CISA

Computational Immune Synapse Analysis.

Installation

This is a py script that can be imported. To use, copy the CISA.py to your working directory.

Example

See example notebook using core A5 from the published melanoma IMC dataset by Hoch et al., 2022

Hoch, Tobias, et al. "Multiplexed imaging mass cytometry of the chemokine milieus in melanoma characterizes features of the response to immunotherapy." Science immunology 7.70 (2022): eabk1692.

Dependency

This is a Python 3 package. The core functions use only numpy, pandas, scipy and skimage.

This package takes pixelated masks for cell segmentation and cell boundaries. However, it is recommanded to use shapely and rasterio to manipulate vectorized cell masks (not demonstrated).

Usage

Input (required):

• img_dict: {marker: 2d image}. Input single channel images as a dictionary.

• cell_df: dataframe of cell information where each row is a cell. The only required columns are cell_id and cell_type, with colnames customizable as optional input.

• cellmask: 2D int ndarray. Image mask of cells. Pixel values should be integers that matches the cell IDs in cell_df.

• boundmask: 2D int ndarray. Image mask of cell boundaries. Pixel values should be integers that matches cell ID in cell_df.

• neighbor_classes: list of neighboring classes. Other classes are not included in the result dataframe, but overlaping

• expressing_markers: list of markers that are expressed in the center cell types. Their background level will be calculated as the average level in other cell types. All other markers that are not listed but included in img_dict will be treated as expressed in the center cell types and their background level will be calculated as the average level in center cell types.

Input (optional):

• cell_list: list of cell IDs of center cells to compute CISA. If None, all cells are considered.

• target_classes: list of center cell types. Default: ['T'].

• n_bootstrap: number of bootstraps for the null model. Default: 1000

• e_width: width of dilation to calculate overlapping. Default: 2 pixels.

• bound_width: width of cell boundary dilation width before calculating overlapping. 0 = use raw bound. Default: 0 pixel.

• dropna: Bool, if drops invalid synapses. Default: True

• class_col: the colname for the cell type column in cell_df. Default: 'Class'

• cell_id_col: the colname for the cell ID column in cell_df. Default: 'Cell'

• keep_cols: list of colnames that should be kept in the output dataframe. Default: []

• excluded: list of cell between which and the center cells the contacted regions should be completely ignored in calculation. One example is to reduce the influence of signals from neighboring same type of cells. Default: ['T'].

Output

Output is a dataframe where each row corresponds to a center-neighbor cell pair.

For each marker m in the input, there will be several columns:

{m}_CISA is the calculated synapse strength between the two cells.

{m}_c and {m}_nc: Sum of pixel intensity in the contacting (c) and non-contacting (nc) boundary areas.

{m}_miu and {m}_sigma: Mean and std of the synapses under the null model where a random connected pixls along the cell boundary are selected as the contacting area.

{m}_z and {m}_p: The z-score and p-value for the observed synapse strength under the null model.

Notes and other information

Image preprocessing, such as de-noising, signal transformation and normalization, is determined by the user.

The running time varies by computation power but the example data can be run within 30 mins with the default parameters. A large bootstrap number may significantly increase the running time. In most cases, the CISA score (as in {m}_CISA column) is sufficient to characterize the behaviour.

The code have been tested on CentOS Linux 7, Windows 11 and macOS 13.4.