Spatial multi-omics of human skin reveals KRAS and inflammatory responses to spaceflight

Abstract

Spaceflight can change metabolic, immunological, and biological homeostasis and cause skin rashes and irritation, yet the molecular basis remains unclear. To investigate the impact of short-duration spaceflight on the skin, we conducted skin biopsies on the Inspiration4 crew members before (L-44) and after (R + 1) flight. Leveraging multi-omics assays including GeoMx™ Digital Spatial Profiler, single-cell RNA/ATAC-seq, and metagenomics/metatranscriptomics, we assessed spatial gene expressions and associated microbial and immune changes across 95 skin regions in four compartments: outer epidermis, inner epidermis, outer dermis, and vasculature. Post-flight samples showed significant up-regulation of genes related to inflammation and KRAS signaling across all skin regions. These spaceflight-associated changes mapped to specific cellular responses, including altered interferon responses, DNA damage, epithelial barrier disruptions, T-cell migration, and hindered regeneration were located primarily in outer tissue compartments. We also linked epithelial disruption to microbial shifts in skin swab and immune cell activity to PBMC single-cell data from the same crew and timepoints. Our findings present the inaugural collection and examination of astronaut skin, offering insights for future space missions and response countermeasures.

Fig. 1. Experimental design and overall pre- and post-timepoint comparisons.

a Experimental design and workflow with representative tissue staining images (created with BioRender.com), b Uniform Manifold Approximation and Projection (UMAP) of all ROIs collected, c Volcano plot of overall post- vs. pre-spaceflight DEGs (using DESeq2 method), d Pathway enrichment analysis comparing DEGs from pre- and post-spaceflight skin tissues, visualizing normalized enrichment scores of MSigDB Hallmark pathways, and e Cell proportion comparisons between pre- and post-spaceflight samples (ns non-significant, *p ≤ 0.05, **p ≤ 0.01, ***p ≤ 0.001, and ****p ≤ 0.0001 by Wilcoxon test, two-sided; boxplot shows median/horizontal line inside the box, the interquartile range/box boundaries, whiskers extending to 1.5 times the interquartile range, and outliers as individual points outside the whiskers; exact p values are included in the Source Data). Source data are provided as a Source Data file.

Fig. 2. Region-specific gene expression profile and cellular composition changes.

a Volcano plot showing DEGs by ROI types—OE, IE, OD, and VA respectively; the number of DEGs were determined by cutoffs of adjusted p value < 0.1 and |log2FC| > 0.5 (using DESeq2 method), b UpSet plots comparing the intersections of region-specific DEGs, c Hallmark, non-germline gene set enrichment analysis across four ROI types; NES Normalized Enrichment Scores; Arrow indicates tissue locations, where OE is the outermost layer and VA is the innermost layer. Source data are provided as a Source Data file.

Fig. 3. Cellular microenvironment changes from spaceflight by tissue region.

a Gene expression changes of interest, b fold change of proportions in post-flight samples relative to pre-flight samples, by compartments, c cell type correlation matrix changes. Black boxes represent undetermined spots (due to minimal cell counts); boxes with X marks represent correlations that did not pass statistical testing (p value < 0.05, Pearson correlation, two-sided). Source data are provided as a Source Data file.

Fig. 4. Skin metagenomics and transcriptomics analysis for skin-microbiota interaction map.

a PCA across all metagenomic and metatranscriptomic (bacterial and viral reads) relative abundance features and all crew members pre- and post-flight, b Total number of bacterial and viral species with nonzero counts, c Relative abundances by sample and timepoint, grouped by family, d Changes in relative abundance before and after spaceflight, grouped by genus; statistically significant or previously reported microbes are visualized (two-sided Wilcoxon test across four crew members was performed to compare means between pre- and post-flight samples and to obtain p values, and error bars represent the standard error of the mean), and e Correlation across relative abundance of bacterial phyla identified by metagenomics data and known barrier/immune genes associated with skin diseases and disruptions. Source data are provided as a Source Data file.

Fig. 5. Changes in the skin immune system relates to peripheral blood immune cell changes.

a Notable cytokine changes and locations from (a) skin transcriptomics data by region and, b cytokine assay from serum samples (sig. Indicates overall statistical significance of the cytokine levels in the postflight samples relative to the preflight samples, where red indicates significantly increased, and green means stable/no change; two-sided Wilcoxon test was done with the p value cutoff of 0.05), c Comparison of DEGs between PBMC multiome data and spatial transcriptomics data from VA ROIs, d Dot plots visualizing mRNA transcript expression levels (left) and gene activity score from ATAC signals (right), where preflight samples were collected 44 days before launch (L-44) and postflight samples were collected 1, 45, and 82 days post return (R + 1, R + 45, and R + 82, respectively), e Flight and cell type specific gene signature enrichment in spatial data by timepoint and ROI types, f gene signature enrichment analysis using gene signatures built from skin disease-related gene expression profiles; two-sided Wilcoxon test across four crew members and 95 ROIs was performed to obtain p value, where *p ≤ 0.05 and **p ≤ 0.01, and error bars represents standard deviation of the mean. Source data are provided as a Source Data file.