Ontogenic gene expression profiles in pig hepatogenesis
Liver is a key organ required for development and growth, whether fetal or post partum. Selection for greater litter size in swine has resulted in increased variability in piglet weight/litter, and greater weight piglets thrive better postnatally. To better understand hepatogenesis, comprehensive ontogenic gene expression profiling was performed to determine baseline expression patterns associated with liver development. Liver was collected from the greatest weight fetus of four distinct unilaterally hysterectomized:ovariectimized gilts at prenatal day (PND) 37, 50, 70, and 110. Total RNA was isolated, mRNA amplified and subjected to microarray analysis (Agilent). Three comparisons were performed: PND50 vs. PND37, PND70 vs. PND50 and PND110 vs. PND70; only genes whose expression, between at least two time points, was ± 1.5 fold with P < 0.05 were characterized further. A total of 6061 annotated genes exhibited altered expression: 648 down- and 341 up-regulated at PND37 vs. PND50; 666 down- and 1399 up-regulated at PND70 vs. PND50; and 1564 down- and 1443 up-regulated at PND110 vs. PND70. Thirty-five transcripts were selected for validation by absolute quantitative PCR; they clustered into five functional categories: i) hematopoietic and early liver function, ii) extracellular matrix, iii) hepatocyte function, iv) biliary function, and v) transcription factors. To examine changes elicited following parturition, liver was collected from greatest weight piglets at post partum day (PPD) 1 (n=3) and 2 (n=4) and analyzed along with fetal samples. The expression of hematopoietic genes (e.g. coproporphyrinogen oxidase) was high at PND37 and declined by mid-gestation. Similarly, the expression of markers of undifferentiated hepatoblasts (e.g. Cbp/P300-Interacting Transactivator 1) was high in early gestation and decreased below the detection threshold post partum. In contrast, the abundance of extracellular matrix genes (e.g. hevin) peaked perinatally. Hepatocyte serum or enzyme transcripts increased gradually with a maximum induction at PPD2 vs. PND37 (e.g alcohol dehydrogenase, a 2266-fold increase). Likewise, the expression of inhibin beta B, involved in biliary duct morphogenesis, increased 188-fold. Transcription factors (e.g. hepatocyte nuclear factors) exhibited small variation during gestation but were significantly elevated perinatally. More impressive was the up-regulation (79-fold) of Kruppel-like factor 9 at and beyond PND 110 vs. earlier time points. The study identified commonalities and differences of expression profiles to those observed in other species; this should help scientists navigate new routes of investigation in liver cell function in swine. Future studies will examine dysregulation of these genes in runt or slow growing pigs.
Keywords: Liver, Development, Pig