ABSTRACT
AIMS: Thymic stromal lymphopoietin (TSLP) plays an important role in inflammatory diseases and is over-expressed in human atherosclerotic artery specimens. The present study investigated the role of TSLP in platelet activation and thrombosis models in vitro and in vivo, as well as the underlying mechanism and signaling pathway. METHODS AND RESULTS: Western blotting and flow cytometry demonstrated that the TSLP receptor was expressed on murine platelets. According to flow cytometry, platelet stimulation with TSLP induced platelet degranulation and integrin αIIbß3 activation. A TSLPR deficiency caused defective platelet aggregation, defective platelet secretion and markedly blunted thrombus growth in perfusion chambers at both low and high shear rates. TSLPR KO mice exhibited defective carotid artery thrombus formation after exposure to FeCl3. TSLP increased Akt phosphorylation, an effect that was abrogated by the PI3K inhibitors wortmannin and LY294002. The PI3K inhibitors further diminished TSLP-induced platelet activation. TSLP-mediated platelet degranulation, integrin αIIbß3 activation and Akt phosphorylation were blunted in platelets that lacked the TSLP receptor. CONCLUSION: This study demonstrated that the functional TSLPR was surface-expressed on murine platelets. The inflammatory cytokine TSLP triggered platelet activation and thrombus formation via TSLP-dependent PI3K/Akt signaling, which suggests an important role for TSLP in linking vascular inflammation and thrombo-occlusive diseases.
Subject(s)
Blood Platelets/metabolism , Cytokines/pharmacology , Immunoglobulins/metabolism , Platelet Activation/drug effects , Platelet Aggregation/drug effects , Receptors, Cytokine/metabolism , Signal Transduction/drug effects , Androstadienes/pharmacology , Animals , Blood Platelets/drug effects , Carotid Artery Thrombosis/chemically induced , Carotid Artery Thrombosis/metabolism , Carotid Artery Thrombosis/pathology , Chlorides/toxicity , Chromones/pharmacology , Disease Models, Animal , Ferric Compounds/toxicity , Humans , Immunoglobulins/deficiency , Immunoglobulins/genetics , Mice , Mice, Inbred C57BL , Mice, Knockout , Morpholines/pharmacology , P-Selectin/metabolism , Phosphatidylinositol 3-Kinases/metabolism , Phosphoinositide-3 Kinase Inhibitors , Phosphorylation/drug effects , Platelet Glycoprotein GPIIb-IIIa Complex/metabolism , Proto-Oncogene Proteins c-akt/metabolism , Receptors, Cytokine/deficiency , Receptors, Cytokine/genetics , Wortmannin , Thymic Stromal LymphopoietinABSTRACT
AIMS: We generated thymic stromal lymphopoietin R-chain deficient apolipoprotein E-double knockout (ApoE-TSLPR DKO) mice to directly explore the role of thymic stromal lymphopoietin (TSLP) in atherogenesis. METHODS AND RESULTS: Both thymic stromal lymphopoietin (TSLP) and its receptor are expressed in atherosclerotic aortas of apolipoprotein E knockout (ApoE KO) mice. Serum thymic stromal lymphopoietin (TSLP) is markedly increased in apolipoprotein E knockout (ApoE KO) mice fed with a high fat diet (HFD). Arterial lesion formation was significantly decreased in thymic stromal lymphopoietin R-chain deficient apolipoprotein E-double knockout (ApoE-TSLPR DKO) mice compared with apolipoprotein E knockout (ApoE KO) mice. Bone marrow chimera studies indicated reduced lesions in apolipoprotein E knockout (ApoE KO) mice which received the bone marrow of thymic stromal lymphopoietin R-chain deficient apolipoprotein E-double knockout (ApoE-TSLPR DKO) mice as well as in TSLPR KO mice which received bone marrow of ApoE-TSLPR DKO mice. Compared with apolipoprotein E knockout (ApoE KO) mice, IFN-γ secretion by activated T cells was increased but IL-4 expression was reduced in thymic stromal lymphopoietin R-chain deficient apolipoprotein E-double knockout (ApoE-TSLPR DKO) mice. Consisted with these results, the mRNA of IFN-γ was increased but IL-4 was reduced in root. These findings suggest that a reduction in atherosclerotic lesions in thymic stromal lymphopoietin R-chain deficient apolipoprotein E-double knockout (ApoE-TSLPR DKO) mice may not be due to a Th1/Th2 imbalance. On the other hand, the number of Th17 cells, the secretion of IL-17A by activated CD4(+) T cells and the mRNA expression of IL-17A in root were decreased in thymic stromal lymphopoietin R-chain deficient apolipoprotein E-double knockout (ApoE-TSLPR DKO) mice. Notably, the number of regulatory T cell expression of IL-10 was increased in thymic stromal lymphopoietin R-chain deficient apolipoprotein E-double knockout (ApoE-TSLPR DKO) mice. CONCLUSIONS: Collectively, our data suggest that activating thymic stromal lymphopoietin (TSLP) promotes atherosclerosis by inducing Th17/Treg imbalance through thymic stromal lymphopoietin/thymic stromal lymphopoietin R-receptor (TSLP/TSLPR) signal way in apolipoprotein E-deficient mice fed with HFD model.
Subject(s)
Atherosclerosis/immunology , Immunoglobulins/genetics , Receptors, Cytokine/genetics , T-Lymphocytes, Regulatory/immunology , Th17 Cells/immunology , Animals , Aorta, Thoracic/pathology , Apolipoproteins E/genetics , Apolipoproteins E/metabolism , Atherosclerosis/metabolism , Atherosclerosis/pathology , Cytokines/metabolism , Diet, High-Fat/adverse effects , Immunoglobulins/metabolism , Lipid Metabolism , Male , Mice, Inbred C57BL , Mice, Knockout , Plaque, Atherosclerotic/immunology , Plaque, Atherosclerotic/metabolism , Plaque, Atherosclerotic/pathology , Receptors, Cytokine/metabolism , T-Lymphocytes, Regulatory/metabolism , Th17 Cells/metabolismABSTRACT
BACKGROUND: Macrophage death in advanced lesion has been confirmed to play an important role in plaque instability. However, the mechanism underlying lesion macrophage death still remains largely unknown. METHODS AND RESULTS: Immunohistochemistry showed that caspase-1 activated in advanced lesion and co-located with macrophages and TUNEL positive reaction. In in-vitro experiments showed that ox-LDL induced caspase-1 activation and this activation was required for ox-LDL induced macrophages lysis, IL-1ß and IL-18 production as well as DNA fragmentation. Mechanism experiments showed that CD36 and NLRP3/caspase-1/pathway involved in ox-LDL induced macrophage pyroptosis. CONCLUSION: Our study here identified a novel cell death, pyroptosis in ox-LDL induced human macrophage, which may be implicated in lesion macrophages death and play an important role in lesion instability.
Subject(s)
Caspase 1/metabolism , Lipoproteins, LDL/pharmacology , Macrophages/cytology , Macrophages/drug effects , Plaque, Atherosclerotic/immunology , CARD Signaling Adaptor Proteins , CD36 Antigens/metabolism , Carrier Proteins/metabolism , Cell Death/drug effects , Cytoskeletal Proteins/metabolism , DNA Fragmentation/drug effects , Enzyme Activation/drug effects , Humans , Interleukin-18/biosynthesis , Interleukin-1beta/biosynthesis , Macrophages/enzymology , Macrophages/metabolism , NLR Family, Pyrin Domain-Containing 3 Protein , Plaque, Atherosclerotic/genetics , Plaque, Atherosclerotic/metabolism , Reactive Oxygen Species/metabolismABSTRACT
BACKGROUND: Adaptive immunity plays a critical role in atherosclerosis and T helper 17 (Th17) cells, a new T-cell lineage, are recently reported to be involved in atherosclerosis. However, the mechanism underlying Th17 inflammation in atherosclerosis remains largely unknown. Thymic stromal lymphopoietin (TSLP) is a novel IL-7-like cytokine and mainly responsible for Th2 inflammation in many inflammatory diseases. METHODS AND RESULTS: Immunohistochemistry showed that TSLP over-expressed in human atherosclerotic lesion and could be induced by ox-LDL in human vascular smooth muscle cells (HVSMCs) and human umbilical vein endothelial cells (HUVECs). TSLP, in turn, could activate dendritic cells (DCs) to differentiate Th17 inflammation in naive CD4(+) T cells. CONCLUSION: TSLP induced by ox-LDL could promote Th17 immune response in vitro, which may be implicated in Th17 inflammation in atherosclerosis.
