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首页> 外文期刊>Fish & Shellfish Immunology >Fish cell cultures as in vitro models of inflammatory responses elicited by immunostimulants. Expression of regulatory genes of the innate immune response
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Fish cell cultures as in vitro models of inflammatory responses elicited by immunostimulants. Expression of regulatory genes of the innate immune response

机译：鱼细胞培养物是免疫刺激剂引起的炎症反应的体外模型。先天免疫应答调节基因的表达

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We report the differential expression of various genes related to the regulation of the innate immune responses, including pro-inflammatory (IL-1 beta 1, IL-8, TNF-alpha 1, TNF-alpha 2) and immune-suppressing (IL-10) cytokines, interferon-induced Mx-1 protein, enzymes regulating nitric oxide (inducible nitric oxide synthase, arginase-2) and eicosanoid (COX-2) production, and Toll-like pathogen pattern-recognition receptors TLR-3, TLR-5 and TLR-9, in two lympho-haematopoietic stromal cell lines derived from the spleen (trout splenic stroma, TSS) and the pronephros (trout pronephric stroma-2, TPS-2) of rainbow trout (Oncorhynchus mykiss), as well as in primary cultures of rainbow trout head kidney macrophages, after their exposure to the well-known immunostimulants LPS, levamisole and poly I:C. Although there were differences in the responses between the two stromal cell lines, using reverse transcription followed by real time polymerase chain reaction (RT-qPCR) we demonstrated that exposure to the immunostimulants, particularly poly I:C and LPS, resulted in significant changes in the expression of the immunoregulatory genes in the two stromal cell lines in many cases their responses resembling in fold change magnitudes and in response profiles to those observed in the primary macrophage cultures. Exposure to poly I:C and, with lower fold change values, to LPS produced upregulation of the pro- (IL-1 beta, IL-8, TNF-alpha) and anti-inflammatory (IL-10) cytokine genes, as well as of the Mx-1 gene. Furthermore, the immunostimulation elicited the upregulation of COX-2, iNOS and arginase-2 genes in the cell lines. Likewise, the TSS and TPS-2 cell lines significantly upregulated the expression of TLR-3, TLR-5 and TLR-9 genes after exposure to the immunostimulants, thus explaining the ability of the stromal cells to recognise and respond to the immunostimulants. Such results give support to an important role of lympho-haematopoietic stromal cells in the development and control of pro-inflammatory responses in fish. The upregulation of genes of pro-inflammatory cytokines and of mediators of the innate immune responses correlates well with the previously demonstrated functional capacities, including phagocytosis, microbicidal activity and NO production, exhibited by the TSS and TPS-2 stromal cell lines when exposed to the same immunostimulants. On the other hand, the expression of immunosuppressing genes (IL-10, COX-2 and arginase-2) demonstrate that the lympho-haematopoietic stromal cells are also able to contribute to the control of inflammatory responses. This study reinforce the possibility of using histotypic cell cultures, as those formed by the TSS and TPS-2 cell lines, formed by heterogeneous cell populations that partially replicates the cell-cell and cell-extracellular matrix interactions, to develop cost-effective and repetitive in vitro systems for the screening of immunostimulant candidates for aquaculture, as they are able to replicate in vitro immune regulatory networks occurring in vivo. (C) 2013 Elsevier Ltd. All rights reserved.
机译：我们报告了与先天免疫反应的调节相关的各种基因的差异表达，包括促炎性（IL-1 beta 1，IL-8，TNF-alpha 1，TNF-alpha 2）和免疫抑制（IL- 10）细胞因子，干扰素诱导的Mx-1蛋白，调节一氧化氮的酶（诱导型一氧化氮合酶，精氨酸酶2）和类花生酸（COX-2）的产生，以及Toll样病原体模式识别受体TLR-3，TLR- 5和TLR-9，分别来自虹鳟（Oncorhynchus mykiss）的脾脏（鳟鱼脾基质，TSS）和前ph（鳟鱼肾基质2，TPS-2）以及两种淋巴造血基质细胞系，以及在虹鳟鱼头肾巨噬细胞的原代培养物中，暴露于众所周知的免疫刺激剂LPS，左旋咪唑和聚I：C后，就可以了。尽管两种基质细胞系之间的反应存在差异，但通过逆转录随后进行实时聚合酶链反应（RT-qPCR），我们证明了暴露于免疫刺激剂（特别是poly I：C和LPS）会导致在许多情况下，免疫调节基因在两个基质细胞系中的表达与原始巨噬细胞培养物中观察到的响应相似，是倍数变化幅度和响应曲线。暴露于poly I：C以及倍数变化值较低的LPS，也会导致pro-（IL-1 beta，IL-8，TNF-alpha）和抗炎性（IL-10）细胞因子基因上调与Mx-1基因相同。此外，免疫刺激引起细胞系中COX-2，iNOS和精氨酸酶-2基因的上调。同样，在暴露于免疫刺激剂后，TSS和TPS-2细胞系显着上调了TLR-3，TLR-5和TLR-9基因的表达，从而解释了基质细胞识别和响应免疫刺激剂的能力。这些结果支持了淋巴造血基质细胞在鱼类促炎反应的发展和控制中的重要作用。 TSS和TPS-2基质细胞系暴露于TPS和TPS-2基质细胞系后，促炎细胞因子基因和先天性免疫应答介质的上调与TSS和TPS-2基质细胞系表现出的先前证明的功能能力（包括吞噬作用，杀微生物活性和NO产生）密切相关。相同的免疫刺激剂。另一方面，免疫抑制基因（IL-10，COX-2和精氨酸酶-2）的表达表明，淋巴造血基质细胞也能够促进炎症反应的控制。这项研究增强了使用组织型细胞培养物的可能性，例如由TSS和TPS-2细胞系形成的，由部分复制细胞-细胞和细胞-细胞外基质相互作用的异质细胞群体形成的培养物，以开发具有成本效益和重复性的用于筛选水产养殖免疫刺激候选物的体外系统，因为它们能够复制体内发生的体外免疫调节网络。 （C）2013 Elsevier Ltd.保留所有权利。

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