Abstract:

Objective　To observe the effect of platelet-rich plasma (PRP) on the apoptosis of rat nucleus pulposus cells induced by interleukin-1β (IL-1β). Methods　This study was from July 2023 to October 2024. The PRP was extracted from the SD rats' blood by the two-step centrifugation method, and was used for the subsequent experiments. The rat nucleus pulposus cells were routinely cultured in vitro. The nucleus pulposus cells were treated by different concentrations of PRP for different periods of time, and the cell viability test kit was used to detect the nucleus pulposus cells' activity. The degeneration model of the nucleus pulposus cells was established by IL-1β induction. The cells were randomly divided into a control group (without the treatment of IL-1β and PRP), an IL-1β group (adding 10 μg/L lL-1β solution), and a PRP group (adding 10 μg/L IL-1β solution and PRP). After 72 hours' intervention, the expression levels of Bcl-2 associated X protein (Bax), B-cell lymphoma/leukemia 2 (Bcl-2), and cysteine aspartic protease-3 (Caspase-3) were detected by the Western blotting. The expressions of Bax, Bcl-2, and Caspase-3 mRNA were detected by the quantitative real-time polymerase chain reaction (qRT-PCR). One-way analysis of variance and LSD-t test were used to analyze the data. Results　The western blotting test results show that the expressions of Bax and Caspase-3 protein and the ratio of Bax/Bcl-2 in the IL-1β group were higher than those in the control group (2.43±0.22 vs. 1.04±0.07, 1.96±0.09 vs. 1.06±0.12, and 3.76±0.34 vs. 1.00±0.10), while the expression of Bcl-2 protein was lower (0.65±0.05 vs. 1.05±0.17), with statistical differences (all P<0.05). The expressions of Bax and Caspase-3 proteins and the ratio of Bax/Bcl-2 in the PRP group were higher than those in the IL-1β group, while the expression of Bcl-2 protein was lower, with statistical differences (all P<0.05). The results of qRT-PCR showed that the expressions of Bax and Caspase-3 mRNA in the IL-1β group were higher than those in the control group (1.15±0.12 vs. 1.00±0.03 and 1.22±0.09 vs. 1.00±0.11), while the expression of Bcl-2 mRNA was lower (0.51±0.05 vs. 1.00±0.04), with statistical differences (all P<0.05). The expressions of Bax and Caspase-3 mRNA in the PRP group were lower than those in the IL-1β group, while the expression of Bcl-2 mRNA were higher, with statistical differences (all P<0.05). Conclusions　PRP can inhibit the apoptosis and inflammatory response of nucleus pulposus cells induced by IL-1β, enhance cell activity, and effectively delay the progression of intervertebral disc degeneration.

Key words:

Platelet-rich plasma, Interleukin-1β, Inflammatory response, Apoptosis, Nucleus pulposus cells, Intervertebral disc degeneration, Animal experiment

摘要：

目的　探讨富血小板血浆（PRP）对白细胞介素-1β（IL-1β）诱导大鼠髓核细胞凋亡的影响。方法　研究时间为2023年7月至2024年10月。采取SD大鼠血液，使用两步离心法提取PRP，用于后续实验。常规培养大鼠髓核细胞。采用不同浓度PRP对髓核细胞进行不同时间点的处理，用细胞活力检测试剂盒检测髓核细胞的活性。通过IL-1β诱导建立髓核细胞退变模型，随机分为3组：对照组（未经IL-1β和PRP处理）、IL-1β组（加入10 μg/L IL-1β溶液）和PRP组（加入10 μg/L IL-1β溶液和PRP）。干预72 h后，Western blot检测髓核组织中Bcl-2家族相关x蛋白（Bax）、B细胞淋巴瘤/白血病2基因（Bcl-2）以及半胱氨酸天冬氨酸蛋白酶-3蛋白（Caspase-3）表达水平；实时荧光定量聚合酶链式反应（qRT-PCR）检测Bax、Bcl-2以及Caspase-3 mRNA的表达。采用单因素方差分析及LSD-t检验进行数据分析。结果　Western blot检测结果显示，IL-1β组的Bax和Caspase-3蛋白表达量、Bax/Bcl-2均高于对照组（2.43±0.22比1.04±0.07、1.96±0.09比1.06±0.12、3.76±0.34比1.00±0.10），Bcl-2蛋白表达量低于对照组（0.65±0.05比1.05±0.17），差异均有统计学意义（均P<0.05）。PRP组的Bax和Caspase-3蛋白表达、Bax/Bcl-2低于IL-1β组，Bcl-2蛋白表达量高于IL-1β组，差异均有统计学意义（均P<0.05）；qRT-PCR检测结果显示，IL-1β组的Bax和Caspase-3 mRNA表达量均高于对照组（1.15±0.12比1.00±0.03、1.22±0.09比1.00±0.11），Bcl-2 mRNA表达量低于对照组（0.51±0.05比1.00±0.04），差异均有统计学意义（均P<0.05）。PRP组的Bax和Caspase-3 mRNA表达量低于IL-1β组，Bcl-2 mRNA表达量高于IL-1β组，差异均有统计学意义（均P<0.05）。结论　PRP可能抑制IL-1β诱导的髓核细胞凋亡和炎症反应，增强细胞活性，有效延缓椎间盘退变疾病进展。

关键词:

富血小板血浆, 白细胞介素-1β, 炎症反应, 细胞凋亡, 髓核细胞, 椎间盘退变, 动物实验