Sulforaphane suppresses TNF-α-mediated activation of NF-kB and induces apoptosis through activation of reactive oxygen species-dependent caspase-3

Abstract

Sulforaphane (SFN) is a biologically active compound extracted from cruciferous vegetables, and possessing potent anti-cancer and anti-inflammatory activities. Here, we show that tumor necrosis factor-α (TNF-α), in combination with a sub-toxic dose of SFN, significantly triggered apoptosis in TNF-α-resistant leukemia cells (THP-1, HL60, U937, and K562), which was associated with caspase activity and poly (ADP-ribose)-polymerase cleavage. We also report that SFN non-specifically inhibited TNF-α-induced NF-κB activation through the inhibition of IκBα phosphorylation, IκBα degradation, and p65 nuclear translocation. This inhibition correlated with the suppression of NF-κB-dependent genes involved in anti-apoptosis (IAP-1, IAP-2, XIAP, Bcl-2, and Bcl-xL), cell proliferation (c-Myc, COX-2, and cyclin D1), and metastasis (VEGF and MMP-9). These effects suggest that SFN inhibits TNF-α-induced NF-κB activation through the suppression of IκBα degradation, leading to reduced expression of NF-κB-regulated gene products. Combined treatment with SFN and TNF-α was also accompanied by the generation of reactive oxygen species (ROS). Pre-treatment with N-acetyl-L-cysteine significantly attenuated the combined treatment-induced ROS generation and caspase-3-dependent apoptosis, implying the involvement of ROS in this type of cell death. In conclusion, the results of the present study indicate that SFN suppresses TNF-α-induced NF-kB activity and induces apoptosis through activation of ROS-dependent caspase-3.

Sulforaphane (SFN) 은 cruciferous vegetables에서 추출한 물질로 항암 · 항염증 효능을 가지고 있다고 알려져 있다. 이 논문에서는 tumor necrosis factor-α (TNF- α) 와 sub-toxic 농도의 SFN을 함께 처리하였을 때, TNF-α-resistant 인체 백혈병 세포주인 THP-1, HL60, U937과 K562에 세포사멸을 효과적으로 유도함을 확인하였다. 또한, SFN의 처리는 TNF-α 에 의해 유도되는 NF-κB의 활성을 억제하였다. 이러한 현상은 IκBα의 인산화 억제, IκBα의 분해와 p65의 핵으로의 이동을 억제함으로써 나타난 결과였다. 또한 SFN은 NF-κB에 의해 유도되는 anti-apoptosis proteins (IAP-1, IAP-2, XIAP, Bcl-2와 Bcl-xL), cell proliferation proteins (c-Myc, COX-2와 cyclin D1)과 metastasis proteins (VEGF와 MMP9)의 발현을 억제하였다. 그리고 SFN과 TNF-α의 동시처리는 reactive oxygen species (ROS)을 생성시켰다. 항산화제인 N-acetyl-_(L)-cystein의 전 처리시 SFN과 TNF-α 에 의해 유도되는 ROS의 생성과 caspase-3 의존적인 apoptosis가 상당히 감소하는 것을 확인하였다. 결과를 종합해 보면, SFN은 IκBα의 분해를 억제 함으로서 TNF-α 에 의해 유도되는 NF-κB의 활성을 억제한다고 보여지며, 이를 통해 NF-κB에 의해 조절 받는 유전자 산물의 발현량을 감소시키고, ROS 의존적인 caspase-3의 활성화를 통해 세포사멸을 유도한다고 사료된다.