Journal: bioRxiv

Article Title: Postnatal Reprogramming Shapes Human Intestinal Epithelial Immune Competency

doi: 10.64898/2026.05.05.722861

Figure Lengend Snippet: A. Schematic of intestinal developmental timepoints analysed by single cell multiomics and in situ spatial transcriptomics (ST). Created with BioRender. B. Overview of single cell multiomics and ST cohorts; each point represents an individual donor. C. UMAP of integrated single cell multiomics data showing all cells, coloured by broad cell type and age group. Colour scheme consistent with D. D. Pseudobulk principal component analysis (PCA) of epithelial, stromal and immune compartments identified by single cell multiomics. Data were downsampled to equal cell numbers and coloured by age group. E. Heatmaps of promoter accessibility and gene expression for development-associated genes in ileal epithelium; values are mean-scaled per sample. Age group colour scheme consistent with D. F. Representative terminal ileum section (3 months) profiled by MERSCOPE ST, coloured by broad cell type. G. Scaled Wasserstein distance from a location-matched fetal anchor plotted against log2-transformed post-conceptual weeks (PCW). Each point represents an individual donor. LOESS fits are shown, highlighting the inflection and peak of the trajectories. H. Integrated transcriptomic atlas combining scRNA-seq data with datasets from Elmentaite et al. 2021 and Fawkner-Corbett et al. 2021; each point represents a donor. I. Performance of a multilayer perceptron (MLP) regressor on the validation dataset. Predicted age corresponds to the median across cells per donor. Points are coloured by location and sized by cell number. J. Volcano plots showing top genes (left) and regulons (right) from the colonic ElasticNet model. Axes indicate signed feature importance and change in mean absolute error (MAE) following in silico perturbation.

Article Snippet: The MERSCOPE Vizgen Platform was used for high-resolution subcellular spatial transcriptomics, following the manufacturer’s instructions for resistant tissue with modifications (Vizgen, MERSCOPE, 91600112, Rev B).

Techniques: Single Cell, In Situ, Spatial Transcriptomics, Gene Expression, Transformation Assay, Biomarker Discovery, In Silico

Journal: bioRxiv

Article Title: Postnatal Reprogramming Shapes Human Intestinal Epithelial Immune Competency

doi: 10.64898/2026.05.05.722861

Figure Lengend Snippet: A. Flow cytometry gating strategy for isolation of stromal (CD45⁻) and immune (CD45⁺) compartments. B. UMAP of integrated MERSCOPE ST data, coloured by broad cell type, age group and location. C. Pseudobulk principal component analysis (PCA) of RNA and peak assays across epithelial, stromal and immune compartments identified by single cell multiomics, coloured by age group. D. UMAP of integrated single cell multiomics data, coloured by location. Paired UMAP overlay with cell cluster and age is shown in . E. UMAP of metacells derived from integrated single cell multiomics data, coloured by cell type. F. UMAP overlays showing gene expression (top), promoter accessibility (middle) and enhancer accessibility (bottom) for marker genes (MKI67, ADAMDEC1, F3). G. UMAP overlay showing cell type annotations and regulon activity in scDOGMA-seq data. Transcription factor regulons mark distinct lineages, including PHOX2B and SOX10 (enteric nervous system), HAND1 (mesenchymal), IRF8 (immune), PAX4 (endocrine) and CDX2 (epithelial), whereas KLF6 and ETS2 show broad activity across cell types.

Article Snippet: The MERSCOPE Vizgen Platform was used for high-resolution subcellular spatial transcriptomics, following the manufacturer’s instructions for resistant tissue with modifications (Vizgen, MERSCOPE, 91600112, Rev B).

Techniques: Flow Cytometry, Isolation, Single Cell, Derivative Assay, Gene Expression, Marker, Activity Assay

Journal: bioRxiv

Article Title: Postnatal Reprogramming Shapes Human Intestinal Epithelial Immune Competency

doi: 10.64898/2026.05.05.722861

Figure Lengend Snippet: A. Circular heatmap of predicted cell-type proportions from single cell multiomics. Age effects were modelled using quasibinomial generalized linear models with a natural cubic spline for age; predicted proportions are z-score normalized across age groups and annotated by broad compartments. B. Circular heatmap of predicted cell-type proportions from MERSCOPE ST data, z-score normalized across age groups and annotated by broad compartments. C. Representative MERSCOPE ST sections from 9 PCW (top) and 3 months (bottom) terminal ileum, coloured by broad cell type (left) and epithelial and T cell clusters (right). D. Spatial expression of CTNNB1 in MERSCOPE ST sections at 9 PCW and 4 weeks.

Article Snippet: The MERSCOPE Vizgen Platform was used for high-resolution subcellular spatial transcriptomics, following the manufacturer’s instructions for resistant tissue with modifications (Vizgen, MERSCOPE, 91600112, Rev B).

Techniques: Single Cell, Expressing

Journal: bioRxiv

Article Title: Postnatal Reprogramming Shapes Human Intestinal Epithelial Immune Competency

doi: 10.64898/2026.05.05.722861

Figure Lengend Snippet: A. Top, overview of diet conditions in single cell multiomics cohorts; each point represents a donor, coloured by diet and shaped by location. Bottom, scatterplot with LOESS-fitted line showing metabolic pathway scores in ileal samples, coloured by pathway. B. UMAP of ileal (top) and colonic (bottom) epithelial cells in a joint RNA–ATAC embedding, coloured by cell type, age group (as in A) and crypt–villus differentiation pseudotime. C. Relationship between mean chronological age (by age group) and crypt–villus differentiation score across epithelial cell clusters, coloured by location. Distributions show stem, differentiated, fetal and adult cNMF module usage across chronology (age group) and differentiation. D. Predicted proportions of epithelial cell types in ileum (top) and colon (bottom), estimated using sccomp and visualized as ribbon plots; colours correspond to cell types in B. E. Generalized additive model (GAM)-fitted trends of promoter and enhancer accessibility per sample and gene expression for selected genes across age in ileal epithelium. Shaded areas indicate +/- 1 standard error of the GAM fitted mean. F. Bubble plot showing age-associated regulon activity across ileal epithelial cells. G. Violin plots showing fetal gene module scores (left) and chromatin accessibility module scores (right) in colonic epithelium. H. Spatial expression of the fetal marker TTR in MERSCOPE ST sections at 9 PCW (left), 5 days (middle) and 13 months (right). I. Volcano plot showing differential expression between fetal and paediatric colonic stem cells in single cell multiomics data. Differential expression was assessed using DESeq2 (Wald test with Benjamini–Hochberg correction). Points are coloured by significance and effect size (red, |log2FC| > 1 and −log10P > 1; blue, |log2FC| < 1 and −log10P > 1; yellow, |log2FC| > 1 and −log10P < 1; grey, not significant). J. Density plots of LGR5 gene expression (left) and promoter accessibility (right) in colonic epithelial cells. K. Coverage and violin plots showing promoter accessibility and gene expression of stem cell marker genes in ileal epithelium, coloured by mean crypt–villus differentiation score per cell type.

Article Snippet: The MERSCOPE Vizgen Platform was used for high-resolution subcellular spatial transcriptomics, following the manufacturer’s instructions for resistant tissue with modifications (Vizgen, MERSCOPE, 91600112, Rev B).

Techniques: Single Cell, Gene Expression, Activity Assay, Expressing, Marker, Quantitative Proteomics

Journal: bioRxiv

Article Title: Postnatal Reprogramming Shapes Human Intestinal Epithelial Immune Competency

doi: 10.64898/2026.05.05.722861

Figure Lengend Snippet: A. Predicted proportions of immune cell types across age groups in MERSCOPE ST data, estimated using sccomp and visualized as ribbon plots (myeloid, ILC, T cells, NK cells, B cells and plasma cells). Colours correspond to cell types in B. B. Representative MERSCOPE ST sections at 9 PCW, 2 weeks, 4 weeks and 2 months, coloured by cell type. C. Representative immunohistochemistry (IHC) images across developmental stages (fetal, newborn, 1 month, 5 months and 1 year) stained for HLA-DR. Scale bars, 100 μm. D. Heatmaps of epithelial–neighbourhood correlations across developmental stages. Pearson correlations between epithelial gene expression programmes and neighbouring cytokine-related signals were computed in terminal ileum samples, averaged within age groups (fetal, <1 month), and visualized by hierarchical clustering using a consistent colour scale. E. Proportion of S1/S2 CXCL8⁺ fibroblasts across age groups in single cell multiomics data. F. Bubble plot showing marker gene expression and regulon activity in S1/S2 CXCL8⁺ fibroblasts in single cell multiomics data. G. Cell–cell communication networks inferred using CellChat from single cell multiomics data. Left, circle plot showing interaction strength between epithelial, immune and fibroblast populations in fetal and neonatal (<2 months) samples. Right, chord diagram highlighting ligand–receptor interactions in CXCL, TGFβ and BMP signalling pathways. H. Cluster–neighbour co-localisation network from MERSCOPE ST data highlighting interactions among S1/S2 CXCL8⁺ fibroblasts, IL1B⁺CXCL8⁺ myeloid cells, REG1B/DUOX2⁺ enterocytes, S100A8/9⁺ myeloid cells and epithelial macrophages. Edges represent co-localisation strength (threshold > 0.02). Node size reflects cell abundance and colour denotes cell type. Data are from terminal ileum samples aged <1 month. I. Proportion of intraepithelial immune cells in chelated epithelial crypt single cell multiomics data across age groups. J. Bubble plot showing cytokine and immune state marker gene expression in intraepithelial immune cells; point size indicates the proportion of each cell type within age groups.

Article Snippet: The MERSCOPE Vizgen Platform was used for high-resolution subcellular spatial transcriptomics, following the manufacturer’s instructions for resistant tissue with modifications (Vizgen, MERSCOPE, 91600112, Rev B).

Techniques: Clinical Proteomics, Immunohistochemistry, Staining, Gene Expression, Single Cell, Marker, Activity Assay

Journal: bioRxiv

Article Title: Postnatal Reprogramming Shapes Human Intestinal Epithelial Immune Competency

doi: 10.64898/2026.05.05.722861

Figure Lengend Snippet: A. Representative MERSCOPE ST sections coloured by inferred spatial niches. TI-terminal ileum. B. Heatmap of cell type–niche enrichment for a representative terminal ileum sample (ABB, 2 weeks). Values are shown as log2-transformed estimates, with statistical significance indicated by asterisks (FDR < 0.05, ** < 0.01, *** < 0.001; n.s., not significant). C. Cluster co-localisation networks within the crypt niche across developmental stages. Networks include clusters with total interaction proportion ≥ 0.2 and edges with weight ≥ 0.1. Node size reflects cell abundance, edge thickness indicates co-localisation strength, and node colour denotes cell compartment.

Article Snippet: The MERSCOPE Vizgen Platform was used for high-resolution subcellular spatial transcriptomics, following the manufacturer’s instructions for resistant tissue with modifications (Vizgen, MERSCOPE, 91600112, Rev B).

Techniques: Transformation Assay

Journal: bioRxiv

Article Title: Postnatal Reprogramming Shapes Human Intestinal Epithelial Immune Competency

doi: 10.64898/2026.05.05.722861

Figure Lengend Snippet: A. Proportion of selected clusters in MERSCOPE ST data across age groups; points represent individual donors and bars indicate mean proportions. B. Heatmaps of epithelial–neighbourhood correlations across developmental stages. Pearson correlations between epithelial gene expression programmes and neighbouring cytokine-related signals were computed in terminal ileum samples, averaged within age groups (>1 month), and visualized by hierarchical clustering using the same colour scale as . C. Bubble plot showing expression of cytokines and receptors across broad cell types in each age group. D. Age-associated changes in the myeloid compartment. (i) UMAP of myeloid cells coloured by cluster identity. (ii) Proportion of IL1B⁺ M1 macrophages within the myeloid compartment. (iii) Differential cluster abundance comparing <1 month versus fetal (top) and >1 month versus <1 month (bottom), assessed using MiloR. E. Box plots showing the distribution of T cell states, inferred using StarCAT, in intraepithelial lymphocytes from crypt-chelated epithelium and in CD45⁺ T cells. CD8 Trm, CD8⁺ tissue-resident memory T cells; TEMRA, terminally differentiated effector memory T cells; gdT, γδ T cells.

Article Snippet: The MERSCOPE Vizgen Platform was used for high-resolution subcellular spatial transcriptomics, following the manufacturer’s instructions for resistant tissue with modifications (Vizgen, MERSCOPE, 91600112, Rev B).

Techniques: Gene Expression, Expressing