《Gut microbe》发表茯苓多糖通过肠道菌群调控改善酒精性肝脂肪变性研究

《Gut Microbes》的一项研究系统阐明了茯苓不溶性多糖（WIP）通过多靶点调控肠道微生物群落缓解酒精性肝脂肪变性（AHS）的机制，并首次揭示了肠道共生真菌——季也蒙迈耶氏酵母（Meyerozyma guilliermondii）通过生物合成前列腺素E2（PGE2）加重肝损伤的新病理机制。研究显示，在酒精性肝脂肪变性模型中，茯苓不溶性多糖（WIP）口服干预能显著改善肝脏炎症损伤和脂肪堆积。其作用机制包括：1. 调节肠道细菌群落：提高厚壁菌门/变形菌门比值，显著增加毛螺菌科（特别是瘤胃梭菌属和未分类梭菌）丰度；2. 抑制真菌过度增殖：有效抑制乙醇诱导的肠道真菌过度生长；3. 激活保护性信号通路：通过激活PPAR-γ信号减轻结肠上皮炎症，营造肠道低氧环境，从而抑制真菌和变形菌的过度生长。通过培养组学和ITS测序技术，研究团队在AHS小鼠粪便中发现共生真菌季也蒙迈耶氏酵母的异常增殖。将该菌株定植于无菌小鼠后，可显著加重AHS表型。机制深入研究表明，该真菌能通过花生四烯酸的生物转化生成PGE2，而肝脏中这种由肠道真菌诱导的PGE2产生被证实是慢性AHS的重要致病机制之一。该研究不仅证实了茯苓多糖通过协同调控肠道细菌和真菌群落发挥肝保护作用，也为开发针对肠道微生物谱系的酒精性肝病治疗策略提供了新的理论依据和实践方向。

研究背景

酒精性肝病（ALD）已成为全球最常见的慢性肝病之一。更重要的是，依据世界卫生组织在2018年发布的《全球酒精与健康报告》，2016年因酒精导致的死亡占全球死亡总数的5.3%。近年来，越来越多的证据表明，肠道菌群失调与ALD的发生发展之间存在不可忽视且具有因果关系的关联。长期饮酒会损伤肠道屏障完整性并导致肠道微生物组成改变。由于肠道细菌对乙醇高度敏感，酒精摄入会显著影响肠道细菌群落。

肠道细菌、真菌和病毒之间的相互作用对维持肠道微生态平衡至关重要。多项研究显示，厚壁菌门中的专性厌氧菌在抑制变形菌门潜在致病菌的扩增以及限制共生真菌在肠道定植方面发挥重要作用。这些发现以及酒精诱导的细菌失调和真菌过度生长导致肝脏炎症和脂肪沉积的证据，提示酒精导致的厚壁菌门减少可能是ALD发病机制的主要因素之一。因此，专门促进厚壁菌门细菌生长的干预措施被寄予治疗ALD的期望。早期研究已证实，能够刺激乳酸杆菌和双歧杆菌生长的益生元低聚果糖可改善小鼠的酒精性肝损伤。作为可食用且具药用价值的真菌，茯苓（Wolfiporia cocos）的块体在传统中医中因其利尿、镇静、补益等功效被广泛使用。我们早期的研究表明，口服茯苓的水不溶性多糖（WIP）能够增加ob/ob小鼠厚壁菌门中产丁酸菌的丰度，这提示其可能对ALD具有潜在益处。WIP是一种（1→3）‑β‑D‑葡聚糖，平均分子量为4.486 × 10⁶ Da，已通过核磁共振（NMR）和SEC‑RI‑MALLS等方法鉴定。目前尚未有研究探讨茯苓对ALD的作用。

研究内容

在本研究中，我们以酒精性肝脂肪变性小鼠模型为实验对象，展示了WIP对酒精诱导的肝脂肪堆积和炎症的治疗效果，证实了WIP对酒精导致的肠道菌群失调的改善作用，并揭示了共生酵母Meyerozyma guilliermondii 与ALD的关联以及真菌诱导的PGE₂在ALD发展中的贡献。

研究结果

Figure 1. Oral treatment with WIP alleviates chronic ethanol feeding-induced hepatic injury and steatosis. (a) Experimental design. (b) The level of plasma alanine aminotransferase (ALT). (c) The level of plasma aspartate aminotransferase (AST). (d) The plasma levels of alkaline phosphatase (ALP). (e) The level of plasma lactate dehydrogenase (LDH). (f) Liver index. (g) The level of hepatic triglyceride (TG). (h) The level of hepatic total cholesterol (TC). (i) The expression of TNF-α in liver. (j) Representative picture of liver sections stained with oil-red. (k) Representative picture of liver sections with MCP-1 immunofluorescence staining. (b-h) N = 10 per group, (i) N = 5 per group, (j-k) N = 3 per group. Control: mice received isocaloric liquid diet instead of ethanol. Alcohol: mice fed with ethanol diet. WIP: mice fed with an ethanol diet supplemented with a water-insoluble polysaccharide from W. cocos. Data are presented as the mean ± standard error of the mean (SEM).

Figure 2. WIP treatment ameliorates the ethanol-induced gut dysbiosis. (a) OTU Venn diagram. (b) Weighted uniFrac-based principal coordinates analysis. (c) Shannon index. Bacterial taxonomic profiling of intestinal bacteria from different groups at the phylum (d) and family level (e). (f, g) Linear discriminant analysis (LDA) scores derived from LEfSe analysis. (h-k) Differentially abundant bacterial genera. The relative expression of occludin-1 (l) and ZO-1 (m) in colon. (n) The level of plasma lipopolysaccharide (LPS). (o) Total fungi in feces assessed by qPCR. The relative expression of ppar-γ (p) and nos2 (q) and colon TNF-α (r) and IL-Iβ (s). (a-k and n) N = 8 per group, (l-s) N = 5 per group. Control: mice received an isocaloric liquid diet instead of ethanol. Alcohol: mice fed with ethanol diet. WIP: mice fed with an ethanol diet supplemented with a water-insoluble polysaccharide from W. cocos. Data are presented as the mean ± standard error of the mean (SEM).

Figure 3. Identification of Meyerozyma guilliermondii as a casual fungus for AHS. (a) Fungi isolated from feces by aerobic culture-dependent approach. The number in the parentheses represented the strains obtained for each identified fungus. (b) Level of Meyerozyma guilliermondii (Mg) in fecal samples based on aerobic culture-dependent approach. (c) Abundance of Meyerozyma in Cecal contents based on ITS1 sequencing. (d) Experimental design. (e) The level of plasma alanine aminotransferase (ALT). (f) The level of plasma aspartate aminotransferase (AST). (g) The level of hepatic triglyceride (TG). (h) Representative picture of liver sections stained with oil-red. (i) The level of plasma triglyceride (TG). (j) The level of hepatic total cholesterol (TC). (k) The level of TNF-α in the liver. (l) The level of plasma β-glucan. (c) N = 4–5 per group, (e-g and i-l) N = 7–9 per group, (h) N = 3 per group. Ampho B: amphotericin B. Data are presented as the mean ± standard error of the mean (SEM).

Figure 4. Contribution of fungi-induced PGE2 to alcoholic hepatic steatosis. The level of PGE2 (a), the expression of EP2 (b), EP4 (c) and cxcl1 (d) in liver of ethanol-fed mice treated with WIP. The level of PGE2 (e), the expression of EP2 (f), EP4 (g) and cxcl1 (h) in liver of ethanol-fed mice treated amphotericin B (ampho B) or caspofungin. The level of PGE2 (i), the expression of EP2 (j), EP4 (k) and cxcl1 (l) in liver of the fungi-free mice treated with live M. guilliermondii (Mg). The level of ALT (m), hepatic TG (n), the expression of EP2 (o), EP4 (p) and cxcl1 (q) in liver after oral PGE2. (a and m-n) N = 9–10 per group, (b-d, f-h, j-l and o-q) N = 5 per group, (e and i) N = 7–9 per group. Control: mice received the isocaloric liquid diet instead of ethanol. Alcohol: mice fed with ethanol diet. WIP: mice fed with an ethanol diet supplemented with a water-insoluble polysaccharide from W. cocos. Ampho B: amphotericin B. Data are presented as the mean ± standard error of the mean (SEM).

研究结论

本研究表明，茯苓（Wolfporia cocos）来源的水不溶性多糖（WIP）可通过调节酒精性肝脂肪变性（AHS）小鼠的肠道微生物群，有效改善其肝脏炎性损伤与脂肪堆积。口服WIP能显著提高厚壁菌门与变形菌门的比值，增加毛螺菌科细菌的丰度（，并抑制乙醇诱导的真菌过度生长。WIP处理可激活过氧化物酶体增殖物激活受体-γ（PPAR-γ）信号通路，减轻结肠上皮细胞的炎症反应，促进肠道内低氧环境形成，从而抑制肠道真菌与变形菌门细菌的过度生长。此外，通过培养法和内转录间隔区（ITS）测序，我们发现酒精性肝脂肪变性小鼠粪便中共生真菌吉列蒙迪梅耶酵母菌（Meyerozyma guilliermondii）的数量大幅增加。将该酵母菌接种到无菌真菌小鼠体内，会加剧酒精性肝脂肪变性的病理特征。研究还发现，吉列蒙迪梅耶酵母菌可通过花生四烯酸的生物转化生成前列腺素E2（PGE2）。进一步研究证实，肠道真菌（吉列蒙迪梅耶酵母菌）诱导肝脏产生PGE2，是慢性酒精性肝脂肪变性的致病机制之一。本研究证实，调控肠道微生物群（细菌和真菌）可作为缓解酒精性肝病的一种有效替代策略。

https://doi.org/10.1080/19490976.2020.1830693

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