Effects of l-Tryptophan and 1,25-Dihydroxycholecalciferol on Proliferation and Osteogenic Differentiation of Mesenchymal Stem Cells Isolated from the Compact Bones of Broilers and Layers.

ABSTRACT

Poultry is vulnerable to bone problems throughout their lives or production period due to rapid growth in broilers and the active laying cycle in layers. The calcium-sensing receptor (CaSR) is important in calcium and bone metabolism. The objective of this study was to investigate the effect of the CaSR ligand (l-Trp) and 1,25-dihydroxycholecalciferol (1,25OHD3) on the regulation of proliferation and osteogenic differentiation of chicken mesenchymal stem cells (MSCs) isolated from the compact bones of 14-day-old Ross 308 chickens and Dekalb pullets, which can provide cell-based evidence for the prevention or alleviation of skeletal disorders in the poultry industry. First, the dose- (0, 0.5, 1, 2, 5, 10, and 15 mM) and time-effects (0, 7, and 14 days) of l-Trp on the proliferation and osteogenic differentiation in chicken MSCs were investigated. The 5 mM l-Trp had a balanced effect between proliferation and osteogenic differentiation in broiler and layer MSCs when differentiated for 7 days. The broiler and layer MSCs were then treated with (1) osteogenic medium, osteogenic medium supplemented with (2) 1 nM 1,25OHD3, (3) 2.5 mM Ca2+, (4) 2.5 mM Ca2+ + 5 mM l-Trp and (5) 2.5 mM Ca2+ + 5 mM l-Trp + 1 µM NPS-2143, separately for 7 days. Results showed that the 5 mM l-Trp significantly inhibited the proliferation of broiler and layer MSCs on day 7 (P < 0.05), but 1 nM 1,25OHD3 significantly promoted the proliferation of layer MSCs (P < 0.05). Only the 2.5 mM Ca2+ + 5 mM l-Trp group significantly increased the mineralization process during osteogenic differentiation (P < 0.05), and this treatment also significantly upregulated the mRNA expression of the vitamin D receptor (VDR), ß-catenin, and osteogenesis genes in broiler MSCs (P < 0.05). The osteogenic differentiation process in layer MSCs was faster than that in broiler MSCs. In layer MSCs, Ca2+ alone significantly facilitated mineralization and ALP activity after 7-day osteogenic differentiation (P < 0.05). However, the 5 mM l-Trp significantly inhibited the differentiation and mineralization process by downregulating the mRNA expression of CaSR, VDR, ß-catenin, and osteogenic genes (P < 0.05) in layer MSCs. Taken together, l-Trp and 1,25OHD3 can regulate proliferation and osteogenic differentiation in both broiler and layer MSCs depending on the dose, treatment time, and cell proliferation and differentiation stages.