High resolution spatially resolved transcriptomic atlas of kidney injury and repair by RNA hybridization-based in situ sequencing
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ABSTRACT:
ORGANISM(S): Mus musculus synthetic construct
PROVIDER: GSE227046 | GEO | 2024/01/17
REPOSITORIES: GEO
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Project description:Spatially resolved transcriptomics technologies allow for the measurement of gene expression in situ. We applied direct RNA hybridization-based in situ sequencing (ISS, Cartana) to compare male and female healthy mouse kidneys and the male kidneys injury and repair timecourse of ischemic reperfusion injury (IRI). A pre-selected panel of 200 genes were used to identify the dynamics of cell states and their spatial distributions during injury and repair. We developed a new computational pipeline, CellScopes, for the rapid analysis, multi-omic integration and visualization of spatially resolved transcriptomic datasets. The resulting atlas allowed us to resolve distinct kidney niches, dynamic alterations in cell state over the course of injury and repair and cell-cell interactions between leukocytes and kidney parenchyma. Projection of snRNA-seq dataset from the same injury and repair samples allowed us to impute the spatial localization of genes not directly measured by Cartana.
2024-01-17 | GSE227045 | GEO
Project description:Spatially resolved transcriptomics technologies allow for the measurement of gene expression in situ. We applied direct RNA hybridization-based in situ sequencing (ISS, Cartana) to compare male and female healthy mouse kidneys and the male kidneys injury and repair timecourse of ischemic reperfusion injury (IRI). A pre-selected panel of 200 genes were used to identify the dynamics of cell states and their spatial distributions during injury and repair. We developed a new computational pipeline, CellScopes, for the rapid analysis, multi-omic integration and visualization of spatially resolved transcriptomic datasets. The resulting atlas allowed us to resolve distinct kidney niches, dynamic alterations in cell state over the course of injury and repair and cell-cell interactions between leukocytes and kidney parenchyma. Projection of snRNA-seq dataset from the same injury and repair samples allowed us to impute the spatial localization of genes not directly measured by Cartana.
2024-01-17 | GSE227044 | GEO
Project description:Visualization of the transcriptome in situ has proven to be a valuable tool in exploring single-cell RNA-sequencing data, providing an additional spatial dimension to investigate multiplexed gene expression, cell types, disease architecture or even data driven discoveries. In situ sequencing (ISS) method based on padlock probes and rolling circle amplification has been used to spatially resolve gene transcripts in tissue sections of various origins. Here, we describe the next iteration of ISS, HybISS, hybridization-based in situ sequencing. Modifications in probe design allows for a new barcoding system via sequence-by-hybridization chemistry for improved spatial detection of RNA transcripts. Due to the amplification of probes, amplicons can be visualized with standard epifluorescence microscopes for high-throughput efficiency and the new sequencing chemistry removes limitations bound by sequence-by-ligation chemistry of ISS. HybISS design allows for increased flexibility and multiplexing, increased signal-to-noise, all without compromising throughput efficiency of imaging large fields of view. Moreover, the current protocol is demonstrated to work on human brain tissue samples, a source that has proven to be difficult to work with image-based spatial analysis techniques. Overall, HybISS technology works as a targeted amplification detection method for improved spatial transcriptomic visualization, and importantly, with an ease of implementation.
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Project description:We optimized and validated a female bilateral ischemia reperfusion injury model. Using the 10X Genomics Visium Spatial Gene Expression solution, we generated spatial maps of gene expression across the injury and repair time course, and applied two open-source computational tools, Giotto and SPOTlight, to increase resolution and measure cell-cell interaction dynamics. An ischemia time of 34 minutes in a female murine model resulted in comparable injury to males across the time course of injury and repair. We report increased resolution of cell and gene expression with Giotto, a computational toolbox for spatial data analysis. Using a seeded non-negative matrix regression (SPOTlight) to deconvolute the dynamic landscape of cell-cell interactions, we find that injured proximal tubule cells are characterized by increasing macrophage and lymphocyte interactions even at 6 weeks after injury, consistent with a pro-inflammatory role for this cell state. In this transcriptomic atlas, we defined region-specific and injury-induced loss of differentiation markers and their re-expression during repair, as well as region-specific injury and repair transcriptional responses. Lastly, we created a data visualization web application for the scientific community to explore these results (http://humphreyslab.com/SingleCell/).
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