| Direct Reprogramming of Mouse Embryonic Fibroblasts to Conventional Type 1 Dendritic Cells by Enforced Expression of Transcription Factors
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Author:
Date:
2020-05-20
[Abstract] Ectopic expression of transcription factor combinations has been recently demonstrated to reprogram differentiated somatic cells towards the dendritic cell (DC) lineage without reversion to a multipotent state. DCs have the ability to induce potent and long-lasting adaptive immune responses. In particular, conventional type 1 DCs (cDC1s) excel on antigen cross-presentation, a critical step for inducing CD8+ T cell cytotoxic responses. The rarity of naturally occurring cDC1s and lack of in vitro methodologies for the generation of pure cDC1 populations strongly hinders the study of cDC1 lineage specification and function. Here, we describe a protocol for the generation of induced DCs (iDCs) by lentiviral-mediated expression of the transcription factors PU.1, IRF8 and ...
[摘要] [摘要] 转录因子组合的异位表达最近被证明可以将分化的体细胞重编程为树突状细胞（DC）谱系，而不会回复到多能状态。DC具有诱导有效和持久的适应性免疫应答的能力。在特定的常规类型1的DC（cDC1s）练成抗原交叉呈递，用于诱导CD8的关键步骤+ T细胞的细胞毒性应答。天然存在的cDC1的稀有性和缺乏用于生成纯cDC1群体的体外方法论，严重阻碍了对cDC1谱系规格和功能的研究。在这里，我们描述了用于生成感应DC（iDC）的协议 慢病毒介导的转录因子PU.1，IRF8和BATF3在小鼠胚胎成纤维细胞中的表达。iDC 在9天内获得DC形态，cDC1表型和转录特征。使用此协议生成的iDC 具有对炎症刺激，吞噬死细胞，将抗原加工并交叉呈递给CD8 + T细胞的功能。DC重新编程提供了一个简单易处理的系统，可以生成大量的cDC1类细胞用于高内涵筛选，从而开辟了新途径，可以更好地了解cDC1的规格和功能。将来，在成纤维细胞中忠实诱导cDC1命运可能会导致产生患者特定的疫苗接种DC。
[背景技术树突状细胞（DC）是专业的抗原呈递细胞，专门用于识别，加工和呈递T细胞抗原，在诱导适应性免疫应答和免疫记忆中起关键作用（Me rad 等，2013）。DC可以分为4个主要子集：浆细胞样DC（pDC ），大量1型干扰素的产生者，循环单核细胞衍生的单核细胞衍生DC 和常规DC（cDC ...
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| A Modified Semisolid Clonal Culture for Identification of B-1 and B-2 Progenitor Colony Forming Ability of Mouse Embryonic Hemogenic Endothelial Cells
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Author:
Date:
2020-05-05
[Abstract] The search for the origin of the first hematopoietic stem cells (HSCs) in the mouse embryo has been a hot topic in the field of developmental hematopoiesis. Detecting lymphoid potential is one of the supportive evidence to show the definitive hematopoietic activity of HSCs. However, the first B-lymphoid potential in the mouse embryos are reported to be biased to innate-like B-1 cell lineage that can develop from hemogenic endothelial cells (HECs) independently of HSCs. On the other hand, conventional adaptive immune B cells (B-2) cells are considered to be exclusively derived from HSCs. Therefore, segregating B-1 and B-2 progenitor potential is important to understand the developmental process of HSCs that are also produced from HECs through intermediate precursors referred to as ...
[摘要] [摘要 ] 在搜索起源的第一造血干细胞（HSCs）在小鼠胚胎一直是一个热门话题在该领域发展造血功能。检测淋巴潜力是一个支持性证据显示权威造血活动的造血干细胞然而，据报道，小鼠胚胎中的第一个B淋巴样电位偏向于先天性B-1细胞谱系，该谱系可以从造血内皮细胞（HEC）脱离HSC发育而来。 B细胞（B-2）细胞被认为仅来自HSC。因此，分离B-1和B-2祖细胞的潜力对于理解也由HEC通过中间前体（也称为HEC）产生的HSC的发育过程非常重要。 HECs和pre-HSCs均显示pre-HSCs显示内皮表面表型并需要基质支持以检测其造血活性。利用基质细胞培养后再进行改良的半固体克隆培养的方法使我们能够检测B-1的菌落形成单位数量/ B-2祖细胞最初源自HEC / HSC之前的细胞，将反映B-1偏倚或多谱系繁殖的HSC的潜力。
[背景 ] 半固体克隆培养（甲基纤维素集落形成测定法）是检测造血祖细胞数量的传统方法。一个集落被认为是来自单个祖细胞（克隆来源），添加的细胞因子在细胞的形成中起着重要作用。产量菌落，例如Epo增强了红细胞的菌落形成单位（CFU-E）或红细胞的爆发形成单位（BFU-E），而G-CSF / GM-CSF将增强CFU-G（粒细胞），M ...
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