Quantitative morphology of induced phenotypes in cultured cells and tissues

我们继续与Ko博士(LG-NIA)合作，使用转录因子操作来描述在小鼠胚胎干细胞中诱导的发育途径。我们能够建立一种仅基于形态的分化分析，使用没有特定标记物的相位对比成像。我们能够证明，分化可以随着时间的推移而被跟踪，分化细胞之间的形态相似性是在早期建立的，并在分化的几天内保持不变。一个重要的里程碑是，我们能够建立一种条件，在这种条件下，细胞系在几个独立的实验中显示出一致的基因功能分组。此外，我们能够扩展这一分析以比较56个细胞系的形态。这些结果正在准备出版。 我们继续研究线虫寿命期间形态年龄-状态转变的分子基础。在之前发表的工作中，我们使用WND-Charm来识别线虫不同的形态衰老状态。我们使用这项技术根据过渡期中的年龄状态对蠕虫进行分类，在过渡期中，老龄化人口在第一阶段和第二阶段被平均分配给个体。这些蠕虫在基因、年龄、生长条件和视觉外观上是相同的，并且只能使用WND-Charm将其分类为年龄状态。对这两个亚群进行的微阵列实验显示，数百个基因的表达谱发生了显著变化。通过将我们的基因列表与线虫中其他衰老研究的基因列表进行比较，我们能够识别出我们研究中独有的几个基因家族和功能群，以及之前已经确定的几个重要的衰老基因。这项研究的结果正在准备发表。 在这项研究的下一阶段，我们将开始使用RNAi文库筛选我们的基因列表，以改变阶段I-阶段II的转变时间。

We have continued our work in collaboration with Dr. Minoru Ko (LG-NIA) to characterize developmental pathways induced in mouse embryonic stem cells using transcription factor manipulation. We were able to establish a differentiation assay based on morphology alone, using phase-contrast imaging without specific markers. We were able to show that differentiation can be tracked over time, and that morphological similarities between differentiating cells are established early and maintained for several days of differentiation. An important milestone is that we were able to establish a conditions where cell lines showed consistent grouping by gene function over several independent experiments. Additionally, we were able to expand this analysis to compare the morphologies of 56 cell lines. These results are being prepared for publication. We have continued work characterizing the molecular basis of morphological age-state transitions during the C. elegans life-span. In previous published work we used WND-CHARM to identify distinct morphological aging states in C. elegans. We used this technique to sort worms based on their age state during a transition period where an aging population is evenly divided between individuals in Stage I and Stage II. The worms were identical genetically, by chronological age, by growth conditions, and by visual appearance, and could only be sorted into age-states using WND-CHARM. Micro-array experiments performed on these two sub-populations revealed several hundred genes with significantly altered expression profiles. By comparing our gene lists with those from other aging studies in C. elegans, we were able to identify several gene families and functional groups that were unique to our study, as well as several important aging genes that were identified previously. The results of this study are being prepared for publication. In the next stage of this study, we will begin to screen through our gene list using RNAi libraries to shift the timing of the Stage I - Stage II transition.