Data from: Enhanced resistance to Listeria infection in mice surviving sepsis: The role of lipid metabolism and myeloid cell reprogramming
Data files
Nov 21, 2025 version files 53.53 KB
Abstract
This dataset contains raw and processed lipidomics data from an experiment investigating the lipid mechanisms of CD11b+ myeloid cells in mice surviving sepsis, through a murine cecal ligation and puncture (CLP) model. Single-cell RNA sequencing data of splenic myeloid cells are accessible in the GEO database (GSE249839). The data encompass untargeted plasma lipidomics, confocal microscopy of lipid droplets, quantitative RT-PCR, and survival and bacterial burden outcomes following Listeria monocytogenes infection. Lipid species were quantified using LC-MS/MS, and lipid class composition was analyzed. This dataset includes normalized lipid abundance tables, instrument settings, and group metadata.
Dataset DOI: 10.5061/dryad.0p2ngf2cj
Description of the data and file structure
This dataset contains raw and processed data supporting the manuscript: "Enhanced resistance to Listeria infection in mice surviving sepsis: the role of lipid metabolism and myeloid cell reprogramming."
EXPERIMENTAL GROUPS
Group: Sham
- Surgical exposure without ligation or puncture.
Group: CLP (Sepsis Survivors)
- Cecal ligation and puncture performed; samples taken 4–12 weeks post-surgery.
Files and variables
FILE: RE_Figure_1_Suvial_and_bacterial_load.csv
Description: Survival (%) and CFU counts (log-transformed) in spleen and liver for sham and CLP mice 3 days post-infection.
Figure 1B: Monocyte Analysis
This section contains data on the percentage (%, left) and absolute counts (number of cells x 10^5) of monocytes. Each row represents a single mouse.
Variables
Sham: The monocyte percentage or absolute count for an individual mouse in the sham control group.
CLP: The monocyte percentage or absolute count for an individual mouse in the CLP sepsis model group.
Figure 1C: Survival Data
This section contains survival data for mice used to generate a Kaplan-Meier survival curve. Note the stacked layout: The data for the Sham group is listed first, followed by the data for the CLP group.
The time point at which survival status was assessed for an individual mouse (0-3 days).
Value 1: Indicates the animal was alive on the specified day.
Value 0: Indicates the animal was deceased or culled on the specified day.
Variables
Time (days): The day post-procedure (0, 1, 2, or 3) on which the survival status was recorded.
Sham: The survival status for a mouse in the sham group on the corresponding day (1 = alive; 0 = deceased/culled).
CLP: The survival status for a mouse in the CLP group on the corresponding day (1 = alive; 0 = deceased/culled).
Figure 1D: Bacterial Load
This section contains bacterial load data from the liver (left) and spleen (right) of mice following infection. Each row represents a single mouse. Unit: CFU (Colony-Forming Units), a measure used to quantify the number of viable bacteria per liver or spleen.
Variables
Sham: The bacterial load (CFU) for an individual mouse in the sham group.
CLP: The bacterial load (CFU) for an individual mouse in the CLP group.
FILE: RE_Figure_2_Imaging_and_Cytokine_Data.csv
Description: This file contains the raw data for generating panels 2B and 2D in Figure 2. The data for each panel is organized as a distinct table within the same CSV file, separated by empty rows.
Figure 2B: Colocalized Voxel Analysis
Description: This table contains quantitative data from imaging analysis. Each value represents the total count of colocalized voxels in each field, which is a measure of the spatial overlap between two different signals in microscopy images (Imaris). Each row represents a single mouse/sample.
Unit: Count of voxels (dimensionless number)
Variables
Sham: The total count of colocalized voxels for a sample from the sham group.
CLP: The total count of colocalized voxels for a sample from the CLP group.
Figure 2D: Interferon-gamma (IFNγ) Concentration
This table contains concentration measurements of the cytokine Interferon-gamma (IFNγ) from serum. Each row represents a single mouse.
Unit: pg/mL
Variables
Sham: The IFNγ concentration (pg/mL) for an individual mouse in the sham group.
CLP: The IFNγ concentration (pg/mL) for an individual mouse in the CLP group.
FILE: Figure_3_scRNAseq_DEGs.csv
- Differential gene expression results for CD11b+Ly6Chigh cells (CLP vs. sham), including log2 fold-change, p-values, and FDR-adjusted q-values.
Variables
gene_name: The official symbol for the differentially expressed gene.
log2FoldChange: The log2 of the fold change in expression (CLP relative to sham).
pvalue: The nominal p-value for the differential expression test.
padj: The p-value adjusted for multiple comparisons using FDR.
FILE: Figure_3_qPCR_validation.csv
- RT-qPCR expression values for genes LPL, ABCA1, and ABCA4. Normalized to GAPDH using the 2^−ΔΔCt method.
Variables
LPL Sham: Relative expression of the LPL gene in a mouse from the sham group.
LPL CLP: Relative expression of the LPL gene in a mouse from the CLP group.
ABCA1 Sham: Relative expression of the ABCA1 gene in a mouse from the sham group.
ABCA1 CLP: Relative expression of the ABCA1 gene in a mouse from the CLP group.
ABCA4 Sham: Relative expression of the ABCA4 gene in a mouse from the sham group.
ABCA4 CLP: Relative expression of the ABCA4 gene in a mouse from the CLP group.
FILE: Figure_4_lipidomics_data.csv
- Normalized plasma concentration for lipid species (PS, PI, PG, PC), with mouse IDs and group labels.
Variables
PS Sham: Normalized concentration of the PS lipid class in a mouse from the sham group (nmol/mL).
PS CLP: Normalized concentration of the PS lipid class in a mouse from the CLP group (nmol/mL).
PG Sham: Normalized concentration of the PG lipid class in a mouse from the sham group (nmol/mL).
PG CLP: Normalized concentration of the PG lipid class in a mouse from the CLP group (nmol/mL).
PC Sham: Normalized concentration of the PC lipid class in a mouse from the sham group (nmol/mL).
PC CLP: Normalized concentration of the PC lipid class in a mouse from the CLP group (nmol/mL).
PI Sham: Normalized concentration of the PI lipid class in a mouse from the sham group (nmol/mL).
PI CLP: Normalized concentration of the PI lipid class in a mouse from the CLP group (nmol/mL).
PI36:2 Sham: Normalized concentration of the PI(36:2) lipid species in a mouse from the sham group (nmol/mL).
PI36:2 CLP: Normalized concentration of the PI(36:2) lipid species in a mouse from the CLP group (nmol/mL).
PI36:1 Sham: Normalized concentration of the PI(36:1) lipid species in a mouse from the sham group (nmol/mL).
PI36:1 CLP: Normalized concentration of the PI(36:1) lipid species in a mouse from the CLP group (nmol/mL).
FILE: RE_Supplementary_Table_1._Lipid_species.csv
Description: pooled concentration of lipid species of mouse plasma (nmol/mL).
Variables
Samples: The name of the lipid species being measured in that row.
S1 to S6: The concentration for Sham group samples 1 through 6.
C1 to C6: The concentration for CLP group samples 1 through 6.
FILE: RE_Supplementary_Table_2._Lipid_class.csv
Description: Calculated concentrations from the lipidomics data (nmol/mL).
Variables
Samples: The name of the lipid class being measured in that row.
S1 to S6: The total concentration for the lipid class in Sham group samples 1 through 6.
C1 to C6: The total concentration for the lipid class in CLP group samples 1 through 6.
Code/software
The dataset can be viewed and analyzed using the following free or open-source software:
Data are provided in standard formats (CSV, XLSX, TXT). For re-analysis, tools such as GraphPad Prism, Seurat (R), MetaboAnalyst, and Microsoft Excel were used in the original analysis.
GraphPad Prism (v9.5.1) – for statistical tests, survival curves, bar plots, and scatter plots.
MetaboAnalyst (v6.0, web-based) – for lipidomics data analysis.
Used for normalization, Pareto scaling, volcano plots, PLS-DA, and heatmaps.
Input: Normalized lipid intensity values from LC-MS analysis (CSV format)
Microsoft Excel – Used to organize and summarize raw data from lipidomics, qPCR, CFU counts, and flow cytometry; processed data include embedded metadata and sample group identifiers for clarity.
Animal Model and Experimental Design
Male C57BL/6J mice (8 weeks old) were used to model sepsis via cecal ligation and puncture (CLP), conducted under isoflurane anesthesia. Postoperative care included buprenorphine analgesia, subcutaneous resuscitation fluids, and one dose of imipenem/cilastatin. Sham controls underwent laparotomy without cecal manipulation. Mice were monitored daily, and moderate sepsis severity resulted in 20–30% mortality within 3–5 days. CLP survivors were studied at 4 and 12 weeks post-surgery.
Listeria monocytogenes Infection and Bacterial Load Analysis
At 12 weeks post-CLP or sham surgery, mice were intravenously challenged with 1×10⁷ CFU of Listeria monocytogenes (Lm). Survival was monitored for 7 days. For bacterial load quantification, mice were injected with 2×10⁶ CFU and sacrificed at 72 hours post-infection. Liver and spleen tissues were harvested, homogenized, serially diluted, plated on Brain Heart Infusion (BHI) agar, and incubated at 37 °C overnight. Colony-forming units (CFU) were normalized per gram of tissue.
Flow Cytometry and Cell Sorting
Splenocytes were harvested 4 weeks post-CLP and stained for CD11b, Ly6C, and Ly6G markers. CD11b⁺Ly6ChighLy6G⁻ monocytes were sorted using BD FACSAria II. Downstream assays included qRT-PCR, scRNA-seq, and immunofluorescence.
Single-Cell RNA Sequencing (scRNA-seq)
Sorted CD11b⁺Ly6Chigh cells were barcoded with TotalSeq™ hashtags and processed with Chromium Single Cell 3′ GEM, Library & Gel Bead Kit v3.1 (10x Genomics). Sequencing was performed on an Illumina HiSeq platform. Data processing included Cell Ranger pipeline alignment to the GRCm38 genome, UMAP-based clustering, and differential gene expression (DEG) analysis via Seurat (v5.0.1). DEGs with log2 fold change > 2 and FDR < 0.05 were used for KEGG pathway enrichment.
Immunohistochemistry and Lipid Droplet Analysis
Spleens were cryosectioned (10 μm) and stained for CD11b (Alexa Fluor 488) and LipidTOX (Deep Red or Green). Confocal images were acquired on a Zeiss LSM900 with Airyscan. LD accumulation in CD11b⁺ myeloid cells was quantified using Imaris software’s 3D colocalization module.
Plasma Lipidomics
Plasma was collected 4 weeks post-CLP following 2-hour fasting. Lipids were extracted from 20 μL plasma using methylene chloride:methanol:isopropanol (25:10:65 v/v/v + 0.1% BHT) and spiked with SPLASH® Lipidomix®. UHPLC was performed using a Waters CSH C18 column with a TripleTOF 5600 MS (AB Sciex). Data were normalized to internal standards and analyzed using MetaboAnalyst 6.0. Significant lipid shifts were determined via PLS-DA and t-tests. Lipids were annotated via LIPID MAPS and NIST references.
Quantitative RT-PCR
RNA from sorted monocytes was extracted with Direct-zol (Zymo), reverse transcribed, and pre-amplified using TaqMan PreAmp Master Mix. qPCR was performed using TaqMan assays for Lpl, Abca1, and Abca4, normalized to Gapdh expression using the2 -ΔΔCt method.
Statistical Analysis
Data are expressed as mean ± SEM. Comparisons between two groups used two-tailed unpaired Student’s t-tests or Mann–Whitney U tests where appropriate. Log-rank tests were used for survival analysis. A p-value < 0.05 was considered statistically significant.