Blood platelets, well known for their role in hemostasis, are abnormally activated in patients suffering from systemic lupus erythematosus (SLE), but also from other immunomediated diseases (scleroderma, vasculitis, myositis, Gougerot-Sjögren's and rheumatoid arthritis) in cases of high disease activity. Once activated, platelets express adhesion molecules such as P-selectin on their surface, enabling them to interact physically with immune cells. In a recent work, we identified that activated platelets from lupus patients interact with regulatory T cells and block their regulatory function, thus participating in the deregulated activation of the immune system in SLE. In addition, inhibition of platelet-immune cell interactions by an anti-P-selectin antibody improved LES symptoms in two mouse models.
The aim of this work is to investigate other potential platelet-immune cell interactions in patients with SLE, in comparison with other autoimmune diseases (systemic scleroderma, ANCA vasculitides, inflammatory myositis, Gougerot-Sjögren syndrome and rheumatoid arthritis).
This study could lead to a better understanding of the role of platelets in the pathophysiology of autoimmune diseases, identify new biomarkers of activity, and assess the potential of new therapeutic avenues in these diseases, such as platelet targeting.
Patients between 18 and 70 years of age
Patient affiliated to a health insurance scheme (beneficiary or beneficiary's beneficiary)
Patient able to understand the aims and risks of research
Patient having signed and dated an informed consent form
Patient in exclusion period (determined by a previous or current study)
Inability to give patient informed consent (patient in emergency or immediate life-threatening situation)
Patient under court protection
Patient under guardianship or curatorship
Patients with systemic auto-immune disease such as : * Systemic lupus * Anca-associated vasculitis * Sjögren's disease * Rheumatoid arthritis * Systemic sclerosis
Cohort
Prospective
Not listed
The primary endpoint will be the percentage of circulating aggregates between platelets and immune cells according to disease activity, assessed by flow cytometry.
Phenotypic impact of platelet/immune cell interaction.
The impact of platelet/immune cell interaction will be assessed using flow cytometry on patient samples. Using a spectral cytometer (Aurora, Cytek) and a multi-color panel (45 colors), we will study the phenotype and degree of activation of different immune subpopulations according to whether or not they interact with platelets. This analysis will be carried out several times during the different visits, and mirrored with the disease activity criteria studied. Activation molecule expression will be analyzed using the same methods as for the main criterion (comparison of the patient with himself/herself using longitudinal samples).
Transcriptomic impact of platelet/immune cell interaction.
To understand the impact of platelet/immune cell interaction at a finer level, we will sort several immune subpopulations (neutrophils, plasmacytoid dendritic cells, regulatory T lymphocytes) aggregated (CD41+) or not (CD41-) to platelets from patient samples by flow cytometry. Total RNA will be isolated from these subpopulations and sequenced by RNAsequencing to assess the impact of the interaction on the immune cell transcriptome. New techniques such as single-cell RNAseq will be used to assess differences between aggregates and non-aggregates.
Measurement of biomarkers of disease activity / predictive of disease flare-ups.
Identifying the mechanisms by which platelets impact immune cells could enable the development of biomarkers of disease activity or prediction of inflammatory flare-ups. These could be molecules of platelet origin (P-selectin, ß-thrombomodulin) or molecules of immune origin produced following interactions with platelets (cytokines, DAMPs). These molecules will be measured using commercial ELISA kits, and analyzed by the same methods as the primary endpoint (comparison of the patient with himself/herself by longitudinal sampling).
Identification of genetic polymorphisms
Identification of genetic polymorphisms (identified by next-generation sequencing) predisposing to platelet activation and targeted sequencing of genes predisposing to platelet activation (FCGR2A,SELP genes) or immune cell activation in inflammatory diseases (SELPG; CD40LG genes).
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