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山东大学学报（医学版） ›› 2017, Vol. 55 ›› Issue (9): 46-52.doi: 10.6040/j.issn.1671-7554.0.2017.030

• 基础医学 • 上一篇    下一篇

自噬对125I-anti-TLR5同种移植排斥靶向显像的影响

赵姗姗,李晓梅,梁婷,张超,宋静,侯桂华   

  1. 山东大学基础医学院生物医学同位素研究中心, 山东 济南 250012
  • 收稿日期:2017-01-10 出版日期:2017-09-10 发布日期:2017-09-10
  • 通讯作者: 侯桂华. E-mail:ghhou@sdu.edu.cn E-mail:ghhou@sdu.edu.cn
  • 基金资助:
    国家自然科学基金(81371601)

Impact of autophagy on targeting imaging of allorejection with 125I-anti-TLR5

ZHAO Shanshan, LI Xiaomei, LIANG Ting, ZHANG Chao, SONG Jing, HOU Guihua   

  1. Research Center of Biomedical Isotope, School of Basic Medicine, Shandong University, Jinan 250012, Shandong, China
  • Received:2017-01-10 Online:2017-09-10 Published:2017-09-10

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摘要： 目的 探讨自噬对放射性碘125标记抗Toll样受体5(125I-anti-TLR5)靶向同种移植排斥显像的影响及特点。 方法 建立小鼠皮肤同种移植模型,分为对照组(无药物处理)、雷帕霉素组、3-甲基腺嘌呤(3MA)组和联合组(雷帕霉素+3MA处理)。采用Iodogen法常规制备125I-anti-TLR5,测定其在生理盐水和血清中的稳定性;体外根据浓度不同将细胞分为对照组、雷帕霉素组(10 ng/mL)、3MA组(10 mmol/L)和联合组(10 ng/mL雷帕霉素+10 mmol/L3MA),研究125I-anti-TLR5与处理细胞的结合与解离;在移植后9 d,在小鼠同种皮肤移植模型尾静脉注射125I-anti-TLR5,72 h后进行生物学分布研究,24、48、72 h进行全身磷屏自显影显像,分析T/NT比值(靶与非靶比值);采用病理及免疫组化染色法分析移植皮片Beclin1/TLR5的表达。 结果 125I-anti-TLR5标记率高,稳定性好。与雷帕霉素组相比,联合组细胞与125I-anti-TLR5结合率与解离率无明显改变。注射125I-anti-TLR5后72 h,标记物主要经肝肾代谢。移植皮片放射性浓聚,雷帕霉素组和联合组T/NT 比值显著高于对照组(P<0.001, P<0.001)。全身动态磷屏放射自显影显像显示,各组48 h移植皮片即可显像,其中雷帕霉素组移植皮片放射性浓聚最明显(P<0.001);雷帕霉素组和联合组72 h显像明显,放射性活度比均显著高于对照组(P<0.001)。移植后12 d对照组移植皮片炎症细胞浸润明显,而雷帕霉素组与联合组炎性浸润减少;对照组及雷帕霉素组自噬分子Beclin1表达增高,而联合组无明显降低;雷帕霉素组及联合组TLR5 表达无显著变化。 结论 自噬抑制对雷帕霉素作用下TLR5表达及125I-anti-TLR5靶向同种移植排斥显像无明显影响,125I-anti-TLR5可用于免疫耐受状态下同种移植排斥显像。

关键词: 放射性碘125, Toll样受体5, 自噬, 磷屏自显影显像, 同种移植

Abstract: Objective To evaluate the effect of autophagy on targeting imaging of allotransplanted skin graft by 125I-anti-TLR5. Methods Mouse alloskin transplantation models were established and divided into 4 groups: control group, rapamycin group, 3-Methyladenine(3MA)group, and rapamycin+3MA group. Anti-TLR5 was labeled with 125I by Iodogen method, and stability in normal saline and serum was determined respectively. Spleen cells were divided into control group, rapamycin group(10 ng/mL), 3MA group(10 mmol/L)and rapamycin+3MA group(10 ng/mL rapamycin+10 mmol/L 3MA). Then the uptake and dissociation of 125I-anti-TLR5 were studied. 125I-anti-TLR5 was injected to transplantation models through tail veins on day 9 after allotransplantation. Biodistribution at 72 h after 山 东 大 学 学 报 (医 学 版)55卷9期 -赵姗姗,等.自噬对125I-anti-TLR5同种移植排斥靶向显像的影响 \=-125I-anti-TLR5 injection and whole-body phosphor-autoradiography at 24 h, 48 h, and 72 h were performed. T/NT ratio(target to non-target)was analyzed. Beclin1/TLR5 expression was detected with histological and immunohistochemical staining. Results 125I-anti-TLR5 was successfully obtained with good labeling rate and stability. The uptake and dissociation of tracer were not apparently changed in rapamycin+3MA group compared with rapamycin group. Biodistribution studies showed that 125I-anti-TLR5 was mainly metabolized through the liver and kidney, and high radioactivity in grafted skin was detected. The T/NT ratio in rapamycin group and rapamycin+3MA group was significantly higher than that in control group(P<0.001, P<0.001). Whole body phosphor-screen autoradioimaging showed clear skin grafts radioactivity-imaging in rapamycin group at 48 h compared with the control group(P<0.001). Radioactivity activity ratio of rapamycin group and rapamycin+3MA group remarkably increased(P<0.001)at 72 h. Allograft histology assay showed that acute inflammatory infiltration was decreased in rapamycin group and rapamycin+3MA group. The expression of Beclin1 was not significantly decreased in rapamycin+3MA group compared with the control group and rapamycin group. TLR5 expression was not significantly changed in rapamycin group and rapamycin+3MA group. Conclusion Autophagy has no significant impact on TLR5 expression and 125I-anti-TLR5 targeted allograft rejection imaging under rapamycin application and 125I-anti-TLR5 could be used for allograft imaging under rapamycin-induced tolerance.

Key words: Toll like receptor 5, Iodine 125, Autophagy, Phosphor auto-radio imaging, Allotransplantation

中图分类号: 

  • R817.33
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