人脐血源基质细胞经SDF-1/CD31通路阻抑ITP小鼠B淋巴细胞活性的研究

At present, there is no consistent treatment for refractory or relapsed ITP. We have found that human umbilical cord blood-derived stromal cells (hUCBDSCs), can recover the number of platelets in ITP mice . However，the potential mechanism need to be explored. In the preliminary study, the hUCBDSCs can secret higher level of SDF-1 than the bone marrow stromal cells from the ITP mice. Secondly, the expression of CD31 in B cells from ITP is increased after cocultured with hUCBDSCs. We hypothesized that hUCBDSCs can secret SDF-1, regulate the expression of CD31 in B cells from the ITP mice suppress the activity of B cells. In this schedule, the ITP mice model and a co-culturing model of hUCBDSCs and B cells will be established. The level of SDF-1 secreted by hUCBDSCs, the level of CD31 expressed by B cells, the proliferation of B cells, secreted platelet-associated antibody and related molecular expression will be detected. At the same time, the gain-of-function and the loss-of-function technologies will be used to verify the hypothesis. This study would supply theoretical basis for ITP treatment with hUCBDSCs, emphasisly from the angle of the cellular immunity .

难治复发性ITP的治疗，临床尚无统一意见。我们前期发现，运用人脐血源基质细胞hUCBDSCs可以促进ITP小鼠模型体内血小板的恢复，具体机制尚不清楚。进一步实验显示hUCBDSCs以分泌高水平SDF-1为特征，可上调共培养ITP中B细胞CD31的表达。结合文献，提出“高表达SDF-1人脐血源基质细胞经CD31调控ITP中B淋巴细胞活性”的假设。据此，本课题拟构建ITP小鼠模型，构建ITP-B淋巴细胞/hUCBDSCs transwell培养模型，检测hUCBDSCs分泌表达SDF-1的水平；检测共培养后B淋巴细胞CD31的表达以及B细胞增殖、分泌PAIgA，IgM，IgG等抗体表达情况；通过敲低SDF-1，CD31检验SHP，CD38，CD98hc等相关分子表达，进一步验证该假设。本研究着重从免疫调节的角度阐释“hUCBDSCs促进血小板恢复”这一理论，为难治复发性ITP的治疗提供新思路。

本课题假设高表达SDF-1 人脐血源基质细胞经CD31 抑制B 细胞活性，最终促进ITP巨核细胞成熟，减少血小板破坏。研究的主要内容包括观察人脐血源基质细胞通过SDF-1/ CD31 这条信号通路来调控ITP 中B 淋巴细胞的增殖、分化，抑制B 细胞免疫反应，减少ITP 血小板抗体的形成，促进ITP 小鼠血小板的恢复。希望使用细胞治疗方法为ITP 的治疗提供新的思路，同时也从免疫调节的角度为“hUCBDSCs促进血小板恢复”这一细胞治疗手段提供理论依据和实验基础。到目前为止，本课题已经按照计划完成大部分实验，现将本课题重要结果及进展汇报如下：.1. 完成了人脐源基质细胞的分离，培养，扩增(hUCBDSCs，缩写hU)。.2. 成功构建了ITP小鼠模型。.3. 人脐血源基质细胞回输能够促进ITP小鼠血小板的恢复。.4. 完成了小鼠B淋巴细胞分离培养.5．B细胞/hUCBDSCsTranswell共培养体系的建立.6. 发现MSCs比ITP-MSCs分泌较多的SDF-1。 .7. HUCBDSCs 促进ITP小鼠B淋巴细胞CD31 的表达.8. HUCBDSCs减少ITP 小鼠B细胞IgG分泌