Distance Analysis: oralcavity-codex-czi

Analyze and visualize cell-to-nearest-endothelial-cell distance distributions for the oralcavity-codex-czi dataset.

import numpy as np
import pandas as pd
import os
import json
import requests
import shutil
import matplotlib.pyplot as plt
import seaborn as sns
from sklearn.preprocessing import normalize
import plotly.express as px

from _cde_compute_edges_from_nodes import *

pd.set_option('display.max_columns', None)
pd.set_option('display.max_rows', None)

# suppress warnings
import warnings
warnings.filterwarnings("ignore")

basepath = "/u/yashjain/hra-cell-distance-analysis/data"
dataset_dir = "oralcavity-codex-czi"
data_filedir = os.path.join("data-processed-nodes-with-harmonized-cell-types", dataset_dir)
output_edge_dir = os.path.join("data-processed-edges", dataset_dir)
figures_output_dir = "generated-figures"

# Function to load your data
def load_data(path, edges=False):
    if edges:
        column_names = ['cell_id', 'x1', 'y1', 'z1', 'x2', 'y2', 'z2']
        data = pd.read_csv(path, header=None, names=column_names)
    else:
        data = pd.read_csv(path)
    return data

# Function to read all files ending with "-nodes.csv" in the `data_filedir` directory into a single DataFrame. 
# Another additional column `Dataset` is added to identify the dataset name which comes from the filename before the `-nodes.csv` suffix.

# Additionally, function reads all files ending with "-edges.csv" in the `output_edge_dir` directory into a single DataFrame. 
# Three additional columns are added "Dataset", "Anchor Cell Type", and "Anchor Cell Type Level" to identify the dataset name, anchor cell type, and anchor cell type level respectively which come from the filename before the `.csv` suffix.
# The three additional columns are created by splitting the filename on the `-` character, and extracting the relevant parts.
# On splitting, the first part is the dataset name, second part is the anchor cell type level, and third part is the anchor cell type, and last part is the `edges` suffix.
# When reading files, check if the file has the correct format (i.e., ends with `-edges.csv`).

# Additionally, the function merges the edges DataFrame with the nodes DataFrame to get the cell type information for the anchor cells.
# This is done by reading the corresponding nodes file from the `data_filedir` directory for each edges file, and merging it with the edges DataFrame on the `cell_id` column.
# The merged DataFrame contains the edges with additional columns for the cell type information.

# The function returns three DataFrames:
# 1. `merged_nodes`: DataFrame containing all nodes with an additional column `Dataset`.
# 2. `merged_edges`: DataFrame containing all edges with additional columns `Dataset`, `Anchor Cell Type`, and `Anchor Cell Type Level`.
# 3. `merged_nodes_for_all_edges`: DataFrame containing all edges with additional columns `Dataset`, `Anchor Cell Type`, `Anchor Cell Type Level`, and the cell type information for cells.
def read_all_edge_datasets(basepath, data_filedir, output_edge_dir):
    all_nodes_files = []
    all_edges_files = []
    all_nodes_edges_files = []
    for file in os.listdir(os.path.join(basepath, output_edge_dir)):
        if file.endswith("-edges.csv"):
            file_path = os.path.join(basepath, output_edge_dir, file)
            dataset_name, anchor_cell_type_level, anchor_cell_type = file.replace("-edges.csv", "").split('-')
            edges_df = load_data(file_path, edges=False)
            edges_df['Dataset'] = dataset_name
            edges_df['Anchor Cell Type'] = anchor_cell_type
            edges_df['Anchor Cell Type Level'] = anchor_cell_type_level
            edges_df.rename(columns={"distance": "Distance"}, inplace=True) # Rename column "distance" to "Distance".
            all_edges_files.append(edges_df)

            # Read the corresponding nodes file from data_filedir to get the cell type information
            nodes_file_path = os.path.join(basepath, data_filedir, f"{dataset_name}-nodes.csv")
            nodes_df = load_data(nodes_file_path)
            nodes_df['Dataset'] = dataset_name
            all_nodes_files.append(nodes_df)

            # Add a new 'cell_id' column to nodes_df
            nodes_df['cell_id'] = range(len(nodes_df))
            # Set 'cell_id' column as index for nodes_df
            nodes_df.set_index('cell_id', inplace=True)
            # Merge edges_df with nodes_df to get the cell type information for the anchor cells
            edges_nodes_df = pd.merge(edges_df, nodes_df[['Level Three Cell Type', 'Level Two Cell Type', 'Level One Cell Type']], how='left', left_on='cell_id', right_index=True)
            all_nodes_edges_files.append(edges_nodes_df)

    
    merged_edges = pd.concat(all_edges_files, ignore_index=True)
    merged_nodes = pd.concat(all_nodes_files, ignore_index=True)
    merged_nodes_for_all_edges = pd.concat(all_nodes_edges_files, ignore_index=True) 

    return merged_nodes, merged_edges, merged_nodes_for_all_edges

def create_directory(directory):
    if not os.path.exists(directory):
        os.makedirs(directory)
        print(f"Directory '{directory}' created successfully.")
    else:
        print(f"Directory '{directory}' already exists.")

Get initial statistics and identify endothelial cell categories for dataset.

df_all_nodes, df_all_edges, df_all_edges_with_cell_types = read_all_edge_datasets(basepath, data_filedir, output_edge_dir)

df_all_nodes.head(5)

	x	y	Original Cell Type	Level Three Cell Type	Level Three CL Label	Level Three CL ID	CL_Match/3	Level Two Cell Type	Level Two CL Label	Level Two CL ID	CL_Match/2	Level One Cell Type	Level One CL Label	Level One CL ID	CL_Match/1	Dataset
0	2466.85	580.95	Others	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	Buccal_Mucosa_And_Minor_Salivary_Glands_1
1	2421.30	629.05	Others	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	Buccal_Mucosa_And_Minor_Salivary_Glands_1
2	2459.00	681.80	Others	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	Buccal_Mucosa_And_Minor_Salivary_Glands_1
3	2418.20	700.00	Others	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	Buccal_Mucosa_And_Minor_Salivary_Glands_1
4	2444.70	768.30	Treg	t cell:regulatory	regulatory T cell	CL:0000815	skos:exactMatch	t cell	T cell	CL:0000084	skos:exactMatch	immune cell	leukocyte	CL:0000738	skos:exactMatch	Buccal_Mucosa_And_Minor_Salivary_Glands_1

# Print the total number of unique cell types per dataset. Compute separately for each cell type column (Level One Cell Type, Level Two Cell Type, Level Three Cell Type, Original Cell Type).
print("Total number of unique cell types per cell type annnotation level:")
unique_cell_types = {
    'Original Cell Type': df_all_nodes['Original Cell Type'].nunique(),
    'Level Three Cell Type': df_all_nodes['Level Three Cell Type'].nunique(),
    'Level Two Cell Type': df_all_nodes['Level Two Cell Type'].nunique(),
    'Level One Cell Type': df_all_nodes['Level One Cell Type'].nunique()
}
for cell_type, count in unique_cell_types.items():
    print(f"{cell_type}: {count}")

Total number of unique cell types per cell type annnotation level:
Original Cell Type: 39
Level Three Cell Type: 28
Level Two Cell Type: 20
Level One Cell Type: 6

# Save the unique cell types containing "endothelial" in name per cell type column (Level One Cell Type, Level Two Cell Type, Level Three Cell Type, Original Cell Type) to a dictionary where the key is the level and the value is a list of unique cell types.
endothelial_cell_types = {
    'Original Cell Type': df_all_nodes[df_all_nodes['Original Cell Type'].str.contains("endothelial", case=False, na=False)]['Original Cell Type'].unique().tolist(),
    'Level Three Cell Type': df_all_nodes[df_all_nodes['Level Three Cell Type'].str.contains("endothelial", case=False, na=False)]['Level Three Cell Type'].unique().tolist(),
    'Level Two Cell Type': df_all_nodes[df_all_nodes['Level Two Cell Type'].str.contains("endothelial", case=False, na=False)]['Level Two Cell Type'].unique().tolist(),
    'Level One Cell Type': df_all_nodes[df_all_nodes['Level One Cell Type'].str.contains("endothelial", case=False, na=False)]['Level One Cell Type'].unique().tolist()
}

print("\nEndothelial cell types per cell type annotation level:")
for level, cell_types in endothelial_cell_types.items():
    print(f"\n{level}:")
    for cell in cell_types:
        print(f"  - {cell}")

Endothelial cell types per cell type annotation level:

Original Cell Type:
  - Vascular Endothelial Cells
  - Lymphatic Endothelial Cells

Level Three Cell Type:
  - endothelial cell of vascular tree
  - endothelial cell of lymphatic vessel

Level Two Cell Type:
  - endothelial cell of vascular tree
  - endothelial cell of lymphatic vessel

Level One Cell Type:
  - endothelial cell

type_field_list = ["Level Three Cell Type", "Level Two Cell Type", "Level One Cell Type"] # Skipping Original Cell Type as it is not a hierarchical level.

# Define the anchor cell type (type of endothelial cell) for each level in type_field_list based on available categories in the previous cell. The distance analysis at all three levels will be limited to the specified anchor cell type.
anchor_cell_type_dict = {
    'Level Three Cell Type': 'endothelial cell of vascular tree',
    'Level Two Cell Type': 'endothelial cell of vascular tree',
    'Level One Cell Type': 'endothelial cell'
}

Process datasets to add region information to Nodes files.

# Create a dictionary to map oral cavity regions to correct region names.
region_map = {'Buccal_Mucosa_And_Minor_Salivary_Glands_1':'Buccal Mucosa And Minor Salivary Glands',
 'Buccal_Mucosa_And_Minor_Salivary_Glands_2':'Buccal Mucosa And Minor Salivary Glands', 
 'Gingiva_ID1_shiny':'Gingiva',
 'Gingiva_ID5_shiny':'Gingiva', 
 'Parotid_1':'Parotid', 
 'Parotid_2':'Parotid', 
 'Parotid_3':'Parotid', 
 'Parotid_4':'Parotid',
 'Submandibular_1':'Submandibular', 
 'Submandibular_2':'Submandibular', 
 'Tongue_3':'Tongue', 
 'Tongue__1':'Tongue', 
 'Tongue__2':'Tongue'
}

df_all_nodes.head()

	x	y	Original Cell Type	Level Three Cell Type	Level Three CL Label	Level Three CL ID	CL_Match/3	Level Two Cell Type	Level Two CL Label	Level Two CL ID	CL_Match/2	Level One Cell Type	Level One CL Label	Level One CL ID	CL_Match/1	Dataset
0	2466.85	580.95	Others	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	Buccal_Mucosa_And_Minor_Salivary_Glands_1
1	2421.30	629.05	Others	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	Buccal_Mucosa_And_Minor_Salivary_Glands_1
2	2459.00	681.80	Others	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	Buccal_Mucosa_And_Minor_Salivary_Glands_1
3	2418.20	700.00	Others	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	Buccal_Mucosa_And_Minor_Salivary_Glands_1
4	2444.70	768.30	Treg	t cell:regulatory	regulatory T cell	CL:0000815	skos:exactMatch	t cell	T cell	CL:0000084	skos:exactMatch	immune cell	leukocyte	CL:0000738	skos:exactMatch	Buccal_Mucosa_And_Minor_Salivary_Glands_1

# Iterate through the df_all_data dataframe to create new column "Unique Region" based on the "Dataset" column.
# The "Unique Region" column is created by mapping the region names based on the full dataset name.
df_all_nodes['Unique Region'] = df_all_nodes['Dataset'].map(region_map)
# df_all_nodes['Unique Region'] = df_all_nodes['Dataset'].str.split('-').str[1].map(region_map)

# Check if the new columns are created correctly.
df_all_nodes[['Dataset', 'Unique Region']].head(5)

	Dataset	Unique Region
0	Buccal_Mucosa_And_Minor_Salivary_Glands_1	Buccal Mucosa And Minor Salivary Glands
1	Buccal_Mucosa_And_Minor_Salivary_Glands_1	Buccal Mucosa And Minor Salivary Glands
2	Buccal_Mucosa_And_Minor_Salivary_Glands_1	Buccal Mucosa And Minor Salivary Glands
3	Buccal_Mucosa_And_Minor_Salivary_Glands_1	Buccal Mucosa And Minor Salivary Glands
4	Buccal_Mucosa_And_Minor_Salivary_Glands_1	Buccal Mucosa And Minor Salivary Glands

# Print all unique regions in the data.
print("\nUnique Regions in the data:")
print(df_all_nodes['Unique Region'].unique())

# Print the total number of unique regions.
print(f"Total number of unique regions: {df_all_nodes['Unique Region'].nunique()}")

# Print number of unique datasets per unique region.
print("\nNumber of unique datasets per unique region:")
for region in df_all_nodes['Unique Region'].unique():
    num_datasets = df_all_nodes[df_all_nodes['Unique Region'] == region]['Dataset'].nunique()
    print(f"{region}: {num_datasets}")

Unique Regions in the data:
['Buccal Mucosa And Minor Salivary Glands' 'Gingiva' 'Parotid'
 'Submandibular' 'Tongue']
Total number of unique regions: 5

Number of unique datasets per unique region:
Buccal Mucosa And Minor Salivary Glands: 2
Gingiva: 2
Parotid: 4
Submandibular: 2
Tongue: 3

Process datasets to add region information to Edges files.

df_all_edges.head(5)

	cell_id	x1	y1	z1	x2	y2	z2	Distance	Dataset	Anchor Cell Type	Anchor Cell Type Level
0	0	2466.85	580.950	0	2600.55	598.700	0	134.873098	Buccal_Mucosa_And_Minor_Salivary_Glands_1	endothelial cell of vascular tree	Level Three Cell Type
1	759	2595.45	407.850	0	2689.10	404.285	0	93.717830	Buccal_Mucosa_And_Minor_Salivary_Glands_1	endothelial cell of vascular tree	Level Three Cell Type
2	782	2599.45	413.290	0	2689.10	404.285	0	90.101124	Buccal_Mucosa_And_Minor_Salivary_Glands_1	endothelial cell of vascular tree	Level Three Cell Type
3	840	2588.60	414.555	0	2689.10	404.285	0	101.023378	Buccal_Mucosa_And_Minor_Salivary_Glands_1	endothelial cell of vascular tree	Level Three Cell Type
4	869	2583.50	419.190	0	2689.10	404.285	0	106.646702	Buccal_Mucosa_And_Minor_Salivary_Glands_1	endothelial cell of vascular tree	Level Three Cell Type

# Process the edge data to create new columns "Unique Region" based on the "Dataset" column, similar to how it was done for the node data.
df_all_edges['Unique Region'] = df_all_edges['Dataset'].map(region_map)

# Check if the new columns are created correctly.
df_all_edges[['Dataset', 'Unique Region']].head(5)

	Dataset	Unique Region
0	Buccal_Mucosa_And_Minor_Salivary_Glands_1	Buccal Mucosa And Minor Salivary Glands
1	Buccal_Mucosa_And_Minor_Salivary_Glands_1	Buccal Mucosa And Minor Salivary Glands
2	Buccal_Mucosa_And_Minor_Salivary_Glands_1	Buccal Mucosa And Minor Salivary Glands
3	Buccal_Mucosa_And_Minor_Salivary_Glands_1	Buccal Mucosa And Minor Salivary Glands
4	Buccal_Mucosa_And_Minor_Salivary_Glands_1	Buccal Mucosa And Minor Salivary Glands

# Print all unique regions in the data.
print("\nUnique Regions in the data:")
print(df_all_edges['Unique Region'].unique())

# Print the total number of unique regions.
print(f"Total number of unique regions: {df_all_edges['Unique Region'].nunique()}")

# Print number of unique datasets per unique region.
print("\nNumber of unique datasets per unique region:")
for region in df_all_edges['Unique Region'].unique():
    num_datasets = df_all_edges[df_all_edges['Unique Region'] == region]['Dataset'].nunique()
    print(f"{region}: {num_datasets}")

Unique Regions in the data:
['Buccal Mucosa And Minor Salivary Glands' 'Gingiva' 'Parotid'
 'Submandibular' 'Tongue']
Total number of unique regions: 5

Number of unique datasets per unique region:
Buccal Mucosa And Minor Salivary Glands: 2
Gingiva: 2
Parotid: 4
Submandibular: 2
Tongue: 3

df_all_edges_with_cell_types['Unique Region'] = df_all_edges_with_cell_types['Dataset'].map(region_map)

# Check if the new columns are created correctly.
df_all_edges_with_cell_types[['Dataset', 'Unique Region']].head(5)

	Dataset	Unique Region
0	Buccal_Mucosa_And_Minor_Salivary_Glands_1	Buccal Mucosa And Minor Salivary Glands
1	Buccal_Mucosa_And_Minor_Salivary_Glands_1	Buccal Mucosa And Minor Salivary Glands
2	Buccal_Mucosa_And_Minor_Salivary_Glands_1	Buccal Mucosa And Minor Salivary Glands
3	Buccal_Mucosa_And_Minor_Salivary_Glands_1	Buccal Mucosa And Minor Salivary Glands
4	Buccal_Mucosa_And_Minor_Salivary_Glands_1	Buccal Mucosa And Minor Salivary Glands

df_all_nodes.head(1)

	x	y	Original Cell Type	Level Three Cell Type	Level Three CL Label	Level Three CL ID	CL_Match/3	Level Two Cell Type	Level Two CL Label	Level Two CL ID	CL_Match/2	Level One Cell Type	Level One CL Label	Level One CL ID	CL_Match/1	Dataset	Unique Region
0	2466.85	580.95	Others	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	unknown cell	cell:unknown	CL:0000000	skos:narrowMatch	Buccal_Mucosa_And_Minor_Salivary_Glands_1	Buccal Mucosa And Minor Salivary Glands

df_all_edges.head(1)

	cell_id	x1	y1	z1	x2	y2	z2	Distance	Dataset	Anchor Cell Type	Anchor Cell Type Level	Unique Region
0	0	2466.85	580.95	0	2600.55	598.7	0	134.873098	Buccal_Mucosa_And_Minor_Salivary_Glands_1	endothelial cell of vascular tree	Level Three Cell Type	Buccal Mucosa And Minor Salivary Glands

df_all_edges_with_cell_types.head(1)

	cell_id	x1	y1	z1	x2	y2	z2	Distance	Dataset	Anchor Cell Type	Anchor Cell Type Level	Level Three Cell Type	Level Two Cell Type	Level One Cell Type	Unique Region
0	0	2466.85	580.95	0	2600.55	598.7	0	134.873098	Buccal_Mucosa_And_Minor_Salivary_Glands_1	endothelial cell of vascular tree	Level Three Cell Type	unknown cell	unknown cell	unknown cell	Buccal Mucosa And Minor Salivary Glands

Node Analysis

# Plot number of cells per cell type in the same plot. Color by cell type and unique region. Output figure saved in existing `figures_output_dir`.
def plot_cells_per_celltype(df, type_field, output_dir):
    plt.figure(figsize=(20, 8))
    plt.rcParams["svg.fonttype"] = 'none'  # to store text as text, not as path
    sns.countplot(data=df, x=type_field, palette='Spectral', hue='Unique Region')
    plt.title(f'Number of Cells per {type_field} in `{dataset_dir}`')
    plt.xticks(rotation=90)
    plt.tight_layout()
    plt.savefig(os.path.join(output_dir, f'{dataset_dir}_cells_per_celltype_{type_field}.png'), dpi=300,
                bbox_inches='tight',
                pad_inches=0.5)
    plt.savefig(os.path.join(output_dir, f'{dataset_dir}_cells_per_celltype_{type_field}.svg'), dpi=300,
                bbox_inches='tight',
                pad_inches=0.5)
    plt.legend(title='Unique Region', bbox_to_anchor=(0.85, 1), loc='upper left')
    plt.xlabel(type_field)

    # For numbers on y-axis, do not use scientific notation.
    plt.ticklabel_format(style='plain', axis='y')
    # Set y-axis label
    plt.ylabel('Number of Cells')
    plt.grid(axis='y', linestyle='--', alpha=0.7)
    plt.tight_layout()
    # Show the plot
    plt.show()
    plt.close()
for type_field in type_field_list:
    plot_cells_per_celltype(df_all_nodes, type_field, os.path.join(basepath, figures_output_dir))

Distance Analysis

# Get mean, median, minimum, maximum distance per unique region per anchor cell type.
df_distance_stats = df_all_edges_with_cell_types.groupby(['Unique Region', 'Anchor Cell Type', 'Anchor Cell Type Level']).agg(
    mean_distance=('Distance', 'mean'),
    median_distance=('Distance', 'median'),
    min_distance=('Distance', 'min'),
    max_distance=('Distance', 'max')
).reset_index()
# Print the first few rows of the distance statistics DataFrame.
df_distance_stats

	Unique Region	Anchor Cell Type	Anchor Cell Type Level	mean_distance	median_distance	min_distance	max_distance
0	Buccal Mucosa And Minor Salivary Glands	endothelial cell	Level One Cell Type	32.788252	22.583456	2.350532	199.920297
1	Buccal Mucosa And Minor Salivary Glands	endothelial cell of lymphatic vessel	Level Three Cell Type	66.570639	56.315739	2.400521	199.999181
2	Buccal Mucosa And Minor Salivary Glands	endothelial cell of lymphatic vessel	Level Two Cell Type	66.570639	56.315739	2.400521	199.999181
3	Buccal Mucosa And Minor Salivary Glands	endothelial cell of vascular tree	Level Three Cell Type	41.164970	28.331828	2.350532	199.920297
4	Buccal Mucosa And Minor Salivary Glands	endothelial cell of vascular tree	Level Two Cell Type	41.164970	28.331828	2.350532	199.920297
5	Gingiva	endothelial cell	Level One Cell Type	46.035025	35.977739	0.006517	199.932087
6	Gingiva	endothelial cell of lymphatic vessel	Level Three Cell Type	118.082543	121.622867	0.466601	199.993077
7	Gingiva	endothelial cell of lymphatic vessel	Level Two Cell Type	118.082543	121.622867	0.466601	199.993077
8	Gingiva	endothelial cell of vascular tree	Level Three Cell Type	46.818074	36.442616	0.006517	199.932087
9	Gingiva	endothelial cell of vascular tree	Level Two Cell Type	46.818074	36.442616	0.006517	199.932087
10	Parotid	endothelial cell	Level One Cell Type	27.287024	22.853827	0.004528	199.996540
11	Parotid	endothelial cell of lymphatic vessel	Level Three Cell Type	102.361408	100.972133	0.056988	199.999212
12	Parotid	endothelial cell of lymphatic vessel	Level Two Cell Type	102.361408	100.972133	0.056988	199.999212
13	Parotid	endothelial cell of vascular tree	Level Three Cell Type	28.225257	23.337365	0.004528	199.996540
14	Parotid	endothelial cell of vascular tree	Level Two Cell Type	28.225257	23.337365	0.004528	199.996540
15	Submandibular	endothelial cell	Level One Cell Type	30.266208	23.193534	2.540177	199.943992
16	Submandibular	endothelial cell of lymphatic vessel	Level Three Cell Type	102.248432	99.812887	2.555386	199.999931
17	Submandibular	endothelial cell of lymphatic vessel	Level Two Cell Type	102.248432	99.812887	2.555386	199.999931
18	Submandibular	endothelial cell of vascular tree	Level Three Cell Type	30.935485	23.700211	2.540177	199.943992
19	Submandibular	endothelial cell of vascular tree	Level Two Cell Type	30.935485	23.700211	2.540177	199.943992
20	Tongue	endothelial cell	Level One Cell Type	37.316385	25.104034	2.438237	199.998506
21	Tongue	endothelial cell of lymphatic vessel	Level Three Cell Type	66.095612	56.164157	2.815138	199.994900
22	Tongue	endothelial cell of lymphatic vessel	Level Two Cell Type	66.095612	56.164157	2.815138	199.994900
23	Tongue	endothelial cell of vascular tree	Level Three Cell Type	48.574001	32.080680	2.438237	199.999050
24	Tongue	endothelial cell of vascular tree	Level Two Cell Type	48.574001	32.080680	2.438237	199.999050

Level One Cell Type Analysis

# Get mean, median, minimum, maximum distance per cell type in all unique regions.
cell_type_level = 'Level One Cell Type'
df_all_edges_with_cell_type_level = df_all_edges_with_cell_types[(df_all_edges_with_cell_types['Anchor Cell Type Level'] == cell_type_level) & (df_all_edges_with_cell_types['Anchor Cell Type'] == anchor_cell_type_dict[cell_type_level])]

df_distance_stats_cell_type_level = df_all_edges_with_cell_type_level.groupby([cell_type_level, 'Unique Region']).agg(
    mean_distance=('Distance', 'mean'),
    median_distance=('Distance', 'median'),
    min_distance=('Distance', 'min'),
    max_distance=('Distance', 'max')
).reset_index()
df_distance_stats_cell_type_level

	Level One Cell Type	Unique Region	mean_distance	median_distance	min_distance	max_distance
0	epithelial cell	Buccal Mucosa And Minor Salivary Glands	47.227324	39.939347	2.451530	199.035857
1	epithelial cell	Gingiva	44.140615	34.768144	0.010330	199.836547
2	epithelial cell	Parotid	26.585115	22.840156	0.017954	199.947587
3	epithelial cell	Submandibular	23.907224	20.577384	2.755449	199.833787
4	epithelial cell	Tongue	85.775049	79.378846	2.815138	199.990006
5	immune cell	Buccal Mucosa And Minor Salivary Glands	24.581746	18.289273	2.400521	198.825866
6	immune cell	Gingiva	52.854768	40.612195	0.029228	199.932087
7	immune cell	Parotid	29.196625	22.425217	0.004528	199.996540
8	immune cell	Submandibular	27.443867	22.173182	2.549510	190.454122
9	immune cell	Tongue	26.795128	18.708086	2.800446	199.876143
10	mesenchymal cell	Buccal Mucosa And Minor Salivary Glands	25.865052	18.751667	2.510976	190.343295
11	mesenchymal cell	Gingiva	41.454674	34.134295	0.127369	196.036100
12	mesenchymal cell	Parotid	24.717051	20.026233	0.009735	199.030058
13	mesenchymal cell	Submandibular	42.638145	32.175398	2.540177	199.943992
14	mesenchymal cell	Tongue	35.806762	26.742008	2.530316	199.998506
15	neural cell	Parotid	24.032759	20.332039	0.124848	191.137047
16	unknown cell	Buccal Mucosa And Minor Salivary Glands	37.841632	25.530619	2.350532	199.920297
17	unknown cell	Gingiva	46.744610	36.358142	0.006517	199.814472
18	unknown cell	Parotid	30.671119	24.682478	0.015375	199.882778
19	unknown cell	Submandibular	31.722377	25.000625	2.720294	199.821445
20	unknown cell	Tongue	27.503129	20.065954	2.438237	199.924628

# Get top five and bottom five cell types with respect to mean distance in each unique region separately.
def get_top_bottom_cell_types_by_mean(df, cell_type_level, unique_region, top_n=5):
    # Filter the DataFrame for the specified unique region and cell type level
    df_filtered = df[df['Unique Region'] == unique_region]

    # Group by the specified cell type level and calculate mean distance
    df_grouped = df_filtered.groupby(cell_type_level).agg(mean_distance=('Distance', 'mean')).reset_index()
    
    # Sort by mean distance to get top and bottom cell types
    df_sorted = df_grouped.sort_values(by='mean_distance', ascending=False)
    
    # Get top N and bottom N cell types
    top_cell_types = df_sorted.head(top_n)
    bottom_cell_types = df_sorted.tail(top_n)
    
    return top_cell_types, bottom_cell_types

# Get top and bottom cell types for each unique region in the dataset.
unique_regions = df_all_edges_with_cell_type_level['Unique Region'].unique()
for region in unique_regions:
    top_bottom = get_top_bottom_cell_types_by_mean(df_all_edges_with_cell_type_level, cell_type_level, region)
    print(f"\nTop 5 cell types in {region}:")
    print(top_bottom[0])
    print(f"\nBottom 5 cell types in {region}:")
    print(top_bottom[1])

Top 5 cell types in Buccal Mucosa And Minor Salivary Glands:
  Level One Cell Type  mean_distance
0     epithelial cell      47.227324
3        unknown cell      37.841632
2    mesenchymal cell      25.865052
1         immune cell      24.581746

Bottom 5 cell types in Buccal Mucosa And Minor Salivary Glands:
  Level One Cell Type  mean_distance
0     epithelial cell      47.227324
3        unknown cell      37.841632
2    mesenchymal cell      25.865052
1         immune cell      24.581746

Top 5 cell types in Gingiva:
  Level One Cell Type  mean_distance
1         immune cell      52.854768
3        unknown cell      46.744610
0     epithelial cell      44.140615
2    mesenchymal cell      41.454674

Bottom 5 cell types in Gingiva:
  Level One Cell Type  mean_distance
1         immune cell      52.854768
3        unknown cell      46.744610
0     epithelial cell      44.140615
2    mesenchymal cell      41.454674

Top 5 cell types in Parotid:
  Level One Cell Type  mean_distance
4        unknown cell      30.671119
1         immune cell      29.196625
0     epithelial cell      26.585115
2    mesenchymal cell      24.717051
3         neural cell      24.032759

Bottom 5 cell types in Parotid:
  Level One Cell Type  mean_distance
4        unknown cell      30.671119
1         immune cell      29.196625
0     epithelial cell      26.585115
2    mesenchymal cell      24.717051
3         neural cell      24.032759

Top 5 cell types in Submandibular:
  Level One Cell Type  mean_distance
2    mesenchymal cell      42.638145
3        unknown cell      31.722377
1         immune cell      27.443867
0     epithelial cell      23.907224

Bottom 5 cell types in Submandibular:
  Level One Cell Type  mean_distance
2    mesenchymal cell      42.638145
3        unknown cell      31.722377
1         immune cell      27.443867
0     epithelial cell      23.907224

Top 5 cell types in Tongue:
  Level One Cell Type  mean_distance
0     epithelial cell      85.775049
2    mesenchymal cell      35.806762
3        unknown cell      27.503129
1         immune cell      26.795128

Bottom 5 cell types in Tongue:
  Level One Cell Type  mean_distance
0     epithelial cell      85.775049
2    mesenchymal cell      35.806762
3        unknown cell      27.503129
1         immune cell      26.795128

# Get top five and bottom five cell types with respect to median distance in each unique region separately.
def get_top_bottom_cell_types_by_median(df, cell_type_level, unique_region, top_n=5):
    # Filter the DataFrame for the specified unique region and cell type level
    df_filtered = df[df['Unique Region'] == unique_region]

    # Group by the specified cell type level and calculate median distance
    df_grouped = df_filtered.groupby(cell_type_level).agg(median_distance=('Distance', 'median')).reset_index()

    # Sort by median distance to get top and bottom cell types
    df_sorted = df_grouped.sort_values(by='median_distance', ascending=False)

    # Get top N and bottom N cell types
    top_cell_types = df_sorted.head(top_n)
    bottom_cell_types = df_sorted.tail(top_n)
    
    return top_cell_types, bottom_cell_types

# Get top and bottom cell types for each unique region in the dataset.
unique_regions = df_all_edges_with_cell_type_level['Unique Region'].unique()
for region in unique_regions:
    top_bottom = get_top_bottom_cell_types_by_median(df_all_edges_with_cell_type_level, cell_type_level, region)
    print(f"\nTop 5 cell types in {region}:")
    print(top_bottom[0])
    print(f"\nBottom 5 cell types in {region}:")
    print(top_bottom[1])

Top 5 cell types in Buccal Mucosa And Minor Salivary Glands:
  Level One Cell Type  median_distance
0     epithelial cell        39.939347
3        unknown cell        25.530619
2    mesenchymal cell        18.751667
1         immune cell        18.289273

Bottom 5 cell types in Buccal Mucosa And Minor Salivary Glands:
  Level One Cell Type  median_distance
0     epithelial cell        39.939347
3        unknown cell        25.530619
2    mesenchymal cell        18.751667
1         immune cell        18.289273

Top 5 cell types in Gingiva:
  Level One Cell Type  median_distance
1         immune cell        40.612195
3        unknown cell        36.358142
0     epithelial cell        34.768144
2    mesenchymal cell        34.134295

Bottom 5 cell types in Gingiva:
  Level One Cell Type  median_distance
1         immune cell        40.612195
3        unknown cell        36.358142
0     epithelial cell        34.768144
2    mesenchymal cell        34.134295

Top 5 cell types in Parotid:
  Level One Cell Type  median_distance
4        unknown cell        24.682478
0     epithelial cell        22.840156
1         immune cell        22.425217
3         neural cell        20.332039
2    mesenchymal cell        20.026233

Bottom 5 cell types in Parotid:
  Level One Cell Type  median_distance
4        unknown cell        24.682478
0     epithelial cell        22.840156
1         immune cell        22.425217
3         neural cell        20.332039
2    mesenchymal cell        20.026233

Top 5 cell types in Submandibular:
  Level One Cell Type  median_distance
2    mesenchymal cell        32.175398
3        unknown cell        25.000625
1         immune cell        22.173182
0     epithelial cell        20.577384

Bottom 5 cell types in Submandibular:
  Level One Cell Type  median_distance
2    mesenchymal cell        32.175398
3        unknown cell        25.000625
1         immune cell        22.173182
0     epithelial cell        20.577384

Top 5 cell types in Tongue:
  Level One Cell Type  median_distance
0     epithelial cell        79.378846
2    mesenchymal cell        26.742008
3        unknown cell        20.065954
1         immune cell        18.708086

Bottom 5 cell types in Tongue:
  Level One Cell Type  median_distance
0     epithelial cell        79.378846
2    mesenchymal cell        26.742008
3        unknown cell        20.065954
1         immune cell        18.708086

# Calculate regional variability
def calculate_regional_variability(df_all_edges_with_cell_type_level, cell_type_level):
    """    Calculate regional variability for distances in the given DataFrame.
    """
    regional_variability = df_all_edges_with_cell_type_level.groupby('Unique Region')['Distance'].agg([
        ('mean', 'mean'),
        ('std', 'std')
    ]).round(2)

    # Add CV as percentage
    regional_variability['CV (%)'] = (regional_variability['std'] / regional_variability['mean'] * 100).round(1)

    print("\nRegional Variability Analysis:")
    print("Mean: Average distance in each region")
    print("Std: Standard deviation of distances")
    print("CV: Coefficient of Variation (std/mean * 100%)")
    print(regional_variability)

    # Calculate variability for each cell type
    cell_type_variability = df_all_edges_with_cell_type_level.groupby(cell_type_level)['Distance'].agg([
        ('mean', 'mean'),
        ('std', 'std')
    ]).round(2)

    # Add CV as percentage
    cell_type_variability['CV (%)'] = (cell_type_variability['std'] / cell_type_variability['mean'] * 100).round(1)

    print("\nCell Type Variability Analysis (sorted by CV):")
    print(cell_type_variability.sort_values('CV (%)', ascending=False))

calculate_regional_variability(df_all_edges_with_cell_type_level, cell_type_level)

Regional Variability Analysis:
Mean: Average distance in each region
Std: Standard deviation of distances
CV: Coefficient of Variation (std/mean * 100%)
                                          mean    std  CV (%)
Unique Region                                                
Buccal Mucosa And Minor Salivary Glands  32.79  30.43    92.8
Gingiva                                  46.04  35.92    78.0
Parotid                                  27.29  20.14    73.8
Submandibular                            30.27  25.72    85.0
Tongue                                   37.32  34.85    93.4

Cell Type Variability Analysis (sorted by CV):
                      mean    std  CV (%)
Level One Cell Type                      
epithelial cell      31.24  27.49    88.0
unknown cell         33.97  29.68    87.4
immune cell          28.45  24.84    87.3
mesenchymal cell     35.45  30.49    86.0
neural cell          24.03  18.50    77.0

# Define the standard region sequence for plots
regions = ['Gingiva', 'Submandibular', 'Parotid', 'Tongue', 'Buccal Mucosa And Minor Salivary Glands']

# Generate Violin Plot
def plot_violin_cells_per_celltype(df_all_edges_with_cell_type_level, cell_type_level, output_dir, density_norm='area'):
    sns.set_style("whitegrid")
    sns.set_context("notebook", rc={"grid.linewidth": 2})
    plt.figure(figsize=(10, 6))
    plt.rcParams["svg.fonttype"] = 'none'  # to store text as text, not as path

    sns.violinplot(data=df_all_edges_with_cell_type_level, x=cell_type_level, y="Distance", density_norm=density_norm, common_norm=True, cut=0, inner="box", split=False, palette='Spectral', alpha=.9)

    sns.set_theme(style="whitegrid")
    sns.set_context("paper")


    font_size = 10
    plt.legend(fontsize=font_size)

    plt.title(f'Violin Plot of distances by {cell_type_level} (Density Normalization: {density_norm})', fontsize=font_size)

    plt.xlabel(f'{cell_type_level}', fontsize=font_size)
    plt.ylabel('Distance (\u03bcm)', fontsize=font_size)

    # Increase font size for all text in the figure
    plt.xticks(fontsize=font_size)
    plt.xticks(rotation=90)
    plt.yticks(fontsize=font_size)

    plt.tight_layout()

    plt.savefig(os.path.join(output_dir, f'{dataset_dir}_violin_cells_per_celltype_{cell_type_level}.png'), dpi=300,
                bbox_inches='tight',
                pad_inches=0.5)
    plt.savefig(os.path.join(output_dir, f'{dataset_dir}_violin_cells_per_celltype_{cell_type_level}.svg'), dpi=300,
                bbox_inches='tight',
                pad_inches=0.5)
    plt.show()

plot_violin_cells_per_celltype(df_all_edges_with_cell_type_level, cell_type_level, os.path.join(basepath, figures_output_dir), density_norm='area')

# Boxplots of distribution of distances by cell type and region.
def plot_distance_distribution_boxplots_by_region(df_all_edges_with_cell_type_level, cell_type_level, output_dir):
    plt.figure(figsize=(16, 8))
    plt.rcParams["svg.fonttype"] = 'none'  # to store text as text, not as path
    # Create categorical type with only the regions that exist in the data
    available_regions = [r for r in regions if r in df_all_edges_with_cell_type_level['Unique Region'].unique()]
    df_all_edges_with_cell_type_level['Unique Region'] = pd.Categorical(
        df_all_edges_with_cell_type_level['Unique Region'],
        categories=available_regions,
        ordered=True
    )

    # Make box plot.
    sns.boxplot(data=df_all_edges_with_cell_type_level, x=cell_type_level, y='Distance', hue='Unique Region', showfliers=False, palette='Spectral') # viridis or Spectral palette for better color distinction
    font_size = 10
    plt.xticks(rotation=90, ha='right', fontsize=font_size)
    plt.yticks(fontsize=font_size)
    plt.title(f'Distribution of distances by {cell_type_level} and region', fontsize=font_size)
    plt.xlabel(f'{cell_type_level}', fontsize=font_size)
    plt.ylabel('Distance (\u03bcm)', fontsize=font_size)
    plt.legend(bbox_to_anchor=(1, 1), loc='upper left')
    plt.tight_layout()

    plt.savefig(os.path.join(output_dir, f'{dataset_dir}_distance_distribution_boxplots_by_region_{cell_type_level}.png'), dpi=300,
                    bbox_inches='tight',
                    pad_inches=0.5)
    plt.savefig(os.path.join(output_dir, f'{dataset_dir}_distance_distribution_boxplots_by_region_{cell_type_level}.svg'), dpi=300,
                    bbox_inches='tight',
                    pad_inches=0.5)
    plt.show()

plot_distance_distribution_boxplots_by_region(df_all_edges_with_cell_type_level, cell_type_level, os.path.join(basepath, figures_output_dir))

# Boxplots of distribution of distances by cell type and region.
def plot_distance_distribution_heatmap(df_all_edges_with_cell_type_level, cell_type_level, output_dir):
    pivot_data = df_all_edges_with_cell_type_level.pivot_table(
    values='Distance',
    index=cell_type_level,
    columns='Unique Region',
    aggfunc='median'
    )

    plt.figure(figsize=(15, 10))
    plt.rcParams["svg.fonttype"] = 'none'  # to store text as text, not as path
    sns.heatmap(pivot_data, annot=True, fmt='.1f', cmap='Spectral')
    plt.title(f'Heatmap of median distances by {cell_type_level}', fontsize=12)

    font_size = 10
    plt.xticks(rotation=90, ha='right', fontsize=font_size)
    plt.yticks(fontsize=font_size)

    plt.xlabel('Unique Region', fontsize=font_size)
    plt.ylabel(f'{cell_type_level}', fontsize=font_size)
    
    plt.tight_layout()

    plt.savefig(os.path.join(output_dir, f'{dataset_dir}_distance_distribution_heatmap_{cell_type_level}.png'), dpi=300,
                    bbox_inches='tight',
                    pad_inches=0.5)
    plt.savefig(os.path.join(output_dir, f'{dataset_dir}_distance_distribution_heatmap_{cell_type_level}.svg'), dpi=300,
                    bbox_inches='tight',
                    pad_inches=0.5)
    plt.show()

plot_distance_distribution_heatmap(df_all_edges_with_cell_type_level, cell_type_level, os.path.join(basepath, figures_output_dir))

# Generate Violin Plot per unique region in both small intestine and large intestine. Create for all 8 regions as 8 subplots.
def plot_violin_plots_all_regions(df_all_edges_with_cell_type_level, cell_type_level, output_dir, density_norm="area"):
    sns.set_style("whitegrid")
    sns.set_context("notebook", rc={"grid.linewidth": 1})
    plt.rcParams["svg.fonttype"] = 'none'  # to store text as text, not as path
    font_size = 10
    fig, axs = plt.subplots(5, 1, figsize=(10, 20)) # Adjusted figsize for horizontal layout
    fig.suptitle(f'Distance distribution per {cell_type_level} in `{dataset_dir}` (density normalization = {density_norm})', fontsize=font_size, y=1)

    # Keep the sequence of Cell Types consistent across plots.
    cell_types = sorted(df_all_edges_with_cell_type_level[cell_type_level].unique())

    # Create a color palette based on the number of unique classes
    color_palette = sns.color_palette("Spectral", n_colors=len(cell_types))

    # Create a dictionary mapping class to color
    class_color_dict = dict(zip(cell_types, color_palette))

    for i, region in enumerate(regions):
        data_reg = df_all_edges_with_cell_type_level[df_all_edges_with_cell_type_level['Unique Region'] == region]
        sns.violinplot(data=data_reg, x=cell_type_level, y="Distance", density_norm=density_norm, common_norm=True, cut=0, inner="box", split=False, palette=class_color_dict, alpha=.9, ax=axs[i], hue=cell_type_level, legend=False, order=cell_types, fill=True)
        axs[i].set_title(region, fontsize=font_size)
        axs[i].set_xlabel('', fontsize=font_size)
        axs[i].set_ylabel('Distance (\u03bcm)', fontsize=font_size)
        # axs[i].tick_params(axis='x', labelrotation=90, labelsize=font_size)
        # only show xtick labels for the last subplot
        if i < len(regions) - 1:
            axs[i].set_xticklabels([])
        else:
            axs[i].set_xticklabels(cell_types, fontsize=font_size, rotation=90, ha='right')
        # axs[i].set_ylim(0, data_reg['Distance'].max() * 1.1)  # Set y-limits to be consistent across all plots
        axs[i].tick_params(axis='both', labelsize=font_size)

    # Use fig.text for precise label positioning
    fig.figure.text(0.5, -0.02, f'{cell_type_level}', ha='center', va='bottom', fontsize=font_size)

    plt.tight_layout()

    plt.savefig(os.path.join(output_dir, f'{dataset_dir}_violin_plots_all_regions_{cell_type_level}.png'), dpi=300,
                    bbox_inches='tight',
                    pad_inches=0.5)
    plt.savefig(os.path.join(output_dir, f'{dataset_dir}_violin_plots_all_regions_{cell_type_level}.svg'), dpi=300,
                    bbox_inches='tight',
                    pad_inches=0.5)
    
    plt.show()

plot_violin_plots_all_regions(df_all_edges_with_cell_type_level, cell_type_level, os.path.join(basepath, figures_output_dir), density_norm="count") # density_norm="count" or "area" can be used based on preference.

Level Two Cell Type Analysis

# Get mean, median, minimum, maximum distance per cell type in all unique regions.
cell_type_level = 'Level Two Cell Type'
df_all_edges_with_cell_type_level = df_all_edges_with_cell_types[(df_all_edges_with_cell_types['Anchor Cell Type Level'] == cell_type_level) & (df_all_edges_with_cell_types['Anchor Cell Type'] == anchor_cell_type_dict[cell_type_level])]

df_distance_stats_cell_type_level = df_all_edges_with_cell_type_level.groupby([cell_type_level, 'Unique Region']).agg(
    mean_distance=('Distance', 'mean'),
    median_distance=('Distance', 'median'),
    min_distance=('Distance', 'min'),
    max_distance=('Distance', 'max')
).reset_index()
df_distance_stats_cell_type_level

	Level Two Cell Type	Unique Region	mean_distance	median_distance	min_distance	max_distance
0	adipocyte	Gingiva	38.921484	31.576881	1.472765	198.358252
1	b cell	Buccal Mucosa And Minor Salivary Glands	23.196496	22.353669	2.804015	98.284549
2	b cell	Gingiva	38.913598	29.370338	1.271453	198.729570
3	b cell	Parotid	22.649857	19.314031	0.004528	195.393503
4	b cell	Submandibular	49.468888	45.465234	3.304921	184.012207
5	b cell	Tongue	23.064363	15.471976	3.696282	119.542137
6	dendritic cell	Buccal Mucosa And Minor Salivary Glands	23.820644	19.333714	2.753634	176.453535
7	dendritic cell	Gingiva	53.327090	40.886464	0.225297	199.932087
8	dendritic cell	Parotid	21.289953	19.328978	0.050563	154.155788
9	dendritic cell	Tongue	49.187117	32.105607	3.162278	199.890432
10	endothelial cell of lymphatic vessel	Buccal Mucosa And Minor Salivary Glands	31.266348	23.013311	3.036445	188.564690
11	endothelial cell of lymphatic vessel	Gingiva	51.530766	47.521442	8.561243	143.755112
12	endothelial cell of lymphatic vessel	Parotid	27.392054	19.534840	0.253150	181.675452
13	endothelial cell of lymphatic vessel	Submandibular	23.177030	17.112130	3.412477	121.602436
14	endothelial cell of lymphatic vessel	Tongue	44.692616	34.740934	4.208622	199.544105
15	epithelial cell	Buccal Mucosa And Minor Salivary Glands	69.005527	63.242607	3.006659	199.616482
16	epithelial cell	Gingiva	49.671535	37.195439	1.752795	198.377561
17	epithelial cell	Parotid	39.288368	29.149386	0.468023	199.229374
18	epithelial cell	Tongue	116.777509	126.256991	3.323402	199.987850
19	fibroblast	Buccal Mucosa And Minor Salivary Glands	35.795373	25.141972	2.510976	199.696019
20	fibroblast	Gingiva	42.361864	34.901074	0.127369	198.612028
21	fibroblast	Parotid	26.934849	21.545359	0.009735	199.030058
22	fibroblast	Submandibular	43.476570	33.036249	2.540177	199.943992
23	fibroblast	Tongue	52.144835	37.926541	2.530316	199.999050
24	gland epithelium cell	Gingiva	38.872536	31.698348	0.029614	199.675802
25	gland epithelium cell	Parotid	26.186293	22.583455	0.017954	199.567157
26	gland epithelium cell	Submandibular	25.000346	21.156766	2.773536	199.833787
27	keratinocyte	Gingiva	53.520737	42.797847	0.010330	199.836547
28	keratinocyte	Parotid	29.006264	23.250918	0.042787	199.947587
29	langerhans cell	Gingiva	35.348131	26.245455	0.353423	166.645525
30	langerhans cell	Parotid	21.515277	18.679420	0.118568	151.121503
31	macrophage	Buccal Mucosa And Minor Salivary Glands	25.882874	20.110103	2.995413	175.639439
32	macrophage	Gingiva	43.967814	35.395149	0.362944	196.376846
33	macrophage	Parotid	27.069626	22.910843	0.100367	196.269036
34	macrophage	Submandibular	30.757152	23.417995	3.103627	190.454122
35	macrophage	Tongue	34.170755	22.358667	2.800446	199.962253
36	mast cell	Gingiva	45.118725	32.004082	0.283616	198.023632
37	mast cell	Parotid	55.949381	42.159777	0.497809	164.482230
38	muscle cell	Parotid	26.374298	22.262380	0.246935	196.110438
39	myoepithelial cell	Gingiva	33.597314	25.924170	0.499390	195.159907
40	myoepithelial cell	Parotid	31.860121	26.601561	0.027328	199.863760
41	myoepithelial cell	Submandibular	23.571161	20.641130	2.755449	136.143821
42	natural killer cell	Buccal Mucosa And Minor Salivary Glands	46.769140	37.025667	3.381198	195.615196
43	natural killer cell	Gingiva	41.823514	29.657322	2.560580	189.288209
44	natural killer cell	Parotid	63.315905	54.364997	0.139128	198.587437
45	natural killer cell	Submandibular	25.280999	20.705518	3.148412	158.294820
46	natural killer cell	Tongue	47.527685	42.494294	3.064719	197.964770
47	neuroglial cell/neuron	Parotid	24.591434	20.583996	0.124848	191.137047
48	neutrophil	Buccal Mucosa And Minor Salivary Glands	30.028958	21.579446	2.554408	194.583330
49	neutrophil	Gingiva	46.213377	32.192202	0.029228	199.542077
50	neutrophil	Parotid	44.659693	34.148935	18.784102	87.823018
51	neutrophil	Submandibular	21.857157	19.521302	2.968164	164.314371
52	neutrophil	Tongue	34.321570	22.459853	3.085855	199.231084
53	perivascular cell	Parotid	21.881249	18.707282	0.068190	166.700706
54	t cell	Buccal Mucosa And Minor Salivary Glands	38.797999	28.289618	3.026549	198.825866
55	t cell	Gingiva	61.869930	49.247660	0.094913	199.784633
56	t cell	Parotid	40.798968	31.956611	0.043842	199.996540
57	t cell	Submandibular	31.282386	27.361812	2.549510	187.306834
58	t cell	Tongue	60.348191	45.091157	2.951694	199.994831
59	unknown cell	Buccal Mucosa And Minor Salivary Glands	42.374333	28.831450	2.350532	199.920297
60	unknown cell	Gingiva	47.466793	36.770012	0.006517	199.814472
61	unknown cell	Parotid	32.154130	25.431573	0.015375	199.882778
62	unknown cell	Submandibular	32.777522	25.710163	2.720294	199.821445
63	unknown cell	Tongue	33.290598	23.558730	2.438237	199.998125

# Get top and bottom cell types for each unique region in the dataset.
unique_regions = df_all_edges_with_cell_type_level['Unique Region'].unique()
for region in unique_regions:
    top_bottom = get_top_bottom_cell_types_by_mean(df_all_edges_with_cell_type_level, cell_type_level, region)
    print(f"\nTop 5 cell types in {region}:")
    print(top_bottom[0])
    print(f"\nBottom 5 cell types in {region}:")
    print(top_bottom[1])

Top 5 cell types in Buccal Mucosa And Minor Salivary Glands:
   Level Two Cell Type  mean_distance
3      epithelial cell      69.005527
6  natural killer cell      46.769140
9         unknown cell      42.374333
8               t cell      38.797999
4           fibroblast      35.795373

Bottom 5 cell types in Buccal Mucosa And Minor Salivary Glands:
                    Level Two Cell Type  mean_distance
2  endothelial cell of lymphatic vessel      31.266348
7                            neutrophil      30.028958
5                            macrophage      25.882874
1                        dendritic cell      23.820644
0                                b cell      23.196496

Top 5 cell types in Gingiva:
                     Level Two Cell Type  mean_distance
14                                t cell      61.869930
7                           keratinocyte      53.520737
2                         dendritic cell      53.327090
3   endothelial cell of lymphatic vessel      51.530766
4                        epithelial cell      49.671535

Bottom 5 cell types in Gingiva:
      Level Two Cell Type  mean_distance
0               adipocyte      38.921484
1                  b cell      38.913598
6   gland epithelium cell      38.872536
8         langerhans cell      35.348131
11     myoepithelial cell      33.597314

Top 5 cell types in Parotid:
    Level Two Cell Type  mean_distance
12  natural killer cell      63.315905
9             mast cell      55.949381
14           neutrophil      44.659693
16               t cell      40.798968
3       epithelial cell      39.288368

Bottom 5 cell types in Parotid:
       Level Two Cell Type  mean_distance
13  neuroglial cell/neuron      24.591434
0                   b cell      22.649857
15       perivascular cell      21.881249
7          langerhans cell      21.515277
1           dendritic cell      21.289953

Top 5 cell types in Submandibular:
  Level Two Cell Type  mean_distance
0              b cell      49.468888
2          fibroblast      43.476570
9        unknown cell      32.777522
8              t cell      31.282386
4          macrophage      30.757152

Bottom 5 cell types in Submandibular:
                    Level Two Cell Type  mean_distance
6                   natural killer cell      25.280999
3                 gland epithelium cell      25.000346
5                    myoepithelial cell      23.571161
1  endothelial cell of lymphatic vessel      23.177030
7                            neutrophil      21.857157

Top 5 cell types in Tongue:
   Level Two Cell Type  mean_distance
3      epithelial cell     116.777509
8               t cell      60.348191
4           fibroblast      52.144835
1       dendritic cell      49.187117
6  natural killer cell      47.527685

Bottom 5 cell types in Tongue:
                    Level Two Cell Type  mean_distance
2  endothelial cell of lymphatic vessel      44.692616
7                            neutrophil      34.321570
5                            macrophage      34.170755
9                          unknown cell      33.290598
0                                b cell      23.064363

# Get top and bottom cell types for each unique region in the dataset.
unique_regions = df_all_edges_with_cell_type_level['Unique Region'].unique()
for region in unique_regions:
    top_bottom = get_top_bottom_cell_types_by_median(df_all_edges_with_cell_type_level, cell_type_level, region)
    print(f"\nTop 5 cell types in {region}:")
    print(top_bottom[0])
    print(f"\nBottom 5 cell types in {region}:")
    print(top_bottom[1])

Top 5 cell types in Buccal Mucosa And Minor Salivary Glands:
   Level Two Cell Type  median_distance
3      epithelial cell        63.242607
6  natural killer cell        37.025667
9         unknown cell        28.831450
8               t cell        28.289618
4           fibroblast        25.141972

Bottom 5 cell types in Buccal Mucosa And Minor Salivary Glands:
                    Level Two Cell Type  median_distance
2  endothelial cell of lymphatic vessel        23.013311
0                                b cell        22.353669
7                            neutrophil        21.579446
5                            macrophage        20.110103
1                        dendritic cell        19.333714

Top 5 cell types in Gingiva:
                     Level Two Cell Type  median_distance
14                                t cell        49.247660
3   endothelial cell of lymphatic vessel        47.521442
7                           keratinocyte        42.797847
2                         dendritic cell        40.886464
4                        epithelial cell        37.195439

Bottom 5 cell types in Gingiva:
    Level Two Cell Type  median_distance
0             adipocyte        31.576881
12  natural killer cell        29.657322
1                b cell        29.370338
8       langerhans cell        26.245455
11   myoepithelial cell        25.924170

Top 5 cell types in Parotid:
    Level Two Cell Type  median_distance
12  natural killer cell        54.364997
9             mast cell        42.159777
14           neutrophil        34.148935
16               t cell        31.956611
3       epithelial cell        29.149386

Bottom 5 cell types in Parotid:
                     Level Two Cell Type  median_distance
2   endothelial cell of lymphatic vessel        19.534840
1                         dendritic cell        19.328978
0                                 b cell        19.314031
15                     perivascular cell        18.707282
7                        langerhans cell        18.679420

Top 5 cell types in Submandibular:
  Level Two Cell Type  median_distance
0              b cell        45.465234
2          fibroblast        33.036249
8              t cell        27.361812
9        unknown cell        25.710163
4          macrophage        23.417995

Bottom 5 cell types in Submandibular:
                    Level Two Cell Type  median_distance
3                 gland epithelium cell        21.156766
6                   natural killer cell        20.705518
5                    myoepithelial cell        20.641130
7                            neutrophil        19.521302
1  endothelial cell of lymphatic vessel        17.112130

Top 5 cell types in Tongue:
                    Level Two Cell Type  median_distance
3                       epithelial cell       126.256991
8                                t cell        45.091157
6                   natural killer cell        42.494294
4                            fibroblast        37.926541
2  endothelial cell of lymphatic vessel        34.740934

Bottom 5 cell types in Tongue:
  Level Two Cell Type  median_distance
1      dendritic cell        32.105607
9        unknown cell        23.558730
7          neutrophil        22.459853
5          macrophage        22.358667
0              b cell        15.471976

calculate_regional_variability(df_all_edges_with_cell_type_level, cell_type_level)

Regional Variability Analysis:
Mean: Average distance in each region
Std: Standard deviation of distances
CV: Coefficient of Variation (std/mean * 100%)
                                          mean    std  CV (%)
Unique Region                                                
Buccal Mucosa And Minor Salivary Glands  41.16  37.14    90.2
Gingiva                                  46.82  36.63    78.2
Parotid                                  28.23  21.27    75.3
Submandibular                            30.94  26.10    84.4
Tongue                                   48.57  44.56    91.7

Cell Type Variability Analysis (sorted by CV):
                                       mean    std  CV (%)
Level Two Cell Type                                       
dendritic cell                        37.36  37.48   100.3
macrophage                            32.67  30.91    94.6
fibroblast                            44.68  39.07    87.4
unknown cell                          36.94  31.98    86.6
mast cell                             46.79  40.10    85.7
keratinocyte                          40.74  34.66    85.1
neutrophil                            25.91  21.90    84.5
t cell                                44.40  37.45    84.3
langerhans cell                       23.62  19.63    83.1
endothelial cell of lymphatic vessel  40.28  33.01    82.0
muscle cell                           26.37  21.33    80.9
b cell                                32.21  25.19    78.2
neuroglial cell/neuron                24.59  19.04    77.4
natural killer cell                   47.10  35.86    76.1
adipocyte                             38.92  28.83    74.1
perivascular cell                     21.88  16.15    73.8
myoepithelial cell                    27.09  19.05    70.3
gland epithelium cell                 26.30  17.81    67.7
epithelial cell                       94.96  57.84    60.9

plot_violin_cells_per_celltype(df_all_edges_with_cell_type_level, cell_type_level, os.path.join(basepath, figures_output_dir), density_norm='area')

plot_distance_distribution_boxplots_by_region(df_all_edges_with_cell_type_level, cell_type_level, os.path.join(basepath, figures_output_dir))

plot_distance_distribution_heatmap(df_all_edges_with_cell_type_level, cell_type_level, os.path.join(basepath, figures_output_dir))

plot_violin_plots_all_regions(df_all_edges_with_cell_type_level, cell_type_level, os.path.join(basepath, figures_output_dir), density_norm="count") # Or, density_norm="count" or "area" based on preference.

Level Three Cell Type Analysis

# Get mean, median, minimum, maximum distance per cell type in all unique regions.
cell_type_level = 'Level Three Cell Type'
df_all_edges_with_cell_type_level = df_all_edges_with_cell_types[(df_all_edges_with_cell_types['Anchor Cell Type Level'] == cell_type_level) & (df_all_edges_with_cell_types['Anchor Cell Type'] == anchor_cell_type_dict[cell_type_level])]

df_distance_stats_cell_type_level = df_all_edges_with_cell_type_level.groupby([cell_type_level, 'Unique Region']).agg(
    mean_distance=('Distance', 'mean'),
    median_distance=('Distance', 'median'),
    min_distance=('Distance', 'min'),
    max_distance=('Distance', 'max')
).reset_index()
df_distance_stats_cell_type_level

	Level Three Cell Type	Unique Region	mean_distance	median_distance	min_distance	max_distance
0	acinar cell of salivary gland	Gingiva	36.739726	30.877582	0.029614	198.327899
1	acinar cell of salivary gland	Parotid	26.180957	22.977472	0.017954	199.271997
2	acinar cell of salivary gland	Submandibular	25.430226	22.379846	3.106445	198.710172
3	adipocyte	Gingiva	38.921484	31.576881	1.472765	198.358252
4	b cell	Buccal Mucosa And Minor Salivary Glands	23.196496	22.353669	2.804015	98.284549
5	b cell	Gingiva	38.913598	29.370338	1.271453	198.729570
6	b cell	Parotid	24.002071	20.842505	0.004528	194.426806
7	b cell	Submandibular	49.468888	45.465234	3.304921	184.012207
8	b cell	Tongue	23.064363	15.471976	3.696282	119.542137
9	dendritic cell	Buccal Mucosa And Minor Salivary Glands	23.820644	19.333714	2.753634	176.453535
10	dendritic cell	Gingiva	53.327090	40.886464	0.225297	199.932087
11	dendritic cell	Parotid	21.289953	19.328978	0.050563	154.155788
12	dendritic cell	Tongue	49.187117	32.105607	3.162278	199.890432
13	endothelial cell of lymphatic vessel	Buccal Mucosa And Minor Salivary Glands	31.266348	23.013311	3.036445	188.564690
14	endothelial cell of lymphatic vessel	Gingiva	51.530766	47.521442	8.561243	143.755112
15	endothelial cell of lymphatic vessel	Parotid	27.392054	19.534840	0.253150	181.675452
16	endothelial cell of lymphatic vessel	Submandibular	23.177030	17.112130	3.412477	121.602436
17	endothelial cell of lymphatic vessel	Tongue	44.692616	34.740934	4.208622	199.544105
18	epithelial cell	Buccal Mucosa And Minor Salivary Glands	69.005527	63.242607	3.006659	199.616482
19	epithelial cell	Gingiva	49.671535	37.195439	1.752795	198.377561
20	epithelial cell	Parotid	39.288368	29.149386	0.468023	199.229374
21	epithelial cell	Tongue	116.777509	126.256991	3.323402	199.987850
22	epithelial cell:ductal	Gingiva	42.560340	33.553722	0.238018	199.675802
23	epithelial cell:ductal	Parotid	26.200793	21.407199	0.037888	199.567157
24	epithelial cell:ductal	Submandibular	24.609401	20.199691	2.773536	199.833787
25	fibroblast	Buccal Mucosa And Minor Salivary Glands	35.795373	25.141972	2.510976	199.696019
26	fibroblast	Gingiva	42.361864	34.901074	0.127369	198.612028
27	fibroblast	Parotid	25.985495	20.910105	0.009735	199.030058
28	fibroblast	Submandibular	45.426189	35.755874	2.540177	199.943992
29	fibroblast	Tongue	52.144835	37.926541	2.530316	199.999050
30	glial cell/neuron	Parotid	24.591434	20.583996	0.124848	191.137047
31	keratinocyte	Gingiva	53.520737	42.797847	0.010330	199.836547
32	keratinocyte	Parotid	29.006264	23.250918	0.042787	199.947587
33	langerhans cell	Gingiva	35.348131	26.245455	0.353423	166.645525
34	langerhans cell	Parotid	21.515277	18.679420	0.118568	151.121503
35	macrophage	Buccal Mucosa And Minor Salivary Glands	25.882874	20.110103	2.995413	175.639439
36	macrophage	Gingiva	43.967814	35.395149	0.362944	196.376846
37	macrophage	Parotid	33.360746	28.845995	3.541892	123.118246
38	macrophage	Submandibular	30.757152	23.417995	3.103627	190.454122
39	macrophage	Tongue	34.170755	22.358667	2.800446	199.962253
40	mast cell	Gingiva	45.118725	32.004082	0.283616	198.023632
41	mast cell	Parotid	55.949381	42.159777	0.497809	164.482230
42	monocyte	Parotid	26.785779	22.590948	0.100367	196.269036
43	mural cell	Parotid	21.881249	18.707282	0.068190	166.700706
44	muscle cell:skeletal	Parotid	26.374298	22.262380	0.246935	196.110438
45	myoepithelial cell	Gingiva	33.597314	25.924170	0.499390	195.159907
46	myoepithelial cell	Parotid	31.860121	26.601561	0.027328	199.863760
47	myoepithelial cell	Submandibular	23.571161	20.641130	2.755449	136.143821
48	myofibroblast	Parotid	32.692357	25.995240	2.804015	184.105520
49	myofibroblast	Submandibular	37.834254	27.126555	2.890069	199.916633
50	natural killer cell	Buccal Mucosa And Minor Salivary Glands	46.769140	37.025667	3.381198	195.615196
51	natural killer cell	Gingiva	41.823514	29.657322	2.560580	189.288209
52	natural killer cell	Parotid	63.315905	54.364997	0.139128	198.587437
53	natural killer cell	Submandibular	25.280999	20.705518	3.148412	158.294820
54	natural killer cell	Tongue	47.527685	42.494294	3.064719	197.964770
55	neutrophil	Buccal Mucosa And Minor Salivary Glands	30.028958	21.579446	2.554408	194.583330
56	neutrophil	Gingiva	46.213377	32.192202	0.029228	199.542077
57	neutrophil	Parotid	44.659693	34.148935	18.784102	87.823018
58	neutrophil	Submandibular	21.857157	19.521302	2.968164	164.314371
59	neutrophil	Tongue	34.321570	22.459853	3.085855	199.231084
60	plasma cell	Parotid	21.626882	17.710427	0.038203	195.393503
61	t cell	Parotid	25.638718	21.043274	0.043842	184.558144
62	t cell:cd4+	Buccal Mucosa And Minor Salivary Glands	38.785521	29.526032	3.226841	189.937523
63	t cell:cd4+	Gingiva	43.033934	32.807020	0.094913	196.281616
64	t cell:cd4+	Parotid	48.489673	36.242813	1.426787	198.546619
65	t cell:cd4+	Submandibular	40.572532	37.560784	3.342529	187.306834
66	t cell:cd4+	Tongue	56.815946	42.660169	2.951694	199.931813
67	t cell:cd8+	Buccal Mucosa And Minor Salivary Glands	33.406082	24.008696	3.083018	196.505044
68	t cell:cd8+	Gingiva	64.303943	52.379107	0.150396	199.784633
69	t cell:cd8+	Parotid	44.306240	33.225023	0.106672	199.996540
70	t cell:cd8+	Submandibular	23.920693	21.025193	2.549510	136.845259
71	t cell:cd8+	Tongue	78.353407	70.255983	4.000000	199.529259
72	t cell:regulatory	Buccal Mucosa And Minor Salivary Glands	40.665551	30.033398	3.026549	198.825866
73	t cell:regulatory	Parotid	38.177654	32.661321	3.010399	198.026242
74	t cell:regulatory	Submandibular	33.265373	30.085399	2.798660	186.292868
75	t cell:regulatory	Tongue	54.034886	38.850515	3.112073	199.994831
76	thymocyte:cd4-intermediate cd8+ double-positive	Parotid	44.202736	37.229592	2.683282	199.631642
77	thymocyte:cd4-intermediate cd8+ double-positive	Submandibular	26.009947	22.290413	3.658210	117.123919
78	thymocyte:cd4-intermediate cd8+ double-positive	Tongue	62.981557	43.618355	5.628943	194.287583
79	unknown cell	Buccal Mucosa And Minor Salivary Glands	42.374333	28.831450	2.350532	199.920297
80	unknown cell	Gingiva	47.466793	36.770012	0.006517	199.814472
81	unknown cell	Parotid	32.154130	25.431573	0.015375	199.882778
82	unknown cell	Submandibular	32.777522	25.710163	2.720294	199.821445
83	unknown cell	Tongue	33.290598	23.558730	2.438237	199.998125

# Get top and bottom cell types for each unique region in the dataset.
unique_regions = df_all_edges_with_cell_type_level['Unique Region'].unique()
for region in unique_regions:
    top_bottom = get_top_bottom_cell_types_by_mean(df_all_edges_with_cell_type_level, cell_type_level, region)
    print(f"\nTop 5 cell types in {region}:")
    print(top_bottom[0])
    print(f"\nBottom 5 cell types in {region}:")
    print(top_bottom[1])

Top 5 cell types in Buccal Mucosa And Minor Salivary Glands:
   Level Three Cell Type  mean_distance
3        epithelial cell      69.005527
6    natural killer cell      46.769140
11          unknown cell      42.374333
10     t cell:regulatory      40.665551
8            t cell:cd4+      38.785521

Bottom 5 cell types in Buccal Mucosa And Minor Salivary Glands:
                  Level Three Cell Type  mean_distance
2  endothelial cell of lymphatic vessel      31.266348
7                            neutrophil      30.028958
5                            macrophage      25.882874
1                        dendritic cell      23.820644
0                                b cell      23.196496

Top 5 cell types in Gingiva:
                   Level Three Cell Type  mean_distance
16                           t cell:cd8+      64.303943
8                           keratinocyte      53.520737
3                         dendritic cell      53.327090
4   endothelial cell of lymphatic vessel      51.530766
5                        epithelial cell      49.671535

Bottom 5 cell types in Gingiva:
            Level Three Cell Type  mean_distance
1                       adipocyte      38.921484
2                          b cell      38.913598
0   acinar cell of salivary gland      36.739726
9                 langerhans cell      35.348131
12             myoepithelial cell      33.597314

Top 5 cell types in Parotid:
   Level Three Cell Type  mean_distance
17   natural killer cell      63.315905
11             mast cell      55.949381
21           t cell:cd4+      48.489673
18            neutrophil      44.659693
22           t cell:cd8+      44.306240

Bottom 5 cell types in Parotid:
   Level Three Cell Type  mean_distance
1                 b cell      24.002071
13            mural cell      21.881249
19           plasma cell      21.626882
9        langerhans cell      21.515277
2         dendritic cell      21.289953

Top 5 cell types in Submandibular:
   Level Three Cell Type  mean_distance
1                 b cell      49.468888
4             fibroblast      45.426189
10           t cell:cd4+      40.572532
7          myofibroblast      37.834254
12     t cell:regulatory      33.265373

Bottom 5 cell types in Submandibular:
                   Level Three Cell Type  mean_distance
3                epithelial cell:ductal       24.609401
11                           t cell:cd8+      23.920693
6                     myoepithelial cell      23.571161
2   endothelial cell of lymphatic vessel      23.177030
9                             neutrophil      21.857157

Top 5 cell types in Tongue:
                              Level Three Cell Type  mean_distance
3                                   epithelial cell     116.777509
9                                       t cell:cd8+      78.353407
11  thymocyte:cd4-intermediate cd8+ double-positive      62.981557
8                                       t cell:cd4+      56.815946
10                                t cell:regulatory      54.034886

Bottom 5 cell types in Tongue:
                   Level Three Cell Type  mean_distance
2   endothelial cell of lymphatic vessel      44.692616
7                             neutrophil      34.321570
5                             macrophage      34.170755
12                          unknown cell      33.290598
0                                 b cell      23.064363

# Get top and bottom cell types for each unique region in the dataset.
unique_regions = df_all_edges_with_cell_type_level['Unique Region'].unique()
for region in unique_regions:
    top_bottom = get_top_bottom_cell_types_by_median(df_all_edges_with_cell_type_level, cell_type_level, region)
    print(f"\nTop 5 cell types in {region}:")
    print(top_bottom[0])
    print(f"\nBottom 5 cell types in {region}:")
    print(top_bottom[1])

Top 5 cell types in Buccal Mucosa And Minor Salivary Glands:
   Level Three Cell Type  median_distance
3        epithelial cell        63.242607
6    natural killer cell        37.025667
10     t cell:regulatory        30.033398
8            t cell:cd4+        29.526032
11          unknown cell        28.831450

Bottom 5 cell types in Buccal Mucosa And Minor Salivary Glands:
                  Level Three Cell Type  median_distance
2  endothelial cell of lymphatic vessel        23.013311
0                                b cell        22.353669
7                            neutrophil        21.579446
5                            macrophage        20.110103
1                        dendritic cell        19.333714

Top 5 cell types in Gingiva:
                   Level Three Cell Type  median_distance
16                           t cell:cd8+        52.379107
4   endothelial cell of lymphatic vessel        47.521442
8                           keratinocyte        42.797847
3                         dendritic cell        40.886464
5                        epithelial cell        37.195439

Bottom 5 cell types in Gingiva:
            Level Three Cell Type  median_distance
0   acinar cell of salivary gland        30.877582
13            natural killer cell        29.657322
2                          b cell        29.370338
9                 langerhans cell        26.245455
12             myoepithelial cell        25.924170

Top 5 cell types in Parotid:
                              Level Three Cell Type  median_distance
17                              natural killer cell        54.364997
11                                        mast cell        42.159777
24  thymocyte:cd4-intermediate cd8+ double-positive        37.229592
21                                      t cell:cd4+        36.242813
18                                       neutrophil        34.148935

Bottom 5 cell types in Parotid:
                   Level Three Cell Type  median_distance
3   endothelial cell of lymphatic vessel        19.534840
2                         dendritic cell        19.328978
13                            mural cell        18.707282
9                        langerhans cell        18.679420
19                           plasma cell        17.710427

Top 5 cell types in Submandibular:
   Level Three Cell Type  median_distance
1                 b cell        45.465234
10           t cell:cd4+        37.560784
4             fibroblast        35.755874
12     t cell:regulatory        30.085399
7          myofibroblast        27.126555

Bottom 5 cell types in Submandibular:
                  Level Three Cell Type  median_distance
8                   natural killer cell        20.705518
6                    myoepithelial cell        20.641130
3               epithelial cell:ductal         20.199691
9                            neutrophil        19.521302
2  endothelial cell of lymphatic vessel        17.112130

Top 5 cell types in Tongue:
                              Level Three Cell Type  median_distance
3                                   epithelial cell       126.256991
9                                       t cell:cd8+        70.255983
11  thymocyte:cd4-intermediate cd8+ double-positive        43.618355
8                                       t cell:cd4+        42.660169
6                               natural killer cell        42.494294

Bottom 5 cell types in Tongue:
   Level Three Cell Type  median_distance
1         dendritic cell        32.105607
12          unknown cell        23.558730
7             neutrophil        22.459853
5             macrophage        22.358667
0                 b cell        15.471976

calculate_regional_variability(df_all_edges_with_cell_type_level, cell_type_level)

Regional Variability Analysis:
Mean: Average distance in each region
Std: Standard deviation of distances
CV: Coefficient of Variation (std/mean * 100%)
                                          mean    std  CV (%)
Unique Region                                                
Buccal Mucosa And Minor Salivary Glands  41.16  37.14    90.2
Gingiva                                  46.82  36.63    78.2
Parotid                                  28.23  21.27    75.3
Submandibular                            30.94  26.10    84.4
Tongue                                   48.57  44.56    91.7

Cell Type Variability Analysis (sorted by CV):
                                                  mean    std  CV (%)
Level Three Cell Type                                                
dendritic cell                                   37.36  37.48   100.3
macrophage                                       32.99  31.34    95.0
myofibroblast                                    36.72  33.92    92.4
t cell:cd8+                                      46.75  41.83    89.5
fibroblast                                       45.47  39.46    86.8
unknown cell                                     36.94  31.98    86.6
mast cell                                        46.79  40.10    85.7
keratinocyte                                     40.74  34.66    85.1
neutrophil                                       25.91  21.90    84.5
langerhans cell                                  23.62  19.63    83.1
t cell:regulatory                                43.48  36.08    83.0
t cell                                           25.64  21.19    82.6
endothelial cell of lymphatic vessel             40.28  33.01    82.0
muscle cell:skeletal                             26.37  21.33    80.9
monocyte                                         26.79  20.86    77.9
t cell:cd4+                                      47.81  37.01    77.4
glial cell/neuron                                24.59  19.04    77.4
plasma cell                                      21.63  16.71    77.3
natural killer cell                              47.10  35.86    76.1
thymocyte:cd4-intermediate cd8+ double-positive  42.88  32.57    76.0
epithelial cell:ductal                           26.18  19.56    74.7
adipocyte                                        38.92  28.83    74.1
mural cell                                       21.88  16.15    73.8
b cell                                           37.32  26.94    72.2
myoepithelial cell                               27.09  19.05    70.3
acinar cell of salivary gland                    26.36  16.89    64.1
epithelial cell                                  94.96  57.84    60.9

plot_violin_cells_per_celltype(df_all_edges_with_cell_type_level, cell_type_level, os.path.join(basepath, figures_output_dir), density_norm='area')

plot_distance_distribution_boxplots_by_region(df_all_edges_with_cell_type_level, cell_type_level, os.path.join(basepath, figures_output_dir))

plot_distance_distribution_heatmap(df_all_edges_with_cell_type_level, cell_type_level, os.path.join(basepath, figures_output_dir))

plot_violin_plots_all_regions(df_all_edges_with_cell_type_level, cell_type_level, os.path.join(basepath, figures_output_dir), density_norm="count") # Or, density_norm="count" or "area" based on preference.