| A 3D Culture System of Human Immortalized Myometrial Cells
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Author:
Date:
2016-10-20
[Abstract] Myometrium forms the middle layer of the uterus and is mainly composed of the smooth muscle cells. The cells in vitro are usually grown in a single layer (2-dimensional; 2D) format, whereas in vivo cells are structured in an extracellular matrix scaffolding that allows the cells to communicate and respond to environmental cues. We have developed human myometrium and leiomyoma 3-dimensional (3D) culture, wherein the cells retain their molecular characteristics and respond to environmental cues (Malik and Catherino, 2012; Malik et al., 2014).
[摘要] 子宫肌层形成子宫的中间层，主要由平滑肌细胞组成。细胞在体外通常生长在单层（2维; 2D）格式中，而体内细胞在细胞外基质支架中结构化，其允许细胞沟通和响应环境线索。我们已经开发了人子宫肌瘤和平滑肌瘤3维（3D）培养物，其中细胞保留其分子特征并响应环境线索（Malik和Catherino，2012; Malik等人，2014）。 br /> [背景] 在过去十年中，随着更多实验室从使用人工2D格式的细胞培养转移到3D细胞培养模型系统，观察到一定的转变，其中细胞生长在允许它们附着并获得更生理结构的基质中。该模型系统为细胞提供更自然的分化状态，并且培养的细胞在体内形成组织样环境。这是一个详细的协议为myometrium 3D细胞培养生长胶原-1矩阵，修改从Malik和Catherino（2012年）。
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| Cell Tracer Violet and CellTracker Red CMTPX Staining of Purified Mature Plasmodium-infected Red Blood Cells
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Author:
Date:
2016-08-05
[Abstract] Efficient staining methods to identify Plasmodium-infected red blood cells (iRBCs) are crucial to discriminate precisely the immune cells responsible for their elimination from circulation. Here, we describe the protocol for the purification of iRBCs and their subsequent staining with the vital dyes Cell Tracer Violet (CTV) or CellTracker Red CMTPX (CMTPX), both of which readily diffuse into cells and bind covalently to intracellular amines. The iRBCs stained by using this protocol were used in ex vivo phagocytosis assays, to determine the ability of splenic dendritic cells of phagocytizing these parasites (Borges da Silva et al., 2015).
[摘要] Efficient staining methods to identify Plasmodium-infected red blood cells (iRBCs) are crucial to discriminate precisely the immune cells responsible for their elimination from circulation. Here, we describe the protocol for the purification of iRBCs and their subsequent staining with the vital dyes Cell Tracer Violet (CTV) or CellTracker Red CMTPX (CMTPX), both of which readily diffuse into cells and bind covalently to intracellular amines. The iRBCs stained by using this protocol were used in ex vivo phagocytosis assays, to determine the ability of splenic dendritic cells of phagocytizing ...
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| In vivo DCs Depletion with Diphtheria Toxin and MARCO+/MOMA1+ Cells Depletion with Clodronate Liposomes in B6.CD11c-DTR Mice
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Author:
Date:
2016-08-05
[Abstract] To evaluate precisely the relative roles of different splenic phagocytic cells during an immune response, efficient methods for the depletion of specific populations are needed. Here, we describe the protocols for the depletion of splenic dendritic cells (DCs) by human diphtheria toxin (DTx) treatment in target mice (which express the human DTx receptor in all CD11c+ DCs) and for the specific depletion of MARCO+/MOMA-1+ marginal zone macrophages (MZMΦs) with clodronate liposomes (ClLip) treatment (when a small dose of ClLip is ministered, MZMΦs preferentially uptake ClLip, and clodronate is released inside those cells causing apoptosis-mediated cell death). These protocols are adaptations from previous works (Jung et al., 2002; McGaha et al ...
[摘要] 为了准确地评估不同的脾吞噬细胞在免疫应答期间的相对作用，需要用于消耗特定群体的有效方法。 在这里，我们描述了通过人类白喉毒素（DTx）治疗在目标小鼠（其在所有CD11c + DC中表达人类DTx受体）中脾脏树突状细胞（DCs） 用氯膦酸脂质体（ClLip）处理（当小剂量的ClLip被分配时，MZMΦs优先摄取）的MARCO + /MOMA-1 + 边缘区巨噬细胞（MZMΦ） ClLip和氯膦酸盐在那些细胞内释放，引起凋亡介导的细胞死亡）。 这些方案是来自先前作品的改编（Jung等人，2002; McGaha等人，2011），并且用于评价DC和MZMΦ的各自的作用 在实验性血液阶段疟疾感染的急性期期间（Borges da Silva等人，2015）。
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