低氧诱导的滋养细胞自噬对DCs生物物理特性及其功能的影响与机制

Pregnancy is a complicated and fine physiological process. The appropriate modulation of numerous biological phenomena involved in implantation and placentation during the early pregnancy is pivotal to a successful pregnancy . Hypoxia is one of the most unique characteristics in the early pregnancy microenvironment, and trophoblastic cells, which give rise to responses directly to hypoxia and play great roles as "educated cells" in mediating other immune cells, such as DCs. But, the mechanisms how hypoxia affects trophoblastic cells and further modulates the differentiation and function of DCs should be elucidated to clarify local physiological properties. Previous researches have confirmed that some stress conditions such as the exhaust oxygen can induce autophagia which participates in a lot of physiological and pathological processes. In this study, we will exploit GFP-LC3-transfected trophoblast line HTR-8/SVneo, and study the effects and mechamisms of hypoxia on autophagia of trophoblastic cells, then further explore the effect of trophoblast cells autophagia change on the differentiation and functions of DCs from biological and physics characteristics. These studies will further broaden views in implantation and explain the regulations and related mechanisms about differentiation and functions of DCs from a new perspective,which are hopeful to provide novel molecular targets and strategies based on the preparations and applications of DCs vaccines.

妊娠是机体一个复杂的生理过程。尤其是孕早期，在胚泡植入及胎盘形成过程中涉及了诸多受精细调控的生物学现象，从而维持妊娠的成功。低氧张力是妊娠早期特征性微环境之一，滋养细胞是低氧感受的直接"应答"细胞，也是调控其他免疫细胞如DCs的重要"驯化"细胞。但低氧如何调控滋养细胞进而影响DCs分化与功能目前尚不清楚。文献已证实细胞在氧耗竭等应激条件下可诱导自噬，并可通过自噬参与调控机体的许多生理及病理生理过程。本项目拟利用GFP-LC3重组质粒稳定转染的人滋养细胞系HTR-8/SVneo细胞，研究低氧对其自噬的影响及机制，并进一步探讨滋养层细胞自噬改变后，对DCs细胞生物物理特性的影响及调控DCs分化及功能的机制。研究结果将进一步丰富妊娠植入过程的机制；并可拓宽免疫学研究领域的视角，从细胞生物物理特性方面解析DCs的分化及其功能的调控与机制，将为基于DCs的疫苗制备及应用提供新的靶点。

母体对半同种异体胎儿的免疫耐受是成功妊娠的关键，母-胎界面是胚胎和母体直接接触的前沿，是母-胎免疫调节网络的核心。母-胎界面微环境主要由胎儿来源的滋养细胞、母体来源的蜕膜基质细胞、蜕膜腺上皮细胞以及蜕膜免疫细胞所组成。低氧张力是妊娠早期特征性微环境之一，滋养细胞是低氧感受的直接“应答”细胞，但低氧如何调控滋养细胞进而影响母胎界面免疫细胞的分化与功能目前尚不清楚。此外，研究表明自噬对于妊娠早期的胚胎发育至关重要。本研究首次探讨了低氧对人滋养层细胞自噬的影响，结果发现低氧显著诱导滋养层细胞系HTR-8/SVneo自噬发生。我们利用基因表达谱芯片、自噬相关PCR-Array分析了低氧调节的差异基因表达，利用GO、聚类及Pathway分析技术绘制自噬关键调控网络图。我们重点探讨了PTEN在调节滋养层细胞自噬当中的作用，发现低氧通过PTEN抑制PI3K/Akt/mTOR通路进而促进自噬发生。另外，我们通过测序技术对低氧及自噬抑制剂3-MA处理的HTR-8/SVneo的全基因组及miRNA进行了测序分析。通过将滋养层细胞与外周血CD14+单核细胞共培养，我们发现滋养层细胞抑制单核细胞向树突状细胞（DCs）的分化和成熟，诱导产生类似蜕膜DCs的表型（DC-SIGN+CD14+CD1a−），HTR-8共培养的DCs诱导T细胞增殖、活化的能力显著降低，但可显著促进Treg的分化。此外，我们的研究发现滋养细胞与外周血CD14+单核细胞共培养，可通过MCP-1诱导其向髓样抑制细胞（MDSCs）分化。我们的研究为进一步理解妊娠早期的生理及病理生理现象，寻找新的病理性妊娠的防治策略提供思路，为揭示母胎界面髓系细胞的分化和功能及母胎界面免疫耐受的调控机制提供新的思路。