Tonggyu-tang, a traditional Korean medicine, suppresses pro-inflammatory cytokine production through inhibition of MAPK and NF-κB activation in human mast cells and keratinocytes

Abstract

Background: Allergic diseases including allergic rhinitis, asthma, and atopic dermatitis are increasing worldwide. Common medications used to treat these inflammatory disorders are anti-histamines and corticosteroids, but they have their own limitations such as short duration and severe side effects. Thus, interest in complementary and alternative medicine is continually growing. Here, we investigate the anti-inflammatory mechanisms of Tonggyu-tang (TGT), a traditional Korean medicine that has been used to treat patients with allergic nasal disorders.

Methods: We measured mRNA expressions and production of pro-inflammatory cytokines such as interleukin (IL)-4, IL-6, IL-8 and tumor necrosis factor alpha (TNF-α) by RT-PCR and ELISA assays in HMC-1 (human mast cell line-1) and HaCaT cells, immortalized human keratinocytes. Moreover, we evaluated the effect of TGT on two major inflammation-related pathways, mitogen activated protein kinase (MAPK) and NF-κB signaling pathway in these two cells.

Results: Our results revealed that that TGT significantly reduced the expression and production of inflammatory cytokines such as IL-4, IL-6, IL-8, and TNF-α in the agonist-treated HMC-1 and HaCaT cells. We also found that TGT suppressed MAPK signaling pathway including extracellular signal-regulated kinase (ERK), p38 mitogen-activated protein kinase (p38), and c-Jun N-terminal kinase (JNK) as well as NF-κB pathway, which are known to regulate inflammatory cytokine expression.

Conclusion: Taken together, our results demonstrate that TGT inhibits expression of pro-inflammatory cytokines by suppressing MAPK and NF-kB pathway in both mast cells and keratinocytes, suggesting the potential use of TGT in treating allergic inflammatory diseases.

Keywords: Anti-inflammation; HaCaT; Hmc-1; Mapk; NF-κB; Tonggyu-tang.

Fig. 1

Effect of TGT on expression of pro-inflammatory cytokines in PI stimulated HMC-1 cells. a PI-stimulated HMC-1 cells were treated with or without different doses of TGT for 24 h. Cytokine mRNA levels were measured by RT-PCR. b The density of each band was calculated with image processing program, Image J, and represented as bar graphs. *; compared to control. #; compared to stimulated cells. Control; HMC-1 with PI (−), Stimulation; HMC-1 with PI (+)

Fig. 2

Effect of TGT on the release of pro-inflammatory cytokines in HMC-1 cells. PI-stimulated HMC-1 cells were treated with or without different doses of TGT for 24 h. The levels of secreted pro-inflammatory cytokines in the cell culture supernatant were measured by ELISA. *; compared to control. #; compared to stimulated cells. Control; HMC-1 with PI (−), Stimulation; HMC-1 with PI (+)

Fig. 3

Effect of TGT on the protein expressions of MAPK and NF-κB signaling pathways in HMC-1 cells. PI-stimulated HMC-1 cells were treated with or without different doses of TGT for 24 h. The levels of protein expressions of (a) MAPK signaling pathway and (b) NF-κB signaling pathway in the whole cell lysates of HMC-1 cells were measured by western blot assays

Fig. 4

Effect of TGT on expression of pro-inflammatory cytokines in LPS stimulated HaCaT cells. a LPS-stimulated HaCaT cells were treated with or without different doses of TGT for 24 h. Cytokine mRNA levels were measured by RT-PCR. b The density of each band was calculated with image processing program, Image J, and represented as bar graph. *; compared to control. #; compared to stimulated cells. Control; HaCaT with LPS (−), Stimulation; HaCaT with LPS (+)

Fig. 5

Effect of TGT on the release of pro-inflammatory cytokines in HaCaT cells. LPS-stimulated HaCaT cells were treated with or without different doses of TGT for 24 h. The levels of secreted pro-inflammatory cytokines in the cell culture supernatant were measured by ELISA. *; compared to control. #; compared to stimulated cells. Control; HaCaT with LPS (−), Stimulation; HaCaT with LPS (+)