| A Reliable Assay to Evaluate the Virulence of Aspergillus nidulans Using the Alternative Animal Model Galleria mellonella (Lepidoptera)
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Author:
Date:
2017-06-05
[Abstract] The greater wax moth Galleria mellonella has emerged as an effective heterologous host to study fungal pathogenesis and the efficacy of promising antifungal drugs (Mylonakis et al., 2005; Li et al., 2013). Here, a methodology describing the Aspergillus nidulans infection in G. mellonella larvae, along with insect survival analysis, is reported. This protocol allowed the distinction between virulent A. nidulans strains (such as TNO2A3), which induced high larval mortality rates, to those in which gene deletion was accompanied by reduced pathogenicity such as ∆gcsA and ∆sdeA (Fernandes et al., 2016).
[摘要] 作为一种有效的异源宿主，越来越多的蜡蛾已经出现了一种有效的异源宿主，用于研究真菌发病机制和有希望的抗真菌药物的功效（Mylonakis等人，2005; Li＆ et al。2013）。这里，描述了一种描述构巢曲霉感染的方法。 mellonella 幼虫与昆虫存活分析一起报道。该协议允许区分有毒的 A。造成高幼虫死亡率的构巢组织菌株（如TNO2A3）与基因缺失伴随着致病性降低的病例如ΔGCA和ΔSAA（Fernandes 等人，2016）。
背景 -G。 mellonella 是一种廉价的模型，易于处理，其先天免疫反应与哺乳动物免疫系统分享功能相似性。此外，感染真菌突变菌株的幼虫和小鼠表现出相似的存活率（Brennan等人，2002）。因此，幼虫构成了一种方便的动物宿主，用于在真菌发病机理分析中代替脊椎动物的使用。尽管昆虫模型具有所有优点，但只有少数报告显示了G中曲霉菌感染的作用。蜡螟。该协议描述了一种有效的方法，用于分析G中的构巢曲霉发病机制。 mellonella 幼虫。
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| Heterochronic Pellet Assay to Test Cell-cell Communication in the Mouse Retina
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Author:
Date:
2017-02-05
[Abstract] All seven retinal cell types that make up the mature retina are generated from a common, multipotent pool of retinal progenitor cells (RPCs) (Wallace, 2011). One way that RPCs know when sufficient numbers of particular cell-types have been generated is through negative feedback signals, which are emitted by differentiated cells and must reach threshold levels to block additional differentiation of that cell type. A key assay to assess whether negative feedback signals are emitted by differentiated cells is a heterochronic pellet assay in which early stage RPCs are dissociated and labeled with BrdU, then mixed with a 20-fold excess of dissociated differentiated cells. The combined cells are then re-aggregated and cultured as a pellet on a membrane for 7-10 days in vitro. During ...
[摘要] 构成成熟视网膜的所有七种视网膜细胞类型都是由普通的多能视网膜祖细胞池（RPC）产生的（Wallace，2011）。已经产生足够数量的特定细胞类型的RPC知道的一种方式是通过负反馈信号，其由分化细胞发射并且必须达到阈值水平以阻止该细胞类型的额外分化。评估负反馈信号是否由分化细胞发出的关键测定是异源沉淀测定，其中早期RPC被解离并用BrdU标记，然后与20倍过量的解离的分化细胞混合。然后将组合的细胞再次聚集并在细胞膜上培养7-10天。在这段时间内，RPC将会分化，BrdU + RPC的命运可以使用细胞类型特异性标记进行评估。开发这种沉淀测定的研究人员最初表明，当两种细胞类型混合在一起时，新生儿RPC与胚胎RPC相比，在加速进度条件下产生杆（Watanabe和Raff，1990; Watanabe等，1997）。我们已经使用这种测定来证明我们发现作为视网膜神经节细胞（RGC）分化的负调节物的声刺猬（Shh）促进RPC增殖（Jensen和Wallace，1997; Ringuette等，2014）。最近我们修改了异质性沉淀测定法，以评估视网膜无长突细胞的反馈信号的作用，将转化生长因子β2（Tgfβ2）鉴定为负反馈信号，并将Pten作为Tgfβ2应答的调节剂（Ma et al。，2007 ; ...
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