目的:骨质疏松是世界范围内最常见的代谢性骨病。骨量的减少主要伴随成骨细胞数量和活性的减少。过氧化物酶(PRDXs)是检测极低过氧化物水平的蛋白质,并充当调节氧化信号的传感器,从而调节各种细胞功能。本研究旨在评价PRDX1和雌激素对成骨细胞生物学行为的影响,包括它们的增殖和分化。
方法:用去卵巢(OVX)小鼠建立骨质疏松模型,并进行形态学和免疫组织化学分析。使用细胞计数试剂盒-8,定量逆转录聚合酶链反应,在小鼠MC3T3-E1前成骨细胞中进行Prdx1基因敲除和过表达,以评估增殖和成骨分化。蛋白质印迹(WB),茜素红S染色,等。结果:OVX小鼠出现骨质疏松,PRDX1表达增加。体外实验表明,在成骨细胞成骨分化过程中,PRDX1表达降低,而COL1和RUNX2的表达增加。在Prdx1淘汰赛之后,成骨细胞增殖减少;Runx2、ALP、COL1增加;矿化增加。然而,Prdx1过表达后,成骨细胞增殖增强,而成骨分化和矿化被抑制。雌激素抑制H2O2诱导的成骨细胞分化减少和PRDX1表达增加。WB显示,当LY294002抑制AKT信号通路时,MC3T3-E1细胞中p-AKT1、p-P65和PRDX1蛋白水平降低。然而,当吡咯烷二硫代氨基甲酸酯(PDTC)抑制NF-κB信号通路时,p-AKT1和PRDX1的表达没有变化,除了p-P65表达显著降低.此外,PDTC逆转了RUNX2、ALP、和COL1由PRDX1过表达引起。
结论:PRDX1促进成骨细胞增殖,抑制成骨分化。雌激素通过影响成骨细胞中PRDX1的表达来调节成骨细胞的分化,其作用与AKT1/NF-κB信号通路有关。
OBJECTIVE: Osteoporosis is the most common metabolic bone disease worldwide. The decrease in bone mass is primarily accompanied by a decrease in the number and activity of osteoblasts. Peroxiredoxins (PRDXs) are proteins that detect extremely low peroxide levels and act as sensors that regulate oxidation signals, thereby regulating various cellular functions. This study aimed to evaluate the effects of PRDX1 and estrogen on the biological behavior of osteoblasts, including their proliferation and differentiation.
METHODS: Ovariectomized (OVX) mice were used to establish a model of osteoporosis and perform morphological and immunohistochemical analyses. Prdx1 gene knockout and overexpression were performed in mouse MC3T3-E1 pre-osteoblasts to assess proliferation and osteogenic differentiation using the cell counting kit-8, quantitative reverse transcription polymerase chain reaction, western blotting (WB), Alizarin Red S staining, etc. RESULTS: The OVX mice exhibited osteoporosis and PRDX1 expression increased. In vitro experiments showed that during the osteogenic differentiation of osteoblasts, PRDX1 expression decreased, while the expression of COL1 and RUNX2 increased. After Prdx1 knockout, the proliferation of osteoblasts decreased; expression of Runx2, ALP, and COL1 increased; and mineralization increased. However, after Prdx1 overexpression, osteoblast proliferation was enhanced, whereas osteogenic differentiation and mineralization were inhibited. Estrogen inhibits the H2O2-induced decrease in osteoblastic differentiation and increase in PRDX1 expression. WB revealed that when LY294002 inhibited the AKT signaling pathway, the levels of p-AKT1, p-P65, and PRDX1 protein in MC3T3-E1 cells decreased. However, when pyrrolidine dithiocarbamate (PDTC) inhibited the NF-κB signaling pathway, the expression of p-AKT1 and PRDX1 did not change except for a significant reduction of p-P65 expression. Furthermore, PDTC reversed the decreased expression of RUNX2, ALP, and COL1 caused by PRDX1 overexpression.
CONCLUSIONS: PRDX1 promotes the proliferation of osteoblasts and inhibits osteogenic differentiation. Estrogen regulated osteoblastic differentiation by affecting the expression of PRDX1 in osteoblasts, and the effect is related to the AKT1/NF-κB signaling pathway.