Research Paper Volume 13, Issue 10 pp 14355—14371

κ-opioid receptor stimulation alleviates rat vascular smooth muscle cell calcification via PFKFB3-lactate signaling

Effects of κ-OR stimulation on the expression of osteogenic proteins in VSMCs treated with lactate. (A) Representative blot images of osteogenic differentiation-associated proteins. (B) Quantitative analysis of RUN2 protein expression using densitometry. (C) Quantitative analysis of BMP2 protein expression using densitometry. (D) Quantitative analysis of SM22a protein expression using densitometry. (E) Calcium nodules were stained with Alizarin red. Red nodules indicate calcium deposition. Scale bar = 50 μm. (F) Quantification of mineralization (mean ± SEM; n = 6). (G) ALP activity was detected using ALP activity kits. Data were normalized using log10 and analyzed using one-way ANOVA tests; U, U50,488H; N, nor-BNI; Data obtained from quantitative densitometry were presented as means ± SEM. n=5 in each group. *P **P #P ##P $P $$P

Figure 5. Effects of κ-OR stimulation on the expression of osteogenic proteins in VSMCs treated with lactate. (A) Representative blot images of osteogenic differentiation-associated proteins. (B) Quantitative analysis of RUN2 protein expression using densitometry. (C) Quantitative analysis of BMP2 protein expression using densitometry. (D) Quantitative analysis of SM22a protein expression using densitometry. (E) Calcium nodules were stained with Alizarin red. Red nodules indicate calcium deposition. Scale bar = 50 μm. (F) Quantification of mineralization (mean ± SEM; n = 6). (G) ALP activity was detected using ALP activity kits. Data were normalized using log10 and analyzed using one-way ANOVA tests; U, U50,488H; N, nor-BNI; Data obtained from quantitative densitometry were presented as means ± SEM. n=5 in each group. *P < 0.05 versus the control group, **P < 0.01 versus the control group, #P < 0.05 versus the β-GP group, ##P < 0.01 versus the β-GP group, $P < 0.05 versus the β-GP+U group, $$P < 0.01 versus the β-GP+U group.