北柴胡多糖对D-gal诱导的HK-2细胞氧化损伤的保护作用

doi:10.13481/j.1671-587X.20220109

吉林大学学报(医学版) ›› 2022, Vol. 48 ›› Issue (1): 65-73.doi: 10.13481/j.1671-587X.20220109

北柴胡多糖对D-gal诱导的HK-2细胞氧化损伤的保护作用

侯俊宇¹,李明慧¹,许梦然²,葛俊宏²,申野³(),李坦³,孙新²()

^1.北华大学基础医学院病理生理学教研室，吉林吉林 132013
^2.吉林医药学院药学院生物制药教研室，吉林吉林 132013
^3.北华大学公共卫生学院社会医学与卫生教研室，吉林吉林 132013

收稿日期:2021-04-13 出版日期:2022-01-28 发布日期:2022-01-17
通讯作者: 申野,孙新 E-mail:56134761@qq.com;sunxinbh@126.com
作者简介:侯俊宇（1994－），男，吉林省四平市人，在读硕士研究生，主要从事抗衰老药物作用机制方面的研究。
基金资助:
吉林省科技厅重点实验室项目(20200201178JC);吉林省发改委自主创新能力项目(2021C007);吉林医药学院博士基金项目(JYBS2021026LK);吉林医药学院横向科研项目(2021-HX002);吉林省吉林市科技局科技创新发展计划项目(201830827)

Protective effect of Bupleurum chinense polysaccharide on oxidative damage of HK-2 cells induced by D-gal

Junyu HOU¹,Minghui LI¹,Mengran XU²,Junhong GE²,Ye SHEN³(),Tan LI³,Xin SUN²()

^1.Department of Pathophysiology，School of Basic Medical Science，Beihua University，Jilin 132013，China
^2.Staff Office of Biopharmaceutical，School of Pharmacy，Jilin Medical University，Jilin 132013，China
^3.Department of Social Medicine and Hygiene，School of Public Health，Beihua University，Jilin 132013，China

Received:2021-04-13 Online:2022-01-28 Published:2022-01-17
Contact: Ye SHEN,Xin SUN E-mail:56134761@qq.com;sunxinbh@126.com

摘要/Abstract

摘要： 目的

研究北柴胡多糖（BCP）体外抗氧化活性，阐明其对D-半乳糖（D-gal）诱导的人肾脏近曲小管HK-2细胞的保护作用。

方法

采用热水浸提法提取BCP，检测其抗氧化活性，即体外羟自由基清除率、总抗氧化能力和抗超氧阴离子活性，并与维生素E的体外抗氧化活性进行比较。采用不同浓度BCP和D-gal分别作用于HK-2细胞24 h，采用MTT法检测细胞增殖率，确定BCP和D-gal最适作用浓度。采用D-gal构建人肾脏近曲小管HK-2细胞氧化损伤模型， HK-2细胞分为对照组、D-gal组和不同浓度（25、50、100、200及400 mg·L^－1）BCP+D-gal组，采用MTT法检测各组细胞增殖率。HK-2细胞分为对照组、^{D-gal组、BCP+D-gal组和维生素E+D-gal组，采用衰老相关β-半乳糖苷酶（SA-β-gal）染色实验观察各组HK-2细胞SA-β-gal染色情况并计算衰老细胞百分率。}

结果

抗氧化活性检测，与相同浓度维生素E组比较，不同浓度BCP组羟自由基清除率明显升高（P<0.05），总抗氧化能力明显增强（P<0.05），5、10和15 g·L^－1 BCP组抗超氧阴离子活性明显增加（P<0.05），且不同浓度BCP组体外抗氧化活性均高于同等浓度维生素E组。与对照组比较，处理HK-2细胞24 h，25~400 mg·L^－1 BCP组HK-2细胞增殖率呈浓度依赖性升高（P<0.05），且200 mg·L^－1 BCP组HK-2细胞增殖率最高。与对照组比较，不同浓度D-gal组HK-2细胞增殖率呈时间-浓度依赖性降低，在处理条件为20 g·L^－1、48 h时，细胞增殖率降低至对照组的64.77%。与对照组比较，D-gal组HK-2 细胞增殖率明显降低（P<0.01）；与D-gal组比较，100~400 mg·L^－1 BCP+D-gal组HK-2细胞增殖率呈浓度依赖性升高（P<0.05或P<0.01）。与对照组比较，D-gal组SA-β-gal染色阳性细胞数明显增多，衰老细胞百分率明显升高（P<0.05）；与D-gal组比较，BCP+D-gal组和维生素E+D-gal组SA-β-gal染色阳性细胞数明显减少，衰老细胞百分率明显降低（P<0.05）。

结论

BCP具有较强的体外抗氧化活性，可通过抑制细胞衰老保护D-gal诱导的HK-2细胞氧化损伤。

关键词: 北柴胡多糖, 人肾脏近曲小管细胞, 氧化损伤, 细胞衰老

Abstract: Objective

To study the antioxidant activity in vitro of Bupleurum chinense polysaccharides （BCP），and to clarify its protective effect on D-galactose （D-gal）-induced proximal convolve tubule HK-2 cells of human kidney.

Methods

BCP was extracted by hot water extraction method. The antioxidant activities of BCP including the hydroxyl radical scavenging rate，the total antioxidant capacity and the anti-superoxide anion activities were detected，and compared to the antioxidant activities of vitamin E.The HK-2 cells were treated with different concentrations of BCP and D-gal，respectively，and the proliferation rates of HK-2 cells were detected；the optimum concentrations of BCP and D-gal were confinmed.D-gal was used to establish the oxidative damage model of proximal convolve tubule of human kidney.The HK-2 cells were divided into control group，D-gal group，and 25，50，100，200，400 mg·L^－1 BCP+D-gal groups；MTT method was used to detect the proliferation rates.The HK-2 cells were divided into control group，D-gal group，BCP+D-gal group and vitamin E+D-gal group；senescence-associated β-galactosidase（SA-β-Gal） staining was used observe the SA-β-gal staining of the HK-2 cells in various groups and the percentages of senescent the HK-2 groups cells were calculated.

Results

The antioxidant activity detection results showed that compared with vitamin E group with the same concentration， the scavenging rates of hydroxyl free radical in different concentrations of BCP groups were increased（P<0.05）， and the total antioxidant capacitie were increased（P<0.05）； the anti-superoxide anion activities in 5，10，and 15 g·L^－1 BCP groups were significantly increased（P<0.05）， and the antioxidant levels in vitro in different concentrations of BCP groups were higher than those in vitamin E groups with the same concentration. Compared with control group，the proliferation rates of HK-2 cels in 25－400 mg·L^－１BCP groups were increased in a concentration-dependent manner after treated for 24 h， and the proliferation rate of HK-2 cells in 200 mg·L^－1BCP group was the highest （P<0.05）. Compared with control group， the proliferation rates of HK-2 cells in different concentrations of D-gal groups were decreased in a time-dose dependent manner； when the treatment conditions were 20 g·L^－1 and 48 h， the proliferation rate of cells was decreased to 64.77% of that in control group （P<0.05）.Compared with control group，the proliferation rate of HK-2 cells in D-gal group was significantly decreased（P<0.05）； compared with D-gal group， the proliferation rates of HK-2 cell in 100－400 mg·L^－1BCP group increased in a dose-dependent manner （P<0.05 or P<0.01）. Compared with control group， the number of SA-β-gal positive staining cells in D-gal group was significantly increased（P<0.05）， and the percentage of senescent cells was increased （P<0.05）； compared with D-gal group， the number of SA-β-gal positive staining cells in BCP+D-gal group and vitamin E+D-gal group was significantly reduced（P<0.05）， and the percentages of senescent cells were decreased （P<0.05）.

Conclusion

BCP has a strong antioxidant activity in vitro and protects the oxidative damage of HK-2 cells induced by D-gal by inhibiting cell senescence.

Key words: Bupleurum chinense polysaccharide, Human kidney proximal tubule cells, Oxidative damage, Senescence

中图分类号:

R285.5

侯俊宇,李明慧,许梦然,葛俊宏,申野,李坦,孙新. 北柴胡多糖对D-gal诱导的HK-2细胞氧化损伤的保护作用[J]. 吉林大学学报(医学版), 2022, 48(1): 65-73.

Junyu HOU,Minghui LI,Mengran XU,Junhong GE,Ye SHEN,Tan LI,Xin SUN. Protective effect of Bupleurum chinense polysaccharide on oxidative damage of HK-2 cells induced by D-gal[J]. Journal of Jilin University(Medicine Edition), 2022, 48(1): 65-73.

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Group	Proliferation rate of HK-2 cells
Group	(t/h) 6	12	24	48
Control	100.00±2.97	100.00±2.08	100.00±0.02	100.00±0.04
D-gal(g·L^－1) 5	93.59±3.94^*	92.39±1.88^*	90.87±2.02^*	85.28±3.54^*
10	89.97±1.64^*	87.59±1.35^*	82.72±2.27^*	75.90±2.04^*
20	85.82±2.18^*	85.92±1.68^*	74.41±1.57^*	64.77±1.31^*
40	81.03±2.79^*	81.01±1.24^*	68.42±1.60^*	58.24±3.44^*
80	75.13±2.16^*	71.06±0.93^*	58.92±0.78^*	37.08±1.07^*
100	69.64±3.20^*	64.30±1.27^*	53.36±1.09^*	27.81±0.81^*

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北柴胡多糖对D-gal诱导的HK-2细胞氧化损伤的保护作用

Protective effect of Bupleurum chinense polysaccharide on oxidative damage of HK-2 cells induced by D-gal

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