方格星虫酶解超滤组分对顺铂所致HK-2人肾小管上皮细胞损伤的保护作用研究  

Protective effect of ultrafiltration fractions of Sipunculus nudus enzymatic hydrolysate on cisplatin-induced HK-2 human renal tubular epithelial cell damages

作　　者：陶素素 彭志兰 陈晖 李茹 刘华锋 杨陈 戚怡 TAO Susu;PENG Zhilan;CHEN Hui;LI Ru;LIU Huafeng;YANG Chen;QI Yi(School of Pharmacy,Guangdong Medical University,Zhanjiang 524000,China;Institute of Marine Medicine,Guangdong Medical University,Zhanjiang 524000,China;Third Institute of Oceanography,Ministry of Natural Resources,Xiamen 361000,China;Institute of Nephropathy,Affiliated Hospital of Guangdong Medical University,Zhanjiang 524000,China)

机构地区：[1]广东医科大学药学院,广东湛江524000 [2]广东医科大学海洋医药研究院,广东湛江524000 [3]自然资源部第三海洋研究所,福建厦门361000 [4]广东医科大学附属医院肾病研究所,广东湛江524000

出　　处：《中国海洋药物》2024年第2期41-52,共12页Chinese Journal of Marine Drugs

基　　金：南海海洋生物制品产业化关键技术研发与创新示范平台项目(2019A01005);广东省医学科研基金项目(A2021190)资助。

摘　　要：目的探究方格星虫酶解产物对顺铂诱导的HK-2人肾小管上皮细胞损伤的改善作用。方法方格星虫酶解后的离心液逐级通过50 kDa和2.5 kDa超滤膜后,获得>50 kDa(F-Ⅰ),2.5 kDa~50 kDa(F-Ⅱ),<2.5 kDa(F-Ⅲ)3个超滤组分,分析其营养成分、分子量排布和氨基酸组成;在顺铂诱导HK-2细胞损伤模型上,评价各超滤组分对顺铂损伤的HK-2细胞活力、细胞凋亡以及细胞内ROS的产生的影响;采用Western blotting探究其中活性最优F-Ⅰ超滤组分对NF-κB信号通路的影响。结果营养成分和分子量排布分析发现,F-Ⅰ超滤组分中总糖所占的比例最高,以干基计约占48.59%,逐级超滤可以提高超滤组分中多肽的含量,F-Ⅲ组分主要为<2.5 kDa的小分子肽。氨基酸组成分析发现3个超滤组分氨基酸组成齐全,疏水性氨基酸含量高。细胞实验结果表明3个超滤组分均能显著提高顺铂损伤后的HK-2细胞的细胞活力,抑制细胞凋亡和减少细胞内活性氧ROS的累积。其中F-Ⅰ组分缓解顺铂对HK-2细胞损伤的效果最好,能抑制NF-κB信号通路p65和Iκb-α蛋白的磷酸化。结论方格星虫的3个酶解超滤组分均对顺铂损伤后的HK-2细胞具有保护作用。相较于其他两个组分,含有较高多糖的F-Ⅰ组分具有更好的改善作用,可能机制与F-Ⅰ组分能更好抑制ROS的积累、细胞凋亡以及NF-κB信号通路有关。Objective To investigate the protective effect of ultrafiltration fractions of Sipunculus nudus enzymatic hydrolysate on cisplatin-induced HK-2 cell damages.Methods Sipunculus nudus was hydrolyzed by trypsase,then gradually ultra-filtrated by 50 kDa and 2.5 kDa membrane with molecular weight cutoffs to separate three ultrafiltration fractions,including>50 kDa(F-Ⅰ),2.5 kDa-50 kDa(F-Ⅱ)and<2.5 kDa(F-Ⅲ).The nutritional ingredients and the amino acid composition of three fractions were further analysed.The ameliorative effects of three fractions were tested in a cisplatin-damaged renal toxicity model by evaluating its cell viability,cell apoptosis and intracellular ROS production level in HK-2 cells.The effect of the most active F-Ⅰultrafiltration component on the NF-κB signaling pathway was investigated by Western blotting.Results The nutrient composition analysis showed that the proportion of polysaccharides in F-Ⅰultrafiltration fraction was the highest,about 48.59%,and the content of peptides in ultrafiltration fractions could be increased by gradually ultrafiltration,the main component of F-Ⅲwas peptides with molecular weight below<2.5 kDa.Three ultrafiltration fractions had complete amino acid composition and high hydrophobic amino acid content.Cell experiment showed that three fractions significantly improved the cell viability of HK-2 cells,inhibited its cell apoptosis and reduced the accumulation of intracellular reactive oxygen species(ROS)in HK-2 cells after cisplatin exposure.F-Ⅰhad the best effect on cisplatin-induced damages of HK-2 cells,as well as inhibited the phosphorylation of p65 and Iκb-αproteins in the NF-κB signaling pathway.Conclusion The effect of F-Ⅰfraction on cisplatin-induced cell damages was significantly better than that of other fractions,which possibly related to better regulating ROS accumulation,cell apoptosis and NF-κB signaling pathway in HK-2 cells.

关 键 词：方格星虫 超滤组分 NF-ΚB信号通路 顺铂 人肾小管上皮细胞 

分 类 号：R931.77[医药卫生—生药学]