Solid phase enzyme immunoassays for the quantification of serum amyloid P (SAP) and complement component 3 (C3) proteins in acute-phase mouse sera.
ABSTRACT Simple, reliable and sensitive enzyme immunoassays have been developed for the quantification of the mouse acute-phase SAP and C3 proteins. The ELISA systems were validated using sera from mice injected with S. dysenteriae endotoxin, and detected 500 pg protein/ml. The assays use 96-well microtitre plates permitting rapid processing of a large number of samples.
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ABSTRACT: Macrophage differentiation and polarization is influenced by, and act on, many processes associated with autoimmunity. However, the molecular mechanisms underlying macrophage polarization in systemic lupus erythematosus (SLE) remain largely debated. We previously demonstrated that macrophage M2b polarization conferred by activated lymphocyte-derived (ALD)-DNA immunization could initiate and propagate murine lupus nephritis. Serum amyloid P component (SAP), a conserved acute-phase protein in mice, has been reported to bind to DNA and modulate immune responses. In this study, murine SAP was shown to promote macrophage-mediated ALD-DNA uptake through binding to ALD-DNA (SAP/ALD-DNA). Moreover, macrophage phenotypic switch from a proinflammatory M2b phenotype induced by ALD-DNA alone to an anti-inflammatory M2a phenotype stimulated with SAP/ALD-DNA were found because of PI3K/Akt-ERK signaling activation. Both in vivo SAP supplements and adoptive transfer of ex vivo programmed M2a macrophages induced by SAP/ALD-DNA into SLE mice could efficiently alleviate lupus nephritis. Importantly, increased IL-10 secretion, accompanied by anti-inflammatory effect exerted by M2a macrophages, was found to predominantly impede macrophage M2b polarization. Furthermore, neutralization of IL-10 notably reduced the suppressive effect of M2a macrophages. Our results demonstrate that binding of SAP to ALD-DNA could switch macrophage phenotypic polarization from proinflammatory M2b to anti-inflammatory M2a via PI3K/Akt-ERK signaling activation, thus exerting protective and therapeutic interventions on murine lupus nephritis. These data provide a possible molecular mechanism responsible for modulation of macrophage polarization in the context of lupus nephritis and open a new potential therapeutic avenue for SLE.The Journal of Immunology 08/2011; 187(4):1764-77. · 5.52 Impact Factor
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ABSTRACT: Preeclampsia is a major obstetric problem defined by new-onset hypertension and proteinuria associated with compromised placental perfusion. Although activation of the complement system is increased in preeclampsia compared to normal pregnancy, it remains unclear whether excess complement activation is a cause or consequence of placental ischemia. Therefore, we hypothesized that complement activation is critical for placental ischemia-induced hypertension. We employed the reduced utero-placental perfusion pressure (RUPP) model of placental ischemia in the rat to induce hypertension in the third trimester and evaluated the effect of inhibiting complement activation with a soluble recombinant form of an endogenous complement regulator, human complement receptor 1 (sCR1; CDX-1135). On day 14 of a 21-day gestation, rats received either RUPP or Sham surgery and 15mg/kg/day sCR1 or saline intravenously on days 14-18. Circulating complement component 3 decreased and complement activation product C3a increased in RUPP vs. Sham (p<0.05), indicating complement activation had occurred. Mean arterial pressure (MAP) measured on day 19 increased in RUPP vs. Sham rats (109.8±2.8mmHg vs. 93.6±1.6mmHg). Treatment with sCR1 significantly reduced elevated MAP in RUPP rats (98.4±3.6mmHg, p<0.05) and reduced C3a production. Vascular endothelial growth factor (VEGF) decreased in RUPP compared to Sham rats, and the decrease in VEGF was not affected by sCR1 treatment. Thus, these studies have identified a mechanistic link between complement activation and the pregnancy complication of hypertension apart from free plasma VEGF and have identified complement inhibition as a potential treatment strategy for placental ischemia-induced hypertension in preeclampsia.Molecular Immunology 05/2013; 56(1-2):91-97. · 2.65 Impact Factor
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ABSTRACT: Whether interleukin (IL)-6 contributes to tumor necrosis factor (TNF)-induced lethal shock or whether, on the contrary, it is part of a protective feedback system, remains unresolved. Here, we report experiments with IL-6 gene-disrupted mice (IL-60/0). We have tested the susceptibility of these to TNF-induced metabolic changes and lethality in different models, and compared the results with those obtained with IL-6+/+ wild-type mice. We studied the response to TNF in three different models: (i) murine TNF administration; (ii) TNF in galactosamine (GalN)-sensitized mice; (iii) TNF in Bacillus Calmette-Guérin-sensitized mice. We observed no significant difference between the two types of mice in any of the three models. Furthermore, IL-60/0 mice could be equally well desensitized (by IL-1) to TNF/GalN-induced lethality and tolerized to TNF-induced shock as IL-6+/+ mice. We also observed that, in response to turpentine, TNF or IL-1, IL-60/0 mice produced significantly less acute phase proteins (APP) than IL-6+/+ mice. In IL-60/0 mice, less corticosterone was induced by TNF than in the control mice, while the response to adrenocorticotropic hormone was the same. The results indicate that IL-6 is not contributing in a major way to the pathogenesis leading to TNF-induced shock, and that neither IL-6 nor the APP studied are essential for a protective feedback system.European Journal of Immunology 08/1994; 24(9):2237 - 2242. · 4.97 Impact Factor