Panaxynol, a bioactive component of American ginseng, targets macrophages and suppresses colitis in mice
ABSTRACT
Ulcerative colitis has a significant impact on the quality of life for the patients, and can substantially increase the risk of colon cancer in patients suffering long-term. Conventional treatments provide only modest relief paired with a high risk of side effects, while complementary and alternative medicines can offer safe and effective options. Over the past decade, we have shown that both American ginseng and its hexane fraction (HAG) have anti-oxidant and anti-inflammatory properties that can suppress mouse colitis and prevent colitis-associated colon cancer. With the goal of isolating a single active compound, we further fractionated HAG, and found the most abundant molecule in this fraction was the polyacetylene, panaxynol (PA). After isolating and characterizing PA, we tested the efficacy of PA in the treatment and prevention of colitis in mice and studied the mechanism of action. We demonstrate here that PA effectively treats colitis in a Dextran Sulfate Sodium mouse model by targeting macrophages for DNA damage and apoptosis. This study provides additional mechanistic evidence that American ginseng can be used for conventional treatment of colitis and other diseases associated with macrophage dysfunction.
INTRODUCTION
Inflammatory bowel diseases (IBDs), including ulcerative colitis (UC) and Crohn’s disease (CD), are debilitating illnesses that significantly affect patients’ lifestyle and carry a high colon cancer risk. IBD prevalence is particularly high in North America and Europe (affecting 3.8 million people), with an economic burden of $30–$45 billion [1–4]. Of note, incidence has been increasing for both males and females over the past 20 years [5]. Frustratingly, conventional treatments of IBD patients have modest outcomes with 20% of patients not responding to anti-TNFα antagonists [6], and toxicity leads to dangerous side effects. As such, about half of all IBD patients (millions) turn to complementary and alternative medicines (CAMs). Although CAMs have been used for thousands of years, there is a gap in our knowledge of the mechanisms that support their effectiveness. Understanding these mechanisms will not only lead to standardized and more efficient treatment for IBD outside of toxic FDA-approved drugs but will also better our understanding of the potential applications of CAMs for other diseases with similar mechanisms.
Inflammation generally occurs as an acute response to an injury and infection. This response is initiated by the activation of sentinel immune cells, such as macrophages and dendritic cells that reside around the injured or infected area, which then release chemokines and cytokines and may further recruit other immune cells [7–9]. Control of the immune response to infection is essential to preventing it from becoming a chronic condition. This control is done through apoptosis of immune cells via the tumor suppressor protein 53 (p53) [10, 11]. Low p53 levels in macrophages were observed to be a cause for higher expression of NF-κB-targeted, pro-inflammatory cytokines such as interleukin-6 (IL-6) and tumor necrosis factor (TNF), which are involved in chronic inflammation such as IBD [12–14].
The natural herb, American ginseng (Panax quinquefolius; AG), improves mental performance and detrimental end points associated with diseases, such as cardiovascular disease, diabetes, and influenza [15, 16]. Over the past decade, we have shown that AG has anti-oxidant and anti-inflammatory properties and is able to suppress mouse colitis and prevent colon cancer associated with colitis [11, 17, 18]. Using bioassay-guided fractionation, we have shown that a hexane fraction of AG was particularly potent in this capacity [19–21].
Polyacetylenes are a distinct group of naturally occurring products, whose numerous pharmacological properties have been recognized [22, 23]. PA ([3(R)-(9Z)-heptadeca-1, 9-dien-4, 6-diyn-3-ol]; falcarinol) is a bioactive member of this family. It has been identified in both traditional herbal medicines, such as AG, and dietary plants, e. g., carrots, celery, and fennel [24]. Interestingly, PA has been shown to have anti-cancer properties [24–27] and neuroprotective effects [28–30]. However, there remains an unanswered question regarding PA’s potential as an anti-inflammatory molecule and, therefore, its capacity to suppress chronic inflammatory diseases, such as UC. Here, we hypothesize that the most abundant single molecule ingredient of HAG is the active component of reducing inflammation in a mouse model and that this molecule targets macrophages (mФ) for apoptosis resulting in the suppression of colitis in mice.
RESULTS
Panaxynol is the most abundant and a potent anti-inflammatory molecule in AG
We have previously shown that AG and HAG are effective in the treatment of colitis and prevention of colon cancer [11, 17–21]. We have also demonstrated that fatty acids and polyacetylenes are both components in AG and HAG [19]. In moving forward, to better understand the active components of HAG, we sub-fractionated this fraction of AG using liquid chromatography with UV/diode array detection (LC-UV/DAD) (Figure 1A). Fraction 1 (< 10% of the whole HAG) contains multiple minor components including two minor polyacetylenes tentatively identified based on UV spectra (Figure 1B). Fraction 2 (30% of HAG) contains two major polyacetylenes, panaxydiol (peak1) and panaxydol (peak 2), and four minor polyacetylenes tentatively identified based on UV spectra (Figure 1C). Fraction 3 (24% of HAG) contains a major polyacetylene, PA (peak 3), and a fatty acid, linolenic acid (peak 4) (Figure 1D). Fraction 4 (27% of HAG) contains linoleic acid (peak 5) and no detectable polyacetylenes (Figure 1E). F5 (10%) contains minor fatty acids including saturates, and no polyacetylenes (Figure 1F).
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