目的 研究膽南星Arisaema Cum Bile多糖組分拮抗高熱驚厥模型大鼠神經(jīng)炎癥作用的可能機(jī)制。方法 采用水提醇沉法制備膽南星多糖，并通過色譜、光譜等方法對(duì)其進(jìn)行結(jié)構(gòu)表征分析；通過ip脂多糖聯(lián)合熱水浴建立高熱驚厥模型，隨機(jī)分為對(duì)照組、模型組、丙戊酸鈉（200 mg/kg）組和膽南星高、低劑量（200、100 mg/kg）組，每組10只。記錄每組大鼠驚厥潛伏期，驚厥持續(xù)時(shí)間和驚厥級(jí)別，并于末次造模2 h后測量大鼠肛溫；蘇木素-伊紅（hematoxylin eosin，HE）染色評(píng)估海馬組織神經(jīng)元病理變化；ELISA法檢測血清中腫瘤壞死因子-α（tumor necrosis factor-α，TNF-α）、白細(xì)胞介素-1β（interleukin-1β，IL-1β）、IL-6及海馬組織中γ-氨基丁酸（γ-aminobutyricacid，GABA）、谷氨酸（glutamic acid，Glu）、NOD樣受體熱蛋白結(jié)構(gòu)域3（NOD like receptor family pyrin domain containing 3，NLRP3）和高遷移率族蛋白B1（high mobility group box 1 protein，HMGB1）水平；采用超高效液相色譜-四級(jí)桿飛行時(shí)間質(zhì)譜（ultra performance liquid chromatography tandem quadrupole time of flight mass spectrometry，UPLC-Q-TOF-MS/MS）對(duì)大鼠的血清進(jìn)行代謝組學(xué)分析。結(jié)果 分離制備得到的膽南星多糖中總糖、蛋白質(zhì)和糖醛酸質(zhì)量分?jǐn)?shù)分別為79.4%、2.96%和2.35%；氣相色譜-質(zhì)譜聯(lián)用技術(shù)（gas chromatography-mass spectrometry，GC-MS）測定膽南星多糖主要由阿拉伯糖、木糖、甘露糖、葡萄糖和半乳糖組成，其物質(zhì)的量比分別為0.030∶0.260∶0.115∶0.741∶0.088；傅里葉變換紅外光譜儀（Fouriertransform infrared spectrometer，F(xiàn)T-IR）光譜表明多糖含有吡喃環(huán)；與模型組比較，膽南星多糖高劑量組大鼠肛溫降低（P＜0.01）且膽南星多糖高、低劑量組均可顯著延長驚厥潛伏期（P＜0.01），縮短驚厥持續(xù)時(shí)間（P＜0.01）；對(duì)海馬神經(jīng)元細(xì)胞具有保護(hù)作用，升高海馬組織GABA含量，同時(shí)降低血清中TNF-α、IL-1β和IL-6與海馬組織中Glu、NLRP3及HMGB1含量（P＜0.05、0.01）；代謝組學(xué)結(jié)果表明，膽南星多糖可顯著回調(diào)16種生物標(biāo)志物的表達(dá)，共涉及花生四烯酸代謝、氨基酸代謝及嘧啶代謝等6條代謝通路徑。結(jié)論 膽南星多糖組分對(duì)高熱驚厥大鼠表現(xiàn)出顯著的拮抗作用，其作用機(jī)制可能與解熱、保護(hù)海馬神經(jīng)元、抑制腦內(nèi)炎癥發(fā)生及調(diào)節(jié)代謝紊亂有關(guān)。

Objective To investigate the possible mechanisms by which polysaccharides of Dannanxing (Arisaema Cum Bile) inhibit neuroinflammation in febrile seizure rat model. Methods The polysaccharides of Arisaema Cum Bile were prepared by a combination of water extraction and alcohol precipitation, and its primary structure was characterized by chromatographic and spectral methods. Febrile seizure model was established by intraperitoneal injection of lipopolysaccharide combined with hot water bath. The rats were randomly divided into five groups, including control group, model group, sodium valproate group (200 mg/kg), Arisaema cum bile polysaccharides high- and low-dose (200, 100 mg/kg) groups, with ten rats in each group. The convulsive latency, duration and grade of convulsion were recorded in each group, and the rectal temperature of rats was monitored 2 h after the last modeling. Hematoxylin eosin (HE) staining was used to evaluate the pathological changes of hippocampal neurons. The levels of tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), IL-6 in serum and γ-aminobutyricacid (GABA), glutamic acid (Glu), NOD like receptor family pyrin domain containing 3 (NLRP3) and high mobility group box 1 protein (HMGB1) in hippocampus tissues were detected by ELISA. The metabolomics analysis of rat serum based on ultra performance liquid chromatography tandem quadrupole time of flight mass spectrometry (UPLC-Q-TOF-MS) was performed. Results The total sugar, protein and glucuronic acid contents of the Arisaema Cum Bile polysaccharides obtained from the isolation preparation were 79.4%, 2.96% and 2.35%, respectively. The results of monosaccharide composition based on gas chromatography-mass spectrometry (GC-MS) exhibited that the polysaccharides of Arisaema Cum Bile were mainly composed of arabinose, xylose, mannose, glucose and galactose with molar mass ratios of 0.030∶0.260∶0.115∶0.741∶0.088, respectively. Fouriertransform infrared spectrometer (FT-IR) spectroscopy indicated that the polysaccharides contained pyran ring. Compared with the model group, the rectal temperature of rats in the high-dose group of Arisaema Cum Bile polysaccharide was decreased (P < 0.01), and both high and low dose groups of Arisaema Cum Bile polysaccharide significantly prolonged the convulsion latency (P < 0.01) and shortened the duration of convulsions (P < 0.01). The high-dose Arisaema cum bile polysaccharides supplementation showed a protective effect on hippocampal neuronal cells, elevated GABA content in hippocampal tissues, and simultaneously reduced the content of TNF-α, IL-1β, IL-6 in serum, Glu, NLRP3 and HMGB1 in hippocampal tissues (P < 0.05, 0.01). The metabolomics results showed that Arisaema Cum Bile polysaccharides could significantly reverse the expression of 16 biomarkers, which involved six metabolic pathways including arachidonic acid metabolism, amino acid metabolism and pyrimidine metabolism. Conclusion The polysaccharide fraction of Arisaema Cum Bile showed significant antagonistic effects on febrile seizure rats, and its underlying mechanism may be closely related to lowering body temperature, protecting hippocampal neurons, inhibiting neuroinflammation in brain and regulating metabolic disorders.

R285.5

2022年全國名老中醫(yī)藥專家傳承工作室建設(shè)項(xiàng)目（國中醫(yī)藥人教函[2022]75號(hào)）;第七批全國老中醫(yī)藥專家學(xué)術(shù)經(jīng)驗(yàn)繼承工作項(xiàng)目（國中醫(yī)藥人教函[2022]76號(hào)）;黑龍江省“頭雁”團(tuán)隊(duì)支持項(xiàng)目（黑龍江省頭雁行動(dòng)領(lǐng)導(dǎo)小組文件[2019]5號(hào)）