Effects of different culture media and feeder cells on the culture of Echinococcus multilocularis in vitro

doi:10.3969/j.issn.2095-4344.1254

Abstract

Abstract:

BACKGROUND: The cystic culture of Echinococcus multilocularis protoscolex in vitro is the experimental basis for extracting and studying its original cells. D-MEM and RPMI-1640 are the most suitable commercial media for in vitro culture of Echinococcus multilocularis, and HeLa cells and rat hepatoma cells (ATCC No. CRL-1600) are also commonly used feeder cells.
OBJECTIVE: A culture model of Echinococcus multilocularis in vitro was established to observe the growth of Echinococcus multilocularis in different feeder cells (HeLa cells and rat hepatoma cells (ATCC No.CRL-1600) and different media (D-MEM (High and Low glucose type) and RPMI-1640) containing 10% fetal bovine serum.
METHODS: HeLa cells and rat hepatoma cells (ATCC No. CRL-1600) were pre-cultured. The Echinococcus multilocularis metacestode was extracted from mice infected with Echinococcus multilocularis for more than 6 months (provided by the Laboratory Animal Center of the First Affiliated Hospital of Shihezi University, Xinjiang Uygur Autonomous Region) and co-cultured with different feeder cells and culture media. The experiment has been approved by the First Affiliated Hospital of School of Medicine, Shihezi University, approval number: 2015-018-01 on April 7, 2017. The experiment was divided into six groups according to the combination of feeder cells and medium. Each group was added 1x106 feeder cells, 50 mL medium containing 10% fetal bovine serum and 5 000 protoscolexes. Group I: HeLa cells and 1640 medium; group II: HeLa cells and D-MEM (high glucose type) medium; group III: hepatocellular carcinoma cells and 1640 medium; group IV: hepatocellular carcinoma cells and D-MEM (high glucose type) medium; group V: HeLa cells and D-MEM (low glucose type) medium; group VI: hepatocellular carcinoma cells and D-MEM (low glucose type) medium. The survival, growth and development of the protoscolex were observed. The rate of protoscolex vesicle formation was calculated every 7 days, and the average diameter of vesicle was measured using microscope micrometer.
RESULTS AND CONCLUSION: (1) Protoscolex could survive for a long time in all four combinations. The vesicle diameter ranged from 1 to 6 mm. (2) On day 56, the cyst formation rate and average vesicle diameter of protocephalus were the best in rat hepatocellular carcinoma cells and D-MEM (high sugar type) culture medium group (P < 0.05), the cyst formation rate was (72.08±1.79)% and the vesicle diameter was (3.379±0.199) mm. (3) Quantitative analysis of vesicle fluid protein showed that the contents of vesicle fluid protein in six groups were lower than those of vesicles in vivo, and the group IV was higher than those in the other five groups (P < 0.05). (4) These results indicate that under the same conditions, the vesicle size and quantity of D-MEM medium are better than those of the other two media. Rat hepatocellular carcinoma cells (ATCC No. CRL-1600) are better than HeLa cells in size and number of vesicles as feeder cells. The cyst formation rate and vesicle size of D-MEM and rat hepatocellular carcinoma cells (ATCC No. CRL-1600) are the best, which can be used as the optimal choice for obtaining Echinococcus multilocularis vesicles in vitro.

中国组织工程研究杂志出版内容重点：组织构建；骨细胞；软骨细胞；细胞培养；成纤维细胞；血管内皮细胞；骨质疏松；组织工程

Key words: Echinococcu, culture in vitro, culture medium, feeder cells, vesicles, glucose, cyst fluid, diameter, cyst formation rate, protein quantification

CLC Number:

R456
R372

Han Zhenyang, Deng Zi, Wang Zeyu, Zhang Shijie . Effects of different culture media and feeder cells on the culture of Echinococcus multilocularis in vitro[J]. Chinese Journal of Tissue Engineering Research, 2019, 23(19): 3080-3089.

Figures/Tables 2

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