中国寄生虫学与寄生虫病杂志 ›› 2021, Vol. 39 ›› Issue (4): 443-448.doi: 10.12140/j.issn.1000-7423.2021.04.004

• 论著 • 上一篇    下一篇

三维适形调强放疗对大鼠继发性股骨细粒棘球蚴感染的疗效研究

周文正1(), 孙俊刚2, 赵喜滨2, 曹力1,*()   

  1. 1 新疆医科大学第一附属医院骨科中心,乌鲁木齐 830001
    2 新疆维吾尔自治区人民医院骨科,乌鲁木齐 830001
  • 收稿日期:2021-03-02 修回日期:2021-05-18 出版日期:2021-08-30 发布日期:2021-08-11
  • 通讯作者: 曹力
  • 作者简介:周文正(1978-)男,硕士研究生,从事骨棘球蚴病治疗的研究。E-mail: 1975255404@qq.com
  • 基金资助:
    新疆维吾尔自治区科技支疆项目(201391129);新疆维吾尔自治区自然科学基金项目(2012211B34)

Therapeutic effect of intensity modulated radiation therapy on secondary femur infection with Echinococcus granulosus in rats

ZHOU Wen-zheng1(), SUN Jun-gang2, ZHAO Xi-bin2, CAO Li1,*()   

  1. 1 Orthopedics Center, the First Affiliated Hospital of Xinjiang Medical University,Urumqi 830001, China
    2 Orthopedics Center, People’s Hospital of Xinjiang Uygur Autonomous Region, Urumqi 830001, China
  • Received:2021-03-02 Revised:2021-05-18 Online:2021-08-30 Published:2021-08-11
  • Contact: CAO Li
  • Supported by:
    Xinjiang Uygur Autonomous Region Science and Technology Support Project(201391129);Natural Science Foundation of Xinjiang Uygur Autonomous Region(2012211B34)

摘要:

目的 探讨三维适形调强放疗(IMRT)对大鼠继发性股骨细粒棘球蚴感染的疗效及对包囊的杀灭机制。方法 选取股骨继发性细粒棘球蚴感染Wistar雄性大鼠75只,其中50只大鼠为IMRT组,以IMRT方式40 Gy剂量照射左侧股骨棘球蚴病灶区,25只为对照组,不做照射。于放射治疗后1周,取大鼠左、右股骨细粒棘球蚴包囊,提取生发层细胞,采用Hoechst 33258荧光染色观察细胞核的形态变化,免疫组化法检测凋亡相关基因Caspase-3和p53的表达情况;ELISA检测IMRT组和对照组大鼠血清中γ干扰素(IFN-γ)、肿瘤坏死因子-α(TNF-α)、白细胞介素4(IL-4)和IL-10细胞因子水平差异;流式细胞术检测两组大鼠脾淋巴细胞CD4+/CD8+的比值。结果 IMRT治疗后1周,IMRT组大鼠血清中Th1细胞因子IFN-γ、TNF-α的吸光度(A490值)分别为322.45 ± 94.42、28.76 ± 6.23,均高于对照组的277.82 ± 65.48、19.63 ± 4.76(P < 0.05);Th2细胞因子IL-4的A490值为11.02 ± 2.55,低于对照组的13.22 ± 3.13(P < 0.05),IL-10的A490值为16.57 ± 3.02,与对照组(20.51 ± 3.17)的差异无统计学意义(P > 0.05)。Hoechst 33258荧光染色可观察到IMRT组大鼠左侧股骨细粒棘球蚴包囊的生发层出现细胞凋亡现象,而右侧生发层细胞正常。免疫组化检测结果显示,IMRT组大鼠左侧股骨细粒棘球蚴包囊的生发层细胞凋亡相关基因Caspase-3和p53大量表达,而右侧生发层细胞未出现。流式细胞术检测结果显示,IMRT组大鼠血清脾淋巴细胞 CD4+/CD8+的比值为2.96 ± 1.90,高于对照组的1.65 ± 1.31,差异有统计学意义(P < 0.05)。结论 IMRT可促进大鼠继发性股骨细粒棘球蚴包囊生发层细胞的凋亡,其作用机制一方面可能是通过调节凋亡相关蛋白Caspase-3和p53而实现,另一方面可能是与射线激发宿主对骨棘球蚴有效的免疫应答有关。

关键词: 细粒棘球蚴, 感染, 三维适形调强放疗, 股骨, 细胞凋亡, T淋巴细胞的亚群

Abstract:

Objective To observe the therapeutic effect of intensity modulated radiation therapy (IMRT) on secondary femur infection with Echinococcus granulosus in rats. Methods Seventy-five male Wistar rats with secondary femur infection of Echinococcus metacestode were used in this study. They were divided into the IMRT group (n = 50) and the control group (n = 25). In the IMRT group, the left thighbone of the rat was irradiated at irradiation dose of 40 Gy. The rats in the control group received no treatment. E. granulosus cysts were obtained from the left and right thighbones respectively, from which the germinal layer cells were extracted. Hoechst 33258 staining was performed to observe morphological changes of nuclei, and immunohistochemistry was conducted to detect the expression of apoptosis-related Caspase-3 and p53. ELISA was used to detect the difference of interferon γ (IFN-γ), tumor necrosis factor-α (TNF-α), interleukin-4 (IL-4) and IL-10 cytokine levels in the mice serum of IMRT group and control group. Flow cytometry was used to detect the CD4+/CD8+ ratio in splenic lymphocytes. Results One week after IMRT treatment, the absorbance (A490 value) of Th1 cytokines IFN-γ and TNF-α in the rat serum of the IMRT group were 322.45 ± 94.42 and 28.76 ± 6.23, respectively, which were higher than those of the control group (277.82 ± 65.48 and 19.63 ± 4.76) (P < 0.05). The A490 value of Th2 cytokine IL-4 was 11.02 ± 2.55, which was lower than that of the control group (13.22 ± 3.13) (P < 0.05). The IL-10 A490 value was 16.57 ± 3.02, which showed no statistical difference from the control group (20.51 ± 3.17) (P > 0.05). Hoechst 33258 fluorescent staining showed apoptosis of the germinal layer cells of the left femoral hydatid cysts of the IMRT group, while the right germinal layer cells were normal. The results of immunohistochemistry showed abundant expression of Caspase-3 and p53 in the germinal layer cells of the left femoral hydatid cyst of rats in the IMRT group, but not in the right germinal cells. The results of flow cytometry showed that the CD4+/CD8+ ratio in the IMRT group (2.96 ± 1.90) was much higher than that in the control group (1.65±1.31)(P < 0.05). Conclusion IMRT can induce apoptosis of the germinal layer cells of the femoral hydatid cyst in rats. The mechanisms may involve the regulation of Caspase-3 and p53, and the radiation triggering host immune responses to the parasites.

Key words: Echinococcus granulosus, Infection, Intensity modulated radiation therapy, Femur, Apoptosis, T lymphocyte subsets

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