Protective Effect of Ginsenosides Rg<sub>1</sub> and Rb<sub>1</sub> Against Intestinal Epithelial Barrier Injury Induced by Lipopolysaccharide <italic style="font-style: italic">in Vitro</italic>

Tian CHEN; Bo-ye LI; Bo-yang YU; Jin-ning YANG; Qin HU; Ying CHEN

doi:10.13422/j.cnki.syfjx.20220705

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Protective Effect of Ginsenosides Rg₁ and Rb₁ Against Intestinal Epithelial Barrier Injury Induced by Lipopolysaccharide in Vitro

更新时间：2022-03-07

- Protective Effect of Ginsenosides Rg₁ and Rb₁ Against Intestinal Epithelial Barrier Injury Induced by Lipopolysaccharide in Vitro
- Chinese Journal of Experimental Traditional Medical Formulae Vol. 28, Issue 7, Pages: 64-72(2022)
- 作者机构：
  
  1.北京工业大学环境与生命学部，北京 100124
  2.中国中医科学院中药研究所，北京 100700
- 作者简介：
- 基金信息：
- DOI：10.13422/j.cnki.syfjx.20220705
  CLC： R2-0;R22;R285.5;R33
- Received：12 November 2021，
  
  Published Online：26 January 2022，
  
  Published：05 April 2022
- 稿件说明：
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陈天,李博野,于渤洋等.人参皂苷Rg₁、Rb₁对脂多糖体外诱导肠上皮屏障损伤的保护作用[J].中国实验方剂学杂志,2022,28(07):64-72.

CHEN Tian,LI Bo-ye,YU Bo-yang,et al.Protective Effect of Ginsenosides Rg₁ and Rb₁ Against Intestinal Epithelial Barrier Injury Induced by Lipopolysaccharide in Vitro[J].Chinese Journal of Experimental Traditional Medical Formulae,2022,28(07):64-72.
陈天,李博野,于渤洋等.人参皂苷Rg₁、Rb₁对脂多糖体外诱导肠上皮屏障损伤的保护作用[J].中国实验方剂学杂志,2022,28(07):64-72. DOI： 10.13422/j.cnki.syfjx.20220705.

CHEN Tian,LI Bo-ye,YU Bo-yang,et al.Protective Effect of Ginsenosides Rg₁ and Rb₁ Against Intestinal Epithelial Barrier Injury Induced by Lipopolysaccharide in Vitro[J].Chinese Journal of Experimental Traditional Medical Formulae,2022,28(07):64-72. DOI： 10.13422/j.cnki.syfjx.20220705.

摘要

目的

基于人髓系白血病单核细胞（THP-1）与肠上皮细胞（Caco-2）共培养体系，研究人参皂苷Rg

和人参皂苷Rb

对脂多糖（LPS）诱导的THP-1细胞炎症因子释放的影响，及其对THP-1细胞活化致Caco-2细胞炎性损伤的保护作用。

方法

首先制备THP-1与Caco-2细胞共培养微流控芯片，实验分为空白组、LPS组和给药组。空白组细胞正常培养；LPS组在上层Caco-2细胞形成单层屏障后，在下层THP-1细胞中加入LPS（1 mg·L

-1

）；给药组在LPS组的基础上在THP-1细胞中分别加入33 mg·L

-1

的人参皂苷Rg

和人参皂苷Rb

。THP-1细胞与Caco-2细胞共培养24 h后采用异硫氰酸荧光素-葡聚糖（FITC-Dextran）示踪法检测下层芯片通道中的FITC-Dextran荧光值。THP-1细胞实验分为空白组、LPS组、给药组。空白组THP-1细胞正常培养；LPS组在THP-1细胞中加入LPS（1 mg·L

-1

）；给药组在LPS组的基础上分别加入相应剂量的人参皂苷Rg

和人参皂苷Rb

（11、33、100 mg·L

-1

）。细胞培养24 h后细胞增殖与活性检测（CCK-8）检测THP-1细胞活性，实时荧光定量聚合酶链式反应（Real-time PCR）检测THP-1细胞炎性细胞因子白细胞介素-6（IL-6）、白细胞介素-1

（IL-1

）、肿瘤坏死因子-

（TNF-

）表达。Caco-2细胞实验分为空白组、LPS组、给药组。空白组Caco-2细胞正常培养；其他组将第二部分THP-1细胞实验中对应组的细胞上清置换于Caco-2细胞中，继续培养24 h后CCK-8检测Caco-2细胞活性，Real-time PCR检测Caco-2细胞炎性细胞因子IL-6、IL-8、TNF-

及紧密连接蛋白封闭蛋白（Occludin）表达，蛋白免疫印迹法（Western blot）检测Caco-2细胞紧密连接蛋白Occludin的表达。

结果

在THP-1与Caco-2细胞共培养体系中，与LPS组比较，人参皂苷Rg

和Rb

均能有效保护LPS诱导肠上皮屏障通透性升高（

0.01）。Rg1和Rb1拮抗LPS诱导的THP-1细胞IL-6、IL-1

、TNF-

炎性细胞因子表达升高（

0.05）。经Rg

和Rb

处理的THP-1细胞上清与Caco-2细胞共培养后，与LPS组比较，显著降低Caco-2细胞IL-6、IL-8、TNF-

炎性细胞因子表达（

0.01），上调紧密连接蛋白Occludin表达。

结论

在THP-1与Caco-2细胞共培养体外模拟肠道上皮屏障功能模型中，人参皂苷Rg

和Rb

通过调节THP-1细胞释放炎性细胞因子，进而调控Caco-2细胞的炎性反应和细胞屏障完整性，在LPS诱导的体外肠上皮屏障损伤中发挥保护作用。

Abstract

Objective

To investigate the effects of ginsenoside Rg

and ginsenoside Rb

on the release of inflammatory factors of human myeloid leukemia monocytes （THP-1） induced by lipopolysaccharide （LPS） and their protective effects on the inflammatory injury of intestinal epithelial cells （Caco-2） induced by THP-1 cell activation based on the co-culture system of THP-1 and Caco-2.

Method

Firstly，the microfluidic chip of co-culture of THP-1 and Caco-2 cells was prepared. In the experiment， a blank group， an LPS group， and drug intervention groups were set up.The cells in the blank group were cultured conventionally. In the LPS group，LPS （1 mg·L

-1

） was added to the lower THP-1 cells after the upper Caco-2 cells formed a monolayer barrier. On the basis of the LPS group， 33 mg·L

-1

ginsenoside Rg

and 33 mg·L

-1

ginsenoside Rb

were added to THP-1 cells respectively. After the co-culture of THP-1 cells and Caco-2 cells for 24 hours， the fluorescein isothiocyanate （FITC）-Dextran fluorescence value in the lower chip channel was detected by FITC-Dextran tracer method. A blank group， an LPS group，and drug intervention groups were set up in the THP-1 cell experiment. THP-1 cells in the blank group were cultured conventionally. In the LPS group， LPS （1 mg·L

-1

） was added to THP-1 cells.Ginsenoside Rg

and ginsenoside Rb

of the corresponding doses （11，33，100 mg·L

-1

） were added to the drug intervention groups respectively on the basis of the LSP group. After 24 hours of cell culture， the activity of THP-1 cells was detected by cell counting kit-8 （CCK-8）. Real-time quantitative polymerase chain reaction （Real-time PCR） was used to detect the expression of inflammatory cytokines such as interleukin-6 （IL-6）， interleukin-1

（IL-1

）， and tumor necrosis factor

（TNF）-

of THP-1 cells. A blank group， an LPS group， and drug intervention groups were set up in the Caco-2 cell experiment. Caco-2 cells in the blank group were cultured conventionally， and in other groups， the corresponding cell supernatant in the second part of the THP-1 cell experiment was employed in Caco-2 cells. After 24 hours of cell culture，the activity of Caco-2 cells was detected by CCK-8. Real-time PCR was used to detect the expression of IL-6，interleukin-8 （IL-8）， TNF

-α

， and Occludin

of Caco-2 cells. The expression of tight junction protein Occludin in Caco-2 cells was detected by Western blot.

Result

Both ginsenoside Rg

and ginsenoside Rb

could effectively protect LPS-induced intestinal epithelial barrier permeability in the co-culture system of THP-1 and Caco-2 cells （

0.01）. Ginsenosides Rg

and Rb

antagonized LPS-induced increased expression of IL-6，IL-1

， and TNF

-α

in THP-1 cells （

0.05）. When the supernatant of THP-1 cells treated with ginsenosides Rg

and Rb

was co-cultured with Caco-2 cells， the expression of IL-6，IL-8， and TNF

-α

in Caco-2 cells was significantly reduced （

0.01）， and the expression of tight junction protein Occludin was up-regulated.

Conclusion

In the co-culture system of THP-1 and Caco-2 cells simulating the intestinal epithelial barrier function

in vitro

，ginsenosides Rg

and Rb

play a protective role against LPS-induced intestinal epithelial barrier injury by regulating the release of inflammatory cytokines by THP-1 cells， thereby regulating the inflammatory response and cell barrier integrity of Caco-2 cells.

关键词

Keywords

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⁰

Protective Effect of Ginsenosides Rg1 and Rb1 Against Intestinal Epithelial Barrier Injury Induced by Lipopolysaccharide in Vitro

DOI：10.13422/j.cnki.syfjx.20220705

摘要

Abstract

关键词

Keywords

references

Protective Effect of Ginsenosides Rg₁ and Rb₁ Against Intestinal Epithelial Barrier Injury Induced by Lipopolysaccharide in Vitro