Aloe-emodin Attenuates Staphylococcus aureus Pathogenicity by Interfering With the Oligomerization of α-Toxin

ORIGINAL RESEARCH published: 15 May 2019 doi: 10.3389/fcimb.2019.00157 Aloe-emodin Attenuates Staphylococcus aureus Pathogenicity by Interfering With the Oligomerization of α-Toxin Lanxiang Jiang 1 , Tian Yi 2 , Ziying Shen 3 , Zihao Teng 1* and Jianfeng Wang 1,2* 1 Department of Dermatology, Second Hospital of Jilin University, Jilin University, Changchun, China, 2 Key Laboratory of Zoonosis Research, Ministry of Education, Institute of Zoonosis, College of Animal Science, Jilin University, Changchun, China, 3 Laboratory Animal Center, College of Animal Sciences, Jilin University, Changchun, China Edited by: Yinduo Ji, University of Minnesota Twin Cities, United States Reviewed by: Marat R. Sadykov, University of Nebraska Medical Center, United States William Schwan, University of Wisconsin–La Crosse, United States *Correspondence: Zihao Teng Jianfeng Wang Specialty section: This article was submitted to Molecular Bacterial Pathogenesis, a section of the journal Frontiers in Cellular and Infection Microbiology Received: 01 March 2019 Accepted: 26 April 2019 Published: 15 May 2019 Citation: Jiang L, Yi T, Shen Z, Teng Z and Wang J (2019) Aloe-emodin Attenuates Staphylococcus aureus Pathogenicity by Interfering With the Oligomerization of α-Toxin. Front. Cell. Infect. Microbiol. 9:157. doi: 10.3389/fcimb.2019.00157 α-toxin, an essential virulence factor secreted by Staphylococcus aureus (S. aureus), is a critical exotoxin in multiple infections. In this study, we found that aloe-emodin (AE), a natural compound lacking anti-S. aureus activity, could inhibit the hemolytic activity of α-toxin. Oligomerization assays, molecular dynamics simulations, and fluorescence-quenching analyses were used to determine the mechanism of this inhibition. The oligomerization of α-toxin was restricted by the engagement of AE with K110, T112, and M113 of the toxin, which eventually resulted in inhibition of the hemolytic activity. Lactate dehydrogenase and live/dead assays demonstrated that AE decreased the injury of human lung epithelial cells (A549) and mouse lung macrophages (MH-S) mediated by S. aureus. Furthermore, treatment with AE showed robust protective effects in mice infected by S. aureus. These findings suggest that AE effectively inhibited the pore-forming activity of α-toxin and showed a protective effect against S. aureus virulence in vitro and in vivo, which may provide a new strategy and new antibacterial agent for clinical treatment of S. aureus infections. Keywords: Staphylococcus aureus, α-toxin, aloe-emodin, antibiotic-resistant, pneumonia INTRODUCTION Staphylococcus aureus (S. aureus), one of the most common pathogens, is a critical cause of many local and systemic infections ranging from pneumonia, sepsis, and andocarditis, to osteomyelitis. Among these diseases, S. aureus pneumonia is one of the most serious ventilator-associated infections, with ∼10–25% mortality and, for some secondary infections, the rate can even reach 75% (Gillet et al., 2002; Del Giudice et al., 2011; Li et al., 2011; Chastre et al., 2014). Although lactams, aminoglycosides, tetracyclines, sulfonamides, and other major antimicrobial drugs have been commonly used in the past century, we still cannot effectively inhibit the S. aureus pneumonia observed in the clinic most of the time. However, the abuse of antibiotics has led to many resistant strains, and treatment of S. aureus infections has cost at least 450 million dollars due to the increasing resistance (Parvizi et al., 2010; Song et al., 2010). In Europe, ∼10–25% of S. aureus isolated from hospitals were observed to be methicillin-resistant S. aureus (MRSA), and the proportion has reached 50% in some regions (Commun, 2011). Even worse, the proportion of MRSA appeared to reach the highest level in years in parts of east Asia, such as Taiwan and South Korea, with an average rate of 77.6% (Chen and Huang, 2014). Since the twentieth century, Frontiers in Cellular and Infection Microbiology | www.frontiersin.org 1 May 2019 | Volume 9 | Article 157 Jiang et al. Attenuation of S. aureus Pathogenicity by Aloe-emodin MIC Determination the multiresistance of MRSA has become more complicated, which typically results in a delay in clinical treatment (Mendes et al., 2013). Currently, vancomycin is the most commonly used drug to treat MRSA-associated pneumonia (Wunderink et al., 2003). However, the sensitivity of MRSA to vancomycin has been gradually decreasing for years and, given the current trends, the time required for the spread of resistant strains is much less than the time required for research and application of a new medicine. Accordingly, no treatments may be available for MRSA pneumonia in the future, and we need a new treatment strategy to replace the old antibiotic use regimens. Several studies have reported that targeting virulence factors typically results in weak pathogenicity of pathogens, suggesting that this may be a promising strategy in the treatment of S. aureus pneumonia (Qiu et al., 2012a,b; Wang et al., 2016). During infection, a variety of virulence factors are secreted for invasion and colonization, including exotoxin and surfaceassociated protein (Vandenesch et al., 2012). α-toxin is one of the most important exotoxins produced by S. aureus and plays a key role in the course of multiple diseases as a pore-forming protein. It is a 33.2 kDa water-soluble monomer encoded by hla and can oligomerize into a 232.4 kDa membrane-inserted heptamer that penetrates the membrane (Gouaux, 1998; Nguyen and Kamio, 2004). The oligomer comprises seven monomers and consists of three major domains, including the cap domain, the rim domain, and the stem domain, which forms the transmembrane channel (Gouaux et al., 1994; Song et al., 1996). Many types of mammalian cells, including monocytes, erythrocytes, macrophages, and epithelial cells, are sensitive to α-toxin (Gouaux, 1998; Nygaard et al., 2012). For S. aureus pneumonia, studies have reported the destructive effect of α-toxin on the air-blood barrier, and a mutant strain lacking α-toxin showed decreased toxicity in animal models (McElroy et al., 1999; Xu et al., 2015). Therefore, targeting α-toxin is a promising therapeutic strategy for S. aureus infections, particularly MRSA pneumonia. Aloe-emodin [AE; 1,8-dihydroxy-3-(hydroxymethyl)anthraquinone] (Figure 1A) is a common active compound derived from the leaves of Aloe vera and Rheum officinale (Dutta et al., 2007) that has been reported to possess antimicrobial, antiviral, and hepatoprotective activities (Eshun and He, 2004) as well as anticancer activity toward hepatoma cells, lung squamous cell carcinoma, and neuroectodemal tumors (Pecere et al., 2000; Lee, 2001; Kuo et al., 2002). In this study, we observed that AE can inhibit the hemolytic activity of S. aureus without decreasing the expression of α-toxin. In addition, we evaluated the protective effect of AE against MRSA in vitro and in vivo. The MIC of AE against USA30 (...truncated)