桂郁金多糖对HO致人脐静脉内皮细胞损伤的保护作用及机制

俞献文; 唐丽清; 覃琴; 冯柏林; 杨秀芬

doi:10.13422/j.cnki.syfjx.20182333

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桂郁金多糖对HO致人脐静脉内皮细胞损伤的保护作用及机制

药理 | 更新时间：2024-09-23

- 桂郁金多糖对HO致人脐静脉内皮细胞损伤的保护作用及机制
- Protective Effect and Mechanism of Polysaccharides from Curcumae Kwangsiensis Radix on HUVECs Injuried by HO
- 中国实验方剂学杂志 2018年24卷第23期页码：146-152
- 作者机构：
- 作者简介：
- 基金信息：
  
  国家自然科学基金项目（81260652）;广西自然科学基金重点项目（2015GXNSFDA139023）;“八桂学者”工程专项经费和硕士研究生科研创新项目（YJSZ201702，YCSW2018168）
- DOI：10.13422/j.cnki.syfjx.20182333
  中图分类号： R285
- 纸质出版日期：2018
- 稿件说明：
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俞献文, 唐丽清, 覃琴, 等. 桂郁金多糖对HO致人脐静脉内皮细胞损伤的保护作用及机制[J]. 中国实验方剂学杂志, 2018,24(23):146-152.

YU Xian-wen, TANG Li-qing, QIN Qin, et al. Protective Effect and Mechanism of Polysaccharides from Curcumae Kwangsiensis Radix on HUVECs Injuried by HO[J]. Chinese journal of experimental traditional medical formulae, 2018, 24(23): 146-152.
俞献文, 唐丽清, 覃琴, 等. 桂郁金多糖对HO致人脐静脉内皮细胞损伤的保护作用及机制[J]. 中国实验方剂学杂志, 2018,24(23):146-152. DOI： 10.13422/j.cnki.syfjx.20182333.

YU Xian-wen, TANG Li-qing, QIN Qin, et al. Protective Effect and Mechanism of Polysaccharides from Curcumae Kwangsiensis Radix on HUVECs Injuried by HO[J]. Chinese journal of experimental traditional medical formulae, 2018, 24(23): 146-152. DOI： 10.13422/j.cnki.syfjx.20182333.

摘要

目的：通过体外实验研究不同质量浓度桂郁金多糖对过氧化氢（H2O2）致人脐静脉内皮细胞损伤的的保护作用及机制。方法：对人脐静脉内皮细胞（HUVECs）进行体外培养，用不同浓度H2O2损伤HUVECs，采用细胞增殖/毒性检测试剂盒（CCK-8），确定H2O2最佳损伤浓度为500 μmol·L-1。然后给予不同浓度的桂郁金多糖对损伤的HUVECs进行保护，用CCK-8，确定62.5，125，250 mg·L-1作为桂郁金多糖的低、中、高质量浓度组。采用CCK-8法检测桂郁金多糖对H2O2损伤HUVECs细胞活力的影响；采用试剂盒测定乳酸脱氢酶（LDH），超氧化物歧化酶（SOD），谷胱甘肽过氧化物酶（GSH-Px），丙二醛（MDA）；通过酶联免疫吸附测定（ELISA）检测细胞中肿瘤坏死因子-α（TNF-α），白细胞介素-6（IL-6）的含量表达。采用Hoechst 33258荧光染色法对细胞凋亡情况进行观察；并采用实时荧光定量聚合酶链式反应（Real-time PCR）测定细胞中B淋巴细胞瘤-2（Bcl-2），Bcl-2相关X蛋白（Bax），半胱氨酸蛋白酶-3（Caspase-3）mRNA的表达情况。结果： CCK-8检测结果表明，给药时间24 h后，桂郁金多糖在15.625~500 mg·L-1，能够促进细胞增殖，而浓度增大后，桂郁金多糖则对细胞有一定的抑制作用。与模型组比较，桂郁金多糖能够使LDH，MDA，IL-6，TNF-α水平明显减少（P<0.05，P<0.01），使SOD和GSH-Px的含量明显增加（P<0.05，P<0.01）。Hoechst 33258荧光染色结果表明，与空白组比较，模型组HUVECs细胞核呈亮蓝色荧光，细胞质内可见浓染致密的颗粒块状荧光，细胞密度降低；与模型组比较，桂郁金多糖给药组的蓝色荧光强度降低，细胞凋亡典型形态减少。Real-time PCR实验结果显示，与空白组比较，模型组减少Bcl-2 mRNA的表达，增加Bax，Caspase-3 mRNA的表达（P<0.05，P<0.01）；与模型组比较，桂郁金多糖给药组能够增加Bcl-2 mRNA的表达（P<0.05，P<0.01），减少Bax，Caspase-3 mRNA的表达（P<0.05，P<0.01）。结论：桂郁金多糖通过抗氧化和抑制细胞凋亡等途径对H2O2损伤的人脐静脉内皮细胞进行保护。

Abstract

Objective:The in vitro study of different doses of polysaccharides from Curcumae Kwangsiensis Radix on the protective effect and mechanism of H2O2-induced injury of HUVECs. Method:Protective effects of polysaccharides from Curcumae Kwangsiensis Radix on H2O2-induced injury of human umbilical vein endothelial cells (HUVECs) and its mechanism were studied. HUVECs were cultured in vitro

HUVECs were injured with different concentrations of H2O2

and the optimal concentration of H2O2 was 500 μmol·L-1 by cell counting kit-8(CCK-8) method. Then

different concentrations of polysaccharides from Curcumae Kwangsiensis Radix were used to protect the damaged HUVECs. The CCK-8 method was used to determine 62.5

125

250 mg·L-1 as the low

medium and high-dose groups. Effect of CCK-8 method was used to detect the polysaccharides from Curcumae Kwangsiensis Radix H2O2 on the injury of HUVECs cell activity. Lactic dehydrogenase (LDH)

superoxide dismutase (SOD)

glutathione-peroxidase (GSH-Px)

malondialdehyde (MDA) were determined by kit. The levels of tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) in cells were detected by enzyme-linked immunosorbent assay (ELISA). Hoechst 33258 fluorescence staining was used to observe the cell apoptosis. The protein expressions of B-cell lymphoma-2 (Bcl-2)

Bcl-2-associated X (Bax) and cysteine aspartate-specific protease-3 (Caspase-3) genes were detected by real-time fluorescence quantitative polymerase chain reaction (Real-time PCR). Result:The result of CCK-8 assay showed after administration for 24 h

polysaccharides from Curcumae Kwangsiensis Radix within the concentration range of 15.625-500 mg·L-1 can promote cell proliferation; as the concentration increased

polysaccharides from Curcumae Kwangsiensis Radix had a certain inhibitory effect on cells. Compared with the model group

polysaccharides from Curcumae Kwangsiensis Radix could significantly reduce the content of LDH

MDA

IL-6

TNF-α(P<0.05

P<0.01)

while significantly increased the contents of SOD and GSH-Px(P<0.05

P<0.01). Hoechst 33258 fluorescence staining results showed that compared with the normal group

the nucleus of HUVECs showed bright blue fluorescence in the model group

dense and dense granular lumps of fluorescence were observed in the cytoplasm

and the cell density was decreased. Compared with the model group

the blue fluorescence intensity decreased

and the typical morphological apoptosis decreased. The Real\|time PCR results showed that compared with the normal group

the model group decreased the expression of Bcl-2 and increased the expression of Bax and Caspase-3(P<0.05

P<0.01). Compared with the model group

polysaccharides from Curcumae Kwangsiensis Radix group could increase the expression of Bcl-2(P<0.05

P<0.01)

while reduce Bax

Caspase-3 expression(P<0.05

P<0.01). Conclusion:polysaccharides from Curcumae Kwangsiensis Radix have a protective effect on human umbilical vein endothelial cells injured by H2O2 by resisting oxidation and inhibiting cell apoptosis.

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