Evaluation of the potential of 4 micro algae extracts to modulate the immune response and metabolic state in intestinal epithelium cells. Evaluation of the potential of 4 micro algae extracts to modulate the immune response and metabolic state in intestinal epithelium cells

Micro algae's are used as alternative protein source in human and animal diets. Besides micro algae contain substantial amounts of proteins they also contain a high concentration of, often unique, biological and chemical substances with potential to induce beneficial and health promoting effects in humans and animals. This study was set up to evaluate the potential of these substances to improve (intestinal) health. The effect of extracts prepared from 3 monocultures of micro algae's (Chlorella vulgaris [C], Haematococcus pluvialis [H], and Spirulina platensis [S]) and a mixed culture of micro algae's (AM; a mixture of Scenedesmus sp. and Chlorella sp. ) was studied in the presence and absence of the enterotoxigenic bacterium Escherichia coli k99 strain (ETEC, [E]) as an in vitro challenge. The E.coli-k99 strain with adhesion factor F41 (41/32) was isolated from a mastitis-infected udder. Gene expression was measured in cultured intestinal porcine epithelium cells (IPECJ2 cell line) after 2 and 6 hours incubation with C, H, and S extracts, and after 6 hours with the AM extract, using “whole genome” porcine microarrays. Gene expression profiles were analysed using functional bioinformatics programs to provide insight in the biological processes induced by micro algae extracts. Overall design: IPECJ2 cells were grown in 2 cm2 wells for 7 days at 37 ºC and 5% CO2 using 1:1 DMEM/Ham’s F10 1:1 medium supplemented with 5% FCS without antibiotics. For all tests, confluent monolayers were washed twice with medium without FCS (hereafter denoted as medium) and incubated for 1 hour with this medium. Hereafter, the medium was discarded and algae extracts suspended in medium were added to wells with IPECJ2 monolayers and incubated for a period of 2 and 6 hours in the absence and presence of ETEC (±10 cfu per cell). All incubations were tested in duplicate and for each type of additive duplicate control wells containing no additive (only culture medium) were incubated for 2 and 6 hours. After incubation total RNA was extracted. RNA's extracted from replicates were pooled (biological replicates) and this pool was hybridized in duplicate (technical replicates). IPECJ2 enterocyte cell line derived from the jejunum of piglets, host-feed interaction

微藻（micro algae）可作为人类与动物膳食中的替代蛋白质来源。除富含蛋白质外，微藻还含有高浓度、通常具有独特性的生物与化学物质，这类物质有望对人类及动物产生有益的健康促进作用。本研究旨在评估这类物质改善（肠道）健康的潜力。
本研究探究了3株单培养微藻（普通小球藻（Chlorella vulgaris）[C]、雨生红球藻（Haematococcus pluvialis）[H]、钝顶螺旋藻（Spirulina platensis）[S]）以及1株混合培养微藻（AM：栅藻属（Scenedesmus）物种与小球藻属（Chlorella）物种的混合物）的提取物，在存在与不存在肠毒素性大肠杆菌K99菌株（enterotoxigenic Escherichia coli k99 strain，ETEC[E]，作为体外攻毒模型）的条件下的生物学效应。本次实验使用的带有黏附因子F41（41/32）的大肠杆菌K99菌株，分离自患乳腺炎的乳腺组织。
本研究采用猪全基因组微阵列技术，对培养的猪肠道上皮细胞（IPEC-J2细胞系）进行基因表达量检测：分别在与C、H、S提取物共孵育2小时、6小时后取样，以及与AM提取物共孵育6小时后取样。通过功能生物信息学工具分析基因表达谱，以解析微藻提取物所诱导的生物学过程。
实验整体设计如下：将IPEC-J2细胞接种于2 cm²培养孔中，于37℃、5% CO₂条件下培养7天，所用培养基为1:1混合的DMEM与Ham’s F10培养基，添加5%胎牛血清（Fetal Calf Serum, FCS）且不含抗生素。所有实验开始前，将汇合的单层细胞用不含FCS的培养基（后续简称"基础培养基"）洗涤两次，随后用该培养基孵育1小时。弃去培养基后，将悬浮于基础培养基中的微藻提取物加入IPEC-J2单层细胞培养孔中，分别在存在或不存在ETEC（每细胞约10 cfu）的条件下孵育2小时与6小时。所有孵育实验均设置重复孔；同时，针对每一类添加物，均设置不含添加物（仅含基础培养基）的重复对照孔，分别孵育2小时与6小时。孵育结束后提取总RNA。将生物学重复样本的RNA混合后，设置技术重复进行杂交实验。IPEC-J2细胞系为源自仔猪空肠的肠上皮细胞系，用于宿主-饲料互作研究。