Protection Effect of Essential Oil from  Rhizome on ox-LDL-induced HAECs Injury via NOS Signal

XU Yi-ni; YANG Hong; LI Chen; LINGHU Ke-gang; WEN Bo; SHEN Xiang-chun; ZHANG Yan-yan

doi:10.13422/j.cnki.syfjx.2017150143

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Protection Effect of Essential Oil from Rhizome on ox-LDL-induced HAECs Injury via NOS Signal

更新时间：2024-01-04

- Protection Effect of Essential Oil from Rhizome on ox-LDL-induced HAECs Injury via NOS Signal
- Chinese Journal of Experimental Traditional Medical Formulae Vol. 23, Issue 15, Pages: 143-147(2017)
- 作者机构：
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- DOI：10.13422/j.cnki.syfjx.2017150143
  CLC： R285
- Published：2017
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XU Yi-ni, YANG Hong, LI Chen, et al. Protection Effect of Essential Oil from Rhizome on ox-LDL-induced HAECs Injury via NOS Signal[J]. Chinese journal of experimental traditional medical formulae, 2017, 23(15): 143-147.
DOI：

XU Yi-ni, YANG Hong, LI Chen, et al. Protection Effect of Essential Oil from Rhizome on ox-LDL-induced HAECs Injury via NOS Signal[J]. Chinese journal of experimental traditional medical formulae, 2017, 23(15): 143-147. DOI： 10.13422/j.cnki.syfjx.2017150143.

摘要

目的：研究艳山姜挥发油（EOFAZ）对氧化低密度脂蛋白（ox-LDL）诱导的人主动脉内皮细胞（HAECs）损伤的保护作用。方法：体外传代培养HAECs，EOFAZ保护作用研究分为7组，分别为空白组（无血清ECM），模型组（200 mg·L-1 ox-LDL），EOFAZ高质量浓度组（200 mg·L-1 ox-LDL+100 μg·L-1 EOFAZ），EOFAZ低质量浓度组（200 mg·L-1 ox-LDL+10 μg·L-1 EOFAZ），阿司匹林组（Asp，200 mg·L-1 ox-LDL+2.5×10-4 mol·L-1 Asp），卡维地洛组（Car，200 mg·L-1 ox-LDL+1×10-6 mol·L-1 Car），阿托伐他汀钙组（Atorv，200 mg·L-1 ox-LDL+1×10-6 mol·L-1 Atorv）；一氧化氮合酶（NOS）信号研究分为5组，分别为空白组（无血清ECM），模型组（200 mg·L-1 ox-LDL），EOFAZ组（200 mg·L-1 ox-LDL+100 μg·L-1 EOFAZ），NOS抑制剂组（200 mg·L-1 ox-LDL+100 μmol·L-1 L-NAME或300 μmol·L-1 L-NMMA），EOFAZ加NOS抑制剂组（200 mg·L-1 ox-LDL+100 μg·L-1 EOFAZ+100 μmol·L-1 L-NAME或300 μmol·L-1 L-NMMA）。噻唑蓝（MTT）法分析细胞存活率，酶标仪法及Griess试剂法分别检测培养上清液中乳酸脱氢酶（LDH）活性和一氧化氮（NO）含量。化学法检测内皮型一氧化氮合酶（eNOS）和诱导型一氧化氮合酶（iNOS）活性。实时荧光定量聚合酶链式反应（Real-time PCR）检测iNOS及eNOS mRNA表达水平。结果：与模型组比较，EOFAZ明显提高ox-LDL诱导损伤的HAECs的细胞存活率，抑制LDH外漏及iNOS活力（P<0.05，P<0.01），促进NO及eNOS的产生（P<0.05， P<0.01）。Real-time PCR分析表明，EOFAZ以及相关抑制剂显著下调iNOS mRNA表达水平（P<0.05，P<0.01），上调eNOS mRNA表达水平（P<0.05）。结论：EOFAZ对ox-LDL诱导损伤的HAECs具有保护作用，其作用机制与调控eNOS和iNOS的表达水平有关。

Abstract

Objective: To investigate the protective effects of essential oil from Alpinia zerumbet rhizome(EOFAZ) on oxidized low density lipoprotein (ox-LDL)-induced human aortic endothelial cells (HAECs) injury. Method: HAECs were subcultured in vitro

and the experiment on EOFAZ protective effect was randomly divided into 7 groups as following: blank control group (serum free ECM)

model group (ox-LDL

200 mg·L-1 ox-LDL)

EOFAZ high dose group (200 mg·L-1 ox-LDL+100 μg·L-1 EOFAZ)

EOFAZ low dose group (200 mg·L-1 ox-LDL+10 μg·L-1 EOFAZ)

Aspirin group (200 mg·L-1 ox-LDL+2.5 × 10-4 mol·L-1 Aspirin)

Carvedilol group (200 mg·L-1 ox-LDL+1×10-6mol·L-1 Carvedilol)

and Atorvastatin Calcium group (200 mg·L-1 ox-LDL+1×10-6mol·L-1 Atorvastatin Calcium). Experiment on NOS signals was divided into 5 groups as following: blank control group (serum free ECM)

model group (ox-LDL

200 mg·L-1 ox-LDL)

EOFAZ group (200 mg·L-1 ox-LDL+100 μg·L-1 EOFAZ)

NOS inhibitor group (200 mg·L-1 ox-LDL+100 μmol·L-1 L-NAMEor 300 μmol·L-1 L-NMMA)

and EOFAZ+NOS inhibitor group(200 mg·L-1 ox-LDL+100 μg·L-1 EOFAZ+100 μmol·L-1 L-NAME or 300 μmol·L-1 L-NMMA). 3-(4

5-Dimethyl-2-thiazolyl)-2

5-diphenyl-2-H-tetrazolium bromide(MTT) method was used to analyze the cell survival ratio; lactate dehydrogenase(LDH) activity and nitric oxide(NO) content in culture supernatant were determined by enzyme labelling method and Griess kit method. Endothelial nitric oxide synthase(eNOS) and inducible nitric oxide synthase(iNOS) activities were determined by chemical methods. Quantitative Real-time polymerase chain reaction(Real-time PCR) was used to detect the mRNA expression of iNOS and eNOS. Result: MTT results showed that EOFAZ could significantly improve the survival rate of ox-LDL-induced injury HAECs

inhibit the increase of LDH and iNOS activity as compared with the model group (P<0.05

P<0.01)

and simultaneously increase the release of NO and eNOS in medium(P<0.05

P<0.01). Real-time PCR analysis indicated that EOFAZ and related inhibitors could down-regulate iNOS mRNA expression (P<0.05

P<0.01) and up-regulate eNOS mRNA expression (P<0.05). Conclusion: EOFAZ could protect against HAECs injury induced by ox-LDL

and this mechanism was associated with regulating eNOS and iNOS expression.

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