OxLDL sensitizes platelets for increased formation of extracellular vesicles capable of finetuning macrophage gene expression

Katariina Maaninka; Maarit Neuvonen; Erja Kerkelä; Kati Hyvärinen; Mari Palviainen; Masood Kamali-Moghaddam; Antonio Federico; Dario Greco; Saara Laitinen; Katariina Öörni; Pia RM Siljander

doi:10.1016/j.ejcb.2023.151311

OxLDL sensitizes platelets for increased formation of extracellular vesicles capable of finetuning macrophage gene expression

Katariina Maaninka
, Maarit Neuvonen
, Erja Kerkelä
, Kati Hyvärinen
, Mari Palviainen
, Masood Kamali-Moghaddam
, Antonio Federico
, Dario Greco
, Saara Laitinen
, Katariina Öörni
, Pia RM Siljander^*

^*Corresponding author for this work

Research output: Contribution to journal › Article › Scientific › peer-review

12 Citations (Scopus)

10 Downloads (Pure)

Abstract

Platelet extracellular vesicles (PEVs) generated upon platelet activation may play a role in inflammatory pathologies such as atherosclerosis. Oxidized low-density lipoprotein (oxLDL), a well-known contributor to atherogenesis, activates platelets and presensitizes them for activation by other agonists. We studied the effect of oxLDL on the secretion, composition, and inflammatory functions of PEVs using contemporary EV analytics. Platelets were activated by co-stimulation with thrombin (T) and collagen (C) ± oxLDL and characterized by high-resolution flow cytometry, nanoparticle tracking analysis, proximity extension assay, western blot, and electron microscopy. The effect of PEVs on macrophage differentiation and functionality was examined by analyzing macrophage surface markers, cytokine secretion, and transcriptome. OxLDL upregulated TC-induced formation of CD61⁺, P-selectin⁺ and phosphatidylserine⁺ PEVs. Blocking the scavenger receptor CD36 significantly suppressed the oxLDL+TC-induced PEV formation, and HDL caused a slight but detectable suppression. The inflammatory protein cargo differed between the PEVs from stimulated and unstimulated platelets. Both oxLDL+TC- and TC-induced PEVs enhanced macrophage HLA-DR and CD86 expression and decreased CD11c expression as well as secretion of several cytokines. Pathways related to cell cycle and regulation of gene expression, and immune system signaling were overrepresented in the differentially expressed genes between TC PEV -treated vs. control macrophages and oxLDL+TC PEV -treated vs. control macrophages, respectively. In conclusion, we speculate that oxLDL and activated platelets contribute to proatherogenic processes by increasing the number of PEVs that provide an adhesive and procoagulant surface, contain inflammatory mediators, and subtly finetune the macrophage gene expression.

Original language	English
Article number	151311
Number of pages	15
Journal	EUROPEAN JOURNAL OF CELL BIOLOGY
Volume	102
Issue number	2
DOIs	https://doi.org/10.1016/j.ejcb.2023.151311
Publication status	Published - Jun 2023
Publication type	A1 Journal article-refereed

Funding

We thank the services of University of Helsinki: EV Core in FIMM Technology Centre supported by the HiLIFE and Biocenter Finland for performing electron microscopy work and Electron Microscopy Unit of the Institute of Biotechnology for providing the facilities. We also thank Core facility Sequencing Unit at FIMM Technology Centre supported by the University of Helsinki and Biocenter Finland, and PLA and Single Cell Facility Swedish SciLifeLab for PEA analysis. Dr. Matti Jauhianen is thanked for kindly providing HDL and the anti-ApoB monoclonal antibody for the experiments. This work was supported by the Academy of Finland [grants 287089 (PRMS and MN), 330486 (PRMS and MP), 332761 (DG and AF), and 332564 (KÖ)]; Business Finland [grant EVE (PRMS and KM)]; Magnus Ehrnrooth Foundation (PRMS); Finnish Foundation for Cardiovascular Research (MN and KÖ); Swedish Research Council [grant 2020–02258 (MKM)]; Medicinska Understödsföreningen Liv och Hälsa rf (PRMS); and Novo Nordisk Foundation [grant NNF19OC0057411 (KÖ)]. The funders had no role in the design of the study, in the collection, analysis, or interpretation of the data, in the writing of the manuscript, or in the decision to publish the results.

Keywords

Atherosclerosis
Extracellular vesicle
Macrophage
Oxidized low-density lipoprotein
Platelet
Transcriptome

Publication forum classification

Publication forum level 1

ASJC Scopus subject areas

Pathology and Forensic Medicine
Histology
Cell Biology

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10.1016/j.ejcb.2023.151311Licence: CC BY-NC-ND

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https://urn.fi/URN:NBN:fi:tuni-202304123622Licence: CC BY-NC-ND

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Maaninka, K., Neuvonen, M., Kerkelä, E., Hyvärinen, K., Palviainen, M., Kamali-Moghaddam, M., Federico, A., Greco, D., Laitinen, S., Öörni, K., & Siljander, P. RM. (2023). OxLDL sensitizes platelets for increased formation of extracellular vesicles capable of finetuning macrophage gene expression. EUROPEAN JOURNAL OF CELL BIOLOGY, 102(2), Article 151311. https://doi.org/10.1016/j.ejcb.2023.151311

@article{244034a2503848deaab5f3ff001bc951,

title = "OxLDL sensitizes platelets for increased formation of extracellular vesicles capable of finetuning macrophage gene expression",

abstract = "Platelet extracellular vesicles (PEVs) generated upon platelet activation may play a role in inflammatory pathologies such as atherosclerosis. Oxidized low-density lipoprotein (oxLDL), a well-known contributor to atherogenesis, activates platelets and presensitizes them for activation by other agonists. We studied the effect of oxLDL on the secretion, composition, and inflammatory functions of PEVs using contemporary EV analytics. Platelets were activated by co-stimulation with thrombin (T) and collagen (C) ± oxLDL and characterized by high-resolution flow cytometry, nanoparticle tracking analysis, proximity extension assay, western blot, and electron microscopy. The effect of PEVs on macrophage differentiation and functionality was examined by analyzing macrophage surface markers, cytokine secretion, and transcriptome. OxLDL upregulated TC-induced formation of CD61+, P-selectin+ and phosphatidylserine+ PEVs. Blocking the scavenger receptor CD36 significantly suppressed the oxLDL+TC-induced PEV formation, and HDL caused a slight but detectable suppression. The inflammatory protein cargo differed between the PEVs from stimulated and unstimulated platelets. Both oxLDL+TC- and TC-induced PEVs enhanced macrophage HLA-DR and CD86 expression and decreased CD11c expression as well as secretion of several cytokines. Pathways related to cell cycle and regulation of gene expression, and immune system signaling were overrepresented in the differentially expressed genes between TC PEV -treated vs. control macrophages and oxLDL+TC PEV -treated vs. control macrophages, respectively. In conclusion, we speculate that oxLDL and activated platelets contribute to proatherogenic processes by increasing the number of PEVs that provide an adhesive and procoagulant surface, contain inflammatory mediators, and subtly finetune the macrophage gene expression.",

keywords = "Atherosclerosis, Extracellular vesicle, Macrophage, Oxidized low-density lipoprotein, Platelet, Transcriptome",

author = "Katariina Maaninka and Maarit Neuvonen and Erja Kerkel{\"a} and Kati Hyv{\"a}rinen and Mari Palviainen and Masood Kamali-Moghaddam and Antonio Federico and Dario Greco and Saara Laitinen and Katariina {\"O}{\"o}rni and Siljander, \{Pia RM\}",

note = "Funding Information: We thank the services of University of Helsinki: EV Core in FIMM Technology Centre supported by the HiLIFE and Biocenter Finland for performing electron microscopy work and Electron Microscopy Unit of the Institute of Biotechnology for providing the facilities. We also thank Core facility Sequencing Unit at FIMM Technology Centre supported by the University of Helsinki and Biocenter Finland, and PLA and Single Cell Facility Swedish SciLifeLab for PEA analysis. Dr. Matti Jauhianen is thanked for kindly providing HDL and the anti-ApoB monoclonal antibody for the experiments. Funding Information: This work was supported by the Academy of Finland [grants 287089 (PRMS and MN), 330486 (PRMS and MP), 332761 (DG and AF), and 332564 (K{\"O})]; Business Finland [grant EVE (PRMS and KM)]; Magnus Ehrnrooth Foundation (PRMS); Finnish Foundation for Cardiovascular Research (MN and K{\"O}); Swedish Research Council [grant 2020–02258 (MKM)]; Medicinska Underst{\"o}dsf{\"o}reningen Liv och H{\"a}lsa rf (PRMS); and Novo Nordisk Foundation [grant NNF19OC0057411 (K{\"O})]. The funders had no role in the design of the study, in the collection, analysis, or interpretation of the data, in the writing of the manuscript, or in the decision to publish the results. Publisher Copyright: {\textcopyright} 2023 The Authors",

year = "2023",

month = jun,

doi = "10.1016/j.ejcb.2023.151311",

language = "English",

volume = "102",

journal = "EUROPEAN JOURNAL OF CELL BIOLOGY",

issn = "0171-9335",

publisher = "Elsevier GmbH",

number = "2",

}

Maaninka, K, Neuvonen, M, Kerkelä, E, Hyvärinen, K, Palviainen, M, Kamali-Moghaddam, M, Federico, A , Greco, D, Laitinen, S, Öörni, K & Siljander, PRM 2023, 'OxLDL sensitizes platelets for increased formation of extracellular vesicles capable of finetuning macrophage gene expression', EUROPEAN JOURNAL OF CELL BIOLOGY, vol. 102, no. 2, 151311. https://doi.org/10.1016/j.ejcb.2023.151311

TY - JOUR

T1 - OxLDL sensitizes platelets for increased formation of extracellular vesicles capable of finetuning macrophage gene expression

AU - Maaninka, Katariina

AU - Neuvonen, Maarit

AU - Kerkelä, Erja

AU - Hyvärinen, Kati

AU - Palviainen, Mari

AU - Kamali-Moghaddam, Masood

AU - Federico, Antonio

AU - Greco, Dario

AU - Laitinen, Saara

AU - Öörni, Katariina

AU - Siljander, Pia RM

N1 - Funding Information: We thank the services of University of Helsinki: EV Core in FIMM Technology Centre supported by the HiLIFE and Biocenter Finland for performing electron microscopy work and Electron Microscopy Unit of the Institute of Biotechnology for providing the facilities. We also thank Core facility Sequencing Unit at FIMM Technology Centre supported by the University of Helsinki and Biocenter Finland, and PLA and Single Cell Facility Swedish SciLifeLab for PEA analysis. Dr. Matti Jauhianen is thanked for kindly providing HDL and the anti-ApoB monoclonal antibody for the experiments. Funding Information: This work was supported by the Academy of Finland [grants 287089 (PRMS and MN), 330486 (PRMS and MP), 332761 (DG and AF), and 332564 (KÖ)]; Business Finland [grant EVE (PRMS and KM)]; Magnus Ehrnrooth Foundation (PRMS); Finnish Foundation for Cardiovascular Research (MN and KÖ); Swedish Research Council [grant 2020–02258 (MKM)]; Medicinska Understödsföreningen Liv och Hälsa rf (PRMS); and Novo Nordisk Foundation [grant NNF19OC0057411 (KÖ)]. The funders had no role in the design of the study, in the collection, analysis, or interpretation of the data, in the writing of the manuscript, or in the decision to publish the results. Publisher Copyright: © 2023 The Authors

PY - 2023/6

Y1 - 2023/6

N2 - Platelet extracellular vesicles (PEVs) generated upon platelet activation may play a role in inflammatory pathologies such as atherosclerosis. Oxidized low-density lipoprotein (oxLDL), a well-known contributor to atherogenesis, activates platelets and presensitizes them for activation by other agonists. We studied the effect of oxLDL on the secretion, composition, and inflammatory functions of PEVs using contemporary EV analytics. Platelets were activated by co-stimulation with thrombin (T) and collagen (C) ± oxLDL and characterized by high-resolution flow cytometry, nanoparticle tracking analysis, proximity extension assay, western blot, and electron microscopy. The effect of PEVs on macrophage differentiation and functionality was examined by analyzing macrophage surface markers, cytokine secretion, and transcriptome. OxLDL upregulated TC-induced formation of CD61+, P-selectin+ and phosphatidylserine+ PEVs. Blocking the scavenger receptor CD36 significantly suppressed the oxLDL+TC-induced PEV formation, and HDL caused a slight but detectable suppression. The inflammatory protein cargo differed between the PEVs from stimulated and unstimulated platelets. Both oxLDL+TC- and TC-induced PEVs enhanced macrophage HLA-DR and CD86 expression and decreased CD11c expression as well as secretion of several cytokines. Pathways related to cell cycle and regulation of gene expression, and immune system signaling were overrepresented in the differentially expressed genes between TC PEV -treated vs. control macrophages and oxLDL+TC PEV -treated vs. control macrophages, respectively. In conclusion, we speculate that oxLDL and activated platelets contribute to proatherogenic processes by increasing the number of PEVs that provide an adhesive and procoagulant surface, contain inflammatory mediators, and subtly finetune the macrophage gene expression.

AB - Platelet extracellular vesicles (PEVs) generated upon platelet activation may play a role in inflammatory pathologies such as atherosclerosis. Oxidized low-density lipoprotein (oxLDL), a well-known contributor to atherogenesis, activates platelets and presensitizes them for activation by other agonists. We studied the effect of oxLDL on the secretion, composition, and inflammatory functions of PEVs using contemporary EV analytics. Platelets were activated by co-stimulation with thrombin (T) and collagen (C) ± oxLDL and characterized by high-resolution flow cytometry, nanoparticle tracking analysis, proximity extension assay, western blot, and electron microscopy. The effect of PEVs on macrophage differentiation and functionality was examined by analyzing macrophage surface markers, cytokine secretion, and transcriptome. OxLDL upregulated TC-induced formation of CD61+, P-selectin+ and phosphatidylserine+ PEVs. Blocking the scavenger receptor CD36 significantly suppressed the oxLDL+TC-induced PEV formation, and HDL caused a slight but detectable suppression. The inflammatory protein cargo differed between the PEVs from stimulated and unstimulated platelets. Both oxLDL+TC- and TC-induced PEVs enhanced macrophage HLA-DR and CD86 expression and decreased CD11c expression as well as secretion of several cytokines. Pathways related to cell cycle and regulation of gene expression, and immune system signaling were overrepresented in the differentially expressed genes between TC PEV -treated vs. control macrophages and oxLDL+TC PEV -treated vs. control macrophages, respectively. In conclusion, we speculate that oxLDL and activated platelets contribute to proatherogenic processes by increasing the number of PEVs that provide an adhesive and procoagulant surface, contain inflammatory mediators, and subtly finetune the macrophage gene expression.

KW - Atherosclerosis

KW - Extracellular vesicle

KW - Macrophage

KW - Oxidized low-density lipoprotein

KW - Platelet

KW - Transcriptome

U2 - 10.1016/j.ejcb.2023.151311

DO - 10.1016/j.ejcb.2023.151311

M3 - Article

AN - SCOPUS:85150831950

SN - 0171-9335

VL - 102

JO - EUROPEAN JOURNAL OF CELL BIOLOGY

JF - EUROPEAN JOURNAL OF CELL BIOLOGY

IS - 2

M1 - 151311

ER -

OxLDL sensitizes platelets for increased formation of extracellular vesicles capable of finetuning macrophage gene expression

Abstract

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