*Article* **Juglone Suppresses LPS-induced Inflammatory Responses and NLRP3 Activation in Macrophages**

#### **Nam-Hun Kim, Hong-Ki Kim, Ji-Hak Lee, Seung-Il Jo, Hye-Min Won, Gyeong-Seok Lee, Hyoun-Su Lee, Kung-Woo Nam, Wan-Jong Kim and Man-Deuk Han \***

Department of Life Science and Biotechnology, College of Natural Sciences, Soonchunhyang University, Asan, Chuncheongnam-do 31538, Korea; sem5603@gmail.com (N.-H.K.); pgswkt@naver.com (H.-K.K.); marong932@naver.com (J.-H.L.); 309cho@naver.com (S.-I.J.); whm1223@naver.com (H.-M.W.); ssm4914@sch.ac.kr (G.-S.L.); lhs21c77@hanmail.net (H.-S.L.); kwnam1@sch.ac.kr (K.-W.N.); wjkim56@sch.ac.kr (W.-J.K.)

**\*** Correspondence: mdhan@sch.ac.kr; Tel.: +82-41-530-4702

Academic Editors: Raffaele Pezzani and Sara Vitalini Received: 19 June 2020; Accepted: 5 July 2020; Published: 7 July 2020

**Abstract:** The NLRP3 (NACHT, LRR and PYD domains-containing protein 3) inflammasome has been implicated in a variety of diseases, including atherosclerosis, neurodegenerative diseases, and infectious diseases. Thus, inhibitors of NLRP3 inflammasome have emerged as promising approaches to treat inflammation-related diseases. The aim of this study was to explore the effects of juglone (5-hydroxyl-1,4-naphthoquinone) on NLRP3 inflammasome activation. The inhibitory effects of juglone on nitric oxide (NO) production were assessed in lipopolysaccharide (LPS)-stimulated J774.1 cells by Griess assay, while its effects on reactive oxygen species (ROS) and NLRP3 ATPase activity were assessed. The expression levels of NLRP3, caspase-1, and pro-inflammatory cytokines (IL-1β, IL-18) and cytotoxicity of juglone in J774.1 cells were also determined. Juglone was non-toxic in J774.1 cells when used at 10 μM (*p* < 0.01). Juglone treatment inhibited the production of ROS and NO. The levels of NLRP3 and cleaved caspase-1, as well as the secretion of IL-1β and IL-18, were decreased by treatment with juglone in a concentration-dependent manner. Juglone also inhibited the ATPase activities of NLRP3 in LPS/ATP-stimulated J774.1 macrophages. Our results suggested that juglone could inhibit inflammatory cytokine production and NLRP3 inflammasome activation in macrophages, and should be considered as a therapeutic strategy for inflammation-related diseases.

**Keywords:** juglone; NLRP3 inflammasome; caspase-1; IL-1β; IL-18
