Establishment and evaluation of the primary cultured tibial osteoblast model of broiler chicks

Osteoblasts are considered as a major factor contributing to bone development and mineralization, however, few studies have been done to establish and evaluate the primary cultured tibial osteoblast model of broiler chicks. Therefore, in the present study, two experiments were conducted to establish and evaluate the primary cultured tibial osteoblast model of broiler chicks. In experiment 1, osteoblasts were isolated from the tibia of one-day-old Arbor Acre male broiler chicks using the explant method and identified through the cell morphology, alkaline phosphatase (ALP) and alizarin red staining. Experiment 2 was carried out to evaluate the vitality and mineralization of primary cultured tibial osteoblasts of broilers on days 4, 8, 12, 16, 20, 24, 28 and 32 after incubation, respectively. The results from experiment 1 demonstrated that primary cultured tibial osteoblasts of broilers showed a spindle-shaped, triangular or polygonal morphology. More than 95% of the cells were stained blue-black after ALP staining, and mineralized nodules were formed after 4 days of continuous incubation. in experiment 2, lactate dehydrogenase (LDH) activity stayed at a relatively stabilized level although incubation time affected (P=0.0012) it during the whole culture period. Additionally, incubation time affected (P≤0.0001) the number and proportion of the area of mineralized nodules. They increased linearly and quadratically (P<0.04) with the increase of incubation time, and remained at a stabilized level from 24 to 32 days of incubation. The estimates of the optimal incubation time were 17 and 26 days based on the best fitted broken-line or quadratic models (P<0.0001) of the number and proportion of the area of mineralized nodules, respectively. These results indicate that the primary cultured tibial osteoblast model of broilers has been established successfully by the explant method, and it showed typical osteoblast morphology and characteristics of ALP activity and mineralization, and could maintain a relatively stabilized vitality from 4 to 32 days of incubation; and the optimal incubation time of primary tibial osteoblasts was 17 to 26 days. Therefore, it could be used to further study the underlying mechanisms of bone development and mineralization of broiler chicks.