探討天麻水萃物於小鼠亞慢性溫和社交挫敗壓力模式中對於腸腦軸線之調節作用

Abstract

近年來，腸被喻為第二個腦，腸道菌相研究更逐漸受到重視，包括其於神經傳遞物質之調控 (如血清素生成、色胺酸代謝等)，且已有多篇研究指出腸腦軸線與神經相關疾病具有密切關聯性且被認為可直接影響情緒。憂鬱症是全球近年來備受關注的神經障礙疾病 (common mental disorder) 之一，根據世界衛生組織 (World Health Organization) 於 2017 年之統計，全球有高達 3 億的人罹患憂鬱症，且遍佈各年齡層，其中更有 2.6 億的患者合併焦慮症，顯見憂鬱症的預防及治療已不容忽視。近年來有多篇文獻指出傳統中草藥對憂鬱症之改善效果，其中天麻 (Gastrodia elata Blume) 屬於一種多年生草本植物，具有抗癲癇、抗腫瘤、抗氧化、抗老化、免疫調節、記憶改善、神經保護、神經可塑性、抗焦慮、抗憂鬱的功效。本實驗室過去多年著手於天麻水萃物對於憂鬱症相關之研究，透過不同的憂鬱症模式，如慢性不可預期溫和壓力、強迫游泳試驗與慢性社交挫敗模式證實天麻水萃物對於憂鬱症具有預防與減緩效果，但未進一步探討天麻在腸腦軸線中所扮演之角色。故本研究將以亞慢性溫和社交挫敗模式與廣效型抗生素的介入下，探討天麻水萃物 (water extract of Gastrodia elata Blume, WGE) 對於壓力誘導之類憂鬱與焦慮行為的影響，並進一步探討腸腦軸線之關係與其相關機制。結果顯示，經社交挫敗壓力之小鼠會產生社交迴避現象，且伴隨焦慮、活動能力與探索性降低。在生化數值方面，血清皮質固酮經壓力後顯著增加 (p < 0.0001)，腦部前額葉皮質 indoleamine 2,3-dioxygenase (IDO) 蛋白質表現量亦顯著增加 (p < 0.01)；大腸組織中犬尿胺酸 (kynurenine, KYN) 生成率顯著增加 (p < 0.05)、血清素 (serotonin, 5-HT) 生成率則顯著降低 (p < 0.005)，前述兩者轉換率顯著增加，推測經壓力刺激下，色胺酸轉而趨向生成 KYN，且其代謝途徑的異常與發炎反應具有正相關。而給予 WGE 則可改善小鼠之社交迴避現象與活動能力，有效降低血清壓力荷爾蒙皮質固酮濃度 (p < 0.05)，顯著減少大腸 KYN 生成率 (p < 0.01)。另外，在廣效性抗生素的介入後，發現小鼠會過度活動且伴隨類焦慮行為，其前額葉皮質 5-HT 生成率顯著增加 (p < 0.05)，5-HT 轉換率顯著減少 (p < 0.05)。而在給予 WGE 後，發現可顯著改善前額葉皮質因抗生素所導致的 5-HT 代謝異常 (p < 0.05)。此外，在腸道菌分析中，小鼠在 WGE 介入 14 天後發現腸道菌多樣性顯著增加 (p < 0.05)。而在壓力試驗後，壓力組菌相與控制組顯著不同 (p < 0.05)。以 Pearson’s correlation 分析得知 WGE 的介入可部分改善菌相組成，包括增加 Bacteriodales S24-7 group (F), Rumminococcaceae (F), Lachnospiraceae (F), Clostridiales vadinBB60 (F), Rumminococcaceae UCG-014 (G), Rumminococcaceae UCG-005 (G) 和 Lachnospiraceae NK4A136 (G) 之菌數，推測WGE可能藉由抑制發炎或增加腸道短鏈脂肪酸生成，進而減緩壓力所造成的行為和生化數值之異常。綜合上述結果，WGE 可藉由改善發炎反應、5-HT 和 KYN 之代謝途徑，達到抗憂鬱及抗焦慮之功效。

Gut has been known as the second brain and affects the brain via various pathways such as neurotransmitter modulation. While the gut-brain axis is assumed to play an important role within the gut microbiota and emotion. According to WHO, depression is a common mental disorder, the leading cause of disability in the whole world. Statistics as of 2017 showed that over 300 million people suffered from depression, not only the olders, but also the youngster. Among them, more than half also suffered from anxiety, occupied more than 264 million people. Studies found that tranditional Chinese herbal medicine has potential to be a good and effective complementary therapy. Gastrodia elata Blume, is prospected to have bioactive functions such as anticonvulsion, antitumor, antioxidant, antiaging, immune modulation, memory improvement, neuroprotection, neuroplasticity, antianxiety and antidepressant. Our labortary previously showed that water extract of Gastrodia elata Blume (WGE) exerts antidepression-like effect in forced swimming test, unpredictable chronic mild stress, and chronic social defeat stress model. However, the impact of WGE on gut in subchronic and mild social defeat stress (sCSDS) model has not been studied. Thus, I investigated the latent gut-brain linked mechanisms of WGE in relieving stress-induced depression-like behaviors which may result from the tryptophan (TRP) metabolism in mice subjected to sCSDS model. Results showed that sCSDS induced both social avoidance and depression-like behaviors such as anxiety, low locomotion and exploration. In the biochemical analysis showned significantly increased of serum corticosterone (CORT) (p < 0.0001), prefrontal cortex indoleamine 2,3-dioxygenase (IDO) protein expression (p < 0.01) and colon kynurenine (KYN) production rate (p < 0.05), while significantly decreased serotonin (5-HT) production rate (p < 0.005). While treatment with WGE, significantly reversed the depression-like phenotypes, such as social avoidance and stress index, serum CORT (p < 0.05) induced by sCSDS. Also, WGE significantly normalised the deviant metabolic of KYN in colon cause by defeat stress (p < 0.01). The administration of antibiotics elevated body weights and induced high locomotion and anxiety behavior of mice, while significantly increase 5-HT production rate (p < 0.05) and decrease 5-HT turnover rate (p < 0.05) in prefrontal cortex. In addition of WGE administration, significantly normalised the deviant metabolic of 5-HT cause by antibiotics (p < 0.05). In general, high levels of 5-HT in brain induced by sCSDS with antibiotics may linked to anxiety behavior, while WGE treatment could reverse it. By analyzing the fecal microbiota, I found that 14 days of WGE treatment increased the fecal diversity (p < 0.05). After the 10-day social defeat, stress groups had a significantly different microbiome status compared with control (p < 0.05), while WGE ameliorated part of them, such as Bacteriodales S24-7 group (F), Rumminococcaceae (F), Lachnospiraceae (F), Clostridiales vadinBB60 (F), Rumminococcaceae UCG-014 (G), Rumminococcaceae UCG-005 (G) and Lachnospiraceae NK4A136 (G), implying that WGE may inhibit the inflammation and promote short chain fatty acids production in gut. Thus, WGE possessed antidepressive and anxiolytic effects via modulating the inflammation, serotonin and kynurenine pathway.