α-半乳糖神经酰胺活化肺iNKT细胞诱导气道免疫耐受及其机制探讨

Invariant natural killer T (iNKT) cells display substantial functional heterogeneity, and there is growing evidence, particularly in mouse models of asthma, showing that interactions between iNKT cells and regulatory T cells (Treg cells) play an important role in induction of tolerance in many different settings to lead to the prevention of diverse disease conditions such as autoimmunity, type 1 diabetes and transplant rejection. In our previous study, α-galactosylceramide (α-GalCer) administration via intraperitoneal injection can activate iNKT cells in the lung from normal wild-type BALB/c mice, but the activation of iNKT cells alone cannot induce airway inflammation without ovalbumin immunization and challenge. Meanwhile, the number of CD4+FoxP+Treg cells significantly increase in the lung, as accompanied with the activation of lung iNKT cells by intraperitoneal injection of α-GalCer in normal wild-type mice. In addition, it was found that intraperitoneal injection of α-GalCer can not results in increase of the lung CD4+FoxP+Treg cells in CD1d knockout-BALB/c mice, which lack iNKT cells, compared with wild-type BALB/c mice. It is well known that Treg cells play a vital role in inducing and maintaining airway immune tolerance, and thereby preventing Th2 response. As a result, it was hypothesized that α-GalCer-activated lung iNKT cells generate CD4+Foxp3+ regulatory T cells to induce airway tolerance. In our preliminary study, α-GalCer-activated lung iNKT cells can significantly enhance the concentration of interleukin-2 (IL-2) in bronchial alveolar lavage fluid (BALF) and the expression of IL-2 mRNA in the lung tissue from wild-type BALB/c mice, compared with CD1d knockout-BALB/c mice. IL-2 can drive the development of CD4+FoxP3+ regulatory T cells, which have suppresser function and mediate immune tolerance. As such, it was hypothesized that α-GalCer-activated lung iNKT cells generate CD4+Foxp3+ regulatory T cells through interleukin-2. For the purpose, wild-type BALB/c mice, CD1d knockout-BALB/c mice, Jα18 knockout-C57BL/6 mice, CNS1 knockout-BALB/c mice, IL-2 knockout-BALB/c mice and humanized mice of immune reconstitution using severe combined immunodeficiency (SCID) mice were used as experiment animals to investigate that α-GalCer-activated lung iNKT cells generate CD4+Foxp3+ regulatory T cells to induce airway tolerance through interleukin-2. Therefore, a better understanding of this immunoregulatory network will add to the ability to optimize iNKT cell-based therapeutics.

课题组发现α-半乳糖神经酰胺（α-GalCer）活化肺恒定自然杀伤（iNK）T细胞不诱导野生型小鼠产生气道炎症，但小鼠肺CD4+FoxP3+T细胞数量增加，同时α-GalCer不诱导缺乏iNKT细胞的CD1d KO小鼠肺CD4+FoxP3+T细胞数量增加。由于调节性T细胞可以诱导气道免疫耐受，防止机体发生Th2反应，故假设α-GalCer活化肺iNKT细胞通过诱导调节性T细胞介导气道免疫耐受。课题组初步发现α-GalCer活化肺iNKT细胞时，小鼠肺IL-2表达水平增加。由于IL-2可以诱导产生调节性T细胞，故假设α-GalCer活化肺iNKT细胞通过IL-2诱导机体产生调节性T细胞。为此，以野生型小鼠，CD1d、Jα18、CNS1 和IL-2 KO小鼠及人源化免疫重建SCID小鼠为实验动物，探讨α-半乳糖神经酰胺活化肺iNKT细胞是否通过IL-2诱导机体产生调节性T细胞介导气道免疫耐受。

课题组发现α-半乳糖神经酰胺（α-GalCer）活化肺恒定自然杀伤（iNK）T细胞不诱导野生型小鼠产生气道炎症，但小鼠肺CD4+FoxP3+T细胞数量增加；α-GalCer活化肺iNKT细胞时，小鼠肺IL-2表达水平增加。由于调节性T细胞可以诱导气道免疫耐受，防止机体发生Th2反应，而IL-2可以诱导产生调节性T细胞，故假设α-GalCer活化肺iNKT细胞通过IL-2诱导机体产生调节性T细胞，进一步诱导气道免疫耐受。为此，探讨：(1) α-半乳糖神经酰胺活化肺iNKT细胞是否诱导小鼠发生气道免疫耐受；(2) α-半乳糖神经酰胺活化肺 iNKT 细胞诱导小鼠发生气道免疫耐受是否依赖肺调节性 T 细胞的产生；(3) α-半乳糖神经酰胺活化肺iNKT细胞诱导小鼠产生Treg细胞是否依赖白介素-2。结果发现：(1)腹腔注射α-半乳糖神经酰胺可以诱导无致敏原致敏和激发的野生型BALB/c小鼠肺调节性T细胞数量明显增加，这与α-半乳糖神经酰胺活化肺iNKT细胞产生白介素-2有关；(2)过敏原致敏前腹腔注射α-半乳糖神经酰胺活化肺iNKT细胞产生白介素-2，进一步诱导肺调节性T细胞数量增加和功能增强，从而抑制哮喘小鼠肺树突状细胞免疫原性成熟和Th2反应。本课题探索α-半乳糖神经酰胺活化肺 iNKT细胞通过白介素-2诱导产生调节性T细胞，进一步介导小鼠气道免疫耐受的形成。本课题发现α-半乳糖神经酰胺活化iNKT细胞通过白介素-2诱导产生调节性T细胞，进一步介导小鼠气道免疫耐受的形成，提出诱导气道免疫耐受的一条免疫调节途径。α-半乳糖神经酰胺可以活化小鼠和人类iNKT细胞，小鼠和人类使用时耐受性好，因此该课题提出α-半乳糖神经酰胺活化肺 iNKT细胞诱导气道免疫耐受及其机制可以为寻找人类支气管哮喘治疗的新靶点提供科学依据。

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