Expression profiles of parathyroid glands from healthy pigs challenged by fetal and postnatal dietary interventions
Expression profiles of parathyroid glands from healthy pigs challenged by fetal and postnatal dietary interventions
The experiment was approved by the Animal Welfare Committee of the Leibniz Institute for Farm Animal Biology (FBN) and was licensed by the Ethics Committee of the federal state of Mecklenburg-Western Pomerania, Germany (LALLF 7221.3-1-053/15). At day 120 of life, the pigs were stunned by electronarcosis and subsequently exsanguinated in the experimental facility of the Leibniz Institute for Farm Animal Biology. The slaughtering comprised five batches of 15-17 pigs. The PTGs were dissected from the cranial thymus near the carotid bifurcation, snap frozen in liquid nitrogen and stored at -80 °C.  Fourteen nulliparous German Landrace sows were randomly assigned to one of three dietary treatment groups. Early on adaptation period (10 days) throughout gestation (115 days) and lactation (28 days), the sows were fed iso-energetic standard corn-barley based diet supplemented with Group#1, recommended amounts of P (n=4); Group#2, reduced P content (n=5); Group#3, higher P content (n=5). Water was supplied ad libitum. The sows were artificially inseminated with semen from two German Landrace boars. Pregnant sows were transferred to individual farrowing pens at day 109, where the sows remained with their litter during the lactation period. No cross-fostering was applied. Notably, the offspring of both German Landrace boars is balanced across all dietary groups. An equal number of piglets from each group were assigned to one of three dietary treatments containing medium- (recommended), low- or high-P content supplemented to standard starter, weaning and fattening diets. Water was supplied ad libitum. Pigs were kept in groups of 8-10 individuals in pens equipped with concrete floor.   Tissue was grounded in liquid nitrogen. The total of the left and right PTGs of each animal were used for RNA isolation using TRI reagent according to the manufacturer's protocols (Sigma-Aldrich). Subsequently, Baseline-ZERO DNase treatment was applied (Biozym). Samples were purified using the column-based NucleoSpin RNA II-Kit (Macherey-Nagel). The quantification of RNA was performed with a NanoDrop ND-2000 (Peqlab). The RNA integrity numbers (RIN) were assessed by a 2100 Bioanalyzer instrument (Agilent Technologies) with resulting RIN of 6.5-8.8. The RNA was stored at -80 °C until use. The PTG samples were validated by the qPCR of PTH expression.   RNA libraries were generated from total RNAs according to the TruSeq Stranded mRNA protocol (Illumina) and the quality was validated using a Agilent DNA-1000 chip kit (Bioanalyzer 2100).