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The exposure of autoantigens by microparticles underlies the formation of potent inflammatory components: the microparticle-associated immune complexes.

Cloutier N, Tan S, Boudreau LH, Cramb C, Subbaiah R, Lahey L, Albert A, Shnayder R, Gobezie R, Nigrovic PA, Farndale RW, Robinson WH, Brisson A, Lee DM, Boilard E - EMBO Mol Med (2012)

Bottom Line: Rather, mpICs display autoantigens vimentin and fibrinogen, and recognition of these targets by anti-citrullinated peptide antibodies contributes to the production of mpICs.Functionally, platelet mpICs are highly pro-inflammatory, eliciting leukotriene production by neutrophils.Taken together, our data suggest a unique role for platelet MPs as autoantigen-expressing elements capable of perpetuating formation of inflammatory ICs.

View Article: PubMed Central - PubMed

Affiliation: Faculté de Médecine de l'Université Laval, Centre de Recherche en Rhumatologie et Immunologie, Centre de Recherche du Centre Hospitalier Universitaire de Québec, Québec, Québec, Canada.

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Platelet MPs form mpICs independently of CD32aA,B. The expression of CD32a by platelet MPs was determined by hs-FCM and Western blot analysis (inset in A). The platelet MPs were generated in vitro upon platelet GPVI stimulation (A) and the endogenous MPs in RA SF were detected using Annexin-V (B). Blue: isotype; grey: specific CD32a labeling (n = 3).C. FSC-PMT and SSC profiles of the fluorescent platelets (top panel) and platelet MPs (bottom panel) (n = 10). The relative dimensions are presented according to size-defined microsphere calibrations.D–G. Results obtained following the incubation of exogenous fluorescent platelet MPs in RA and PA SF (n = 10 RA and n = 18 PA). (D) Representative FSC-PMT and SSC plots obtained using RA (top) and PA SF (bottom). Two subpopulations (subpop) (1 and 2) were detected in RA and only one in PA SF (1). (E,F) hs-FCM evaluation of the presence of IgG on surface of MPs upon incubation in RA (E) and PA SF (F) using Cy5-conjugated anti-IgG antibody. The four-quadrant gates were positioned according to the isotypic controls. (E) MPs (in green) are IgG− and show dimensions ranging from 100 to 300 nm (lower inset) while the mpICs (in blue) have dimensions ranging from 700 to 3000 nm (upper inset). (D,E) The relative dimensions are presented according to size-defined microsphere calibrations. (G) hs-FCM quantifications of the mpICs were determined in absence (1st and 3rd column) or presence (2nd column) of CD32a blocking antibody (1st and 2nd column p = 0.4478 (unpaired t-test), 1st and 3rd column ***p < 0.0001). Statistical analyses are presented under the graphs. Data are mean ± SEM.
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fig04: Platelet MPs form mpICs independently of CD32aA,B. The expression of CD32a by platelet MPs was determined by hs-FCM and Western blot analysis (inset in A). The platelet MPs were generated in vitro upon platelet GPVI stimulation (A) and the endogenous MPs in RA SF were detected using Annexin-V (B). Blue: isotype; grey: specific CD32a labeling (n = 3).C. FSC-PMT and SSC profiles of the fluorescent platelets (top panel) and platelet MPs (bottom panel) (n = 10). The relative dimensions are presented according to size-defined microsphere calibrations.D–G. Results obtained following the incubation of exogenous fluorescent platelet MPs in RA and PA SF (n = 10 RA and n = 18 PA). (D) Representative FSC-PMT and SSC plots obtained using RA (top) and PA SF (bottom). Two subpopulations (subpop) (1 and 2) were detected in RA and only one in PA SF (1). (E,F) hs-FCM evaluation of the presence of IgG on surface of MPs upon incubation in RA (E) and PA SF (F) using Cy5-conjugated anti-IgG antibody. The four-quadrant gates were positioned according to the isotypic controls. (E) MPs (in green) are IgG− and show dimensions ranging from 100 to 300 nm (lower inset) while the mpICs (in blue) have dimensions ranging from 700 to 3000 nm (upper inset). (D,E) The relative dimensions are presented according to size-defined microsphere calibrations. (G) hs-FCM quantifications of the mpICs were determined in absence (1st and 3rd column) or presence (2nd column) of CD32a blocking antibody (1st and 2nd column p = 0.4478 (unpaired t-test), 1st and 3rd column ***p < 0.0001). Statistical analyses are presented under the graphs. Data are mean ± SEM.

Mentions: Human platelets express the IC receptor FcγRIIA (CD32a; Huang et al, 2011; Parren et al, 1992). We thus postulated that CD32a present on platelet-derived MPs may contribute to formation of mpICs. In this set of experiments, we initially assessed CD32a expression by platelet-derived MPs. Since platelets promptly release MPs following stimulation of the collagen receptor glycoprotein VI (GPVI) (Boilard et al, 2010; Knight et al, 1999), and since GPVI deficiency in mice leads to reduction of the severity of arthritis (Boilard et al, 2010), we generated MPs via this arthritis-relevant stimulus. Here, we find that platelet-derived MPs retain CD32a (Fig 4A). Examination of MPs in RA SF demonstrates they too harbour CD32a (Fig 4B).


The exposure of autoantigens by microparticles underlies the formation of potent inflammatory components: the microparticle-associated immune complexes.

Cloutier N, Tan S, Boudreau LH, Cramb C, Subbaiah R, Lahey L, Albert A, Shnayder R, Gobezie R, Nigrovic PA, Farndale RW, Robinson WH, Brisson A, Lee DM, Boilard E - EMBO Mol Med (2012)

Platelet MPs form mpICs independently of CD32aA,B. The expression of CD32a by platelet MPs was determined by hs-FCM and Western blot analysis (inset in A). The platelet MPs were generated in vitro upon platelet GPVI stimulation (A) and the endogenous MPs in RA SF were detected using Annexin-V (B). Blue: isotype; grey: specific CD32a labeling (n = 3).C. FSC-PMT and SSC profiles of the fluorescent platelets (top panel) and platelet MPs (bottom panel) (n = 10). The relative dimensions are presented according to size-defined microsphere calibrations.D–G. Results obtained following the incubation of exogenous fluorescent platelet MPs in RA and PA SF (n = 10 RA and n = 18 PA). (D) Representative FSC-PMT and SSC plots obtained using RA (top) and PA SF (bottom). Two subpopulations (subpop) (1 and 2) were detected in RA and only one in PA SF (1). (E,F) hs-FCM evaluation of the presence of IgG on surface of MPs upon incubation in RA (E) and PA SF (F) using Cy5-conjugated anti-IgG antibody. The four-quadrant gates were positioned according to the isotypic controls. (E) MPs (in green) are IgG− and show dimensions ranging from 100 to 300 nm (lower inset) while the mpICs (in blue) have dimensions ranging from 700 to 3000 nm (upper inset). (D,E) The relative dimensions are presented according to size-defined microsphere calibrations. (G) hs-FCM quantifications of the mpICs were determined in absence (1st and 3rd column) or presence (2nd column) of CD32a blocking antibody (1st and 2nd column p = 0.4478 (unpaired t-test), 1st and 3rd column ***p < 0.0001). Statistical analyses are presented under the graphs. Data are mean ± SEM.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3569640&req=5

fig04: Platelet MPs form mpICs independently of CD32aA,B. The expression of CD32a by platelet MPs was determined by hs-FCM and Western blot analysis (inset in A). The platelet MPs were generated in vitro upon platelet GPVI stimulation (A) and the endogenous MPs in RA SF were detected using Annexin-V (B). Blue: isotype; grey: specific CD32a labeling (n = 3).C. FSC-PMT and SSC profiles of the fluorescent platelets (top panel) and platelet MPs (bottom panel) (n = 10). The relative dimensions are presented according to size-defined microsphere calibrations.D–G. Results obtained following the incubation of exogenous fluorescent platelet MPs in RA and PA SF (n = 10 RA and n = 18 PA). (D) Representative FSC-PMT and SSC plots obtained using RA (top) and PA SF (bottom). Two subpopulations (subpop) (1 and 2) were detected in RA and only one in PA SF (1). (E,F) hs-FCM evaluation of the presence of IgG on surface of MPs upon incubation in RA (E) and PA SF (F) using Cy5-conjugated anti-IgG antibody. The four-quadrant gates were positioned according to the isotypic controls. (E) MPs (in green) are IgG− and show dimensions ranging from 100 to 300 nm (lower inset) while the mpICs (in blue) have dimensions ranging from 700 to 3000 nm (upper inset). (D,E) The relative dimensions are presented according to size-defined microsphere calibrations. (G) hs-FCM quantifications of the mpICs were determined in absence (1st and 3rd column) or presence (2nd column) of CD32a blocking antibody (1st and 2nd column p = 0.4478 (unpaired t-test), 1st and 3rd column ***p < 0.0001). Statistical analyses are presented under the graphs. Data are mean ± SEM.
Mentions: Human platelets express the IC receptor FcγRIIA (CD32a; Huang et al, 2011; Parren et al, 1992). We thus postulated that CD32a present on platelet-derived MPs may contribute to formation of mpICs. In this set of experiments, we initially assessed CD32a expression by platelet-derived MPs. Since platelets promptly release MPs following stimulation of the collagen receptor glycoprotein VI (GPVI) (Boilard et al, 2010; Knight et al, 1999), and since GPVI deficiency in mice leads to reduction of the severity of arthritis (Boilard et al, 2010), we generated MPs via this arthritis-relevant stimulus. Here, we find that platelet-derived MPs retain CD32a (Fig 4A). Examination of MPs in RA SF demonstrates they too harbour CD32a (Fig 4B).

Bottom Line: Rather, mpICs display autoantigens vimentin and fibrinogen, and recognition of these targets by anti-citrullinated peptide antibodies contributes to the production of mpICs.Functionally, platelet mpICs are highly pro-inflammatory, eliciting leukotriene production by neutrophils.Taken together, our data suggest a unique role for platelet MPs as autoantigen-expressing elements capable of perpetuating formation of inflammatory ICs.

View Article: PubMed Central - PubMed

Affiliation: Faculté de Médecine de l'Université Laval, Centre de Recherche en Rhumatologie et Immunologie, Centre de Recherche du Centre Hospitalier Universitaire de Québec, Québec, Québec, Canada.

Show MeSH
Related in: MedlinePlus