scholarly journals Giardia duodenalis Induces Proinflammatory Cytokine Production in Mouse Macrophages via TLR9-Mediated p38 and ERK Signaling Pathways

2021 ◽  
Vol 9 ◽  
Author(s):  
Xudong Pu ◽  
Xin Li ◽  
Lili Cao ◽  
Kaiming Yue ◽  
Panpan Zhao ◽  
...  
Keyword(s):  
Inflammatory Response ◽  
Signaling Pathways ◽  
Proinflammatory Cytokines ◽  
Giardia Duodenalis ◽  
Erk Signaling ◽  
Enzyme Linked Immunosorbent Assay ◽  
Dependent Manner ◽  
Mouse Macrophages ◽  
Host Inflammatory Response

Giardia duodenalis, also known as Giardia lamblia or Giardia intestinalis, is an important opportunistic, pathogenic, zoonotic, protozoan parasite that infects the small intestines of humans and animals, causing giardiasis. Several studies have demonstrated that innate immunity-associated Toll-like receptors (TLRs) are critical for the elimination of G. duodenalis; however, whether TLR9 has a role in innate immune responses against Giardia infection remains unknown. In the present study, various methods, including reverse transcriptase–quantitative polymerase chain reaction, Western blot, enzyme-linked immunosorbent assay, immunofluorescence, inhibitor assays, and small-interfering RNA interference, were utilized to probe the role of TLR9 in mouse macrophage-mediated defenses against G. lamblia virus (GLV)–free or GLV-containing Giardia trophozoites. The results revealed that in G. duodenalis–stimulated mouse macrophages, the secretion of proinflammatory cytokines, including interleukin 6 (IL-6), tumor necrosis factor α (TNF-α), and IL-12 p40, was enhanced, concomitant with the significant activation of TLR9, whereas silencing TLR9 attenuated the host inflammatory response. Notably, the presence of GLV exacerbated the secretion of host proinflammatory cytokines. Moreover, G. duodenalis stimulation activated multiple signaling pathways, including the nuclear factor κB p65 (NF-κB p65), p38, ERK, and AKT pathways, the latter three in a TLR9-dependent manner. Additionally, inhibiting the p38 or ERK pathway downregulated the G. duodenalis–induced inflammatory response, whereas AKT inhibition aggravated this process. Taken together, these results indicated that G. duodenalis may induce the secretion of proinflammatory cytokines by activating the p38 and ERK signaling pathways in a TLR9-dependent manner in mouse macrophages. Our in vitro findings on the mechanism underlying the TLR9-mediated host inflammatory response may help establish the foundation for an in-depth investigation of the role of TLR9 in the pathogenicity of G. duodenalis.

scholarly journals Giardia duodenalis extracellular vesicles regulate the proinflammatory immune response in mouse macrophages in vitro via the MAPK, AKT and NF-κB pathways

2021 ◽  
Vol 14 (1) ◽  
Author(s):  
Panpan Zhao ◽  
Lili Cao ◽  
Xiaocen Wang ◽  
Jianhua Li ◽  
Jingquan Dong ◽  
...  
Keyword(s):  
Immune Response ◽  
Inflammatory Response ◽  
Signaling Pathways ◽  
Innate Immune Response ◽  
Proinflammatory Cytokines ◽  
Giardia Duodenalis ◽  
Innate Immune ◽  
Tnf Α ◽  
Mouse Macrophages

Abstract Background Giardia duodenalis is an extracellular protozoan parasite that causes giardiasis in mammals. The presentation of giardiasis ranges from asymptomatic to severe diarrhea, and the World Health Organization lists it in the Neglected Diseases Initiative. Extracellular vesicles (EVs) are a key mediator of intracellular communication. Although previous studies have shown that G. intestinalis can regulate a host’s innate immune response, the role of G. intestinalis EVs (GEVs) in triggering a G. intestinalis-induced innate immune response remains to be further explored. Methods In this study, GEVs, G. intestinalis and GEVs + G. intestinalis were inoculated into macrophages, respectively. The transcription and secretion levels of proinflammatory cytokines, including interleukin (IL)-1β, IL-6 and tumor necrosis factor alpha (TNF-α), were measured using real-time quantitative PCR (qPCR) and enzyme-linked immunosorbent assays (ELISAs). The phosphorylation levels of the MAPK, AKT and NF-κB signaling pathways in GEV-stimulated mouse macrophages were examined using western blotting and immunofluorescence methods. The roles of activated pathways in the GEV-triggered inflammatory response were determined using inhibition assays, western blotting and ELISAs. Results The results showed that pretreatment with GEVs enhanced with G. intestinalis (GEVs + G. intestinalis) induced IL-1β, IL-6 and TNF-α transcription and secretion from mouse macrophages compared to stimulation with either GEVs or G. intestinalis alone. Inoculation of mouse macrophages with GEVs upregulated the phosphorylation levels of the p38 MAPK, p44/42 MAPK (Erk1/2), AKT and NF-κB signaling pathways and led to the nuclear translocation of NF-κB p65. Blocking the activated p38, Erk and NF-κB signaling pathways significantly downregulated the secretion of proinflammatory cytokines, and blocking the activated AKT signaling pathway demonstrated reverse effects. Conclusions The results of this study reveal that GEVs can enhance G. intestinalis-induced inflammatory response levels in mouse macrophages through activation of the p38, ERK and NF-κB signaling pathways. The role of GEVs in regulating host cell immune responses may provide insights into exploring the underlying mechanisms in G. intestinalis–host interactions. Graphical abstract

scholarly journals Role of MDA5 in regulating CXCL10 expression induced by TLR3 signaling in human rheumatoid fibroblast-like synoviocytes

2021 ◽  
Author(s):  
Tatsuro Saruga ◽  
Tadaatsu Imaizumi ◽  
Shogo Kawaguchi ◽  
Kazuhiko Seya ◽  
Tomoh Matsumiya ◽  
...  
Keyword(s):  
Inflammatory Responses ◽  
Neutralizing Antibody ◽  
Poly I:C ◽  
Enzyme Linked Immunosorbent Assay ◽  
Dependent Manner ◽  
Inflammatory Reactions ◽  
Polyinosinic:Polycytidylic Acid ◽  
Tlr3 Signaling ◽  
Fibroblast Like Synoviocytes

AbstractC-X-C motif chemokine 10 (CXCL10) is an inflammatory chemokine and a key molecule in the pathogenesis of rheumatoid arthritis (RA). Melanoma differentiation-associated gene 5 (MDA5) is an RNA helicase that plays a role in innate immune and inflammatory reactions. The details of the regulatory mechanisms of CXCL10 production and the precise role of MDA5 in RA synovitis have not been fully elucidated. The aim of this study was to examine the role of MDA5 in regulating CXCL10 expression in cultured human rheumatoid fibroblast-like synoviocytes (RFLS). RFLS was stimulated with Toll-like receptor 3 (TLR3) ligand polyinosinic:polycytidylic acid (poly I:C), a synthetic double-stranded RNA mimetic. Expression of interferon beta (IFN-β), MDA5, and CXCL10 was measured by real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR), western blotting, and enzyme-linked immunosorbent assay. A neutralizing antibody of IFN-β and siRNA-mediated MDA5 knockdown were used to determine the role of these molecules in regulating CXCL10 expression downstream of TLR3 signaling in RFLS. Poly I:C induced IFN-β, MDA5, and CXCL10 expression in a concentration- and time-dependent manner. IFN-β neutralizing antibody suppressed the expression of MDA5 and CXCL10, and knockdown of MDA5 decreased a part of CXCL10 expression (p < 0.001). The TLR3/IFN-β/CXCL10 axis may play a crucial role in the inflammatory responses in RA synovium, and MDA5 may be partially involved in this axis.

scholarly journals Inflammasomes inMycobacterium tuberculosis-Driven Immunity

2017 ◽  
Vol 2017 ◽  
pp. 1-9 ◽  
Author(s):  
Sebastian Wawrocki ◽  
Magdalena Druszczynska
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Inflammatory Response ◽  
Immune Responses ◽  
Proinflammatory Cytokines ◽  
Immune Cells ◽  
Protein Complexes ◽  
Inflammasome Activation ◽  
Host Immune Responses ◽  
Multimeric Protein

The development of effective innate and subsequent adaptive host immune responses is highly dependent on the production of proinflammatory cytokines that increase the activity of immune cells. The key role in this process is played by inflammasomes, multimeric protein complexes serving as a platform for caspase-1, an enzyme responsible for proteolytic cleavage of IL-1βand IL-18 precursors. Inflammasome activation, which triggers the multifaceted activity of these two proinflammatory cytokines, is a prerequisite for developing an efficient inflammatory response against pathogenicMycobacterium tuberculosis(M.tb). This review focuses on the role of NLRP3 and AIM2 inflammasomes inM.tb-driven immunity.

scholarly journals Abnormally expressed miR-23b in Chinese Mongolian at high cardiovascular risk may contribute to monocyte/macrophage inflammatory reaction in atherosclerosis

2018 ◽  
Vol 38 (6) ◽  
Author(s):  
Li-ping He ◽  
Xing-sheng Zhao ◽  
Le-ping He
Keyword(s):  
Cardiovascular Risk ◽  
Inflammatory Response ◽  
Low Density Lipoprotein ◽  
Density Lipoprotein ◽  
Nuclear Factor Κb ◽  
Luciferase Reporter ◽  
Enzyme Linked Immunosorbent Assay ◽  
High Cardiovascular Risk ◽  
Peripheral Blood Mononuclear

Background: The prevalence of coronary heart disease (CHD) appears to be high among Chinese Mongolians. MiR-23b has been proven to play a key role in atherosclerosis. The expression and role of miR-23b in the Mongolians at high cardiovascular risk were explored in the present study. Methods: Forty cases of blood samples from the Mongolians at high cardiovascular risk were enrolled in the present study. The expression of miR-23b was quantified by quantitative real-time PCR. To induce monocytes differentiation into macrophages, HP-1 cells were cultured with phorbol 12-myristate 13-acetate. The level of inflammatory markers was determined by the enzyme-linked immunosorbent assay. The interaction between miR-23b and A20 was explored by the dual luciferase reporter assay. Results: The expression of miR-23b in the Mongolian at high cardiovascular risk was higher than that in healthy Mongolian volunteers. Decrease in ATP-binding cassette transporter A1 caused by miR-23b is responsible for TC accumulation in the Mongolian at high cardiovascular risk. MiR-23b enhanced the oxidized low-density lipoprotein (oxLDL)-induced inflammatory response of THP-1 derived macrophage. MiR-23b regulated nuclear factor-κB (NF-κB) pathway through targeting A20. MiR-23b mediated oxLDL-induced inflammatory response of peripheral blood mononuclear cell in the Mongolian at high cardiovascular risk. Conclusion MiR-23b enhanced oxLDL-induced inflammatory response of macrophages in the Mongolian at high cardiovascular risk through the A20/NF-κB signaling pathway, and thus contributing to atherosclerosis.

scholarly journals Effect of Anthocyanin-Rich Extract of Sour Cherry for Hyperglycemia-Induced Inflammatory Response and Impaired Endothelium-Dependent Vasodilation

Nutrients ◽  
2020 ◽  
Vol 12 (11) ◽  
pp. 3373
Author(s):  
Arnold Markovics ◽  
Attila Biró ◽  
Andrea Kun-Nemes ◽  
Mónika Éva Fazekas ◽  
Anna Anita Rácz ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Gene Expression ◽  
Diabetes Mellitus ◽  
Endothelial Dysfunction ◽  
Inflammatory Response ◽  
Proinflammatory Cytokines ◽  
Sour Cherry ◽  
The Elderly ◽  
Human Umbilical Cord ◽  
Dependent Manner

Diabetes mellitus (DM)-related morbidity and mortality are steadily rising worldwide, affecting about half a billion people worldwide. A significant proportion of diabetic cases are in the elderly, which is concerning given the increasing aging population. Proper nutrition is an important component in the effective management of diabetes in the elderly. A plethora of active substances of plant origin exhibit potency to target the pathogenesis of diabetes mellitus. The nutraceutical and pharmaceutical effects of anthocyanins have been extensively studied. In this study, the effect of Hungarian sour cherry, which is rich in anthocyanins, on hyperglycemia-induced endothelial dysfunction was tested using human umbilical cord vein endothelial cells (HUVECs). HUVECs were maintained under both normoglycemic (5 mM) and hyperglycemic (30 mM) conditions with or without two concentrations (1.50 ng/µL) of anthocyanin-rich sour cherry extract. Hyperglycemia-induced oxidative stress and inflammatory response and damaged vasorelaxation processes were investigated by evaluating the level of reactive oxygen species (ROS) and gene expression of four proinflammatory cytokines, namely, tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), interleukin-8 (IL-8), and interleukin-1α (IL-1α), as well as the gene expression of nitric oxide synthase (NOS) endothelin-1 (ET-1) and endothelin-converting enzyme-1 (ECE-1). It was found that hyperglycemia-induced oxidative stress was significantly suppressed by anthocyanin-rich sour cherry extract in a concentration-dependent manner. The gene expression of the tested proinflammatory cytokines increased under hyperglycemic conditions but was significantly reduced by both 1 and 50 ng/µL anthocyanin-rich sour cherry extract. Further, although increased ET-1 and ECE-1 expression due to hyperglycemia was reduced by anthocyanin-rich sour cherry extract, NOS expression was increased by the extract. Collectively, these data suggest that anthocyanin-rich sour cherry extract could alleviate hyperglycemia-induced endothelial dysfunction due to its antioxidant, anti-inflammatory, and vasorelaxant effects.

scholarly journals Caffeic Acid Prevented LPS-Induced Injury of Primary Bovine Mammary Epithelial Cells through Inhibiting NF-κB and MAPK Activation

2019 ◽  
Vol 2019 ◽  
pp. 1-12 ◽  
Author(s):  
Mingjiang Liu ◽  
Guoqing Fang ◽  
Shaojie Yin ◽  
Xin Zhao ◽  
Chi Zhang ◽  
...  
Keyword(s):  
Epithelial Cells ◽  
Cell Viability ◽  
Signaling Pathways ◽  
Caffeic Acid ◽  
Proinflammatory Cytokines ◽  
Mammary Epithelial Cells ◽  
Mammary Epithelial ◽  
Signal Pathways ◽  
Dependent Manner ◽  
Bovine Mammary Epithelial Cells

In our previous study, lipopolysaccharide (LPS) significantly reduced the cell viability of primary bovine mammary epithelial cells (bMEC) leading to cell apoptosis, which were prevented by caffeic acid (CA) through inhibiting NF-κB activation and reducing proinflammatory cytokine expression. While the underlying mechanism remains unclear, here, we determined that LPS induced the extensive microstructural damage of bMEC, especially the mitochondria and endoplasmic reticulum. Then, the obvious reduction of mitochondrial membrane potential and expression changes of apoptosis-associated proteins (Bcl-2, Bax, and casepase-3) indicated that apoptosis signaling through the mitochondria should be responsible for the cell viability decrease. Next, the high-throughput cDNA sequencing (RNA-Seq) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were employed to verify that the MAPK and JAK-STAT signaling pathways also were the principal targets of LPS. Following, the critical proteins (ERK, JNK, p38, and c-jun) of the MAPK signaling pathways were activated, and the release of proinflammatory cytokines (TNF-α, IL-1β, IL-6, and IL-8) regulated by NF-κB and MAPKs was significantly increased, which can promote a cascade of inflammation that induces cell injury and apoptosis. Meanwhile, CA significantly inhibited the activation of MAPKs and the release of proinflammatory cytokines in a dose-dependent manner, which were similar to its effects on the NF-κB activation that we previously published. So we concluded that CA regulates the proteins located in the upstream of multiple cell signal pathways which can reduce the LPS-induced activation of NF-κB and MAPKs, thus weakening the inflammatory response and maintaining cell structure and function, which accordingly inhibit apoptosis.

scholarly journals Binding of Brucella protein, Bp26, to select extracellular matrix molecules

2019 ◽  
Vol 20 (1) ◽  
Author(s):  
Yasmin ElTahir ◽  
Amna Al-Araimi ◽  
Remya R. Nair ◽  
Kaija J. Autio ◽  
Hongmin Tu ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Type I Collagen ◽  
Host Cells ◽  
Mouse Serum ◽  
Enzyme Linked Immunosorbent Assay ◽  
Target Cells ◽  
Type I ◽  
Dependent Manner ◽  
Biolayer Interferometry

Abstract Background Brucella is a facultative intracellular pathogen responsible for zoonotic disease brucellosis. Little is known about the molecular basis of Brucella adherence to host cells. In the present study, the possible role of Bp26 protein as an adhesin was explored. The ability of Brucella protein Bp26 to bind to extracellular matrix (ECM) proteins was determined by enzyme-linked immunosorbent assay (ELISA) and biolayer interferometry (BLI). Results ELISA experiments showed that Bp26 bound in a dose-dependent manner to both immobilized type I collagen and vitronectin. Bp26 bound weakly to soluble fibronectin but did not bind to immobilized fibronectin. No binding to laminin was detected. Biolayer interferometry showed high binding affinity of Bp26 to immobilized type I collagen and no binding to fibronectin or laminin. Mapping of Bp26 antigenic epitopes by biotinylated overlapping peptides spanning the entire sequence of Bp26 using anti Bp26 mouse serum led to the identification of five linear epitopes. Collagen and vitronectin bound to peptides from several regions of Bp26, with many of the binding sites for the ligands overlapping. The strongest binding for anti-Bp26 mouse serum, collagen and vitronectin was to the peptides at the C-terminus of Bp26. Fibronectin did not bind to any of the peptides, although it bound to the whole Bp26 protein. Conclusions Our results highlight the possible role of Bp26 protein in the adhesion process of Brucella to host cells through ECM components. This study revealed that Bp26 binds to both immobilized and soluble type I collagen and vitronectin. It also binds to soluble but not immobilized fibronectin. However, Bp26 does not bind to laminin. These are novel findings that offer insight into understanding the interplay between Brucella and host target cells, which may aid in future identification of a new target for diagnosis and/or vaccine development and prevention of brucellosis.

scholarly journals Leptin Upregulates the Expression of β3-Integrin, MMP9, HB-EGF, and IL-1β in Primary Porcine Endometrium Epithelial Cells In Vitro

2020 ◽  
Vol 17 (18) ◽  
pp. 6508
Author(s):  
Hongfang Wang ◽  
Jinlian Fu ◽  
Aiguo Wang
Keyword(s):  
Signaling Pathways ◽  
Embryo Implantation ◽  
Mapk Signaling ◽  
Dependent Manner ◽  
Reproductive Disorders ◽  
Β3 Integrin ◽  
Epithelium Cells ◽  
Dose Dependent

Obesity has become a global health problem. Research suggests that leptin, a hormone that responds to fat deposition, may be involved in mammalian reproduction; however, its precise role in embryo implantation is poorly understood. Here, primary porcine endometrium epithelium cells (PEECs) were cultured in vitro and used to evaluate the regulatory role of different leptin levels on β3-integrin, MMP9, HB-EGF, and IL-1β, which are, respectively, involved in four critical steps of embryo implantation. Results showed that only 0.01 nM leptin significantly improved β3-integrin mRNA expression (p < 0.05). MMP9 and HB-EGF mRNA expressions were upregulated by 0.10–10.00 nM leptin (p < 0.05). The IL-1β expression level was only increased by 10.00 nM leptin (p < 0.05). β3-integrin, MMP9, HB-EGF, and IL-1β mRNA and protein have a similar fluctuant response to increased leptin. Leptin’s influence on β3-integrin, MMP9, HB-EGF, and IL-1β disappeared when the JAK2, PI(3)K, or MAPK signaling pathways were blocked, respectively. In conclusion, leptin affected porcine implantation by regulating the expression of β3-integrin, MMP9, HB-EGF, and IL-1β in a dose-dependent manner. The signaling pathways of JAK2, PI(3)K, and MAPK may participate in this regulatory process. These findings will contribute to further understanding the mechanisms of reproductive disorders in obesity.

scholarly journals Role of vaspin in porcine ovary: effect on signaling pathways and steroid synthesis via GRP78 receptor and protein kinase A†

2020 ◽  
Vol 102 (6) ◽  
pp. 1290-1305 ◽  
Author(s):  
Patrycja Kurowska ◽  
Ewa Mlyczyńska ◽  
Monika Dawid ◽  
Joelle Dupont ◽  
Agnieszka Rak
Keyword(s):  
Protein Kinase ◽  
Western Blot ◽  
Protein Kinase A ◽  
Signaling Pathways ◽  
Dependent Manner ◽  
Steroid Synthesis ◽  
Ovarian Cells ◽  
Vitro Effect ◽  
Kinase A

Abstract Vaspin, visceral-adipose-tissue-derived serine protease inhibitor, is involved in the development of obesity, insulin resistance, inflammation, and energy metabolism. Our previous study showed vaspin expression and its regulation in the ovary; however, the role of this adipokine in ovarian cells has never been studied. Here, we studied the in vitro effect of vaspin on various kinase-signaling pathways: mitogen-activated kinase (MAP3/1), serine/threonine kinase (AKT), signal transducer and activator of transcription 3 (STAT3) protein kinase AMP (PRKAA1), protein kinase A (PKA), and on expression of nuclear factor kappa B (NFKB2) as well as on steroid synthesis by porcine ovarian cells. By using western blot, we found that vaspin (1 ng/ml), in a time-dependent manner, increased phosphorylation of MAP3/1, AKT, STAT3, PRKAA1, and PKA, while it decreased the expression of NFKB2. We observed that vaspin, in a dose-dependent manner, increased the basal steroid hormone secretion (progesterone and estradiol), mRNA and protein expression of steroid enzymes using real-time PCR and western blot, respectively, and the mRNA of gonadotropins (FSHR, LHCGR) and steroids (PGR, ESR2) receptors. The stimulatory effect of vaspin on basal steroidogenesis was reversed when ovarian cells were cultured in the presence of a PKA pharmacological inhibitor (KT5720) and when GRP78 receptor was knocked down (siRNA). However, in the presence of insulin-like growth factor type 1 and gonadotropins, vaspin reduced steroidogenesis. Thus, vaspin, by activation of various signaling pathways and stimulation of basal steroid production via GRP78 receptor and PKA, could be a new regulator of porcine ovarian function.

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