BIO407 Group4 Image processing of live cells, treated with DFX

## Study: This data is part of the practical course BIO407 2025 at the University of Zurich, titled 'Advanced Microscopy: From preparation to data and visualization'. The effects of Deferasirox (DFX) on mitochondria morphology were examined using different microscopy techniques. The data is originally from the paper: Gottwald EM, Schuh CD, Dr點ker P, Haenni D, Pearson A, Ghazi S, Bugarski M, Polesel M, Duss M, Landau EM, Kaech A, Ziegler U, Lundby AKM, Lundby C, Dittrich PS, Hall AM. The iron chelator Deferasirox causes severe mitochondrial swelling without depolarization due to a specific effect on inner membrane permeability. Sci Rep. 2020 Jan 31;10(1):1577. doi: 10.1038/s41598-020-58386-9. PMID: 32005861; PMCID: PMC6994599. ## Study Component: Job: Image Processing Task: 4) Timelapse of mitochondria in deferasirox (DFX) treated cells Timelapse data was analyzed: Mitochondria were segmented, their morphology and signal intensity were measured over time, and plotted. ## Biosample: Opossum kidney (OK) cells (kind gift from the group of Prof O. Devuyst (Physiology, University of Zurich)) ## Specimen: DFX treatment: 200uM Dyes: -Mitochondria-GFP BacMam 2.0 -TMRM (mitochondrial membrane potential dependent dye) ## Image Acquisition: Images were acquired using a Leica SP8 inverse STED 3x. Acquired channels: Channel 1: Mitochondria-GFP, 488nm excitation, 493nm-548nm emission Channel 2: TMRM, 553nm excitation, 564nm-650nm emission Objective: HC PL APO CS2 100x/1.40 OIL ## Image Data: Raw timelapse data: 151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression.lif Metadata of raw timelapse data: metadata_raw_data.txt Segmentation masks of mitochondria: GFP: 151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression_Mask_Channel1.tif TMRM: 151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression_Mask_Channel2.tif FIJI macro, documenting the analysis steps: Fiji_analysis.ijm Mean circularity of segmented mitochondria at each timepoint measured with GFP signal (Circ.) and Mean GFP signal intensity of segmented mitochondria at each timepoint (Mean): Channel1_Measurements.csv Mean circularity of segmented mitochondria at each timepoint measured with TMRM signal (Circ.) and Mean TMRM signal intensity of segmented mitochondria at each timepoint (Mean): Channel2_Measurements.csv Plot of mean mitochondria circularity over time: GFP: Channel1_Circularity.png TMRM: Channel2_Circularity.png Plot of mean GFP signal in mitochondria over time: Channel1_MeanIntensity.png Plot of mean TMRM signal in mitochondria over time: Channel2_MeanIntensity.png ## Image Correlation 151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression_Mask_Channel1.tif and 151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression_Mask_Channel2.tif contain the segmentation masks for the timelapse in 151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression.lif ## Image Analysis We used FIJI to perform a segmentation of the mitochondria. Individual steps can be found in the FIJI Macro. (Fiji_analysis.ijm) The segmentation masks were analyzed in FIJI and the 'Analyze Particles' command was used to measure the circularity and mean signal intensity of all segmented mitochondria. The mean measurements were then plotted for each timepoint.## 研究背景： 本数据集隶属于苏黎世大学2025学年BIO407实践课程，课程主题为《高级显微镜学：从样品制备到数据获取与可视化》。本研究通过多种显微镜技术探究了地拉罗司（Deferasirox，DFX）对线粒体形态的影响。本数据集的原始来源为以下学术论文： Gottwald EM, Schuh CD, Dräger P, Haenni D, Pearson A, Ghazi S, Bugarski M, Polesel M, Duss M, Landau EM, Kaech A, Ziegler U, Lundby AKM, Lundby C, Dittrich PS, Hall AM. 铁螯合剂地拉罗司通过特异性影响线粒体内膜通透性引发严重线粒体肿胀但不影响膜电位. Sci Rep. 2020 Jan 31;10(1):1577. doi: 10.1038/s41598-020-58386-9. PMID: 32005861; PMCID: PMC6994599. ## 研究组成部分： 任务类型：图像处理 具体任务：4）地拉罗司（DFX）处理细胞的线粒体延时成像分析 对延时成像数据开展了如下分析：对线粒体进行分割，随时间维度测量其形态特征与信号强度，并绘制相关可视化图表。 ## 生物样品： 负鼠肾（OK）细胞（由苏黎世大学生理学系O·德武伊斯特教授课题组惠赠） ## 样品处理： 地拉罗司（DFX）处理浓度：200μM 所用染料： - 线粒体-GFP BacMam 2.0 - TMRM（线粒体膜电位依赖性染料） ## 图像采集： 图像采集使用Leica SP8倒置STED 3x显微镜 采集通道： 通道1：线粒体-GFP，激发波长488nm，发射波长范围493nm-548nm 通道2：TMRM，激发波长553nm，发射波长范围564nm-650nm 物镜：HC PL APO CS2 100x/1.40 OIL ## 图像数据集： 原始延时成像数据： 151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression.lif 原始延时成像数据元数据： metadata_raw_data.txt 线粒体分割掩码： GFP通道：151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression_Mask_Channel1.tif TMRM通道：151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression_Mask_Channel2.tif 记录分析步骤的FIJI宏脚本： Fiji_analysis.ijm 基于GFP信号测量的各时间点分割线粒体平均圆度（Circ.）及分割线粒体的平均GFP信号强度（Mean）： Channel1_Measurements.csv 基于TMRM信号测量的各时间点分割线粒体平均圆度（Circ.）及分割线粒体的平均TMRM信号强度（Mean）： Channel2_Measurements.csv 线粒体平均圆度随时间变化图表： GFP通道：Channel1_Circularity.png TMRM通道：Channel2_Circularity.png 线粒体平均GFP信号强度随时间变化图表： Channel1_MeanIntensity.png 线粒体平均TMRM信号强度随时间变化图表： Channel2_MeanIntensity.png ## 图像关联说明： 文件151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression_Mask_Channel1.tif与151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression_Mask_Channel2.tif包含对应151106_slide1_bacmam20_PPC_7473_Esther_DH_Progression.lif中延时成像的分割掩码。 ## 图像分析流程： 我们使用FIJI软件完成线粒体分割，具体操作步骤可参见FIJI宏脚本Fiji_analysis.ijm。通过FIJI软件对分割掩码进行分析，调用「分析粒子」命令测量所有分割线粒体的圆度与平均信号强度，并针对每个时间点绘制平均测量值的可视化图表。