In Vitro Effect of Alcohol Extracts of 5 Traditional Chinese Medicines on Leukocyte Immune Activity in Peripheral Blood of Nile Tilapia
CHENG Min1,2,3, LI Zhongqin1,2,3, ZHAI Shaowei1,2,3, LAI Xiaojian1,2,3, YANG Qiuhua3,4, JIANG Xinglong1,2,3
1.Fisheries College, Jimei University, Xiamen 361021, China; 2.Eel Engineering Research Centre ofthe Modern Industry Technology, Ministry of Education, Xiamen 361021, China; 3.Engineering Technology ResearchCentre of Fishery Medicine in Xiamen, Xiamen 361021, China; 4.Fujian Fisheries Research Institute, Xiamen 361021, China
Abstract:Chinese gall (Galla Chinensis), Chinese pulsaiilla root (Radix Pulsatillae), garden burnet root (Radix Sanguisorbae), fructus chebulae (Terminalia chebula), and giant knotweed rhizome (Rhizoma Polygoni Cuspidati) were powdered and sieved through a 160 mesh sieve, and then mixed with 60% aqueous solution of ethanol at the ratio of material to liquid 1∶10,finally ultrasonicated by an ultrasonic pulverizer for 1 hour. The filtrate was collected by filtration through a filter paper, and concentrated in a multi-sample parallel evaporator to obtain a Chinese medicine extract. About 0.5 mL of blood was collected from the caudal vein of Nile tilapia (Oreochromis niloticus) using a sterile syringe with 0.2 mL of anticoagulant, and mixed with L-15 cell medium at a ratio of 1∶9 by volume to form plasma. The Chinese herbal medicine extract was incubated with the Chinese herbal medicine extract alone and in combination and the effects of Chinese herbal medicine extract on the respiratory burst activity and leukocyte phagocytic activity in peripheral blood leukocytes of Nile tilapia were determined by nitrogen blue tetrazolium reduction method and flow cytometry. Single use of Chinese gall, Chinese pulsaiilla root, garden burnet root, fructus chebulae, and giant knotweed rhizome was shown to lead to significantly enhance the leukocyte oxygenation burst activity by 128.68%, 123.96%, 28.37%, 44.44%, and 78.93%, and the phagocytic rate elevate by 44.04%, 44.66%, 95.00%, 10.35% and 64.93%, respectively. In the double-use experiment of two traditional Chinese medicines, the combination showing synergistic effect on the leukocyte oxygen respiratory burst activity was Chinese pulsaiilla root + garden burnet root, with the combined respiratory burst index of 1.28. There were 4 combinations showing synergistic effects on phagocytic activity of leukocyte, including Chinese pulsaiilla root + giant knotweed rhizome, fructus chebulae + giant knotweed rhizome, Chinese gall + giant knotweed rhizome, and Chinese pulsaiilla root + fructus chebulae, with the phagocytic combined index of 11.45, 6.20, 3.03, and 1.06, respectively.
程敏, 李忠琴, 翟少伟, 赖晓健, 杨求华, 江兴龙. 5味中药醇提物对尼罗罗非鱼白细胞免疫活性的作用[J]. 水产科学, 2020, 39(2): 160-167.
CHENG Min, LI Zhongqin, ZHAI Shaowei, LAI Xiaojian, YANG Qiuhua, JIANG Xinglong. In Vitro Effect of Alcohol Extracts of 5 Traditional Chinese Medicines on Leukocyte Immune Activity in Peripheral Blood of Nile Tilapia. Fisheries Science, 2020, 39(2): 160-167.
[1]孙晓飞. 棕点石斑鱼中草药免疫增强剂的筛选及其非特异性免疫增强效果研究[D].海口:海南大学,2014. [2]陈秋莹,詹剑华.中药免疫调节作用及其研究进展[J].江西医药,2019,54(2):181-184. [3]孟彬,孙敬锋,吕爱军.发酵中草药在水产养殖中的应用[J].水产科学,2018,37(3):421-426. [4]陈丽婷,郇志利,王晓清,等.中草药添加剂在水产养殖中的应用研究进展[J].水产科学,2014,33(3):190-194. [5]李梅芳,张文杰,毛芝娟,等.投喂中草药对大黄鱼几种免疫酶活性的影响[J].水产科学,2014,33(11):718-722. [6]谢丽玲,曹俊辉,杨素霞,等.中草药作为饲料添加剂养殖罗非鱼试验[J].水产科学,2009,28(1):11-14. [7]张照红,林旋,张伟妮,等.复方中草药对奥尼罗非鱼血液非特异性免疫功能的影响[J].水产科学,2011,30(1):1-5. [8]杨雨生,徐家欢,张红霞,等.黄颡鱼免疫增强剂的研究现状和进展[J].水产科技情报,2018,45(6):331-338. [9]林紫薇,汤菊芬,简纪常,等.复方中草药在罗非鱼养殖中的安全性评价[J].广东海洋大学学报,2017,37(3):43-48. [10]石芳芳,李成华,宋林生,等.用流式细胞仪测定扇贝血细胞吞噬活性[J].生物技术通报,2006(增刊):430-433. [11]戴小连. 褐点石斑鱼天然免疫增强剂筛选及黄芪对其免疫功能影响研究[D].海口:海南大学,2012. [12]周春源,旺建伟.中药提取分离技术的研究进展[J].黑龙江中医药,2016,45(2):68-69. [13]王德强,佟延南,李芳远,等.中草药联用对罗非鱼无乳链球菌作用的研究[J].水产养殖,2017,38(1):47-51. [14]高桂平. 日粮中添加4种传统中药方剂对吉富罗非鱼幼鱼的相关非特异性免疫和生长指标的影响[D].南宁:广西大学,2018. [15]解清,孙璇,闫赖赖,等.Percoll密度梯度离心法在血液组分分离及元素测定中的应用[J].中国卫生检验杂志,2015,25(7):954-958. [16]谢昆,黄民昆.鱼类外周血淋巴细胞的分离和体外培养[J].黑龙江畜牧兽医,2011(3):138-142. [17]孙裔雷. 闪光鲟中草药免疫增强剂的筛选及其非特异性免疫增强效果的研究[D].南京:南京农业大学,2015. [18]贺政新,陈晶,王宪灵,等.流式细胞术检测全血白细胞吞噬白念珠菌方法的建立和应用[J].细胞与分子免疫学杂志,2015,31(4):470-473. [19]李进寿,陈军,唐艳霞,等.利用流式细胞仪技术研究杂色鲍血细胞的吞噬率[J].台湾海峡,2010,29(4):460-465,593. [20]李超,陈明,王瑞,等.罗非鱼外周血白细胞分离与血细胞指数研究[J].大连水产学院学报,2009,24(增刊):10-14. [21]Jian J C,Wu Z H. Influences of traditional Chinese medicine on non-specific immunity of Jian carp (Cyprinus carpio var.Jian)[J].Fish & Shellfish Immunology,2004,16(2):185-191. [22]刘敏,潘俊丽,张士璀.鱼类吞噬细胞和吞噬作用的研究进展[J].鲁东大学学报:自然科学版,2010,26(2):167-172,182. [23]林紫薇.一种复方中草药在小鼠及罗非鱼中的安全性评价[D].湛江:广东海洋大学,2017. [24]Ardo' L, Yin G J, Xu P, et al.Chinese herbs (Astragalus membranaceus and Lonicera japonica) and boron enhance the non-specific immune response of Nile tilapia (Oreochromis niloticus) and resistance against Aeromonas hydrophila[J].Aquaculture,2008,275(1/4):26-33.