Browsing by Author "Anfossi, Renatto"

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    Interferon-β decreases LPS-induced neutrophil recruitment to cardiac fibroblasts
    (2023) Anfossi, Renatto; Vivar, Raul; Ayala, Pedro; Gonzalez-Herrera, Fabiola; Espinoza-Perez, Claudio; Osorio, Jose Miguel; Roman-Torres, Mauricio; Bolivar, Samir; Diaz-Araya, Guillermo
    Introduction: Cardiac fibroblasts (CF) are crucial cells in damaged heart tissues, expressing TLR4, IFN-receptor and responding to lipopolysaccharide (LPS) and interferon-beta (IFN-beta) respectively. While CF interact with immune cells; however, their relationship with neutrophils remains understudied. Additionally, theimpact of LPS and IFN-beta on CF-neutrophil interaction is poorly understood.Methods: Isolated CF from adult rats were treated with LPS, with or without IFN-beta. This study examined IL-8 secretion, ICAM-1 and VCAM-1 expression, and neutrophil recruitment, as well as their effects on MMPs activity.Results: LPS triggered increased IL-8 expression and secretion, along with elevated ICAM-1 and VCAM-1 expression, all of which were blocked by TAK-242. Pre-treatment with IFN-beta countered these LPS effects. LPS treated CF showed higher neutrophil recruitment (migration and adhesion) compared to unstimulated CF, an effect prevented by IFN-beta. Ruxolitinib blocked these IFN-beta anti-inflammatory effects, implicating JAK signaling. Analysis of culture medium zymograms from CF alone, and CF-neutrophils interaction, revealed that MMP2 was mainly originated from CF, while MMP9 could come from neutrophils. LPS and IFN-beta boosted MMP2 secretion by CF. MMP9 activity in CF was low, and LPS or IFN-beta had no significant impact. Pre-treating CF with LPS, IFN-beta, or both before co-culture with neutrophils increased MMP2. Neutrophil co-culture increased MMP9 activity, with IFN-beta pre-treatment reducing MMP9 compared to unstimulated CF.Conclusion: In CF, LPS induces the secretion of IL-8 favoring neutrophils recruitment and these effects were blocked by IFN-. The results highlight that CF-neutrophil interaction appears to influence the extracellular matrix through MMPs activity modulation.
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    Lipopolysaccharide Activates Toll-Like Receptor 4 and Prevents Cardiac Fibroblast-to-Myofibroblast Differentiation
    (HUMANA PRESS INC, 2017) Bolivar, Samir; Santana, Roxana; Ayala, Pedro; Landaeta, Rodolfo; Boza, Pia; Humeres, Claudio; Vivar, Raul; Munoz, Claudia; Pardo, Viviana; Fernandez, Samuel; Anfossi, Renatto; Diaz Araya, Guillermo
    Bacterial lipopolysaccharide (LPS) is a known ligand of Toll-like receptor 4 (TLR4) which is expressed in cardiac fibroblasts (CF). Differentiation of CF to cardiac myofibroblasts (CMF) is induced by transforming growth factor-beta 1 (TGF-beta 1), increasing alpha-smooth muscle actin (alpha-SMA) expression. In endothelial cells, an antagonist effect between LPS-induced signaling and canonical TGF-beta 1 signaling was described; however, it has not been studied whether in CF and CMF the expression of alpha-SMA induced by TGF-beta 1 is antagonized by LPS and the mechanism involved. In adult rat CF and CMF, alpha-SMA, ERK1/2, Akt, NF-kappa beta, Smad3, and Smad7 protein levels were determined by western blot, TGF-beta isoforms by ELISA, and alpha-SMA stress fibers by immunocytochemistry. CF and CMF secrete the three TGF-beta isoforms, and the secretion levels of TGF-beta 2 was affected by LPS treatment. In CF, LPS treatment decreased the protein levels of alpha-SMA, and this effect was prevented by TAK-242 (TLR4 inhibitor) and LY294002 (Akt inhibitor), but not by BAY 11-7082 (NF-kappa beta inhibitor) and PD98059 (ERK1/2 inhibitor). TGF-beta 1 increased alpha-SMA protein levels in CF, and LPS prevented partially this effect. In addition, in CMF alpha-SMA protein levels were decreased by LPS treatment, which was abolished by TAK-242. Finally, in CF LPS decreased the p-Smad3 phosphorylation and increased the Smad7 protein levels. LPS treatment prevents the CF-to-CMF differentiation and reverses the CMF phenotype induced by TGF-beta 1, through decreasing p-Smad3 and increasing Smad7 protein levels.