脂多糖注射液致大鼠亚急性脑 Microhemorrhages
Summary
我们提出了一种诱导和检测杜勒大鼠 LPS 注射液引起的中草药的实验方案, 可用于今后中草药的发病机制研究。
Abstract
脑 microhemorrhages (中草药) 在老年患者中是常见的, 与各种神经精神障碍相关。中草药的病因是复杂的, 神经炎症经常被观察作为共同发生。本文介绍了一种由脂多糖 (LPS) 注射液诱导的亚急性中草药大鼠模型, 以及一种检测中草药的方法. 系统性 LPS 注射液相对简单、经济、经济实惠。LPS 注射液的一个主要优点是它的稳定性导致炎症。通过观察、苏木精和伊红 (he) 染色、Perl 普鲁士染色、埃文斯蓝 (EB) 双重标记和磁共振成像-易感性加权成像 (MRI-中草药) 技术, 可以检测到 LPS 注射液引起的。最后, 本报告还讨论了其他开发中草药动物模型的方法, 包括它们的优点和/或缺点。
Introduction
经典脑 microhemorrhages (中草药) 是指血液降解产物的微小血管周围沉积物, 如含铁血黄素从脑1中的红细胞。根据鹿特丹扫描研究, 中草药可以发现在近17.8% 的人60–69岁和38.3% 在那些80年2。中草药在老年人中的患病率较高, 中草药的积累与认知和神经精神功能障碍之间的相关性已建立3,4。最近报道了中草药的几种动物模型, 包括 IV 型胶原酶立体定向注射液5、应用转基因6、β n-甲基氨基-l-丙氨酸暴露7、高血压8等诱发的啮齿动物模型,由全身炎症诱发的中草药是最接受的选择之一。费舍尔等。9首次使用的 LPS 来源于鼠伤寒沙门氏菌, 开发出一种急性中草药小鼠模型。随后, 同一组报告了使用相同方法2的亚急性中草药小鼠模型的发展。
LPS 被认为是一个标准化的炎症刺激通过腹腔注射。先前的研究证实, LPS 注射液可能导致神经炎症, 在动物模型2,10中大量的小胶质细胞和星形胶质细胞活化反应。此外, 神经炎症活化激活与中草药数量之间有正相关关系, 已建立2、10。在前人研究的基础上, 通过腹腔注射 LPS, 建立了中草药大鼠模型。
检测技术的进步导致了中草药研究调查的数量增加。最广泛公认的检测中草药的方法包括通过苏木精和伊红 (he) 染色检测红血球, 普鲁士蓝染色9检测铁铁, 通过免疫荧光法检测埃文斯蓝 (EB) 沉积物成像和7.0 特斯拉磁共振成像-易感性加权成像 (MRI-社会工作)10。本研究旨在开发一种中草药的筛选方法。
Protocol
这里描述的所有方法都已获得中国人民解放军陆军总医院动物护理和使用委员会 (ACUC) 的批准。 1. 材料 LPS 注射液的制备 将25毫升蒸馏水添加到25毫克的 LPS 粉末中, 从鼠伤寒到最终浓度为1毫克/毫升。将注射剂储存在4摄氏度的无菌管中。注意: LPS 是有毒的。 在正常0.9% 盐水溶液中制备 2% eb 溶液, 以保持 eb 注射液在工作浓度。 <p …
Representative Results
中草药可以使用各种方法检测到。最接受的方法包括: (1) 表面中草药的总观测和评估 (如图 1、上部板所示);(2) he 染色检测红细胞 (图2A、上部板) 或普鲁士染色检测红细胞裂解产生的铁 (图 2A、下部板);(3) eb 双重染色检测从 BBB 泄漏中提取的 eb 沉积物 (图 3, 左面板);(4) MRI-中草药 hypointe…
Discussion
中草药的研究在过去几年中有所增加。然而, 中草药的机制仍然不清楚, 促使科学家建立模拟这种特殊情况的动物模型。例如, 霍夫曼等。开发了缺氧诱导的中草药小鼠模型, 表明中草药是由缺氧和脑血管自动调节12的破坏引起的。路透社等。5在 APP23-transgenic 小鼠中建立了中草药模型, 表明脑淀粉样血管病 (CAA) 在中草药病因中起着重要作用. 费舍尔<em…
Divulgaciones
The authors have nothing to disclose.
Acknowledgements
我们感谢健丰老师和首都医科大学的同事在核磁共振成像方面的指导。我们还感谢宜昌第二人民医院神经病学系提供技术支持。
Materials
| LPS | Sigma-Aldrich | L-2630 | for inflammation induction |
| EB | Sigma-aldrich | E2129 | for EB leakage detection |
| DAPI dying solution | Servicbio | G1012 | count medium for IF |
| Perl’s Prussian staining | Solarbio | G1424 | Kit for Prussian staining |
| HE staining | Solarbio | G1120 | Kit for HE staining |
| chloral hydrate | Sigma-Aldrich | 47335U | For anesthesia |
| phosphate buffer saline (PBS) | Solarbio | P1022 | a kind of buffer solution commonly used in experiment |
| 0.9% saline solution | Hainan DonglianChangfu Pharmaceutical Co., Ltd., China | solution for perfusion | |
| paraformaldehyde | Sigma-Aldrich | 158127 | a kind of solution commonly used for fixation |
| 20% sucrose solution | Solarbio | G2461 | a kind of solution commonly used for fixation |
| 30% sucrose solution | Solarbio | G2460 | a kind of solution commonly used for fixation |
| vet ointment | Solcoseryl eye gel, Bacel, Switzerland | for rat's eyes protection |
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Citar este artículo
Li, D., Zhào, H., Wei, W., Liu, N., Dr. Huang, Y. Sub-acute Cerebral Microhemorrhages Induced by Lipopolysaccharide Injection in Rats. J. Vis. Exp. (140), e58423, doi:10.3791/58423 (2018).