ILC2细胞经CCL27/CCR10信号对急性放射性损伤的调控作用研究

Acute radiation-induced skin injury (ARD) is the most common complication in the process of cancer radiotherapy. Group 2 of innate lymphoid cells (ILC2) are members of the large ILC family. It has been demonstrated that activation of ILC2 triggers immune response to epithelial damage, followed by leading to skin inflammation diseases. Additionally, one newly report showed that activation of ILC2 was correlation with wound healing of skin. On the other hand, CCL27, a member of the CC chemokine subfamily, is the ligand that specifically binds CC chemokine receptor CCR10. The binding of CCL27 and CCR10 (CCL27/CCR10 signaling) may attenuate skin inflammation through regulating lymphocytes to maintain equilibrium of skin immune. However, up to now, whether ILC2 affect the occurrence and severity of ARD and the effect of ICL2 on ARD by CCL27/CCR10 signaling still remain unclear. In this project, two transgene mice, including Rag1-/-C57BL/6 mice and CCR10+/EGFP C57BL/6 mice, will be used in the process of experiment. After local skin of mice is exposed to irradiation, the occurrence and severity of ARD will be observed. Activating factors to ILC2 and type 2 cytokines secreted from ILC2, meanwhile, are measured by ELISA and Western blot. Furthermore, activation and percentage of ILC2 are analyzed through flow cytometry in order to evaluate the relationship of ILC2 and ARD. As certificating influence of ILC2 on ARD, numbers and percentage of ILC2 are further analyzed along with antibody blocking activation of ILC2. And then ARD severity will be evaluated in local skin of irradiated mice and activating factors to ILC2 and type 2 cytokines will be measured as well. Subsequently, effect of CCL27/CCR10 signaling on ARD will be investigated. Rag1-/-C57BL/6 mice are crossed to CCR10+/EGFP C57BL/6 mice for the generation of Rag1-/-CCR10+/EGFP mice and Rag1-/-CCR10 EGFP/EGFP mice. Such two mice are irradiated to observe the occurrence and severity of ARD. Contents of CCL27 and relative cytokines in the irradiated local skin are measured; meanwhile, numbers and percentage of CCR10+ILC2 are analyzed. Consequently, it will be revealed that, in this project, the occurrence and severity of ARD may be regulated through CCL27/CCR10 signaling on ILC2. It may be available to protect normal tissue, especially skin, in the process of radiotherapy.

急性放射性皮肤损伤（ARD）是最常见的放疗并发症。研究表明，固有淋巴细胞ILC2与皮肤炎性疾病的发生密切相关，并调节皮肤创伤的愈合。CCL27/CCR10趋化信号可调节淋巴细胞在皮肤免疫中的平衡，降低炎性反应。本研究拟采用Rag1-/-C57BL/6和CCR10+/EGFPC57BL/6转基因鼠，进行局部照射，观察ARD的发生和分级，并测定ILC2的活化因子和分泌的2类细胞因子含量，通过流式细胞术分析ILC2的活化情况。同时，用抗体封闭ILC2表面受体阻止其活化后，观察ILC2变化对ARD的影响。进而对2种转基因鼠交配后的子代Rag1-/-CCR10+/EGFP和Rag1-/-CCR10 EGFP/EGFPC57BL/6鼠进行照射，检测皮肤内CCL27含量和ILC2上下游细胞因子的变化，分析CCR10+ILC2的活化情况，以了解ILC2细胞经CCL27/CCR10信号对ARD的调控作用。

急性放射性皮肤损伤（ARD）是最常见的放疗并发症。研究表明，固有淋巴细胞ILC2与皮肤炎性疾病的发生密切相关，并调节皮肤创伤的愈合。CCL27/CCR10趋化信号可调节淋巴细胞在皮肤免疫中的平衡，降低炎性反应。本研究以CCR10+/EGFPC57BL/6和CCR10EGFP/EGFPC57BL/6转基因鼠为研究对象，分别以5Gy单次照射和6×5Gy分割累计照射小鼠皮肤组织，观察照射后放射性皮肤损伤的发生和分级，采用ELISA方法检测小鼠皮肤和血液中CCL27，IL-1β和TNF-α的浓度，并同时取部分皮肤组织进行HE染色，观察照射组与未照射组的炎症程度。同时检测皮肤内CCL27含量和ILC2上下游细胞因子的变化，分析CCR10+ILC2的活化情况，以了解ILC2细胞经CCL27/CCR10信号对ARD的调控作用。构建了免疫联体小鼠模型，检测辐照对远端分泌的影响。同时，采集癌症放疗患者外周血，流式检测外周血中T淋巴细胞、B淋巴细胞、NK细胞和固有淋巴细胞放疗前后的变化水平。结果发现，CCR10-/-小鼠在照射后表现出更多的皮肤炎症和更严重的病理变化。脾脏和血液中不同基因型小鼠的ILCs无显着差异。单次照射能够诱导CCR10敲除小鼠分泌CCL27并同时促进IL-1β的升高。分割照射能够造成小鼠皮肤角质形成细胞受损，进而使其分泌CCL27减少，但IL-1β的分泌增加明显。在放疗患者外周血中，T淋巴细胞、B淋巴细胞、NK细胞在放疗后均显著降低，放射治疗后ILC2在ILCs中所占比例较放疗前升高。而ILC1和ILC3在放疗前后变化不明显。本研究结果表明ILC2可以通过CCR10调节放射性皮炎的发生发展，可能是降低放射性皮炎严重程度的潜在治疗靶点。