芍药苷调节LKB1-AMPK信号通路对LPS诱导的牙周膜干细胞凋亡和成骨分化的影响

doi:10.3969/j.issn.1006-6233.2023.01.001

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Abstract Objective: To investigate the influences of paeoniflorin (PF) on lipopolysaccharide (LPS)-induced apoptosis and osteogenic differentiation of periodontal ligament stem cells (PDLSCs) by regulating liver kinase B1 (LKB1)-adenylate-activated protein kinase (AMPK) signaling pathway. Methods: PDLSCs in logarithmic growth phase were collected and separated into control group, LPS group (10μg/mL), LPS+PF low concentration (PF-L, 10μg/mL LPS+5μmol/L PF) group, LPS+PF medium concentration (PF-M, 10μg/mL LPS+10μmol/L PF) group, LPS+PF high concentration (PF-H, 10μg/mL LPS+20μmol/L PF) group, and LPS+PF-H +LKB1 inhibitor (radicicol, 10μg/mL LPS+20μmol/L PF+10μmol/L radicicol) group. ELISA was used to detect the levels of inflammatory factors [interleukin (IL)-1β, tumor necrosis factor-α (TNF-α), IL-6] in cells; alizarin red staining was used to observe the changes of mineralized nodules; alkaline phosphatase (ALP) was used to detect the cell osteogenic differentiation; cell apoptosis was detected by flow cytometry; the expression levels of osteogenic genes (OPN, RUNX2 and OCN) were detected by qRT-PCR; Western blot was used to detect the expression levels of LKB1-AMPK pathway protein and apoptosis protein (caspase-1). Results: Compared with the control group, the mineralized nodules were reduced in the LPS group, the apoptosis rate, the expression of caspase-1, the contents of TNF-α, IL-1β and IL-6 increased obviously, the level of ALP, the expression of OPN, RUNX2 and OCN, and the phosphorylation levels of LKB1 and AMPK decreased obviously (P<0.05); compared with the LPS group, the mineralized nodules were reduced in the LPS+PF-L group, the LPS+PF-M group, and the LPS+PF-H group, the apoptosis rate, the expression of caspase-1, the contents of TNF-α, IL-1β and IL-6 decreased obviously, the level of ALP, the expression of OPN, RUNX2 and OCN, and the phosphorylation levels of LKB1 and AMPK increased obviously, all were dose-dependent (P<0.05); however, radicicol reversed the effects of PF-H on LPS-induced apoptosis and osteogenic differentiation of PDLSCs (P<0.05). Conclusion: PF inhibits LPS-induced apoptosis of PDLSCs and promotes osteogenic differentiation by up-regulating LKB1-AMPK signaling pathway.

Key words： Paeoniflorin LKB1-AMPK signaling pathway Lipopolysaccharide Periodontal ligament stem cells Apoptosis Osteogenic differentiation

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http://www.hbyxzzs.cn/EN/10.3969/j.issn.1006-6233.2023.01.001 OR http://www.hbyxzzs.cn/EN/Y2023/V29/I1/1

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