마황 추출물의 in vitro 간세포 염증반응 유도

In vitro hepatocyte inflammation by Ephedra sinica extracts

Abstract

본 연구에서는 마황 70% 에탄올 추출물을 HepG2 세포에 $0.001-100{\mu}g/mL$의 농도로 처리하여 세포사멸, 사이토카인 분비, 세포 내 지방 축적 정도를 측정함으로써 마황의 간독성 기전을 in vitro 실험을 통해 살펴보았다. 마황 추출물 처리에 의해 $5-100{\mu}g/mL$의 농도에서 세포 사멸이 관찰되었다(p<0.05). 마황 추출물 처리에 의해 염증성 사이토카인인 IL-8과 M-CSF의 분비가 각각 0.05-100, $0.5-100{\mu}g/mL$의 농도에서 유의적으로 촉진되었으며, 세포 내 지방 축적은 $0.01-100{\mu}g/mL$의 처리 농도에서 유의적으로 증가하였다(p<0.05). 본 실험에서 마황 추출물은 IL-8 및 M-CSF와 같은 염증성 사이토카인의 분비를 촉진시키고, 간세포에 지방을 축적시킴으로써 간세포에 염증을 유발하는 것으로 나타났다. 이러한 형태의 간독성은 세포 사멸과 같은 심각한 독성을 유발하는 농도보다 10-500배 낮은 농도에서 관찰되어 저농도의 마황섭취에 의해 간염과 같은 간독성이 유발될 수 있음을 시사한다.

In this study, the in vitro hepatotoxic mechanism of Ephedra sinica (ma-huang) was investigated by measuring the degree of cell death, secretion of cytokine, and fat accumulation by treating HepG2 cells with 70% ethanolic extracts of ma-huang. Cell death was observed at concentrations of around $5-100{\mu}g/mL$ by treatment with ma-huang extracts (p<0.05). The secretion of interleukin 8 (IL-8) and macrophage colony-stimulating factor (M-CSF), which are inflammatory cytokines, were significantly promoted at concentrations of around 0.05-100 and $0.5-100{\mu}g/mL$, respectively (p<0.05). In this experiment, it was shown that the extracts of ma-huang stimulate the secretion of inflammatory cytokines, such as IL-8 and M-CSF, and lead to fat accumulation in the hepatocytes, thereby causing inflammation of the hepatocytes. Hepatotoxicity was observed at around 10-500 times lower concentration than the concentration required to cause serious toxicity, such as cell death, suggesting that hepatic toxicity (hepatitis) may be induced at a low dose.

Fig. 1. HepG2 cell viability by ma-huang extract.

Fig. 2. LDH release of HepG2 cells by ma-huang extract.

Fig. 3. IL-8 secretion of HepG2 cells by ma-huang extract.

Fig. 4. M-CSF secretion of HepG2 cells by ma-huang extract.

Fig. 5. Lipid accumulation of HepG2 cells by amiodarone (A) or ma-huang extract (B).

Fig. 6. Confocal imaging of nile red binding in HepG2 cells after exposure to control (A), amiodarone (B) or ma-huang extract (C).