Expression of RUNX2/LAPTM5 in MC3T3-E1 osteoblastic cells with induced mineralization

Abstract

To investigate the association of the expressions of RUNX2/LAPTM5 with osteogenesis and lysosomes in osteoblastic cells during mineralization induction. MC3T3- E1 cells cultured in osteogenic induction medium was examined for mineralization and osteogenic differentiation using Alizarin red staining and alkaline phosphatase (ALP) staining, respectively. RT-qPCR and Western blotting were used to detect the mRNA and protein expressions of Runx2 and LAPTM5 in the cells during osteogenic induction for 5 days. The effects of overexpression and interference of RUNX2/ LAPTM5 on the expressions of ALP and osteocalcin (OCN) in the cells were examined with Western blotting. MC3T3- E1 cells cultured in osteogenic induction medium showed an increased number of mineralized nodules over time, and the size of the mineralized nodules increased as the culture time extended; the number of purple-blue granules stained by ALP also increased gradually with time. RT-qPCR and Western blotting showed that the expressions of RUNX2 and LAPTM5 in the cells increased progressively during osteogenic mineralization (P < 0.001). Overexpression and interference of RUNX2 obviously affected LAPTM5 expression in the cells (P < 0.05); modulation of LAPTM5 expression did not significantly affect RUNX2 expression but caused significant changes in ALP and OCN expressions (P < 0.01). RUNX2 /LAPTM5 may participate in the regulation of osteoblast differentiation, and RUNX2 may be involved in the regulation of LAPTM5 expression. RUNX2 /LAPTM5 may play a mediating role in the process of osteogenic mineralization involving lysosomes.