Dietary Calcium and Phosphate Levels Affect Bone Development and Marrow Adipose Tissue Deposition in Neonatal Pigs

A deficit in bone development during early-life has been associated with increased likelihood of developing osteoporosis later in life. Dietary calcium and phosphate play a pivotal role in bone development, which is largely orchestrated by the cellular activities: osteogenesis, ostoclastogenesis and adipogenesis. This study aims to determine the impact of different concentrations of dietary calcium and phosphate supplementation on bone development and mesenchymal stem cell (MSC) programming. Twenty-seven newborn neonatal pigs (n= 27, 24±6h) were randomly assigned into three groups and received a milk replacer diet for 16 days. Diets were isonitrogenous, isocaloric, but had varying concentrations of calcium and phosphate. The diets were designed to be either a 25% deficient, adequate, or 25% in excess (NRC standard) of Ca and phosphate requirements. Bone mineral content, serum mineral and hormone concentrations and activity of mesenchymal stem cells were determined. Serum phosphate concentrations were significantly higher in pigs receiving the adequate diet compared to those fed the deficient diet on day 8, but lower among pigs fed the excess diet compared to those fed the deficient diet on day 16 (P<0.05). Serum FGF23 concentrations were greater among the pigs fed the deficient diet than those fed the excess diet on day 16 (77.21±1.90 pg/mL vs. 70.38±1.90 pg/mL, P<0.05). Pigs receiving the diet containing excess Ca and phosphate had significantly higher bone ash and bone volume as compared to the deficient group. We determined in vivo proliferation of the MSC with BrdU, and there was greater cell proliferation in pigs fed the adequate diet than in those fed the deficient diet (8.9%±0.7 vs. 6.3%±0.7, P<0.05). Calcium and phosphate deficiency shifted MSC towards adipogenic differentiation, as indicated by both Oil Red O staining and increased gene expression of key adipogenic factors such as PPARγ2, FABP4, and LPL. Meanwhile, osteogenesis in the deficient group was reduced as compared to adequate group, which was illustrated by lower expression of master osteogenic regulator osteocalcin. This study demonstrates the importance of even minor deficiencies in neonatal calcium and phosphate intake and their impact on bone mineral acquisition and in shaping MSC activity and lineage allocation.