Dataset Information

The anti-inflammatory effects of asiatic acid in lipopolysaccharide-stimulated human corneal epithelial cells.

ABSTRACT:

Aim

To investigate the anti-inflammatory effects of asiatic acid (AA) on lipopolysaccharide (LPS)-induced inflammatory response in human corneal epithelial cells (HCECs).

Methods

Cell viability was measured using a cell counting kit-8 (CCK-8) assay. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to determine the mRNA expression of interleukin-8 (IL-8), interleukin-6 (IL-6), interleukin-1β (IL-1β), tumor necrosis factor-alpha (TNF-α), and transforming growth factor-β (TGF-β) in HCECs. Intracellular reactive oxygen species (ROS) was measured using the ROS assay kit. Glutathione (GSH) concentration was measured using the total GSH assay kit. Akt1 and Akt phosphorylation (p-Akt1) levels were measured by Western blotting and immunofluorescence.

Results

AA induced toxicity at high concentrations and significantly stimulated the proliferation of HCECs at concentrations of 20 µmol/L for 1h. LPS at concentrations of 300 ng/mL for 1h significantly stimulated the mRNA expression of IL-8, IL-6, IL-1β, TNF-α, and TGF-β in HCECs, while the stimulation effects were significantly inhibited by AA (20 µmol/L). In addition, AA was found to decrease the content of ROS, increase GSH generation, and also inhibit LPS-induced p-Akt in HCECs.

Conclusion

AA decreases the generation of inflammatory factors IL-8, IL-6, IL-1β, TNF-α, and TGF-β in LPS-stimulated HCECs. AA significantly inhibites the intracellular concentrations of ROS and increases GSH generation. AA also inhibites LPS-induced p-Akt in HCECs. These findings reveal that AA has anti-inflammation effects in LPS-stimulated HCECs.

SUBMITTER: Chen H

PROVIDER: S-EPMC5313538 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Publications

The anti-inflammatory effects of asiatic acid in lipopolysaccharide-stimulated human corneal epithelial cells.

Chen Hao H Hua Xiao-Min XM Ze Bai-Chen BC Wang Bin B Wei Li L

International journal of ophthalmology 20170218 2

<h4>Aim</h4>To investigate the anti-inflammatory effects of asiatic acid (AA) on lipopolysaccharide (LPS)-induced inflammatory response in human corneal epithelial cells (HCECs).<h4>Methods</h4>Cell viability was measured using a cell counting kit-8 (CCK-8) assay. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to determine the mRNA expression of interleukin-8 (IL-8), interleukin-6 (IL-6), interleukin-1β (IL-1β), tumor necrosis factor-alpha (TNF-α), and transforming growth fa ...[more]

PMID: 28251074

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Project description:Inflammation and oxidative stress are essential for the pathogenesis of fulminant hepatic failure (FHF). Asiatic acid (AA), which is a pentacyclic triterpene that widely occurs in various vegetables and fruits, has been reported to possess antioxidant and anti-inflammatory properties. In this study, we investigated the protective effects of AA against lipopolysaccharide (LPS) and d-galactosamine (GalN)-induced FHF and the underlying molecular mechanisms. Our findings suggested that AA treatment effectively protected against LPS/d-GalN-induced FHF by lessening the lethality; decreasing the alanine transaminase and aspartate aminotransferase levels, interleukin (IL)-1β, IL-6, and tumor necrosis factor-α production, malondialdehyde formation, myeloperoxidase level and reactive oxygen species generation (i.e., H2O2, NO, and [Formula: see text]), and increasing the glutathione and superoxide dismutase contents. Moreover, AA treatment significantly inhibited mitogen-activated protein kinase (MAPK) and nuclear factor-kappa B (NF-κB) signaling pathway activation via the partial induction of programmed cell death 4 (PDCD4) protein expressions, which are involved in inflammatory responses. Furthermore, AA treatment dramatically induced the expression of the glutamate-cysteine ligase modifier subunit, the glutamate-cysteine ligase catalytic subunit, heme oxygenase-1, and NAD (P) H: quinoneoxidoreductase 1 (NQO1), which are largely dependent on activation of the nuclear factor-erythroid 2-related factor 2 (Nrf2) through the induction of AMP-activated protein kinase (AMPK) and glycogen synthase kinase-3β (GSK3β) phosphorylation. Accordingly, AA exhibited protective roles against LPS/d-GalN-induced FHF by inhibiting oxidative stress and inflammation. The underlying mechanism may be associated with the inhibition of MAPK and NF-κB activation via the partial induction of PDCD4 and upregulation of Nrf2 in an AMPK/GSK3β pathway activation-dependent manner.

Project description:PURPOSE: To investigate the modulatory effect of rat bone marrow mesenchymal stem cells (MSC) on human corneal epithelial cells (HCE-T) stimulated with pro-inflammatory cytokines interferon gamma (IFN-γ) and tumor necrosis factor alpha (TNF-α) in an in vitro co-cultured model. METHODS: HCE-T alone and co-cultured with MSC were stimulated with IFN-γ/TNF for 24 and 48 hours or left untreated. The expression of intracellular adhesion molecule (ICAM)-1, human leukocyte antigen ABC, DR and G (HLA-ABC, HLA-DR, HLA-G) were investigated by flow cytometry. Subcellular localization of nuclear factor-kappa B (NF-κB) and expression of indoleamine 2,3-dioxygenase (IDO) were assessed by immunofluorescence staining and western blot. The concentration of transforming growth factor beta 1 (TGF-β1) in the conditioned media from different cultures was evaluated by enzyme-linked immunosorbent assay. NF-κB and TGF-β1 signaling pathway blocking experiments were performed to analyze associations between the expression of cell surface molecules and the NF-κB transcription pathway, and the expression of IDO and TGF-β1 signaling pathway. RESULTS: IFN-γ/TNF treatment significantly up-regulated expression of ICAM-1, HLA-ABC, and induced de novo expression of HLA-DR and IDO on HCE-T cultured alone, while HLA-G expression remained unaffected. Up-regulation was significantly inhibited by co-culture with MSC. Increased TGF-β1 secretion was detected in 48 h IFN-γ/TNF-stimulated MSC monocultures and HCE-T/MSC co-cultures. MSC attenuated the activation of cytokine-induced NF-κB and IDO induction. Blockade of NF-κB transcription pathway by BMS-345541 significantly reduced the up-regulation of ICAM-1, HLA-ABC, HLA-DR and IDO expression, while blockade of TGF-β1 signaling pathways reversed the modulatory effect of MSC on IDO expression. CONCLUSIONS: MSC reduced the expression of adhesion and immunoregulatory molecules on pro-inflammatory cytokine-stimulated HCE-T via the NF-κB transcription pathway. MSC attenuated expression of IDO through both NF-κB transcription and TGF-β1 signaling pathways. Co-culture of HCEC with MSC therefore provides a useful in vitro model to study the anti-inflammatory properties of MSC on corneal epithelium.

Dataset Information

The anti-inflammatory effects of asiatic acid in lipopolysaccharide-stimulated human corneal epithelial cells.

Aim

Methods

Results

Conclusion

Publications

The anti-inflammatory effects of asiatic acid in lipopolysaccharide-stimulated human corneal epithelial cells.

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