Fig 1.
High glucose concentration promoted osteoblast differentiation and increased histone lactylation levels in C2C12 cells.
C2C12 cells were incubated for 72 (a right, b, c) or 24 (a left, d) hours after addition of BMP-2 (150 ng/ml) into medium with a high (4500 mg/L) or low (900 mg/L) glucose concentration. Amounts of (a) lactate, and (b) ALP staining and activity are shown. (c) Osteoblast differentiation marker genes were determined using real-time RT-PCR. (d) Expression of lactyl lysine after addition of BMP-2 was evaluated by western blotting. Data are expressed as mean ± SD (n = 3–5). *, **Significantly different from control group (*p <0.05, **p <0.01).
Fig 2.
Inhibition of LDHA reduced osteoblast differentiation and lactyl lysine levels.
C2C12 cells were incubated for 72 (a right, b, c) or 24 (a left, d) hours after addition of BMP-2 with oxamate. Amounts of (a) lactate, and (b) ALP staining and activity are shown. (c) Osteoblast differentiation marker genes were determined using real-time RT-PCR. (d) Expression of lactyl lysine after addition of BMP-2 and oxamate was evaluated by western blotting. Data are expressed as mean ± SD (n = 3). *, **Significantly different from control group (*p <0.05, **p <0.01). NS, not significant.
Fig 3.
Lactate recovered suppression of osteoblast differentiation and decreased histone lactylation levels caused by low glucose.
C2C12 cells were incubated for 72 (a right, b, c) or 24 (a left, d) hours after placing in medium with a high or low glucose concentration in the absence or presence of lactate. Amounts of (a) lactate, and (b) ALP staining and activity are shown. (c) Osteoblast differentiation marker genes were determined using real-time RT-PCR. (d) Expression of lactyl lysine under the same conditions was evaluated by western blotting. Data are expressed as mean ± SD (n = 3–4). *, **Significantly different from control group (*p <0.05, **p <0.01). ND: not detected.
Fig 4.
Ep300 siRNA suppressed osteoblast differentiation and decreased histone lactylation levels.
Following introduction of Ep300 siRNA, C2C12 cells were cultured for 72 (a right, b, c) or 24 (a left, d) hours after addition of BMP-2 to the medium. Amounts of (a) lactate, and (b) ALP staining and activity are shown. (c) Osteoblast differentiation marker genes were determined using real-time RT-PCR. (d) Expression of lactyl lysine under the same conditions was evaluated by western blotting. Data are expressed as mean ± SD (n = 3–5). *, **Significantly different from control group (*p <0.05, **p <0.01). NS, not significant.