目的:探讨髓母细胞瘤(MB)中4种亚型CD8+T细胞浸润的特点,分析CD8+T细胞浸润与预后的关系,研究C-X-C基序趋化因子配体11(CXCL11)及其受体在CD8+T细胞浸润肿瘤中的作用,为探索MB的免疫治疗提供必要的临床病理依据。
方法:在研究中,从2012年至2019年的多个医学中心选取48例临床MB样本(4种亚型各12例)。在NanoStringPanCancerIO360TM面板(NanoStringTechnologies)上进行48个临床样品的肿瘤的转录组学分析。福尔马林固定的免疫组织化学(IHC)染色,使用CD8一级抗体对MB的石蜡包埋切片进行分析,以分析MB四种亚型中CD8+T细胞的差异量。通过生物信息学分析,研究CD8+T细胞浸润与患者预后的关系以及不同亚型MB中各种趋化因子的表达差异。通过双重免疫荧光染色验证MB中CD8+T细胞表面CXCR3受体的表达,探讨了CD8+T细胞浸润肿瘤的分子机制。
结果:WNT亚型MB中CD8+T细胞的特征指数相对较高,提示WNT亚型中CD8+T细胞的数量明显高于其他三种亚型,通过使用R2在线数据分析平台,通过CD8免疫组织化学染色和基因表达Omnibus(GEO)数据库分析证实了这一点。CD8+T细胞浸润的增加与患者生存率呈正相关。CXCL11在WNT亚型MB中的表达程度显著高于其他三种亚型。免疫荧光染色显示CXCL11受体的存在,CXCR3在CD8+T细胞表面,提示CD8+T细胞可能被CXCL11通过CXCR3吸引到MB微环境。
结论:与其他亚型相比,CD8+T细胞在WNT亚型MB中浸润更多。其机制可能与CXCL11-CXCR3趋化因子系统的激活有关,肿瘤中CD8+T细胞浸润较多的患者预后较好。这一发现可能为MB中CD8+T细胞浸润的调控机制提供必要的临床病理基础,并为未来MB的免疫治疗提供了新的潜在治疗靶点。
OBJECTIVE: To investigate the characteristics of the CD8+ T cells infiltration from the 4 subtypes in
medulloblastoma (MB), to analyze the relationship between CD8+ T cells infiltration and prognosis, to study the function of C-X-C motif chemokine ligand 11 (CXCL11) and its receptor in CD8+ T cells infiltration into tumors and to explore the potential mechanism, and to provide the necessary clinicopathological basis for exploring the immunotherapy of MB.
METHODS: In the study, 48 clinical MB samples (12 cases in each of 4 subtypes) were selected from the multiple medical center from 2012 to 2019. The transcriptomics analysis for the tumor of 48 clinical samples was conducted on the NanoString PanCancer IO360TM Panel (NanoString Technologies). Immunohistochemistry (IHC) staining of formalin-fixed, paraffin-embedded sections from MB was carried out using CD8 primary antibody to analyze diffe-rential quantities of CD8+ T cells in the MB four subtypes. Through bioinformatics analysis, the relationship between CD8+T cells infiltration and prognosis of the patients and the expression differences of various chemokines in the different subtypes of MB were investigated. The expression of CXCR3 receptor on the surface of CD8+T cells in MB was verified by double immunofluorescence staining, and the underlying molecular mechanism of CD8+T cells infiltration into the tumor was explored.
RESULTS: The characteristic index of CD8+T cells in the WNT subtype of MB was relatively high, suggesting that the number of CD8+T cells in the WNT subtype was significantly higher than that in the other three subtypes, which was confirmed by CD8 immunohistochemical staining and Gene Expression Omnibus (GEO) database analysis by using R2 online data analysis platform. And the increase of CD8+T cells infiltration was positively correlated with the patient survival. The expression level of CXCL11 in the WNT subtype MB was significantly higher than that of the other three subtypes. Immunofluorescence staining showed the presence of CXCL11 receptor, CXCR3, on the surface of CD8+T cells, suggesting that the CD8+T cells might be attracted to the MB microenvironment by CXCL11 through CXCR3.
CONCLUSIONS: The CD8+T cells infiltrate more in the WNT subtype MB than other subtypes. The mechanism may be related to the activation of CXCL11-CXCR3 chemokine system, and the patients with more infiltration of CD8+T cells in tumor have better prognosis. This finding may provide the necessary clinicopathological basis for the regulatory mechanism of CD8+T cells infiltration in MB, and give a new potential therapeutic target for the future immunotherapy of MB.