Establishment of co-culture model of mouse osteoblasts and osteoclasts

Objective To establish a co-culture system of mouse osteoblasts and osteoclasts for further investigating the interaction function of them.Methods Primary osteoblasts and osteoclasts are obtained respectively from the calvarias of newborn mice and long bones(femur and tibia)of adult mice(4-8 weeks old).An indirect contact co-culture system is set up by inserting a glass sheet sticking bone marrow mononuclear phagocytes into culture dish which had inoculated osteoblasts for 24h.The resistant acid phosphatase(TRAP)staining and bone resorption essay were used to identify the osteoclasts after 3 days of co-culture.The marked genes of osteoclast,MMP-9,TRAP and Cathepsin K,were detected by RT-PCR.Results The TRAP positive multinuclear cells emerged after 5 days of co-culture and the number of TRAP positive multinuclear cells reached the maximum after 13 days.Bone resorption essay indicated that resorption pits formed on bone slices after 7 days of co-culture and the area of pits gradually increased with the period of co-culture extending.The marked gene of osteoclast of TRAP expressed after 3 days of co-culture,but the MMP-9 and Cathepsin K were detected after 5 days.Conclusions The co-culture system can induce osteoclasts which degraded bone actively.This model can provide foundation for the research on the interaction function of osteoblasts and osteoclasts.