Revealing fine-scale cellular heterogeneity among spatial context and the functional and structural foundations of tissue architecture is fundamental within biological research and pharmacology. Unlike traditional approaches involving single molecules or bulk omics, cutting-edge, spatially resolved transcriptomics techniques offer near-single-cell or even subcellular resolution within tissues. Massive information across higher dimensions along with position-coordinating labels can better map the whole 3D transcriptional landscape of tissues. In this review, we focus on developments and strategies in spatially resolved transcriptomics, compare the cell and gene throughput and spatial resolution in detail for existing methods, and highlight the enormous potential in biomedical research.
Although complex inflammatory-like alterations are observed around the amyloid plaques of Alzheimer's disease (AD), little is known about the molecular changes and cellular interactions that characterize this response. We investigate here, in an AD mouse model, the transcriptional changes occurring in tissue domains in a 100-μm diameter around amyloid plaques using spatial transcriptomics. We demonstrate early alterations in a gene co-expression network enriched for myelin and oligodendrocyte genes (OLIGs), whereas a multicellular gene co-expression network of plaque-induced genes (PIGs) involving the complement system, oxidative stress, lysosomes, and inflammation is prominent in the later phase of the disease. We confirm the majority of the observed alterations at the cellular level using in situ sequencing on mouse and human brain sections. Genome-wide spatial transcriptomics analysis provides an unprecedented approach to untangle the dysregulated cellular network in the vicinity of pathogenic hallmarks of AD and other brain diseases.
The mechanisms underlying the selective degeneration of motor neurones in amyotrophic lateral sclerosis (ALS) are poorly understood. The aim of this study was to implement spatially resolved RNA sequencing in human post mortem cortical tissue from an ALS patient harbouring the C9orf72 hexanucleotide repeat expansion to identify dysregulated transcripts that may account for differential vulnerabilities of distinct (i) cell types and (ii) brain regions in the pathogenesis of ALS.
Using spatial transcriptomics (ST) we analysed the transcriptome of post mortem brain tissue, with spatial resolution down to 100 μm. Validation of these findings was then performed using BaseScope, an adapted, in situ hybridization technique with single-transcript single-cell-resolution, providing extensive regional and cell-type specific confirmation of these dysregulated transcripts. The validation cohort was then extended to include multiple post mortem brain regions and spinal cord tissue from an extended cohort of C9orf72, sporadic ALS (sALS) and SOD1 ALS cases.
We identified sixteen dysregulated transcripts of proteins that have roles within six disease-related pathways. Furthermore, these complementary molecular pathology techniques converged to identify two spatially dysregulated transcripts, GRM3 and USP47, that are commonly dysregulated across sALS, SOD1 and C9orf72 cases alike.
This study presents the first description of ST in human post mortem cortical tissue from an ALS patient harbouring the C9orf72 hexanucleotide repeat expansion. These data taken together highlight the importance of preserving spatial resolution, facilitating the identification of genes whose dysregulation may in part underlie regional susceptibilities to ALS, crucially highlighting potential therapeutic and diagnostic targets.
Identifying genes that display spatial expression patterns in spatially resolved transcriptomic studies is an important first step toward characterizing the spatial transcriptomic landscape of complex tissues. Here we present a statistical method, SPARK, for identifying spatial expression patterns of genes in data generated from various spatially resolved transcriptomic techniques. SPARK directly models spatial count data through generalized linear spatial models. It relies on recently developed statistical formulas for hypothesis testing, providing effective control of type I errors and yielding high statistical power. With a computationally efficient algorithm, which is based on penalized quasi-likelihood, SPARK is also scalable to datasets with tens of thousands of genes measured on tens of thousands of samples. Analyzing four published spatially resolved transcriptomic datasets using SPARK, we show it can be up to ten times more powerful than existing methods and disclose biological discoveries that otherwise cannot be revealed by existing approaches.
Spatially resolved transcriptomic techniques allow the characterization of spatial organization of cells in tissues, which revolutionize the studies of tissue function and disease pathology. New strategies for detecting spatial gene expression patterns are emerging, and spatially resolved transcriptomic data are accumulating rapidly. However, it is not convenient for biologists to exploit these data due to the diversity of strategies and complexity in data analysis. Here, we present SpatialDB, the first manually curated database for spatially resolved transcriptomic techniques and datasets. The current version of SpatialDB contains 24 datasets (305 sub-datasets) from 5 species generated by 8 spatially resolved transcriptomic techniques. SpatialDB provides a user-friendly web interface for visualization and comparison of spatially resolved transcriptomic data. To further explore these data, SpatialDB also provides spatially variable genes and their functional enrichment annotation. SpatialDB offers a repository for research community to investigate the spatial cellular structure of tissues, and may bring new insights into understanding the cellular microenvironment in disease. SpatialDB is freely available at https://www.spatialomics.org/SpatialDB.
To advance our understanding of the genetic programs that drive cell and tissue specialization, it is necessary to obtain a comprehensive overview of gene expression patterns. Here, we have used spatial transcriptomics to generate high-resolution, anteroposterior gene expression maps of C. elegans males and hermaphrodites. To explore these maps, we have developed computational methods for discovering region- and tissue-specific genes. We have found extensive sex-specific gene expression differences in the germline and sperm and discovered genes that are specifically expressed in the male reproductive tract. These include a group of uncharacterized genes that encode small secreted proteins that are required for male fertility. We conclude that spatial gene expression maps provide a powerful resource for identifying tissue-specific gene functions in C. elegans. Importantly, we found that expression maps from different animals can be precisely aligned, enabling transcriptome-wide comparisons of gene expression patterns.
Considerable progress in sequencing technologies makes it now possible to study the genomic and transcriptomic landscape of single cells. However, to better understand the complexity of multicellular organisms, we must devise ways to perform high-throughput measurements while preserving spatial information about the tissue context or subcellular localization of analysed nucleic acids. In this Innovation article, we summarize pioneering technologies that enable spatially resolved transcriptomics and discuss how these methods have the potential to extend beyond transcriptomics to encompass spatially resolved genomics, proteomics and possibly other omic disciplines.
Transcriptome analysis enables the study of gene expression in human tissues and is a valuable tool to characterise liver function and gene expression dynamics during liver disease, as well as to identify prognostic markers or signatures, and to facilitate discovery of new therapeutic targets. In contrast to whole tissue RNA sequencing analysis, single-cell RNA-sequencing (scRNA-seq) and spatial transcriptomics enables the study of transcriptional activity at the single cell or spatial level. ScRNA-seq has paved the way for the discovery of previously unknown cell types and subtypes in normal and diseased liver, facilitating the study of rare cells (such as liver progenitor cells) and the functional roles of non-parenchymal cells in chronic liver disease and cancer. By adding spatial information to scRNA-seq data, spatial transcriptomics has transformed our understanding of tissue functional organisation and cell-to-cell interactions in situ. These approaches have recently been applied to investigate liver regeneration, organisation and function of hepatocytes and non-parenchymal cells, and to profile the single cell landscape of chronic liver diseases and cancer. Herein, we review the principles and technologies behind scRNA-seq and spatial transcriptomic approaches, highlighting the recent discoveries and novel insights these methodologies have yielded in both liver physiology and disease biology.
Spatial and molecular characteristics determine tissue function, yet high-resolution methods to capture both concurrently are lacking. Here, we developed high-definition spatial transcriptomics, which captures RNA from histological tissue sections on a dense, spatially barcoded bead array. Each experiment recovers several hundred thousand transcript-coupled spatial barcodes at 2-μm resolution, as demonstrated in mouse brain and primary breast cancer. This opens the way to high-resolution spatial analysis of cells and tissues.
The stereotyped spatial architecture of the brain is both beautiful and fundamentally related to its function, extending from gross morphology to individual neuron types, where soma position, dendritic architecture, and axonal projections determine their roles in functional circuitry. Our understanding of the cell types that make up the brain is rapidly accelerating, driven in particular by recent advances in single-cell transcriptomics. However, understanding brain function, development, and disease will require linking molecular cell types to morphological, physiological, and behavioral correlates. Emerging spatially resolved transcriptomic methods promise to fill this gap by localizing molecularly defined cell types in tissues, with simultaneous detection of morphology, activity, or connectivity. Here, we review the requirements for spatial transcriptomic methods toward these goals, consider the challenges ahead, and describe promising applications.