Views: 23 Author: WenyiZhang、YahuiGuo、YuliangCheng、WeirongYao、HeQian Publish Time: 2022-12-28 Origin: Neural Regeneration Research 18(4): p869-874, April 2023.
Alzheimer's disease (AD) is a neurodegenerative disease with insidious onset and slow progression. The main clinical symptoms include amnesia, memory loss, progressive language impairment, cognitive impairment, and accompanying neuropsychiatric abnormalities. Currently, AD threatens the health of about 50 million people worldwide, and it is estimated that the prevalence of AD will rise to 131 million by 2050. The neuropathological hallmarks of AD are the deposition of extracellular amyloid plaques and the formation of intracellular neurofibrillary tangles, which further lead to synapse loss and neuronal death. Over the past few decades, a variety of pathological hypotheses have been proposed, including β-amyloid deposition, neurofibrillary tangles, choline dysfunction, oxidative stress, neuroinflammation, and others. Despite tremendous efforts and investments targeting these AD-related pathologies, there are still no effective drugs to treat them. Poor drug development outcomes suggest that etiology, the right time to intervene, and the right target are key factors in drug discovery.
Recent evidence suggests that the gut microbiota plays a role in the pathogenesis of AD. There were different abundances of bacteria, including Bacteroidetes, Firmicutes, and Bifidobacteria, between healthy people and AD patients. Another clinical study reported decreased levels of rectal Eubacteria and increased abundance of Escherichia/Shigella in amyloid-positive patients, which may be related to the systemic inflammatory state in cognitively deficient patients. Gut microbiota disturbances are often accompanied by compromised gut barrier (leaky gut), elevated plasma pro-inflammatory microbial metabolites lipopolysaccharide (LPS) and bacterial amyloid, ultimately leading to systemic inflammation. Previous studies have shown that systemic administration of LPS leads to long-term elevation of Aβ in the hippocampus of mice. Furthermore, LPS activates microglia and induces the expression of pro-inflammatory cytokines in the brain by modulating the toll-like receptor 4 (TLR4)/nuclear factor-κB (NF-κB) signaling pathway. Given the emerging role of the gut microbiome in AD, nutritional interventions may offer a potential opportunity to modulate neurodegenerative disease.
Hydrangea (Sparassis. crispa), commonly known as "cauliflower mushroom", is a valuable edible and medicinal mushroom that contains a variety of bioactive components. Hydrangea polysaccharide (SCP), the main component of this fungus, has multiple pharmacological activities, including anti-angiogenic, anti-tumor, and immunomodulatory effects. In a previous study, a neutral polysaccharide isolated from Hydrangea, named SCP-1, had been shown to be protective against H2O2-induced oxidative stress in mouse hippocampal HT22 cells. Structural information shows that the main chain of galactoglucan contains (1→6)-β-D-Glcp, (1→3)-β-D-Glcp and (1→6)-α-D-Galp, side Chains include (1→3)-β-D-Glcp, (1→4)-β-D-Glcp, T-α-L-Fucp and T-β-D-Glcp. Several galactoglucans isolated from other edible mushrooms, such as Flammulina velutipes, improve cognitive function in different rodent models of aging. Notably, polysaccharides cannot cross the blood-brain barrier to directly affect brain function. However, multiple studies have shown that polysaccharides are a useful dietary source that gut microbiota in the large intestine can use to produce metabolites, some of which may indirectly activate immune cells and microglia in the brain. In particular, short-chain fatty acids, as the main metabolites of intestinal flora, can not only directly provide energy for intestinal epithelial cells, but also alleviate related host diseases. They are important regulators that protect the central nervous system through the "microbiota-gut-brain" (MGB) axis. Therefore, it was hypothesized that administration of SCP-1 could improve cognitive deficits via the MGB axis in a mouse model of AD.
Mice were weighed, classified according to body weight, and divided into four groups using a random number table, 10 in each group: (i) control group (Con), intragastrically administered distilled water, subcutaneously injected with normal saline; (ii) AD Group (Mod), intragastric administration of AlCl3 (20/kg/d), subcutaneous injection of galactose D-Gal (120 mg/kg/d), lasted for 8 weeks; (iii-iv) SCPL group and SCPH group, from the 5th At the beginning of the week, the mice were fed with SCP-1 at doses of 25 mg/kg/d and 100 mg/kg/d, once a day. The body weight of each mouse was recorded weekly. Four weeks after SCP-1 administration, fecal samples were collected and stored at −80◦C until microbiological analysis. These mice were then subjected to behavioral tests.
Figure 1. Protective effect of SCP-1 on memory deficits in D-Gal/AlCl3 mice. (A) Experimental procedures for animal handling. (B) Representative tracking trajectories of mice in training trials.
SCP-1 treatment ameliorated the decline in learning and memory in AD mice in the water maze test and passive avoidance test. In addition, SCP-1 treatment significantly reversed neuropathological changes such as neuronal atrophy and Aβ deposition in the hippocampus, which was consistent with the results of other natural polysaccharides in the same mouse model. Neurotransmitters are chemical massagers that facilitate the transmission of signals between neurons across synapses, thereby affecting brain function. Some neurotransmitters, such as Ach, Glu, GABA and 5-HT, play a key role in the pathogenesis of AD. The cholinergic neurotransmitter Ach is closely related to the learning and memory process of the brain. The deterioration of the cholinergic system may partly contribute to the behavioral abnormalities and deposition of toxic plaques in AD. Together with acetylcholine, excessive accumulation of extracellular Glu leads to activation of NMDA receptors, which leads to excitotoxicity during AD progression. GABA is an important inhibitory neurotransmitter that is decreased in AD. 5-HT has been found to play a role in cognitive processes in AD such as mood disorders, stress and schizophrenia. In the current study, these neurotransmitters were significantly altered in the hippocampus of AD mice. Interestingly, intervention with SCP-1 restored hippocampal concentrations of these neuroactive messengers to normal levels. Although 5-HT is an important neuroactive molecule, no changes were observed in the brains of AD mice after SCP-1 administration, suggesting that the serotonergic system may not be the regulatory target of SCP-1 in AD. These results suggest that SCP-1 treatment ameliorated cognitive dysfunction in AD mouse brains, suppressed Aβ accumulation, and modulated the levels of specific neurotransmitters.
Figure 2. Effects of SCP-1 treatment on D-Gal/AlCl3-induced histopathological changes, amyloidosis and neurotransmitters in mice. (A) Representative images of HE staining in the DG and CA1 regions of the hippocampus. (B) Representative immunohistochemical images of Aβ deposition in the DG and CA1 regions of the hippocampus.
Gut microbiota is largely dependent on dietary fiber or polysaccharides as energy sources, and dietary polysaccharides play important roles in gut microbial ecology and host health. Recent studies have shown that changes in gut microbiota during AD pathogenesis may be alleviated by prebiotic treatment. This study reveals D-Gal/AlCl3-induced changes in the gut microbiota of AD mice at the phylum level, characterized by lower levels of Firmicutes and higher levels of Bacteroidetes, which is consistent with human studies and animal experiments unanimous. The depletion of Firmicutes was closely associated with the enrichment of Aβ accumulation in the brain, implying that reduced levels of Firmicutes may affect the development of AD. Although the high-dose SCP-1 intervention somewhat reversed this trend, no differences were found between the Mod and SCP-1 intervention groups. We also found that specific AD-associated genera were altered in AD mice but not normalized by SCP-1 intervention, suggesting irreversible changes in certain gut taxa during AD pathogenesis. The analysis showed that SCP-1 intervention significantly promoted the growth of intestinal bacteria in the intestine and promoted the synthesis of butyrate in the large intestine. A large number of metabolites produced by gut microbiota may directly or indirectly affect brain function. Among them, SCFAs such as butyrate, propionate, and acetate are thought to regulate the host immune system and play key roles in central nervous system function. Ample evidence has shown that SCFAs are not only energy substrates for host cells, but also multifunctional molecules that mediate diverse physiological roles, such as maintaining intestinal integrity, improving cognitive function, and modulating immune responses. In particular, butyrate achieves neuroprotection by modulating neurotrophic factors for cell growth, inhibiting microglial activation, and increasing the permeability of the blood-brain barrier. In this study, SCP-1 intervention significantly increased the fecal SCFAs concentrations of AD mice, especially the levels of acetate and butyrate. These results suggest that SCFAs produced by gut microbiota play an important role in the protective effect of SCP-1 in AD mice.
Comprehensive intestinal flora analysis showed that high-dose treatment with SCP-1 inhibited the growth of Escherichia/Shigella in AD mice and promoted the growth of Eubacterium ventricular flora. This finding is consistent with clinical studies suggesting that increased abundance of the pathogenic bacteria Escherichia/Shigella may be associated with peripheral inflammation in patients with cerebral amyloidosis and cognitive deficits. Furthermore, the Eubacterium compartment group was inversely correlated with the inflammatory mediators IL-6 and IL-8. Dysbiosis of intestinal flora may lead to increased intestinal barrier permeability, trigger systemic inflammation, damage the blood-brain barrier, promote neuroinflammation, and ultimately exacerbate neurodegeneration in AD. Significant reductions in serum pro-inflammatory cytokines (IL-6, TNF-α, and IL-1β) were observed in the SCP-1 high-dose group. Consistent with the results of peripheral inflammation, SCP-1 intervention significantly improved the expression of tight junction proteins in the colon, accompanied by a decrease in serum LPS levels.
Figure 3. Correlation between TLR4-NF-kB-mediated brain signaling pathways and intestinal flora imbalance. Red arrows indicate positive correlations, and blue arrows indicate negative correlations. A solid edge indicates a strong and significant correlation, and a dashed line indicates an insignificant correlation.
AD进展的另一个潜在机制是神经炎症，其中小胶质细胞和星形胶质细胞包围神经元并导致炎症。肠道炎症和肠道屏障功能的损害使细菌源性病原体和内毒素LPS更易进入循环系统，导致神经炎症甚至神经紊乱。目前，SCP-1干预抑制了AD小鼠大脑中小胶质细胞和星形胶质细胞的激活，并下调了IL-6、IL-1β和TNF-α的水平。TLR4是一种跨膜蛋白，主要在小胶质细胞中表达，可触发先天性宿主免疫。LPS是一种典型的细菌病原体相关分子模式，可能通过激活TLR4/NF-κB信号通路释放炎症分子，如IL-6和TNF-α。Aβ是另一种TLR配体，通过类似于LPS的途径促进神经炎症。NF-κB是与炎症和免疫反应密切相关的重要转录因子。它以非活性形式存在于与κB抑制剂结合的细胞质中。TLR配体或细胞因子激活信号转导导致κB抑制剂激酶快速磷酸化，进而促进NF-κB的释放和激活。为了进一步研究SCP-1干预改善认知缺陷和神经炎症的信号通路，检测了海马组织中TLR4、NF-κB和磷酸化NF-κB的蛋白表达，结果表明SCP-1处理显著抑制了AD小鼠海马TLR4和NF-kB的表达。
Figure 4. Effect of SCP-1 on D-Gal/AlCl3-induced neuroinflammation in mice. Representative images of (A) Iba1 and (B) GFAP immunohistochemistry in the brain.
In summary, SCP-1 can prevent the occurrence of AD, which may involve the regulation of intestinal flora and the suppression of inflammation, which will provide potential application value for the prevention of AD.
