Running ONTraC on a simulated dataset

This dataset was generated using the R package dyngen and consists 2 lineages, 10 cell types, and 1,000 cells. Lineage 1 cells exhibit a strong correlation between the spatial distribution (radius) and simulated time, while lineage 2 cells have a random spatial distribution.

Simulated dataset introduction

Generate the simulated data using Dyngen

library(tidyverse)
library(dyngen)

set.seed(1)

backbone <- backbone_bifurcating()
config <- initialise_model(
   backbone = backbone,
   num_tfs = nrow(backbone$module_info),
   num_targets = 500,
   num_hks = 500,
   verbose = FALSE
)

config <- initialise_model(
   backbone = backbone,
   num_cells = 1000,
   num_tfs = nrow(backbone$module_info),
   num_targets = 50,
   num_hks = 50,
   verbose = FALSE,
   download_cache_dir = tools::R_user_dir("dyngen", "data"),
   simulation_params = simulation_default(
      total_time = 1000,
      census_interval = 2,
      ssa_algorithm = ssa_etl(tau = 300/3600),
      experiment_params = simulation_type_wild_type(num_simulations = 10)
   )
)

model <- generate_tf_network(config)
model <- generate_feature_network(model)
model <- generate_kinetics(model)
model <- generate_gold_standard(model)
model <- generate_cells(model)
model <- generate_experiment(model)

  • Add noise. Due to the random nature of the noise, the simulated data will be slightly different each time.

library(SingleCellExperiment)

sce <- as_sce(model)

colData(sce)$sim_time <- colData(sce)$sim_time + rnorm(nrow(colData(sce)), mean = 0, sd = 0.01 * colData(sce)$sim_time)
colData(sce)$sim_time <- ifelse(colData(sce)$sim_time == 0, colData(sce)$sim_time + runif(sum(colData(sce)$sim_time == 0), 0.01, 1), colData(sce)$sim_time)
colData(sce)

write.csv(sce@assays@data@listData$counts, "gene_expression_matrix_of_simulated_data.csv")

library(scater)

sce <- readRDS("simulation_data_with_noise.rds")

set.seed(8)
sce <- logNormCounts(sce)
sce <- runPCA(sce)
sce <- runUMAP(sce)
reducedDims(sce)

  • Create umap_df

umap_coords <- reducedDims(sce)$UMAP
umap_df <- as.data.frame(umap_coords)
umap_df$time <- colData(sce)$sim_time

  • Assign the lineage

umap_df$trajectory_1 <- ifelse((umap_df$UMAP2 > 0) | (umap_df$UMAP2 < 0 & umap_df$UMAP1 < 0),
'true', 'false')

  • Create the ‘trajectory_2’ column

umap_df$trajectory_2 <- ifelse(umap_df$UMAP2 <= 0 & umap_df$UMAP1 > 0,
'true', 'false')

umap_df$lineage <- ifelse(umap_df$trajectory_1 == 'true', 1,
ifelse(umap_df$trajectory_2 == 'true', 2, NA))

umap_df$trajectory_1 <- NULL
umap_df$trajectory_2 <- NULL

  • Assign the cell type

umap_df <- umap_df[order(umap_df$lineage, umap_df$time), ]
cell_types <- c(rep(LETTERS[1:7], each = 100), rep(LETTERS[8:10], each = c(100, 100, 99)))
umap_df$Cell_Type <- cell_types

umap_df$Cell_Type <- NA

time_list <- as.list(umap_df$time)

num_cell_types_lineage_1 <- 7  # A-G
num_cell_types_lineage_2 <- 3  # H-J

cell_types_lineage_1 <- LETTERS[1:num_cell_types_lineage_1]
cell_types_lineage_2 <- LETTERS[(num_cell_types_lineage_1 + 1):(num_cell_types_lineage_1 + num_cell_types_lineage_2)]

cells_per_cell_type_lineage_1 <- ceiling(nrow(umap_df[umap_df$lineage == 1, ]) / num_cell_types_lineage_1)
cells_per_cell_type_lineage_2 <- ceiling(nrow(umap_df[umap_df$lineage == 2, ]) / num_cell_types_lineage_2)

# Assign cell types
for (lineage in unique(umap_df$lineage)) {
if (lineage == 1) {
   # Sort the data frame by time within lineage 1
   umap_df_lineage_1 <- umap_df[umap_df$lineage == 1, ]
   umap_df_lineage_1 <- umap_df_lineage_1[order(umap_df_lineage_1$time), ]

   # A-G
   umap_df_lineage_1$Cell_Type <- rep(cell_types_lineage_1, each = cells_per_cell_type_lineage_1)[1:nrow(umap_df_lineage_1)]

   umap_df[umap_df$lineage == 1, ] <- umap_df_lineage_1
} else if (lineage == 2) {
   umap_df_lineage_2 <- umap_df[umap_df$lineage == 2, ]
   umap_df_lineage_2 <- umap_df_lineage_2[order(umap_df_lineage_2$time), ]

   # H-J
   umap_df_lineage_2$Cell_Type <- rep(cell_types_lineage_2, each = cells_per_cell_type_lineage_2)[1:nrow(umap_df_lineage_2)]

   umap_df[umap_df$lineage == 2, ] <- umap_df_lineage_2
}
}

umap_df$time <- sapply(time_list, function(x) unlist(x))

  • Assign the spatial coordinates

umap_df$r <- NA
umap_df$theta <- NA
umap_df$x <- NA
umap_df$y <- NA

# Define the number of cells for each lineage
num_cells_lineage_1 <- sum(umap_df$lineage == 1)
num_cells_lineage_2 <- sum(umap_df$lineage == 2)

# r for lineage 1 is directly inherited from the time point
# r for lineage 2 is randomized
umap_df$r[umap_df$lineage == 1] <- sqrt(umap_df$time[umap_df$lineage == 1]) * 30
umap_df$r[umap_df$lineage == 2] <- runif(num_cells_lineage_2, min = 0, max = 1000)

# theta for lineage 1 is between 40-50
# theta for lineage 2 is between 0-90
umap_df$theta[umap_df$lineage == 1] <- runif(num_cells_lineage_1, 0, 360)
umap_df$theta[umap_df$lineage == 2] <- runif(num_cells_lineage_2, 0, 360)
umap_df$x <- umap_df$r * cos(umap_df$theta * pi / 180)
umap_df$y <- umap_df$r * sin(umap_df$theta * pi / 180)

  • Prepare the input file for ONTraC

export_df <- umap_df[c("Cell_Type","x","y","time","r","lineage")]
export_df <- data.frame(Cell_ID = rownames(export_df), export_df, row.names = NULL)
export_df <- data.frame(export_df[,1], Sample = "Simulation", export_df[, -1])
names(export_df)[1] <- "Cell_ID"
colnames(export_df)[6] <- "sim_time"

  • Save the input file for ONTraC

write.csv(export_df, "simulated_dataset.csv", row.names = FALSE)

Running ONTraC

If your default shell is not Bash, please adjust this code.

ONTraC will run on CPU if CUDA is not available.

conda activate ONTraC
ONTraC --meta-input full_simulation_data_with_noise.csv \
--NN-dir simulation_NN \
--GNN-dir simulation_GNN \
--NT-dir simulation_NT \
--device cuda --epochs 1000 -s 42 --lr 0.03 --hidden-feats 4 -k 6 \
--modularity-loss-weight 0.3 --regularization-loss-weight 0.1 \
--purity-loss-weight 300 --beta 0.03 2>&1 | tee simulation.log

Results visualization

Please see the Visualizations tutorials for details.

  • Loading results

from ONTraC.analysis.data import AnaData
from optparse import Values

options = Values()
options.NN_dir = 'simulation_NN'
options.GNN_dir = 'simulation_GNN'
options.NT_dir = 'simulation_NT'
options.log = 'simulation.log'
options.reverse = True  # Set it to False if you don't want reverse NT score
options.output = None  # We save the output figure by our self here
ana_data = AnaData(options)

  • Spatial cell type distribution

from ONTraC.analysis.cell_type import plot_spatial_cell_type_distribution_dataset_from_anadata

cell_type_pal = {'A': '#7CAE00',
               'B': '#00BC5A',
               'C': '#00C0B3',
               'D': '#00B4F0',
               'E': '#8E92FF',
               'F': '#EA6AF1',
               'G': '#FF64B0',
               'H': '#C42F5D',
               'I': '#A45900',
               'J': '#6A7300'}



fig, axes = plot_spatial_cell_type_distribution_dataset_from_anadata(ana_data = ana_data,
               palette=cell_type_pal)
fig.savefig('figures/Spatial_cell_type.png', dpi=150)
Simulated spatial cell types

  • Cell-level NT score spatial distribution

from ONTraC.analysis.spatial import plot_cell_NT_score_dataset_from_anadata

fig, ax = plot_cell_NT_score_dataset_from_anadata(ana_data=ana_data)
fig.savefig('cell_level_NT_score.png', dpi=300)
Cell level NT score