目的 明確當(dāng)歸通過調(diào)節(jié)鞘脂代謝發(fā)揮對皮質(zhì)酮（CORT）損傷的PC12細(xì)胞的保護作用及機制。方法 制備當(dāng)歸水提物，LC-MS法進(jìn)行成分鑒定；當(dāng)歸水提物含藥血清（ASWEMS）的制備：將12只SPF級雄性SD大鼠隨機分為空白血清（給予0.9%氯化鈉溶液）組和ASWEMS（生藥量7.5 g·kg^-1）組，連續(xù)ig給藥6 d，采血管腹主動脈采血，分離血清后同組混合，56℃水浴加熱30 min后經(jīng)0.22 μm微孔濾膜濾過除菌。利用CORT誘導(dǎo)PC12細(xì)胞構(gòu)建損傷模型，將PC12細(xì)胞分為：對照（5%空白血清）組、模型組（400 μmol·L^-1 CORT＋5%空白血清）、ASWEMS組（400 μmol·L^-1 CORT＋5% ASWEMS）、神經(jīng)酰胺合成酶抑制劑伏馬菌素（FB1，400 μmol·L^-1 CORT＋5%空白血清＋10 μmol·L^-1 FB1）組，處理24 h。通過CCK-8法檢測細(xì)胞存活率；采用Annexin FITC/PI流式細(xì)胞術(shù)檢測細(xì)胞凋亡率；基于超高效液相色譜-四極桿-飛行時間質(zhì)譜（UPLC-Q-TOF-MS）脂質(zhì)組學(xué)技術(shù)檢測PC12細(xì)胞脂質(zhì)代謝輪廓變化及鞘脂類代謝物相對含量；采用ELISA檢測PC12細(xì)胞神經(jīng)酰胺（Cer）含量。結(jié)果 LC-MS法鑒定出當(dāng)歸水提物17種化學(xué)成分。與模型組比較，ASWEMS和FB1均可顯著回調(diào)CORT損傷PC12細(xì)胞所引起的細(xì)胞存活率下降（P＜0.001），細(xì)胞凋亡率增加（P＜0.001）。鞘脂類代謝物是引起對照組與模型組差異貢獻(xiàn)較大的代謝物，與對照組比較，CORT干預(yù)24 h后，PC12細(xì)胞中的二氫鞘氨醇（Dhsph）類、植物鞘氨醇（PhytoSph）類代謝物相對含量顯著降低（P＜0.001），鞘磷脂（SM）類代謝物相對含量顯著升高（P＜0.001），Cer相對含量及絕對含量均顯著升高（P＜0.01）；與模型組比較，給予ASWEMS、FB1干預(yù)后各代謝物含量具有不同程度的回調(diào)（P＜0.01、0.001）。結(jié)論 ASWEMS對CORT誘導(dǎo)的PC12細(xì)胞損傷具有保護作用，其機制與調(diào)節(jié)鞘脂代謝通路、抑制細(xì)胞凋亡有關(guān)。

Objective To clarify the protective effect of Angelica sinensis water extract medicated serum (ASWEMS) on PC12 cells damaged by corticosterone (CORT) through the regulation of sphingolipid metabolism. Methods Preparation of Angelica sinensis water extract and identification of its components by LC-MS method. Preparation of Angelica sinensis water extract medicated serum (ASWEMS): Twelve healthy male SD rats were randomly divided into blank serum (received 0.9% sodium chloride solution) group and ASWEMS (crude drug dose of 7.5 g·kg^-1) group, acclimated for one week, and then intragastrically administered ASWEMS for six consecutive days. Blood was collected from the abdominal aorta of the rats, and the serum was separated and mixed within the same group. The serum was then heated at 56 ℃ for 30 min and filtered through a 0.22 μm microfiltration membrane to remove bacteria. A cell injury model was constructed by treating PC12 cells with CORT, with the cells divided into control (5% control serum), model (400 μmol·L^-1 CORT＋5% control serum), ASWEMS (400 μmol·L^-1 CORT＋5% ASWEMS) and fumonisin (FB1, 400 μmol·L^-1 CORT＋5% control serum ＋10 μmol·L·L^-1 FB1) groups, which subjected to the treatment for 24 h. The cell survival rate was detected by CCK-8 method. Annexin FITC/PI flow cytometry was used to detect apoptosis rate. Changes in lipid metabolism profile and the relative content of sphingolipid metabolites in PC12 cells were determined by ultra-high performance liquid chromatography with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS). The contents of Ceramide (Cer) in PC12 cells was detected by ELISA. Results LC-MS method identified 17 chemical components in Angelica sinensis water extract. Compared with the model group, ASWEMS and FB1 could significantly reverse the decreased cell viability and increased apoptosis rate caused by CORT injury in PC12 cells (P <0.001). Sphingolipids metabolites were the metabolites that contributed more to the difference between the control group and the model group. Compared with the control group, the relative content of Dhsph-type and PhytoSph-type metabolites in PC12 cells was significantly lower after CORT intervention for 24 h (P <0.001), while the relative content of SM-type metabolites was significantly higher (P <0.001), and the relative content and absolute content of Cer were both significantly higher (P <0.01). Compared with the model group, the contents of all metabolites were significantly adjusted after ASWEMS or FB1 intervention (P <0.01, 0.001). Conclusion ASWEMS has a protective effect against CORT-induced PC12 cell injury through regulating sphingolipid metabolism pathway and inhibiting apoptosis.

R285.5

國家自然科學(xué)基金項目（82004502）;山西省應(yīng)用基礎(chǔ)研究計劃項目（202403021211187）;經(jīng)方扶陽山西省重點實驗室項目（CP‐SY202304）;山西省重點研發(fā)項目（202102130501010）