[Kcnq1ot1 promotes osteogenic differentiation and suppresses osteoclast differentiation].

OBJECTIVE

To investigate the regulatory role of long non-coding RNA Kcnq1ot1 in osteoclast differentiation, osteogenic differentiation and osteoporosis.

METHODS

The expression of lnc-Kcnq1ot1, Bglap, Runx2, Alp, Bsp, Nfatc1, Mmp9, Ctsk and Oscar were detected by real-time quantitative PCR (qRT-PCR) in the femoral bones from mouse models of postmenopausal osteoporosis (ovariectomized mice, =8), disuse osteoporosis (induced by tail suspension, =14) and agerelated osteoporosis (18-month-old mice, =8), and also in MC3T3-E1 cells during osteoblast differentiation and in murine bone marrow-derived macrophages (BMMs) and RAW264.7 cells during osteoclast differentiation. MC3T3-E1 cells with lncKcnq1ot1 knockdown by lentivirus infection were induced to differentiate into osteoblasts using osteogenic induction medium, and the expression of lnc-Kcnq1ot1, Alp and Bglap was detected with qRT-PCR and ALP activity was assessed with ALP staining. BMMs and RAW264.7 cells were transfected with siRNAs targeting lnc-Kcnq1ot1 and stimulated with RANKL and/or M-CSF, and the expression of lnc-Kcnq1ot1, Ctsk and Oscar was detected by qRT-PCR, and TRAP activity was assessed by TRAP staining. The subcellular localization of lnc-Kcnq1ot1 in MC3T3-E1 and RAW264.7 cells was determined using cell fractionation followed by qRT-PCR.

RESULTS

The expression of lnc-Kcnq1ot1 was significantly upregulated during osteoblast differentiation but downregulated in the bone tissues of osteoporotic mice and during osteoclast differentiation ( < 0.05). Silencing lnc-Kcnq1ot1 obviously decreased the expression of Bglap and Alp ( < 0.05) and attenuated osteogenic medium-induced osteoblast differentiation. Knockdown of lnc-Kcnq1ot1 also promoted the expression of Ctsk and Oscar ( < 0.05) and aggravated RANKL-induced osteoclast differentiation. The results of cell fractionation and qRT-PCR demonstrated that lnc-Kcnq1ot1 was located mainly in the nuclei of MC3T3-E1 and RAW264.7 cells.

CONCLUSIONS

Our data demonstrate that lnc-Kcnq1ot1 promotes osteogenic differentiation and alleviates osteoclast differentiation, suggesting the potential of lnc-Kcnq1ot1 as a therapeutic target against osteoporosis.