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. 2016 Nov 18;11(11):e0166642.
doi: 10.1371/journal.pone.0166642. eCollection 2016.

Chlorogenic Acid Combined with Lactobacillus plantarum 2142 Reduced LPS-Induced Intestinal Inflammation and Oxidative Stress in IPEC-J2 Cells

Orsolya Palócz  1 , Erzsébet Pászti-Gere  1 , Péter Gálfi  1 , Orsolya Farkas  1
Affiliations

Chlorogenic Acid Combined with Lactobacillus plantarum 2142 Reduced LPS-Induced Intestinal Inflammation and Oxidative Stress in IPEC-J2 Cells

Orsolya Palócz et al. PLoS One. .

Abstract

This study was carried out to investigate protective effect of chlorogenic acid against lipopolysaccharide-induced inflammation and oxidative stress in intestinal epithelial cells. As a marker of inflammatory response, IL-6, IL-8, TNF-α mRNA and protein levels, furthermore, COX-2 mRNA level were followed up. Intracellular redox status and extracellular H2O2 level were also monitored by two fluorescent assays (DCFH-DA, Amplex Red). Moreover, the effect of gut microbiota metabolites in the above mentioned processes was taken into account in our model using Lactobacillus plantarum 2142 bacterial strain. Our data revealed that chlorogenic acid had significant lowering effect on the inflammatory response. Treatment with chlorogenic acid (25-50 μM) significantly decreased gene expression and concentration of proinflammatory cytokines IL-6 and IL-8 compared to LPS-treated cells. COX-2 and TNF-α mRNA levels were also reduced. Furthermore, chlorogenic acid reduced the level of reactive oxygen species in IPEC-J2 cells. Simultaneous application of chlorogenic acid and Lactobacillus plantarum 2142 supernatant resulted protective effect against LPS-induced inflammation and oxidative stress as well.

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Conflict of interest statement

The authors have declared that no competing interests exist.

Figures

Fig 1
Fig 1. Structure of chlorogenic acid.
Fig 2
Fig 2. Rate of living IPEC-J2 cells after chlorogenic acid treatment.
Cell viability was measured by Neutral Red uptake method. Effect of chlorogenic acid on cell viability was studied in a dose-(CGA concentrations: 25 μM, 50 μM and 100 μM) and in a time-dependent manner (treatment time: 1 h, 4 h and 24 h). Data are shown as means + standard deviations. CGA: chlorogenic acid.
Fig 3
Fig 3. Relative gene expression of TNF-α in IPEC-J2 cells exposed to LPS treatment (at 10 μg/ml; 1 h treatment).
Effect of CGA (25 μM, 50 μM), Lp 2142 SCS (13.3%) and combined treatment (CGA 25 μM and Lp 2142 SCS on the TNF-α mRNA levels (n = 3-4/group; *P<0.05). Data are shown as means + standard deviations. CGA: chlorogenic acid, Lp 2142: Lactobacillus plantarum 2142.
Fig 4
Fig 4. (a) Relative gene expression and (b) concentration of IL-6 in IPEC-J2 cells exposed to LPS treatment (at 10 μg/ml; 1 h treatment).
Effect of CGA (25 μM, 50 μM), Lp 2142 SCS (13.3%) and combined treatment (CGA 25 μM and Lp 2142 SCS on the IL-6 mRNA levels (n = 3-4/group; *P< 0.05). Data are shown as means + standard deviations. CGA: chlorogenic acid, Lp 2142: Lactobacillus plantarum 2142.
Fig 5
Fig 5. (a) Relative gene expression and (b) concentration of IL-8 in IPEC-J2 cells exposed to LPS treatment (at 10 μg/ml; 1 h treatment).
Effect of CGA (25 μM, 50 μM), Lp 2142 SCS (13.3%) and combined treatment (CGA 25 μM and Lp 2142 SCS on the IL-8 mRNA levels (n = 3-4/group; *P< 0.05). Data are shown as means + standard deviations. CA: chlorogenic acid, Lp 2142: Lactobacillus plantarum 2142.
Fig 6
Fig 6. Relative gene expression of COX-2 in IPEC-J2 cells exposed to LPS treatment (at 10 μg/ml; 1 h treatment).
Effect of CGA (25 μM, 50 μM), Lp 2142SCS (13.3%) and combined treatment (CGA 25 μM and Lp 2142SCS on the COX-2 mRNA levels (n = 3-4/group; *P< 0.05). Data are shown as means + standard deviations. CGA: chlorogenic acid, Lp 2142: Lactobacillus plantarum 2142.
Fig 7
Fig 7. Level of extracellular H2O2 in IPEC-J2 cells exposed to CGA treatment (25 and 50 μM; 24 h).
Effect of CGA on the relative extracellular H2O2 levels (n = 3-4/group; *P< 0.05). Fluorescence measurement was performed by Amplex Red method. Fluorescence was detected 24 h after CGA treatment. Data are shown as means + standard deviations. CGA: chlorogenic acid, Lp 2142: Lactobacillus plantarum 2142.
Fig 8
Fig 8. Intracellular redox status in IPEC-J2 cells exposed to CGA treatment (25 and 50 μM; 24 h).
Effect of CGA on the relative intracellular ROS levels (n = 3-4/group; *P< 0.05). Fluorescence measurement was performed by DCFH-DA method. Fluorescence was detected 24 h after CGA treatment. Data are shown as means + standard deviations. CGA: chlorogenic acid, Lp 2142: Lactobacillus plantarum 2142.
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