Abstract
Background
and Purpose: Polysaccharides from Panax ginseng C. A. Meyer (P.
ginseng) are the main active component and exhibit significant
intestinal anti-inflammatory activity. However, the unclear therapeutic
mechanism of ginseng polysaccharide hinders the application for medicine
or functional food.
Experimental Approach: In this study, a polysaccharide
was isolated from P. ginseng (GP). The primary structure and morphology
of GP were studied by HPLC, FT-IR spectra, and scanning electron
microscopy (SEM). Further, its intestinal anti-inflammatory activity and
its mechanism of function were evaluated in experimental systems using
DSS-induced rats, fecal microbiota transplantation (FMT), and
LPS-stimulated HT-29 cells.
Key Results: Results showed that GP restored
mTOR-dependent autophagic dysfunction via modulating the structure of
gut microbiota and blocking the TLR4-MyD88 pathway. Consequently, active
autophagy suppressed inflammation through the inhibition of NF-κB,
oxidative stress, and the release of cytokines. Conclusion
and Implications: Therefore, our research provided a rationale for
future investigations into the relationship between microbiota and
autophagy via TLR4 and revealed the therapeutic potential of GP for
inflammatory bowel disease.
Keywords: Panax ginseng C. A. Meyer; Polysaccharide;
Intestinal inflammation; Gut microbiota; Autophagy.
Introduction
Inflammatory bowel disease (IBD) consists of a group of disorders
including Crohn’s disease (CD) and
ulcerative colitis (UC) and is a kind
of recurrent, refractory gastrointestinal
disease (Rooks &
Garrett, 2016). At present, the incidence of IBD increases yearly.
Inflammatory infiltration, redox imbalance, and gut microbiota dysbiosis
are involved in the initiation, development, and exacerbation of
intestinal inflammatory diseases. Intestinal epithelium plays an
important role in maintaining gut homeostasis and is the main defense
against pathogen invasion (Barker, 2014).
It has been reported that the regulation of gut microbiota can alleviate
the inflammatory response induced by dextran sodium sulphate (DSS) in
mice. Lipopolysaccharide (LPS) is the main component of the cell walls
of many Gram-negative bacteria. The inseparable
relationship between microbiota dysbiosis, LPS
content and abnormal immune response has been reported previously
(Miller, Choi, Wiesner, & Bae, 2012;
Rogers, Moore, & Cohen, 1985). Toll-like
receptor 4 (TLR4), which is abundantly expressed in intestinal
epithelial cells, is a gene coding receptor for bacterial LPS
(Di Lorenzo et al., 2015;
Medzhitov, Preston-Hurlburt, & Janeway,
1997). An abnormal microbiome, especially with microbes that produce
LPS, triggers intestinal inflammation, and TLR4 might be the initial
point of microbial interaction. Activated TLR4 recruits the downstream
molecule MyD88 to trigger the phosphorylation of MAPKs
(Q. Wang, Dziarski, Kirschning, Muzio, &
Gupta, 2001), and it is indispensable in orchestrating the secretion of
inflammatory cytokines and oxidative stress response throughout the
initiation, development, and exacerbation of IBD
(Russell et al., 2016;
Tan, Gao, Guo, & Huang, 2014;
Verkade et al., 2016).
Autophagy, a highly conserved process that evolved in eukaryotes, is
involved in maintaining organism homeostasis via lysosome-mediated
self-digestion and recycling of organelles and
proteins (Dikic & Elazar, 2018;
Mizushima, 2018). Cells trigger autophagy
under various stress, such as exposure to toxic environments,
starvation, and ischemiareperfusion
(Netea-Maier, Plantinga, van de Veerdonk,
Smit, & Netea, 2016). It has been reported that autophagy dysfunction
increased susceptibility to inflammatory intestinal diseases
(François et al., 2013;
Hang, Lapaquette, Bringer, &
Darfeuille-Michaud, 2013; Kuenzig et al.,
2017; Strisciuglio et al., 2013).
Repairing hampered autophagy normalized redox imbalance, increased the
clearance of intracellular bacteria, and alleviated inflammation in
intestinal mucosa (Schwerd et al., 2016);
thus, it has become a new target of clinical drug development for IBD.
Mounting evidence suggests the inseparable association between autophagy
impairment and inflammation injury
(Deretic, Saitoh, & Akira, 2013;
Santeford et al., 2016;
M. Zhou et al., 2018). mTOR, a highly
conserved serine/threonine protein kinase, negatively regulates
autophagy upstream, and it has demonstrated great autophagy activating
and inflammation attenuating efficacy in silencing mTOR
(Cosin-Roger et al., 2017). Interestingly,
TLR4-MyD88-MAPK is one of the important pathways in mTOR regulation, and
recent research clearly revealed the relationship between them
(M. Zhou et al., 2018). Accumulating
evidence indicates that autophagy and inflammation are linked by
reciprocal regulation through the microflora-TLR4-mTOR axis.
Mechanistically, microbiota dysbiosis activates the TLR4-MyD88-MAPK
pathway, which is followed by the phosphorylation of mTOR to inhibit
autophagy, thereby aggravating inflammatory injury and oxidative stress
(M. Zhou et al., 2018). Although the
roles of microflora in modulating inflammation and autophagy have had
increased attention in regard to IBD, the development of related drugs
is still in its infancy.
Panax ginseng C. A. Meyer
(P. ginseng) has been widely used
as an herb and functional food in the world
(Li & Ji, 2018). Polysaccharide of P.
ginseng has obvious beneficial effects, including gut microbiota
regulation, intestinal mucosal barrier protection, autophagy promotion,
and alleviation of inflammation and oxidative stress
(Kim, Kim, & Park, 2020). However, the
exact target and mechanism of the P. ginseng polysaccharide in gut
microbiota and autophagy modulation is not well understood. Fecal
microbiota transplantation (FMT) is one of the most effective ways to
regulate the gut microbiota and can potentially reveal the function of
microbiota and establish the causal relationship between flora and
disease (Khoruts, 2018). At present, the
use of FMT with P. ginseng polysaccharide to treat diseases through
intestinal flora is still unknown. Therefore, the present study aims to
purify crude polysaccharides from P. ginseng and to evaluate its gut
microbiota and intestinal anti-inflammatory activity in dextran sulfate
sodium (DSS)-induced rats with FMT. The mechanism of reciprocal
regulation between inflammation and autophagy was estimated by
LPS-induced inflammatory intestinal mucosal cells (HT-29 cells).
Additionally, the changes in cytokines, reactive
oxygen species (ROS), and autophagic proteins were detected in order to
investigate the protective mechanism of the microbiota-autophagy
relationship. Our study provides a reliable theoretical basis for the
application of ginseng polysaccharide as a functional food material in
the treatment of intestinal diseases.