J Nutr Health.  2016 Aug;49(4):241-246. 10.4163/jnh.2016.49.4.241.

Aloe-emodin inhibits Pam₃CSK₄-induced MAPK and NF-κB signaling through TLR2 in macrophages

Affiliations
  • 1Department of Food Science and Nutrition, College of Natural Science, Soonchunhyang University, Asan 31538, Korea. soon56@sch.ac.kr
  • 2Department of Food & Nutrition Education, Graduate School of Education, Soonchunhyang University, Asan 31538, Korea.

Abstract

PURPOSE
Aloe-emodin (AE), an ingredient of aloe, is known to exhibit anti-inflammatory activities. However, little is known about the underlying molecular mechanisms of its inflammatory modulatory activity in vitro. In the present study, we investigated the anti-inflammatory potential of AE using Pam₃CSK₄-stimulated macrophages.
METHODS
RAW 264.7 macrophages were treated with AE (0~20 mM) for 1 h, followed by treatment with Pam₃CSK₄ for 1 h. After incubation, mRNA expression levels of cytokines were measured. The effect of AE on TLR2-related molecules was also investigated in Pam₃CSK₄-stimulated RAW 264.7 macrophages.
RESULTS
AE attenuated Pam₃CSKâ‚„-stimulated expression of proinflammatory cytokines, including tumor necrosis factor-α (TNF-α), interleukin-6 (IL-6), and interleukin-1β (IL-1β) in RAW 264.7 macrophages. Two concentrations of AE (10 µM and 20 µM) effectively reduced mRNA expression of TLR2 by 41.18% and 54.43%, respectively, compared to that in control cells (p < 0.05). AE also decreased nuclear factor-kappa B (NF-κB) activation and mitogen-activated protein kinase (MAPK) phosphorylation. Phosphorylation levels of ERK1/2, p38, and JNK were markedly reduced by 20 µM AE. In particular, AE decreased phosphorylation of ERK in a dose-dependent manner in Pam₃CSKâ‚„-stimulated RAW 264.7 macrophages.
CONCLUSION
Our data indicate that AE exerts its anti-inflammatory effect by suppressing TLR2-mediated activation of NF-κB and MAPK signaling pathways in macrophages.

Keyword

Aloe-emodin (AE); macrophage; mitogen-activated protein kinase (MAPK); nuclear factorkappa B (NF-κB); Toll-like receptor 2 (TLR2)

MeSH Terms

Aloe
Cytokines
In Vitro Techniques
Interleukin-6
Macrophages*
Necrosis
Phosphorylation
Protein Kinases
RNA, Messenger
Cytokines
Interleukin-6
Protein Kinases
RNA, Messenger

Figure

  • Fig. 1 Effect of AE on Pam3CSK4-induced TNF-α mRNA expression in RAW 264.7 macrophages. RAW 264.7 macrophages were pretreated with 0~20 µM AE for 1 h. The cells were further stimulated with Pam3CSK4 (1 µg/mL). After 1 h, the TNF-α mRNA expression was analyzed in an agarose gel.

  • Fig. 2 Effect of AE on the Pam3CSK4-induced TNF-α (A), IL-6 (B), and IL-1β (C) mRNA expression in RAW 264.7 macrophages. RAW 264.7 macrophages were pretreated with 0~20 µM AE for 1 h and further stimulated with Pam3CSK4 (1 µg/mL). After 1 h, TNF-α, IL-6, and IL-1β mRNA expression was determined. The data are expressed as a fold induction compared with the vehicle-treated cells. The values are the mean ± SD (n = 3). *Significantly different from the Pam3CSK4-stimulated control (p < 0.05).

  • Fig. 3 Effect of AE on the Pam3CSK4-induced TLR2 mRNA expression in RAW 264.7 macrophages. RAW 264.7 macrophages were pretreated with 0~20 µM AE for 1 h and further stimulated with Pam3CSK4 (1 µg/mL). After 1 h, TLR2 mRNA expression was determined. The data are expressed as a fold induction relative to the vehicle-treated cells. The values are the mean ± SD (n = 4). *Significantly different from the Pam3CSK4-stimulated control, p < 0.05.

  • Fig. 4 Effect of AE on the NF-κB activation (A) and MAPK expression (B) in RAW 264.7 macrophages. RAW 264.7 macrophages were pretreated with 0~20 µM AE for 1 h and further stimulated with Pam3CSK4 (1 µg/mL). After 1 h, NF-κB activity was determined (A), and cell lysates were analyzed by MAPK immunoblotting (B). The NF-κB activity data are expressed as % activation compared with the vehicle-treated cells. The values are the mean ± SD (n = 4). *Significantly different from the Pam3CSK4-stimulatedcontrol, p < 0.05.


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