DOI: 10.4121/c8025882-83fb-4d4f-bf20-80789cb5e776.v1
title: Data underlying the publication: Inter-Spheroid proximity and matrix remodelling determine CAF mediated cancer cell invasion

description: <strong>README</strong>Data Underlying: Inter-spheroid Proximity and Matrix Remodelling Determines CAF-mediated Cancer Cell InvasionStatus: Manuscript submitted and currently under review<br><strong>Overview</strong>This dataset supports the manuscript titled “Inter-spheroid Proximity and Matrix Remodelling Determines CAF-mediated Cancer Cell Invasion”. This work investigates the interaction between Cancer Associated Fibroblast (CAFs) and Luminal A breast cancer cell (MCF7s) spheroids in collagen matrices, with the objective of elucidating the interaction mechanisms that enable CAFs to promote cancer cell invasion. Timelapse imaging and analysis reveals a systematic interaction pattern between the two cell types. This dataset includes raw and processed data from 3D spheroid invasion assays of MCF7 breast cancer cell spheroids (mCherry-labelled), 19TT Cancer-Associated Fibroblast (CAF) spheroids (H2B-GFP-labeled), and heterospheroids consisting of a 1:1 ratio of MCF7 (mCherry-labelled) and 19TT Cancer-Associated Fibroblast (CAF) cells (H2B-GFP-labeled). All experiments were conducted in 3 mg/mL type 1 Collagen matrices. For invasion assays, experiments were run for a duration of 48h (Figs 2, 5, and 6) and 24h for MMP inhibition experiments (Figs 3 and 4). Imaging was performed using time-lapse fluorescence microscopy with reflection imaging to visualize collagen structure.<br><strong>Folder Structure</strong>Main folders are named Data_Fig1, Data_Fig2 through Data_Fig6. Within these folders, subfolders include Datasets, Fig Images, Graphs, and Movies. The Datasets folder contains the raw files used to analyse and generate images and graphs. Datasets used in Fig 1 (IF_pCK_aSMA and PSR_FL_slide_scans) can only be opened using the QuPath software. Datasets for the rest of the figures can be opened using FIJI/ImageJ2. Dataset folders are further divided into sub-folders by condition (e.g. Control, BB94). The Fig Images folder contains images used in manuscript figures. The Graphs folder contains vector image files of graphs (.pdf), Excel files (.xlsx) with data values, and Prism Graphpad files (.pzfx). The Movies folder contains video files (.avi) for datasets used in figures.Sample Analysis contains subfolders with sample analysis files and intermediates. Each type of analysis is shown once using data from a specific figure. For example, the Fig2C_Expansion_Analysis folder contains two datasets, intermediates, and results for the dArea graphs in Fig2C.Figure Files includes the graphical abstract, main text figures, and supporting submission figures in vector format (.pdf) and Keynote format (.key).Submission Videos contains all movies linked to the manuscript, organized by figure.Scripts contains all MATLAB codes used to analyse and quantify data. These include scripts for collagen fibre orientation analysis (colfibre_orientation.m, colfibre_mcf.m), CAF invasion index quantification (invasive_index.m), and collagen void fraction and pixel intensity measurement (Pxq_VF.m). To use these scripts, extract individual channels from composite tiff images and save them as image sequences via ImageJ. Scripts are modular, annotated, and require MATLAB’s Image Processing Toolbox. Some use custom fibre analysis functions. Custom functions saved in this folder include CAF_center.m, Centroid_3d.m, Circfit.m, Histwc.m, Imcrop.m, and PDF_function3.m. Supporting Information contains datasets and figure files used for supporting figures. The folder structure mirrors the main data folders. FigS2_CAF02-03-06 contains .mrxs files for CAF02, CAF03, and CAF06 tumors. Graphs are saved as Excel (.xlsx) and Prism Graphpad (.pzfx) files.Text contains various manuscript-related documents including the main manuscript (.docx), figure captions (.docx), electronic supporting information (.docx), declaration of competing interests (.docx), statement of significance (.docx), and supporting information text (.docx).Filetypes included in the dataset are text files (.docx), Excel spreadsheets (.xlsx) containing quantified data, image files (.tiff) of raw datasets and figures, MATLAB scripts (.m), native graph files (.pzfx) from Prism, slide scanner image files (.mrxs), Keynote files (.key), and high-resolution vector images (.pdf).