旨在探讨日本血吸虫(Schistosoma japonicum)胞外囊泡(Extracellular vesicles，EVs)的提取方法及特性。本研究利用尺寸排阻法收集SjEVs，通过透射电镜、蛋白质谱对SjEVs的形态学、蛋白谱进行解析并对鉴定到的蛋白进行生物信息学分析。将PKH67绿色荧光标记的SjEVs与人肝星状细胞LX-2共孵育，观察细胞摄取SjEVs的情况，并对细胞中虫源miRNAs及细胞内纤维化因子进行检测，初步探讨SjEVs对肝星状细胞的影响。结果表明:SjEVs呈圆形或椭圆形囊泡结构，内含EVs标志蛋白。生物信息学分析结果显示，SjEVs蛋白可能与细胞进程、代谢等途径密切相关。SjEVs与人肝星状细胞LX-2共孵育结果显示，LX-2细胞可成功摄取SjEVs，且定量反转录PCR(RT-qPCR)结果显示LX-2细胞中部分虫源miRNAs显著升高，纤维化因子α-平滑肌肌动蛋白(α-SMA)、Ⅰ 型胶原纤维 α1(Collagen type Ⅰ alpha 1，Col1α1)和 Ⅲ 型胶原纤维 α1(Collagen type Ⅲ alpha 1，Col3α1)极显著升高，提示肝星状细胞活化。综上，本研究为解析SjEVs在宿主肝纤维化中的作用提供了依据，为进一步探究SjEVs的潜在功能奠定了基础。
Extraction and Identification of Schistosoma japonicum Extracellular Vesicles and Its Effect on Human Hepatic Stellate Cells
To explore the isolation technique and characterization of extracellular vesicles(EVs)of Schistosoma japonicum,in the present study,the size exclusion method was utilized to collect SjEVs,and the morphology and protein profiles of SjEVs were analyzed by transmission electron microscopy and protein mass spectrometry,respectively.Bioinformatics analysis was conducted on all identified proteins.PKH67 green fluorescence labeled SjEVs were co-incubated with human hepatic stellate cell line(LX-2)to observe the uptake of SjEVs and the expression of Sj-miRNAs and the fibrotic factors in the cells were also detected.The results showed that SjEVs presented vesicular structures and contained EVs marker proteins.Bioinformatics analysis showed that SjEVs proteins might be closely related to cellular processes and metabolism pathways.Co-incubation of SjEVs with LX-2 cells revealed that SjEVs could be successfully absorbed,and significantly increased expression of Sj-miRNAs and fibrotic factors were also observed.In summary,this study provides a theoretical basis for analyzing the potential functions of SjEVs,with a view to laying a foundation for further elucidation of schistosome-host interactions.
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