Trans-caryophyllene inhibits amyloid β (Aβ) oligomer-induced neuroinflammation in BV-2 microglial cells
Introduction
Alzheimer's disease (AD) is the most common progressive neurodegenerative disorder among aging people worldwide. One of the major pathological hallmarks of AD is extra-neuronal accumulation of beta-amyloid (Aβ) [46]. Aβ is the major component of senile plaques and recent studies have shown that AD-related pathologies are caused primarily by oligomeric forms of the Aβ peptide [35]. A neuro-inflammatory response from the central nervous system (CNS) is a well-known feature of AD. These responses are interceded by the activation of microglia, the resident immune cells of the CNS [47]. Microglia are activated around the amyloid plaques in the AD brain [47]. It is well-known that oligomeric forms of Aβ serve as an inflammatory stimulus for neuronal cells. Neuronal death is caused by the release of several mediators, including inflammatory cytokines such as interleukin (IL)-1β, IL-6, and tumor necrosis factor-α (TNF-α) [2]. In addition, excessive production of reactive oxygen species (ROS), nitric oxide (NO), proteolytic enzymes, and complement factors are also involved in neuronal death in Alzheimer's disease [13]. It is well-known that microglia-mediated neuroinflammation plays a critical role in the pathogenesis of AD [50]. Microglia can identify and bind to Aβ oligomers and fibrils through receptors present on the cell surface [6]. Treatment with oligomeric Aβ stimulates the activation of microglia, which leads to the production of various pro-inflammatory cytokines, chemokines, prostaglandin E2 (PGE2), NO, ROS, and matrix metalloproteinases (MMPs), thus causing direct neuronal cell damage and furthering the pathogenesis of AD [1]. Excessive release of these factors causes neurotoxic effects and is correlated with deteriorated Aβ pathology in post-mortem AD brains and the brains of transgenic pathological rodent models [36]. One of the Toll-like receptors, Toll-like receptor 4 (TLR4) acts as an inflammation mediator. TLR4 signaling has been shown to be involved in the progression of AD [26]. It is possible that the TLR4/NF-кB signaling pathway plays a major role in Aβ-induced neuro-inflammation [28]. Multiple lines of evidence have demonstrated that nonsteroidal anti-inflammatory drugs attenuate neuro-inflammation, suggesting that inhibition of the release of pro-inflammatory mediators may offer promising therapeutic benefits for AD [14]. Intervention with drugs targeting modulation of microglia and productions of proinflammatory cytokines has become an important strategy for subverting the course of the disease [49].
Trans-caryophyllene (TC), a major component in essential oils derived from various species of medicinal plants, including clove stem oil from Syzygium aromaticum, rosemary essential oil from Rosmarinus officinalis, and hempseed oil from Cannabis sativa [17], [18], [30], [42], has been reported to possess many pharmacological properties [29]. It has been reported to exert an anti-spasmodic effect in rat tracheal smooth muscle [43]. Importantly, a recent in vivo study demonstrated that systemic treatment with TC reduced the production of cytokines, such as tumor necrosis factor-α (TNF-α), induced by intraplantar injection of carrageenan in rats, which suggests that TC may represent an important tool for the management and/or treatment of inflammatory diseases [22]. In addition, the neuroprotective effects of TC were identified in a rodent model of cerebral ischemic injury based on the finding that activation of cortical CB2R by TC ameliorates ischemic injury, potentially through modulation of AMPK/CREB signaling [12]. A recent study reported that TC could be used preventatively or therapeutically to block the development and progression of clinical and neurological signs of experimental autoimmune encephalomyelitis (EAE), which was correlated with the hindrance of immune cell activation, neuroinflammation, and demyelinating processes in the central nervous system (CNS) [3]. Interestingly, TC has been identified as a functional nonpsychoactive ligand of cannabinoid receptor type 2 (CB2R) [17]. An elevated level of CB2R has been reported in certain brain regions in human AD patients, and is mainly expressed in the microglia that surround senile plaques [44]. Notably, specific CB2R agonists have been reported to attenuate neuro-inflammation associated with the pathological progression of AD [4]. These findings suggest that TC may be able to exert a protective effect against Aβ-induced neuroinflammation in AD. In this study, our results demonstrated that TC could inhibit Aβ 1–42 oligomer-induced neuro-inflammation in BV-2 microglial cells.
Section snippets
Preparation of Aβ1–42 solution
To generate soluble oligomers, Aβ1–42 peptide (American Peptide Co., USA) or a scrambled Aβ42–1 peptide was dissolved in 1,1,1,3,3,3-hexafluoro-2-propanol (HFIP). Vacuum evaporation was carried out for 24 h under a fume hood to remove HFIP. The peptide film was dissolved to make 5 mM stock followed by sonication for 10 min. HFIP/DMSO and DMSO were used as vehicle controls. Scrambled Aβ42–1 peptide was used as a control for Aβ1–42. Successful oligomerization of Aβ1–42 was evaluated by Tris-Tricine
Results
The molecular structure of TC is shown in Fig. 1A. To determine the cellular compatibility of TC with BV-2 cells, LDH assay was performed at different TC concentrations ranging from 1 to 100 μM. TC exerted a toxic effect on cultured BV-2 cells at concentrations above 50 μM (Fig. 1B). As shown, this effect was 6 fold at 100 μM what it was at 50 μM. In addition, treatment with TC only at concentrations of 10, 25, and 50 μM did not affect the basal levels of TNF-α, IL-6, and IL-1β section, the
Discussion
AD is the most common clinically recognized form of dementia. The fundamental pathogenesis of AD remains controversial. However, increasing evidence suggests that Aβ is a critical factor in the development and progression of AD (Coomaraswamy et al., 2010). Aβ exerts both direct and indirect neurotoxicity [23]. In terms of direct toxicity, Aβ can directly cause neuronal cell death. In terms of indirect toxicity, Aβ can induce microglial cells to produce inflammatory and toxic factors that then
Acknowledgement
This work has been funded by:
1. National Natural Science Foundation of China (NO, 81671183, 81171171).
2. Science and Technology Development Foundation of Shandong Province (NO.2014GSF118117).
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