Department of Neurology, Henry Ford Hospital, Detroit, MI, United States.
Department of Physics, Oakland University, Rochester, MI, United States.
Front Immunol. 2019 Nov 26;10:2747. doi: 10.3389/fimmu.2019.02747. eCollection 2019.
Microparticles (MPs, ~size between 0.1 and 1 mm) are lipid encased containers derived from intact cells which contain antigen from the parent cells. MPs are involved in intercellular communication and regulate inflammation. Stroke increases secretion of brain derived MP (BDMP) which activate macrophages/microglia and induce neuroinflammation. Lactadherin (Milk fat globule-EGF factor-8) binds to anionic phospholipids and extracellular matrices, promotes apoptotic cell clearance and limits pathogenic antigen cross presentation. In this study, we investigate whether BDMP affects stroke-induced neuroinflammation and whether Lactadherin treatment reduces stroke initiated BDMP-induced neuroinflammation, thereby improving functional outcome after stroke. Middle aged (8-9 months old) male C57BL/6J mice were subjected to distal middle cerebral artery occlusion (dMCAo) stroke, and BDMPs were extracted from ischemic brain 24 h after dMCAo by ultracentrifugation. Adult male C57BL/6J mice were subjected to dMCAo and treated via tail vein injection at 3 h after stroke with: (A) +PBS ( = 5/group); (B) +BDMPs (1.5 × 10, = 6/group); (C) +Lactadherin (400 μg/kg, = 5/group); (D) ++Lactadherin ( = 6/group). A battery of neurological function tests were performed and mice sacrificed for immunostaining at 14 days after stroke. Blood plasma was used for Western blot assay. Our data indicate: (1) treatment of Stroke with BDMP significantly increases lesion volume, neurological deficits, blood brain barrier (BBB) leakage, microglial activation, inflammatory cell infiltration (CD45, microglia/macrophages, and neutrophils) into brain, inflammatory factor (TNFα, IL6, and IL1β) expression in brain, increases axon/white matter (WM) damage identified by decreased axon and myelin density, and increases inflammatory factor expression in the plasma when compared to PBS treated stroke mice; (2) when compared to PBS and BDMP treated stroke mice, Lactadherin and BDMP+Lactadherin treatment significantly improves neurological outcome, and decreases lesion volume, BBB leakage, axon/WM injury, inflammatory cell infiltration and inflammatory factor expression in the ischemic brain, respectively. Lactadherin treatment significantly increases anti-inflammatory factor (IL10) expression in ischemic brain and decreases IL1β expression in plasma compared to PBS and BDMP treated stroke mice, respectively. BDMP increases neuroinflammation and aggravates ischemic brain damage after stroke. Thus, Lactadherin exerts anti-inflammatory effects and improves the clearance of MPs to reduce stroke and BDMP induced neurological deficits.
微粒（MPs，~大小为 0.1 至 1 毫米）是源自完整细胞的脂质包裹容器，其中包含来自母细胞的抗原。MPs 参与细胞间通讯并调节炎症。中风会增加脑源性 MP（BDMP）的分泌，激活巨噬细胞/小胶质细胞并诱导神经炎症。乳贴蛋白（Milk fat globule-EGF factor-8）与阴离子磷脂和细胞外基质结合，促进凋亡细胞清除并限制致病性抗原交叉呈递。在这项研究中，我们研究了 BDMP 是否会影响中风引起的神经炎症，以及乳贴蛋白治疗是否会减少中风引发的 BDMP 诱导的神经炎症，从而改善中风后的功能结果。中年（8-9 个月大）雄性 C57BL/6J 小鼠接受远端大脑中动脉闭塞（dMCAo）中风，中风后 24 小时通过超速离心从缺血性大脑中提取 BDMP。成年雄性 C57BL/6J 小鼠接受 dMCAo，并在中风后 3 小时通过尾静脉注射以下药物进行治疗：（A）+PBS（每组=5 只）；（B）+BDMPs（1.5×10 ，每组=6 只）；（C）+乳贴蛋白（400μg/kg，每组=5 只）；（D）++乳贴蛋白（每组=6 只）。进行一系列神经功能测试，并在中风后 14 天处死小鼠进行免疫染色。用 Western blot 测定法测定血浆。我们的数据表明：（1）BDMP 治疗中风会显著增加病变体积、神经功能缺损、血脑屏障（BBB）渗漏、小胶质细胞活化、炎性细胞浸润（CD45、小胶质细胞/巨噬细胞和中性粒细胞）入脑、炎性因子（TNFα、IL6 和 IL1β）在脑中的表达，减少轴突/白质（WM）损伤，表现为轴突和髓鞘密度降低，并增加血浆中的炎性因子表达与 PBS 治疗的中风小鼠相比；（2）与 PBS 和 BDMP 治疗的中风小鼠相比，乳贴蛋白和 BDMP+乳贴蛋白治疗可显著改善神经功能，减少病变体积、BBB 渗漏、轴突/WM 损伤、炎性细胞浸润和缺血性大脑中的炎性因子表达，分别。与 PBS 和 BDMP 治疗的中风小鼠相比，乳贴蛋白治疗可显著增加缺血性大脑中的抗炎因子（IL10）表达并降低血浆中的 IL1β 表达。BDMP 增加中风后的神经炎症并加重缺血性脑损伤。因此，乳贴蛋白发挥抗炎作用并促进 MPs 的清除，以减少中风和 BDMP 引起的神经功能缺损。
