Deer antler peptide modulation of the SLC7A11/GPX4 axis inhibits dexamethasone-induced ferroptosis in osteoblasts

doi:10.12307/2025.619

Abstract

Abstract: BACKGROUND: Steroid-induced osteonecrosis of the femoral head is closely related to ferroptosis in osteoblasts. Deer antler peptide can promote the survival and functional establishment of osteoclasts by inhibiting ferroptosis in osteoblasts, and has the potential to treat steroid-induced osteonecrosis of the femoral head, but its regulatory mechanism of ferroptosis in osteoblasts has not yet been clarified.
OBJECTIVE: To investigate the mechanism by which deer antler peptide inhibits dexamethasone-induced ferroptosis in osteoblasts.
METHODS: (1) Different concentration gradients of antler peptide and dexamethasone were used to intervene in MC3T3-E1 14 cells, and the cell activity was detected by cell counting kit-8 method to determine the effect concentration of antler peptide and dexamethasone. (2) MC3T3-E1 14 cells treated with dexamethasone (800 μmol/L) were intervened with different concentrations of gradient antler polypeptide, which were then divided into blank control group, dexamethasone group and dexamethasone+antler peptide group. Cell counting kit-8 method was used to calculate the effects of different concentrations of antler polypeptide on the proliferation of MC3T3-E1 14 cells. (3) Glutathione, superoxide dismutase, malondialdehyde, lipid peroxide, cellular iron, and reactive oxygen species levels in the blank control group, dexamethasone group and dexamethasone+antler peptide group were detected using kits. The protein expressions of glutathione peroxidase 4 and solute carrier family 7 member 11 were detected by western blot to verify the pathway by which antler polypeptide inhibits ferroptosis.
RESULTS AND CONCIUSION: After cell activity was detected by cell counting kit-8 assay, antler peptide (10 mg/mL) and dexamethasone (800 μmol/L) were selected to treat MC3T3-E1 14 cells for 24 hours in subsequent experiments. After treatment with dexamethasone, malondialdehyde, lipid peroxide, cellular iron and reactive oxygen species levels were all increased (P < 0.01), while glutathione content and superoxide dismutase activity were decreased and the protein expression of glutathione peroxidase 4 and solute carrier family 7 member 11 were also decreased (P < 0.05-0.01). After antler peptide intervention, the changes in the above indexes were obviously reversed (P < 0.05-0.01). To conclude, antler peptide may inhibit ferroptosis in osteoblasts by regulating the glutathione peroxidase 4/solute carrier family 7 member 11 axis, and thereby exert a therapeutic role in steroid-induced osteonecrosis of the femoral head.

中国组织工程研究杂志出版内容重点：组织构建；骨细胞；软骨细胞；细胞培养；成纤维细胞；血管内皮细胞；骨质疏松；组织工程

Key words: antler peptide, dexamethasone, steroid-induced osteonecrosis of the femoral head, inhibition, osteoblast, ferroptosis

CLC Number:

Shao Xuekun, Wang Cheng, Wang Yi, Wang Ping, Qiu Zhuoya, Wang Xinru, Sun Tiefeng. Deer antler peptide modulation of the SLC7A11/GPX4 axis inhibits dexamethasone-induced ferroptosis in osteoblasts[J]. Chinese Journal of Tissue Engineering Research, 2025, 29(14): 2875-2881.

Figures/Tables 4

2.1 确定鹿角多肽和地塞米松干预 MC3T3-E1 14细胞的浓度通过 CCK-8 实验探究鹿角多肽和地塞米松干预 MC3T3-E1 14细胞的适宜浓度：①鹿角多肽浓度筛选结果显示：大于 5 mg/mL的鹿角多肽干预 MC3T3-E1 14细胞，24 h后细胞形态及密度与对照组(0 mg/mL鹿角多肽)相比差异有显著性意义(P < 0.05)，见图1A。②地塞米松浓度筛选结果显示：大于800 μmol/L的地塞米松干预MC3T3-E1 14细胞后，可以明显看出细胞增殖被抑制，与对照组(0 μmol/L地塞米松)相比差异有显著性意义(P < 0.01)，见图1B。因此，后续实验选择以 800 μmol/L浓度的地塞米松干预MC3T3-E1 14细胞并处理 24 h。 2.2 不同浓度梯度鹿角多肽对地塞米松(800 μmol/L)干预后 MC3T3-E1 14细胞的影响与空白对照组相比，经地塞米松单独处理的地塞米松组细胞活性显著被抑制，随着加入鹿角多肽的质量浓度增加，细胞存活率随之上升，10 mg/mL的鹿角多肽增殖率最为明显，且与空白对照组相比差异有显著性意义(P < 0.01)；20 mg/mL的鹿角多肽较10 mg/mL的鹿角多肽相比增殖率有所下降。经此筛选，确定后续实验选择以鹿角多肽(10 mg/mL)和地塞米松 (800 μmol/L)干预MC3T3-E1 14细胞处理24 h。见图2。 2.3 鹿角多肽和地塞米松干预 MC3T3-E1 14细胞后对铁死亡相关指标的影响为探究鹿角多肽对地塞米松处理过的细胞相关铁死亡的影响，对铁死亡相关指标进行了验证。与空白对照组相比，地塞米松组的谷胱甘肽含量及超氧化物歧化酶活性显著降低，地塞米松+鹿角多肽组的谷胱甘肽含量及超氧化物歧化酶活性相比地塞米松组有所升高；与空白对照组相比，地塞米松组的丙二醛、细胞铁、脂质过氧化物含量显著增加，地塞米松+鹿角多肽组的丙二醛、细胞铁、脂质过氧化物含量相比地塞米松组明显降低。见图3。 2.4 鹿角多肽和地塞米松干预 MC3T3-E1 14细胞后对活性氧的影响倒置荧光显微镜结果显示，与空白对照组相比，地塞米松组细胞内的活性氧增加，促进了细胞铁死亡；地塞米松+鹿角多肽组细胞内的活性氧有所降低，细胞铁死亡有所减少，见图4。 2.5 Western blot 结果鹿角多肽可以增加细胞内铁死亡标记物GPX4的表达水平，Western blot结果显示，鹿角多肽可以逆转GPX4和SLC7A11的下降，提示鹿角多肽可能通过SLC7A11/GPX4通路发挥对激素性股骨头坏死的治疗作用，见图5。"

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