Single-cell RNA sequencing data analysis based on non-uniform ε-neighborhood network.

Ischemic stroke (IS) is a leading cause of adult disability worldwide. The inflammatory processes involved are complex, making it challenging to fully understand the pathological mechanisms of IS. Phagocytosis plays an important role in eliminating neurotoxic or damaged neurons resulting from inflammatory responses. This study employed bioinformatics methods to analyze single-cell RNA sequencing (scRNA-seq) data to investigate the cell types and molecular biological processes involved in IS. scRNA-seq data for IS were obtained from the Gene Expression Omnibus (GEO). Following sample screening and reprocessing, 5,582 single cells were identified from healthy controls and patients with IS. Uniform manifold approximation and projection (UMAP) was utilized to further explore the cellular composition in IS. Functional enrichment analysis of differentially expressed genes was conducted to identify transcriptional regulators, whereas cell developmental trajectories were predicted to uncover potential cell fate decisions. iTALK was employed to identify potential ligand-receptor axes within the cell-type immune microenvironment of IS. Based on scRNA-seq data analysis, we identified four cell types and their associated subclusters, along with genes exhibiting significant differential expression within these subclusters. Phagocytosis was significantly enriched in cell types linked to IS, while the differentiation trajectories of subpopulations in IS was different. Additionally, multiple receptor-ligand axes were identified, indicating diverse interactions within the immune microenvironment of IS. This study demonstrated that phagocytosis in IS cell types critically influences disease progression. It also predicted the trajectories of infarct cells. These findings provide valuable insights into the molecular and cellular mechanisms underlying IS and highlight potential pathways for therapeutic intervention.

Crustacean aquaculture is expected to be a major source of fishery commodities in the near future. Hemocytes are key players of the immune system in shrimps; however, their classification, maturation, and differentiation are still under debate. To date, only discrete and inconsistent information on the classification of shrimp hemocytes has been reported, showing that the morphological characteristics are not sufficient to resolve their actual roles. Our present study using single-cell RNA sequencing revealed six types of hemocytes of Marsupenaeus japonicus based on their transcriptional profiles. We identified markers of each subpopulation and predicted the differentiation pathways involved in their maturation. We also predicted cell growth factors that might play crucial roles in hemocyte differentiation. Different immune roles among these subpopulations were suggested from the analysis of differentially expressed immune-related genes. These results provide a unified classification of shrimp hemocytes, which improves the understanding of its immune system.

BackgroundThis study aimed to use single-cell RNA-seq (scRNA-seq) to discover marker genes in endothelial cells (ECs) and construct a prognostic model for glioblastoma multiforme (GBM) patients in combination with traditional high-throughput RNA sequencing (bulk RNA-seq).MethodsBulk RNA-seq data was downloaded from The Cancer Genome Atlas (TCGA) and The China Glioma Genome Atlas (CGGA) databases. 10x scRNA-seq data for GBM were obtained from the Gene Expression Omnibus (GEO) database. The uniform manifold approximation and projection (UMAP) were used for downscaling and cluster identification. Key modules and differentially expressed genes (DEGs) were identified by weighted gene correlation network analysis (WGCNA). A non-negative matrix decomposition (NMF) algorithm was used to identify the different subtypes based on DEGs, and multivariate cox regression analysis to model the prognosis. Finally, differences in mutational landscape, immune cell abundance, immune checkpoint inhibitors (ICIs)-associated genes, immunotherapy effects, and enriched pathways were investigated between different risk groups.ResultsThe analysis of scRNA-seq data from eight samples revealed 13 clusters and four cell types. After applying Fisher’s exact test, ECs were identified as the most important cell type. The NMF algorithm identified two clusters with different prognostic and immunological features based on DEGs. We finally built a prognostic model based on the expression levels of four key genes. Higher risk scores were significantly associated with poorer survival outcomes, low mutation rates in IDH genes, and upregulation of immune checkpoints such as PD-L1 and CD276.ConclusionWe built and validated a 4-gene signature for GBM using 10 scRNA-seq and bulk RNA-seq data in this work.

A crucial component of intuitive data visualization is presenting a hierarchical tree structure with interactive functions. For example, single-cell transcriptomics studies may generate gene expression values with developmental trajectories or cell lineage structures. Two common visualization methods, t-Distributed Stochastic Neighbor Embedding (t-SNE) and Uniform Manifold Approximation and Projection (UMAP), require two separate figures to depict the distribution of cell types and gene expression data, with low-dimension projections that may not capture the hierarchical structures among cells. Here, we present a JavaScript framework and an interactive web app named Collapsible Tree, which presents values jointly with interactive, expandable, and collapsible lineage structures. For example, the Collapsible Tree presents cellular states and gene expression from single-cell transcriptomics within a single hierarchical plot, enabling comparisons of gene expression across lineages and subtle patterns between sub-lineages. Our framework can facilitate the exploration of complicated value patterns that are not evident in UMAP or t-SNE plots. The Collapsible Tree web interface is available at https://collapsibletree.data2in.net. The JavaScript library source code is available at https://github.com/data2intelligence/collapsible_tree.

Visualization is widely used to help explore and interpret high dimensional single cell (sc) omics data, such as scRNA-seq expression data. In particular, uniform manifold approximation and projection (UMAP) has become nearly ubiquitous in scientific publications that apply single cell omics technologies. Some experts have expressed concerns that the global cell-cell relationship, especially the spatial distances among cell clusters in a dataset, may not be faithfully depicted in a 2-dimensional (2D) UMAP. To help users to better appreciate this issue with their own data, we created an online server for the community to upload their single cell data and interactively make UMAPs with different hyper-parameters to witness how the distribution of cell clusters changes. The server thus can help promote proper usages of UMAP, especially to avoid the common pitfalls in misinterpretation of inter-cluster relationships in single cell studies.Availability and ImplementationShinyUMAP is freely available as an online Shiny server implemented in Python athttps://scviewer.shinyapps.io/shinyUMAP/.

Single-cell transcriptome sequencing, often referred to as single-cell RNA sequencing (scRNA-seq), is used to measure gene expression at the single-cell level and provides a higher resolution of cellular differences than bulk RNA-seq. With more detailed and accurate information, scRNA-seq will greatly promote the understanding of cell functions, disease progression, and treatment response. Although the scRNA-seq experimental protocols have been improved very quickly, many challenges in the scRNA-seq data analysis still need to be overcome. In this chapter, we focus on the introduction and discussion of the research status in the field of scRNA-seq data normalization and cluster analysis, which are the two most important challenges in the scRNA-seq data analysis. Particularly, we present a protocol to discover and validate cancer stem cells (CSCs) using scRNA-seq. Suggestions have also been made to help researchers rationally design their scRNA-seq experiments and data analysis in their future studies.

Learning from potentially infinite and high-dimensional data streams poses significant challenges in the classification task. For instance, k-Nearest Neighbors (kNN) is one of the most often used algorithms in the data stream mining area that proved to be very resource-intensive when dealing with high-dimensional spaces. Uniform Manifold Approximation and Projection (UMAP) is a novel manifold technique and one of the most promising dimension reduction and visualization techniques in the non-streaming setting because of its high performance in comparison with competitors. However, there is no version of UMAP that copes with the challenging context of streams. To overcome these restrictions, we propose a batch-incremental approach that pre-processes data streams using UMAP, by producing successive embeddings on a stream of disjoint batches in order to support an incremental kNN classification. Experiments conducted on publicly available synthetic and real-world datasets demonstrate the substantial gains that can be achieved with our proposal compared to state-of-the-art techniques.

Advances in single-cell technologies have enabled high-resolution dissection of tissue composition. Several tools for dimensionality reduction are available to analyze the large number of parameters generated in single-cell studies. Recently, a nonlinear dimensionality-reduction technique, uniform manifold approximation and projection (UMAP), was developed for the analysis of any type of high-dimensional data. Here we apply it to biological data, using three well-characterized mass cytometry and single-cell RNA sequencing datasets. Comparing the performance of UMAP with five other tools, we find that UMAP provides the fastest run times, highest reproducibility and the most meaningful organization of cell clusters. The work highlights the use of UMAP for improved visualization and interpretation of single-cell data.

Single-cell RNA-seq (scRNA-seq) is a powerful tool for analyzing heterogeneous and functionally diverse cell population. Visualizing scRNA-seq data can help us effectively extract meaningful biological information and identify novel cell subtypes. Currently, the most popular methods for scRNA-seq visualization are principal component analysis (PCA) and t-distributed stochastic neighbor embedding (t-SNE). While PCA is an unsupervised dimension reduction technique, t-SNE incorporates cluster information into pairwise probability, and then maximizes the Kullback–Leibler divergence. Uniform Manifold Approximation and Projection (UMAP) is another recently developed visualization method similar to t-SNE. However, one limitation with UMAP and t-SNE is that they can only capture the local structure of the data, the global structure of the data is not faithfully preserved. In this manuscript, we propose a semisupervised principal component analysis (ssPCA) approach for scRNA-seq visualization. The proposed approach incorporates cluster-labels into dimension reduction and discovers principal components that maximize both data variance and cluster dependence. ssPCA must have cluster-labels as its input. Therefore, it is most useful for visualizing clusters from a scRNA-seq clustering software. Our experiments with simulation and real scRNA-seq data demonstrate that ssPCA is able to preserve both local and global structures of the data, and uncover the transition and progressions in the data, if they exist. In addition, ssPCA is convex and has a global optimal solution. It is also robust and computationally efficient, making it viable for scRNA-seq cluster visualization.

This study compares the effectiveness of various dimensionality reduction (DR) techniques-including Principal Component Analysis (PCA), Uniform Manifold Approximation and Projection (UMAP), Multidimensional Scaling (MDS), and t-Distributed Stochastic Neighbor Embedding (t-SNE)-combined with a Support Vector Machine (SVM) classifier for a diagnostic task. The results showed that applying DR significantly enhanced performance over the baseline SVM (Accuracy: 0.68). Specifically, the fusion of t-SNE and an Autoencoder achieved the highest accuracy at 94 %, outperforming all other configurations. This demonstrates that leveraging advanced non-linear DR and feature learning methods provides a superior and highly accurate framework for classification.

The complexity of scRNA-sequencing datasets highlights the urgent need for enhanced clustering and visualization methods. Here, we propose Stardust, an iterative, force-directed graph layout algorithm that enables the simultaneous embedding of cells and marker genes. Stardust, for the first time, allows a single-stop visualization of cells and marker genes on a single 2D map. While Stardust provides its own visualization pipeline, it can be plugged in with state-of-the-art methods such as Uniform Manifold Approximation and Projection (UMAP) and t-Distributed Stochastic Neighbor Embedding (t-SNE). We benchmarked Stardust against popular visualization and clustering tools on both scRNA-seq and spatial transcriptomics datasets. In all cases, Stardust performs competitively in identifying and visualizing cell types in an accurate and spatially coherent manner.

Wavelet analysis has been recognized as a cutting‐edge and promising tool in the fields of signal processing and data analysis. However, application of wavelet‐based method in single‐cell RNA sequencing (scRNA‐seq) data is little known. Here, we present M‐band wavelet‐based imputation of scRNA‐seq matrix and multi‐view clustering of cells. We applied combination of M‐band wavelet analysis and uniform manifold approximation and projection (UMAP) to a panel of single cell sequencing datasets by breaking up the data matrix into a trend (low frequency or low resolution) component and M‐1 fluctuation (high frequency or high resolution) components. Our hybrid analysis enables us to examine multi‐view clustering of cell types, identity, and functional states. Distinct to standard scRNA‐seq workflow, our wavelet‐based approach is a new addition to resolve the notorious chaotic sparsity of scRNA‐seq matrix and to uncover rare cell types with a fine‐resolution.

Unsupervised clustering is a fundamental step of single-cell RNA-sequencing (scRNA-seq) data analysis. This issue has inspired several clustering methods to classify cells in scRNA-seq data. However, accurate prediction of the cell clusters remains a substantial challenge. In this study, we propose a new algorithm for scRNA-seq data clustering based on Sparse Optimization and low-rank matrix factorization (scSO). We applied our scSO algorithm to analyze multiple benchmark datasets and showed that the cluster number predicted by scSO was close to the number of reference cell types and that most cells were correctly classified. Our scSO algorithm is available at https://github.com/QuKunLab/scSO. Overall, this study demonstrates a potent cell clustering approach that can help researchers distinguish cell types in single- scRNA-seq data.

Numerical modelling increasingly generates massive, high-dimensional spatio-temporal datasets. Exploring such datasets relies on effective visualization. This study presents a generic workflow to (i) project high-dimensional spatio-temporal data on a two-dimensional (2D) plane accurately (ii) compare dimensionality reduction techniques (DRTs) in terms of resolution and computational efficiency (iii) represent 2D projection spatially using a 2D perceptually uniform background color map. Machine learning (ML) based DRTs for data visualization i.e., principal component analysis (PCA), generative topographic mapping (GTM), t-distributed stochastic neighbor embedding (t-SNE) and uniform manifold approximation and projection (UMAP) are compared in terms of accuracy, resolution and computational efficiency to handle massive datasets. The accuracy of visualization is evaluated using a quality metric based on a co-ranking framework. The workflow is applied to an output of an Australian Water Resource Assessment (AWRA) model for Tasmania, Australia. The dataset consists of daily time series of nine components of the water balance at a 5 km grid cell resolution for the year 2017. The case study shows that PCA allows rapid visualization of global data structures, while t-SNE and UMAP allows more accurate representation of local trends. Furthermore, UMAP is computationally more efficient than t-SNE and least affected by the outliers compared to GTM.

In the analysis of single-cell RNA-sequencing (scRNA-seq) data, how to effectively and accurately identify cell clusters from a large number of cell mixtures is still a challenge. Low-rank representation (LRR) method has achieved excellent results in subspace clustering. But in previous studies, most LRR-based methods usually choose the original data matrix as the dictionary. In addition, the methods based on LRR usually use spectral clustering algorithm to complete cell clustering. Therefore, there is a matching problem between the spectral clustering method and the affinity matrix, which is difficult to ensure the optimal effect of clustering. Considering the above two points, we propose the DLNLRR method to better identify the cell type. First, DLNLRR can update the dictionary during the optimization process instead of using the predefined fixed dictionary, so it can realize dictionary learning and LRR learning at the same time. Second, DLNLRR can realize subspace clustering without relying on spectral clustering algorithm, that is, we can perform clustering directly based on the low-rank matrix. Finally, we carry out a large number of experiments on real single-cell datasets and experimental results show that DLNLRR is superior to other scRNA-seq data analysis algorithms in cell type identification.