IFN-γ Inhibits the Suppressive Effects of PGE2 on the Production of Tumor Necrosis Factor-α by Mouse Macrophages

Tumor necrosis factor-α (TNF-α) has become known as a central mediator of responses to endotoxin, rheumatoid diseases, and other forms of inflammation. Current investigations indicate that the production of TNF-α is controlled by other mediators, including interferon-γ (IFN-γ) and prostaglandin E₂ (PGE₂). In the present study, we investigated the regulatory effects of IFN-γ and/or PGE₂ on LPS-induced TNF-α production and mRNA expression in mouse peritoneal macrophages using the enzyme immunoassay and Northern blot analysis, respectively. In response to 10 ng/ml of LPS, TNF-α production reached a maximum at approximately 4 hrs, followed by rapid decline. At the molecular level, TNF-α mRNA accumulated rapidly after LPS exposure, reaching a peak by 3 hr, and declined more rapidly than did the production of TNF-α. Exposure of macrophages to 100 U/ml of IFN-γ caused an increase in both the TNF-α production and mRNA expression induced by LPS. Exogenous PGE₂ caused a dose dependent reduction in LPS-induced TNF-α mRNA accumulation as well as TNF-α production. Macrophages primed with IFN-γ showed the reduced responsiveness to the suppressive effect of PGE₂ on the production of TNF-α and the accumulation of TNF-α mRNA. These findings indicate that the suppressive effects induced by PGE₂ on the accumulation of TNF-α mRNA as well as the production of TNF-α can be reduced by the pretreatment of macrophages with IFN-γ. These studies demonstrate the role of IFN-γ as an immunomodulating compound that may effectively regulate TNF-α production by modulation of macrophage responsiveness to PGE₂.     

The Influence of Retinoic Acid and Retinoic Acid Derivatives on β2 Integrins and L-Selectin Expression in HL-60 Cells In Vitro

A decreased expression of the 2-integrin CD11b molecules on peripheral neutrophils from patients with pustular psoriasis occurred during treatment with retinoid compounds. Since this effect could not be mimicked in vitro with isolated peripheral neutrophils, the effect of retinoid compounds on cell differentiation was investigated. The promyelocytic cell line, HL60, was used to study what effect different retinoid compounds had on the cell surface expression of CD11b and L-selectin (CD62L) molecules, complement-mediated phagocytosis, adhesion and the oxidative burst. Retinoid-differentiated cells showed a significantly lower expression of CD11b and CD62L, and a decreased phagocytosis and oxidative burst compared to DMSO-differentiated HL60 cells or peripheral blood neutrophils. The diminished expression of 2-integrins or L-selectin did not affect their adhesion to non-activated or lipopolysaccharide-activated endothelial cells in vitro but may however affect adhesion to vascular endothelium under shear forces during blood flow. These results suggest that retinoid treatment could affect several early steps in the inflammatory process.     

PD-L1 Regulates Inflammation in LPS-Induced Lung Epithelial Cells and Vascular Endothelial Cells by Interacting with the HIF-1α Signaling Pathway

Inflammation - Sepsis-induced lung injury was the most common cause of death in patients. This study aimed to investigate whether PD-L1 regulates the inflammation in LPS-induced lung epithelial...     

Polyinosinic-Polycytidylic Acid Induces the Expression of GRO-α in BEAS-2B Cells

Growth-related oncogene protein-α (GRO-α)/CXCLl is a chemokine that activates neutrophils and plays an important role in inflammatory reactions. Polyinosinic-polycytidylic acid (poly IC) is a synthetic double-stranded RNA (dsRNA), which is a ligand for Toll-like receptor-3. Poly IC mimics viral infection when applied to cells and induces inflammatory and immune responses. In the present study, we found the induction of GRO-α in BEAS-2B bronchial epithelial cells treated with poly IC. Pretreatment of cells with 2-aminopurine, an inhibitor for dsRNA-dependent protein kinase (PKR), inhibited the expression of GRO-α-induced by poly IC. Overexpression of interferon-regulatory factor-3 (IRF-3) or retinoic-acid inducible gene-I (RIG-I) enhanced the induction of GRO-α by poly IC. PKR, IRF-3, and RIG-I may be involved in the poly IC-induced expression of GRO-α in BEAS-2B cells. Airway viral infection may elicit GRO-α expression in the bronchial epithelium, which may be implicated in inflammatory and immune reactions.     

Effects of Gamma-Aminobutyric Acid on the Hering–Breuer Inspiration-Inhibiting Reflex

Acute experiments on rats were performed to study the effects of intraventricular microinjections of gamma-aminobutyric acid (GABA) on the volume-time parameters of external respiration and the inspiration-inhibiting Hering–Breuer reflex. The state of this reflex before and after GABA administration was assessed in terms of the extent of changes in the duration and amplitude of inspiratory oscillations in intrathoracic pressure in response to end-expiratory occlusion of the trachea. Administration of 20 μM GABA into the lateral ventricles of the brain decreased the minute ventilation (due to reductions in the respiratory frequency and respiratory volume), weakened respiratory muscle contractions, and decreased the peak airflow rate on inspiration and expiration. The response to end-expiratory occlusion decreased significantly after administration of GABA, demonstrating the involvement of GABAergic mechanisms in mediating the inspiration-inhibiting Hering–Breuer reflex.     

C-Reactive Protein Induces TNF-α Secretion by p38 MAPK–TLR4 Signal Pathway in Rat Vascular Smooth Muscle Cells

Atherosclerosis is a chronic inflammatory disease. C-reactive protein (CRP) not only is an inflammatory marker but also regulates the expressions of other inflammatory cytokines associated with the pathogenesis of atherosclerosis. Toll-like receptor 4 (TLR4) also contributes to atherogenesis via transducting inflammatory signals. Herein, our studies focused on characterizing the effect of CRP on tumor necrosis factor α (TNF-α) production and TLR4-related molecular mechanisms in rat vascular smooth muscle cells (VSMCs). The results showed that CRP stimulated VSMCs to secrete TNF-α and enhanced TLR4 expression in a time-concentration-dependent manner. TLR4 knockdown significantly inhibited CRP-induced TNF-α generation, and p38 mitogen-activated protein kinase (MAPK) blocker SB203580 depressed TLR4 expression and TNF-α production initiated by CRP in VSMCs. The data demonstrate that CRP triggers an inflammatory response in rat VSMCs by inducing TNF-α secretion, which is mediated by p38 MAPK–TLR4 signaling pathway.     

Effect of Thalidomide on the Expression of TNF-α m-RNA and Synthesis of TNF-α in Cells from Leprosy Patients with Reversal Reaction

Hypersensitivity reactions called reversal reaction (RR) and erythema nodosum leprosum (ENL) occur in leprosy. They are characterized by an increase in tumor necrosis factor-alpha (TNF-α). Thalidomide is an effective treatment for ENL but not RR. Its effectiveness in ENL is attributed to inhibition of TNF-α, and this does not explain its failure to treat RR. We assessed thalidomide's effect on TNF-α in RR. Mononuclear cells from RR and non-RR patients and healthy individuals were treated with thalidomide and M.leprae (AFB), a cytosol fraction of M. leprae or Dharmendra lepromin. Thalidomide suppressed TNF-α, but when some RR patients' cells were stimulated with AFB, it enhanced TNF-α.     

The Effects of Cyclic Stretching on the Proliferation of Lung Adenocarcinoma Cells in Vitro

Using FX-4000 strain unit, the prolieration in human lung adenocarcinoma A549 cells that underwent mechanical strain of different waveform、frequency and duration were studied. Image analysis revealed that cellular proliferation rate(PR) reduced significantly after cells were subjected to square wave with 0～20% elongation at frequency 30,40,50 and 60 cycles/min for 2h. The PR had no distinct difference at heart wave , triangle wave and sine wave group compared with control. It is concluded that square wave and higher frequency play an important role in inhibiting A549 cells proliferation.     

Retinoic Acid Receptor γ-Dependent Signaling Cooperates with BMP2 to Induce Osteoblastic Differentiation of C2C12 Cells

All-trans retinoic acid and bone morphogenetic protein 2 (BMP2) synergistically induced an alkaline phosphatase (ALP) activity, one of the osteoblastic differentiation markers, and promoted the extracellular matrix calcification in a myoblastic C2C12 cell culture system. The induced ALP mRNA was not suppressed in the presence of a protein synthesis inhibitor, suggesting that the de novo protein synthesis does not influence this induction. There are three isotypes for the retinoic acid receptor (RARα, RARβ, RARγ). Both the ALP activity and the extracellular matrix calcification were inhibited by the addition of the specific siRNA for RARγ, but not by that for RARα or RARβ. When the effects of the RAR subtype-specific agonists on the ALP activity in the presence of BMP2 were examined, the RARγ-specific agonist was the most effective. The ALP activity induced by any RAR subtype-specific agonist was inhibited by the addition of the specific siRNA for RARγ, but not by that for RARα or RARβ. These results suggest that a RARγ-dependent functional crosstalk is present between the retinoic acid and BMP2 signaling to induce osteogenic transdifferentiation in myoblastic C2C12 cells.     

Effects of Low-Dose γ-Irradiation on the Counts of Immunocompetent Cells in a Model Experiment on Primates

Cellular immunity was studied by flow cytometry with Becton Dickinson monoclonal antibodies in clinically healthy Macaca mulatta males before and after low-dose exposure to ionizing radiation. It was shown that T and B cells are radiosensitive, B cells being more sensitive, which is seen from a significant drop of their count. Natural killers are radioresistant. The count of immunocompetent cells recovers sooner after single compared to fractionated irradiation in the same summary dose.     

Retinoic Acid Enhances the Production of IL-10 While Reducing the Synthesis of IL-12 and TNF-α from LPS-Stimulated Monocytes/Macrophages

Vitamin A and its metabolites, e.g., all trans-retinoic acid (atRA) and 9-cis-retinoic acid have attracted considerable attention as compounds that have a broad range of immune modulating effects on both humoral and cellular immune responses. The cellular and molecular mechanisms that underlie the effects of retinoids on the immune system remain to be more clearly defined. These immune modulating effects of atRA may be mediated by cytokines elaborated by monocytes and other cell types. To further understand the mechanism(s) by which retinoids affect the immune response, we examined the effects of atRA on several proinflammatory and immune modulating cytokines produced by monocytes. The effects of atRA on LPS-induced mRNA expression of IL-10, IL-12p40, TNF-α, IL-18, and TGF-β in the THP-1 monocyte/macrophage cell line and in cord blood mononuclear cells were measured by competitive RT-PCR. The ELISPOT was employed to evaluate IL-10 and TNF-α protein production enumerating the number of IL-10 and TNF-α producing cells. The addition of atRA to cell cultures potentiated the LPS-induced IL-10 mRNA expression and the number of IL-10 secreting cells from THP-1 cells and cord blood mononuclear cells. In contrast, the addition of atRA inhibited the LPS-induced TNF-α and IL-12p40 mRNA expression, and the number of ELISPOT positive cells for TNF-α. atRA did not change the LPS-induced mRNA expression of IL-18 and TGF-β. These results suggest that atRA may have multiple effects on LPS-induced monocyte/macrophage derived cytokines. While atRA downregulated the proinflammatory cytokines, e.g., IL-12 and TNF-α, the production of an immune modulating cytokine, IL-10 was enhanced by atRA. The effects of atRA on these cytokines may play an important role in the modulation of the immune and inflammatory responses.     

Effects of propofol on P2X7 receptors and the secretion of tumor necrosis factor-α in cultured astrocytes

Upon CNS injury, adenosine-5′-triphosphate is released and acts on P2X7 receptors, which might influence many cytokines secretion from glial cells and, in turn, affects the survival of neurons. Propofol, an intravenous anesthetic, has been shown to provide neuroprotective effect. However, the effect of propofol on astrocyte-associated processes remains to be clarified. In this study, we investigated the effects of propofol on P2X7 activity in astrocytes and tumor necrosis factor-α (TNF-α) secretion from these cells and thereby to infer the possible role(s) of glial P2X7 receptors in propofol neural protective effects. Whole-cell patch clamp results showed that in clinically relevant concentrations (3.3, 10 or 33 μM), propofol increased the P2X7 current amplitudes significantly and propofol in 10 μM extended the inactivation times of P2X7 receptors. Enzyme-linked immunosorbent assay showed that propofol increased the secretion of TNF-α from astrocytes in high concentration (300 μM), while inhibited in clinically relevant concentration (10 μM). Both of these effects were not influenced by Brilliant blue G. These results suggest that in clinically relevant concentrations, propofol increases the activity of P2X7 receptors in activated astrocytes, but this does not contribute to the downregulation of the secretion of TNF-α.     

Effects of Laminins 332 and 411 on the Epithelial—Mesenchymal Status of Colorectal Cancer Cells

Bulletin of Experimental Biology and Medicine - The effects of laminins 332 and 411 (LM-332 and LM-411) on the epithelial—mesenchymal transformation of colorectal cancer cells (lines HT-29,...     

Killing Effects of IFN R−/− Mouse NK Cells Activated by HN Protein of NDV on Mouse Hepatoma Cells and Possible Mechanism with Syk and NF-κB

The objective of this article is to evaluate whether the tumoricidal activity of mouse IFN R^−/− nature killer (NK) cells is induced by Newcastle disease virus hemagglutinin-neuraminidase (NDV-HN) stimulation, and to investigate what is the mechanism of the HN-stimulated NK cells to kill mouse hepatoma cell line in vitro. The mouse IFN R^−/− NK cells were stimulated for 16 hr with 500 ng/mL NDV-HN in 1640 medium. Quantify the cytotoxic activities of NK cells against mouse hepatoma cells (Hepa1-6) by flow cytometry. Granzymes B (GrB) and Fas/FasL concentrations in the supernatants of IFN R^−/− NK cells medium were determined by specific ELISA assay. The expression of cell surface GrB and Fas was determined by Western blot. NDV-HN stimulation enhanced tumoricidal activity of IFN R^−/− NK cells toward Hepa1-6 in vitro. Treating with anti-HN neutralizing mAb induced significant decline in the cytotoxicity of IFN R^−/− NK cells toward Hepa1-6 cell line (P < 0.05). After treating with anti-HN protein (1 μL/mL), Syk-specific inhibitor Herbimycin A(250 ng/mL) and NF-κB inhibitor PDTC (500 ng/mL) downregulated the tumoricidal activity of HN-stimulated IFN R^−/− NK cells (P < 0.05). Moreover, significant suppressions in the production of GrB and Fas/FasL were observed in HN-stimulated IFN R^−/− NK cells (P < 0.05). Thus, we concluded that killer activation receptors pathway is involved in the IFN-γ-independent GrB and Fas/FasL expression of NDV-HN-stimulated IFN R^−/− NK cells, and these are activated by Syk and NF-κB. Anat Rec, 302:1718–1725, 2019. © 2019 The Authors. The Anatomical Record published by Wiley Periodicals, Inc. on behalf of American Association for Anatomy     

Simvastatin Inhibits IL-1β-Induced Apoptosis and Extracellular Matrix Degradation by Suppressing the NF-kB and MAPK Pathways in Nucleus Pulposus Cells

Statins are widely used hypocholesterolemic drugs that block the mevalonate pathway. Some studies have shown that statins may have the potential to inhibit intervertebral disk (IVD) degeneration (IDD). Interleukin (IL)-1β, a catabolic cytokine, is a key regulator of IDD. This study aimed to investigate the mechanism underlying the effect of simvastatin on IDD. The viability of nucleus pulposus (NP) cells was determined by the methyl-thiazolyl-tetrazolium (MTT) assay. The apoptosis of NP cells was measured by flow cytometric analysis, terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL), and western blotting of relevant apoptotic proteins. The protein levels of catabolic factors and anabolic factors were determined by western blotting. The cells were stimulated with IL-1β in the absence or presence of simvastatin to investigate the effects on matrix metalloproteinase (MMP)-3, MMP-13, a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS)-4, ADAMTS-5, type II collagen, and aggrecan expression. Our findings indicate that simvastatin considerably inhibited IL-1β-induced apoptosis in NP cells. We also found that simvastatin attenuated IL-1β-induced expression and MMP-3, MMP-13, ADAMTS-4, and ADAMTS-5 activities and also reduced the decrease in type II collagen and aggrecan expression. In addition, simvastatin considerably suppressed the nuclear translocation and activation of nuclear factor-kappa B (NF-KB) by inhibiting p65 phosphorylation and translocation and blocking inhibitor kB-α degradation. It also inhibited MAPK pathway activation by blocking c-Jun N-terminal kinase (JNK), p38, and ERK phosphorylation. The results of our study revealed that simvastatin is a potential agent for IDD prevention and treatment.