Regulatory T cell as a restorative therapy for ischemic stroke

 描述（由申请人提供）：白色物质（WM）病变的特征是髓鞘和产生髓鞘的少突胶质细胞（OL）的丢失，是卒中后功能障碍的主要原因，但直到最近才在治疗研究中得到广泛认识。 在这里，我们建议纠正这个领域的差距，集中在WM的完整性和它的调制缺血性脑免疫反应。已知不同表型的活化小胶质细胞/巨噬细胞决定脑损伤后OL细胞的命运和WM的完整性。具体地说，“交替激活”的M2表型对于WM保存和修复是必不可少的，因为M2细胞解决局部炎症，清除破裂的髓鞘，并提供促进WM修复的营养因子。CD 4 + CD 25+调节性T细胞（TCRs）是负调节免疫应答的特化T细胞亚群。我们最近的研究表明，过继性Treg治疗通过靶向卒中后炎症失调和神经血管破坏发挥早期神经保护作用。然而，目前尚不清楚TdR是否对WM完整性也有有益影响。最近，我们发现Treg条件培养基刺激小胶质细胞向M2表型极化，并且M2小胶质细胞在体外增强OL存活并促进OPC分化。这些令人兴奋的结果表明，TdR可以保持WM的完整性。我们获得了进一步有希望的数据，显示1）在再灌注2 h时Treg转移减少了WM损伤的程度，并改善了短暂大脑中动脉闭塞（tMCAO）后至少28天的感觉运动功能; 2）卒中后Treg治疗导致IL-10的长期升高，IL-10是一种主要的Treg衍生的细胞因子，对WM修复很重要; 3）Treg处理促进了tMCAO后WM和灰质中小胶质细胞/巨噬细胞的M2极化。此外，我们通过全身注射白细胞介素（IL）-2/IL-2抗体复合物（IL-2/IL- 2Ab）成功地诱导了中风后循环中Tcl 3的稳健增加，这是一种在体内扩增Tcl 3的既定方法。我们证明了IL-2/IL-2Ab诱导的Treg扩增减少了tMCAO后7天的髓鞘丢失并改善了感觉运动功能。目前的提案将进一步探索TdR对脑卒中后WM损伤和修复的影响，并开发体内Treg扩增作为促进WM完整性和增强脑卒中后恢复的新策略。待检验的中心假设是Tcl 4通过以IL-10依赖性方式使小胶质细胞/巨噬细胞向M2表型极化来促进WM完整性和卒中后的长期恢复。提出了三个具体目标：目标1。检验卒中后Treg治疗改善长期功能恢复并促进WM完整性的假设。目标2.检验Treg衍生的IL-10将小胶质细胞/巨噬细胞极化向M2表型转移，从而促进卒中后WM完整性的假设。目标3：验证卒中后IL-2/IL-2Ab治疗的THBE体内扩增在减少长期WM损伤和改善卒中后神经恢复方面有效的假设。

 DESCRIPTION (provided by applicant): White matter (WM) lesions, characterized by the loss of myelin and myelin-producing oligodendrocytes (OLs), are a major cause of functional disability after stroke but have not been widely appreciated in therapeutic studies until recently. Here we propose to rectify this gap in the field by focusing on WM integrity and its modulation by immune responses in the ischemic brain. Activated microglia/macrophages of distinct phenotypes are known to determine OL cell fate and WM integrity after brain injuries. Specifically, the "alternatively activated" M2 phenotype is essential for WM preservation and repair because M2 cells resolve local inflammation, clear broken myelin sheaths, and provide trophic factors that promote WM repair. CD4+CD25+ regulatory T cells (Tregs) are a specialized subpopulation of T cells that negatively regulate immune responses. Our recent study demonstrated that adoptive Treg therapy exerted early neuroprotection by targeting inflammatory dysregulation and neurovascular disruption after stroke. However, it is not known whether Tregs also have a beneficial effect on WM integrity. Recently, we discovered that Treg-conditioned media stimulates microglial polarization toward the M2 phenotype, and M2 microglia enhance OL survival and promote OPC differentiation in vitro. These exciting results suggest that Tregs can preserve WM integrity. We obtained further promising data showing that 1) Treg transfer at 2h of reperfusion reduced the extent of WM injury and improved sensorimotor functions for at least 28d after transient middle cerebral artery occlusion (tMCAO); 2) Post-stroke Treg treatment resulted in a long-lasting elevation of IL-10, a major Treg-derived cytokine that is important for WM repair; 3) Treg treatment promoted M2 polarization of microglia/macrophages in both WM and gray matter after tMCAO. Furthermore, we have successfully induced a robust increase of Tregs in the circulation after stroke by systemic injection of interleukin (IL)-2/IL-2 antibody complex (IL-2/IL- 2Ab), an established approach to expand Tregs in vivo. We demonstrated that IL-2/IL-2Ab-induced Treg expansion reduces myelin loss 7d after tMCAO and improves sensorimotor functions. The current proposal will further explore the effects of Tregs on WM injury and repair after stroke and develop in vivo Treg expansion as a novel strategy to promote WM integrity and enhance post-stroke recovery. The central hypothesis to be tested is that Tregs promote WM integrity and long-term recovery after stroke by polarizing microglia/macrophages toward the M2 phenotype in an IL-10 dependent manner. Three specific aims are proposed: Aim 1. Test the hypothesis that Treg treatment after stroke improves long-term functional recovery and promotes WM integrity. Aim 2. Test the hypothesis that Treg-derived IL-10 shifts microglia/macrophage polarization towards the M2 phenotype, thereby promoting WM integrity after stroke. Aim 3. Test the hypothesis that in vivo expansion of Tregs with post-stroke IL-2/IL-2Ab treatment is effective in reducing long-term WM injury and improving neurological recovery after stroke.