脾多肽对化疗所致血小板减少症雌鼠的疗效及促血小板生成的机制研究

The therapeutic effect of lienal polypeptide on female mice model of chemotherapy-induced thrombocytopenia and the mechanism of thrombopoiesis

摘要:

目的 研究脾多肽注射液对化疗所致血小板减少症的作用及其血小板生成机制。方法 将85只健康昆明雌鼠随机分为正常组、模型组、重组人血小板生成素（rhTPO）阳性对照组、脾多肽注射液低剂量组、脾多肽注射液高剂量组。模型组、给药组第1天单次腹腔注射70 mg/ kg 卡铂复制血小板减少症模型，正常组注射生理盐水。第2天起，不同药物连续干预15 d，并于化疗前以及化疗后第2～16天隔天尾部取血计数血小板；化疗后第8天采用瑞氏-吉姆萨染色骨髓巨核细胞，显微镜观察其形态和数量，采用流式细胞术检测骨髓巨核细胞百分率，酶联免疫吸附试验检测血清干细胞因子（SCF）、促血小板生成素（TPO）水平。结果 各组雌鼠化疗前及化疗后2 d、4 d、6 d、8 d、10 d、12 d、14 d、16 d血小板计数比较，经重复测量设计的方差分析，结果 ①不同时间点血小板计数变化有差异（F =22.413，P =0.000）；②各组血小板计数有差异（F =6.822，P =0.006）；③各组血小板计数变化趋势有差异（F =6.326，P =0.008）。镜下观察巨核细胞细胞核及胞体较其他细胞大，且随着巨核细胞的成熟，核多为不规则形态，胞质愈丰富。细胞染色后，深紫色部分为核，浅紫色部分为胞质。模型组骨髓巨核细胞数量和百分率低于正常组（P <0.05），rhTPO阳性对照组高于模型组（P <0.05），脾多肽注射液高剂量组高于模型组（P <0.05）。模型组SCF、rhTPO水平高于正常组（P <0.05），脾多肽注射液高剂量组SCF水平低于模型组（P <0.05），rhTPO阳性对照组rhTPO水平低于模型组（P <0.05）。结论 60 mg/（kg·d）脾多肽注射液能有效改善卡铂化疗所致雌鼠血小板减少症，可能与调节血清SCF因子恢复至正常水平及上调骨髓巨核细胞数量有关。

Abstract:

Objective To reveal the effects of lienal polypeptide injection on thrombocytopenia induced by chemotherapy and the mechanism of thrombopoiesis.Methods A total of 85 healthy Kunming mice were randomly divided into normal group, model group, positive control (recombinant human thrombopoietin, rhTPO) group, and low-dose and high-dose lienal polypeptide group. The normal group was injected with normal saline, and the other groups were given a single dose of 70 mg/kg carboplatin via intraperitoneal injection to establish the chemotherapy-induced thrombocytopenia model. From the following day, different groups of mice were treated with corresponding drugs for 15 consecutive days. The blood was taken from the tail vein to determine the platelet count before chemotherapy and every other day from the 2nd to 16th day after chemotherapy. On the 8th day after chemotherapy, bone marrow megakaryocytes were observed and counted under the Wright-Giemsa stain. The percentages of megakaryocytes were detected by flow cytometry. The serum levels of stem cell factor (SCF) and thrombopoietin (TPO) were detected by enzyme-linked immunosorbent assay.Results The platelet count of female mice in each group before chemotherapy and 2 d, 4 d, 6 d, 8 d, 10 d, 12 d, 14 d, and 16 d after chemotherapy was compared and analyzed via the repeated measures ANOVA. The results demonstrated that there were differences in platelet count at different time points (F = 22.413, P = 0.000) and among the groups (F = 6.822, P = 0.006), and that there were differences in the change trends of platelet count among the groups (F = 6.326, P = 0.008). Under the microscope, megakaryocytes have larger nuclei and the size of megakaryocytes was greater than other cells. As megakaryocytes became mature, most of the nuclei were irregular in shape, and the cytoplasm was abundant. After staining, the dark purple part characterized the nucleus and the light purple part characterized the cytoplasm. The number and percentage of bone marrow megakaryocytes in the model group were lower than those of the normal group (P <0.05), and those in the positive control group and high-dose lienal polypeptide group were both higher compared with the model group (P < 0.05). The levels of SCF and rhTPO in the model group were higher than those in the normal group (P < 0.05), and the level of SCF in the high-dose lienal polypeptide group was lower than that in the model group (P < 0.05). Besides, the level of rhTPO in the positive control group was lower than that in the model group (P < 0.05).Conclusions Lienal polypeptide injection at a dose of 60 mg/(kg·d) is effective for ameliorating the thrombocytopenia induced by carboplatin chemotherapy in female mice, which may be related to the normalization of the serum level of SCF and the up-regulation of bone marrow megakaryocytes.