Transcriptional Regulation of Genes Involved in Calcium and Phosphate Metabolism in Neonatal Pigs Fed with Different Levels of Dietary Calcium and Phosphate

Dietary calcium and phosphate intake during the neonatal period may be of critical importance for lifetime skeletal development. In older animals, calcium and phosphate homeostasis is orchestrated by intestinal absorption, bone turnover, and kidney excretion, which are regulated by several hormones such as parathyroid hormone (PTH), calcitonin, fibroblast growth factor 23 (FGF23) and calcitriol [1,25(OH)₂D₃]. In older animals, the PTH/calcitriol system appears to be the major regulator of calcium and phosphate homeostasis, however, in neonates this system is not fully developed. We hypothesized that dietary calcium and phosphate concentrations would affect the expression of genes involved in calcium and phosphate homeostasis in neonatal pigs. Twenty-seven neonatal pigs (24±6h old) were randomly assigned into one of three treatment groups. Groups received a milk replacer diet for 16d that contained calcium and phosphate at a 25% deficiency, adequate, or 25% excess (NRC standard). At the completion of the feeding trial, samples of liver and kidney were obtained for total RNA isolation. Gene expression of vitamin D receptor (VDR), parathyroid hormone receptor (PTHR), type II sodium-dependent phosphate cotransporter (NPT2), vitamin D-activating enzyme 1α-hydroxylase (Cyp27B1), and 1,25-dihydroxy vitamin D3-24-hydroxylase (Cyp24A1) were determined by Real-Time PCR. Data, adjusted by housekeeping gene ribosomal protein L4 (RPL4), were normalized back to the deficient group, and then analyzed by ANOVA using the Glimmix procedure of SAS 9.4. Both PTHR and NPT2 were significantly higher in the calcium and phosphate deficient group than the adequate group (PTHR:1.01±0.07 vs 0.75±0.08; NPT2:1.01±0.09 vs 0.65±0.09; P<0.05). No significant differences were detected between the excess group and any other two groups. Similarly, expression of Cyp27B1 was significantly higher in the deficient group than both adequate and excess groups (Deficient: 1.10±0.13; Adequate: 0.60±0.13; Excess 0.55±0.13). These results provide some clues into the homeostatic regulation of calcium and phosphate in neonates. Based on these results, even a small deficiency in dietary calcium and phosphate intake can initiate transcriptional changes of key regulatory gene pathways to maintain homeostatic conditions.