补肾生血药对化疗早期骨髓细胞的影响

Effect of Bushen Shengxue Decoction on bone marrow cells in the early stage of chemotherapy

目的 观察补肾生血药对化疗早期骨髓细胞的影响。方法 选取65只SPF级6~8周龄体重(20±2)g雄性BALB/c小鼠,按随机数字表法将其分为空白组、模型组、阳性对照组、补肾生血药高剂量组、补肾生血药低剂量组,每组13只。预防给药5 d,补肾生血药高(47.0 g/kg)、低(23.5 g/kg)剂量组采用补肾生血药灌胃,其余组灌胃等量蒸馏水。预防给药结束后,模型组、阳性对照组、补肾生血药高及低剂量组均腹腔注射环磷酰胺(100 mg/kg),空白组腹腔注射等量生理盐水,连续3 d。同时进行干预用药,阳性对照组皮下注射重组人粒细胞集落刺激因子(30μg/kg),其余组与预防给药阶段相同,连续5 d。干预结束后,收集各组骨髓细胞,观察细胞超微结构,检测骨髓中肿瘤坏死因子-α(TNF-α)、转化生长因子-β(TGF-β)、γ干扰素(IFN-γ)含量,检测骨髓细胞JAK激酶2(JAK2)、信号转导及转录激活因子5 (STAT5)的mRNA及JAK2、STAT5及磷酸化STAT5 (P-STAT5)蛋白表达情况。结果 空白组骨髓有核细胞分布密集,核膜、胞膜完整,细胞器结构清晰。模型组有核细胞稀少,见大量细胞碎片,其余各组有核细胞数不同程度增多,胞体减小,细胞器尚好。模型组TNF-α、TGF-β、IFN-γ含量高于空白组,补肾生血药高、低剂量组TNF-α、TGF-β、IFN-γ含量低于模型组,补肾生血药高剂量组TGF-β、IFN-γ含量低于补肾生血药低剂量组,差异均有统计学意义(P <0.05或P <0.01)。与空白组比较,模型组JAK2、STAT5 m RNA表达下调;与模型组比较,补肾生血药高、低剂量组STAT5 mRNA表达上调;与补肾生血药低剂量组比较,补肾生血药高剂量组JAK2 mRNA表达下调,差异均有统计学意义(P <0.05或P <0.01)。模型组JAK2、P-STAT5、STAT5蛋白表达水平低于空白组,补肾生血药高、低剂量组JAK2、P-STAT5蛋白表达水平高于模型组,补肾生血药高剂量组P-STAT5蛋白表达水平低于补肾生血药低剂量组,差异均有统计学意义(P <0.05或P <0.01)。结论 补肾生血药可减轻化疗早期骨髓细胞损伤,其机制与降低造血负调控因子含量,正调控JAK2-STAT5通路有关。

Objective To observe the effect of Bushen Shengxue Decoction on bone marrow cells in the early stage of chemotherapy. Methods Sixty-five male SPF class BALB/c mice aged six to eight weeks with body weight of(20±2) g were selected and divided into blank group, model group, positive control group, high dose Bushen Shengxue Decoction group, and low dose Bushen Shengxue Decoction group according to random number table method, with 13 mice in each group. For five days of preventive administration, the high(47.0 g/kg) and low(23.5 g/kg) dose Bushen Shengxue Decoction groups were given intragastric administration, while the other groups were given intragastric administration with equal amount of distilled water. After preventive administration, Cyclophosphamide(100 mg/kg) was intraperitoneally injected into model group, positive control group, high and low dose Bushen Shengxue Decoction groups, and the same amount of normal saline was intraperitoneally injected into blank group for consecutive three days. At the same time, the positive control group was subcutaneously injected with recombinant human granulocyte colony stimulating factor(30 μg/kg), and the other groups were given the same prophylactic administration for consecutive five days. After the intervention, bone marrow cells of each group were collected, and ultrastructure of the cells was observed. The contents of tumor necrosis factor-α(TNF-α), transforming growth factor-β(TGF-β), and interferon-γ(IFN-γ) in bone marrow were detected. The mRNA expression of JAK kinase 2(JAK2), signal transduction and transcription activator 5(STAT5), and the protein expression of JAK2, STAT5, and phosphorylated STAT5(P-STAT5)in bone marrow cells were detected. Results In blank group, the distribution of nucleated cells was dense, the nuclear membrane and cell membrane were intact, and the organelle structure was clear. There were few nuclear cells in the model group and a large number of cell fragments. The number of nuclear cells in the other groups increased to varying degrees, while the cell body decreased and the organelles were good. The contents of TNF-α, TGF-β, and IFN-γ in the model group were higher than those in the blank group, the contents of TNF-α, TGF-β, and IFN-γ in the high and low dose Bushen Shengxue Decoction groups were lower than those in the model group, and the contents of TGF-β and IFN-γ in the high dose Bushen Shengxue Decoction group were lower than those in the low dose Bushen Shengxue Decoction group, the differences were all statistically significant(P < 0.05 or P < 0.01). Compared with blank group, the m RNA expression of JAK2 and STAT5 in model group was down-regulated. Compared with model group, STAT5 mRNA expression in the high and low dose Bushen Shengxue Decoction groups was up-regulated. Compared with the low dose Bushen Shengxue Decoction group, the expression of JAK2 mRNA in the high dose Bushen Shengxue Decoction group was down-regulated, the differences were all statistically significant(P < 0.05 or P < 0.01). The expression levels of JAK2, P-STAT5, and STAT5 protein in the model group were lower than those in the blank group, the expression levels of JAK2 and P-STAT5 protein in the high and low dose Bushen Shengxue Decoction groups were higher than those in the model group, the expression level of P-STAT5 protein in the high dose Bushen Shengxue Decoction group was lower than that in the low dose Bushen Shengxue Decoction group, the differences were statistically significant(P < 0.05 or P <0.01). Conclusion Bushen Shengxue Decoction can alleviate bone marrow cell damage in the early stage of chemotherapy, and the mechanism is related to the reduction of hematopoietic negative regulatory factor content and positive regulation of JAK2-STAT5 pathway.