Yan Ping, Zhu Alec, Liao Fan, Xiao Qingli, Kraft Andrew, Gonzales Ernie, Perez Ron, Greenberg Steven M, Holtzman David, Lee Jin-Moo
Department of Neurology and the Hope Center for Neurological Disorders, Washington University School of Medicine, St. Louis, Missouri, USA.
Philip Kistler Stroke Research Center, Department of Neurology, Massachusetts General Hospital and Harvard Medical School, Boston, MA, USA.
Stroke. 2015 Jun;46(6):1633-1640. doi: 10.1161/STROKEAHA.115.008582. Epub 2015 May 5.
Cerebral amyloid angiopathy (CAA) is a common cause of recurrent intracerebral hemorrhage in the elderly. Previous studies have shown that CAA induces inflammation and expression of matrix metalloproteinase-2 and matrix metalloproteinase-9 (gelatinases) in amyloid-laden vessels. Here, we inhibited both using minocycline in CAA mouse models to determine whether spontaneous intracerebral hemorrhage could be reduced.
Tg2576 (n=16) and 5xFAD/ApoE4 knockin mice (n=16), aged 17 and 12 months, respectively, were treated with minocycline (50 mg/kg, IP) or saline every other day for 2 months. Brains were extracted and stained with X-34 (to quantify amyloid), Perls' blue (to quantify hemorrhage), and immunostained to examined β-amyloid peptide load, gliosis (glial fibrillary acidic protein [GFAP], Iba-1), and vascular markers of blood-brain barrier integrity (zonula occludins-1 [ZO-1] and collagen IV). Brain extracts were used to quantify mRNA for a variety of inflammatory genes.
Minocycline treatment significantly reduced hemorrhage frequency in the brains of Tg2576 and 5xFAD/ApoE4 mice relative to the saline-treated mice, without affecting CAA load. Gliosis (GFAP and Iba-1 immunostaining), gelatinase activity, and expression of a variety of inflammatory genes (matrix metalloproteinase-9, NOX4, CD45, S-100b, and Iba-1) were also significantly reduced. Higher levels of microvascular tight junction and basal lamina proteins were found in the brains of minocycline-treated Tg2576 mice relative to saline-treated controls.
Minocycline reduced gliosis, inflammatory gene expression, gelatinase activity, and spontaneous hemorrhage in 2 different mouse models of CAA, supporting the importance of matrix metalloproteinase-related and inflammatory pathways in intracerebral hemorrhage pathogenesis. As a Food and Drug Administration-approved drug, minocycline might be considered for clinical trials to test efficacy in preventing CAA-related intracerebral hemorrhage.
脑淀粉样血管病（CAA）是老年人复发性脑出血的常见病因。既往研究表明，CAA可诱导载有淀粉样蛋白的血管发生炎症以及基质金属蛋白酶-2和基质金属蛋白酶-9（明胶酶）的表达。在此，我们在CAA小鼠模型中使用米诺环素同时抑制这两者，以确定是否能减少自发性脑出血。
分别对17月龄的Tg2576小鼠（n = 16）和12月龄的5xFAD/ApoE4基因敲入小鼠（n = 16）每隔一天给予米诺环素（50 mg/kg，腹腔注射）或生理盐水，持续2个月。提取脑组织，用X-34染色（用于量化淀粉样蛋白）、Perls蓝染色（用于量化出血），并进行免疫染色以检测β-淀粉样肽负荷、胶质增生（胶质纤维酸性蛋白[GFAP]、离子钙结合衔接分子1[Iba-1]）以及血脑屏障完整性的血管标志物（闭合蛋白-1[ZO-1]和IV型胶原）。用脑组织提取物对多种炎症基因的mRNA进行定量分析。
与生理盐水处理的小鼠相比，米诺环素治疗显著降低了Tg2576和5xFAD/ApoE4小鼠脑内的出血频率，且不影响CAA负荷。胶质增生（GFAP和Iba-1免疫染色）、明胶酶活性以及多种炎症基因（基质金属蛋白酶-9、NADPH氧化酶4[NOX4]、白细胞共同抗原[CD45]、S-100β蛋白和Iba-1）的表达也显著降低。与生理盐水处理的对照组相比，米诺环素处理的Tg2576小鼠脑内微血管紧密连接蛋白和基膜蛋白水平更高。
米诺环素可减轻2种不同CAA小鼠模型中的胶质增生、炎症基因表达、明胶酶活性以及自发性出血，这支持了基质金属蛋白酶相关途径和炎症途径在脑出血发病机制中的重要性。作为一种经美国食品药品监督管理局批准的药物，米诺环素可考虑用于临床试验，以测试其预防CAA相关脑出血的疗效。
