Liver transcriptome profiling reveals that dietary DHA and EPA levels influence suites of genes involved in metabolism, redox homeostasis, and immune function in Atlantic salmon (Salmo salar)

The optimal dietary requirement of omega-3 long-chain polyunsaturated fatty acids (ω3 LC-PUFA), namely docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA), for Atlantic salmon that promotes optimal growth and health warrants careful investigation. We used 44K microarrays to study the influence of increasing levels of dietary DHA + EPA (0, 1.0 and 1.4% of the diet, as formulated) in the presence of high linoleic acid (LA) on Atlantic salmon growth and liver transcriptome. After a 14-week feeding trial, Atlantic salmon fed diet ω3LC0 (i.e. 0% of DHA + EPA) showed significantly lower final weight and weight gain, and higher feed conversion ratio compared with ω3LC1.0 and ω3LC1.4 diet groups. The microarray experiment identified 55 and 77 differentially expressed probes (Rank Products analyses; PFP < 10%) in salmon fed diets ω3LC1.4 and ω3LC1.0 compared with those fed diet ω3LC0, respectively. The comparison between ω3LC1.4 and ω3LC1.0 revealed 134 differentially expressed probes. Atlantic salmon were fed the three experimental diets (ω3LC0, ω3LC1, and ω3LC1.4; four tanks per diet) for 14 weeks. Liver samples were taken from 5 fish from each tank at week 16. A universal reference design was used for the microarray experiment. For the test samples, RNA was used from individual livers of fish from the 3 treatment groups. For each treatment group we used 8 biological replicates (2 fish from each of 4 tanks). All test samples were labeled with Cy5. The common reference was a pool of 24 RNA samples from livers of fish from all individuals invovled in microarray experiment. The common reference was labeled with Cy3. Each individual test sample was hybridized together with the common reference sample on an array, so the experiment consisted of 24 arrays. Please note that sample titles represent 'diet_tank number_individual fish number' for each sample. variable: other: ω3LC0: ω3LC0_T8_2, ω3LC0_T8_3, ω3LC0_T23_1, ω3LC0_T23_3, ω3LC0_T31_2, ω3LC0_T31_4, ω3LC0_T35_1, ω3LC0_T35_3 variable: other: ω3LC1: ω3LC1_T7_1, ω3LC1_T7_3, ω3LC1_T20_1 ,ω3LC1_T20_2, ω3LC1_T34_1, ω3LC1_T34_3, ω3LC1_T38_2, ω3LC1_T38_3 variable: other: ω3LC1.4: ω3LC1.4_T3_3, ω3LC1.4_T3_4, ω3LC1.4_T19_2, ω3LC1.4_T19_3, ω3LC1.4_T26_1, ω3LC1.4_T26_2, ω3LC1.4_T33_1, ω3LC1.4_T33_2 repeat: biological replicate: ω3LC0_T8_2, ω3LC0_T8_3, ω3LC0_T23_1, ω3LC0_T23_3, ω3LC0_T31_2, ω3LC0_T31_4, ω3LC0_T35_1, ω3LC0_T35_3 repeat: biological replicate: ω3LC1_T7_1, ω3LC1_T7_3, ω3LC1_T20_1 ,ω3LC1_T20_2, ω3LC1_T34_1, ω3LC1_T34_3, ω3LC1_T38_2, ω3LC1_T38_3 repeat: biological replicate: ω3LC1.4_T3_3, ω3LC1.4_T3_4, ω3LC1.4_T19_2, ω3LC1.4_T19_3, ω3LC1.4_T26_1, ω3LC1.4_T26_2, ω3LC1.4_T33_1, ω3LC1.4_T33_2

大西洋鲑（Atlantic salmon）对Omega-3长链多不饱和脂肪酸（ω3 LC-PUFA）——即二十二碳六烯酸（DHA）与二十碳五烯酸（EPA）——的最优膳食需求量，该物质可促进机体最佳生长与健康，仍有待开展系统性严谨探究。本研究采用44K微阵列（44K microarrays）技术，探究在高亚油酸（LA）环境下，递增剂量的膳食DHA+EPA（按配方分别为日粮的0、1.0%及1.4%占比）对大西洋鲑生长性能与肝脏转录组的影响。
经过14周的饲喂试验后，与饲喂ω3LC1.0及ω3LC1.4日粮的组别相比，饲喂ω3LC0日粮（即DHA+EPA占比为0%）的大西洋鲑终末体重、增重显著更低，且饲料转化率更高。通过秩积分析（Rank Products analyses，PFP < 10%）鉴定得出：相较于饲喂ω3LC0日粮的鲑鱼，饲喂ω3LC1.4与ω3LC1.0日粮的组别分别存在55个与77个差异表达探针；而ω3LC1.4与ω3LC1.0组间的比较则鉴定出134个差异表达探针。
本试验设置3组日粮处理（ω3LC0、ω3LC1及ω3LC1.4），每组设置4个养殖单元，饲喂周期为14周。于第16周时，从每个养殖单元中选取5尾鱼采集肝脏样本。微阵列实验采用通用参考设计：测试样本采用各处理组个体肝脏的总RNA，每个处理组设置8个生物学重复（即从4个养殖单元中各选取2尾鱼）。所有测试样本均以Cy5荧光标记；通用参考样本为参与本次微阵列实验的所有鱼类的肝脏RNA混合样品，以Cy3荧光标记。每个测试样本均与通用参考样本共同杂交于一张微阵列芯片，本试验共计使用24张芯片。
请注意，每个样本的命名格式为「日粮组_养殖单元编号_个体鱼编号」。
变量：其他：
ω3LC0组样本：ω3LC0_T8_2、ω3LC0_T8_3、ω3LC0_T23_1、ω3LC0_T23_3、ω3LC0_T31_2、ω3LC0_T31_4、ω3LC0_T35_1、ω3LC0_T35_3
变量：其他：
ω3LC1组样本：ω3LC1_T7_1、ω3LC1_T7_3、ω3LC1_T20_1、ω3LC1_T20_2、ω3LC1_T34_1、ω3LC1_T34_3、ω3LC1_T38_2、ω3LC1_T38_3
变量：其他：
ω3LC1.4组样本：ω3LC1.4_T3_3、ω3LC1.4_T3_4、ω3LC1.4_T19_2、ω3LC1.4_T19_3、ω3LC1.4_T26_1、ω3LC1.4_T26_2、ω3LC1.4_T33_1、ω3LC1.4_T33_2
生物学重复信息：
ω3LC0组生物学重复：ω3LC0_T8_2、ω3LC0_T8_3、ω3LC0_T23_1、ω3LC0_T23_3、ω3LC0_T31_2、ω3LC0_T31_4、ω3LC0_T35_1、ω3LC0_T35_3
ω3LC1组生物学重复：ω3LC1_T7_1、ω3LC1_T7_3、ω3LC1_T20_1、ω3LC1_T20_2、ω3LC1_T34_1、ω3LC1_T34_3、ω3LC1_T38_2、ω3LC1_T38_3
ω3LC1.4组生物学重复：ω3LC1.4_T3_3、ω3LC1.4_T3_4、ω3LC1.4_T19_2、ω3LC1.4_T19_3、ω3LC1.4_T26_1、ω3LC1.4_T26_2、ω3LC1.4_T33_1、ω3LC1.4_T33_2