Establishment and characterization of an intestinal epithelial cell line from Japanese flounder (Paralichthys olivaceus)
Establishment and characterization of an intestinal epithelial cell line from Japanese flounder (Paralichthys olivaceus)
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DOI：
10.1016/j.aquaculture.2023.740325
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发表时间：
2023-11-11
期刊：
影响因子：
4.5
通讯作者：
Xiu，Yunji
中科院分区：
文献类型：
--
作者：
Su，Lin;Guo，Baoshan;Xiu，Yunji
牙鲆（Paralichthys olivaceus）是一种高价值的海水养殖品种。然而，世界范围内的水产养殖业面临着重大挑战，由于比目鱼腹水，一种严重的疾病引起的革兰氏阴性细菌爱德华氏菌感染的鱼类养殖。研究证实，迟缓爱德华氏菌感染比目鱼引起肠道炎症性疾病。肠上皮细胞对病原微生物的识别是炎症反应的主要原因。本研究建立了一种新的比目鱼远端肠（主要是中肠和后肠）细胞系。将该细胞系（命名为PoIEC系）在24 °C下在含有15%胎牛血清的DMEM中维持>1年，并传代培养>55次。PoIEC为鹅卵石样上皮样细胞。透射电镜观察到PoIEC细胞表面有微绒毛，证实了上皮标志物的存在。转染pEGFP-N3质粒的PoIEC发出强烈的绿色荧光信号。这些细胞的转染效率计算为约37%，使该细胞系的一个潜在的工具，为未来的外源基因功能的研究。此外，用LPS和liveE刺激PoIEC。迟发性痴呆，免疫相关基因（MHC Iα、MHC IIα、TNFα、IL-8和IL-1β）的表达谱证实这些细胞识别抗原并激活免疫反应。综上所述，本研究建立了一种新的比目鱼肠道细胞系，该细胞系为研究肠道黏膜免疫、益生植物群和营养物质代谢提供了理想的细胞平台。
The Japanese flounderParalichthys olivaceusis a high-value mariculture species. However, the aquaculture industry worldwide faces significant challenges due to flounder ascites, a serious disease caused by gram-negative bacterium Edwardsiella infection of fish cultures. Studies have confirmed that Edwardsiella tarda infection of flounder causes intestinal inflammatory disease. The recognition of pathogenic microorganisms by intestinal epithelial cells is the primary cause of inflammatory responses. In this study, a new cell line derived from the distal intestine (mainly the mid and posterior intestine) of Japanese flounder was established and characterized. This cell line, designated as the PoIEC line, was maintained in DMEM with 15% fetal bovine serum at 24 °C for >1 year, and subcultured >55 times. The PoIECs were epithelial-like cells with cobblestone-like shapes. Microvilli were observed on the surface of the PoIECs through transmission electron microscopy and the existence of epithelial markers was confirmed. PoIECs transfected with a pEGFP-N3 plasmid emitted an intense green fluorescence signal. Transfection efficiency of these cells was calculated to be approximately 37%, making this cell line a potential tool for future exogenous gene function research. Furthermore, PoIECs were stimulated by LPS and liveE. tarda, and the expression profiles of immune-related genes (MHC Iα,MHC IIα,TNFα,IL-8, andIL-1β) confirmed that these cells recognized antigens and activated immune responses. In conclusion, a new cell line derived from the intestine of Japanese flounder was established, and this cell line was an ideal cellular platform for the study of mucosal immunity, probiotic flora and nutrient metabolism in the fish gut.