Foxp3在Treg介导的HPV16持续感染及宫颈癌免疫逃逸作用中的机制研究

The human papillomavirus (HPV) persistent infection is the major cause of cervical cancer. Immune evasion provides a microenvironment necessary for HPV persistent infection and cervical malformation. When the immune balance shifts towards immune evasion, it facilitates the HPV persistence and may permit the development of cervical cancer. Foxp3 mediated Treg (CD4 + CD25 + regulatory T cells) is an important factor for the immune evasion of HPV, neoplasia and cervical cancer. Our preliminary work suggests a positive correlation between Foxp3 expression and pathological progression of cervical lesion. However, it's mechanism on immune evasion of HPV persistent infection and cervical cancer is unclear. Our investigation based on previous work is designed to explore (1) the association between Treg activation with its activating molecular Foxp3 and HPV16 persistent infection; (2) the relationship between Treg activation with its activating molecular Foxp3 and clinical pathological process of cervical cancer caused by HPV16 persistent infection; (3) In vitro experiment based on CTL cell model and nude mice model, to prove the immune regulatory role of Treg activated by Foxp3 on HPV16 (+) cervical cancer cell function. (4) the different Foxp3 expression induced molecular changes of HPV16 E6 specific Treg (mainly by chip assay on mRNA and cytokines, ChIP-high-throughput sequencing on transcription factors and histone modification). By our investigation, we want to find out the exact effect and mechanism of Foxp3 activated Treg-mediated immune evasion on HPV16 persistent infection and the development of cervical cancer.

人乳头瘤病毒（HPV）持续感染是诱发宫颈癌的主要因素。免疫逃逸为HPV持续感染及肿瘤形成提供了必要微环境。Foxp3介导的Treg（CD4+CD25+调节性T细胞）是促成免疫逃逸的关键因素之一。前期工作提示，Foxp3的表达与宫颈癌病理进程呈正相关，但其介导的免疫逃逸对HPV持续感染及HPV持续感染导致宫颈癌的作用机制尚不清楚。本研究拟在前期工作基础上探讨（1）Treg活化标志分子Foxp3与HPV16持续感染的相关性；（2）Foxp3活化Treg与HPV16持续感染所致宫颈癌临床病理进程的相关性；（3）体外实验研究证实Foxp3活化Treg对HPV16(+)宫颈癌细胞功能的免疫调节作用；（4）研究Foxp3表达差异导致的HPV16 E6特异性Treg细胞模型中的分子改变。通过本研究阐明Foxp3 在Treg介导的HPV16持续感染及宫颈癌发生发展过程中的免疫逃逸作用与机制。

本课题通过检测不同宫颈病变组织中调节性T细胞( Treg)的表达及人乳头瘤病毒HPV基因组整合状态，研究Treg与人乳头瘤病毒整合状态在宫颈病变发病机制中的作用及意义。发现HPV整合型组Treg细胞多于游离型组，这说明Treg在HPV持续感染所致宫颈癌的发生及发展中可能起着重要作用。Treg可以通过抑制机体的特异性免疫作用，促进机体对肿瘤细胞的免疫耐受，帮助肿瘤免疫逃逸，促进肿瘤发生。Foxp3是Treg的功能基因和特异性标记物。课题组进一步研究发现，Foxp3除了在Tregs细胞中表达，还表达于宫颈癌组织。本课题应用免疫组化方法检测不同程度宫颈病变组织中Foxp3、p16INK4a、TLR4、B7-H4及HMGB1蛋白的表达，发现正常宫颈上皮组织、CIN和宫颈癌组织中Foxp3、p16INK4a、TLR4、B7-H4及HMGB1蛋白的表达逐渐增高，宫颈癌组织中Foxp3、p16INK4a、B7-H4的表达水平与宫颈癌的FIGO分期、淋巴结转移、肿瘤大小有关。HMGB1表达水平与宫颈癌的FIGO分期、淋巴结转移及肿瘤分化程度有关。TLR4的表达仅与临床FIGO分期和淋巴结转移情况密切相关。提示Foxp3、p16INK4a、B7-H4、TLR4及HMGB1的表达增高可能是宫颈癌发生发展的机制之一。Foxp3的阳性表达率还与HPV相关，提示Foxp3表达上调与宫颈癌的不良预后相关。我们进一步建立Foxp3沉默细胞系，分别检测沉默Foxp3之后宫颈癌细胞增殖能力、侵袭能力及凋亡的改变。发现沉默Foxp3基因能够显著抑制宫颈癌细胞增殖能力和侵袭能力，明显减少S、G2期的细胞数目，增加宫颈癌细胞的凋亡比率。进一步研究发现，CD4+CD25+Treg细胞可以特异性抑制CD8+CTL细胞杀伤Siha宫颈癌细胞的活性，这种抑制作用主要是通过CD4+CD25+Treg细胞与CD8+CTL 细胞的直接接触和CD4+CD25+Treg细胞分泌的细胞因子IL-10、TGF-β所起的作用。发表文章6篇，其中SCI 4篇，中文2篇。