 11.2 (10. 0-12.0)/x500 field, respectively (P <.001). On day 16 of induction chemotherapy, microvessel density was reduced by 60% (44-66) (P <. 001) in hypoplastic marrows without residual blasts, in contrast to only 17% (0-37) reduction in hypoplastic marrows with >/= 5% residual blasts (P <.001 for the difference between both groups). Bone marrow biopsies taken at the time of complete remission displayed a microvessel density in the same range as the controls. In conclusion, there is evidence of increased microvessel density in the bone marrow of patients with AML, which supports the hypothesis of an important role of angiogenesis in AML. Furthermore, these findings suggest that antiangiogenic therapy might constitute a novel strategy for the treatment of AML. (Blood. 2000;95:2637-2644)
Subject(s)
Bone Marrow/blood supply , Bone Marrow/pathology , Leukemia, Myeloid/pathology , Leukemia, Myeloid/physiopathology , Neovascularization, Pathologic , Acute Disease , Adolescent , Adult , Aged , Aged, 80 and over , Angiogenesis Inhibitors/therapeutic use , Antineoplastic Agents/therapeutic use , Female , Humans , Leukemia, Myeloid/drug therapy , Male , Middle AgedABSTRACT
Recent studies suggest that high plasma levels of tissue-type plasminogen activator (tPA) and its inhibitor (plasminogen activator inhibitor-1, PAI-1) are markers of an increased risk of atherothrombotic ischemic events such as stroke and myocardial infarction. In this prospective study, we measured tPA antigen, PAI-1 antigen and activity, as well as tPA/PAI-1 complex in patients with acute stroke. Stroke subtypes were classified according to the TOAST criteria. From 132 consecutively screened patients, 89 (100%) were enrolled in this study, including 42 patients (47%) with large artery atherosclerosis (LAA), 32 (36%) with small vessel occlusion (SVO), and 15 (17%) with cardioembolism (CE). Nineteen age-matched neurologic patients without manifestations of cerebrovascular disease served as control subjects (CS). Patients with acute stroke had significantly higher plasma levels of tPA antigen (p < 0.001), PAI-1 antigen (p < 0.05) and PAI activity (p < 0.05) than patients in the control group. t-PA antigen, PAI activity and tPA/PAI-1 complex levels were similar regardless of stroke etiology. Only PAI-1 antigen was lower in patients with cardioembolic stroke than in stroke patients with LAA (p < 0.05). Plasma tPA antigen, PAI-1 antigen, and PAI activity are significantly increased in patients with acute ischemic stroke. Except for PAI-1 antigen, this increase appears not to be related to the underlying stroke etiology.
Subject(s)
Brain Ischemia/blood , Infarction, Anterior Cerebral Artery/blood , Infarction, Anterior Cerebral Artery/etiology , Plasminogen Activator Inhibitor 1/blood , Tissue Plasminogen Activator/blood , Acute Disease , Age Factors , Aged , Anticoagulants/administration & dosage , Arteriosclerosis/blood , Brain Ischemia/complications , Brain Ischemia/drug therapy , Carotid Artery, Internal , Female , Humans , Infarction, Middle Cerebral Artery/blood , Infarction, Middle Cerebral Artery/etiology , Intracranial Embolism/blood , Male , Middle Aged , Predictive Value of Tests , Prospective Studies , Vertebrobasilar Insufficiency/blood , Vertebrobasilar Insufficiency/complicationsABSTRACT
UNLABELLED: The plasminogen activator (PA)/plasmin system is involved in various pathological processes that are considered important features of atherogenesis and atherothrombosis. These include the proteolysis of fibrin deposits and extracellular matrix components as well as the induction of cell migration and mitogenesis. Tissue-type PA (TPA) is a key enzyme mediating plasminogen to plasmin conversion. TPA plasma concentrations are elevated in patients with advanced atherosclerosis and correlate with an increased risk for myocardial infarction and stroke. In this study, we have analysed the content and expression of TPA in human coronary arteries and their relation to the presence and severity of atherosclerotic lesions. METHODS: Segments of coronary arteries obtained from heart explants (n = 15) were classified by the presence and types of atherosclerotic lesions. TPA was quantitatively determined in protein extracts of intimal and medial layers. In situ hybridization and immunohistochemical analyses were performed on serial sections of representative tissue specimens. RESULTS: PA activity entirely attributable to the presence of active TPA was consistently detected in the protein extracts. Extractable TPA antigen and activity showed a significant graded increase in relation to the presence and severity of atherosclerotic lesions. The ratios of active over total TPA were increased several-fold in extracts of advanced lesions despite a concomitant threefold increase in TPA complexed to its inhibitor PA-1. In macroscopically normal arterial segments and in early lesions, TPA was expressed in the endothelium and in colocalization with vascular smooth muscle cells (VSMCs). In advanced plaques, TPA mRNA was mainly detected in the lateral regions of the fibrous caps in association with migrating VSMCs and in the vicinity of the core areas infiltrated by CD68-positive macrophages. CONCLUSIONS: TPA content and expression is consistently increased in relation to the severity of the lesions in atherosclerotic coronary arteries. This may contribute to plaque destabilization and disruption. Conversely, the increased intramural TPA activity may counteract mural fibrin deposition.
Subject(s)
Coronary Artery Disease/metabolism , Coronary Vessels/metabolism , Tissue Plasminogen Activator/metabolism , Arteries/metabolism , Coronary Artery Disease/pathology , Coronary Vessels/pathology , Humans , Immunohistochemistry , In Situ Hybridization , Plasminogen Activators/metabolismABSTRACT
Urokinase-type plasminogen activator (UPA) has been implicated in a broad spectrum of pathological processes - e.g. cell adhesion, migration and proliferation and matrix remodeling - that are considered important features of atherogenesis and plaque disruption. In this study, we have analyzed the content and expression of UPA in human coronary arteries and its relation to the presence and severity of atherosclerotic lesions. Segments of coronary arteries obtained from human heart explants (n = 15) were classified by the presence and types of atherosclerotic lesions. UPA was quantitatively determined in protein extracts of the intimal and medial layers. In situ hybridization and immunohistochemical analyses were performed on serial sections of representative tissue specimens. UPA was detected in the extracts as pro-UPA, UPA complexed to plasminogen activator inhibitor-1, or as otherwise inactive UPA antigen, but not in the active two-chain form. Both functional and total UPA were increased several-fold in extracts of advanced lesions, while the ratios of functional over total UPA showed the opposite trend suggesting enhanced UPA inactivation and turnover. UPA expression in early atherosclerotic lesions was particularly prominent in areas of proliferating SMCs in the abluminal part of the neointima, whereas in advanced lesions UPA was widely expressed in macrophage-rich areas adjacent to the rims and shoulder regions of the necrotic cores. The results strongly suggest a causal involvement of UPA in coronary atherogenesis and its clinical outcome.