Regulatory T cell as a restorative therapy for ischemic stroke

 描述(申请人提供)：白质(WM)损害，其特征是髓鞘丢失和产生髓鞘的少突胶质细胞(OLs)，是中风后功能障碍的主要原因，但直到最近才在治疗研究中被广泛认识。 在这里，我们建议通过关注WM的完整性及其在缺血脑中的免疫反应调节来纠正这一领域的差距。不同表型的激活的小胶质细胞/巨噬细胞决定了脑损伤后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的保存和修复是必不可少的，因为M2细胞可以解决局部炎症，清除破碎的髓鞘，并提供促进WM修复的营养因子。CD4+CD25+调节性T细胞(Tregs)是一种特殊的T细胞亚群，对免疫反应具有负性调节作用。我们最近的研究表明，过继Treg疗法通过靶向卒中后炎性调节障碍和神经血管破坏而发挥早期神经保护作用。然而，目前还不知道Tregs是否也对WM的完整性有有益的影响。最近，我们发现Treg条件培养液刺激小胶质细胞向M2表型分化，M2小胶质细胞在体外提高了OL的存活率和促进OPC的分化。这些令人兴奋的结果表明，Tregs可以保持WM的完整性。我们获得了进一步的有希望的数据：1)再灌注2小时的Treg转移减少了WM的损伤程度，并至少在28d内改善了感觉运动功能；2)卒中后Treg治疗导致IL-10持续升高，IL-10是一种对WM修复至关重要的Treg衍生细胞因子；3)Treg治疗促进了tMCAO后西区和灰质中小胶质细胞/巨噬细胞的M2极化。此外，通过系统注射IL-2/IL-2抗体复合体(IL-2/IL-2Ab)，我们成功地诱导了中风后循环中Tregs的强劲增加，这是一种已建立的在体内扩大Tregs的方法。我们证明了IL-2/IL-2Ab诱导的Treg扩张减少了tMCAO后7d的髓鞘丢失，并改善了感觉运动功能。目前的建议将进一步探讨Treg在卒中后WM损伤和修复中的作用，并开发体内Treg扩展作为促进WM完整性和促进卒中后恢复的新策略。有待检验的中心假设是，Treg通过以IL-10依赖的方式将小胶质细胞/巨噬细胞极化为M2表型，促进中风后WM的完整性和长期恢复。本研究提出了三个具体目标：目的1.验证卒中后Treg治疗改善长期功能恢复和促进WM完整性的假设。目的2.验证这一假说，即Treg来源的IL-10使小胶质细胞/巨噬细胞极化向M2表型转变，从而促进卒中后WM的完整性。目的3.验证卒中后IL-2/IL-2Ab体内扩增Tregs能有效减轻卒中后长期WM损伤、促进神经功能恢复的假说。

 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.