Defining the lactocrine-sensitive neonatal porcine uterine transcriptome

Authors: RAHMAN, KATHLEEN - Rutgers University; CAMP, MEREDITH - Rutgers University; PRASAD, NRIPESH - Hudsonalpha Institute For Biotechnology; McNeel, Anthony; LEVY, SHAWN - Hudsonalpha Institute For Biotechnology; BARTOL, FRANK - Auburn University; BAGNELL, CAROL - Rutgers University

Submitted to: Society for the Study of Reproduction Annual Meeting
Publication Type: Abstract Only
Publication Acceptance Date: 7/1/2014
Publication Date: 7/19/2014
Citation: Rahman, K.M., Camp, M.E., Prasad, N., McNeel, A.K., Levy, S., Bartol, F.F., Bagnell, C.A. 2014. Defining the lactocrine-sensitive neonatal porcine uterine transcriptome [abstract]. In: Society for the Study of Reproduction, 47th Annual Meeting, 19-23 July 2014, Grand Rapids, MI. Abstract No. 485.

Interpretive Summary:

Technical Abstract: Milk-borne bioactive factors, delivered from mother to nursing offspring via a lactocrine mechanism, affect development of somatic tissues including the uterus. In the pig, lactocrine-sensitive gene expression events associated with the onset of endometrial adenogenesis between birth (postnatal day = PND 0) and PND 2 define the uterine developmental program and can determine developmental trajectory and function. Neither the neonatal porcine uterine developmental transcriptome, nor associated lactocrine-sensitive elements of this transcriptome, have been defined during this period. Here, objectives were to determine effects of age and imposition of the lactocrine-null condition for 48 h from birth by milk replacer feeding, on the uterine transcriptome at PND 2 using whole transcriptome RNA sequencing (RNAseq). Crossbred gilts (Sus scrofa domesticus, n = 4/group) were assigned at birth: (1) to have uteri collected prior to nursing, within 1 h of birth; or to be (2) nursed ad libitum for 48 h, or (3) gavage-fed commercial porcine milk-replacer for 48 h (30ml/kg BW/2h). Uteri were obtained from nursed and replacer-fed gilts at 50 h. For each uterus, total RNA was extracted and both RNA concentration and integrity were estimated. For RNAseq, 500 ng of RNA from each uterus was used for library preparation. Each library was sequenced (paired end) at > 90 million reads per sample, demultiplexed, and raw reads were mapped to the latest pig Sscrofa10.2 build using the Avadis NGS (Strand Scientifics) software package. Quantification was accomplished using TMM normalization and further validated by comparing RNAseq results to those obtained for targeted genes using quantitative RT-PCR. Gene enrichment and functional analyses were accomplished using the Database for Annotation, Visualization and Integrated Discovery (DAVID) and Ingenuity Pathway Analysis (IPA). Uterine gene expression was affected (P < 0.05; corrected for false discovery rate) by both age and treatment. For nursed gilts, 3283 differential gene expression events were identified between birth and PND 2. By contrast, 4662 differential gene expression events were identified between birth and PND 2 for replacer-fed gilts. On PND 2, 896 lactocrine-sensitive genes were differentially expressed in nursed as compared to replacer-fed gilts. Thus, effects of both age and treatment (nursing vs replacer feeding) on uterine gene expression patterns were evident by PND 2. Based on DAVID analyses, the top up- and down-regulated genes affected by age and treatment included genes associated with immune response, cell adhesion, ion transport, and DNA packaging. IPA analyses revealed alterations in multiple signaling pathways associated with age and/or treatment, including those linked to the matrix metalloproteinase family of molecules and the estrogen receptor signaling cascade. Neonatal uterine transcriptomic analyses can be used to refine and focus hypotheses related to identification of cellular, molecular and lactocrine mechanisms regulating endometrial development. [Support: USDA-NIFA 2013-67016-20523.]