Abstract：
BACKGROUND: This study determines the role and mechanism of APS in cyclophosphamide-induced myelosuppression in mice and bone mesenchymal stem cells (BMSCs) cell model.
METHODS: Cy-induced myelosuppression mice and BMSCs cell model were established. Fifty C57BL/6 mice (weighing 20 ± 2 g) were randomly divided into five groups. Femur and tibia samples, bone marrow samples, and blood samples were collected 3 days after the last injection of Cy. Histopathology changes and cell apoptosis were detected. Cell viability, apoptosis, cycle distribution, reactive oxygen species activity, osteogenesis ability, and protein levels were detected. γ-H2AX and senescence-associated β-galactosidase activity expression was detected by immunofluorescence. Cy-induced senescence and Wnt/β-catenin related protein levels were detected using western blotting.
RESULTS: The results showed that APS effectively induced Cy-induced histological injury and cell apoptosis rate. After treated with APS, ROS and ALP levels were significantly increased. In BMSCs, cell viability, apoptosis, and cell cycle distribution were also influenced by APS treatment. Compared with the control group, cell viability was significantly increased, the cell apoptosis rate was decreased while the number of cells remained in the G0-G1 phase was increased. Meanwhile, ROS levels were significantly increased in APS group. Cell senescence and Wnt/β-catenin related protein (γ-H2AX, SA-β-gal, p21, p16, p-β-catenin/ β-catenin, c-Myc, and AXIN2) levels were also altered both in vivo and in vitro. Interestingly, the effects of APS were reversed by BML-284.
CONCLUSIONS: Our results indicate that APS protected Cy-induced myelosuppression through the Wnt/β-catenin pathway and APS is a potential therapeutic drug for Cy-induced myelosuppression.
METHODS: Cy-induced myelosuppression mice and BMSCs cell model were established. Fifty C57BL/6 mice (weighing 20 ± 2 g) were randomly divided into five groups. Femur and tibia samples, bone marrow samples, and blood samples were collected 3 days after the last injection of Cy. Histopathology changes and cell apoptosis were detected. Cell viability, apoptosis, cycle distribution, reactive oxygen species activity, osteogenesis ability, and protein levels were detected. γ-H2AX and senescence-associated β-galactosidase activity expression was detected by immunofluorescence. Cy-induced senescence and Wnt/β-catenin related protein levels were detected using western blotting.
RESULTS: The results showed that APS effectively induced Cy-induced histological injury and cell apoptosis rate. After treated with APS, ROS and ALP levels were significantly increased. In BMSCs, cell viability, apoptosis, and cell cycle distribution were also influenced by APS treatment. Compared with the control group, cell viability was significantly increased, the cell apoptosis rate was decreased while the number of cells remained in the G0-G1 phase was increased. Meanwhile, ROS levels were significantly increased in APS group. Cell senescence and Wnt/β-catenin related protein (γ-H2AX, SA-β-gal, p21, p16, p-β-catenin/ β-catenin, c-Myc, and AXIN2) levels were also altered both in vivo and in vitro. Interestingly, the effects of APS were reversed by BML-284.
CONCLUSIONS: Our results indicate that APS protected Cy-induced myelosuppression through the Wnt/β-catenin pathway and APS is a potential therapeutic drug for Cy-induced myelosuppression.
摘要：
背景：这项研究确定了APS在环磷酰胺诱导的小鼠骨髓抑制和骨髓间充质干细胞（BMSCs）细胞模型中的作用和机制。
方法：建立Cy诱导的骨髓抑制小鼠和BMSCs细胞模型。将50只C57BL/6小鼠(体重20±2g)随机分为5组。股骨和胫骨样本,骨髓样本,最后一次注射Cy后3天收集血样。检测组织病理学改变和细胞凋亡。细胞活力，凋亡，周期分布,活性氧活性，成骨能力，并检测到蛋白质水平。免疫荧光法检测γ-H2AX和衰老相关β-半乳糖苷酶活性表达。使用蛋白质印迹法检测Cy诱导的衰老和Wnt/β-catenin相关蛋白水平。
结果：结果表明，APS可有效诱导Cy诱导的组织学损伤和细胞凋亡率。用APS治疗后，ROS和ALP水平显著升高。在BMSCs中,细胞活力，凋亡，和细胞周期分布也受APS处理的影响。与对照组相比,细胞活力显著增加，细胞凋亡率降低，而保持在G0-G1期的细胞数量增加。同时,APS组ROS水平显著升高。细胞衰老与Wnt/β-catenin相关蛋白（γ-H2AX，SA-β-gal,p21，p16，p-β-连环蛋白/β-连环蛋白，c-Myc,和AXIN2）水平在体内和体外也发生了变化。有趣的是,APS的作用被BML-284逆转。
结论：我们的结果表明，APS通过Wnt/β-catenin通路保护Cy诱导的骨髓抑制，并且APS是Cy诱导的骨髓抑制的潜在治疗药物。
方法：建立Cy诱导的骨髓抑制小鼠和BMSCs细胞模型。将50只C57BL/6小鼠(体重20±2g)随机分为5组。股骨和胫骨样本,骨髓样本,最后一次注射Cy后3天收集血样。检测组织病理学改变和细胞凋亡。细胞活力，凋亡，周期分布,活性氧活性，成骨能力，并检测到蛋白质水平。免疫荧光法检测γ-H2AX和衰老相关β-半乳糖苷酶活性表达。使用蛋白质印迹法检测Cy诱导的衰老和Wnt/β-catenin相关蛋白水平。
结果：结果表明，APS可有效诱导Cy诱导的组织学损伤和细胞凋亡率。用APS治疗后，ROS和ALP水平显著升高。在BMSCs中,细胞活力，凋亡，和细胞周期分布也受APS处理的影响。与对照组相比,细胞活力显著增加，细胞凋亡率降低，而保持在G0-G1期的细胞数量增加。同时,APS组ROS水平显著升高。细胞衰老与Wnt/β-catenin相关蛋白（γ-H2AX，SA-β-gal,p21，p16，p-β-连环蛋白/β-连环蛋白，c-Myc,和AXIN2）水平在体内和体外也发生了变化。有趣的是,APS的作用被BML-284逆转。
结论：我们的结果表明，APS通过Wnt/β-catenin通路保护Cy诱导的骨髓抑制，并且APS是Cy诱导的骨髓抑制的潜在治疗药物。
